A Novel Mutation in OTX2 Causes Combined Pituitary Hormone Deficiency, Bilateral Microphthalmia, and Agenesis of the Left Internal Carotid Artery.

PubMed

Shimada, Aya; Takagi, Masaki; Nagashima, Yuka; Miyai, Kentaro; Hasegawa, Yukihiro

2016-01-01

Mutations in OTX2 cause hypopituitarism, ranging from isolated growth hormone deficiency to combined pituitary hormone deficiency (CPHD), which are commonly detected in association with severe eye abnormalities, including anophthalmia or microphthalmia. Pituitary phenotypes of OTX2 mutation carriers are highly variable; however, ACTH deficiency during the neonatal period is not common in previous reports. We report a novel missense OTX2 (R89P) mutation in a CPHD patient with severe hypoglycemia in the neonatal period due to ACTH deficiency, bilateral microphthalmia, and agenesis of the left internal carotid artery (ICA). We identified a novel heterozygous mutation in OTX2 (c.266G>C, p.R89P). R89P OTX2 showed markedly reduced transcriptional activity of HESX1 and POU1F1 reporters compared with wild-type OTX2. A dominant negative effect was noted only in the transcription analysis with POU1F1 promoter. Electrophoretic mobility shift assay experiments showed that R89P OTX2 abrogated DNA-binding ability. OTX2 mutations can cause ACTH deficiency in the neonatal period. Our study also shows that OTX2 mutations are associated with agenesis of the ICA. To the best of our knowledge, this is the first report of a transcription factor gene mutation, which was identified due to agenesis of the ICA of a patient with CPHD. This study extends our understanding of the phenotypic features, molecular mechanism, and developmental course associated with mutations in OTX2. © 2016 S. Karger AG, Basel.

The m.3291T>C mt-tRNALeu(UUR) mutation is definitely pathogenic and causes multisystem mitochondrial disease

PubMed Central

Yarham, John W.; Blakely, Emma L.; Alston, Charlotte L.; Roberts, Mark E.; Ealing, John; Pal, Piyali; Turnbull, Douglass M.; McFarland, Robert; Taylor, Robert W.

2013-01-01

Mitochondrial tRNA point mutations are important causes of human disease, and have been associated with a diverse range of clinical phenotypes. Definitively proving the pathogenicity of any given mt-tRNA mutation requires combined molecular, genetic and functional studies. Subsequent evaluation of the mutation using a pathogenicity scoring system is often very helpful in concluding whether or not the mutation is causing disease. Despite several independent reports linking the m.3291T>C mutation to disease in humans, albeit in association with several different phenotypes, its pathogenicity remains controversial. A lack of conclusive functional evidence and an over-emphasis on the poor evolutionary conservation of the affected nucleotide have contributed to this controversy. Here we describe an adult patient who presented with deafness and lipomas and evidence of mitochondrial abnormalities in his muscle biopsy, who harbours the m.3291T > C mutation, providing conclusive evidence of pathogenicity through analysis of mutation segregation with cytochrome c oxidase (COX) deficiency in single muscle fibres, underlining the importance of performing functional studies when assessing pathogenicity. PMID:23273904

'Laminopathies': A wide spectrum of human diseases

DOE Office of Scientific and Technical Information (OSTI.GOV)

Worman, Howard J.; Bonne, Gisele; Universite Pierre et Marie Curie-Paris 6, Faculte de medecine, Paris F-75013

2007-06-10

Mutations in genes encoding the intermediate filament nuclear lamins and associated proteins cause a wide spectrum of diseases sometimes called 'laminopathies.' Diseases caused by mutations in LMNA encoding A-type lamins include autosomal dominant Emery-Dreifuss muscular dystrophy and related myopathies, Dunnigan-type familial partial lipodystrophy, Charcot-Marie-Tooth disease type 2B1 and developmental and accelerated aging disorders. Duplication in LMNB1 encoding lamin B1 causes autosomal dominant leukodystrophy and mutations in LMNB2 encoding lamin B2 are associated with acquired partial lipodystrophy. Disorders caused by mutations in genes encoding lamin-associated integral inner nuclear membrane proteins include X-linked Emery-Dreifuss muscular dystrophy, sclerosing bone dysplasias, HEM/Greenberg skeletal dysplasiamore » and Pelger-Huet anomaly. While mutations and clinical phenotypes of 'laminopathies' have been carefully described, data explaining pathogenic mechanisms are only emerging. Future investigations will likely identify new 'laminopathies' and a combination of basic and clinical research will lead to a better understanding of pathophysiology and the development of therapies.« less

Target enrichment and high-throughput sequencing of 80 ribosomal protein genes to identify mutations associated with Diamond-Blackfan anaemia.

PubMed

Gerrard, Gareth; Valgañón, Mikel; Foong, Hui En; Kasperaviciute, Dalia; Iskander, Deena; Game, Laurence; Müller, Michael; Aitman, Timothy J; Roberts, Irene; de la Fuente, Josu; Foroni, Letizia; Karadimitris, Anastasios

2013-08-01

Diamond-Blackfan anaemia (DBA) is caused by inactivating mutations in ribosomal protein (RP) genes, with mutations in 13 of the 80 RP genes accounting for 50-60% of cases. The remaining 40-50% cases may harbour mutations in one of the remaining RP genes, but the very low frequencies render conventional genetic screening as challenging. We, therefore, applied custom enrichment technology combined with high-throughput sequencing to screen all 80 RP genes. Using this approach, we identified and validated inactivating mutations in 15/17 (88%) DBA patients. Target enrichment combined with high-throughput sequencing is a robust and improved methodology for the genetic diagnosis of DBA. © 2013 John Wiley & Sons Ltd.

HCN4 mutation as a molecular explanation on patients with bradycardia and non-compaction cardiomyopathy.

PubMed

Millat, Gilles; Janin, Alexandre; de Tauriac, Olivier; Roux, Antoine; Dauphin, Claire

2015-09-01

A very recent study suggested that HCN4 mutations could be associated with sinusal bradycardia and myocardial non compaction. A French family with 3 affected sisters presenting the same clinical phenotype (sinus bradycardia in combination with non compaction cardiomyopathy (NCCM)) have benefited both from a systematic cardiovascular exploration and molecular investigations. The molecular analysis, performed by NGS sequencing, led to identify only one likely-disease causing variation: p.Gly482Arg on HCN4 gene. Our results confirm the genetic evidence for the involvement of the HCN4 mutations in the combined bradycardia-NCCM phenotype and illustrates that, in front of this combined clinical phenotype, HCN4 mutations has to be suspected. Copyright © 2015 Elsevier Masson SAS. All rights reserved.

Resistance to herbicides caused by single amino acid mutations in acetyl-CoA carboxylase in resistant populations of grassy weeds.

PubMed

Jang, SoRi; Marjanovic, Jasmina; Gornicki, Piotr

2013-03-01

Eleven spontaneous mutations of acetyl-CoA carboxylase have been identified in many herbicide-resistant populations of 42 species of grassy weeds, hampering application of aryloxyphenoxypropionate, cyclohexadione and phenylpyrazoline herbicides in agriculture. IC(50) shifts (resistance indices) caused by herbicide-resistant mutations were determined using a recombinant yeast system that allows comparison of the effects of single amino acid mutations in the same biochemical background, avoiding the complexity inherent in the in planta experiments. The effect of six mutations on the sensitivity of acetyl-CoA carboxylase to nine herbicides representing the three chemical classes was studied. A combination of partially overlapping binding sites of the three classes of herbicides and the structure of their variable parts explains cross-resistance among and between the three classes of inhibitors, as well as differences in their specificity. Some degree of resistance was detected for 51 of 54 herbicide/mutation combinations. Introduction of new herbicides targeting acetyl-CoA carboxylase will depend on their ability to overcome the high degree of cross-resistance already existing in weed populations. © 2013 The Authors. New Phytologist © 2013 New Phytologist Trust.

Mutations in XRCC4 cause primordial dwarfism without causing immunodeficiency.

PubMed

Saito, Shinta; Kurosawa, Aya; Adachi, Noritaka

2016-08-01

In successive reports from 2014 to 2015, X-ray repair cross-complementing protein 4 (XRCC4) has been identified as a novel causative gene of primordial dwarfism. XRCC4 is indispensable for non-homologous end joining (NHEJ), the major pathway for repairing DNA double-strand breaks. As NHEJ is essential for V(D)J recombination during lymphocyte development, it is generally believed that abnormalities in XRCC4 cause severe combined immunodeficiency. Contrary to expectations, however, no overt immunodeficiency has been observed in patients with primordial dwarfism harboring XRCC4 mutations. Here, we describe the various XRCC4 mutations that lead to disease and discuss their impact on NHEJ and V(D)J recombination.

A new SETX mutation producing AOA2 in two siblings.

PubMed

Datta, Neil; Hohler, Anna

2013-09-01

In this paper, we document two cases of a new SETX mutation (820:A>G) combined with an established recessive SETX mutation (5927:T>G) causing ataxia with oculomotor apraxia type 2 (AOA2). The patients had a detailed neurological history and examination performed. Radiological imaging was obtained and genetic analysis was obtained. Both siblings demonstrated healthy and normal growth until adolescence. At that time, slowed speech, hypophonia, dysarthria, extraocular muscle dysfunction and some mild choreiform movements began to appear. Family history included some movement disorder difficulties in second degree relatives. The diagnosis of AOA2 was confirmed by genetic testing. We describe a new SETX gene mutation, which when combined with a recognized SETX mutation results in AOA2. The clinical, radiographic and ancillary testing are described.

Genetic causes of isolated and combined pituitary hormone deficiency.

PubMed

Giordano, Mara

2016-12-01

Research over the last 20 years has led to the elucidation of the genetic aetiologies of Isolated Growth Hormone Deficiency (IGHD) and Combined Pituitary Hormone Deficiency (CPHD). The pituitary plays a central role in growth regulation, coordinating the multitude of central and peripheral signals to maintain the body's internal balance. Naturally occurring mutation in humans and in mice have demonstrated a role for several factors in the aetiology of IGHD/CPHD. Mutations in the GH1 and GHRHR genes shed light on the phenotype and pathogenesis of IGHD whereas mutations in transcription factors such as HESX1, PROP1, POU1F1, LHX3, LHX4, GLI2 and SOX3 contributed to the understanding of CPHD. Depending upon the expression patterns of these molecules, the phenotype may consist of isolated hypopituitarism, or more complex disorders such as septo-optic dysplasia (SOD) and holoprosencephaly. Although numerous monogenic causes of growth disorders have been identified, most of the patients with IGHD/CPHD remain with an explained aetiology as shown by the relatively low mutation detection rate. The introduction of novel diagnostic approaches is now leading to the disclosure of novel genetic causes in disorders characterized by pituitary hormone defects. Copyright © 2016. Published by Elsevier Ltd.

A Dual Phenotype of Periventricular Nodular Heterotopia and Frontometaphyseal Dysplasia in One Patient Caused by a Single FLNA Mutation Leading to Two Functionally Different Aberrant Transcripts

PubMed Central

Zenker, Martin; Rauch, Anita; Winterpacht, Andreas; Tagariello, Andreas; Kraus, Cornelia; Rupprecht, Thomas; Sticht, Heinrich; Reis, André

2004-01-01

Two disorders, periventricular nodular heterotopia (PVNH) and a group of skeletal dysplasias belonging to the oto-palato-digital (OPD) spectrum, are caused by FLNA mutations. They are considered mutually exclusive because of the different presumed effects of the respective FLNA gene mutations, leading to loss of function (PVNH) and gain of function (OPD), respectively. We describe here the first patient manifesting PVNH in combination with frontometaphyseal dysplasia, a skeletal dysplasia of the OPD-spectrum. A novel de novo mutation, 7315C→A in exon 45 of the FLNA gene, was identified. It leads to two aberrant transcripts, one full-length transcript with the point mutation causing a substitution of a highly conserved leucine residue (L2439M) and a second shortened transcript lacking 21 bp due to the creation of an ectopic splice donor site in exon 45. We propose that the dual phenotype is caused by two functionally different, aberrant filamin A proteins and therefore represents an exceptional model case of allelic gain-of-function and loss-of-function phenotypes due to a single mutational event. PMID:14988809

De novo point mutations in patients diagnosed with ataxic cerebral palsy

PubMed Central

Parolin Schnekenberg, Ricardo; Perkins, Emma M.; Miller, Jack W.; Davies, Wayne I. L.; D’Adamo, Maria Cristina; Pessia, Mauro; Fawcett, Katherine A.; Sims, David; Gillard, Elodie; Hudspith, Karl; Skehel, Paul; Williams, Jonathan; O’Regan, Mary; Jayawant, Sandeep; Jefferson, Rosalind; Hughes, Sarah; Lustenberger, Andrea; Ragoussis, Jiannis

2015-01-01

Cerebral palsy is a sporadic disorder with multiple likely aetiologies, but frequently considered to be caused by birth asphyxia. Genetic investigations are rarely performed in patients with cerebral palsy and there is little proven evidence of genetic causes. As part of a large project investigating children with ataxia, we identified four patients in our cohort with a diagnosis of ataxic cerebral palsy. They were investigated using either targeted next generation sequencing or trio-based exome sequencing and were found to have mutations in three different genes, KCNC3, ITPR1 and SPTBN2. All the mutations were de novo and associated with increased paternal age. The mutations were shown to be pathogenic using a combination of bioinformatics analysis and in vitro model systems. This work is the first to report that the ataxic subtype of cerebral palsy can be caused by de novo dominant point mutations, which explains the sporadic nature of these cases. We conclude that at least some subtypes of cerebral palsy may be caused by de novo genetic mutations and patients with a clinical diagnosis of cerebral palsy should be genetically investigated before causation is ascribed to perinatal asphyxia or other aetiologies. PMID:25981959

Exome Sequencing Identifies Mitochondrial Alanyl-tRNA Synthetase Mutations in Infantile Mitochondrial Cardiomyopathy

PubMed Central

Götz, Alexandra; Tyynismaa, Henna; Euro, Liliya; Ellonen, Pekka; Hyötyläinen, Tuulia; Ojala, Tiina; Hämäläinen, Riikka H.; Tommiska, Johanna; Raivio, Taneli; Oresic, Matej; Karikoski, Riitta; Tammela, Outi; Simola, Kalle O.J.; Paetau, Anders; Tyni, Tiina; Suomalainen, Anu

2011-01-01

Infantile cardiomyopathies are devastating fatal disorders of the neonatal period or the first year of life. Mitochondrial dysfunction is a common cause of this group of diseases, but the underlying gene defects have been characterized in only a minority of cases, because tissue specificity of the manifestation hampers functional cloning and the heterogeneity of causative factors hinders collection of informative family materials. We sequenced the exome of a patient who died at the age of 10 months of hypertrophic mitochondrial cardiomyopathy with combined cardiac respiratory chain complex I and IV deficiency. Rigorous data analysis allowed us to identify a homozygous missense mutation in AARS2, which we showed to encode the mitochondrial alanyl-tRNA synthetase (mtAlaRS). Two siblings from another family, both of whom died perinatally of hypertrophic cardiomyopathy, had the same mutation, compound heterozygous with another missense mutation. Protein structure modeling of mtAlaRS suggested that one of the mutations affected a unique tRNA recognition site in the editing domain, leading to incorrect tRNA aminoacylation, whereas the second mutation severely disturbed the catalytic function, preventing tRNA aminoacylation. We show here that mutations in AARS2 cause perinatal or infantile cardiomyopathy with near-total combined mitochondrial respiratory chain deficiency in the heart. Our results indicate that exome sequencing is a powerful tool for identifying mutations in single patients and allows recognition of the genetic background in single-gene disorders of variable clinical manifestation and tissue-specific disease. Furthermore, we show that mitochondrial disorders extend to prenatal life and are an important cause of early infantile cardiac failure. PMID:21549344

Snyder-Robinson Syndrome: Rescuing the Disease-Causing Effect of G56S mutant by Small Molecule Binding

NASA Astrophysics Data System (ADS)

Zhang, Zhe; Martiny, Virginie; Lagorce, David; Alexov, Emil; Miteva, Maria; Clemson University Team; Université Paris Diderot Team

2013-03-01

Snyder-Robinson Syndrome (SRS) is an X-linked mental retardation disorder, which is caused by defects in a particular gene coding for the spermine synthase (SMS) protein. Among the missense mutations known to be disease-causing is the G56S, which is positioned at the interface of the SMS homo-dimer. Previous computational and experimental investigations have shown that G56S mutation destabilizes the homo-dimer and thus greatly reduces the SMS enzymatic activity. In this study, we explore the possibility of mitigating the effect of G56S mutation by binding small molecules to suitable pockets around the mutation site. It is done by combined efforts of molecular dynamics simulations and in silico screening. The binding of selected molecules was calculated to fully compensate the effect of the mutation and rescue the wild type dimer affinity. This work was supported by NIH, NLM grant. No. 1R03LM009748

Identification of new molecular alterations in Fatal Familial Insomnia

USDA-ARS?s Scientific Manuscript database

Fatal Familial Insomnia (FFI) is a rare disease caused by a D178N mutation in combination with methionine (Met) at codon 129 in the mutated allele of PRNP (D178N-129M haplotype). FFI is manifested by sleep disturbances with insomnia, autonomic disorders, hallucinations, delirium, and spontaneous and...

Mutant NDUFS3 subunit of mitochondrial complex I causes Leigh syndrome.

PubMed

Bénit, P; Slama, A; Cartault, F; Giurgea, I; Chretien, D; Lebon, S; Marsac, C; Munnich, A; Rötig, A; Rustin, P

2004-01-01

Respiratory chain complex I deficiency represents a genetically heterogeneous group of diseases resulting from mutations in mitochondrial or nuclear genes. Mutations have been reported in 13 of the 14 subunits encoding the core of complex I (seven mitochondrial and six nuclear genes) and these result in Leigh or Leigh-like syndromes or cardiomyopathy. In this study, a combination of denaturing high performance liquid chromatography and sequence analysis was used to study the NDUFS3 gene in a series of complex I deficient patients. Mutations found in this gene (NADH dehydrogenase iron-sulphur protein 3), coding for the seventh and last subunit of complex I core, were shown to cause late onset Leigh syndrome, optic atrophy, and complex I deficiency. A biochemical diagnosis of complex I deficiency on cultured amniocytes from a later pregnancy was confirmed through the identification of disease causing NDUFS3 mutations in these cells. While mutations in the NDUFS3 gene thus result in Leigh syndrome, a dissimilar clinical phenotype is observed in mutations in the NDUFV2 and NDUFS2 genes, resulting in encephalomyopathy and cardiomyopathy. The reasons for these differences are uncertain.

CDKL5 and ARX mutations in males with early-onset epilepsy.

PubMed

Mirzaa, Ghayda M; Paciorkowski, Alex R; Marsh, Eric D; Berry-Kravis, Elizabeth M; Medne, Livija; Alkhateeb, Asem; Grix, Art; Wirrell, Elaine C; Powell, Berkley R; Nickels, Katherine C; Burton, Barbara; Paras, Andrea; Kim, Katherine; Chung, Wendy; Dobyns, William B; Das, Soma

2013-05-01

Mutations in CDKL5 and ARX are known causes of early-onset epilepsy and severe developmental delay in males and females. Although numerous males with ARX mutations associated with various phenotypes have been reported in the literature, the majority of CDKL5 mutations have been identified in females with a phenotype characterized by early-onset epilepsy, severe global developmental delay, absent speech, and stereotypic hand movements. To date, only 10 males with CDKL5 mutations have been reported. Our retrospective study reports on the clinical, neuroimaging, and molecular findings of 18 males with early-onset epilepsy caused by either CDKL5 or ARX mutations. These 18 patients include eight new males with CDKL5 mutations and 10 with ARX mutations identified through sequence analysis of 266 and 346 males, respectively, at our molecular diagnostic laboratory. Our large dataset therefore expands on the number of reported males with CDKL5 mutations and highlights that aberrations of CDKL5 and ARX combined are an important consideration in the genetic forms of early-onset epilepsy in boys. Copyright © 2013 Elsevier Inc. All rights reserved.

CDKL5 and ARX mutations in males with early-onset epilepsy

PubMed Central

Mirzaa, Ghayda M.; Paciorkowski, Alex R.; Marsh, Eric D.; Berry-Kravis, Elizabeth M.; Medne, Livija; Grix, Art; Wirrell, Elaine C.; Powell, Berkley R.; Nickels, Katherine C.; Burton, Barbara; Paras, Andrea; Kim, Katherine; Chung, Wendy; Dobyns, William B.; Das, Soma

2013-01-01

Mutations in CDKL5 and ARX are known causes of early-onset epilepsy and severe developmental delay in males and females. While numerous males with ARX mutations associated with various phenotypes have been reported in the literature, the majority of CDKL5 mutations have been identified in females with a phenotype characterized by early-onset epilepsy, severe global developmental delay, absent speech, and stereotypic hand movements. To date, only ten males with CDKL5 mutations have been reported. Our retrospective study reports on the clinical, neuroimaging and molecular findings of 18 males with early-onset epilepsy caused by either CDKL5 or ARX mutations. The 18 patients include eight new males with CDKL5 mutations and ten with ARX mutations identified through sequence analysis of 266 and 346 males, respectively, at our molecular diagnostic laboratory. Our large data set therefore expands on the number of reported males with CDKL5 mutations and highlights that aberrations of CDKL5 and ARX combined are an important consideration in the genetic forms of early-onset epilepsy. PMID:23583054

De novo mutations in regulatory elements in neurodevelopmental disorders

PubMed Central

Short, Patrick J.; McRae, Jeremy F.; Gallone, Giuseppe; Sifrim, Alejandro; Won, Hyejung; Geschwind, Daniel H.; Wright, Caroline F.; Firth, Helen V; FitzPatrick, David R.; Barrett, Jeffrey C.; Hurles, Matthew E.

2018-01-01

We previously estimated that 42% of patients with severe developmental disorders carry pathogenic de novo mutations in coding sequences. The role of de novo mutations in regulatory elements affecting genes associated with developmental disorders, or other genes, has been essentially unexplored. We identified de novo mutations in three classes of putative regulatory elements in almost 8,000 patients with developmental disorders. Here we show that de novo mutations in highly evolutionarily conserved fetal brain-active elements are significantly and specifically enriched in neurodevelopmental disorders. We identified a significant twofold enrichment of recurrently mutated elements. We estimate that, genome-wide, 1-3% of patients without a diagnostic coding variant carry pathogenic de novo mutations in fetal brain-active regulatory elements and that only 0.15% of all possible mutations within highly conserved fetal brain-active elements cause neurodevelopmental disorders with a dominant mechanism. Our findings represent a robust estimate of the contribution of de novo mutations in regulatory elements to this genetically heterogeneous set of disorders, and emphasize the importance of combining functional and evolutionary evidence to identify regulatory causes of genetic disorders. PMID:29562236

Structure-functional prediction and analysis of cancer mutation effects in protein kinases.

PubMed

Dixit, Anshuman; Verkhivker, Gennady M

2014-01-01

A central goal of cancer research is to discover and characterize the functional effects of mutated genes that contribute to tumorigenesis. In this study, we provide a detailed structural classification and analysis of functional dynamics for members of protein kinase families that are known to harbor cancer mutations. We also present a systematic computational analysis that combines sequence and structure-based prediction models to characterize the effect of cancer mutations in protein kinases. We focus on the differential effects of activating point mutations that increase protein kinase activity and kinase-inactivating mutations that decrease activity. Mapping of cancer mutations onto the conformational mobility profiles of known crystal structures demonstrated that activating mutations could reduce a steric barrier for the movement from the basal "low" activity state to the "active" state. According to our analysis, the mechanism of activating mutations reflects a combined effect of partial destabilization of the kinase in its inactive state and a concomitant stabilization of its active-like form, which is likely to drive tumorigenesis at some level. Ultimately, the analysis of the evolutionary and structural features of the major cancer-causing mutational hotspot in kinases can also aid in the correlation of kinase mutation effects with clinical outcomes.

Orkambi® and amplifier co-therapy improves function from a rare CFTR mutation in gene-edited cells and patient tissue.

PubMed

Molinski, Steven V; Ahmadi, Saumel; Ip, Wan; Ouyang, Hong; Villella, Adriana; Miller, John P; Lee, Po-Shun; Kulleperuma, Kethika; Du, Kai; Di Paola, Michelle; Eckford, Paul Dw; Laselva, Onofrio; Huan, Ling Jun; Wellhauser, Leigh; Li, Ellen; Ray, Peter N; Pomès, Régis; Moraes, Theo J; Gonska, Tanja; Ratjen, Felix; Bear, Christine E

2017-09-01

The combination therapy of lumacaftor and ivacaftor (Orkambi ® ) is approved for patients bearing the major cystic fibrosis (CF) mutation: ΔF508 It has been predicted that Orkambi ® could treat patients with rarer mutations of similar "theratype"; however, a standardized approach confirming efficacy in these cohorts has not been reported. Here, we demonstrate that patients bearing the rare mutation: c.3700 A>G, causing protein misprocessing and altered channel function-similar to ΔF508-CFTR, are unlikely to yield a robust Orkambi ® response. While  in silico  and biochemical studies confirmed that this mutation could be corrected and potentiated by lumacaftor and ivacaftor, respectively, this combination led to a minor in vitro response in patient-derived tissue. A CRISPR/Cas9-edited bronchial epithelial cell line bearing this mutation enabled studies showing that an "amplifier" compound, effective in increasing the levels of immature CFTR protein, augmented the Orkambi ® response. Importantly, this "amplifier" effect was recapitulated in patient-derived nasal cultures-providing the first evidence for its efficacy in augmenting Orkambi ® in tissues harboring a rare CF-causing mutation. We propose that this multi-disciplinary approach, including creation of CRISPR/Cas9-edited cells to profile modulators together with validation using primary tissue, will facilitate therapy development for patients with rare CF mutations. © 2017 The Authors. Published under the terms of the CC BY 4.0 license.

A novel mutation in the JH4 domain of JAK3 causing severe combined immunodeficiency complicated by vertebral osteomyelitis.

PubMed

Qamar, Farah; Junejo, Samina; Qureshi, Sonia; Seleman, Michael; Bainter, Wayne; Massaad, Michel; Chou, Janet; Geha, Raif S

2017-10-01

JAK3 is a tyrosine kinase essential for signaling downstream of the common gamma chain subunit shared by multiple cytokine receptors. JAK3 deficiency results in T - B + NK - severe combined immune deficiency (SCID). We report a patient with SCID due to a novel mutation in the JAK3 JH4 domain. The function of the JH4 domain remains unknown. This is the first report of a missense mutation in the JAK3 JH4 domain, thereby demonstrating the importance of the JH4 domain of JAK3 in host immunity. Copyright © 2017 Elsevier Inc. All rights reserved.

Chromosomal Effects on Mutability in the P-M System of Hybrid Dysgenesis in DROSOPHILA MELANOGASTER

PubMed Central

Simmons, Michael J.; Raymond, John D.; Laverty, Todd R.; Doll, Rhonda F.; Raymond, Nancy C.; Kocur, Gordon J.; Drier, Eric A.

1985-01-01

Two manifestations of hybrid dysgenesis were studied in flies with chromosomes derived from two different P strains. In one set of experiments, the occurrence of recessive X-linked lethal mutations in the germ cells of dysgenic males was monitored. In the other, the behavior of an X-linked P-element insertion mutation, sn w, was studied in dysgenic males and also in dysgenic females. The chromosomes of one P strain were more proficient at causing dysgenesis in both sets of experiments. However, there was variation among the chromosomes of each strain in regard to the ability to induce lethals or to destabilize snw. The X chromosome, especially when it came from the stronger P strain, had a pronounced effect on both measures of dysgenesis, but in combination with the major autosomes, these effects were reduced. For the stronger P strain, the autosomes by themselves contributed significantly to the production of X-linked lethals and also had large effects on the behavior of snw, but they did not act additively on these two characters. For this strain, the effects of the autosomes on the X-linked lethal mutation rate suggest that only 1/100 P element transpositions causes a recessive lethal mutation. For the weaker P strain, the autosomes had only slight effects on the behavior of snw and appeared to have negligible effects on the X-linked lethal mutation rate. Combinations of chromosomes from either the strong or the weak P strain affected both aspects of dysgenesis in a nonadditive fashion, suggesting that the P elements on these chromosomes competed with each other for transposase, the P-encoded function that triggers P element activity. Age and sex also influenced the ability of chromosomes and combinations of chromosomes to cause dysgenesis. PMID:3934034

Identification of new molecular alterations in Fatal Familial Insomnia.

USDA-ARS?s Scientific Manuscript database

Fatal familial insomnia (FFI) is an autosomal dominant prion disease caused by a D178N mutation in PRNP in combination with methionine (Met) at codon 129 in the mutated allele of the same gene (D178N-129M haplotype). The present study analyzes pathological and molecular features in seven FFI cases c...

Mutations in LAMA1 Cause Cerebellar Dysplasia and Cysts with and without Retinal Dystrophy

PubMed Central

Aldinger, Kimberly A.; Mosca, Stephen J.; Tétreault, Martine; Dempsey, Jennifer C.; Ishak, Gisele E.; Hartley, Taila; Phelps, Ian G.; Lamont, Ryan E.; O’Day, Diana R.; Basel, Donald; Gripp, Karen W.; Baker, Laura; Stephan, Mark J.; Bernier, Francois P.; Boycott, Kym M.; Majewski, Jacek; Parboosingh, Jillian S.; Innes, A. Micheil; Doherty, Dan

2014-01-01

Cerebellar dysplasia with cysts (CDC) is an imaging finding typically seen in combination with cobblestone cortex and congenital muscular dystrophy in individuals with dystroglycanopathies. More recently, CDC was reported in seven children without neuromuscular involvement (Poretti-Boltshauser syndrome). Using a combination of homozygosity mapping and whole-exome sequencing, we identified biallelic mutations in LAMA1 as the cause of CDC in seven affected individuals (from five families) independent from those included in the phenotypic description of Poretti-Boltshauser syndrome. Most of these individuals also have high myopia, and some have retinal dystrophy and patchy increased T2-weighted fluid-attenuated inversion recovery (T2/FLAIR) signal in cortical white matter. In one additional family, we identified two siblings who have truncating LAMA1 mutations in combination with retinal dystrophy and mild cerebellar dysplasia without cysts, indicating that cysts are not an obligate feature associated with loss of LAMA1 function. This work expands the phenotypic spectrum associated with the lamininopathy disorders and highlights the tissue-specific roles played by different laminin-encoding genes. PMID:25105227

De novo point mutations in patients diagnosed with ataxic cerebral palsy.

PubMed

Parolin Schnekenberg, Ricardo; Perkins, Emma M; Miller, Jack W; Davies, Wayne I L; D'Adamo, Maria Cristina; Pessia, Mauro; Fawcett, Katherine A; Sims, David; Gillard, Elodie; Hudspith, Karl; Skehel, Paul; Williams, Jonathan; O'Regan, Mary; Jayawant, Sandeep; Jefferson, Rosalind; Hughes, Sarah; Lustenberger, Andrea; Ragoussis, Jiannis; Jackson, Mandy; Tucker, Stephen J; Németh, Andrea H

2015-07-01

Cerebral palsy is a sporadic disorder with multiple likely aetiologies, but frequently considered to be caused by birth asphyxia. Genetic investigations are rarely performed in patients with cerebral palsy and there is little proven evidence of genetic causes. As part of a large project investigating children with ataxia, we identified four patients in our cohort with a diagnosis of ataxic cerebral palsy. They were investigated using either targeted next generation sequencing or trio-based exome sequencing and were found to have mutations in three different genes, KCNC3, ITPR1 and SPTBN2. All the mutations were de novo and associated with increased paternal age. The mutations were shown to be pathogenic using a combination of bioinformatics analysis and in vitro model systems. This work is the first to report that the ataxic subtype of cerebral palsy can be caused by de novo dominant point mutations, which explains the sporadic nature of these cases. We conclude that at least some subtypes of cerebral palsy may be caused by de novo genetic mutations and patients with a clinical diagnosis of cerebral palsy should be genetically investigated before causation is ascribed to perinatal asphyxia or other aetiologies. © The Author (2015). Published by Oxford University Press on behalf of the Guarantors of Brain.

Structural Modeling Insights into Human VKORC1 Phenotypes

PubMed Central

Czogalla, Katrin J.; Watzka, Matthias; Oldenburg, Johannes

2015-01-01

Vitamin K 2,3-epoxide reductase complex subunit 1 (VKORC1) catalyses the reduction of vitamin K and its 2,3-epoxide essential to sustain γ-carboxylation of vitamin K-dependent proteins. Two different phenotypes are associated with mutations in human VKORC1. The majority of mutations cause resistance to 4-hydroxycoumarin- and indandione-based vitamin K antagonists (VKA) used in the prevention and therapy of thromboembolism. Patients with these mutations require greater doses of VKA for stable anticoagulation than patients without mutations. The second phenotype, a very rare autosomal-recessive bleeding disorder caused by combined deficiency of vitamin K dependent clotting factors type 2 (VKCFD2) arises from a homozygous Arg98Trp mutation. The bleeding phenotype can be corrected by vitamin K administration. Here, we summarize published experimental data and in silico modeling results in order to rationalize the mechanisms of VKA resistance and VKCFD2. PMID:26287237

Screening for duplications, deletions and a common intronic mutation detects 35% of second mutations in patients with USH2A monoallelic mutations on Sanger sequencing.

PubMed

Steele-Stallard, Heather B; Le Quesne Stabej, Polona; Lenassi, Eva; Luxon, Linda M; Claustres, Mireille; Roux, Anne-Francoise; Webster, Andrew R; Bitner-Glindzicz, Maria

2013-08-08

Usher Syndrome is the leading cause of inherited deaf-blindness. It is divided into three subtypes, of which the most common is Usher type 2, and the USH2A gene accounts for 75-80% of cases. Despite recent sequencing strategies, in our cohort a significant proportion of individuals with Usher type 2 have just one heterozygous disease-causing mutation in USH2A, or no convincing disease-causing mutations across nine Usher genes. The purpose of this study was to improve the molecular diagnosis in these families by screening USH2A for duplications, heterozygous deletions and a common pathogenic deep intronic variant USH2A: c.7595-2144A>G. Forty-nine Usher type 2 or atypical Usher families who had missing mutations (mono-allelic USH2A or no mutations following Sanger sequencing of nine Usher genes) were screened for duplications/deletions using the USH2A SALSA MLPA reagent kit (MRC-Holland). Identification of USH2A: c.7595-2144A>G was achieved by Sanger sequencing. Mutations were confirmed by a combination of reverse transcription PCR using RNA extracted from nasal epithelial cells or fibroblasts, and by array comparative genomic hybridisation with sequencing across the genomic breakpoints. Eight mutations were identified in 23 Usher type 2 families (35%) with one previously identified heterozygous disease-causing mutation in USH2A. These consisted of five heterozygous deletions, one duplication, and two heterozygous instances of the pathogenic variant USH2A: c.7595-2144A>G. No variants were found in the 15 Usher type 2 families with no previously identified disease-causing mutations. In 11 atypical families, none of whom had any previously identified convincing disease-causing mutations, the mutation USH2A: c.7595-2144A>G was identified in a heterozygous state in one family. All five deletions and the heterozygous duplication we report here are novel. This is the first time that a duplication in USH2A has been reported as a cause of Usher syndrome. We found that 8 of 23 (35%) of 'missing' mutations in Usher type 2 probands with only a single heterozygous USH2A mutation detected with Sanger sequencing could be attributed to deletions, duplications or a pathogenic deep intronic variant. Future mutation detection strategies and genetic counselling will need to take into account the prevalence of these types of mutations in order to provide a more comprehensive diagnostic service.

Rift Valley Fever Virus MP-12 Vaccine Is Fully Attenuated by a Combination of Partial Attenuations in the S, M, and L Segments

PubMed Central

Hill, Terence E.; Smith, Jennifer K.; Zhang, Lihong; Juelich, Terry L.; Gong, Bin; Slack, Olga A. L.; Ly, Hoai J.; Lokugamage, Nandadeva; Freiberg, Alexander N.

2015-01-01

ABSTRACT Rift Valley fever (RVF) is a mosquito-borne zoonotic disease endemic to Africa and characterized by a high rate of abortion in ruminants and hemorrhagic fever, encephalitis, or blindness in humans. RVF is caused by Rift Valley fever virus (RVFV; family Bunyaviridae, genus Phlebovirus), which has a tripartite negative-stranded RNA genome (consisting of the S, M, and L segments). Further spread of RVF into countries where the disease is not endemic may affect the economy and public health, and vaccination is an effective approach to prevent the spread of RVFV. A live-attenuated MP-12 vaccine is one of the best-characterized RVF vaccines for safety and efficacy and is currently conditionally licensed for use for veterinary purposes in the United States. Meanwhile, as of 2015, no other RVF vaccine has been conditionally or fully licensed for use in the United States. The MP-12 strain is derived from wild-type pathogenic strain ZH548, and its genome encodes 23 mutations in the three genome segments. However, the mechanism of MP-12 attenuation remains unknown. We characterized the attenuation of wild-type pathogenic strain ZH501 carrying a mutation(s) of the MP-12 S, M, or L segment in a mouse model. Our results indicated that MP-12 is attenuated by the mutations in the S, M, and L segments, while the mutations in the M and L segments confer stronger attenuation than those in the S segment. We identified a combination of 3 amino acid changes, Y259H (Gn), R1182G (Gc), and R1029K (L), that was sufficient to attenuate ZH501. However, strain MP-12 with reversion mutations at those 3 sites was still highly attenuated. Our results indicate that MP-12 attenuation is supported by a combination of multiple partial attenuation mutations and a single reversion mutation is less likely to cause a reversion to virulence of the MP-12 vaccine. IMPORTANCE Rift Valley fever (RVF) is a mosquito-transmitted viral disease that is endemic to Africa and that has the potential to spread into other countries. Vaccination is considered an effective way to prevent the disease, and the only available veterinary RVF vaccine in the United States is a live-attenuated MP-12 vaccine, which is conditionally licensed. Strain MP-12 is different from its parental pathogenic RVFV strain, strain ZH548, because of the presence of 23 mutations. This study determined the role of individual mutations in the attenuation of the MP-12 strain. We found that full attenuation of MP-12 occurs by a combination of multiple mutations. Our findings indicate that a single reversion mutation will less likely cause a major reversion to virulence of the MP-12 vaccine. PMID:25948740

Rift Valley Fever Virus MP-12 Vaccine Is Fully Attenuated by a Combination of Partial Attenuations in the S, M, and L Segments.

PubMed

Ikegami, Tetsuro; Hill, Terence E; Smith, Jennifer K; Zhang, Lihong; Juelich, Terry L; Gong, Bin; Slack, Olga A L; Ly, Hoai J; Lokugamage, Nandadeva; Freiberg, Alexander N

2015-07-01

Rift Valley fever (RVF) is a mosquito-borne zoonotic disease endemic to Africa and characterized by a high rate of abortion in ruminants and hemorrhagic fever, encephalitis, or blindness in humans. RVF is caused by Rift Valley fever virus (RVFV; family Bunyaviridae, genus Phlebovirus), which has a tripartite negative-stranded RNA genome (consisting of the S, M, and L segments). Further spread of RVF into countries where the disease is not endemic may affect the economy and public health, and vaccination is an effective approach to prevent the spread of RVFV. A live-attenuated MP-12 vaccine is one of the best-characterized RVF vaccines for safety and efficacy and is currently conditionally licensed for use for veterinary purposes in the United States. Meanwhile, as of 2015, no other RVF vaccine has been conditionally or fully licensed for use in the United States. The MP-12 strain is derived from wild-type pathogenic strain ZH548, and its genome encodes 23 mutations in the three genome segments. However, the mechanism of MP-12 attenuation remains unknown. We characterized the attenuation of wild-type pathogenic strain ZH501 carrying a mutation(s) of the MP-12 S, M, or L segment in a mouse model. Our results indicated that MP-12 is attenuated by the mutations in the S, M, and L segments, while the mutations in the M and L segments confer stronger attenuation than those in the S segment. We identified a combination of 3 amino acid changes, Y259H (Gn), R1182G (Gc), and R1029K (L), that was sufficient to attenuate ZH501. However, strain MP-12 with reversion mutations at those 3 sites was still highly attenuated. Our results indicate that MP-12 attenuation is supported by a combination of multiple partial attenuation mutations and a single reversion mutation is less likely to cause a reversion to virulence of the MP-12 vaccine. Rift Valley fever (RVF) is a mosquito-transmitted viral disease that is endemic to Africa and that has the potential to spread into other countries. Vaccination is considered an effective way to prevent the disease, and the only available veterinary RVF vaccine in the United States is a live-attenuated MP-12 vaccine, which is conditionally licensed. Strain MP-12 is different from its parental pathogenic RVFV strain, strain ZH548, because of the presence of 23 mutations. This study determined the role of individual mutations in the attenuation of the MP-12 strain. We found that full attenuation of MP-12 occurs by a combination of multiple mutations. Our findings indicate that a single reversion mutation will less likely cause a major reversion to virulence of the MP-12 vaccine. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

A novel NHS mutation causes Nance-Horan Syndrome in a Chinese family.

PubMed

Tian, Qi; Li, Yunping; Kousar, Rizwana; Guo, Hui; Peng, Fenglan; Zheng, Yu; Yang, Xiaohua; Long, Zhigao; Tian, Runyi; Xia, Kun; Lin, Haiying; Pan, Qian

2017-01-07

Nance-Horan Syndrome (NHS) (OMIM: 302350) is a rare X-linked developmental disorder characterized by bilateral congenital cataracts, with occasional dental anomalies, characteristic dysmorphic features, brachymetacarpia and mental retardation. Carrier females exhibit similar manifestations that are less severe than in affected males. Here, we report a four-generation Chinese family with multiple affected individuals presenting Nance-Horan Syndrome. Whole-exome sequencing combined with RT-PCR and Sanger sequencing was used to search for a genetic cause underlying the disease phenotype. Whole-exome sequencing identified in all affected individuals of the family a novel donor splicing site mutation (NM_198270: c.1045 + 2T > A) in intron 4 of the gene NHS, which maps to chromosome Xp22.13. The identified mutation results in an RNA processing defect causing a 416-nucleotide addition to exon 4 of the mRNA transcript, likely producing a truncated NHS protein. The donor splicing site mutation NM_198270: c.1045 + 2T > A of the NHS gene is the causative mutation in this Nance-Horan Syndrome family. This research broadens the spectrum of NHS gene mutations, contributing to our understanding of the molecular genetics of NHS.

Recessive mutations in ELOVL4 cause ichthyosis, intellectual disability, and spastic quadriplegia.

PubMed

Aldahmesh, Mohammed A; Mohamed, Jawahir Y; Alkuraya, Hisham S; Verma, Ishwar C; Puri, Ratna D; Alaiya, Ayodele A; Rizzo, William B; Alkuraya, Fowzan S

2011-12-09

Very-long-chain fatty acids (VLCFAs) play important roles in membrane structure and cellular signaling, and their contribution to human health is increasingly recognized. Fatty acid elongases catalyze the first and rate-limiting step in VLCFA synthesis. Heterozygous mutations in ELOVL4, the gene encoding one of the elongases, are known to cause macular degeneration in humans and retinal abnormalities in mice. However, biallelic ELOVL4 mutations have not been observed in humans, and murine models with homozygous mutations die within hours of birth as a result of a defective epidermal water barrier. Here, we report on two human individuals with recessive ELOVL4 mutations revealed by a combination of autozygome analysis and exome sequencing. These individuals exhibit clinical features of ichthyosis, seizures, mental retardation, and spasticity-a constellation that resembles Sjögren-Larsson syndrome (SLS) but presents a more severe neurologic phenotype. Our findings identify recessive mutations in ELOVL4 as the cause of a neuro-ichthyotic disease and emphasize the importance of VLCFA synthesis in brain and cutaneous development. Copyright © 2011 The American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.

A novel mutation in HESX1 causes combined pituitary hormone deficiency without septo optic dysplasia phenotypes.

PubMed

Takagi, Masaki; Takahashi, Mai; Ohtsu, Yoshiaki; Sato, Takeshi; Narumi, Satoshi; Arakawa, Hirokazu; Hasegawa, Tomonobu

2016-04-25

Heterozygous and/or homozygous HESX1 mutations have been reported to cause isolated growth hormone deficiency (IGHD) or combined pituitary hormone deficiency (CPHD), in association with septo optic dysplasia (SOD). We report a novel heterozygous HESX1 mutation in a CPHD patient without SOD phenotypes. The propositus was a one-year-old Japanese girl. Shortly after birth, she was found to be hypoglycemic. She was diagnosed with central adrenal insufficiency based on low cortisol and ACTH at a time of severe hypoglycemia. Further endocrine studies indicated that the patient also had central hypothyroidism and growth hormone deficiency. Using a next-generation sequencing strategy, we identified a novel heterozygous HESX1 mutation, c.326G>A (p.Arg109Gln). Western blotting and subcellular localization revealed no significant difference between wild type and mutant HESX1. Electrophoretic mobility shift assays showed that the mutant HESX1 abrogated DNA-binding ability. Mutant HESX1 was unable to repress PROP1-mediated activation. In conclusion, this study identified Arg109 as a critical residue in the HESX1 protein and extends our understanding of the phenotypic features, molecular mechanism, and developmental course associated with mutations in HESX1. When multiple genes need to be analyzed for mutations simultaneously, targeted sequence analysis of interesting genomic regions is an attractive approach.

Mutations in PROP1 cause familial combined pituitary hormone deficiency.

PubMed

Wu, W; Cogan, J D; Pfäffle, R W; Dasen, J S; Frisch, H; O'Connell, S M; Flynn, S E; Brown, M R; Mullis, P E; Parks, J S; Phillips, J A; Rosenfeld, M G

1998-02-01

Combined pituitary hormone deficiency (CPHD) in man denotes impaired production of growth hormone (GH) and one or more of the other five anterior pituitary hormones. Mutations of the pituitary transcription factor gene POU1F1 (the human homologue of mouse Pit1) are responsible for deficiencies of GH, prolactin and thyroid stimulating hormone (TSH) in Snell and Jackson dwarf mice and in man, while the production of adrenocorticotrophic hormone (ACTH), luteinizing hormone (LH) and follicle stimulating hormone (FSH) is preserved. The Ames dwarf (df) mouse displays a similar phenotype, and appears to be epistatic to Snell and Jackson dwarfism. We have recently positionally cloned the putative Ames dwarf gene Prop1, which encodes a paired-like homeodomain protein that is expressed specifically in embryonic pituitary and is necessary for Pit1 expression. In this report, we have identified four CPHD families with homozygosity or compound heterozygosity for inactivating mutations of PROP1. These mutations in the human PROP1 gene result in a gene product with reduced DNA-binding and transcriptional activation ability in comparison to the product of the murine df mutation. In contrast to individuals with POU1F1 mutations, those with PROP1 mutations cannot produce LH and FSH at a sufficient level and do not enter puberty spontaneously. Our results identify a major cause of CPHD in humans and suggest a direct or indirect role for PROP1 in the ontogenesis of pituitary gonadotropes, as well as somatotropes, lactotropes and caudomedial thyrotropes.

A combination of two truncating mutations in USH2A causes more severe and progressive hearing impairment in Usher syndrome type IIa.

PubMed

Hartel, Bas P; Löfgren, Maria; Huygen, Patrick L M; Guchelaar, Iris; Lo-A-Njoe Kort, Nicole; Sadeghi, Andre M; van Wijk, Erwin; Tranebjærg, Lisbeth; Kremer, Hannie; Kimberling, William J; Cremers, Cor W R J; Möller, Claes; Pennings, Ronald J E

2016-09-01

Usher syndrome is an inherited disorder that is characterized by hearing impairment (HI), retinitis pigmentosa, and in some cases vestibular dysfunction. Usher syndrome type IIa is caused by mutations in USH2A. HI in these patients is highly heterogeneous and the present study evaluates the effects of different types of USH2A mutations on the audiometric phenotype. Data from two large centres of expertise on Usher Syndrome in the Netherlands and Sweden were combined in order to create a large combined sample of patients to identify possible genotype-phenotype correlations. A retrospective study on HI in 110 patients (65 Dutch and 45 Swedish) genetically diagnosed with Usher syndrome type IIa. We used methods especially designed for characterizing and testing differences in audiological phenotype between patient subgroups. These methods included Age Related Typical Audiograms (ARTA) and a method to evaluate the difference in the degree of HI developed throughout life between subgroups. Cross-sectional linear regression analysis of last-visit audiograms for the best hearing ear demonstrated a gradual decline of hearing over decades. The congenital level of HI was in the range of 16-33 dB at 0.25-0.5 kHz, and in the range of 51-60 dB at 1-8 kHz. The annual threshold deterioration was in the range of 0.4-0.5 dB/year at 0.25-2 kHz and in the range of 0.7-0.8 dB/year at 4-8 kHz. Patients with two truncating mutations, including homozygotes for the common c.2299delG mutation, developed significantly more severe HI throughout life than patients with one truncating mutation combined with one nontruncating mutation, and patients with two nontruncating mutations. The results have direct implications for patient counselling in terms of prognosis of hearing and may serve as baseline measures for future (genetic) therapeutic interventions. Copyright © 2016 Elsevier B.V. All rights reserved.

Clinical and functional characterization of TNNT2 mutations identified in patients with dilated cardiomyopathy

PubMed Central

Hershberger, Ray E.; Pinto, Jose Renato; Parks, Sharie B.; Kushner, Jessica D.; Li, Duanxiang; Ludwigsen, Susan; Cowan, Jason; Morales, Ana; Parvatiyar, Michelle S.; Potter, James D.

2009-01-01

Background A key issue for cardiovascular genetic medicine is ascertaining if a putative mutation indeed causes dilated cardiomyopathy (DCM). This is critically important as genetic DCM, usually presenting with advanced, life-threatening disease, may be preventable with early intervention in relatives known to carry the mutation. Methods and Results We recently undertook bidirectional resequencing of TNNT2, the cardiac troponin T gene, in 313 probands with DCM. We identified six TNNT2 protein-altering variants in nine probands, all who had early onset, aggressive disease. Additional family members of mutation carriers were then studied when available. Four of the nine probands had DCM without a family history, and five had familial DCM. Only one mutation (Lys210del) could be attributed as definitively causative from prior reports. Four of the five missense mutations were novel (Arg134Gly, Arg151Cys, Arg159Gln, Arg205Trp), and one was previously reported with hypertrophic cardiomyopathy (Glu244Asp). Based on the clinical, pedigree and molecular genetic data these five mutations were considered possibly or likely disease causing. To further clarify their potential pathophysiologic impact, we undertook functional studies of these mutations in cardiac myocytes reconstituted with mutant troponin T proteins. We observed decreased Ca2+ sensitivity of force development, a hallmark of DCM, in support of the conclusion that these mutations are disease-causing. Conclusions We conclude that the combination of clinical, pedigree, molecular genetic and functional data strengthen the interpretation of TNNT2 mutations in DCM. PMID:20031601

High prevalence of mutations affecting the splicing process in a Spanish cohort with autosomal dominant retinitis pigmentosa

PubMed Central

Ezquerra-Inchausti, Maitane; Barandika, Olatz; Anasagasti, Ander; Irigoyen, Cristina; López de Munain, Adolfo; Ruiz-Ederra, Javier

2017-01-01

Retinitis pigmentosa is the most frequent group of inherited retinal dystrophies. It is highly heterogeneous, with more than 80 disease-causing genes 27 of which are known to cause autosomal dominant RP (adRP), having been identified. In this study a total of 29 index cases were ascertained based on a family tree compatible with adRP. A custom panel of 31 adRP genes was analysed by targeted next-generation sequencing using the Ion PGM platform in combination with Sanger sequencing. This allowed us to detect putative disease-causing mutations in 14 out of the 29 (48.28%) families analysed. Remarkably, around 38% of all adRP cases analysed showed mutations affecting the splicing process, mainly due to mutations in genes coding for spliceosome factors (SNRNP200 and PRPF8) but also due to splice-site mutations in RHO. Twelve of the 14 mutations found had been reported previously and two were novel mutations found in PRPF8 in two unrelated patients. In conclusion, our results will lead to more accurate genetic counselling and will contribute to a better characterisation of the disease. In addition, they may have a therapeutic impact in the future given the large number of studies currently underway based on targeted RNA splicing for therapeutic purposes. PMID:28045043

Structure-Functional Prediction and Analysis of Cancer Mutation Effects in Protein Kinases

PubMed Central

Dixit, Anshuman; Verkhivker, Gennady M.

2014-01-01

A central goal of cancer research is to discover and characterize the functional effects of mutated genes that contribute to tumorigenesis. In this study, we provide a detailed structural classification and analysis of functional dynamics for members of protein kinase families that are known to harbor cancer mutations. We also present a systematic computational analysis that combines sequence and structure-based prediction models to characterize the effect of cancer mutations in protein kinases. We focus on the differential effects of activating point mutations that increase protein kinase activity and kinase-inactivating mutations that decrease activity. Mapping of cancer mutations onto the conformational mobility profiles of known crystal structures demonstrated that activating mutations could reduce a steric barrier for the movement from the basal “low” activity state to the “active” state. According to our analysis, the mechanism of activating mutations reflects a combined effect of partial destabilization of the kinase in its inactive state and a concomitant stabilization of its active-like form, which is likely to drive tumorigenesis at some level. Ultimately, the analysis of the evolutionary and structural features of the major cancer-causing mutational hotspot in kinases can also aid in the correlation of kinase mutation effects with clinical outcomes. PMID:24817905

TGFB2 mutations cause familial thoracic aortic aneurysms and dissections associated with mild systemic features of Marfan syndrome.

PubMed

Boileau, Catherine; Guo, Dong-Chuan; Hanna, Nadine; Regalado, Ellen S; Detaint, Delphine; Gong, Limin; Varret, Mathilde; Prakash, Siddharth K; Li, Alexander H; d'Indy, Hyacintha; Braverman, Alan C; Grandchamp, Bernard; Kwartler, Callie S; Gouya, Laurent; Santos-Cortez, Regie Lyn P; Abifadel, Marianne; Leal, Suzanne M; Muti, Christine; Shendure, Jay; Gross, Marie-Sylvie; Rieder, Mark J; Vahanian, Alec; Nickerson, Deborah A; Michel, Jean Baptiste; Jondeau, Guillaume; Milewicz, Dianna M

2012-07-08

A predisposition for thoracic aortic aneurysms leading to acute aortic dissections can be inherited in families in an autosomal dominant manner. Genome-wide linkage analysis of two large unrelated families with thoracic aortic disease followed by whole-exome sequencing of affected relatives identified causative mutations in TGFB2. These mutations-a frameshift mutation in exon 6 and a nonsense mutation in exon 4-segregated with disease with a combined logarithm of odds (LOD) score of 7.7. Sanger sequencing of 276 probands from families with inherited thoracic aortic disease identified 2 additional TGFB2 mutations. TGFB2 encodes transforming growth factor (TGF)-β2, and the mutations are predicted to cause haploinsufficiency for TGFB2; however, aortic tissue from cases paradoxically shows increased TGF-β2 expression and immunostaining. Thus, haploinsufficiency for TGFB2 predisposes to thoracic aortic disease, suggesting that the initial pathway driving disease is decreased cellular TGF-β2 levels leading to a secondary increase in TGF-β2 production in the diseased aorta.

Homozygous single base deletion in TUSC3 causes intellectual disability with developmental delay in an Omani family.

PubMed

Al-Amri, Ahmed; Saegh, Abeer Al; Al-Mamari, Watfa; El-Asrag, Mohammed E; Ivorra, Jose L; Cardno, Alastair G; Inglehearn, Chris F; Clapcote, Steven J; Ali, Manir

2016-07-01

Intellectual disability (ID) is the term used to describe a diverse group of neurological conditions with congenital or juvenile onset, characterized by an IQ score of less than 70 and difficulties associated with limitations in cognitive function and adaptive behavior. The condition can be inherited or caused by environmental factors. The genetic forms are heterogeneous, with mutations in over 500 known genes shown to cause the disorder. We report a consanguineous Omani family in which multiple individuals have ID and developmental delay together with some variably present features including short stature, microcephaly, moderate facial dysmorphism, and congenital malformations of the toes or hands. Homozygosity mapping combined with whole exome next generation sequencing identified a novel homozygous single base pair deletion in TUSC3, c.222delA, p.R74 fs. The mutation segregates with the disease phenotype in a recessive manner and is absent in 60,706 unrelated individuals from various disease-specific and population genetic studies. TUSC3 mutations have been previously identified as causing either syndromic or non-syndromic ID in patients from France, Italy, Iran and Pakistan. This paper supports the previous clinical descriptions of the condition caused by TUSC3 mutations and describes the seventh family with mutations in this gene, thus contributing to the genetic spectrum of mutations. This is the first report of a family from the Arabian peninsula with this form of ID. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Structure-Based Systematic Isolation of Conditional-Lethal Mutations in the Single Yeast Calmodulin Gene

PubMed Central

Ohya, Y.; Botstein, D.

1994-01-01

Conditional-lethal mutations of the single calmodulin gene in Saccharomyces cerevisiae have been very difficult to isolate by random and systematic methods, despite the fact that deletions cause recessive lethality. We report here the isolation of numerous conditional-lethal mutants that were recovered by systematically altering phenylalanine residues. The phenylalanine residues of calmodulin were implicated in function both by structural studies of calmodulin bound to target peptides and by their extraordinary conservation in evolution. Seven single and 26 multiple Phe -> Ala mutations were constructed. Mutant phenotypes were examined in a haploid cmd1 disrupted strain under three conditions: single copy, low copy, and overexpressed. Whereas all but one of the single mutations caused no obvious phenotype, most of the multiple mutations caused obvious growth phenotypes. Five were lethal, 6 were lethal only in synthetic medium, 13 were temperature-sensitive lethal and 2 had no discernible phenotypic consequences. Overexpression of some of the mutant genes restored the phenotype to nearly wild type. Several temperature-sensitive calmodulin mutations were suppressed by elevated concentration of CaCl(2) in the medium. Mutant calmodulin protein was detected at normal levels in extracts of most of the lethal mutant cells, suggesting that the deleterious phenotypes were due to loss of the calmodulin function and not protein instability. Analysis of diploid strains heterozygous for all combinations of cmd1-ts alleles revealed four intragenic complementation groups. The contributions of individual phe->ala changes to mutant phenotypes support the idea of internal functional redundancy in the symmetrical calmodulin protein molecule. These results suggest that the several phenylalanine residues in calmodulin are required to different extents in different combinations in order to carry out each of the several essential tasks. PMID:7896089

β-Myosin heavy chain variant Val606Met causes very mild hypertrophic cardiomyopathy in mice, but exacerbates HCM phenotypes in mice carrying other HCM mutations.

PubMed

Blankenburg, Robert; Hackert, Katarzyna; Wurster, Sebastian; Deenen, René; Seidman, J G; Seidman, Christine E; Lohse, Martin J; Schmitt, Joachim P

2014-07-07

Approximately 40% of hypertrophic cardiomyopathy (HCM) is caused by heterozygous missense mutations in β-cardiac myosin heavy chain (β-MHC). Associating disease phenotype with mutation is confounded by extensive background genetic and lifestyle/environmental differences between subjects even from the same family. To characterize disease caused by β-cardiac myosin heavy chain Val606Met substitution (VM) that has been identified in several HCM families with wide variation of clinical outcomes, in mice. Unlike 2 mouse lines bearing the malignant myosin mutations Arg453Cys (RC/+) or Arg719Trp (RW/+), VM/+ mice with an identical inbred genetic background lacked hallmarks of HCM such as left ventricular hypertrophy, disarray of myofibers, and interstitial fibrosis. Even homozygous VM/VM mice were indistinguishable from wild-type animals, whereas RC/RC- and RW/RW-mutant mice died within 9 days after birth. However, hypertrophic effects of the VM mutation were observed both in mice treated with cyclosporine, a known stimulator of the HCM response, and compound VM/RC heterozygous mice, which developed a severe HCM phenotype. In contrast to all heterozygous mutants, both systolic and diastolic function of VM/RC hearts was severely impaired already before the onset of cardiac remodeling. The VM mutation per se causes mild HCM-related phenotypes; however, in combination with other HCM activators it exacerbates the HCM phenotype. Double-mutant mice are suitable for assessing the severity of benign mutations. © 2014 American Heart Association, Inc.

A Novel Missense Mutation p.Gly162Glu of the Gene MYL2 Involved in Hypertrophic Cardiomyopathy: A Pedigree Analysis of a Proband.

PubMed

Renaudin, Pauline; Janin, Alexandre; Millat, Gilles; Chevalier, Philippe

2018-04-01

Hypertrophic cardiomyopathy (HCM), a common and clinically heterogeneous disease characterized by unexplained ventricular myocardial hypertrophy, is mostly caused by mutations in sarcomeric genes. Identifying the genetic cause is important for management, therapy, and genetic counseling. A molecular diagnosis was performed on a 51-year-old woman diagnosed with HCM using a next-generation sequencing workflow based on a panel designed for sequencing the most prevalent cardiomyopathy-causing genes. Segregation analysis was performed on the woman's family. A novel myosin regulatory light chain (MYL2) missense variant, NM_000432.3:c485G>A, p.Gly162Glu, was identified and firstly considered as a putative pathogenic mutation. Among the 27 family members tested, 16 were carriers for the MYL2-p.Gly162Glu mutation, of whom 12 with the phenotype were positive. None of the 11 family members without mutation had cardiomyopathy. Genetic analysis combined with a segregation study allowed us to classify this novel MYL2 variation, p.Gly162Glu, as a novel pathogenic mutation leading to a familial form of HCM. Due to absence of fast in vitro approaches to evaluate the functional impact of missense variants on HCM-causing genes, segregation studies remain, when possible, the easiest approach to evaluate the putative pathogenicity of novel gene variants, more particularly missense ones.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Koivisto, U.M.; Viikari, J.S.; Kontula, K.

Two deletions of the low-density lipoprotein (LDL) receptor gene were previously shown to account for about two thirds of all mutations causing familial hypercholesterolemia (FH) in Finland. We screened the DNA samples from a cohort representing the remaining 30% of Finnish heterozygous FH patients by amplifying all the 18 exons of the receptor gene by PCR and searching for DNA variations with the SSCP technique. Ten novel mutations were identified, comprising two nonsense and seven missense mutations as well as one frameshift mutation caused by a 13-bp deletion. A single nucleotide change, substituting adenine for guanidine at position 2533 andmore » resulting in an amino acid change of glycine to aspartic acid at codon 823, was found in DNA samples from 14 unrelated FH probands. This mutation (FH-Turku) affects the sequence encoding the putative basolateral sorting signal of the LDL receptor protein; however, the exact functional consequences of this mutation are yet to be examined. The FH-Turku gene and another point mutation (Leu380{r_arrow}His or FH-Pori) together account for {approximately}8% of the FH-causing genes in Finland and are particularly common among FH patients from the southwestern part of the country (combined, 30%). Primer-introduced restriction analysis was applied for convenient assay of the FH-Turku and FH-Pori point mutations. In conclusion, this paper demonstrates the unique genetic background of FH in Finland and describes a commonly occurring FH gene with a missense mutation closest to the C terminus thus far reported. 32 refs., 5 figs., 2 tabs.« less

Clinical and pathological features of amyotrophic lateral sclerosis caused by mutation in the C9ORF72 gene on chromosome 9p.

PubMed

Stewart, Heather; Rutherford, Nicola J; Briemberg, Hannah; Krieger, Charles; Cashman, Neil; Fabros, Marife; Baker, Matt; Fok, Alice; DeJesus-Hernandez, Mariely; Eisen, Andrew; Rademakers, Rosa; Mackenzie, Ian R A

2012-03-01

Two studies recently identified a GGGGCC hexanucleotide repeat expansion in a non-coding region of the chromosome 9 open-reading frame 72 gene (C9ORF72) as the cause of chromosome 9p-linked amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). In a cohort of 231 probands with ALS, we identified the C9ORF72 mutation in 17 familial (27.4%) and six sporadic (3.6%) cases. Patients with the mutation presented with typical motor features of ALS, although subjects with the C9ORF72 mutation had more frequent bulbar onset, compared to those without this mutation. Dementia was significantly more common in ALS patients and families with the C9ORF72 mutation and was usually early-onset FTD. There was striking clinical heterogeneity among the members of individual families with the mutation. The associated neuropathology was a combination of ALS with TDP-ir inclusions and FTLD-TDP. In addition to TDP-43-immunoreactive pathology, a consistent and specific feature of cases with the C9ORF72 mutation was the presence of ubiquitin-positive, TDP-43-negative inclusions in a variety of neuroanatomical regions, such as the cerebellar cortex. These findings support the C9ORF72 mutation as an important newly recognized cause of ALS, provide a more detailed characterization of the associated clinical and pathological features and further demonstrate the clinical and molecular overlap between ALS and FTD.

Recessive myosin myopathy with external ophthalmoplegia associated with MYH2 mutations.

PubMed

Tajsharghi, Homa; Hammans, Simon; Lindberg, Christopher; Lossos, Alexander; Clarke, Nigel F; Mazanti, Ingrid; Waddell, Leigh B; Fellig, Yakov; Foulds, Nicola; Katifi, Haider; Webster, Richard; Raheem, Olayinka; Udd, Bjarne; Argov, Zohar; Oldfors, Anders

2014-06-01

Myosin myopathies comprise a group of inherited diseases caused by mutations in myosin heavy chain (MyHC) genes. Homozygous or compound heterozygous truncating MYH2 mutations have been demonstrated to cause recessive myopathy with ophthalmoplegia, mild-to-moderate muscle weakness and complete lack of type 2A muscle fibers. In this study, we describe for the first time the clinical and morphological characteristics of recessive myosin IIa myopathy associated with MYH2 missense mutations. Seven patients of five different families with a myopathy characterized by ophthalmoplegia and mild-to-moderate muscle weakness were investigated. Muscle biopsy was performed to study morphological changes and MyHC isoform expression. Five of the patients were homozygous for MYH2 missense mutations, one patient was compound heterozygous for a missense and a nonsense mutation and one patient was homozygous for a frame-shift MYH2 mutation. Muscle biopsy demonstrated small or absent type 2A muscle fibers and reduced or absent expression of the corresponding MyHC IIa transcript and protein. We conclude that mild muscle weakness and ophthalmoplegia in combination with muscle biopsy demonstrating small or absent type 2A muscle fibers are the hallmark of recessive myopathy associated with MYH2 mutations.

Analysis of 24 genes reveals a monogenic cause in 11.1% of cases with steroid-resistant nephrotic syndrome at a single center.

PubMed

Tan, Weizhen; Lovric, Svjetlana; Ashraf, Shazia; Rao, Jia; Schapiro, David; Airik, Merlin; Shril, Shirlee; Gee, Heon Yung; Baum, Michelle; Daouk, Ghaleb; Ferguson, Michael A; Rodig, Nancy; Somers, Michael J G; Stein, Deborah R; Vivante, Asaf; Warejko, Jillian K; Widmeier, Eugen; Hildebrandt, Friedhelm

2018-02-01

Steroid-resistant nephrotic syndrome (SRNS) is the second most frequent cause of end-stage renal disease (ESRD) among patients manifesting at under 25 years of age. We performed mutation analysis using a high-throughput PCR-based microfluidic technology in 24 single-gene causes of SRNS in a cohort of 72 families, who presented with SRNS before the age of 25 years. Within an 18-month interval, we obtained DNA samples, pedigree information, and clinical information from 77 consecutive children with SRNS from 72 different families seen at Boston Children's Hospital (BCH). Mutation analysis was completed by combining high-throughput multiplex PCR with next-generation sequencing. We analyzed the sequences of 18 recessive and 6 dominant genes of SRNS in all 72 families for disease-causing variants. We identified the disease-causing mutation in 8 out of 72 (11.1%) families. Mutations were detected in the six genes: NPHS1 (2 out of 72), WT1 (2 out of 72), NPHS2, MYO1E, TRPC6, and INF2. Median age at onset was 4.1 years in patients without a mutation (range 0.5-18.8), and 3.2 years in those in whom the causative mutation was detected (range 0.1-14.3). Mutations in dominant genes presented with a median onset of 4.5 years (range 3.2-14.3). Mutations in recessive genes presented with a median onset of 0.5 years (range 0.1-3.2). Our molecular genetic diagnostic study identified underlying monogenic causes of steroid-resistant nephrotic syndrome in ~11% of patients with SRNS using a cost-effective technique. We delineated some of the therapeutic, diagnostic, and prognostic implications. Our study confirms that genetic testing is indicated in pediatric patients with SRNS.

Novel mutations responsible for α-thalassemia in Iranian families.

PubMed

Bayat, Nooshin; Farashi, Samaneh; Hafezi-Nejad, Nima; Faramarzi, Negin; Ashki, Mehri; Vakili, Shadi; Imanian, Hashem; Khosravi, Mohsen; Azar-Keivan, Azita; Najmabadi, Hossein

2013-01-01

α-Thalassemia (α-thal) is usually caused by deletions on the α-globin gene cluster and the role of point mutations is less well investigated. In the present study, a total of 1048 individuals with hypochromic microcytic anemia, who did not present the most common α-thal deletions, were referred for α-globin gene DNA sequencing. The nucleotide changes were studied and a total of five new mutations was identified, of which three were located on the α2 gene [codon7 (Lys→Stop), codon 34 (Leu→Pro) and codon 83 (Leu→Arg)] and two on the α1 gene [IVS-I-116 (A>G) and codon 44 (+C)]. These novel mutations not only explain new findings by molecular analysis of the α-globin gene but also have clinical importance due to their changes in α-globin production in means of decreased hemoglobin (Hb) related values. Moreover, considerations of its role in combination with other mutations, and the possibility of causing Hb H (β4) are yet to be studied.

Live-cell imaging of retrograde transport initiation in primary neurons.

PubMed

Nirschl, Jeffrey J; Holzbaur, Erika L F

2016-01-01

Axonal transport is an essential function in neurons, as mutations in either motor proteins or their adaptors cause neurodegeneration. While some mutations cause a complete block in axonal transport, other mutations affect transport more subtly. This is especially true of mutations identified in human patients, many of which impair but do not block motor function in the cell. Dissecting the pathogenic mechanisms of these more subtle mutations requires assays that can tease apart the distinct phases of axonal transport, including transport initiation, sustained/regulated motility, and cargo-specific sorting or delivery. Here, we describe a live-cell photobleaching assay to assess retrograde flux from the distal axon tip, a measure for distal transport initiation. We have previously used this method to show that the CAP-Gly domain of DCTN1 is required for efficient retrograde transport initiation in the distal axon, but it is not required to maintain retrograde flux along the mid-axon (Moughamian & Holzbaur, 2012). This approach has allowed us to examine the effects of disease-causing mutations in the axonal transport machinery, and in combination with other assays, will be useful in determining the mechanisms and regulation of axonal transport in normal and diseased conditions. Copyright © 2016 Elsevier Inc. All rights reserved.

Mutational signatures of DNA mismatch repair deficiency in C. elegans and human cancers.

PubMed

Meier, Bettina; Volkova, Nadezda V; Hong, Ye; Schofield, Pieta; Campbell, Peter J; Gerstung, Moritz; Gartner, Anton

2018-05-01

Throughout their lifetime, cells are subject to extrinsic and intrinsic mutational processes leaving behind characteristic signatures in the genome. DNA mismatch repair (MMR) deficiency leads to hypermutation and is found in different cancer types. Although it is possible to associate mutational signatures extracted from human cancers with possible mutational processes, the exact causation is often unknown. Here, we use C. elegans genome sequencing of pms-2 and mlh-1 knockouts to reveal the mutational patterns linked to C. elegans MMR deficiency and their dependency on endogenous replication errors and errors caused by deletion of the polymerase ε subunit pole-4 Signature extraction from 215 human colorectal and 289 gastric adenocarcinomas revealed three MMR-associated signatures, one of which closely resembles the C. elegans MMR spectrum and strongly discriminates microsatellite stable and unstable tumors (AUC = 98%). A characteristic difference between human and C. elegans MMR deficiency is the lack of elevated levels of N C G > NTG mutations in C. elegans, likely caused by the absence of cytosine (CpG) methylation in worms . The other two human MMR signatures may reflect the interaction between MMR deficiency and other mutagenic processes, but their exact cause remains unknown. In summary, combining information from genetically defined models and cancer samples allows for better aligning mutational signatures to causal mutagenic processes. © 2018 Meier et al.; Published by Cold Spring Harbor Laboratory Press.

Cortical atrophy and hypofibrinogenemia due to FGG and TBCD mutations in a single family: a case report.

PubMed

Stephen, Joshi; Nampoothiri, Sheela; Vinayan, K P; Yesodharan, Dhanya; Remesh, Preetha; Gahl, William A; Malicdan, May Christine V

2018-05-16

Blended phenotypes or co-occurrence of independent phenotypically distinct conditions are extremely rare and are due to coincidence of multiple pathogenic mutations, especially due to consanguinity. Hereditary fibrinogen deficiencies result from mutations in the genes FGA, FGB, and FGG, encoding the three different polypeptide chains that comprise fibrinogen. Neurodevelopmental abnormalities have not been associated with fibrinogen deficiencies. In this study, we report an unusual patient with a combination of two independently inherited genetic conditions; fibrinogen deficiency and early onset cortical atrophy. The study describes a male child from consanguineous family presented with hypofibrinogenemia, diffuse cortical atrophy, microcephaly, hypertonia and axonal motor neuropathy. Through a combination of homozygosity mapping and exome sequencing, we identified bi-allelic pathogenic mutations in two genes: a homozygous novel truncating mutation in FGG (c.554del; p.Lys185Argfs*14) and a homozygous missense mutation in TBCD (c.1423G > A;p.Ala475Thr). Loss of function mutations in FGG have been associated with fibrinogen deficiency, while the c.1423G > A mutation in TBCD causes a novel syndrome of neurodegeneration and early onset encephalopathy. Our study highlights the importance of homozygosity mapping and exome sequencing in molecular prenatal diagnosis, especially when multiple gene mutations are responsible for the phenotype.

Neonatal Diabetes Caused by Mutations in Sulfonylurea Receptor 1: Interplay between Expression and Mg-Nucleotide Gating Defects of ATP-Sensitive Potassium Channels

PubMed Central

Zhou, Qing; Garin, Intza; Castaño, Luis; Argente, Jesús; Muñoz-Calvo, Ma. Teresa; Perez de Nanclares, Guiomar; Shyng, Show-Ling

2010-01-01

Context: ATP-sensitive potassium (KATP) channels regulate insulin secretion by coupling glucose metabolism to β-cell membrane potential. Gain-of-function mutations in the sulfonylurea receptor 1 (SUR1) or Kir6.2 channel subunit underlie neonatal diabetes. Objective: The objective of the study was to determine the mechanisms by which two SUR1 mutations, E208K and V324M, associated with transient neonatal diabetes affect KATP channel function. Design: E208K or V324M mutant SUR1 was coexpressed with Kir6.2 in COS cells, and expression and gating properties of the resulting channels were assessed biochemically and electrophysiologically. Results: Both E208K and V324M augment channel response to MgADP stimulation without altering sensitivity to ATP4− or sulfonylureas. Surprisingly, whereas E208K causes only a small increase in MgADP response consistent with the mild transient diabetes phenotype, V324M causes a severe activating gating defect. Unlike E208K, V324M also impairs channel expression at the cell surface, which is expected to dampen its functional impact on β-cells. When either mutation was combined with a mutation in the second nucleotide binding domain of SUR1 previously shown to abolish Mg-nucleotide response, the activating effect of E208K and V324M was also abolished. Moreover, combination of E208K and V324M results in channels with Mg-nucleotide sensitivity greater than that seen in individual mutations alone. Conclusion: The results demonstrate that E208K and V324M, located in distinct domains of SUR1, enhance transduction of Mg-nucleotide stimulation from the SUR1 nucleotide binding folds to Kir6.2. Furthermore, they suggest that diabetes severity is determined by interplay between effects of a mutation on channel expression and channel gating. PMID:20810569

Synergistic effect of factor VII gene polymorphisms causing mild factor VII deficiency in a case of severe factor X deficiency.

PubMed

Deshpande, Rutuja; Ghosh, Kanjaksha; Shetty, Shrimati

2017-01-01

Congenital combined deficiency of coagulation factors VII and X are mainly attributed to large deletions involving both the genes in chromosome 13 or occasionally due to the coincidental occurrence of independently occurring mutations. We report the molecular basis of congenital combined deficiency of factors VII and X in a 6-year-old female child. Direct DNA sequencing of both factor VII (F7) and factor X (F10) genes showed a novel homozygous missense mutation p.Cys90Tyr (c.307G>A) in exon 4 of F10. No mutations were detected in F7; however, the patient was homozygous for three polymorphic alleles known to be associated with reduced factor VII levels. The present case illustrates the synergistic effect of multiple polymorphisms resulting in phenotypic factor VII deficiency in the absence of a pathogenic mutation.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Greenberg, C.R.; Taylor, C.D.; Haworth, J.C.

The authors have discovered a single homoallelic nucleotide substitution as the putative cause of the perinatal (lethal) form of hypophosphatasia in Canadian Mennonites. Previous linkage and haplotype analysis in this population suggested that a single mutational event was responsible for this autosomal recessive form of hypophosphatasia. The mutation is a guanosine-to-adenosine substitution at nucleotide position 1177 in exon 10 of the tissue nonspecific (liver/bone/kidney) alkaline phosphatase gene. This Gly[sup 317] [yields] Asp mutation segregates exclusively with the heterozygote phenotype previously assigned by biochemical testing (maximum combined lod score of 18.24 at [theta] = 0.00). This putative disease-causing mutation has notmore » been described in controls nor in other non-Mennonite probands with both lethal and nonlethal forms of hypophosphatasia studied to date. This Gly[sup 317] [yields] Asp mutation changes a polar glycine to an acidic aspartate at amino acid position 317 within the highly conserved active site region of the 507-amino-acid polypeptide. Carrier screening for this lethal mutation in a high-risk population is now feasible. 15 refs., 2 figs.« less

The genetic basis of female reproductive disorders: Etiology and clinical testing ☆

PubMed Central

Layman, Lawrence C.

2013-01-01

With the advent of improved molecular biology techniques, the genetic basis of an increasing number of reproductive disorders has been elucidated. Mutations in at least 20 genes cause hypogonadotropic hypogonadism including Kallmann syndrome in about 35–40% of patients. The two most commonly involved genes are FGFR1 and CHD7. When combined pituitary hormone deficiency includes hypogonadotropic hypogonadism as a feature, PROP1 mutations are the most common of the six genes involved. For hypergonadotropic hypogonadism, mutations in 14 genes cause gonadal failure in 15% of affected females, most commonly in FMR1. In eugonadal disorders, activating FSHR mutations have been identified for spontaneous ovarian hyperstimulation syndrome; and WNT4 mutations have been described in mullerian aplasia. For other eugonadal disorders, such as endometriosis, polycystic ovary syndrome, and leiomyomata, specific germline gene mutations have not been identified, but some chromosomal regions are associated with the corresponding phenotype. Practical genetic testing is possible to perform in both hypogonadotropic and hypergonadotropic hypogonadism and spontaneous ovarian hyperstimulation syndrome. However, clinical testing for endometriosis, polycystic ovary syndrome, and leiomyomata is not currently practical for the clinician. PMID:23499866

Absence of mutations in HCRT, HCRTR1 and HCRTR2 in patients with ROHHAD.

PubMed

Barclay, Sarah F; Rand, Casey M; Gray, Paul A; Gibson, William T; Wilson, Richard J A; Berry-Kravis, Elizabeth M; Ize-Ludlow, Diego; Bech-Hansen, N Torben; Weese-Mayer, Debra E

2016-01-15

Rapid-onset obesity with hypothalamic dysfunction, hypoventilation, and autonomic dysregulation (ROHHAD) is a rare pediatric disease of unknown cause. Here, in response to a recent case report describing a ROHHAD patient who suffered from secondary narcolepsy confirmed by an absence of hypocretin-1 in the cerebrospinal fluid, we consider whether the ROHHAD phenotype is owing to one or more mutations in genes specific to hypocretin protein signalling. DNA samples from 16 ROHHAD patients were analyzed using a combination of next-generation and Sanger sequencing to identify exonic sequence variations in three genes: HCRT, HCRTR1, and HCRTR2. No rare or novel mutations were identified in the exons of HCRT, HCRTR1, or HCRTR2 genes in a set of 16 ROHHAD patients. ROHHAD is highly unlikely to be caused by mutations in the exons of the genes for hypocretin and its two receptors. Copyright © 2015 Elsevier B.V. All rights reserved.

Computational and Experimental Approaches to Reveal the Effects of Single Nucleotide Polymorphisms with Respect to Disease Diagnostics

PubMed Central

Kucukkal, Tugba G.; Yang, Ye; Chapman, Susan C.; Cao, Weiguo; Alexov, Emil

2014-01-01

DNA mutations are the cause of many human diseases and they are the reason for natural differences among individuals by affecting the structure, function, interactions, and other properties of DNA and expressed proteins. The ability to predict whether a given mutation is disease-causing or harmless is of great importance for the early detection of patients with a high risk of developing a particular disease and would pave the way for personalized medicine and diagnostics. Here we review existing methods and techniques to study and predict the effects of DNA mutations from three different perspectives: in silico, in vitro and in vivo. It is emphasized that the problem is complicated and successful detection of a pathogenic mutation frequently requires a combination of several methods and a knowledge of the biological phenomena associated with the corresponding macromolecules. PMID:24886813

Gain-of-function mutations in interleukin-7 receptor-α (IL7R) in childhood acute lymphoblastic leukemias

PubMed Central

Shochat, Chen; Tal, Noa; Bandapalli, Obul R.; Palmi, Chiara; Ganmore, Ithamar; te Kronnie, Geertruy; Cario, Gunnar; Cazzaniga, Giovanni; Kulozik, Andreas E.; Stanulla, Martin; Schrappe, Martin; Biondi, Andrea; Basso, Giuseppe; Bercovich, Dani; Muckenthaler, Martina U.

2011-01-01

Interleukin-7 receptor α (IL7R) is required for normal lymphoid development. Loss-of-function mutations in this gene cause autosomal recessive severe combined immune deficiency. Here, we describe somatic gain-of-function mutations in IL7R in pediatric B and T acute lymphoblastic leukemias. The mutations cause either a serine-to-cysteine substitution at amino acid 185 in the extracellular domain (4 patients) or in-frame insertions and deletions in the transmembrane domain (35 patients). In B cell precursor leukemias, the mutations were associated with the aberrant expression of cytokine receptor-like factor 2 (CRLF2), and the mutant IL-7R proteins formed a functional receptor with CRLF2 for thymic stromal lymphopoietin (TSLP). Biochemical and functional assays reveal that these IL7R mutations are activating mutations conferring cytokine-independent growth of progenitor lymphoid cells. A cysteine, included in all but three of the mutated IL-7R alleles, is essential for the constitutive activation of the receptor. This is the first demonstration of gain-of-function mutations of IL7R. Our current and recent observations of mutations in IL7R and CRLF2, respectively suggest that the addition of cysteine to the juxtamembranous domains is a general mechanism for mutational activation of type I cytokine receptors in leukemia. PMID:21536738

Combining Structural Modeling with Ensemble Machine Learning to Accurately Predict Protein Fold Stability and Binding Affinity Effects upon Mutation

PubMed Central

Garcia Lopez, Sebastian; Kim, Philip M.

2014-01-01

Advances in sequencing have led to a rapid accumulation of mutations, some of which are associated with diseases. However, to draw mechanistic conclusions, a biochemical understanding of these mutations is necessary. For coding mutations, accurate prediction of significant changes in either the stability of proteins or their affinity to their binding partners is required. Traditional methods have used semi-empirical force fields, while newer methods employ machine learning of sequence and structural features. Here, we show how combining both of these approaches leads to a marked boost in accuracy. We introduce ELASPIC, a novel ensemble machine learning approach that is able to predict stability effects upon mutation in both, domain cores and domain-domain interfaces. We combine semi-empirical energy terms, sequence conservation, and a wide variety of molecular details with a Stochastic Gradient Boosting of Decision Trees (SGB-DT) algorithm. The accuracy of our predictions surpasses existing methods by a considerable margin, achieving correlation coefficients of 0.77 for stability, and 0.75 for affinity predictions. Notably, we integrated homology modeling to enable proteome-wide prediction and show that accurate prediction on modeled structures is possible. Lastly, ELASPIC showed significant differences between various types of disease-associated mutations, as well as between disease and common neutral mutations. Unlike pure sequence-based prediction methods that try to predict phenotypic effects of mutations, our predictions unravel the molecular details governing the protein instability, and help us better understand the molecular causes of diseases. PMID:25243403

Prediction of Disease Causing Non-Synonymous SNPs by the Artificial Neural Network Predictor NetDiseaseSNP

PubMed Central

Johansen, Morten Bo; Izarzugaza, Jose M. G.; Brunak, Søren; Petersen, Thomas Nordahl; Gupta, Ramneek

2013-01-01

We have developed a sequence conservation-based artificial neural network predictor called NetDiseaseSNP which classifies nsSNPs as disease-causing or neutral. Our method uses the excellent alignment generation algorithm of SIFT to identify related sequences and a combination of 31 features assessing sequence conservation and the predicted surface accessibility to produce a single score which can be used to rank nsSNPs based on their potential to cause disease. NetDiseaseSNP classifies successfully disease-causing and neutral mutations. In addition, we show that NetDiseaseSNP discriminates cancer driver and passenger mutations satisfactorily. Our method outperforms other state-of-the-art methods on several disease/neutral datasets as well as on cancer driver/passenger mutation datasets and can thus be used to pinpoint and prioritize plausible disease candidates among nsSNPs for further investigation. NetDiseaseSNP is publicly available as an online tool as well as a web service: http://www.cbs.dtu.dk/services/NetDiseaseSNP PMID:23935863

Diversity of ARSACS mutations in French-Canadians.

PubMed

Thiffault, I; Dicaire, M J; Tetreault, M; Huang, K N; Demers-Lamarche, J; Bernard, G; Duquette, A; Larivière, R; Gehring, K; Montpetit, A; McPherson, P S; Richter, A; Montermini, L; Mercier, J; Mitchell, G A; Dupré, N; Prévost, C; Bouchard, J P; Mathieu, J; Brais, B

2013-01-01

The growing number of spastic ataxia of Charlevoix-Saguenay (SACS) gene mutations reported worldwide has broadened the clinical phenotype of autosomal recessive spastic ataxia of Charlevoix-Saguenay (ARSACS). The identification of Quebec ARSACS cases without two known SACS mutation led to the development of a multi-modal genomic strategy to uncover mutations in this large gene and explore phenotype variability. Search for SACS mutations by combining various methods on 20 cases with a classical French-Canadian ARSACS phenotype without two mutations and a group of 104 sporadic or recessive spastic ataxia cases of unknown cause. Western blot on lymphoblast protein from cases with different genotypes was probed to establish if they still expressed sacsin. A total of 12 mutations, including 7 novels, were uncovered in Quebec ARSACS cases. The screening of 104 spastic ataxia cases of unknown cause for 98 SACS mutations did not uncover carriers of two mutations. Compounds heterozygotes for one missense SACS mutation were found to minimally express sacsin. The large number of SACS mutations present even in Quebec suggests that the size of the gene alone may explain the great genotypic diversity. This study does not support an expanding ARSACS phenotype in the French-Canadian population. Most mutations lead to loss of function, though phenotypic variability in other populations may reflect partial loss of function with preservation of some sacsin expression. Our results also highlight the challenge of SACS mutation screening and the necessity to develop new generation sequencing methods to ensure low cost complete gene sequencing.

Spastic Paraplegia Type 7 Is Associated with Multiple Mitochondrial DNA Deletions

PubMed Central

Wedding, Iselin Marie; Koht, Jeanette; Tran, Gia Tuong; Misceo, Doriana; Selmer, Kaja Kristine; Holmgren, Asbjørn; Frengen, Eirik; Bindoff, Laurence; Tallaksen, Chantal M. E.; Tzoulis, Charalampos

2014-01-01

Spastic paraplegia 7 is an autosomal recessive disorder caused by mutations in the gene encoding paraplegin, a protein located at the inner mitochondrial membrane and involved in the processing of other mitochondrial proteins. The mechanism whereby paraplegin mutations cause disease is unknown. We studied two female and two male adult patients from two Norwegian families with a combination of progressive external ophthalmoplegia and spastic paraplegia. Sequencing of SPG7 revealed a novel missense mutation, c.2102A>C, p.H 701P, which was homozygous in one family and compound heterozygous in trans with a known pathogenic mutation c.1454_1462del in the other. Muscle was examined from an additional, unrelated adult female patient with a similar phenotype caused by a homozygous c.1047insC mutation in SPG7. Immunohistochemical studies in skeletal muscle showed mosaic deficiency predominantly affecting respiratory complex I, but also complexes III and IV. Molecular studies in single, microdissected fibres showed multiple mitochondrial DNA deletions segregating at high levels (38–97%) in respiratory deficient fibres. Our findings demonstrate for the first time that paraplegin mutations cause accumulation of mitochondrial DNA damage and multiple respiratory chain deficiencies. While paraplegin is not known to be directly associated with the mitochondrial nucleoid, it is known to process other mitochondrial proteins and it is possible therefore that paraplegin mutations lead to mitochondrial DNA deletions by impairing proteins involved in the homeostasis of the mitochondrial genome. These studies increase our understanding of the molecular pathogenesis of SPG7 mutations and suggest that SPG7 testing should be included in the diagnostic workup of autosomal recessive, progressive external ophthalmoplegia, especially if spasticity is present. PMID:24466038

Combination therapy of apatinib with icotinib for primary acquired icotinib resistance in patients with advanced pulmonary adenocarcinoma with EGFR mutation.

PubMed

Xia, Pinghui; Cao, Jinlin; Lv, Xiayi; Wang, Luming; Lv, Wang; Hu, Jian

2018-05-01

Multi-targeted agents represent the next generation of targeted therapies for solid tumors, and patients with acquired resistance to EGFR-tyrosine kinase inhibitors (TKIs) may also benefit from their combination with TKI therapy. Third-generation targeted drugs, such as osimertinib, are very expensive, thus a more economical solution is required. The aim of this study was to explore the use of apatinib combined with icotinib therapy for primary acquired resistance to icotinib in three patients with advanced pulmonary adenocarcinoma with EGFR mutations. We achieved favorable oncologic outcomes in all three patients, with progression-free survival of four to six months. Unfortunately, the patients ultimately had to cease combination therapy because of intolerable adverse effects of hand and foot syndrome and oral ulcers. Combination therapy of apatinib with icotinib for primary acquired resistance to icotinib may be an option for patients with advanced pulmonary adenocarcinoma with EGFR mutations, but physicians must also be aware of the side effects caused by such therapy. © 2018 The Authors. Thoracic Cancer published by China Lung Oncology Group and John Wiley & Sons Australia, Ltd.

[Cetuximab in combination with icotinib overcomes the acquired resistance caused by EGFR T790M mutation in non-small cell lung cancer].

PubMed

Wang, Meng; Zhang, Lianmin; Zhao, Xiaoliang; Liu, Jun; Chen, Yulong; Wang, Changli

2014-09-01

The aim of this study was to investigate the effects of combination of icotinib and cetuximab on the acquired drug resistance caused by T790M mutation of EGFR in NSCLC, and provide experimental evidence for rational treatment of NSCLC. The effects of these two agents on cell proliferation, apoptosis, and EGFR-dependent signaling were evaluated using 3-(4, 5-dimethylthiazol-2-yl)- 5-diphenyltetrazolium bromide (MTT) assay, annexin V staining, and Western blotting. The expression of molecular markers of tumor proliferation PCNA and Ki-67 protein was further examined by immunohistochemistry, and the expression of EGFR-signaling-related proteins in tissue sections taken from H1975 tumor xenografts was assessed by Western blot assay. Sensitivity to EGFR inhibitors was detected in human H1975 tumor xenograft in nude mice. The in vitro experiment showed that the proliferative ability of H1975 cells was inhibited in a dose-dependent manner, along with the increasing doses of cetuximab and icotinib, and the combination of cetuximab with icotinib resulted in a more pronounced growth inhibition of the H1975 cells. The apoptosis rate of H1975 cells after treatment with 0.5 µmol/L icotinib and 1 µg/ml cetuximab was (22.03 ± 2.41)% and that after treatment with 5 µmol/L icotinib and 10 µg/ml cetuximab was (42.75 ± 2.49)%, both were significantly higher than that after treatment with the same dose of icotinib or cetuximab alone (P < 0.05). The nude mouse experiment showed that the transplanted tumor was growing to (614.5 ± 10.8) mm(3) in the blank control group and to (611.2 ± 8.7) mm(3) at 28 days after icotinib treatment, but (30.8 ± 2.0) mm(3) in the cetuximab treatment group and 0 mm(3) in the cetuximab combined with icotinib group. There was a significantly decreased expression of Ki-67 and PCNA proteins and down-regulation of phosphorylation of EGFR signaling-related proteins in the cetuximab combined with icotinib group. The combination of icotinib with cetuximab can exert synergistic inhibitory effect on the acquired drug resistance caused by T790M mutation of EGFR in NSCLC H1975 cells, interrupts the EGFR-downstream signaling pathway, and enhances the anticancer activity of chemotherapeutic drugs. Our results provide further experimental evidence for the clinical studies of combination of icotinib with cetuximab in the treatment of NSCLC patients associated with secondary drug resistance caused by T790M mutation of EGFR.

A new nonsense mutation in the NF1 gene with neurofibromatosis-Noonan syndrome phenotype.

PubMed

Yimenicioğlu, Sevgi; Yakut, Ayten; Karaer, Kadri; Zenker, Martin; Ekici, Arzu; Carman, Kürşat Bora

2012-12-01

Neurofibromatosis-Noonan syndrome is a rare autosomal dominant disorder which combines neurofibromatosis type 1 (NF1) features with Noonan syndrome. NF1 gene mutations are reported in the majority of these patients. Sequence analysis of the established genes for Noonan syndrome revealed no mutation; a heterozygous NF1 point mutation c.7549C>T in exon 51, creating a premature stop codon (p.R2517X), had been demonstrated. Neurofibromatosis-Noonan syndrome recently has been considered a subtype of NF1 and caused by different NF1 mutations. We report the case of a 14-year-old boy with neurofibromatosis type 1 with Noonan-like features, who complained of headache with triventricular hydrocephaly and a heterozygous NF1 point mutation c.7549C>T in exon 51.

Phenotypic patterns of desminopathy associated with three novel mutations in the desmin gene

PubMed Central

Olivé, Montse; Armstrong, Judith; Miralles, Francesc; Pou, Adolf; Fardeau, Michel; Gonzalez, Laura; Martínez, Francesca; Fischer, Dirk; Matos, Juan Antonio Martínez; Shatunov, Alexey; Goldfarb, Lev; Ferrer, Isidre

2016-01-01

Desminopathy represents a subgroup of myofibrillar myopathies caused by mutations in the desmin gene. Three novel disease-associated mutations in the desmin gene were identified in unrelated Spanish families affected by cardioskeletal myopathy. A selective pattern of muscle involvement, which differed from that observed in myofibrillar myopathy resulting from mutations in the myotilin gene, was observed in each of the three families with novel mutations and each of three desminopathy patients with known desmin mutations. Prominent joint retractions at the ankles and characteristic nasal speech were observed early in the course of illness. These findings suggest that muscle imaging in combination with routine clinical and pathological examination may be helpful in distinguishing desminopathy from other forms of myofibrillar myopathy and ordering appropriate molecular investigations. PMID:17418574

A Novel Mutation in ERCC8 Gene Causing Cockayne Syndrome

PubMed Central

Taghdiri, Maryam; Dastsooz, Hassan; Fardaei, Majid; Mohammadi, Sanaz; Farazi Fard, Mohammad Ali; Faghihi, Mohammad Ali

2017-01-01

Cockayne syndrome (CS) is a rare autosomal recessive multisystem disorder characterized by impaired neurological and sensory functions, cachectic dwarfism, microcephaly, and photosensitivity. This syndrome shows a variable age of onset and rate of progression, and its phenotypic spectrum include a wide range of severity. Due to the progressive nature of this disorder, diagnosis can be more important when additional signs and symptoms appear gradually and become steadily worse over time. Therefore, mutation analysis of genes involved in CS pathogenesis can be helpful to confirm the suspected clinical diagnosis. Here, we report a novel mutation in ERCC8 gene in a 16-year-old boy who suffers from poor weight gain, short stature, microcephaly, intellectual disability, and photosensitivity. The patient was born to consanguineous family with no previous documented disease in his parents. To identify disease-causing mutation in the patient, whole exome sequencing utilizing next-generation sequencing on an Illumina HiSeq 2000 platform was performed. Results revealed a novel homozygote mutation in ERCC8 gene (NM_000082: exon 11, c.1122G>C) in our patient. Another gene (ERCC6), which is also involved in CS did not have any disease-causing mutations in the proband. The new identified mutation was then confirmed by Sanger sequencing in the proband, his parents, and extended family members, confirming co-segregation with the disease. In addition, different bioinformatics programs which included MutationTaster, I-Mutant v2.0, NNSplice, Combined Annotation Dependent Depletion, The PhastCons, Genomic Evolutationary Rate Profiling conservation score, and T-Coffee Multiple Sequence Alignment predicted the pathogenicity of the mutation. Our study identified a rare novel mutation in ERCC8 gene and help to provide accurate genetic counseling and prenatal diagnosis to minimize new affected individuals in this family. PMID:28848724

A Novel Mutation in ERCC8 Gene Causing Cockayne Syndrome.

PubMed

Taghdiri, Maryam; Dastsooz, Hassan; Fardaei, Majid; Mohammadi, Sanaz; Farazi Fard, Mohammad Ali; Faghihi, Mohammad Ali

2017-01-01

Cockayne syndrome (CS) is a rare autosomal recessive multisystem disorder characterized by impaired neurological and sensory functions, cachectic dwarfism, microcephaly, and photosensitivity. This syndrome shows a variable age of onset and rate of progression, and its phenotypic spectrum include a wide range of severity. Due to the progressive nature of this disorder, diagnosis can be more important when additional signs and symptoms appear gradually and become steadily worse over time. Therefore, mutation analysis of genes involved in CS pathogenesis can be helpful to confirm the suspected clinical diagnosis. Here, we report a novel mutation in ERCC8 gene in a 16-year-old boy who suffers from poor weight gain, short stature, microcephaly, intellectual disability, and photosensitivity. The patient was born to consanguineous family with no previous documented disease in his parents. To identify disease-causing mutation in the patient, whole exome sequencing utilizing next-generation sequencing on an Illumina HiSeq 2000 platform was performed. Results revealed a novel homozygote mutation in ERCC8 gene (NM_000082: exon 11, c.1122G>C) in our patient. Another gene ( ERCC6 ), which is also involved in CS did not have any disease-causing mutations in the proband. The new identified mutation was then confirmed by Sanger sequencing in the proband, his parents, and extended family members, confirming co-segregation with the disease. In addition, different bioinformatics programs which included MutationTaster, I-Mutant v2.0, NNSplice, Combined Annotation Dependent Depletion, The PhastCons, Genomic Evolutationary Rate Profiling conservation score, and T-Coffee Multiple Sequence Alignment predicted the pathogenicity of the mutation. Our study identified a rare novel mutation in ERCC8 gene and help to provide accurate genetic counseling and prenatal diagnosis to minimize new affected individuals in this family.

Genetic Characterization of the SufJ Frameshift Suppressor in SALMONELLA TYPHIMURIUM

PubMed Central

Bossi, Lionello; Kohno, Tadahiko; Roth, John R.

1983-01-01

A new suppressor of +1 frameshift mutations has been isolated in Salmonella typhimurium. This suppressor, sufJ, maps at minute 89 on the Salmonella genetic map between the argH and rpo(rif) loci, closely linked to the gene for the ochre suppressor tyrU(supM). The suppressor mutation is dominant to its wild-type allele, consistent with the suppressor phenotype being caused by an altered tRNA species. The sufJ map position coincides with that of a threonine tRNA(ACC/U) gene; the suppressor has been shown to read the related fourbase codons ACCU, ACCC, ACCA.—The ability of sufJ to correct one particular mutation depends on the presence of a hisT mutation which causes a defect in tRNA modification. This requirement is allele specific, since other frameshift mutations can be corrected by sufJ regardless of the state of the hisT locus.—Strains carrying both a sufJ and a hisT mutation are acutely sensitive to growth inhibition by uracil; the inhibition is reversed by arginine. This behavior is characteristic of strains with mutations affecting the arginine-uracil biosynthetic enzyme carbamyl phosphate synthetase. The combination of two mutations affecting tRNA structure may reduce expression of the structural gene for this enzyme (pyrA). PMID:6188650

The TERT promoter SNP rs2853669 decreases E2F1 transcription factor binding and increases mortality and recurrence risks in liver cancer.

PubMed

Ko, Eunkyong; Seo, Hyun-Wook; Jung, Eun Sun; Kim, Baek-hui; Jung, Guhung

2016-01-05

A common single-nucleotide polymorphism in the telomerase reverse transcriptase (TERT) promoter, rs2853669 influences patient survival rates and the risk of developing cancer. Recently, several lines of evidence suggest that the rs2853669 suppresses TERT promoter mutation-mediated TERT expression levels and cancer mortality as well as recurrence rates. However, no reports are available on the impact of rs2853669 on TERT expression in hepatocellular carcinoma (HCC) and its association with patient survival. Here, we found that HCC-related overall and recurrence-free survival rates were not associated with TERT promoter mutation individually, but rs2853669 and the TERT promoter mutation in combination were associated with poor survival rates. TERT mRNA expression and telomere fluorescence levels were greater in patients with HCC who had both the combination. The combination caused TERT promoter methylation through regulating the binding of DNA methyltransferase 1 and histone deacetylase 1 to the TERT promoter in HCC cell lines. The TERT expression level was significantly higher in HCC tumor with a methylated promoter than in that with an unmethylated promoter. In conclusion, we demonstrate a substantial role for the rs2853669 in HCC with TERT promoter mutation, which suggests that the combination of the rs2853669 and the mutation indicate poor prognoses in liver cancer.

Rare hereditary cause of chronic pancreatitis in a young male: SPINK1 mutation

PubMed Central

Patel, Janaki; Madan, Arina; Gammon, Amanda; Sossenheimer, Michael; Samadder, Niloy Jewel

2017-01-01

Hereditary chronic pancreatitis associated with a mutation in the serine protease inhibitor, Kazal Type-1 (SPINK-1 gene) is extremely rare. The SPINK1 mutation results in trypsinogen activation which predisposes to chronic pancreatitis predominately when combined with CFTR gene mutations. It presents as either chronic or recurrent acute pancreatitis. Symptom control and management of complications is important. Active surveillance with cross-sectional imaging for pancreatic malignancy in individuals with hereditary pancreatitis is advocated due to individuals being high risk. We present an unusual case of a young male who initially presented with renal colic and was incidentally diagnosed with severe chronic pancreatitis on abdominal imaging, with genetic testing confirming a homozygous SPINK1 mutation. PMID:29515728

Rare hereditary cause of chronic pancreatitis in a young male: SPINK1 mutation.

PubMed

Patel, Janaki; Madan, Arina; Gammon, Amanda; Sossenheimer, Michael; Samadder, Niloy Jewel

2017-01-01

Hereditary chronic pancreatitis associated with a mutation in the serine protease inhibitor, Kazal Type-1 (SPINK-1 gene) is extremely rare. The SPINK1 mutation results in trypsinogen activation which predisposes to chronic pancreatitis predominately when combined with CFTR gene mutations. It presents as either chronic or recurrent acute pancreatitis. Symptom control and management of complications is important. Active surveillance with cross-sectional imaging for pancreatic malignancy in individuals with hereditary pancreatitis is advocated due to individuals being high risk. We present an unusual case of a young male who initially presented with renal colic and was incidentally diagnosed with severe chronic pancreatitis on abdominal imaging, with genetic testing confirming a homozygous SPINK1 mutation.

Some gating potentiators, including VX-770, diminish ΔF508-CFTR functional expression

PubMed Central

Veit, Guido; Avramescu, Radu G.; Perdomo, Doranda; Phuan, Puay-Wah; Bagdany, Miklos; Apaja, Pirjo M.; Borot, Florence; Szollosi, Daniel; Wu, Yu-Sheng; Finkbeiner, Walter E.; Hegedus, Tamas; Verkman, Alan S.; Lukacs, Gergely L.

2015-01-01

Cystic fibrosis (CF) is caused by mutations in the CF transmembrane regulator (CFTR) that result in reduced anion conductance at the apical membrane of secretory epithelia. Treatment of CF patients carrying the G551D gating mutation with the potentiator VX-770 (ivacaftor) largely restores channel activity and has shown substantial clinical benefit. However, most CF patients carry the ΔF508 mutation, which impairs CFTR folding, processing, function, and stability. Studies in homozygous ΔF508 CF patients indicated little clinical benefit of monotherapy with the investigational corrector VX-809 (lumacaftor) or VX-770, whereas combination clinical trials show limited but significant improvements in lung function. We show that VX-770, as well as most other potentiators, reduces the correction efficacy of VX-809 and another investigational corrector, VX-661. To mimic the administration of VX-770 alone or in combination with VX-809, we examined its long-term effect in immortalized and primary human respiratory epithelia. VX-770 diminished the folding efficiency and the metabolic stability of ΔF508-CFTR at the endoplasmic reticulum (ER) and post-ER compartments, respectively, causing reduced cell surface ΔF508-CFTR density and function. VX-770–induced destabilization of ΔF508-CFTR was influenced by second-site suppressor mutations of the folding defect and was prevented by stabilization of the nucleotide-binding domain 1 (NBD1)–NBD2 interface. The reduced correction efficiency of ΔF508-CFTR, as well as of two other processing mutations in the presence of VX-770, suggests the need for further optimization of potentiators to maximize the clinical benefit of corrector-potentiator combination therapy in CF. PMID:25101887

Some gating potentiators, including VX-770, diminish ΔF508-CFTR functional expression.

PubMed

Veit, Guido; Avramescu, Radu G; Perdomo, Doranda; Phuan, Puay-Wah; Bagdany, Miklos; Apaja, Pirjo M; Borot, Florence; Szollosi, Daniel; Wu, Yu-Sheng; Finkbeiner, Walter E; Hegedus, Tamas; Verkman, Alan S; Lukacs, Gergely L

2014-07-23

Cystic fibrosis (CF) is caused by mutations in the CF transmembrane regulator (CFTR) that result in reduced anion conductance at the apical membrane of secretory epithelia. Treatment of CF patients carrying the G551D gating mutation with the potentiator VX-770 (ivacaftor) largely restores channel activity and has shown substantial clinical benefit. However, most CF patients carry the ΔF508 mutation, which impairs CFTR folding, processing, function, and stability. Studies in homozygous ΔF508 CF patients indicated little clinical benefit of monotherapy with the investigational corrector VX-809 (lumacaftor) or VX-770, whereas combination clinical trials show limited but significant improvements in lung function. We show that VX-770, as well as most other potentiators, reduces the correction efficacy of VX-809 and another investigational corrector, VX-661. To mimic the administration of VX-770 alone or in combination with VX-809, we examined its long-term effect in immortalized and primary human respiratory epithelia. VX-770 diminished the folding efficiency and the metabolic stability of ΔF508-CFTR at the endoplasmic reticulum (ER) and post-ER compartments, respectively, causing reduced cell surface ΔF508-CFTR density and function. VX-770-induced destabilization of ΔF508-CFTR was influenced by second-site suppressor mutations of the folding defect and was prevented by stabilization of the nucleotide-binding domain 1 (NBD1)-NBD2 interface. The reduced correction efficiency of ΔF508-CFTR, as well as of two other processing mutations in the presence of VX-770, suggests the need for further optimization of potentiators to maximize the clinical benefit of corrector-potentiator combination therapy in CF. Copyright © 2014, American Association for the Advancement of Science.

MtDNA mutations are a common cause of severe disease phenotypes in children with Leigh syndrome.

PubMed

Naess, Karin; Freyer, Christoph; Bruhn, Helene; Wibom, Rolf; Malm, Gunilla; Nennesmo, Inger; von Döbeln, Ulrika; Larsson, Nils-Göran

2009-05-01

Leigh syndrome is a common clinical manifestation in children with mitochondrial disease and other types of inborn errors of metabolism. We characterised clinical symptoms, prognosis, respiratory chain function and performed extensive genetic analysis of 25 Swedish children suffering from Leigh syndrome with the aim to obtain insights into the molecular pathophysiology and to provide a rationale for genetic counselling. We reviewed the clinical history of all patients and used muscle biopsies in order to perform molecular, biochemical and genetic investigations, including sequencing the entire mitochondrial DNA (mtDNA), the mitochondrial DNA polymerase (POLGA) gene and the surfeit locus protein 1 (SURF1) gene. Respiratory chain enzyme activity measurements identified five patients with isolated complex I deficiency and five with combined enzyme deficiencies. No patient presented with isolated complex IV deficiency. Seven patients had a decreased ATP production rate. Extensive sequence analysis identified eight patients with pathogenic mtDNA mutations and one patient with mutations in POLGA. Mutations of mtDNA are a common cause of LS and mtDNA analysis should always be included in the diagnosis of LS patients, whereas SURF1 mutations are not a common cause of LS in Sweden. Unexpectedly, age of onset, clinical symptoms and prognosis did not reveal any clear differences in LS patients with mtDNA or nuclear DNA mutations.

Clinical spectrum of 4H leukodystrophy caused by POLR3A and POLR3B mutations

PubMed Central

Vanderver, Adeline; van Spaendonk, Rosalina M.L.; Schiffmann, Raphael; Brais, Bernard; Bugiani, Marianna; Sistermans, Erik; Catsman-Berrevoets, Coriene; Kros, Johan M.; Pinto, Pedro Soares; Pohl, Daniela; Tirupathi, Sandya; Strømme, Petter; de Grauw, Ton; Fribourg, Sébastien; Demos, Michelle; Pizzino, Amy; Naidu, Sakkubai; Guerrero, Kether; van der Knaap, Marjo S.; Bernard, Geneviève

2014-01-01

Objective: To study the clinical and radiologic spectrum and genotype–phenotype correlation of 4H (hypomyelination, hypodontia, hypogonadotropic hypogonadism) leukodystrophy caused by mutations in POLR3A or POLR3B. Methods: We performed a multinational cross-sectional observational study of the clinical, radiologic, and molecular characteristics of 105 mutation-proven cases. Results: The majority of patients presented before 6 years with gross motor delay or regression. Ten percent had an onset beyond 10 years. The disease course was milder in patients with POLR3B than in patients with POLR3A mutations. Other than the typical neurologic, dental, and endocrine features, myopia was seen in almost all and short stature in 50%. Dental and hormonal findings were not invariably present. Mutations in POLR3A and POLR3B were distributed throughout the genes. Except for French Canadian patients, patients from European backgrounds were more likely to have POLR3B mutations than other populations. Most patients carried the common c.1568T>A POLR3B mutation on one allele, homozygosity for which causes a mild phenotype. Systematic MRI review revealed that the combination of hypomyelination with relative T2 hypointensity of the ventrolateral thalamus, optic radiation, globus pallidus, and dentate nucleus, cerebellar atrophy, and thinning of the corpus callosum suggests the diagnosis. Conclusions: 4H is a well-recognizable clinical entity if all features are present. Mutations in POLR3A are associated with a more severe clinical course. MRI characteristics are helpful in addressing the diagnosis, especially if patients lack the cardinal non-neurologic features. PMID:25339210

DMD mutation spectrum analysis in 613 Chinese patients with dystrophinopathy.

PubMed

Guo, Ruolan; Zhu, Guosheng; Zhu, Huimin; Ma, Ruiyu; Peng, Ying; Liang, Desheng; Wu, Lingqian

2015-08-01

Dystrophinopathy is a group of inherited diseases caused by mutations in the DMD gene. Within the dystrophinopathy spectrum, Duchenne and Becker muscular dystrophies are common X-linked recessive disorders that mainly feature striated muscle necrosis. We combined multiplex ligation-dependent probe amplification with Sanger sequencing to detect large deletions/duplications and point mutations in the DMD gene in 613 Chinese patients. A total of 571 (93.1%) patients were diagnosed, including 428 (69.8%) with large deletions/duplications and 143 (23.3%) with point mutations. Deletion/duplication breakpoints gathered mostly in introns 44-55. Reading frame rules could explain 88.6% of deletion mutations. We identified seventy novel point mutations that had not been previously reported. Spectrum expansion and genotype-phenotype analysis of DMD mutations on such a large sample size in Han Chinese population would provide new insights into the pathogenic mechanism underlying dystrophinopathies.

Application of molecular imaging combined with genetic screening in diagnosing MELAS, diabetes and recurrent pancreatitis.

PubMed

Zhiping, W; Quwen, L; Hai, Z; Jian, Z; Peiyi, G

2016-01-01

We report molecular imaging combined with gene diagnosis in a family with 7 members who carried an A3243G mutation in mitochondrial tRNA and p.Thr 137 Met in cationic trypsinogen (PRSS1) gene presented with mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes (MELAS), diabetes, and recurrent pancreatitis. DNA sequencing was used to detect and validate mitochondrial DNA and PRSS1. We also verified that mitochondrial heterozygous mutations and c.410 C>T mutation causing p.Thr 137 Met could be detected in oral epithelial cells or in urine sediment cells. In addition, molecular imaging was carried out in the affected family members. In this pedigree, MELAS syndrome accompanied by pancreatitis was an important clinical feature, followed by diabetes. Heteroplasmy of the mtDNA A3243G and c.410 C>T mutation of PRSS1 was found in all tissue samples of these patients, but no mutations were found in 520 normal control and normal individuals of the family. However, based on molecular imaging observations, patients with relatively higher lactate/pyruvate levels had more typical and more severe symptoms, particularly those of pancreatic disease (diabetes or pancreatitis). MELAS syndrome may be associated with pancreatitis. For the diagnosis, it is more reasonable to perform molecular imaging combined with gene diagnosis.

Exome sequencing identifies DYNC2H1 mutations as a common cause of asphyxiating thoracic dystrophy (Jeune syndrome) without major polydactyly, renal or retinal involvement

PubMed Central

Schmidts, Miriam; Arts, Heleen H; Bongers, Ernie M H F; Yap, Zhimin; Oud, Machteld M; Antony, Dinu; Duijkers, Lonneke; Emes, Richard D; Stalker, Jim; Yntema, Jan-Bart L; Plagnol, Vincent; Hoischen, Alexander; Gilissen, Christian; Forsythe, Elisabeth; Lausch, Ekkehart; Veltman, Joris A; Roeleveld, Nel; Superti-Furga, Andrea; Kutkowska-Kazmierczak, Anna; Kamsteeg, Erik-Jan; Elçioğlu, Nursel; van Maarle, Merel C; Graul-Neumann, Luitgard M; Devriendt, Koenraad; Smithson, Sarah F; Wellesley, Diana; Verbeek, Nienke E; Hennekam, Raoul C M; Kayserili, Hulya; Scambler, Peter J; Beales, Philip L; Knoers, Nine VAM; Roepman, Ronald; Mitchison, Hannah M

2013-01-01

Background Jeune asphyxiating thoracic dystrophy (JATD) is a rare, often lethal, recessively inherited chondrodysplasia characterised by shortened ribs and long bones, sometimes accompanied by polydactyly, and renal, liver and retinal disease. Mutations in intraflagellar transport (IFT) genes cause JATD, including the IFT dynein-2 motor subunit gene DYNC2H1. Genetic heterogeneity and the large DYNC2H1 gene size have hindered JATD genetic diagnosis. Aims and methods To determine the contribution to JATD we screened DYNC2H1 in 71 JATD patients JATD patients combining SNP mapping, Sanger sequencing and exome sequencing. Results and conclusions We detected 34 DYNC2H1 mutations in 29/71 (41%) patients from 19/57 families (33%), showing it as a major cause of JATD especially in Northern European patients. This included 13 early protein termination mutations (nonsense/frameshift, deletion, splice site) but no patients carried these in combination, suggesting the human phenotype is at least partly hypomorphic. In addition, 21 missense mutations were distributed across DYNC2H1 and these showed some clustering to functional domains, especially the ATP motor domain. DYNC2H1 patients largely lacked significant extra-skeletal involvement, demonstrating an important genotype–phenotype correlation in JATD. Significant variability exists in the course and severity of the thoracic phenotype, both between affected siblings with identical DYNC2H1 alleles and among individuals with different alleles, which suggests the DYNC2H1 phenotype might be subject to modifier alleles, non-genetic or epigenetic factors. Assessment of fibroblasts from patients showed accumulation of anterograde IFT proteins in the ciliary tips, confirming defects similar to patients with other retrograde IFT machinery mutations, which may be of undervalued potential for diagnostic purposes. PMID:23456818

The heartstrings mutation in zebrafish causes heart/fin Tbx5 deficiency syndrome.

PubMed

Garrity, Deborah M; Childs, Sarah; Fishman, Mark C

2002-10-01

Holt-Oram syndrome is one of the autosomal dominant human "heart-hand" disorders, with a combination of upper limb malformations and cardiac defects. Holt-Oram syndrome is caused by mutations in the TBX5 gene, a member of a large family of T-box transcription factors that play important roles in cell-type specification and morphogenesis. In a screen for mutations affecting zebrafish cardiac function, we isolated the recessive lethal mutant heartstrings, which lacks pectoral fins and exhibits severe cardiac dysfunction, beginning with a slow heart rate and progressing to a stretched, non-functional heart. We mapped and cloned the heartstrings mutation and find it to encode the zebrafish ortholog of the TBX5 gene. The heartstrings mutation causes premature termination at amino acid 316. Homozygous mutant embryos never develop pectoral fin buds and do not express several markers of early fin differentiation. The total absence of any fin bud differentiation distinguishes heartstrings from most other mutations that affect zebrafish fin development, suggesting that Tbx5 functions very early in the pectoral fin induction pathway. Moderate reduction of Tbx5 by morpholino causes fin malformations, revealing an additional early requirement for Tbx5 in coordinating the axes of fin outgrowth. The heart of heartstrings mutant embryos appears to form and function normally through the early heart tube stage, manifesting only a slight bradycardia compared with wild-type siblings. However, the heart fails to loop and then progressively deteriorates, a process affecting the ventricle as well as the atrium. Relative to mammals, fish require lower levels of Tbx5 to produce malformed appendages and display whole-heart rather than atrial-predominant cardiac defects. However, the syndromic deficiencies of tbx5 mutation are remarkably well retained between fish and mammals.

De novo germline and postzygotic mutations in AKT3, PIK3R2 and PIK3CA cause a spectrum of related megalencephaly syndromes

PubMed Central

Rivière, Jean-Baptiste; Mirzaa, Ghayda M.; O’Roak, Brian J.; Beddaoui, Margaret; Alcantara, Diana; Conway, Robert L.; St-Onge, Judith; Schwartzentruber, Jeremy A.; Gripp, Karen W.; Nikkel, Sarah M.; Worthylake, Thea; Sullivan, Christopher T.; Ward, Thomas R.; Butler, Hailly E.; Kramer, Nancy A.; Albrecht, Beate; Armour, Christine M.; Armstrong, Linlea; Caluseriu, Oana; Cytrynbaum, Cheryl; Drolet, Beth A.; Innes, A. Micheil; Lauzon, Julie L.; Lin, Angela E.; Mancini, Grazia M. S.; Meschino, Wendy S.; Reggin, James D.; Saggar, Anand K.; Lerman-Sagie, Tally; Uyanik, Gökhan; Weksberg, Rosanna; Zirn, Birgit; Beaulieu, Chandree L.; Majewski, Jacek; Bulman, Dennis E.; O’Driscoll, Mark; Shendure, Jay; Graham, John M.; Boycott, Kym M.; Dobyns, William B.

2012-01-01

Megalencephaly-capillary malformation (MCAP) and megalencephaly-polymicrogyria-polydactyly-hydrocephalus (MPPH) syndromes are sporadic overgrowth disorders associated with markedly enlarged brain size and other recognizable features1-5. We performed exome sequencing in three families with MCAP or MPPH and confirmed our initial observations in exomes from 7 MCAP and 174 control individuals, as well as in 40 additional megalencephaly subjects using a combination of Sanger sequencing, restriction-enzyme assays, and targeted deep sequencing. We identified de novo germline or postzygotic mutations in three core components of the phosphatidylinositol-3-kinase (PI3K)/AKT pathway. These include two mutations of AKT3, one recurrent mutation of PIK3R2 in 11 unrelated MPPH families, and 15 mostly postzygotic mutations of PIK3CA in 23 MCAP and one MPPH patients. Our data highlight the central role of PI3K/AKT signaling in vascular, limb and brain development, and emphasize the power of massively parallel sequencing in a challenging context of phenotypic and genetic heterogeneity combined with postzygotic mosaicism. PMID:22729224

De novo germline and postzygotic mutations in AKT3, PIK3R2 and PIK3CA cause a spectrum of related megalencephaly syndromes.

PubMed

Rivière, Jean-Baptiste; Mirzaa, Ghayda M; O'Roak, Brian J; Beddaoui, Margaret; Alcantara, Diana; Conway, Robert L; St-Onge, Judith; Schwartzentruber, Jeremy A; Gripp, Karen W; Nikkel, Sarah M; Worthylake, Thea; Sullivan, Christopher T; Ward, Thomas R; Butler, Hailly E; Kramer, Nancy A; Albrecht, Beate; Armour, Christine M; Armstrong, Linlea; Caluseriu, Oana; Cytrynbaum, Cheryl; Drolet, Beth A; Innes, A Micheil; Lauzon, Julie L; Lin, Angela E; Mancini, Grazia M S; Meschino, Wendy S; Reggin, James D; Saggar, Anand K; Lerman-Sagie, Tally; Uyanik, Gökhan; Weksberg, Rosanna; Zirn, Birgit; Beaulieu, Chandree L; Majewski, Jacek; Bulman, Dennis E; O'Driscoll, Mark; Shendure, Jay; Graham, John M; Boycott, Kym M; Dobyns, William B

2012-06-24

Megalencephaly-capillary malformation (MCAP) and megalencephaly-polymicrogyria-polydactyly-hydrocephalus (MPPH) syndromes are sporadic overgrowth disorders associated with markedly enlarged brain size and other recognizable features. We performed exome sequencing in 3 families with MCAP or MPPH, and our initial observations were confirmed in exomes from 7 individuals with MCAP and 174 control individuals, as well as in 40 additional subjects with megalencephaly, using a combination of Sanger sequencing, restriction enzyme assays and targeted deep sequencing. We identified de novo germline or postzygotic mutations in three core components of the phosphatidylinositol 3-kinase (PI3K)-AKT pathway. These include 2 mutations in AKT3, 1 recurrent mutation in PIK3R2 in 11 unrelated families with MPPH and 15 mostly postzygotic mutations in PIK3CA in 23 individuals with MCAP and 1 with MPPH. Our data highlight the central role of PI3K-AKT signaling in vascular, limb and brain development and emphasize the power of massively parallel sequencing in a challenging context of phenotypic and genetic heterogeneity combined with postzygotic mosaicism.

Homozygous STIL Mutation Causes Holoprosencephaly and Microcephaly in Two Siblings

PubMed Central

Mouden, Charlotte; de Tayrac, Marie; Dubourg, Christèle; Rose, Sophie; Carré, Wilfrid; Hamdi-Rozé, Houda; Babron, Marie-Claude; Akloul, Linda; Héron-Longe, Bénédicte; Odent, Sylvie; Dupé, Valérie; Giet, Régis; David, Véronique

2015-01-01

Holoprosencephaly (HPE) is a frequent congenital malformation of the brain characterized by impaired forebrain cleavage and midline facial anomalies. Heterozygous mutations in 14 genes have been identified in HPE patients that account for only 30% of HPE cases, suggesting the existence of other HPE genes. Data from homozygosity mapping and whole-exome sequencing in a consanguineous Turkish family were combined to identify a homozygous missense mutation (c.2150G>A; p.Gly717Glu) in STIL, common to the two affected children. STIL has a role in centriole formation and has previously been described in rare cases of microcephaly. Rescue experiments in U2OS cells showed that the STIL p.Gly717Glu mutation was not able to fully restore the centriole duplication failure following depletion of endogenous STIL protein indicating the deleterious role of the mutation. In situ hybridization experiments using chick embryos demonstrated that expression of Stil was in accordance with a function during early patterning of the forebrain. It is only the second time that a STIL homozygous mutation causing a recessive form of HPE was reported. This result also supports the genetic heterogeneity of HPE and increases the panel of genes to be tested for HPE diagnosis. PMID:25658757

[A study on mutations of the overlapping hepatitis B virus surface and polymerase gene in patients with HBV reinfection after liver transplantations].

PubMed

Song, Hong-li; Shen, Zhong-yang; Wang, Jian; Zheng, Wei-ping; Wang, Zheng-lu

2008-04-01

To investigate the influence of combined hepatitis B immune globulin (HBIG) and lamivudine (LMV) treatment on hepatitis B virus (HBV) surface antigen and polymerase overlapping gene mutations in HBV reinfected liver transplant recipients. From June 2002 to December 2003, 320 patients who underwent liver transplantations due to HBV-related end-stage liver diseases were followed-up for 1.5 to 3 years postoperatively. Fourteen patients developed HBV reinfection. They had LMV before their liver transplantations and had LMV and HBIG after the transplantations to prevent HBV infections. Their serum levels of HBV DNA were measured by polymerase chain reaction. Gene sequencing method was used to analyze HBV genotype and mutations of the S gene. Micro-particle enzyme immunoassay was used to measure the serum concentration of HBIG. (1) There was no obvious difference in the number of amino acid mutation sites in S and P regions before and after the transplantations. (2) The HBV genotypes were B-type (n=2) and C-type (n=12) in the reinfected group before the transplantations, and genotypes after the transplantations remained the same. (3) HBIG concentrations were 0 U/L in 7 patients, less than 100 U/L in 5 patients, and more than 100 U/L in 2 patients. Mutations were detected as I126S, T131N, S143T and G145R in 'a' determinant and L110F, I113S, T160K in up- or down-stream of 'a' determinant. (4) Mutations in S gene caused missense mutation in the surface antigen region. These mutations also caused corresponding missense mutations in the polymerase region. The missense mutation in the polymerase region involved lamivudine mutation sites and other mutation sites. Immunosuppressant therapy has no obvious influence on the numbers of mutations, but it can influence the sites of the mutations. Besides 'a' determinant mutations, there exist mutations in up- or down-streams of 'a' determinant and they may cause HBV reinfection.

Combining Single Strand Oligodeoxynucleotides and CRISPR/Cas9 to Correct Gene Mutations in β-Thalassemia-induced Pluripotent Stem Cells.

PubMed

Niu, Xiaohua; He, Wenyin; Song, Bing; Ou, Zhanhui; Fan, Di; Chen, Yuchang; Fan, Yong; Sun, Xiaofang

2016-08-05

β-Thalassemia (β-Thal) is one of the most common genetic diseases in the world. The generation of patient-specific β-Thal-induced pluripotent stem cells (iPSCs), correction of the disease-causing mutations in those cells, and then differentiation into hematopoietic stem cells offers a new therapeutic strategy for this disease. Here, we designed a CRISPR/Cas9 to specifically target the Homo sapiens hemoglobin β (HBB) gene CD41/42(-CTTT) mutation. We demonstrated that the combination of single strand oligodeoxynucleotides with CRISPR/Cas9 was capable of correcting the HBB gene CD41/42 mutation in β-Thal iPSCs. After applying a correction-specific PCR assay to purify the corrected clones followed by sequencing to confirm mutation correction, we verified that the purified clones retained full pluripotency and exhibited normal karyotyping. Additionally, whole-exome sequencing showed that the mutation load to the exomes was minimal after CRISPR/Cas9 targeting. Furthermore, the corrected iPSCs were selected for erythroblast differentiation and restored the expression of HBB protein compared with the parental iPSCs. This method provides an efficient and safe strategy to correct the HBB gene mutation in β-Thal iPSCs. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Genome-Wide Mutation Rate Response to pH Change in the Coral Reef Pathogen Vibrio shilonii AK1.

PubMed

Strauss, Chloe; Long, Hongan; Patterson, Caitlyn E; Te, Ronald; Lynch, Michael

2017-08-22

Recent application of mutation accumulation techniques combined with whole-genome sequencing (MA/WGS) has greatly promoted studies of spontaneous mutation. However, such explorations have rarely been conducted on marine organisms, and it is unclear how marine habitats have influenced genome stability. This report resolves the mutation rate and spectrum of the coral reef pathogen Vibrio shilonii , which causes coral bleaching and endangers the biodiversity maintained by coral reefs. We found that its mutation rate and spectrum are highly similar to those of other studied bacteria from various habitats, despite the saline environment. The mutational properties of this marine bacterium are thus controlled by other general evolutionary forces such as natural selection and genetic drift. We also found that as pH drops, the mutation rate decreases and the mutation spectrum is biased in the direction of generating G/C nucleotides. This implies that evolutionary features of this organism and perhaps other marine microbes might be altered by the increasingly acidic ocean water caused by excess CO 2 emission. Nonetheless, further exploration is needed as the pH range tested in this study was rather narrow and many other possible mutation determinants, such as carbonate increase, are associated with ocean acidification. IMPORTANCE This study explored the pH dependence of a bacterial genome-wide mutation rate. We discovered that the genome-wide rates of appearance of most mutation types decrease linearly and that the mutation spectrum is biased in generating more G/C nucleotides with pH drop in the coral reef pathogen V. shilonii . Copyright © 2017 Strauss et al.

Mutations in KEOPS-complex genes cause nephrotic syndrome with primary microcephaly.

PubMed

Braun, Daniela A; Rao, Jia; Mollet, Geraldine; Schapiro, David; Daugeron, Marie-Claire; Tan, Weizhen; Gribouval, Olivier; Boyer, Olivia; Revy, Patrick; Jobst-Schwan, Tilman; Schmidt, Johanna Magdalena; Lawson, Jennifer A; Schanze, Denny; Ashraf, Shazia; Ullmann, Jeremy F P; Hoogstraten, Charlotte A; Boddaert, Nathalie; Collinet, Bruno; Martin, Gaëlle; Liger, Dominique; Lovric, Svjetlana; Furlano, Monica; Guerrera, I Chiara; Sanchez-Ferras, Oraly; Hu, Jennifer F; Boschat, Anne-Claire; Sanquer, Sylvia; Menten, Björn; Vergult, Sarah; De Rocker, Nina; Airik, Merlin; Hermle, Tobias; Shril, Shirlee; Widmeier, Eugen; Gee, Heon Yung; Choi, Won-Il; Sadowski, Carolin E; Pabst, Werner L; Warejko, Jillian K; Daga, Ankana; Basta, Tamara; Matejas, Verena; Scharmann, Karin; Kienast, Sandra D; Behnam, Babak; Beeson, Brendan; Begtrup, Amber; Bruce, Malcolm; Ch'ng, Gaik-Siew; Lin, Shuan-Pei; Chang, Jui-Hsing; Chen, Chao-Huei; Cho, Megan T; Gaffney, Patrick M; Gipson, Patrick E; Hsu, Chyong-Hsin; Kari, Jameela A; Ke, Yu-Yuan; Kiraly-Borri, Cathy; Lai, Wai-Ming; Lemyre, Emmanuelle; Littlejohn, Rebecca Okashah; Masri, Amira; Moghtaderi, Mastaneh; Nakamura, Kazuyuki; Ozaltin, Fatih; Praet, Marleen; Prasad, Chitra; Prytula, Agnieszka; Roeder, Elizabeth R; Rump, Patrick; Schnur, Rhonda E; Shiihara, Takashi; Sinha, Manish D; Soliman, Neveen A; Soulami, Kenza; Sweetser, David A; Tsai, Wen-Hui; Tsai, Jeng-Daw; Topaloglu, Rezan; Vester, Udo; Viskochil, David H; Vatanavicharn, Nithiwat; Waxler, Jessica L; Wierenga, Klaas J; Wolf, Matthias T F; Wong, Sik-Nin; Leidel, Sebastian A; Truglio, Gessica; Dedon, Peter C; Poduri, Annapurna; Mane, Shrikant; Lifton, Richard P; Bouchard, Maxime; Kannu, Peter; Chitayat, David; Magen, Daniella; Callewaert, Bert; van Tilbeurgh, Herman; Zenker, Martin; Antignac, Corinne; Hildebrandt, Friedhelm

2017-10-01

Galloway-Mowat syndrome (GAMOS) is an autosomal-recessive disease characterized by the combination of early-onset nephrotic syndrome (SRNS) and microcephaly with brain anomalies. Here we identified recessive mutations in OSGEP, TP53RK, TPRKB, and LAGE3, genes encoding the four subunits of the KEOPS complex, in 37 individuals from 32 families with GAMOS. CRISPR-Cas9 knockout in zebrafish and mice recapitulated the human phenotype of primary microcephaly and resulted in early lethality. Knockdown of OSGEP, TP53RK, or TPRKB inhibited cell proliferation, which human mutations did not rescue. Furthermore, knockdown of these genes impaired protein translation, caused endoplasmic reticulum stress, activated DNA-damage-response signaling, and ultimately induced apoptosis. Knockdown of OSGEP or TP53RK induced defects in the actin cytoskeleton and decreased the migration rate of human podocytes, an established intermediate phenotype of SRNS. We thus identified four new monogenic causes of GAMOS, describe a link between KEOPS function and human disease, and delineate potential pathogenic mechanisms.

Detecting novel genetic mutations in Chinese Usher syndrome families using next-generation sequencing technology.

PubMed

Qu, Ling-Hui; Jin, Xin; Xu, Hai-Wei; Li, Shi-Ying; Yin, Zheng-Qin

2015-02-01

Usher syndrome (USH) is the most common cause of combined blindness and deafness inherited in an autosomal recessive mode. Molecular diagnosis is of great significance in revealing the molecular pathogenesis and aiding the clinical diagnosis of this disease. However, molecular diagnosis remains a challenge due to high phenotypic and genetic heterogeneity in USH. This study explored an approach for detecting disease-causing genetic mutations in candidate genes in five index cases from unrelated USH families based on targeted next-generation sequencing (NGS) technology. Through systematic data analysis using an established bioinformatics pipeline and segregation analysis, 10 pathogenic mutations in the USH disease genes were identified in the five USH families. Six of these mutations were novel: c.4398G > A and EX38-49del in MYO7A, c.988_989delAT in USH1C, c.15104_15105delCA and c.6875_6876insG in USH2A. All novel variations segregated with the disease phenotypes in their respective families and were absent from ethnically matched control individuals. This study expanded the mutation spectrum of USH and revealed the genotype-phenotype relationships of the novel USH mutations in Chinese patients. Moreover, this study proved that targeted NGS is an accurate and effective method for detecting genetic mutations related to USH. The identification of pathogenic mutations is of great significance for elucidating the underlying pathophysiology of USH.

PMS2 monoallelic mutation carriers: the known unknown

PubMed Central

Goodenberger, McKinsey L.; Thomas, Brittany C.; Riegert-Johnson, Douglas; Boland, C. Richard; Plon, Sharon E.; Clendenning, Mark; Ko Win, Aung; Senter, Leigha; Lipkin, Steven M.; Stadler, Zsofia K.; Macrae, Finlay A.; Lynch, Henry T.; Weitzel, Jeffrey N.; de la Chapelle, Albert; Syngal, Sapna; Lynch, Patrick; Parry, Susan; Jenkins, Mark A.; Gallinger, Steven; Holter, Spring; Aronson, Melyssa; Newcomb, Polly A.; Burnett, Terrilea; Le Marchand, Loïc; Pichurin, Pavel; Hampel, Heather; Terdiman, Jonathan P.; Lu, Karen H.; Thibodeau, Stephen; Lindor, Noralane M.

2016-01-01

Germline mutations in MLH1, MSH2, MSH6 and PMS2 have been shown to cause Lynch syndrome. The penetrance for cancer and tumor spectrum has been repeatedly studied and multiple professional societies have proposed clinical management guidelines for affected individuals. Several studies have demonstrated a reduced penetrance for monoallelic carriers of PMS2 mutations compared to the other mismatch repair (MMR) genes, but clinical management guidelines have largely proposed the same screening recommendations for all MMR gene carriers. The authors considered whether enough evidence existed to propose new screening guidelines specific to PMS2 mutation carriers with regard to age of onset and frequency of colonic screening. Published reports of PMS2 germline mutations were combined with unpublished cases from the authors’ research registries and clinical practices, and a discussion of potential modification of cancer screening guidelines was pursued. A total of 234 monoallelic PMS2 mutation carriers from 170 families were included. Approximately 8% of those with CRC were diagnosed under age 30 and each of these tumors presented on the left-side of the colon. As it is currently unknown what causes the early-onset of CRC in some families with monoallelic PMS2 germline mutations, the authors recommend against reducing cancer surveillance guidelines in families found having monoallelic PMS2 mutations in spite of the documented reduced penetrance. PMID:25856668

PMS2 monoallelic mutation carriers: the known unknown.

PubMed

Goodenberger, McKinsey L; Thomas, Brittany C; Riegert-Johnson, Douglas; Boland, C Richard; Plon, Sharon E; Clendenning, Mark; Win, Aung Ko; Senter, Leigha; Lipkin, Steven M; Stadler, Zsofia K; Macrae, Finlay A; Lynch, Henry T; Weitzel, Jeffrey N; de la Chapelle, Albert; Syngal, Sapna; Lynch, Patrick; Parry, Susan; Jenkins, Mark A; Gallinger, Steven; Holter, Spring; Aronson, Melyssa; Newcomb, Polly A; Burnett, Terrilea; Le Marchand, Loïc; Pichurin, Pavel; Hampel, Heather; Terdiman, Jonathan P; Lu, Karen H; Thibodeau, Stephen; Lindor, Noralane M

2016-01-01

Germ-line mutations in MLH1, MSH2, MSH6, and PMS2 have been shown to cause Lynch syndrome. The penetrance of the cancer and tumor spectrum has been repeatedly studied, and multiple professional societies have proposed clinical management guidelines for affected individuals. Several studies have demonstrated a reduced penetrance for monoallelic carriers of PMS2 mutations compared with the other mismatch repair (MMR) genes, but clinical management guidelines have largely proposed the same screening recommendations for all MMR gene carriers. The authors considered whether enough evidence existed to propose new screening guidelines specific to PMS2 mutation carriers with regard to age at onset and frequency of colonic screening. Published reports of PMS2 germ-line mutations were combined with unpublished cases from the authors' research registries and clinical practices, and a discussion of potential modification of cancer screening guidelines was pursued. A total of 234 monoallelic PMS2 mutation carriers from 170 families were included. Approximately 8% of those with colorectal cancer (CRC) were diagnosed before age 30, and each of these tumors presented on the left side of the colon. As it is currently unknown what causes the early onset of CRC in some families with monoallelic PMS2 germline mutations, the authors recommend against reducing cancer surveillance guidelines in families found having monoallelic PMS2 mutations in spite of the reduced penetrance.Genet Med 18 1, 13-19.

Mutations in the evolutionarily highly conserved KEOPS complex genes cause nephrotic syndrome with microcephaly

PubMed Central

Braun, Daniela A.; Rao, Jia; Mollet, Geraldine; Schapiro, David; Daugeron, Marie-Claire; Tan, Weizhen; Gribouval, Olivier; Boyer, Olivia; Revy, Patrick; Jobst-Schwan, Tilman; Schmidt, Johanna Magdalena; Lawson, Jennifer A.; Schanze, Denny; Ashraf, Shazia; Boddaert, Nathalie; Collinet, Bruno; Martin, Gaëlle; Liger, Dominique; Lovric, Svjetlana; Furlano, Monica; Guerrera, I. Chiara; Sanchez-Ferras, Oraly; Menten, Björn; Vergult, Sarah; De Rocker, Nina; Airik, Merlin; Hermle, Tobias; Shril, Shirlee; Widmeier, Eugen; Gee, Heon Yung; Choi, Won-Il; Sadowski, Carolin E.; Pabst, Werner L.; Warejko, Jillian; Daga, Ankana; LeBerre, Tamara Basta; Matejas, Verena; Behnam, Babak; Beeson, Brendan; Begtrup, Amber; Bruce, Malcolm; Ch'ng, Gaik-Siew; Lin, Shuan-Pei; Chang, Jui-Hsing; Chen, Chao-Huei; Cho, Megan T.; Gipson, Patrick E.; Hsu, Chyong-Hsin; Kari, Jameela A.; Ke, Yu-Yuan; Kiraly-Borri, Cathy; Lai, Wai-ming; Lemyre, Emmanuelle; Littlejohn, Rebecca Okasha; Masri, Amira; Moghtaderi, Mastaneh; Nakamura, Kazuyuki; Praet, Marleen; Prasad, Chitra; Prytula, Agnieszka; Roeder, Elizabeth; Rump, Patrick; Schnur, Rhonda E.; Shiihara, Takashi; Sinha, Manish; Soliman, Neveen A; Soulami, Kenza; Sweetser, David A.; Tsai, Wen-Hui; Tsai, Jeng-Daw; Vester, Udo; Viskochil, David H.; Vatanavicharn, Nithiwat; Waxler, Jessica L.; Wolf, Matthias T.F.; Wong, Sik-Nin; Poduri, Annapurna; Truglio, Gessica; Mane, Shrikant; Lifton, Richard P.; Bouchard, Maxime; Kannu, Peter; Chitayat, David; Magen, Daniella; Calleweart, Bert; van Tilbeurgh, Herman; Zenker, Martin; Antignac, Corinne; Hildebrandt, Friedhelm

2018-01-01

Galloway-Mowat syndrome (GAMOS) is a severe autosomal-recessive disease characterized by the combination of early-onset steroid-resistant nephrotic syndrome (SRNS) and microcephaly with brain anomalies. To date, mutations of WDR73 are the only known monogenic cause of GAMOS and in most affected individuals the molecular diagnosis remains elusive. We here identify recessive mutations of OSGEP, TP53RK, TPRKB, or LAGE3, encoding the 4 subunits of the KEOPS complex in 33 individuals of 30 families with GAMOS. CRISPR/Cas9 knockout in zebrafish and mice recapitulates the human phenotype of microcephaly and results in early lethality. Knockdown of OSGEP, TP53RK, or TPRKB inhibits cell proliferation, which human mutations fail to rescue, and knockdown of either gene activates DNA damage response signaling and induces apoptosis. OSGEP and TP53RK molecularly interact and co-localize with the actin-regulating ARP2/3 complex. Furthermore, knockdown of OSGEP and TP53RK induces defects of the actin cytoskeleton and reduces migration rate of human podocytes, an established intermediate phenotype of SRNS. We thus identify 4 novel monogenic causes of GAMOS, describe the first link between KEOPS function and human disease, and delineate potential pathogenic mechanisms. PMID:28805828

High-throughput sequencing of the entire genomic regions of CCM1/KRIT1, CCM2 and CCM3/PDCD10 to search for pathogenic deep-intronic splice mutations in cerebral cavernous malformations.

PubMed

Rath, Matthias; Jenssen, Sönke E; Schwefel, Konrad; Spiegler, Stefanie; Kleimeier, Dana; Sperling, Christian; Kaderali, Lars; Felbor, Ute

2017-09-01

Cerebral cavernous malformations (CCM) are vascular lesions of the central nervous system that can cause headaches, seizures and hemorrhagic stroke. Disease-associated mutations have been identified in three genes: CCM1/KRIT1, CCM2 and CCM3/PDCD10. The precise proportion of deep-intronic variants in these genes and their clinical relevance is yet unknown. Here, a long-range PCR (LR-PCR) approach for target enrichment of the entire genomic regions of the three genes was combined with next generation sequencing (NGS) to screen for coding and non-coding variants. NGS detected all six CCM1/KRIT1, two CCM2 and four CCM3/PDCD10 mutations that had previously been identified by Sanger sequencing. Two of the pathogenic variants presented here are novel. Additionally, 20 stringently selected CCM index cases that had remained mutation-negative after conventional sequencing and exclusion of copy number variations were screened for deep-intronic mutations. The combination of bioinformatics filtering and transcript analyses did not reveal any deep-intronic splice mutations in these cases. Our results demonstrate that target enrichment by LR-PCR combined with NGS can be used for a comprehensive analysis of the entire genomic regions of the CCM genes in a research context. However, its clinical utility is limited as deep-intronic splice mutations in CCM1/KRIT1, CCM2 and CCM3/PDCD10 seem to be rather rare. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

IMHOTEP—a composite score integrating popular tools for predicting the functional consequences of non-synonymous sequence variants

PubMed Central

Knecht, Carolin; Mort, Matthew; Junge, Olaf; Cooper, David N.; Krawczak, Michael

2017-01-01

Abstract The in silico prediction of the functional consequences of mutations is an important goal of human pathogenetics. However, bioinformatic tools that classify mutations according to their functionality employ different algorithms so that predictions may vary markedly between tools. We therefore integrated nine popular prediction tools (PolyPhen-2, SNPs&GO, MutPred, SIFT, MutationTaster2, Mutation Assessor and FATHMM as well as conservation-based Grantham Score and PhyloP) into a single predictor. The optimal combination of these tools was selected by means of a wide range of statistical modeling techniques, drawing upon 10 029 disease-causing single nucleotide variants (SNVs) from Human Gene Mutation Database and 10 002 putatively ‘benign’ non-synonymous SNVs from UCSC. Predictive performance was found to be markedly improved by model-based integration, whilst maximum predictive capability was obtained with either random forest, decision tree or logistic regression analysis. A combination of PolyPhen-2, SNPs&GO, MutPred, MutationTaster2 and FATHMM was found to perform as well as all tools combined. Comparison of our approach with other integrative approaches such as Condel, CoVEC, CAROL, CADD, MetaSVM and MetaLR using an independent validation dataset, revealed the superiority of our newly proposed integrative approach. An online implementation of this approach, IMHOTEP (‘Integrating Molecular Heuristics and Other Tools for Effect Prediction’), is provided at http://www.uni-kiel.de/medinfo/cgi-bin/predictor/. PMID:28180317

Comprehensive Molecular Screening in Chinese Usher Syndrome Patients.

PubMed

Sun, Tengyang; Xu, Ke; Ren, Yanfan; Xie, Yue; Zhang, Xiaohui; Tian, Lu; Li, Yang

2018-03-01

Usher syndrome (USH) refers to a group of autosomal recessive disorders causing deafness and blindness. The objectives of this study were to determine the mutation spectrum in a cohort of Chinese patients with USH and to describe the clinical features of the patients with mutations. A total of 119 probands who were clinically diagnosed with USH were recruited for genetic analysis. All probands underwent ophthalmic examinations. A combination of molecular screening methods, including targeted next-generation sequencing, Sanger-DNA sequencing, and multiplex ligation probe amplification assay, was used to detect mutations. We found biallelic mutations in 92 probands (77.3%), monoallelic mutations in 5 patients (4.2%), and 1 hemizygous mutation in 1 patient (0.8%), resulting in an overall mutation detection rate of 78.2%. Overall, 132 distinct disease-causing mutations involving seven USH (ABHD12, CDH23, GPR98, MYO7A, PCDH15, USH1C, and USH2A) genes; 5 other retinal degeneration genes (CHM, CNGA1, EYS, PDE6B, and TULP1); and 1 nonsyndromic hearing loss gene (MYO15A) were identified, and 78 were novel. Mutations of MYOA7 were responsible for 60% of USH1 families, followed by PCDH15 (20%) and USH1C (10%). Mutations of USH2A accounted for 67.7% of USH2 families, and mutation c.8559-2A>G was the most frequent one, accounting for 19.1% of the identified USH2A alleles. Our results confirm that the mutation spectrum for each USH gene in Chinese patients differs from those of other populations. The formation of the mutation profile for the Chinese population will enable a precise genetic diagnosis for USH patients in the future.

A novel mutation in the MYO7A gene is associated with Usher syndrome type 1 in a Chinese family.

PubMed

He, Xiaoguang; Peng, Qi; Li, Siping; Zhu, Pengyuan; Wu, Chunqiu; Rao, Chunbao; Lin, Jingqi; Lu, Xiaomei

2017-08-01

We aimed to investigate the genetic causes of hearing loss in a Chinese proband with autosomal recessive congenital deafness. The targeted capture of 159 known deafness genes and next-generation sequencing were performed to study the genetic causes of hearing loss in the Chinese family. Sanger sequencing was employed to verify the variant mutations in members of this family. The proband harbored two mutations in the MYO7A gene in the form of compound heterozygosity. She was found to be heterozygous for a novel insertion mutation c.3847_3848 ins TCTG (p.N1285LfsX24) in exon 30 and for the known mutation c.2239_2240delAG (p.R747S fsX16)in exon 19. The novel mutation was absent in the 1000 Genomes Project. These variants were carried in the heterozygous state by the parents and were therefore co-segregated with the genetic disease. Clinical re-assessment, including detailed audiologic and ocular examinations, revealed congenital deafness and retinitis pigmentosa in the proband. Collectively, the combination of audiometric, ophthalmologic and genetic examinations successfully confirmed the phenotype of Usher syndrome type 1 (USH1). This study demonstrates that the novel mutation c.3847_3848insTCTG (p. N1285LfsX24) in compound heterozygosity with c.2239_2240delAG in the MYO7A gene is the main cause of USH1 in the proband. Our study expands the mutational spectrum of MYO7A and provides a foundation for further investigations elucidating the MYO7A-related mechanisms of USH1. Copyright © 2017 Elsevier B.V. All rights reserved.

Novel Mutations in HESX1 and PROP1 Genes in Combined Pituitary Hormone Deficiency.

PubMed

Avbelj Stefanija, Magdalena; Kotnik, Primož; Bratanič, Nina; Žerjav Tanšek, Mojca; Bertok, Sara; Bratina, Nataša; Battelino, Tadej; Trebušak Podkrajšek, Katarina

2015-01-01

The HESX1 gene is essential in forebrain development and pituitary organogenesis, and its mutations are the most commonly identified genetic cause of septo-optic dysplasia (SOD). The PROP1 gene is involved in anterior pituitary cell lineage specification and is commonly implicated in non-syndromic combined pituitary hormone deficiency (CPHD). We aimed to assess the involvement of HESX1 and PROP1 mutations in a cohort of patients with SOD and CPHD. Six patients with sporadic SOD and 16 patients with CPHD from 14 pedigrees were screened for mutations in HESX1 and PROP1 genes by exon sequencing. Half of the CPHD patients had variable associated clinical characteristics, such as hearing loss, orofacial cleft, kidney disorder or developmental delay. Novel variants were evaluated in silico and verified in SNP databases. A novel heterozygous p.Glu102Gly mutation in the HESX1 gene and a novel homozygous p.Arg121Thr mutation in the PROP1 gene were detected in 2 pedigrees with CPHD. A small previously reported deletion in PROP1 c.301_302delAG was detected in a separate patient with CPHD, in heterozygous state. No mutations were identified in patients with SOD. Our results expand the spectrum of mutations implicated in CPHD. The frequency of 15% of the PROP1 mutations in CPHD was low, likely due to the clinical heterogeneity of the cohort. © 2015 S. Karger AG, Basel.

Two Desmin Gene Mutations Associated with Myofibrillar Myopathies in Polish Families

PubMed Central

Berdynski, Mariusz; Sikorska, Agata; Filipek, Slawomir; Redowicz, Maria Jolanta; Kaminska, Anna; Zekanowski, Cezary

2014-01-01

Desmin is a muscle-specific intermediate filament protein which forms a network connecting the sarcomere, T tubules, sarcolemma, nuclear membrane, mitochondria and other organelles. Mutations in the gene coding for desmin (DES) cause skeletal myopathies often combined with cardiomyopathy, or isolated cardiomyopathies. The molecular pathomechanisms of the disease remain ambiguous. Here, we describe and comprehensively characterize two DES mutations found in Polish patients with a clinical diagnosis of desminopathy. The study group comprised 16 individuals representing three families. Two mutations were identified: a novel missense mutation (Q348P) and a small deletion of nine nucleotides (A357_E359del), previously described by us in the Polish population. A common ancestry of all the families bearing the A357_E359del mutation was confirmed. Both mutations were predicted to be pathogenic using a bioinformatics approach, including molecular dynamics simulations which helped to rationalize abnormal behavior at molecular level. To test the impact of the mutations on DES expression and the intracellular distribution of desmin muscle biopsies were investigated. Elevated desmin levels as well as its atypical localization in muscle fibers were observed. Additional staining for M-cadherin, α-actinin, and myosin heavy chains confirmed severe disruption of myofibrill organization. The abnormalities were more prominent in the Q348P muscle, where both small atrophic fibers as well large fibers with centrally localized nuclei were observed. We propose that the mutations affect desmin structure and cause its aberrant folding and subsequent aggregation, triggering disruption of myofibrils organization. PMID:25541946

Mutations in the NHEJ Component XRCC4 Cause Primordial Dwarfism

PubMed Central

Murray, Jennie E.; van der Burg, Mirjam; IJspeert, Hanna; Carroll, Paula; Wu, Qian; Ochi, Takashi; Leitch, Andrea; Miller, Edward S.; Kysela, Boris; Jawad, Alireza; Bottani, Armand; Brancati, Francesco; Cappa, Marco; Cormier-Daire, Valerie; Deshpande, Charu; Faqeih, Eissa A.; Graham, Gail E.; Ranza, Emmanuelle; Blundell, Tom L.; Jackson, Andrew P.; Stewart, Grant S.; Bicknell, Louise S.

2015-01-01

Non-homologous end joining (NHEJ) is a key cellular process ensuring genome integrity. Mutations in several components of the NHEJ pathway have been identified, often associated with severe combined immunodeficiency (SCID), consistent with the requirement for NHEJ during V(D)J recombination to ensure diversity of the adaptive immune system. In contrast, we have recently found that biallelic mutations in LIG4 are a common cause of microcephalic primordial dwarfism (MPD), a phenotype characterized by prenatal-onset extreme global growth failure. Here we provide definitive molecular genetic evidence supported by biochemical, cellular, and immunological data for mutations in XRCC4, encoding the obligate binding partner of LIG4, causing MPD. We report the identification of biallelic mutations in XRCC4 in five families. Biochemical and cellular studies demonstrate that these alterations substantially decrease XRCC4 protein levels leading to reduced cellular ligase IV activity. Consequently, NHEJ-dependent repair of ionizing-radiation-induced DNA double-strand breaks is compromised in XRCC4 cells. Similarly, immunoglobulin junctional diversification is impaired in cells. However, immunoglobulin levels are normal, and individuals lack overt signs of immunodeficiency. Additionally, in contrast to individuals with LIG4 mutations, pancytopenia leading to bone marrow failure has not been observed. Hence, alterations that alter different NHEJ proteins give rise to a phenotypic spectrum, from SCID to extreme growth failure, with deficiencies in certain key components of this repair pathway predominantly exhibiting growth deficits, reflecting differential developmental requirements for NHEJ proteins to support growth and immune maturation. PMID:25728776

Mutagenesis: Interactions with a parallel universe.

PubMed

Miller, Jeffrey H

Unexpected observations in mutagenesis research have led to a new perspective in this personal reflection based on years of studying mutagenesis. Many mutagens have been thought to operate via a single principal mechanism, with secondary effects usually resulting in only minor changes in the observed mutation frequencies and spectra. For example, we conceive of base analogs as resulting in direct mispairing as their main mechanism of mutagenesis. Recent studies now show that in fact even these simple mutagens can cause very large and unanticipated effects both in mutation frequencies and in the mutational spectra when used in certain pair-wise combinations. Here we characterize this leap in mutation frequencies as a transport to an alternate universe of mutagenesis. Copyright © 2018 Elsevier B.V. All rights reserved.

A retrospective study of clinical and mutational findings in 45 Danish families with ectodermal dysplasia.

PubMed

Tiedemann Svendsen, Mathias; Henningsen, Emil; Hertz, Jens Michael; Vestergaard Grejsen, Dorthe; Bygum, Anette

2014-09-01

Ectodermal dysplasias form a complex, nosologic group of diseases with defects in at least 2 ectodermal structures. A retrospective study of patients with ectodermal dysplasia seen at our department over a period of 19 years (1994-2013) was performed. The study population consisted of 67 patients covering 17 different diagnoses. Forty-five families were identified of which 26 were sporadic cases with no affected family members. In 27 tested families a disease-causing mutation was identified in 23 families. Eleven mutations were novel mutations. To our knowledge, we present the first large ectodermal dysplasia cohort focusing on clinical manifestations in combination with mutational analysis. We recommend a nationwide study to estimate the prevalence of the ectodermal dysplasia and to ensure relevant molecular genetic testing which may form the basis of a national ectodermal dysplasia database.

Recessive and dominant mutations in COL12A1 cause a novel EDS/myopathy overlap syndrome in humans and mice

PubMed Central

Zou, Yaqun; Zwolanek, Daniela; Izu, Yayoi; Gandhy, Shreya; Schreiber, Gudrun; Brockmann, Knut; Devoto, Marcella; Tian, Zuozhen; Hu, Ying; Veit, Guido; Meier, Markus; Stetefeld, Jörg; Hicks, Debbie; Straub, Volker; Voermans, Nicol C.; Birk, David E.; Barton, Elisabeth R.; Koch, Manuel; Bönnemann, Carsten G.

2014-01-01

Collagen VI-related myopathies are disorders of connective tissue presenting with an overlap phenotype combining clinical involvement from the muscle and from the connective tissue. Not all patients displaying related overlap phenotypes between muscle and connective tissue have mutations in collagen VI. Here, we report a homozygous recessive loss of function mutation and a de novo dominant mutation in collagen XII (COL12A1) as underlying a novel overlap syndrome involving muscle and connective tissue. Two siblings homozygous for a loss of function mutation showed widespread joint hyperlaxity combined with weakness precluding independent ambulation, while the patient with the de novo missense mutation was more mildly affected, showing improvement including the acquisition of walking. A mouse model with inactivation of the Col12a1 gene showed decreased grip strength, a delay in fiber-type transition and a deficiency in passive force generation while the muscle seems more resistant to eccentric contraction induced force drop, indicating a role for a matrix-based passive force-transducing elastic element in the generation of the weakness. This new muscle connective tissue overlap syndrome expands on the emerging importance of the muscle extracellular matrix in the pathogenesis of muscle disease. PMID:24334604

Parkinson disease: α-synuclein mutational screening and new clinical insight into the p.E46K mutation.

PubMed

Pimentel, Márcia M G; Rodrigues, Fabíola C; Leite, Marco Antônio A; Campos Júnior, Mário; Rosso, Ana Lucia; Nicaretta, Denise H; Pereira, João S; Silva, Delson José; Della Coletta, Marcus V; Vasconcellos, Luiz Felipe R; Abreu, Gabriella M; Dos Santos, Jussara M; Santos-Rebouças, Cíntia B

2015-06-01

Amongst Parkinson's disease-causing genetic factors, missense mutations and genomic multiplications in the gene encoding α-synuclein are well established causes of the disease, although genetic data in populations with a high degree of admixture, such as the Brazilian one, are still scarce. In this study, we conducted a molecular screening of α-synuclein point mutations and copy number variation in the largest cohort of Brazilian patients with Parkinson's disease (n = 549) and also in twelve Portuguese and one Bolivian immigrants. Genomic DNA was isolated from peripheral blood leukocytes or saliva, and the mutational screening was performed by quantitative and qualitative real-time PCR. The only alteration identified was the p.E46K mutation in a 60-year-old man, born in Bolivia, with a familial history of autosomal dominant Parkinson's disease. This is the second family ever reported, in which this rare pathogenic mutation is segregating. The same mutation was firstly described ten years ago in a Spanish family with a neurodegenerative syndrome combining parkinsonism, dementia and visual hallucinations. The clinical condition of our proband reveals a less aggressive phenotype than previously described and reinforces that marked phenotypic heterogeneity is common among patients with Parkinson's disease, even among those carriers sharing the same mutation. Our findings add new insight into the preexisting information about α-synuclein p.E46K, improving our understanding about the endophenotypes associated to this mutation and corroborate that missense alterations and multiplications in α-synuclein are uncommon among Brazilian patients with Parkinson's disease. Copyright © 2015 Elsevier Ltd. All rights reserved.

Stargardt disease: towards developing a model to predict phenotype.

PubMed

Heathfield, Laura; Lacerda, Miguel; Nossek, Christel; Roberts, Lisa; Ramesar, Rajkumar S

2013-10-01

Stargardt disease is an ABCA4-associated retinopathy, which generally follows an autosomal recessive inheritance pattern and is a frequent cause of macular degeneration in childhood. ABCA4 displays significant allelic heterogeneity whereby different mutations can cause retinal diseases with varying severity and age of onset. A genotype-phenotype model has been proposed linking ABCA4 mutations, purported ABCA4 functional protein activity and severity of disease, as measured by degree of visual loss and the age of onset. It has, however, been difficult to verify this model statistically in observational studies, as the number of individuals sharing any particular mutation combination is typically low. Seven founder mutations have been identified in a large number of Caucasian Afrikaner patients in South Africa, making it possible to test the genotype-phenotype model. A generalised linear model was developed to predict and assess the relative pathogenic contribution of the seven mutations to the age of onset of Stargardt disease. It is shown that the pathogenicity of an individual mutation can differ significantly depending on the genetic context in which it occurs. The results reported here may be used to identify suitable candidates for inclusion in clinical trials, as well as guide the genetic counselling of affected individuals and families.

Novel AQP2 mutation causing congenital nephrogenic diabetes insipidus: challenges in management during infancy.

PubMed

Rugpolmuang, Rottanat; Deeb, Asma; Hassan, Yousef; Deekajorndech, Tawatchai; Shotelersuk, Vorasuk; Sahakitrungruang, Taninee

2014-01-01

Congenital nephrogenic diabetes insipidus (NDI) is a rare inherited disorder, mostly caused by AVPR2 mutations. Less than 10% of cases are due to mutations in the aquaporin-2 (AQP2) gene. Diagnosis and management of this condition remain challenging especially during infancy. Here, we report two unrelated patients, a 6-month-old Thai boy and a 5-year-old Emirati girl, with a history of failure to thrive, chronic fever, polydipsia, and polyuria presented in early infancy. The results of water deprivation test were compatible with a diagnosis of NDI. The entire coding regions of the AVPR2 and AQP2 gene were amplified by polymerase chain reaction and sequenced. Patient 1 was homozygous for a novel missense AQP2 mutation p.G96E, inherited from both parents. Patient 2 harbored a previously described homozygous p.T126M mutation in the AQP2 gene. Both patients were treated with a combination of thiazide diuretics and amiloride. Patient 1 developed paradoxical hyponatremia and severe dehydration 2 weeks after medical treatment began. In conclusion, we report a novel mutation of the AQP2 gene and highlight an important role of genetic testing for definite diagnosis. Vigilant monitoring of the fluid status and electrolytes after beginning the therapy is mandatory in infants with NDI.

Stargardt Disease: towards developing a model to predict phenotype

PubMed Central

Heathfield, Laura; Lacerda, Miguel; Nossek, Christel; Roberts, Lisa; Ramesar, Rajkumar S

2013-01-01

Stargardt disease is an ABCA4-associated retinopathy, which generally follows an autosomal recessive inheritance pattern and is a frequent cause of macular degeneration in childhood. ABCA4 displays significant allelic heterogeneity whereby different mutations can cause retinal diseases with varying severity and age of onset. A genotype–phenotype model has been proposed linking ABCA4 mutations, purported ABCA4 functional protein activity and severity of disease, as measured by degree of visual loss and the age of onset. It has, however, been difficult to verify this model statistically in observational studies, as the number of individuals sharing any particular mutation combination is typically low. Seven founder mutations have been identified in a large number of Caucasian Afrikaner patients in South Africa, making it possible to test the genotype–phenotype model. A generalised linear model was developed to predict and assess the relative pathogenic contribution of the seven mutations to the age of onset of Stargardt disease. It is shown that the pathogenicity of an individual mutation can differ significantly depending on the genetic context in which it occurs. The results reported here may be used to identify suitable candidates for inclusion in clinical trials, as well as guide the genetic counselling of affected individuals and families. PMID:23695285

Structural signature of the G719S-T790M double mutation in the EGFR kinase domain and its response to inhibitors

PubMed Central

C., George Priya Doss; B., Rajith; Chakraborty, Chiranjib; N., NagaSundaram; Ali, Shabana Kouser; Zhu, Hailong

2014-01-01

Some individuals with non-small-cell lung cancer (NSCLC) benefit from therapies targeting epidermal growth factor receptor (EGFR), and the characterization of a new mechanism of resistance to the EGFR-specific antibody gefitinib will provide valuable insight into how therapeutic strategies might be designed to overcome this particular resistance mechanism. The G719S and T790M mutations and their combination were involved in causing different conformational redistribution of EGFR. In the present computational study, we analyzed the impact and structural influence of G719S/T790M double mutation (DM) in EGFR with ligand (gefitinib) through molecular dynamic simulation (50 ns) and docking analysis. We observed the escalation in distance between the functional loop and activation loop with respect to T790M mutation compared to the G719S mutation. Furthermore, we confirmed that the G719S mutation causes the ligand to move closer to the hinge region, whereas T790M makes the ligand escape from the binding pocket. Obtained results provide with an explanation for the resistance induced by T790M and a vital clue for the design of drugs to combat gefitinib resistance. PMID:25091415

Recurrent papillary craniopharyngioma with BRAFV600E mutation treated with neoadjuvant-targeted therapy.

PubMed

Rostami, Elham; Witt Nyström, Petra; Libard, Sylwia; Wikström, Johan; Casar-Borota, Olivera; Gudjonsson, Olafur

2017-11-01

Craniopharyngiomas are histologically benign but locally aggressive tumors in the sellar region that may cause devastating neurological and endocrine deficits. They tend to recur following surgery with high morbidity; hence, postoperative radiotherapy is recommended following sub-total resection. BRAFV600E mutation is the principal oncogenic driver in the papillary variant of craniopharyngiomas. Recently, a dramatic tumor reduction has been reported in a patient with BRAFV600E mutated, multiply recurrent papillary craniopharyngioma using a combination therapy of BRAF inhibitor dabrafenib and MEK inhibitor trametinib. Here, we report on near-radical reduction of a growing residual BRAFV600E craniopharyngioma using the same neoadjuvant therapy.

A novel human autoimmune syndrome caused by combined hypomorphic and activating mutations in ZAP-70

PubMed Central

Chan, Alice Y.; Punwani, Divya; Kadlecek, Theresa A.; Cowan, Morton J.; Olson, Jean L.; Mathes, Erin F.; Sunderam, Uma; Man Fu, Shu; Srinivasan, Rajgopal; Kuriyan, John; Brenner, Steven E.; Weiss, Arthur

2016-01-01

A brother and sister developed a previously undescribed constellation of autoimmune manifestations within their first year of life, with uncontrollable bullous pemphigoid, colitis, and proteinuria. The boy had hemophilia due to a factor VIII autoantibody and nephrotic syndrome. Both children required allogeneic hematopoietic cell transplantation (HCT), which resolved their autoimmunity. The early onset, severity, and distinctive findings suggested a single gene disorder underlying the phenotype. Whole-exome sequencing performed on five family members revealed the affected siblings to be compound heterozygous for two unique missense mutations in the 70-kD T cell receptor ζ-chain associated protein (ZAP-70). Healthy relatives were heterozygous mutation carriers. Although pre-HCT patient T cells were not available, mutation effects were determined using transfected cell lines and peripheral blood from carriers and controls. Mutation R192W in the C-SH2 domain exhibited reduced binding to phosphorylated ζ-chain, whereas mutation R360P in the N lobe of the catalytic domain disrupted an autoinhibitory mechanism, producing a weakly hyperactive ZAP-70 protein. Although human ZAP-70 deficiency can have dysregulated T cells, and autoreactive mouse thymocytes with weak Zap-70 signaling can escape tolerance, our patients’ combination of hypomorphic and activating mutations suggested a new disease mechanism and produced previously undescribed human ZAP-70–associated autoimmune disease. PMID:26783323

Mutation Spectrum of the ABCA4 Gene in a Greek Cohort with Stargardt Disease: Identification of Novel Mutations and Evidence of Three Prevalent Mutated Alleles

PubMed Central

Vassiliki, Kokkinou; George, Koutsodontis; Polixeni, Stamatiou; Christoforos, Giatzakis; Minas, Aslanides Ioannis; Stavrenia, Koukoula; Ioannis, Datseris

2018-01-01

Aim To evaluate the frequency and pattern of disease-associated mutations of ABCA4 gene among Greek patients with presumed Stargardt disease (STGD1). Materials and Methods A total of 59 patients were analyzed for ABCA4 mutations using the ABCR400 microarray and PCR-based sequencing of all coding exons and flanking intronic regions. MLPA analysis as well as sequencing of two regions in introns 30 and 36 reported earlier to harbor deep intronic disease-associated variants was used in 4 selected cases. Results An overall detection rate of at least one mutant allele was achieved in 52 of the 59 patients (88.1%). Direct sequencing improved significantly the complete characterization rate, that is, identification of two mutations compared to the microarray analysis (93.1% versus 50%). In total, 40 distinct potentially disease-causing variants of the ABCA4 gene were detected, including six previously unreported potentially pathogenic variants. Among the disease-causing variants, in this cohort, the most frequent was c.5714+5G>A representing 16.1%, while p.Gly1961Glu and p.Leu541Pro represented 15.2% and 8.5%, respectively. Conclusions By using a combination of methods, we completely molecularly diagnosed 48 of the 59 patients studied. In addition, we identified six previously unreported, potentially pathogenic ABCA4 mutations. PMID:29854428

Next-generation sequencing identifies a novel compound heterozygous mutation in MYO7A in a Chinese patient with Usher Syndrome 1B.

PubMed

Wei, Xiaoming; Sun, Yan; Xie, Jiansheng; Shi, Quan; Qu, Ning; Yang, Guanghui; Cai, Jun; Yang, Yi; Liang, Yu; Wang, Wei; Yi, Xin

2012-11-20

Targeted enrichment and next-generation sequencing (NGS) have been employed for detection of genetic diseases. The purpose of this study was to validate the accuracy and sensitivity of our method for comprehensive mutation detection of hereditary hearing loss, and identify inherited mutations involved in human deafness accurately and economically. To make genetic diagnosis of hereditary hearing loss simple and timesaving, we designed a 0.60 MB array-based chip containing 69 nuclear genes and mitochondrial genome responsible for human deafness and conducted NGS toward ten patients with five known mutations and a Chinese family with hearing loss (never genetically investigated). Ten patients with five known mutations were sequenced using next-generation sequencing to validate the sensitivity of the method. We identified four known mutations in two nuclear deafness causing genes (GJB2 and SLC26A4), one in mitochondrial DNA. We then performed this method to analyze the variants in a Chinese family with hearing loss and identified compound heterozygosity for two novel mutations in gene MYO7A. The compound heterozygosity identified in gene MYO7A causes Usher Syndrome 1B with severe phenotypes. The results support that the combination of enrichment of targeted genes and next-generation sequencing is a valuable molecular diagnostic tool for hereditary deafness and suitable for clinical application. Copyright © 2012 Elsevier B.V. All rights reserved.

Neurofibromatosis-Noonan Syndrome: A Possible Paradigm of the Combination of Genetic and Epigenetic Factors.

PubMed

Yapijakis, Christos; Pachis, Nikos; Voumvourakis, Costas

2017-01-01

Neurofibromatosis-Noonan syndrome (NFNS) is a clinical entity possessing traits of autosomal dominant disorders neurofibromatosis type 1 (NF1) and Noonan syndrome (NS). Germline mutations that disrupt the RAS/MAPK pathway are involved in the pathogenesis of both NS and NF1. In light of a studied Greek family, a new theory for etiological pathogenesis of NFNS is suggested. The NFNS phenotype may be the final result of a combination of a genetic factor (a mutation in the NF1 gene) and an environmental factor with the epigenetic effects of muscle hypotonia (such as hydantoin in the reported Greek family), causing hypoplasia of the face and micrognathia.

Left-dominant arrhythmogenic cardiomyopathy in a large family: associated desmosomal or nondesmosomal genotype?

PubMed

Groeneweg, Judith A; van der Zwaag, Paul A; Jongbloed, Jan D H; Cox, Moniek G P J; Vreeker, Arnold; de Boer, Rudolf A; van der Heijden, Jeroen F; van Veen, Toon A B; McKenna, William J; van Tintelen, J Peter; Dooijes, Dennis; Hauer, Richard N W

2013-04-01

Arrhythmogenic cardiomyopathy (AC) is considered a predominantly right ventricular (RV) desmosomal disease. However, left-dominant forms due to desmosomal gene mutations, including PKP2 variant c.419C>T, have been described. Recently, a nondesmosomal phospholamban (PLN) mutation (c.40_42delAGA) has been identified, causing dilated cardiomyopathy and arrhythmias. To gain more insight into pathogenicity of the PKP2 variant c.419C>T by cosegregation analysis of the PKP2 variant c.419C>T vs the PLN mutation c.40_42delAGA. A Dutch family (13 family members, median age 49 years, range 34-71 years) with ventricular tachycardia underwent (1) meticulous phenotypic characterization and (2) screening of 5 desmosomal genes (PKP2, DSC2, DSG2, DSP, JUP) and PLN. Six family members fulfilled 2010 AC Task Force Criteria. Seven had signs of left ventricular (LV) involvement (inverted T waves in leads V4-V6, LV wall motion abnormalities and late enhancement, and reduced LV ejection fraction), including 6 family members with proven AC. The PKP2 variant c.419C>T was found as a single variant in 3 family members, combined with the PLN mutation c.40_42delAGA in 3 others. PLN mutation was found in 9 family members, including the 6 with AC and all 7 with LV involvement. The PLN mutation c.40_42delAGA was found as a single mutation in 6, combined with the PKP2 variant c.419C>T in 3 others. A low-voltage electrocardiogram was seen in 4 of 9 PLN mutation-positive subjects. None of the family members with the single PKP2 variant showed any sign of RV or LV involvement. The PLN mutation c.40_42delAGA cosegregates with AC and with electrocardiographic and structural LV abnormalities. In this family, there was no evidence of disease-causing contribution of the PKP2 variant c.419C>T. Copyright © 2013 Heart Rhythm Society. Published by Elsevier Inc. All rights reserved.

Correction of Hirschsprung-Associated Mutations in Human Induced Pluripotent Stem Cells Via Clustered Regularly Interspaced Short Palindromic Repeats/Cas9, Restores Neural Crest Cell Function.

PubMed

Lai, Frank Pui-Ling; Lau, Sin-Ting; Wong, John Kwong-Leong; Gui, Hongsheng; Wang, Reeson Xu; Zhou, Tingwen; Lai, Wing Hon; Tse, Hung-Fat; Tam, Paul Kwong-Hang; Garcia-Barcelo, Maria-Mercedes; Ngan, Elly Sau-Wai

2017-07-01

Hirschsprung disease is caused by failure of enteric neural crest cells (ENCCs) to fully colonize the bowel, leading to bowel obstruction and megacolon. Heterozygous mutations in the coding region of the RET gene cause a severe form of Hirschsprung disease (total colonic aganglionosis). However, 80% of HSCR patients have short-segment Hirschsprung disease (S-HSCR), which has not been associated with genetic factors. We sought to identify mutations associated with S-HSCR, and used the clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 gene editing system to determine how mutations affect ENCC function. We created induced pluripotent stem cell (iPSC) lines from 1 patient with total colonic aganglionosis (with the G731del mutation in RET) and from 2 patients with S-HSCR (without a RET mutation), as well as RET +/- and RET -/- iPSCs. IMR90-iPSC cells were used as the control cell line. Migration and differentiation capacities of iPSC-derived ENCCs were analyzed in differentiation and migration assays. We searched for mutation(s) associated with S-HSCR by combining genetic and transcriptome data from patient blood- and iPSC-derived ENCCs, respectively. Mutations in the iPSCs were corrected using the CRISPR/Cas9 system. ENCCs derived from all iPSC lines, but not control iPSCs, had defects in migration and neuronal lineage differentiation. RET mutations were associated with differentiation and migration defects of ENCCs in vitro. Genetic and transcriptome analyses associated a mutation in the vinculin gene (VCL M209L) with S-HSCR. CRISPR/Cas9 correction of the RET G731del and VCL M209L mutations in iPSCs restored the differentiation and migration capacities of ENCCs. We identified mutations in VCL associated with S-HSCR. Correction of this mutation in iPSC using CRISPR/Cas9 editing, as well as the RET G731del mutation that causes Hirschsprung disease with total colonic aganglionosis, restored ENCC function. Our study demonstrates how human iPSCs can be used to identify disease-associated mutations and determine how they affect cell functions and contribute to pathogenesis. Copyright © 2017 AGA Institute. Published by Elsevier Inc. All rights reserved.

Non-syndromic hearing loss caused by the dominant cis mutation R75Q with the recessive mutation V37I of the GJB2 (Connexin 26) gene.

PubMed

Kim, Juwon; Jung, Jinsei; Lee, Min Goo; Choi, Jae Young; Lee, Kyung-A

2015-06-19

GJB2 alleles containing two cis mutations have been rarely found in non-syndromic hearing loss. Herein, we present a Korean patient with non-syndromic hearing loss caused by the R75Q cis mutation with V37I, which arose de novo in the father and was inherited by the patient. Biochemical coupling and hemichannel permeability assays were performed after molecular cloning and transfection of HEK293T cells. Student's t-tests or analysis of variance followed by Tukey's multiple comparison test was used as statistical analysis. Biochemical coupling was significantly reduced in connexin 26 (Cx26)-R75Q- and Cx26-V37I-transfected cells, with greater extent in Cx26-R75Q and Cx26-R75Q+V37I cells. Interestingly, our patient and his father with the mutations had more residual hearing compared with patients with the dominant mutation alone. Although the difference in hemichannel activity between R75Q alone and R75Q in combination with V37I failed to reach significance, it is of note that there is a possibility that V37I located upstream of R75Q might have the ability to ameliorate R75Q expression. Our study emphasizes the importance of cis mutations with R75Q, as the gene effect of R75Q can be modulated depending on the type of additional mutation.

Base damage, local sequence context and TP53 mutation hotspots: a molecular dynamics study of benzo[a]pyrene induced DNA distortion and mutability

PubMed Central

Menzies, Georgina E.; Reed, Simon H.; Brancale, Andrea; Lewis, Paul D.

2015-01-01

The mutational pattern for the TP53 tumour suppressor gene in lung tumours differs to other cancer types by having a higher frequency of G:C>T:A transversions. The aetiology of this differing mutation pattern is still unknown. Benzo[a]pyrene,diol epoxide (BPDE) is a potent cigarette smoke carcinogen that forms guanine adducts at TP53 CpG mutation hotspot sites including codons 157, 158, 245, 248 and 273. We performed molecular modelling of BPDE-adducted TP53 duplex sequences to determine the degree of local distortion caused by adducts which could influence the ability of nucleotide excision repair. We show that BPDE adducted codon 157 has greater structural distortion than other TP53 G:C>T:A hotspot sites and that sequence context more distal to adjacent bases must influence local distortion. Using TP53 trinucleotide mutation signatures for lung cancer in smokers and non-smokers we further show that codons 157 and 273 have the highest mutation probability in smokers. Combining this information with adduct structural data we predict that G:C>T:A mutations at codon 157 in lung tumours of smokers are predominantly caused by BPDE. Our results provide insight into how different DNA sequence contexts show variability in DNA distortion at mutagen adduct sites that could compromise DNA repair at well characterized cancer related mutation hotspots. PMID:26400171

Exome Sequencing of 18 Chinese Families with Congenital Cataracts: A New Sight of the NHS Gene

PubMed Central

Sun, Wenmin; Xiao, Xueshan; Li, Shiqiang; Guo, Xiangming; Zhang, Qingjiong

2014-01-01

Purpose The aim of this study was to investigate the mutation spectrum and frequency of 34 known genes in 18 Chinese families with congenital cataracts. Methods Genomic DNA and clinical data was collected from 18 families with congenital cataracts. Variations in 34 cataract-associated genes were screened by whole exome sequencing and then validated by Sanger sequencing. Results Eleven candidate variants in seven of the 34 genes were detected by exome sequencing and then confirmed by Sanger sequencing, including two variants predicted to be benign and the other pathogenic mutations. The nine mutations were present in 9 of the 18 (50%) families with congenital cataracts. Of the four families with mutations in the X-linked NHS gene, no other abnormalities were recorded except for cataract, in which a pseudo-dominant inheritance form was suggested, as female carriers also had different forms of cataracts. Conclusion This study expands the mutation spectrum and frequency of genes responsible for congenital cataract. Mutation in NHS is a common cause of nonsyndromic congenital cataract with pseudo-autosomal dominant inheritance. Combined with our previous studies, a genetic basis could be identified in 67.6% of families with congenital cataracts in our case series, in which mutations in genes encoding crystallins, genes encoding connexins, and NHS are responsible for 29.4%, 14.7%, and 11.8% of families, respectively. Our results suggest that mutations in NHS are the common cause of congenital cataract, both syndromic and nonsyndromic. PMID:24968223

Bioinformatic Analysis of Pathogenic Missense Mutations of Activin Receptor Like Kinase 1 Ectodomain

PubMed Central

Scotti, Claudia; Olivieri, Carla; Boeri, Laura; Canzonieri, Cecilia; Ornati, Federica; Buscarini, Elisabetta; Pagella, Fabio; Danesino, Cesare

2011-01-01

Activin A receptor, type II-like kinase 1 (also called ALK1), is a serine-threonine kinase predominantly expressed on endothelial cells surface. Mutations in its ACVRL1 encoding gene (12q11-14) cause type 2 Hereditary Haemorrhagic Telangiectasia (HHT2), an autosomal dominant multisystem vascular dysplasia. The study of the structural effects of mutations is crucial to understand their pathogenic mechanism. However, while an X-ray structure of ALK1 intracellular domain has recently become available (PDB ID: 3MY0), structure determination of ALK1 ectodomain (ALK1EC) has been elusive so far. We here describe the building of a homology model for ALK1EC, followed by an extensive bioinformatic analysis, based on a set of 38 methods, of the effect of missense mutations at the sequence and structural level. ALK1EC potential interaction mode with its ligand BMP9 was then predicted combining modelling and docking data. The calculated model of the ALK1EC allowed mapping and a preliminary characterization of HHT2 associated mutations. Major structural changes and loss of stability of the protein were predicted for several mutations, while others were found to interfere mainly with binding to BMP9 or other interactors, like Endoglin (CD105), whose encoding ENG gene (9q34) mutations are known to cause type 1 HHT. This study gives a preliminary insight into the potential structure of ALK1EC and into the structural effects of HHT2 associated mutations, which can be useful to predict the potential effect of each single mutation, to devise new biological experiments and to interpret the biological significance of new mutations, private mutations, or non-synonymous polymorphisms. PMID:22028876

Two novel de novo mutations of KRT6A and KRT16 genes in two Chinese pachyonychia congenita pedigrees with fissured tongue or diffuse plantar keratoderma.

PubMed

Du, Zhen-Fang; Xu, Chen-Ming; Zhao, Yan; Liu, Wen-Ting; Chen, Xiao-Ling; Chen, Chun-Yue; Fang, Hong; Ke, Hai-Ping; Zhang, Xian-Ning

2012-01-01

Mutations in the KRT6A or KRT16 gene cause pachyonychia congenita type 1 (PC-1), while mutations in KRT16 or KRT6C underlie focal palmoplantar keratoderma (FPPK). A new classification system of PC has been adopted based on the mutated gene. PC rarely presents the symptoms of diffuse plantar keratoderma. Mutation in the tail domain of keratins is rarely reported. PC combined with fissured tongue has never been described. To investigate the genotype-phenotype correlations between clinical features and gene mutational sites in two unrelated southern Chinese PC pedigrees (one family presented with specific fissured tongue, the other with diffuse plantar keratoderma). The whole coding regions of the KRT6A/KRT16/KRT17/KRT6B genes were amplified and directly sequenced to detect the mutation. To confirm the effect of the IVS8-2A>C mutation in KRT6A at the mRNA level, total RNA from the plantar lesion of a patient was extracted and reverse-transcribed to cDNA for sequence analysis. Two novel de novo mutations, a splice acceptor site variant IVS8-2A>C (p.S487FfsX72) in KRT6A and a heterozygous substitution c.AA373_374GG (p.N125G) within exon 1 of KRT16, were found separately in the two PC families. Genotype-phenotype correlations among PC patients with codon-125 mutation in KRT16 were established, while the phenotypes caused by the IVS8-2A>C mutation in KRT6A need further studies to confirm the rare feature of fissured tongue.

Mutations in GDP-mannose pyrophosphorylase B cause congenital and limb-girdle muscular dystrophies associated with hypoglycosylation of α-dystroglycan.

PubMed

Carss, Keren J; Stevens, Elizabeth; Foley, A Reghan; Cirak, Sebahattin; Riemersma, Moniek; Torelli, Silvia; Hoischen, Alexander; Willer, Tobias; van Scherpenzeel, Monique; Moore, Steven A; Messina, Sonia; Bertini, Enrico; Bönnemann, Carsten G; Abdenur, Jose E; Grosmann, Carla M; Kesari, Akanchha; Punetha, Jaya; Quinlivan, Ros; Waddell, Leigh B; Young, Helen K; Wraige, Elizabeth; Yau, Shu; Brodd, Lina; Feng, Lucy; Sewry, Caroline; MacArthur, Daniel G; North, Kathryn N; Hoffman, Eric; Stemple, Derek L; Hurles, Matthew E; van Bokhoven, Hans; Campbell, Kevin P; Lefeber, Dirk J; Lin, Yung-Yao; Muntoni, Francesco

2013-07-11

Congenital muscular dystrophies with hypoglycosylation of α-dystroglycan (α-DG) are a heterogeneous group of disorders often associated with brain and eye defects in addition to muscular dystrophy. Causative variants in 14 genes thought to be involved in the glycosylation of α-DG have been identified thus far. Allelic mutations in these genes might also cause milder limb-girdle muscular dystrophy phenotypes. Using a combination of exome and Sanger sequencing in eight unrelated individuals, we present evidence that mutations in guanosine diphosphate mannose (GDP-mannose) pyrophosphorylase B (GMPPB) can result in muscular dystrophy variants with hypoglycosylated α-DG. GMPPB catalyzes the formation of GDP-mannose from GTP and mannose-1-phosphate. GDP-mannose is required for O-mannosylation of proteins, including α-DG, and it is the substrate of cytosolic mannosyltransferases. We found reduced α-DG glycosylation in the muscle biopsies of affected individuals and in available fibroblasts. Overexpression of wild-type GMPPB in fibroblasts from an affected individual partially restored glycosylation of α-DG. Whereas wild-type GMPPB localized to the cytoplasm, five of the identified missense mutations caused formation of aggregates in the cytoplasm or near membrane protrusions. Additionally, knockdown of the GMPPB ortholog in zebrafish caused structural muscle defects with decreased motility, eye abnormalities, and reduced glycosylation of α-DG. Together, these data indicate that GMPPB mutations are responsible for congenital and limb-girdle muscular dystrophies with hypoglycosylation of α-DG. Copyright © 2013 The American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.

Compared effects of missense mutations in Very-Long-Chain Acyl-CoA Dehydrogenase deficiency: Combined analysis by structural, functional and pharmacological approaches.

PubMed

Gobin-Limballe, Stéphanie; McAndrew, Ryan P; Djouadi, Fatima; Kim, Jung-Ja; Bastin, Jean

2010-05-01

Very-Long-Chain Acyl-CoA Dehydrogenase deficiency (VLCADD) is an autosomal recessive disorder considered as one of the more common ss-oxidation defects, possibly associated with neonatal cardiomyopathy, infantile hepatic coma, or adult-onset myopathy. Numerous gene missense mutations have been described in these VLCADD phenotypes, but only few of them have been structurally and functionally analyzed, and the molecular basis of disease variability is still poorly understood. To address this question, we first analyzed fourteen disease-causing amino acid changes using the recently described crystal structure of VLCAD. The predicted effects varied from the replacement of amino acid residues lining the substrate binding cavity, involved in holoenzyme-FAD interactions or in enzyme dimerisation, predicted to have severe functional consequences, up to amino acid substitutions outside key enzyme domains or lying on near enzyme surface, with predicted milder consequences. These data were combined with functional analysis of residual fatty acid oxidation (FAO) and VLCAD protein levels in patient cells harboring these mutations, before and after pharmacological stimulation by bezafibrate. Mutations identified as detrimental to the protein structure in the 3-D model were generally associated to profound FAO and VLCAD protein deficiencies in the patient cells, however, some mutations affecting FAD binding or monomer-monomer interactions allowed a partial response to bezafibrate. On the other hand, bezafibrate restored near-normal FAO rates in some mutations predicted to have milder consequences on enzyme structure. Overall, combination of structural, biochemical, and pharmacological analysis allowed assessment of the relative severity of individual mutations, with possible applications for disease management and therapeutic approach. Copyright 2010 Elsevier B.V. All rights reserved.

Two novel mutations in the POU1F1 gene generate null alleles through different mechanisms leading to combined pituitary hormone deficiency.

PubMed

Turton, J P; Strom, M; Langham, S; Dattani, M T; Le Tissier, P

2012-03-01

  Mutations in the POU1F1 gene severely affect the development and function of the anterior pituitary gland and lead to combined pituitary hormone deficiency (CPHD).   The clinical and genetic analysis of a patient presenting with CPHD and functional characterization of identified mutations.   We describe a male patient with extreme short stature, learning difficulties, anterior pituitary hypoplasia, secondary hypothyroidism and undetectable prolactin, growth hormone (GH) and insulin-like growth factor 1 (IGF1), with normal random cortisol.   The POU1F1 coding region was amplified by PCR and sequenced; the functional consequence of the mutations was analysed by cell transfection and in vitro assays.   Genetic analysis revealed compound heterozygosity for two novel putative loss of function mutations in POU1F1: a transition at position +3 of intron 1 [IVS1+3nt(A>G)] and a point mutation in exon 6 resulting in a substitution of arginine by tryptophan (R265W). Functional analysis revealed that IVS1+3nt(A>G) results in a reduction in the correctly spliced POU1F1 mRNA, which could be corrected by mutations of the +4, +5 and +6 nucleotides. Analysis of POU1F1(R265W) revealed complete loss of function resulting from severely reduced protein stability.   Combined pituitary hormone deficiency in this patient is caused by loss of POU1F1 function by two novel mechanisms, namely aberrant splicing (IVS1+3nt (A>G) and protein instability (R265W). Identification of the genetic basis of CPHD enabled the cessation of hydrocortisone therapy without the need for further assessment for evolving endocrinopathy. © 2012 Blackwell Publishing Ltd.

Amyloid precursor protein-induced axonopathies are independent of amyloid-beta peptides.

PubMed

Stokin, Gorazd B; Almenar-Queralt, Angels; Gunawardena, Shermali; Rodrigues, Elizabeth M; Falzone, Tomás; Kim, Jungsu; Lillo, Concepción; Mount, Stephanie L; Roberts, Elizabeth A; McGowan, Eileen; Williams, David S; Goldstein, Lawrence S B

2008-11-15

Overexpression of amyloid precursor protein (APP), as well as mutations in the APP and presenilin genes, causes rare forms of Alzheimer's disease (AD). These genetic changes have been proposed to cause AD by elevating levels of amyloid-beta peptides (Abeta), which are thought to be neurotoxic. Since overexpression of APP also causes defects in axonal transport, we tested whether defects in axonal transport were the result of Abeta poisoning of the axonal transport machinery. Because directly varying APP levels also alters APP domains in addition to Abeta, we perturbed Abeta generation selectively by combining APP transgenes in Drosophila and mice with presenilin-1 (PS1) transgenes harboring mutations that cause familial AD (FAD). We found that combining FAD mutant PS1 with FAD mutant APP increased Abeta42/Abeta40 ratios and enhanced amyloid deposition as previously reported. Surprisingly, however, this combination suppressed rather than increased APP-induced axonal transport defects in both Drosophila and mice. In addition, neuronal apoptosis induced by expression of FAD mutant human APP in Drosophila was suppressed by co-expressing FAD mutant PS1. We also observed that directly elevating Abeta with fusions to the Familial British and Danish Dementia-related BRI protein did not enhance axonal transport phenotypes in APP transgenic mice. Finally, we observed that perturbing Abeta ratios in the mouse by combining FAD mutant PS1 with FAD mutant APP did not enhance APP-induced behavioral defects. A potential mechanism to explain these findings was suggested by direct analysis of axonal transport in the mouse, which revealed that axonal transport or entry of APP into axons is reduced by FAD mutant PS1. Thus, we suggest that APP-induced axonal defects are not caused by Abeta.

Autosomal dominant polycystic kidney disease in a family with mosaicism and hypomorphic allele.

PubMed

Reiterová, Jana; Štekrová, Jitka; Merta, Miroslav; Kotlas, Jaroslav; Elišáková, Veronika; Lněnička, Petr; Korabečná, Marie; Kohoutová, Milada; Tesař, Vladimír

2013-03-15

Autosomal dominant polycystic kidney disease (ADPKD) is the most common form of inherited kidney disease that results in renal failure. ADPKD is a systemic disorder with cysts and connective tissue abnormalities involving many organs. ADPKD caused by mutations in PKD1 gene is significantly more severe than the cases caused by PKD2 gene mutations. The large intra-familial variability of ADPKD highlights a role for genetic background. Here we report a case of ADPKD family initially appearing unlinked to the PKD1 or PKD2 loci and the influence of mosaicism and hypomorphic allele on the variability of the clinical course of the disease. A grandmother with the PKD1 gene mutation in mosaicism (p.Val1105ArgfsX4) and with mild clinical course of ADPKD (end stage renal failure at the age of 77) seemed to have ADPKD because of PKD2 gene mutation. On the other hand, her grandson had a severe clinical course (end stage renal disease at the age of 45) in spite of the early treatment of mild hypertension. There was found by mutational analysis of PKD genes that the severe clinical course was caused by PKD1 gene frameshifting mutation inherited from his father and mildly affected grandmother in combination with inherited hypomorphic PKD1 allele with described missense mutation (p.Thr2250Met) from his clinically healthy mother. The sister with two cysts and with PKD1 hypomorphic allele became the kidney donor to her severely affected brother. We present the first case of ADPKD with the influence of mosaicism and hypomorphic allele of the PKD1 gene on clinical course of ADPKD in one family. Moreover, this report illustrates the role of molecular genetic testing in assessing young related kidney donors for patients with ADPKD.

A Novel Rasopathy Caused by Recurrent De Novo Missense Mutations In PPP1CB Closely Resembles Noonan Syndrome with Loose Anagen Hair

PubMed Central

Gripp, Karen W.; Aldinger, Kimberly A.; Bennett, James T.; Baker, Laura; Tusi, Jessica; Powell-Hamilton, Nina; Stabley, Deborah; Sol-Church, Katia; Timms, Andrew E.; Dobyns, William B.

2016-01-01

Noonan syndrome is a rasopathy caused by mutations in multiple genes encoding components of the RAS/MAPK pathway. Despite its variable phenotype, limited genotype-phenotype correlations exist. Noonan syndrome with loose anagen hair (NS-LAH) is characterized by its distinctive hair anomalies, developmental differences and structural brain abnormalities and is caused by a single recurrent missense SHOC2 mutation. SHOC2 forms a complex with protein phosphatase 1 (PP1C). Protein phosphatases counterbalance kinases and control activation of signaling proteins, such as the mitogen activated protein kinases of the RAS/MAPK pathway. Here we report four patients with de novo missense mutations in protein phosphatase 1 catalytic subunit beta (PPP1CB), sharing a recognizable phenotype. Three individuals had the recurrent PPP1CB c.146G>C, p.Pro49Arg mutation, the fourth had a c.166G>C, p.Ala56Pro change. All had relative or absolute macrocephaly, low-set and posteriorly angulated ears and developmental delay. Slow growing and/or sparse hair and/or an unruly hair texture was present in all. Three individuals had feeding difficulties requiring feeding tubes. One of two males had cryptorchidism, another had pectus excavatum. Short stature was present in three. A female with the recurrent mutation had a Dandy-Walker malformation and optic nerve hypoplasia. Mild ventriculomegaly occurred in all, cerebellar tonsillar ectopia was seen in two and progressed to Chiari 1 malformation in one individual. Based on the combination of phenotypic findings and PPP1CB’s effect on RAF dephosphorylation within the RAS/MAPK pathway, this novel condition can be considered a rasopathy, most similar to NS-LAH. Collectively, these mutations meet the standardized criteria for pathogenicity. PMID:27264673

A Recurrent Mutation in CACNA1G Alters Cav3.1 T-Type Calcium-Channel Conduction and Causes Autosomal-Dominant Cerebellar Ataxia

PubMed Central

Coutelier, Marie; Blesneac, Iulia; Monteil, Arnaud; Monin, Marie-Lorraine; Ando, Kunie; Mundwiller, Emeline; Brusco, Alfredo; Le Ber, Isabelle; Anheim, Mathieu; Castrioto, Anna; Duyckaerts, Charles; Brice, Alexis; Durr, Alexandra; Lory, Philippe; Stevanin, Giovanni

2015-01-01

Hereditary cerebellar ataxias (CAs) are neurodegenerative disorders clinically characterized by a cerebellar syndrome, often accompanied by other neurological or non-neurological signs. All transmission modes have been described. In autosomal-dominant CA (ADCA), mutations in more than 30 genes are implicated, but the molecular diagnosis remains unknown in about 40% of cases. Implication of ion channels has long been an ongoing topic in the genetics of CA, and mutations in several channel genes have been recently connected to ADCA. In a large family affected by ADCA and mild pyramidal signs, we searched for the causative variant by combining linkage analysis and whole-exome sequencing. In CACNA1G, we identified a c.5144G>A mutation, causing an arginine-to-histidine (p.Arg1715His) change in the voltage sensor S4 segment of the T-type channel protein Cav3.1. Two out of 479 index subjects screened subsequently harbored the same mutation. We performed electrophysiological experiments in HEK293T cells to compare the properties of the p.Arg1715His and wild-type Cav3.1 channels. The current-voltage and the steady-state activation curves of the p.Arg1715His channel were shifted positively, whereas the inactivation curve had a higher slope factor. Computer modeling in deep cerebellar nuclei (DCN) neurons suggested that the mutation results in decreased neuronal excitability. Taken together, these data establish CACNA1G, which is highly expressed in the cerebellum, as a gene whose mutations can cause ADCA. This is consistent with the neuropathological examination, which showed severe Purkinje cell loss. Our study further extends our knowledge of the link between calcium channelopathies and CAs. PMID:26456284

Presence of novel compound BCR-ABL mutations in late chronic and advanced phase imatinib sensitive CML patients indicates their possible role in CML progression.

PubMed

Akram, Afia Muhammad; Iqbal, Zafar; Akhtar, Tanveer; Khalid, Ahmed Mukhtar; Sabar, Muhammad Farooq; Qazi, Mahmood Hussain; Aziz, Zeba; Sajid, Nadia; Aleem, Aamer; Rasool, Mahmood; Asif, Muhammad; Aloraibi, Saleh; Aljamaan, Khaled; Iqbal, Mudassar

2017-04-03

BCR-ABL kinase domain (K D ) mutations are well known for causing resistance against tyrosine kinase inhibitors (TKIs) and disease progression in chronic myeloid leukemia (CML). In recent years, compound BCR-ABL mutations have emerged as a new threat to CML patients by causing higher degrees of resistance involving multiple TKIs, including ponatinib. However, there are limited reports about association of compound BCR-ABL mutations with disease progression in imatinib (IM) sensitive CML patients. Therefore, we investigated presence of ABL-K D mutations in chronic phase (n = 41), late chronic phase (n = 33) and accelerated phase (n = 16) imatinib responders. Direct sequencing analysis was used for this purpose. Eleven patients (12.22%) in late-CP CML were detected having total 24 types of point mutations, out of which 8 (72.72%) harbored compound mutated sites. SH2 contact site mutations were dominant in our study cohort, with E355G (3.33%) being the most prevalent. Five patients (45%) all having compound mutated sites, progressed to advanced phases of disease during follow up studies. Two novel silent mutations G208G and E292E/E were detected in combination with other mutants, indicating limited tolerance for BCR-ABL1 kinase domain for missense mutations. However, no patient in early CP of disease manifested mutated ABL-K D . Occurrence of mutations was found associated with elevated platelet count (p = 0.037) and patients of male sex (p = 0.049). The median overall survival and event free survival of CML patients (n = 90) was 6.98 and 5.8 y respectively. The compound missense mutations in BCR-ABL kinase domain responsible to elicit disease progression, drug resistance or disease relapse in CML, can be present in yet Imatinib sensitive patients. Disease progression observed here, emphasizes the need of ABL-K D mutation screening in late chronic phase CML patients for improved clinical management of disease.

Accurate calculation of mutational effects on the thermodynamics of inhibitor binding to p38α MAP kinase: a combined computational and experimental study.

PubMed

Zhu, Shun; Travis, Sue M; Elcock, Adrian H

2013-07-09

A major current challenge for drug design efforts focused on protein kinases is the development of drug resistance caused by spontaneous mutations in the kinase catalytic domain. The ubiquity of this problem means that it would be advantageous to develop fast, effective computational methods that could be used to determine the effects of potential resistance-causing mutations before they arise in a clinical setting. With this long-term goal in mind, we have conducted a combined experimental and computational study of the thermodynamic effects of active-site mutations on a well-characterized and high-affinity interaction between a protein kinase and a small-molecule inhibitor. Specifically, we developed a fluorescence-based assay to measure the binding free energy of the small-molecule inhibitor, SB203580, to the p38α MAP kinase and used it measure the inhibitor's affinity for five different kinase mutants involving two residues (Val38 and Ala51) that contact the inhibitor in the crystal structure of the inhibitor-kinase complex. We then conducted long, explicit-solvent thermodynamic integration (TI) simulations in an attempt to reproduce the experimental relative binding affinities of the inhibitor for the five mutants; in total, a combined simulation time of 18.5 μs was obtained. Two widely used force fields - OPLS-AA/L and Amber ff99SB-ILDN - were tested in the TI simulations. Both force fields produced excellent agreement with experiment for three of the five mutants; simulations performed with the OPLS-AA/L force field, however, produced qualitatively incorrect results for the constructs that contained an A51V mutation. Interestingly, the discrepancies with the OPLS-AA/L force field could be rectified by the imposition of position restraints on the atoms of the protein backbone and the inhibitor without destroying the agreement for other mutations; the ability to reproduce experiment depended, however, upon the strength of the restraints' force constant. Imposition of position restraints in corresponding simulations that used the Amber ff99SB-ILDN force field had little effect on their ability to match experiment. Overall, the study shows that both force fields can work well for predicting the effects of active-site mutations on small molecule binding affinities and demonstrates how a direct combination of experiment and computation can be a powerful strategy for developing an understanding of protein-inhibitor interactions.

Combination of icotinib, surgery, and internal-radiotherapy of a patient with lung cancer severely metastasized to the vertebrae bones with EGFR mutation: a case report.

PubMed

Qu, Li-Li; Qin, Hai-Feng; Gao, Hong-Jun; Liu, Xiao-Qing

2015-01-01

A 48-year-old Chinese female was referred to us regarding EGFR-mutated advanced non-small cell lung cancer, and metastasis to left scapula and vertebrae bones which caused pathological fracture at T8 and T10 thoracic vertebrae. An aggressive combined therapy with icotinib, vertebrae operation, and radioactive particle implantation and immunotherapy was proposed to prevent paraplegia, relieve pain, and control the overall and local tumor lesions. No postoperative symptoms were seen after surgery, and the pain was significantly relieved. Icotinib merited a 31-month partial response with grade 1 diarrhea as its drug-related adverse event. High dose of icotinib was administered after pelvis lesion progression for 3 months with good tolerance. Combination therapy of icotinib, surgery, and internal radiation for metastases of the vertebrae bones from non-small cell lung cancer seems to be a very promising technique both for sufficient pain relief and for local control of the tumor, vertebrae operation can be an encouraging option for patients with EFGR positive mutation and good prognosis indicator.

Optimising the combination dosing strategy of abemaciclib and vemurafenib in BRAF-mutated melanoma xenograft tumours.

PubMed

Tate, Sonya C; Burke, Teresa F; Hartman, Daisy; Kulanthaivel, Palaniappan; Beckmann, Richard P; Cronier, Damien M

2016-03-15

Resistance to BRAF inhibition is a major cause of treatment failure for BRAF-mutated metastatic melanoma patients. Abemaciclib, a cyclin-dependent kinase 4 and 6 inhibitor, overcomes this resistance in xenograft tumours and offers a promising drug combination. The present work aims to characterise the quantitative pharmacology of the abemaciclib/vemurafenib combination using a semimechanistic pharmacokinetic/pharmacodynamic modelling approach and to identify an optimum dosing regimen for potential clinical evaluation. A PK/biomarker model was developed to connect abemaciclib/vemurafenib concentrations to changes in MAPK and cell cycle pathway biomarkers in A375 BRAF-mutated melanoma xenografts. Resultant tumour growth inhibition was described by relating (i) MAPK pathway inhibition to apoptosis, (ii) mitotic cell density to tumour growth and, under resistant conditions, (iii) retinoblastoma protein inhibition to cell survival. The model successfully described vemurafenib/abemaciclib-mediated changes in MAPK pathway and cell cycle biomarkers. Initial tumour shrinkage by vemurafenib, acquisition of resistance and subsequent abemaciclib-mediated efficacy were successfully captured and externally validated. Model simulations illustrate the benefit of intermittent vemurafenib therapy over continuous treatment, and indicate that continuous abemaciclib in combination with intermittent vemurafenib offers the potential for considerable tumour regression. The quantitative pharmacology of the abemaciclib/vemurafenib combination was successfully characterised and an optimised, clinically-relevant dosing strategy was identified.

Evaluation of genotoxic effects caused by extracts of chlorinated drinking water using a combination of three different bioassays.

PubMed

Zeng, Qiang; Zhang, Shao-Hui; Liao, Jing; Miao, Dong-Yue; Wang, Xin-Yi; Yang, Pan; Yun, Luo-Jia; Liu, Ai-Lin; Lu, Wen-Qing

2015-10-15

Potential genotoxic effects of chlorinated drinking water now are of a great concern. In this study, raw water, finished water, and tap water from a water plant in Wuhan, China were collected in two different sampling times of the year (January and July). Genotoxic effects of water extracts were evaluated using a combination of three different bioassays: SOS/umu test, HGPRT gene mutation assay, and micronucleus assay, which were separately used to detect DNA damage, gene mutation, and chromosome aberration. The results of three different bioassays showed that all water samples in January and July induced at least one types of genotoxic effects, of which the DNA-damage effects were all detectable. The levels of DNA-damage effects and gene-mutation effects of finished water and tap water in January were higher than those in July. Chlorination could increase the DNA-damage effects of drinking water in January and the gene-mutation effects of drinking water in both January and July, but did not increase the chromosome-aberration effects of drinking water in both January and July. Our results highlighted the importance of using a combination of different bioassays to evaluate the genotoxicity of water samples in different seasons. Copyright © 2015 Elsevier B.V. All rights reserved.

Combinations of chlorocatechols and heavy metals cause DNA degradation in vitro but must not result in increased mutation rates in vivo.

PubMed

Schweigert, N; Belkin, S; Leong-Morgenthaler, P; Zehnder, A J; Eggen, R I

1999-01-01

Chlorocatechols introduced into the environment directly or as a result of degradation processes are highly toxic, particularly when combined with heavy metals. With in vitro DNA degradation assays, the high reactivity of chlorocatechols combined with heavy metals could be shown, whereby copper was shown to be more active than iron. Structure-activity analysis showed that the degradation potential of the chlorocatechols decreased with an increasing number of chloratoms. The addition of reactive oxygen species scavengers allowed the identification of hydrogen peroxide as an important agent leading to DNA damage in this reaction. The potential of other reactive compounds, however, can neither be determined nor excluded with this approach. Exposure of Escherichia coli and Salmonella typhimurium cultures to the same mixtures of chlorocatechols and copper surprisingly did not lead to an enhanced mutation rate. This phenomenon was explained by doing marker gene expression measurements and toxicity tests with E. coli mutants deficient in oxidative stress defense or DNA repair. In catechol-copper-exposed cultures an increased peroxide level could indeed be demonstrated, but the highly efficient defense and repair systems of E. coli avoid the phenotypical establishment of mutations. Increased mutation rates under chronic exposure, however, cannot be excluded.

A double mutation in exon 6 of the [beta]-hexosaminidase [alpha] subunit in a patient with the B1 variant of Tay-Sachs disease

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ainsworth, P.J.; Coulter-Mackie, M.B.

1992-10-01

The B1 variant form of Tay-Sachs disease is enzymologically unique in that the causative mutation(s) appear to affect the active site in the [alpha] subunit of [beta]-hexosaminidase A without altering its ability to associate with the [beta] subunit. Most previously reported B1 variant mutations were found in exon 5 within codon 178. The coding sequence of the [alpha] subunit gene of a patient with the B1 variant form was examined with a combination of reverse transcription of mRNA to cDNA, PCR, and dideoxy sequencing. A double mutation in exon 6 has been identified: a G[sub 574][yields]C transversion causing a val[submore » 192][yields]leu change and a G[sub 598][yields] A transition resulting in a val[sub 200][yields]met alteration. The amplified cDNAs were otherwise normal throughout their sequence. The 574 and 598 alterations have been confirmed by amplification directly from genomic DNA from the patient and her mother. Transient-expression studies of the two exon 6 mutations (singly or together) in COS-1 cells show that the G[sub 574][yields]C change is sufficient to cause the loss of enzyme activity. The biochemical phenotype of the 574 alteration in transfection studies is consistent with that expected for a B1 variant mutation. As such, this mutation differs from previously reported B1 variant mutations, all of which occur in exon 5. 31 refs., 2 figs., 2 tabs.« less

Contribution of SHANK3 Mutations to Autism Spectrum Disorder

PubMed Central

Moessner, Rainald ; Marshall, Christian R. ; Sutcliffe, James S. ; Skaug, Jennifer ; Pinto, Dalila ; Vincent, John ; Zwaigenbaum, Lonnie ; Fernandez, Bridget ; Roberts, Wendy ; Szatmari, Peter ; Scherer, Stephen W. 

2007-01-01

Mutations in SHANK3, which encodes a synaptic scaffolding protein, have been described in subjects with an autism spectrum disorder (ASD). To assess the quantitative contribution of SHANK3 to the pathogenesis of autism, we determined the frequency of DNA sequence and copy-number variants in this gene in 400 ASD-affected subjects ascertained in Canada. One de novo mutation and two gene deletions were discovered, indicating a contribution of 0.75% in this cohort. One additional SHANK3 deletion was characterized in two ASD-affected siblings from another collection, which brings the total number of published mutations in unrelated ASD-affected families to seven. The combined data provide support that haploinsufficiency of SHANK3 can cause a monogenic form of autism in sufficient frequency to warrant consideration in clinical diagnostic testing. PMID:17999366

Constitutional abnormalities of IDH1 combined with secondary mutations predispose a patient with Maffucci syndrome to acute lymphoblastic leukemia.

PubMed

Hirabayashi, Shinsuke; Seki, Masafumi; Hasegawa, Daisuke; Kato, Motohiro; Hyakuna, Nobuyuki; Shuo, Takuya; Kimura, Shunsuke; Yoshida, Kenichi; Kataoka, Keisuke; Fujii, Yoichi; Shiraishi, Yuichi; Chiba, Kenichi; Tanaka, Hiroko; Kiyokawa, Nobutaka; Miyano, Satoru; Ogawa, Seishi; Takita, Junko; Manabe, Atsushi

2017-12-01

Maffucci syndrome is a nonhereditary disorder caused by somatic mosaic isocitrate dehydrogenase 1 or 2 (IDH1 or IDH2) mutations and is characterized by multiple enchondromas along with hemangiomas. Malignant transformation of enchondromas to chondrosarcomas and secondary neoplasms, such as brain tumors or acute myeloid leukemia, are serious complications. A 15-year-old female with Maffucci syndrome developed B-cell precursor acute lymphoblastic leukemia (BCP-ALL). A somatic mutation in IDH1 was detected in hemangioma and leukemic cells. KRAS mutation and deletion of IKZF1 were detected in leukemic cells. Patients with Maffucci syndrome may, therefore, be at risk of BCP-ALL associated with secondary genetic events that affect lymphocyte differentiation. © 2017 Wiley Periodicals, Inc.

Targeting mutant fibroblast growth factor receptors in cancer.

PubMed

Greulich, Heidi; Pollock, Pamela M

2011-05-01

Fibroblast growth factor receptors (FGFRs) play diverse roles in the control of cell proliferation, cell differentiation, angiogenesis and development. Activating the mutations of FGFRs in the germline has long been known to cause a variety of skeletal developmental disorders, but it is only recently that a similar spectrum of somatic FGFR mutations has been associated with human cancers. Many of these somatic mutations are gain-of-function and oncogenic and create dependencies in tumor cell lines harboring such mutations. A combination of knockdown studies and pharmaceutical inhibition in preclinical models has further substantiated genomically altered FGFR as a therapeutic target in cancer, and the oncology community is responding with clinical trials evaluating multikinase inhibitors with anti-FGFR activity and a new generation of specific pan-FGFR inhibitors. Copyright © 2011. Published by Elsevier Ltd.

Mutations in the NHEJ component XRCC4 cause primordial dwarfism.

PubMed

Murray, Jennie E; van der Burg, Mirjam; IJspeert, Hanna; Carroll, Paula; Wu, Qian; Ochi, Takashi; Leitch, Andrea; Miller, Edward S; Kysela, Boris; Jawad, Alireza; Bottani, Armand; Brancati, Francesco; Cappa, Marco; Cormier-Daire, Valerie; Deshpande, Charu; Faqeih, Eissa A; Graham, Gail E; Ranza, Emmanuelle; Blundell, Tom L; Jackson, Andrew P; Stewart, Grant S; Bicknell, Louise S

2015-03-05

Non-homologous end joining (NHEJ) is a key cellular process ensuring genome integrity. Mutations in several components of the NHEJ pathway have been identified, often associated with severe combined immunodeficiency (SCID), consistent with the requirement for NHEJ during V(D)J recombination to ensure diversity of the adaptive immune system. In contrast, we have recently found that biallelic mutations in LIG4 are a common cause of microcephalic primordial dwarfism (MPD), a phenotype characterized by prenatal-onset extreme global growth failure. Here we provide definitive molecular genetic evidence supported by biochemical, cellular, and immunological data for mutations in XRCC4, encoding the obligate binding partner of LIG4, causing MPD. We report the identification of biallelic mutations in XRCC4 in five families. Biochemical and cellular studies demonstrate that these alterations substantially decrease XRCC4 protein levels leading to reduced cellular ligase IV activity. Consequently, NHEJ-dependent repair of ionizing-radiation-induced DNA double-strand breaks is compromised in XRCC4 cells. Similarly, immunoglobulin junctional diversification is impaired in cells. However, immunoglobulin levels are normal, and individuals lack overt signs of immunodeficiency. Additionally, in contrast to individuals with LIG4 mutations, pancytopenia leading to bone marrow failure has not been observed. Hence, alterations that alter different NHEJ proteins give rise to a phenotypic spectrum, from SCID to extreme growth failure, with deficiencies in certain key components of this repair pathway predominantly exhibiting growth deficits, reflecting differential developmental requirements for NHEJ proteins to support growth and immune maturation. Copyright © 2015 The American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.

A heterozygous putative null mutation in ROM1 without a mutation in peripherin/RDS in a family with retinitis pigmentosa

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sakuma, Hitoshi; Inana, G.; Murakami, Akira

1995-05-20

ROM1 is a 351-amino-acid, 37-kDa outer segment membrane protein of rod photoreceptors. ROM1 is related to peripherin/RDS, another outer segment membrane protein found in both rods and cones. The precise function of ROM1 or peripherin/RDS is not known, but they have been suggested to play important roles in the function and/or structure of the rod photoreceptor outer segment disks. A recent report implicated ROM1 in disease by suggesting that RP can be caused by a heterozygous null mutation in ROM1 but only in combination with another heterozygous mutation in peripherin/RDS. Screening of the ROM1 gene using polymerase chain reaction amplification,more » denaturing gradient gel electrophoresis, and direct DNA sequencing identified the same heterozygous putative null mutation in a family with RP.« less

Exome sequencing reveals a novel Moroccan founder mutation in SLC19A3 as a new cause of early-childhood fatal Leigh syndrome.

PubMed

Gerards, Mike; Kamps, Rick; van Oevelen, Jo; Boesten, Iris; Jongen, Eveline; de Koning, Bart; Scholte, Hans R; de Angst, Isabel; Schoonderwoerd, Kees; Sefiani, Abdelaziz; Ratbi, Ilham; Coppieters, Wouter; Karim, Latifa; de Coo, René; van den Bosch, Bianca; Smeets, Hubert

2013-03-01

Leigh syndrome is an early onset, often fatal progressive neurodegenerative disorder caused by mutations in the mitochondrial or nuclear DNA. Until now, mutations in more than 35 genes have been reported to cause Leigh syndrome, indicating an extreme genetic heterogeneity for this disorder, but still only explaining part of the cases. The possibility of whole exome sequencing enables not only mutation detection in known candidate genes, but also the identification of new genes associated with Leigh syndrome in small families and isolated cases. Exome sequencing was combined with homozygosity mapping to identify the genetic defect in a Moroccan family with fatal Leigh syndrome in early childhood and specific magnetic resonance imaging abnormalities in the brain. We detected a homozygous nonsense mutation (c.20C>A; p.Ser7Ter) in the thiamine transporter SLC19A3. In vivo overexpression of wild-type SLC19A3 showed an increased thiamine uptake, whereas overexpression of mutant SLC19A3 did not, confirming that the mutation results in an absent or non-functional protein. Seventeen additional patients with Leigh syndrome were screened for mutations in SLC19A3 using conventional Sanger sequencing. Two unrelated patients, both from Moroccan origin and one from consanguineous parents, were homozygous for the same p.Ser7Ter mutation. One of these patients showed the same MRI abnormalities as the patients from the first family. Strikingly, patients receiving thiamine had an improved life-expectancy. One patient in the third family deteriorated upon interruption of the thiamine treatment and recovered after reinitiating. Although unrelated, all patients came from the province Al Hoceima in Northern Morocco. Based on the recombination events the mutation was estimated to have occurred 1250-1750 years ago. Our data shows that SLC19A3 is a new candidate for mutation screening in patients with Leigh syndrome, who might benefit from high doses of thiamine and/or biotin. Especially, Moroccan patients with Leigh syndrome should be tested for the c.20C>A founder mutation in SLC19A3.

Mutant p53 expression in fallopian tube epithelium drives cell migration.

PubMed

Quartuccio, Suzanne M; Karthikeyan, Subbulakshmi; Eddie, Sharon L; Lantvit, Daniel D; Ó hAinmhire, Eoghainín; Modi, Dimple A; Wei, Jian-Jun; Burdette, Joanna E

2015-10-01

Ovarian cancer is the fifth leading cause of cancer death among US women. Evidence supports the hypothesis that high-grade serous ovarian cancers (HGSC) may originate in the distal end of the fallopian tube. Although a heterogeneous disease, 96% of HGSC contain mutations in p53. In addition, the "p53 signature," or overexpression of p53 protein (usually associated with mutation), is a potential precursor lesion of fallopian tube derived HGSC suggesting an essential role for p53 mutation in early serous tumorigenesis. To further clarify p53-mutation dependent effects on cells, murine oviductal epithelial cells (MOE) were stably transfected with a construct encoding for the R273H DNA binding domain mutation in p53, the most common mutation in HGSC. Mutation in p53 was not sufficient to transform MOE cells but did significantly increase cell migration. A similar p53 mutation in murine ovarian surface epithelium (MOSE), another potential progenitor cell for serous cancer, was not sufficient to transform the cells nor change migration suggesting tissue specific effects of p53 mutation. Microarray data confirmed expression changes of pro-migratory genes in p53(R273H) MOE compared to parental cells, which could be reversed by suppressing Slug expression. Combining p53(R273H) with KRAS(G12V) activation caused transformation of MOE into high-grade sarcomatoid carcinoma when xenografted into nude mice. Elucidating the specific role of p53(R273H) in the fallopian tube will improve understanding of changes at the earliest stage of transformation. This information can help develop chemopreventative strategies to prevent the accumulation of additional mutations and reverse progression of the "p53 signature" thereby, improving survival rates. © 2015 UICC.

Vacuolar Protein Sorting Genes in Parkinson's Disease: A Re-appraisal of Mutations Detection Rate and Neurobiology of Disease.

PubMed

Gambardella, Stefano; Biagioni, Francesca; Ferese, Rosangela; Busceti, Carla L; Frati, Alessandro; Novelli, Giuseppe; Ruggieri, Stefano; Fornai, Francesco

2016-01-01

Mammalian retromers play a critical role in protein trans-membrane sorting from endosome to the trans-Golgi network (TGN). Recently, retromer alterations have been related to the onset of Parkinson's Disease (PD) since the variant p.Asp620Asn in VPS35 (Vacuolar Protein Sorting 35) was identified as a cause of late onset PD. This variant causes a primary defect in endosomal trafficking and retromers formation. Other mutations in VPS genes have been reported in both sporadic and familial PD. These mutations are less defined. Understanding the specific prevalence of all VPS gene mutations is key to understand the relevance of retromers impairment in the onset of PD. A number of PD-related mutations despite affecting different biochemical systems (autophagy, mitophagy, proteasome, endosomes, protein folding), all converge in producing an impairment in cell clearance. This may explain how genetic predispositions to PD may derive from slightly deleterious VPS mutations when combined with environmental agents overwhelming the clearance of the cell. This manuscript reviews genetic data produced in the last 5 years to re-define the actual prevalence of VPS gene mutations in the onset of PD. The prevalence of p.Asp620Asn mutation in VPS35 is 0.286 of familial PD. This increases up to 0.548 when considering mutations affecting all VPS genes. This configures mutations in VPS genes as the second most frequent autosomal dominant PD genotype. This high prevalence, joined with increased awareness of the role played by retromers in the neurobiology of PD, suggests environmentally-induced VPS alterations as crucial in the genesis of PD.

Pathogenic Parkinson's disease mutations across the functional domains of LRRK2 alter the autophagic/lysosomal response to starvation.

PubMed

Manzoni, Claudia; Mamais, Adamantios; Dihanich, Sybille; McGoldrick, Phillip; Devine, Michael J; Zerle, Julia; Kara, Eleanna; Taanman, Jan-Willem; Healy, Daniel G; Marti-Masso, Jose-Felix; Schapira, Anthony H; Plun-Favreau, Helene; Tooze, Sharon; Hardy, John; Bandopadhyay, Rina; Lewis, Patrick A

2013-11-29

LRRK2 is one of the most important genetic contributors to Parkinson's disease (PD). Point mutations in this gene cause an autosomal dominant form of PD, but to date no cellular phenotype has been consistently linked with mutations in each of the functional domains (ROC, COR and Kinase) of the protein product of this gene. In this study, primary fibroblasts from individuals carrying pathogenic mutations in the three central domains of LRRK2 were assessed for alterations in the autophagy/lysosomal pathway using a combination of biochemical and cellular approaches. Mutations in all three domains resulted in alterations in markers for autophagy/lysosomal function compared to wild type cells. These data highlight the autophagy and lysosomal pathways as read outs for pathogenic LRRK2 function and as a marker for disease, and provide insight into the mechanisms linking LRRK2 function and mutations. Copyright © 2013 The Authors. Published by Elsevier Inc. All rights reserved.

Expanding the spectrum of genetic mutations in antenatal Bartter syndrome type II.

PubMed

Fretzayas, Andreas; Gole, Evangelia; Attilakos, Achilleas; Daskalaki, Anna; Nicolaidou, Polyxeni; Papadopoulou, Anna

2013-06-01

Bartter syndrome (BS) is a group of genetic disorders characterized by hypokalemic metabolic alkalosis, hyponatremia and elevated renin and aldosterone plasma concentrations. BS type II is caused by mutations in the KCNJ1 gene and usually presents with transient hyperkalemia. We report here a novel KCNJ1 mutation in a male neonate, prematurely born after a pregnancy complicated by polyhydramnios. The infant presented with typical clinical and laboratory findings of BS type II, such as hyponatremia, hypochloremic metabolic alkalosis, severe weight loss, elevated renin and aldosterone levels and transient hyperkalemia in the early postnatal period, which were later normalized. Molecular analysis revealed a compound heterozygous mutation in the KCNJ1 gene, consisting of a novel K76E and an already described V315G mutation, both affecting functional domains of the channel protein. Typical manifestations of antenatal BS in combination with hyperkalemia should prompt the clinician to search for mutations in the KCNJ1 gene first. © 2013 The Authors. Pediatrics International © 2013 Japan Pediatric Society.

Mutation Rate Variation is a Primary Determinant of the Distribution of Allele Frequencies in Humans

PubMed Central

Pritchard, Jonathan K.

2016-01-01

The site frequency spectrum (SFS) has long been used to study demographic history and natural selection. Here, we extend this summary by examining the SFS conditional on the alleles found at the same site in other species. We refer to this extension as the “phylogenetically-conditioned SFS” or cSFS. Using recent large-sample data from the Exome Aggregation Consortium (ExAC), combined with primate genome sequences, we find that human variants that occurred independently in closely related primate lineages are at higher frequencies in humans than variants with parallel substitutions in more distant primates. We show that this effect is largely due to sites with elevated mutation rates causing significant departures from the widely-used infinite sites mutation model. Our analysis also suggests substantial variation in mutation rates even among mutations involving the same nucleotide changes. In summary, we show that variable mutation rates are key determinants of the SFS in humans. PMID:27977673

Whole-Exome Sequencing Reveals GPIHBP1 Mutations in Infantile Colitis With Severe Hypertriglyceridemia

PubMed Central

Gonzaga-Jauregui, Claudia; Mir, Sabina; Penney, Samantha; Jhangiani, Shalini; Midgen, Craig; Finegold, Milton; Muzny, Donna M.; Wang, Min; Bacino, Carlos A.; Gibbs, Richard A.; Lupski, James R.; Kellermayer, Richard; Hanchard, Neil A.

2014-01-01

Severe congenital hypertriglyceridemia (HTG) is a rare disorder caused by mutations in genes affecting lipoprotein lipase (LPL) activity. Here we report a 5-week-old Hispanic girl with severe HTG (12,031 mg/dL, normal limit 150 mg/dL) who presented with the unusual combination of lower gastrointestinal bleeding and milky plasma. Initial colonoscopy was consistent with colitis, which resolved with reduction of triglycerides. After negative sequencing of the LPL gene, whole-exome sequencing revealed novel compound heterozygous mutations in GPIHBP1. Our study broadens the phenotype of GPIHBP1-associated HTG, reinforces the effectiveness of whole-exome sequencing in Mendelian diagnoses, and implicates triglycer-ides in gastrointestinal mucosal injury. PMID:24614124

Whole-exome sequencing reveals GPIHBP1 mutations in infantile colitis with severe hypertriglyceridemia.

PubMed

Gonzaga-Jauregui, Claudia; Mir, Sabina; Penney, Samantha; Jhangiani, Shalini; Midgen, Craig; Finegold, Milton; Muzny, Donna M; Wang, Min; Bacino, Carlos A; Gibbs, Richard A; Lupski, James R; Kellermayer, Richard; Hanchard, Neil A

2014-07-01

Severe congenital hypertriglyceridemia (HTG) is a rare disorder caused by mutations in genes affecting lipoprotein lipase (LPL) activity. Here we report a 5-week-old Hispanic girl with severe HTG (12,031 mg/dL, normal limit 150 mg/dL) who presented with the unusual combination of lower gastrointestinal bleeding and milky plasma. Initial colonoscopy was consistent with colitis, which resolved with reduction of triglycerides. After negative sequencing of the LPL gene, whole-exome sequencing revealed novel compound heterozygous mutations in GPIHBP1. Our study broadens the phenotype of GPIHBP1-associated HTG, reinforces the effectiveness of whole-exome sequencing in Mendelian diagnoses, and implicates triglycerides in gastrointestinal mucosal injury.

Adaptor Protein Complex 4 Deficiency Causes Severe Autosomal-Recessive Intellectual Disability, Progressive Spastic Paraplegia, Shy Character, and Short Stature

PubMed Central

Abou Jamra, Rami; Philippe, Orianne; Raas-Rothschild, Annick; Eck, Sebastian H.; Graf, Elisabeth; Buchert, Rebecca; Borck, Guntram; Ekici, Arif; Brockschmidt, Felix F.; Nöthen, Markus M.; Munnich, Arnold; Strom, Tim M.; Reis, Andre; Colleaux, Laurence

2011-01-01

Intellectual disability inherited in an autosomal-recessive fashion represents an important fraction of severe cognitive-dysfunction disorders. Yet, the extreme heterogeneity of these conditions markedly hampers gene identification. Here, we report on eight affected individuals who were from three consanguineous families and presented with severe intellectual disability, absent speech, shy character, stereotypic laughter, muscular hypotonia that progressed to spastic paraplegia, microcephaly, foot deformity, decreased muscle mass of the lower limbs, inability to walk, and growth retardation. Using a combination of autozygosity mapping and either Sanger sequencing of candidate genes or next-generation exome sequencing, we identified one mutation in each of three genes encoding adaptor protein complex 4 (AP4) subunits: a nonsense mutation in AP4S1 (NM_007077.3: c.124C>T, p.Arg42∗), a frameshift mutation in AP4B1 (NM_006594.2: c.487_488insTAT, p.Glu163_Ser739delinsVal), and a splice mutation in AP4E1 (NM_007347.3: c.542+1_542+4delGTAA, r.421_542del, p.Glu181Glyfs∗20). Adaptor protein complexes (AP1-4) are ubiquitously expressed, evolutionarily conserved heterotetrameric complexes that mediate different types of vesicle formation and the selection of cargo molecules for inclusion into these vesicles. Interestingly, two mutations affecting AP4M1 and AP4E1 have recently been found to cause cerebral palsy associated with severe intellectual disability. Combined with previous observations, these results support the hypothesis that AP4-complex-mediated trafficking plays a crucial role in brain development and functioning and demonstrate the existence of a clinically recognizable syndrome due to deficiency of the AP4 complex. PMID:21620353

Whole exome sequencing confirms the clinical diagnosis of Marfan syndrome combined with X-linked hypophosphatemia.

PubMed

Sheng, Xunlun; Chen, Xue; Lei, Bo; Chen, Rui; Wang, Hui; Zhang, Fangxia; Rong, Weining; Ha, Ruoshui; Liu, Yani; Zhao, Feng; Yang, Peizeng; Zhao, Chen

2015-06-04

To determine the genetic lesions and to modify the clinical diagnosis for a Chinese family with significant intrafamilial phenotypic diversities and unusual presentations. Three affected patients and the asymptomatic father were included and received comprehensive systemic examinations. Whole exome sequencing (WES) was performed for mutation detection. Structural modeling test was applied to analyze the potential structural changes caused by the missense substitution. The proband showed a wide spectrum of systemic anomalies, including bilateral ectopia lentis, atrial septal defect, ventricular septal defect, widening of tibial metaphysis with medial bowing, and dolichostenomelia in digits, while her mother and elder brother only demonstrated similar skeletal changes. A recurrent mutation, PHEX p.R291*, was found in all patients, while a de novo mutation, FBN1 p.C792F, was only detected in the proband. The FBN1 substitution was also predicted to cause significant conformational change in fibrillin-1 protein, thus changing its physical and biological properties. Taken together, we finalized the diagnosis for this family as X-linked hypophosphatemia (XLH), and diagnosed this girl as Marfan syndrome combined with XLH, and congenital heart disease. Our study also emphasizes the importance of WES in assisting the clinical diagnosis for complicated cases when the original diagnoses are challenged.

Activation of a cryptic splice site in the mitochondrial elongation factor GFM1 causes combined OXPHOS deficiency☆

PubMed Central

Simon, Mariella T.; Ng, Bobby G.; Friederich, Marisa W.; Wang, Raymond Y.; Boyer, Monica; Kircher, Martin; Collard, Renata; Buckingham, Kati J.; Chang, Richard; Shendure, Jay; Nickerson, Deborah A.; Bamshad, Michael J.; Van Hove, Johan L.K.; Freeze, Hudson H.; Abdenur, Jose E.

2017-01-01

We report the clinical, biochemical, and molecular findings in two brothers with encephalopathy and multi-systemic disease. Abnormal transferrin glycoforms were suggestive of a type I congenital disorder of glycosylation (CDG). While exome sequencing was negative for CDG related candidate genes, the testing revealed compound heterozygous mutations in the mitochondrial elongation factor G gene (GFM1). One of the mutations had been reported previously while the second, novel variant was found deep in intron 6, activating a cryptic splice site. Functional studies demonstrated decreased GFM1 protein levels, suggested disrupted assembly of mitochondrial complexes III and V and decreased activities of mitochondrial complexes I and IV, all indicating combined OXPHOS deficiency. PMID:28216230

A Recessive Mutation Resulting in a Disabling Amino Acid Substitution (T194R) in the LHX3 Homeodomain Causes Combined Pituitary Hormone Deficiency

PubMed Central

Bechtold-Dalla Pozza, Susanne; Hiedl, Stefan; Roeb, Julia; Lohse, Peter; Malik, Raleigh E.; Park, Soyoung; Durán-Prado, Mario; Rhodes, Simon J.

2012-01-01

Background/Aims Recessive mutations in the LHX3 ho-meodomain transcription factor gene are associated with developmental disorders affecting the pituitary and nervous system. We describe pediatric patients with combined pituitary hormone deficiency (CPHD) who harbor a novel mutation in LHX3. Methods Two female siblings from related parents were examined. Both patients had neonatal complications. The index patient had CPHD featuring deficiencies of GH, LH, FSH, PRL, and TSH, with later onset of ACTH deficiency. She also had a hypoplastic anterior pituitary, respiratory distress, hearing impairment, and limited neck rotation. The LHX3 gene was sequenced and the biochemical properties of the predicted altered proteins were characterized. Results A novel homozygous mutation predicted to change amino acid 194 from threonine to arginine (T194R) was detected in both patients. This amino acid is conserved in the DNA-binding homeodomain. Computer modeling predicted that the T194R change would alter the homeodomain structure. The T194R protein did not bind tested LHX3 DNA recognition sites and did not activate the α-glycoprotein and PRL target genes. Conclusion The T194R mutation affects a critical residue in the LHX3 protein. This study extends our understanding of the phenotypic features, molecular mechanism, and developmental course associated with mutations in the LHX3 gene. PMID:22286346

Simultaneous Detection of Both Single Nucleotide Variations and Copy Number Alterations by Next-Generation Sequencing in Gorlin Syndrome

PubMed Central

Morita, Kei-ichi; Naruto, Takuya; Tanimoto, Kousuke; Yasukawa, Chisato; Oikawa, Yu; Masuda, Kiyoshi; Imoto, Issei; Inazawa, Johji; Omura, Ken; Harada, Hiroyuki

2015-01-01

Gorlin syndrome (GS) is an autosomal dominant disorder that predisposes affected individuals to developmental defects and tumorigenesis, and caused mainly by heterozygous germline PTCH1 mutations. Despite exhaustive analysis, PTCH1 mutations are often unidentifiable in some patients; the failure to detect mutations is presumably because of mutations occurred in other causative genes or outside of analyzed regions of PTCH1, or copy number alterations (CNAs). In this study, we subjected a cohort of GS-affected individuals from six unrelated families to next-generation sequencing (NGS) analysis for the combined screening of causative alterations in Hedgehog signaling pathway-related genes. Specific single nucleotide variations (SNVs) of PTCH1 causing inferred amino acid changes were identified in four families (seven affected individuals), whereas CNAs within or around PTCH1 were found in two families in whom possible causative SNVs were not detected. Through a targeted resequencing of all coding exons, as well as simultaneous evaluation of copy number status using the alignment map files obtained via NGS, we found that GS phenotypes could be explained by PTCH1 mutations or deletions in all affected patients. Because it is advisable to evaluate CNAs of candidate causative genes in point mutation-negative cases, NGS methodology appears to be useful for improving molecular diagnosis through the simultaneous detection of both SNVs and CNAs in the targeted genes/regions. PMID:26544948

Simultaneous Detection of Both Single Nucleotide Variations and Copy Number Alterations by Next-Generation Sequencing in Gorlin Syndrome.

PubMed

Morita, Kei-ichi; Naruto, Takuya; Tanimoto, Kousuke; Yasukawa, Chisato; Oikawa, Yu; Masuda, Kiyoshi; Imoto, Issei; Inazawa, Johji; Omura, Ken; Harada, Hiroyuki

2015-01-01

Gorlin syndrome (GS) is an autosomal dominant disorder that predisposes affected individuals to developmental defects and tumorigenesis, and caused mainly by heterozygous germline PTCH1 mutations. Despite exhaustive analysis, PTCH1 mutations are often unidentifiable in some patients; the failure to detect mutations is presumably because of mutations occurred in other causative genes or outside of analyzed regions of PTCH1, or copy number alterations (CNAs). In this study, we subjected a cohort of GS-affected individuals from six unrelated families to next-generation sequencing (NGS) analysis for the combined screening of causative alterations in Hedgehog signaling pathway-related genes. Specific single nucleotide variations (SNVs) of PTCH1 causing inferred amino acid changes were identified in four families (seven affected individuals), whereas CNAs within or around PTCH1 were found in two families in whom possible causative SNVs were not detected. Through a targeted resequencing of all coding exons, as well as simultaneous evaluation of copy number status using the alignment map files obtained via NGS, we found that GS phenotypes could be explained by PTCH1 mutations or deletions in all affected patients. Because it is advisable to evaluate CNAs of candidate causative genes in point mutation-negative cases, NGS methodology appears to be useful for improving molecular diagnosis through the simultaneous detection of both SNVs and CNAs in the targeted genes/regions.

Exome sequencing identifies a novel SMCHD1 mutation in facioscapulohumeral muscular dystrophy 2.

PubMed

Mitsuhashi, Satomi; Boyden, Steven E; Estrella, Elicia A; Jones, Takako I; Rahimov, Fedik; Yu, Timothy W; Darras, Basil T; Amato, Anthony A; Folkerth, Rebecca D; Jones, Peter L; Kunkel, Louis M; Kang, Peter B

2013-12-01

FSHD2 is a rare form of facioscapulohumeral muscular dystrophy (FSHD) characterized by the absence of a contraction in the D4Z4 macrosatellite repeat region on chromosome 4q35 that is the hallmark of FSHD1. However, hypomethylation of this region is common to both subtypes. Recently, mutations in SMCHD1 combined with a permissive 4q35 allele were reported to cause FSHD2. We identified a novel p.Lys275del SMCHD1 mutation in a family affected with FSHD2 using whole-exome sequencing and linkage analysis. This mutation alters a highly conserved amino acid in the ATPase domain of SMCHD1. Subject III-11 is a male who developed asymmetrical muscle weakness characteristic of FSHD at 13 years. Physical examination revealed marked bilateral atrophy at biceps brachii, bilateral scapular winging, some asymmetrical weakness at tibialis anterior and peroneal muscles, and mild lower facial weakness. Biopsy of biceps brachii in subject II-5, the father of III-11, demonstrated lobulated fibers and dystrophic changes. Endomysial and perivascular inflammation was found, which has been reported in FSHD1 but not FSHD2. Given the previous report of SMCHD1 mutations in FSHD2 and the clinical presentations consistent with the FSHD phenotype, we conclude that the SMCHD1 mutation is the likely cause of the disease in this family. Copyright © 2013 Elsevier B.V. All rights reserved.

Novel TMEM67 Mutations and Genotype-phenotype Correlates in Meckelin-related Ciliopathies

PubMed Central

Iannicelli, Miriam; Brancati, Francesco; Mougou-Zerelli, Soumaya; Mazzotta, Annalisa; Thomas, Sophie; Elkhartoufi, Nadia; Travaglini, Lorena; Gomes, Céline; Ardissino, Gian Luigi; Bertini, Enrico; Boltshauser, Eugen; Castorina, Pierangela; D'Arrigo, Stefano; Fischetto, Rita; Leroy, Brigitte; Loget, Philippe; Bonnière, Maryse; Starck, Lena; Tantau, Julia; Gentilin, Barbara; Majore, Silvia; Swistun, Dominika; Flori, Elizabeth; Lalatta, Faustina; Pantaleoni, Chiara; Johannes.Penzien; Grammatico, Paola; Dallapiccola, Bruno; Gleeson, Joseph G.; Attie-Bitach, Tania; Valente, Enza Maria

2010-01-01

Human ciliopathies are hereditary conditions caused by defects of proteins expressed at the primary cilium. Among ciliopathies, Joubert syndrome and related disorders (JSRD), Meckel syndrome (MKS) and nephronophthisis (NPH) present clinical and genetic overlap, being allelic at several loci. One of the most interesting gene is TMEM67, encoding the transmembrane protein meckelin. We performed mutation analysis of TMEM67 in 341 probands, including 265 JSRD representative of all clinical subgroups and 76 MKS fetuses. We identified 33 distinct mutations, of which 20 were novel, in 8/10 (80%) JS with liver involvement (COACH phenotype) and 12/76 (16%) MKS fetuses. No mutations were found in other JSRD subtypes, confirming the strong association between TMEM67 mutations and liver involvement. Literature review of all published TMEM67 mutated cases was performed to delineate genotype-phenotype correlates. In particular, comparison of the types of mutations and their distribution along the gene in lethal versus non lethal phenotypes showed in MKS patients a significant enrichment of missense mutations falling in TMEM67 exons 8 to 15, especially when in combination with a truncating mutation. These exons encode for a region of unknown function in the extracellular domain of meckelin. PMID:20232449

Exome Sequencing Identifies a Founder Frameshift Mutation in an Alternative Exon of USH1C as the Cause of Autosomal Recessive Retinitis Pigmentosa with Late-Onset Hearing Loss

PubMed Central

Khateb, Samer; Zelinger, Lina; Ben-Yosef, Tamar; Crystal-Shalit, Ornit; Gross, Menachem; Banin, Eyal; Sharon, Dror

2012-01-01

We used a combined approach of homozygosity mapping and whole exome sequencing (WES) to search for the genetic cause of autosomal recessive retinitis pigmentosa (arRP) in families of Yemenite Jewish origin. Homozygosity mapping of two arRP Yemenite Jewish families revealed a few homozygous regions. A subsequent WES analysis of the two index cases revealed a shared homozygous novel nucleotide deletion (c.1220delG) leading to a frameshift (p.Gly407Glufs*56) in an alternative exon (#15) of USH1C. Screening of additional Yemenite Jewish patients revealed a total of 16 homozygous RP patients (with a carrier frequency of 0.008 in controls). Funduscopic and electroretinography findings were within the spectrum of typical RP. While other USH1C mutations usually cause Usher type I (including RP, vestibular dysfunction and congenital deafness), audiometric screening of 10 patients who are homozygous for c.1220delG revealed that patients under 40 years of age had normal hearing while older patients showed mild to severe high tone sensorineural hearing loss. This is the first report of a mutation in a known USH1 gene that causes late onset rather than congenital sensorineural hearing loss. The c.1220delG mutation of USH1C accounts for 23% of RP among Yemenite Jewish patients in our cohort. PMID:23251578

Recurrent gain-of-function mutation in PRKG1 causes thoracic aortic aneurysms and acute aortic dissections.

PubMed

Guo, Dong-chuan; Regalado, Ellen; Casteel, Darren E; Santos-Cortez, Regie L; Gong, Limin; Kim, Jeong Joo; Dyack, Sarah; Horne, S Gabrielle; Chang, Guijuan; Jondeau, Guillaume; Boileau, Catherine; Coselli, Joseph S; Li, Zhenyu; Leal, Suzanne M; Shendure, Jay; Rieder, Mark J; Bamshad, Michael J; Nickerson, Deborah A; Kim, Choel; Milewicz, Dianna M

2013-08-08

Gene mutations that lead to decreased contraction of vascular smooth-muscle cells (SMCs) can cause inherited thoracic aortic aneurysms and dissections. Exome sequencing of distant relatives affected by thoracic aortic disease and subsequent Sanger sequencing of additional probands with familial thoracic aortic disease identified the same rare variant, PRKG1 c.530G>A (p.Arg177Gln), in four families. This mutation segregated with aortic disease in these families with a combined two-point LOD score of 7.88. The majority of affected individuals presented with acute aortic dissections (63%) at relatively young ages (mean 31 years, range 17-51 years). PRKG1 encodes type I cGMP-dependent protein kinase (PKG-1), which is activated upon binding of cGMP and controls SMC relaxation. Although the p.Arg177Gln alteration disrupts binding to the high-affinity cGMP binding site within the regulatory domain, the altered PKG-1 is constitutively active even in the absence of cGMP. The increased PKG-1 activity leads to decreased phosphorylation of the myosin regulatory light chain in fibroblasts and is predicted to cause decreased contraction of vascular SMCs. Thus, identification of a gain-of-function mutation in PRKG1 as a cause of thoracic aortic disease provides further evidence that proper SMC contractile function is critical for maintaining the integrity of the thoracic aorta throughout a lifetime. Copyright © 2013 The American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.

Biallelic mutations in BRCA1 cause a new Fanconi anemia subtype.

PubMed

Sawyer, Sarah L; Tian, Lei; Kähkönen, Marketta; Schwartzentruber, Jeremy; Kircher, Martin; Majewski, Jacek; Dyment, David A; Innes, A Micheil; Boycott, Kym M; Moreau, Lisa A; Moilanen, Jukka S; Greenberg, Roger A

2015-02-01

Deficiency in BRCA-dependent DNA interstrand crosslink (ICL) repair is intimately connected to breast cancer susceptibility and to the rare developmental syndrome Fanconi anemia. Bona fide Fanconi anemia proteins, BRCA2 (FANCD1), PALB2 (FANCN), and BRIP1 (FANCJ), interact with BRCA1 during ICL repair. However, the lack of detailed phenotypic and cellular characterization of a patient with biallelic BRCA1 mutations has precluded assignment of BRCA1 as a definitive Fanconi anemia susceptibility gene. Here, we report the presence of biallelic BRCA1 mutations in a woman with multiple congenital anomalies consistent with a Fanconi anemia-like disorder and breast cancer at age 23. Patient cells exhibited deficiency in BRCA1 and RAD51 localization to DNA-damage sites, combined with radial chromosome formation and hypersensitivity to ICL-inducing agents. Restoration of these functions was achieved by ectopic introduction of a BRCA1 transgene. These observations provide evidence in support of BRCA1 as a new Fanconi anemia gene (FANCS). We establish that biallelic BRCA1 mutations cause a distinct FA-S, which has implications for risk counselling in families where both parents harbor BRCA1 mutations. The genetic basis of hereditary cancer susceptibility syndromes provides diagnostic information, insights into treatment strategies, and more accurate recurrence risk counseling to families. ©2014 American Association for Cancer Research.

Immunotherapy holds the key to cancer treatment and prevention in constitutional mismatch repair deficiency (CMMRD) syndrome.

PubMed

Westdorp, Harm; Kolders, Sigrid; Hoogerbrugge, Nicoline; de Vries, I Jolanda M; Jongmans, Marjolijn C J; Schreibelt, Gerty

2017-09-10

Monoallelic germline mutations in one of the DNA mismatch repair (MMR) genes cause Lynch syndrome, with a high lifetime risks of colorectal and endometrial cancer at adult age. Less well known, is the constitutional mismatch repair deficiency (CMMRD) syndrome caused by biallelic germline mutations in MMR genes. This syndrome is characterized by the development of childhood cancer. Patients with CMMRD are at extremely high risk of developing multiple cancers including hematological, brain and intestinal tumors. Mutations in MMR genes impair DNA repair and therefore most tumors of patients with CMMRD are hypermutated. These mutations lead to changes in the translational reading frame, which consequently result in neoantigen formation. Neoantigens are recognized as foreign by the immune system and can induce specific immune responses. The growing evidence on the clinical efficacy of immunotherapies, such as immune checkpoint inhibitors, offers the prospect for treatment of patients with CMMRD. Combining neoantigen-based vaccination strategies and immune checkpoint inhibitors could be an effective way to conquer CMMRD-related tumors. Neoantigen-based vaccines might also be a preventive treatment option in healthy biallelic MMR mutation carriers. Future studies need to reveal the safety and efficacy of immunotherapies for patients with CMMRD. Copyright © 2017 The Author(s). Published by Elsevier B.V. All rights reserved.

Novel mutations in LRP6 highlight the role of WNT signaling in tooth agenesis

PubMed Central

Ludwig, Kerstin U.; Sullivan, Robert; van Rooij, Iris A.L.M.; Thonissen, Michelle; Swinnen, Steven; Phan, Milien; Conte, Federica; Ishorst, Nina; Gilissen, Christian; RoaFuentes, Laury; van de Vorst, Maartje; Henkes, Arjen; Steehouwer, Marloes; van Beusekom, Ellen; Bloemen, Marjon; Vankeirsbilck, Bruno; Bergé, Stefaan; Hens, Greet; Schoenaers, Joseph; Poorten, Vincent Vander; Roosenboom, Jasmien; Verdonck, An; Devriendt, Koen; Roeleveldt, Nel; Jhangiani, Shalini N.; Vissers, Lisenka E.L.M.; Lupski, James R.; de Ligt, Joep; Von den Hoff, Johannes W.; Pfundt, Rolph; Brunner, Han G.; Zhou, Huiqing; Dixon, Jill; Mangold, Elisabeth; van Bokhoven, Hans; Dixon, Michael J.; Kleefstra, Tjitske

2016-01-01

Purpose Here we aimed to identify a novel genetic cause of tooth agenesis (TA) and/or orofacial clefting (OFC) by combining whole exome sequencing (WES) and targeted re-sequencing in a large cohort of TA and OFC patients. Methods WES was performed in two unrelated patients, one with severe TA and OFC and another with severe TA only. After identifying deleterious mutations in a gene encoding the low density lipoprotein receptor-related protein 6 (LRP6), all its exons were re-sequenced with molecular inversion probes, in 67 patients with TA, 1,072 patients with OFC and in 706 controls. Results We identified a frameshift (c.4594delG, p.Cys1532fs) and a canonical splice site mutation (c.3398-2A>C, p.?) in LRP6 respectively in the patient with TA and OFC, and in the patient with severe TA only. The targeted re-sequencing showed significant enrichment of unique LRP6 variants in TA patients, but not in nonsyndromic OFC. From the 5 variants in patients with TA, 2 affect the canonical splice site and 3 were missense variants; all variants segregated with the dominant phenotype and in 1 case the missense mutation occurred de novo. Conclusion Mutations in LRP6 cause tooth agenesis in man. PMID:26963285

The Saccharomyces cerevisiae MLH3 gene functions in MSH3-dependent suppression of frameshift mutations.

PubMed

Flores-Rozas, H; Kolodner, R D

1998-10-13

The Saccharomyces cerevisiae genome encodes four MutL homologs. Of these, MLH1 and PMS1 are known to act in the MSH2-dependent pathway that repairs DNA mismatches. We have investigated the role of MLH3 in mismatch repair. Mutations in MLH3 increased the rate of reversion of the hom3-10 allele by increasing the rate of deletion of a single T in a run of 7 Ts. Combination of mutations in MLH3 and MSH6 caused a synergistic increase in the hom3-10 reversion rate, whereas the hom3-10 reversion rate in an mlh3 msh3 double mutant was the same as in the respective single mutants. Similar results were observed when the accumulation of mutations at frameshift hot spots in the LYS2 gene was analyzed, although mutation of MLH3 did not cause the same extent of affect at every LYS2 frameshift hot spot. MLH3 interacted with MLH1 in a two-hybrid system. These data are consistent with the idea that a proportion of the repair of specific insertion/deletion mispairs by the MSH3-dependent mismatch repair pathway uses a heterodimeric MLH1-MLH3 complex in place of the MLH1-PMS1 complex.

Significance of combined detection of JAK2V617F, MPL and CALR gene mutations in patients with essential thrombocythemia

PubMed Central

Ji, Liying; Qian, Mengyao; Wu, Nana; Wu, Jianmin

2017-01-01

The aim of this study was to analyze the mutation rate of JAK2V617F, MPLW515L/K and CALR genes in adult patients with essential thrombocythemia (ET) and the accuracy of the combined detection by the receiver operating curve. Three hundred and forty-two cases with high-platelets (≥300×109/l) were consecutively selected. The patients were analyzed for routine blood examination, bone marrow biopsy and genetic testing. One hundred and fifty-four cases (45.03%) were diagnosed with ET and 188 cases of secondary thrombocythemia according to the hematopoietic and lymphoid tissue tumor classification standards of 2008. It was found that the mutant type of three genes showed three bands, whereas only one band for wild-type. The JAK2V617F and MPL mutations did not cause a change in the open reading frame and the CALR mutation resulted in its change. The mutation rate of JAK2V617F and CALR in ET group was significantly higher than that in the secondary thrombocythemia group (p<0.05). The positive mutation rate of MPL was only 4.55%. JAK2V617F-positive mutation alone was used to diagnose with ET. The area under the curve (AUC) was 0.721. The sensitivity was 72.4%, the specificity was 79.5% and the cut-off value was 0.25. When CALR-positive mutation alone was used to diagnose ET, the AUC, sensitivity, specificity and cut-off value were 0.664, 68.4, 82.4 and 0.09%, respectively. JAK2V617F combined with CALR mutation were used for diagnosis of ET. The AUC was 0.862, the sensitivity was 85.9%, the specificity was 87.8%, and the cut-off values were 0.21 and 0.07. In conclusion, the positive mutation rate of JAK2V617F and CALR in ET was higher, and the sensitivity, specificity and accuracy of the diagnosis of ET were significantly improved using the detection of JAK2V617F and CALR. PMID:28450924

Significance of combined detection of JAK2V617F, MPL and CALR gene mutations in patients with essential thrombocythemia.

PubMed

Ji, Liying; Qian, Mengyao; Wu, Nana; Wu, Jianmin

2017-03-01

The aim of this study was to analyze the mutation rate of JAK2V617F, MPLW515L/K and CALR genes in adult patients with essential thrombocythemia (ET) and the accuracy of the combined detection by the receiver operating curve. Three hundred and forty-two cases with high-platelets (≥300×10 9 /l) were consecutively selected. The patients were analyzed for routine blood examination, bone marrow biopsy and genetic testing. One hundred and fifty-four cases (45.03%) were diagnosed with ET and 188 cases of secondary thrombocythemia according to the hematopoietic and lymphoid tissue tumor classification standards of 2008. It was found that the mutant type of three genes showed three bands, whereas only one band for wild-type. The JAK2V617F and MPL mutations did not cause a change in the open reading frame and the CALR mutation resulted in its change. The mutation rate of JAK2V617F and CALR in ET group was significantly higher than that in the secondary thrombocythemia group (p<0.05). The positive mutation rate of MPL was only 4.55%. JAK2V617F-positive mutation alone was used to diagnose with ET. The area under the curve (AUC) was 0.721. The sensitivity was 72.4%, the specificity was 79.5% and the cut-off value was 0.25. When CALR-positive mutation alone was used to diagnose ET, the AUC, sensitivity, specificity and cut-off value were 0.664, 68.4, 82.4 and 0.09%, respectively. JAK2V617F combined with CALR mutation were used for diagnosis of ET. The AUC was 0.862, the sensitivity was 85.9%, the specificity was 87.8%, and the cut-off values were 0.21 and 0.07. In conclusion, the positive mutation rate of JAK2V617F and CALR in ET was higher, and the sensitivity, specificity and accuracy of the diagnosis of ET were significantly improved using the detection of JAK2V617F and CALR.

ER stress and basement membrane defects combine to cause glomerular and tubular renal disease resulting from Col4a1 mutations in mice

PubMed Central

Jones, Frances E.; Bailey, Matthew A.; Murray, Lydia S.; Lu, Yinhui; McNeilly, Sarah; Schlötzer-Schrehardt, Ursula; Lennon, Rachel; Sado, Yoshikazu; Brownstein, David G.; Mullins, John J.; Kadler, Karl E.; Van Agtmael, Tom

2016-01-01

ABSTRACT Collagen IV is a major component of basement membranes, and mutations in COL4A1, which encodes collagen IV alpha chain 1, cause a multisystemic disease encompassing cerebrovascular, eye and kidney defects. However, COL4A1 renal disease remains poorly characterized and its pathomolecular mechanisms are unknown. We show that Col4a1 mutations in mice cause hypotension and renal disease, including proteinuria and defects in Bowman's capsule and the glomerular basement membrane, indicating a role for Col4a1 in glomerular filtration. Impaired sodium reabsorption in the loop of Henle and distal nephron despite elevated aldosterone levels indicates that tubular defects contribute to the hypotension, highlighting a novel role for the basement membrane in vascular homeostasis by modulation of the tubular response to aldosterone. Col4a1 mutations also cause diabetes insipidus, whereby the tubular defects lead to polyuria associated with medullary atrophy and a subsequent reduction in the ability to upregulate aquaporin 2 and concentrate urine. Moreover, haematuria, haemorrhage and vascular basement membrane defects confirm an important vascular component. Interestingly, although structural and compositional basement membrane defects occurred in the glomerulus and Bowman's capsule, no tubular basement membrane defects were detected. By contrast, medullary atrophy was associated with chronic ER stress, providing evidence for cell-type-dependent molecular mechanisms of Col4a1 mutations. These data show that both basement membrane defects and ER stress contribute to Col4a1 renal disease, which has important implications for the development of treatment strategies for collagenopathies. PMID:26839400

CAV3 mutations causing exercise intolerance, myalgia and rhabdomyolysis: Expanding the phenotypic spectrum of caveolinopathies.

PubMed

Scalco, Renata Siciliani; Gardiner, Alice R; Pitceathly, Robert D S; Hilton-Jones, David; Schapira, Anthony H; Turner, Chris; Parton, Matt; Desikan, Mahalekshmi; Barresi, Rita; Marsh, Julie; Manzur, Adnan Y; Childs, Anne-Marie; Feng, Lucy; Murphy, Elaine; Lamont, Phillipa J; Ravenscroft, Gianina; Wallefeld, William; Davis, Mark R; Laing, Nigel G; Holton, Janice L; Fialho, Doreen; Bushby, Kate; Hanna, Michael G; Phadke, Rahul; Jungbluth, Heinz; Houlden, Henry; Quinlivan, Ros

2016-08-01

Rhabdomyolysis is often due to a combination of environmental trigger(s) and genetic predisposition; however, the underlying genetic cause remains elusive in many cases. Mutations in CAV3 lead to various neuromuscular phenotypes with partial overlap, including limb girdle muscular dystrophy type 1C (LGMD1C), rippling muscle disease, distal myopathy and isolated hyperCKemia. Here we present a series of eight patients from seven families presenting with exercise intolerance and rhabdomyolysis caused by mutations in CAV3 diagnosed by next generation sequencing (NGS) (n = 6). Symptoms included myalgia (n = 7), exercise intolerance (n = 7) and episodes of rhabdomyolysis (n = 2). Percussion-induced rapid muscle contractions (PIRCs) were seen in five out of six patients examined. A previously reported heterozygous mutation in CAV3 (p.T78M) and three novel variants (p.V14I, p.F41S, p.F54V) were identified. Caveolin-3 immunolabeling in muscle was normal in 3/4 patients; however, immunoblotting showed more than 50% reduction of caveolin-3 in five patients compared with controls. This case series demonstrates that exercise intolerance, myalgia and rhabdomyolysis may be caused by CAV3 mutations and broadens the phenotypic spectrum of caveolinopathies. In our series, immunoblotting was a more sensitive method to detect reduced caveolin-3 levels than immunohistochemistry in skeletal muscle. Patients presenting with muscle pain, exercise intolerance and rhabdomyolysis should be routinely tested for PIRCs as this may be an important clinical clue for caveolinopathies, even in the absence of other "typical" features. The use of NGS may expand current knowledge concerning inherited diseases, and unexpected/atypical phenotypes may be attributed to well-known human disease genes. Copyright © 2016 Elsevier B.V. All rights reserved.

BRAF/KRAS gene sequencing of sebaceous neoplasms after mismatch repair protein analysis.

PubMed

Cornejo, Kristine M; Hutchinson, Lloyd; Deng, April; Tomaszewicz, Keith; Welch, Matthew; Lyle, Stephen; Dresser, Karen; Cosar, Ediz F

2014-06-01

Sebaceous neoplasms are cutaneous markers for the autosomal-dominant Muir-Torre syndrome (MTS). This phenotypic variant of Lynch syndrome (LS) is caused by germline mutations in DNA mismatch repair (MMR) genes. Microsatellite instability or loss of protein expression suggests a mutation or promoter hypermethylation in 1 of the MMR genes. BRAF gene sequencing may help to distinguish between patients with sporadic and LS-associated colorectal carcinomas with loss of MLH1 expression. LS-associated carcinomas are virtually negative for BRAF mutations, but a subset harbors KRAS mutations. The aim of our study was to test sebaceous neoplasms for V600E BRAF or KRAS mutations to determine if these mutations are associated with somatic or germline MMR defects, analogous to colorectal carcinomas. Over a 4-year period, 32 cases comprising 21 sebaceous adenomas, 3 sebaceomas, and 8 sebaceous carcinomas with sufficient material for testing were collected. MMR immunohistochemistry showed that 7 neoplasms had combined loss of MLH1-PMS2, 16 neoplasms had combined loss of MSH2-MSH6, 2 neoplasms had solitary loss of MSH6, and 7 sebaceous neoplasms had intact protein expression. BRAF/KRAS testing revealed all sebaceous neoplasms contained a wild-type BRAF gene. Two (15%) of 13 patients with MTS were found to harbor a KRAS mutation and loss of MLH1 expression. We conclude that a V600E BRAF mutation may not be helpful in distinguishing sporadic from MTS-associated sebaceous neoplasms. Further studies are needed to determine if KRAS mutations are restricted to patients with MTS or are also present in sporadic sebaceous neoplasms. Copyright © 2014 Elsevier Inc. All rights reserved.

A single center analysis of nucleophosmin in acute myeloid leukemia: value of combining immunohistochemistry with molecular mutation analysis.

PubMed

Woolthuis, Carolien M; Mulder, André B; Verkaik-Schakel, Rikst Nynke; Rosati, Stefano; Diepstra, Arjan; van den Berg, Eva; Schuringa, Jan Jacob; Vellenga, Edo; Kluin, Philip M; Huls, Gerwin

2013-10-01

Mutations of nucleophosmin 1 are frequently found in acute myeloid leukemia and lead to aberrant cytoplasmic accumulation of nucleophosmin protein. Immunohistochemical staining is therefore recommended as the technique of choice in front-line screening. In this study, we assessed the sensitivity and specificity of immunohistochemistry on formalin-fixed bone marrow biopsies compared with gold standard molecular analysis to predict nucleophosmin 1 mutation status in 119 patients with acute myeloid leukemia. Discrepant cases were further characterized by gene expression analyses and fluorescence in situ hybridization. A large overlap between both methods was observed. Nevertheless, nine patients demonstrated discordant results at initial screening. Five cases demonstrated nuclear staining of nucleophosmin 1 by immunohistochemistry, but a nucleophosmin 1 mutation by molecular analysis. In two cases this could be attributed to technical issues and in three cases minor subpopulations of myeloblasts had not been discovered initially. All tested cases exhibited the characteristic nucleophosmin-mutated gene expression pattern. Four cases had cytoplasmic nucleophosmin 1 staining and a nucleophosmin-mutated gene expression pattern without a detectable nucleophosmin 1 mutation. In two of these cases we found the chromosomal translocation t(3;5)(q25;q35) encoding the NPM-MLF1 fusion protein. In the other discrepant cases the aberrant cytoplasmic nucleophosmin staining and gene expression could not be explained. In total six patients (5%) had true discordant results between immunohistochemistry and mutation analysis. We conclude that cytoplasmic nucleophosmin localization is not always caused by a conventional nucleophosmin 1 mutation and that in the screening for nucleophosmin 1 abnormalities, most information will be obtained by combining immunohistochemistry with molecular analysis.

A single center analysis of nucleophosmin in acute myeloid leukemia: value of combining immunohistochemistry with molecular mutation analysis

PubMed Central

Woolthuis, Carolien M.; Mulder, André B.; Verkaik-Schakel, Rikst Nynke; Rosati, Stefano; Diepstra, Arjan; van den Berg, Eva; Schuringa, Jan Jacob; Vellenga, Edo; Kluin, Philip M.; Huls, Gerwin

2013-01-01

Mutations of nucleophosmin 1 are frequently found in acute myeloid leukemia and lead to aberrant cytoplasmic accumulation of nucleophosmin protein. Immunohistochemical staining is therefore recommended as the technique of choice in front-line screening. In this study, we assessed the sensitivity and specificity of immunohistochemistry on formalin-fixed bone marrow biopsies compared with gold standard molecular analysis to predict nucleophosmin 1 mutation status in 119 patients with acute myeloid leukemia. Discrepant cases were further characterized by gene expression analyses and fluorescence in situ hybridization. A large overlap between both methods was observed. Nevertheless, nine patients demonstrated discordant results at initial screening. Five cases demonstrated nuclear staining of nucleophosmin 1 by immunohistochemistry, but a nucleophosmin 1 mutation by molecular analysis. In two cases this could be attributed to technical issues and in three cases minor subpopulations of myeloblasts had not been discovered initially. All tested cases exhibited the characteristic nucleophosmin-mutated gene expression pattern. Four cases had cytoplasmic nucleophosmin 1 staining and a nucleophosmin-mutated gene expression pattern without a detectable nucleophosmin 1 mutation. In two of these cases we found the chromosomal translocation t(3;5)(q25;q35) encoding the NPM-MLF1 fusion protein. In the other discrepant cases the aberrant cytoplasmic nucleophosmin staining and gene expression could not be explained. In total six patients (5%) had true discordant results between immunohistochemistry and mutation analysis. We conclude that cytoplasmic nucleophosmin localization is not always caused by a conventional nucleophosmin 1 mutation and that in the screening for nucleophosmin 1 abnormalities, most information will be obtained by combining immunohistochemistry with molecular analysis. PMID:23716555

Biallelic Mutations in MRPS34 Lead to Instability of the Small Mitoribosomal Subunit and Leigh Syndrome.

PubMed

Lake, Nicole J; Webb, Bryn D; Stroud, David A; Richman, Tara R; Ruzzenente, Benedetta; Compton, Alison G; Mountford, Hayley S; Pulman, Juliette; Zangarelli, Coralie; Rio, Marlene; Boddaert, Nathalie; Assouline, Zahra; Sherpa, Mingma D; Schadt, Eric E; Houten, Sander M; Byrnes, James; McCormick, Elizabeth M; Zolkipli-Cunningham, Zarazuela; Haude, Katrina; Zhang, Zhancheng; Retterer, Kyle; Bai, Renkui; Calvo, Sarah E; Mootha, Vamsi K; Christodoulou, John; Rötig, Agnes; Filipovska, Aleksandra; Cristian, Ingrid; Falk, Marni J; Metodiev, Metodi D; Thorburn, David R

2017-08-03

The synthesis of all 13 mitochondrial DNA (mtDNA)-encoded protein subunits of the human oxidative phosphorylation (OXPHOS) system is carried out by mitochondrial ribosomes (mitoribosomes). Defects in the stability of mitoribosomal proteins or mitoribosome assembly impair mitochondrial protein translation, causing combined OXPHOS enzyme deficiency and clinical disease. Here we report four autosomal-recessive pathogenic mutations in the gene encoding the small mitoribosomal subunit protein, MRPS34, in six subjects from four unrelated families with Leigh syndrome and combined OXPHOS defects. Whole-exome sequencing was used to independently identify all variants. Two splice-site mutations were identified, including homozygous c.321+1G>T in a subject of Italian ancestry and homozygous c.322-10G>A in affected sibling pairs from two unrelated families of Puerto Rican descent. In addition, compound heterozygous MRPS34 mutations were identified in a proband of French ancestry; a missense (c.37G>A [p.Glu13Lys]) and a nonsense (c.94C>T [p.Gln32 ∗ ]) variant. We demonstrated that these mutations reduce MRPS34 protein levels and the synthesis of OXPHOS subunits encoded by mtDNA. Examination of the mitoribosome profile and quantitative proteomics showed that the mitochondrial translation defect was caused by destabilization of the small mitoribosomal subunit and impaired monosome assembly. Lentiviral-mediated expression of wild-type MRPS34 rescued the defect in mitochondrial translation observed in skin fibroblasts from affected subjects, confirming the pathogenicity of MRPS34 mutations. Our data establish that MRPS34 is required for normal function of the mitoribosome in humans and furthermore demonstrate the power of quantitative proteomic analysis to identify signatures of defects in specific cellular pathways in fibroblasts from subjects with inherited disease. Copyright © 2017 American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.

Multiple molecular dynamics simulations of human LOX-1 and Trp150Ala mutant reveal the structural determinants causing the full deactivation of the receptor.

PubMed

Iacovelli, Federico; Tucci, Fabio Giovanni; Macari, Gabriele; Falconi, Mattia

2017-10-01

Multiple classical molecular dynamics simulations have been applied to the human LOX-1 receptor to clarify the role of the Trp150Ala mutation in the loss of binding activity. Results indicate that the substitution of this crucial residue, located at the dimer interface, markedly disrupts the wild-type receptor dynamics. The mutation causes an irreversible rearrangement of the subunits interaction pattern that in the wild-type protein allows the maintaining of a specific symmetrical motion of the monomers. The subunits dislocation determines a loss of linearity of the arginines residues composing the basic spine and a consequent alteration of the long-range electrostatic attraction of the substrate. Moreover, the anomalous subunits arrangement observed in the mutated receptor also affects the integrity of the hydrophobic tunnel, actively involved in the short-range hydrophobic recognition of the substrate. The combined effect of these structural rearrangements generates the impairing of the receptor function. © 2017 Wiley Periodicals, Inc.

Cellular and Circuitry Bases of Autism: Lessons Learned from the Temporospatial Manipulation of Autism Genes in the Brain.

PubMed

Hulbert, Samuel W; Jiang, Yong-Hui

2017-04-01

Transgenic mice carrying mutations that cause Autism Spectrum Disorders (ASDs) continue to be valuable for determining the molecular underpinnings of the disorders. Recently, researchers have taken advantage of such models combined with Cre-loxP and similar systems to manipulate gene expression over space and time. Thus, a clearer picture is starting to emerge of the cell types, circuits, brain regions, and developmental time periods underlying ASDs. ASD-causing mutations have been restricted to or rescued specifically in excitatory or inhibitory neurons, different neurotransmitter systems, and cells specific to the forebrain or cerebellum. In addition, mutations have been induced or corrected in adult mice, providing some evidence for the plasticity and reversibility of core ASD symptoms. The limited availability of Cre lines that are highly specific to certain cell types or time periods provides a challenge to determining the cellular and circuitry bases of autism, but other technological advances may eventually overcome this obstacle.

Molecular analysis of brazilian patients with combined pituitary hormone deficiency and orthotopic posterior pituitary lobe reveals eight different PROP1 alterations with three novel mutations.

PubMed

Madeira, Joao Lo; Nishi, Mirian Y; Nakaguma, Marilena; Benedetti, Anna F; Biscotto, Isabela Peixoto; Fernandes, Thamiris; Pequeno, Thiago; Figueiredo, Thalita; Franca, Marcela M; Correa, Fernanda A; Otto, Aline P; Abrão, Milena; Miras, Mirta B; Santos, Silvana; Jorge, Alexander Al; Costalonga, Everlayny F; Mendonca, Berenice B; Arnhold, Ivo Jp; Carvalho, Luciani R

2017-12-01

Mutations in PROP1, HESX1 and LHX3 are associated with combined pituitary hormone deficiency (CPHD) and orthotopic posterior pituitary lobe (OPP). To identify mutations in PROP1, HESX1 and LHX3 in a large cohort of patients with CPHD and OPP (35 Brazilian, two Argentinian). We studied 23 index patients with CPHD and OPP (six familial and 17 sporadic) as well as 14 relatives. PROP1 was sequenced by the Sanger method in all except one sporadic case studied using a candidate gene panel. Multiplex ligation-dependent probe amplification (MLPA) was applied to one familial case in whom PROP1 failed to amplify by PCR. In the 13 patients without PROP1 mutations, HESX1 and LHX3 were sequenced by the Sanger method. We identified PROP1 mutations in 10 index cases. Three mutations were novel: one affecting the initiation codon (c.1A>G) and two affecting splicing sites, c.109+1G>A and c.342+1G>C. The known mutations, c.150delA (p.Arg53Aspfs*112), c.218G>A (p.Arg73His), c.263T>C (p.Phe88Ser) and c.301_302delAG (p.Leu102Cysfs*8), were also detected. MLPA confirmed complete PROP1 deletion in one family. We did not identify HESX1 and LHX3 mutations by Sanger. PROP1 mutations are a prevalent cause of congenital CPHD with OPP, and therefore, PROP1 sequencing must be the first step of molecular investigation in patients with CPHD and OPP, especially in populations with a high frequency of PROP1 mutations. In the absence of mutations, massively parallel sequencing is a promising approach. The high prevalence and diversity of PROP1 mutations is associated with the ethnic background of this cohort. © 2017 John Wiley & Sons Ltd.

CFTR allelic heterogeneity in Mexican patients with cystic fibrosis: implications for molecular screening.

PubMed

Chávez-Saldaña, Margarita; Yokoyama, Emiy; Lezana, José Luis; Carnevale, Alessandra; Macías, Miguel; Vigueras, Rosa M; López, Marisol; Orozco, Lorena

2010-01-01

Cystic fibrosis, the most common autosomal recessive disorder, is caused by defects in the CF transmembrane conductance regulator gene (CFTR) that encodes a chloride channel. To date, over 1,800 mutations have been described related to the causative gene of CF, showing a variable frequency among populations. In a previous extensive analysis of the CFTR locus in 97 Mexican patients, 34 different mutations (75% of CF alleles) were found using several strategies for mutation screening; however, 63% had at least an uncharacterized allele. Despite the combined technologies used, there are still a great number of unknown mutations in the Mexican population. Screening of the CFTR gene to provide additional evidence of the mutational wide spectrum responsible for CF in Mexican patients. In this study, the number of unrelated CF patients was increased to 230, 133 new cases and the 97 previously reported to include 63% with at least an uncharacterized allele. Additional tools were used to improve the detection rate of CF mutations, such as a commercial kit for 36 mutations plus a single chain conformational polymorphism method and DNA sequencing. By using a combination of these strategies we characterized 77.7% of all the CF alleles, resulting in a total of 46 different mutations detected, including the identification of 12 additional mutations (p.R334W, p.A455E, c.3120+1G > A, c.3272-26A > G, c.711+1G > T, p.Q552X, p.W1282X, c.IVS8-5T, p.R1162X and p.R347P, p.D1152H and p.T1036N). Although these 12 mutations have been reported in other populations, they have not yet been reported in Mexican patients. This report shows that Mexico has one of the widest spectra of CFTR mutations worldwide. The knowledge of the ethnic and geographic distribution of CFTR mutations in this population will allow the development of more effective methods for diagnosis and treatment.

Genetic changes that accompanied shifts of low pathogenic avian influenza viruses toward higher pathogenicity in poultry

PubMed Central

Abdelwhab, El-Sayed M; Veits, Jutta; Mettenleiter, Thomas C

2013-01-01

Avian influenza viruses (AIV) of H5 and H7 subtypes exhibit two different pathotypes in poultry: infection with low pathogenic (LP) strains results in minimal, if any, health disturbances, whereas highly pathogenic (HP) strains cause severe morbidity and mortality. LPAIV of H5 and H7 subtypes can spontaneously mutate into HPAIV. Ten outbreaks caused by HPAIV are known to have been preceded by circulation of a predecessor LPAIV in poultry. Three of them were caused by H5N2 subtype and seven involved H7 subtype in combination with N1, N3, or N7. Here, we review those outbreaks and summarize the genetic changes which resulted in the transformation of LPAIV to HPAIV under natural conditions. Mutations that were found directly in those outbreaks are more likely to be linked to virulence, pathogenesis, and early adaptation of AIV. PMID:23863606

A Disease Mutation Causing Episodic Ataxia Type I in the S1 Links Directly to the Voltage Sensor and the Selectivity Filter in Kv Channels.

PubMed

Petitjean, Dimitri; Kalstrup, Tanja; Zhao, Juan; Blunck, Rikard

2015-09-02

The mutation F184C in Kv1.1 leads to development of episodic ataxia type I (EA1). Although the mutation has been said to alter activation kinetics and to lower expression, we show here that the underlying molecular mechanisms may be more complex. Although F184 is positioned in the "peripheral" S1 helix, it occupies a central position in the 3D fold. We show in cut-open oocyte voltage-clamp recordings of gating and ionic currents of the Shaker Kv channel expressed in Xenopus oocytes that F184 not only interacts directly with the gating charges of the S4, but also creates a functional link to the selectivity filter of the neighboring subunit. This link leads to impaired fast and slow inactivation. The effect on fast inactivation is of an allosteric nature considering that fast inactivation is caused by a linked cytosolic ball peptide. The extensive effects of F184C provide a new mechanism underlying EA. Episodic ataxia (EA) is an inherited disease that leads to occasional loss of motor control in combination with variable other symptoms such as vertigo or migraine. EA type I (EA1), studied here, is caused by mutations in a voltage-gated potassium channel that contributes to the generation of electrical signals in the brain. The mechanism by which mutations in voltage-gated potassium channels lead to EA is still unknown and there is no consistent pharmacological treatment. By studying in detail one disease-causing mutation in Kv1.1, we describe a novel molecular mechanism distinct from mechanisms described previously. This mechanism contributes to the understanding of potassium channel function in general and might lead to a better understanding of how EA develops. Copyright © 2015 the authors 0270-6474/15/3512198-09$15.00/0.

Cochlear Implantation in Siblings With Refsum's Disease.

PubMed

Stähr, Kerstin; Kuechler, Alma; Gencik, Martin; Arnolds, Judith; Dendy, Meaghan; Lang, Stephan; Arweiler-Harbeck, Diana

2017-08-01

Whether the origin of severe hearing loss in Refsum's syndrome is caused by cochlear impairment or retrocochlear degeneration remains unclear. This case report aims to investigate hearing performance before and after cochlear implantation to shed light on this question. Also, identification of new mutations causing Refsum's syndrome would be helpful in generating additional means of diagnosis. A family of 4 individuals was subjected to genetic testing. Two siblings (56 and 61 years old) suffered from severe hearing and vision loss and received bilateral cochlear implants. Genetic analysis, audiological outcome, and clinical examinations were performed. One new mutation in the PHYH gene (c.768del63bp) causing Refsum's disease was found. Preoperative distortion product otoacoustic emissions (DPAOEs) were absent. Postoperative speech perception in Freiburger speech test was 100% for bisyllabic words and 85% (patient No. 1) and 65% (patient No. 2), respectively, for monosyllabic words. Five years after implantation, speech perception remained stable for bisyllabic words but showed decreasing capabilities for monosyllabic words. A new mutation causing Refsum's disease is presented. Cochlear implantation in case of severe hearing loss leads to an improvement in speech perception and should be recommended for patients with Refsum's disease, especially when the hearing loss is combined with a severe loss of vision. Decrease of speech perception in the long-term follow-up could indicate an additional retrocochlear degeneration.

Optimising the combination dosing strategy of abemaciclib and vemurafenib in BRAF-mutated melanoma xenograft tumours

PubMed Central

Tate, Sonya C; Burke, Teresa F; Hartman, Daisy; Kulanthaivel, Palaniappan; Beckmann, Richard P; Cronier, Damien M

2016-01-01

Background: Resistance to BRAF inhibition is a major cause of treatment failure for BRAF-mutated metastatic melanoma patients. Abemaciclib, a cyclin-dependent kinase 4 and 6 inhibitor, overcomes this resistance in xenograft tumours and offers a promising drug combination. The present work aims to characterise the quantitative pharmacology of the abemaciclib/vemurafenib combination using a semimechanistic pharmacokinetic/pharmacodynamic modelling approach and to identify an optimum dosing regimen for potential clinical evaluation. Methods: A PK/biomarker model was developed to connect abemaciclib/vemurafenib concentrations to changes in MAPK and cell cycle pathway biomarkers in A375 BRAF-mutated melanoma xenografts. Resultant tumour growth inhibition was described by relating (i) MAPK pathway inhibition to apoptosis, (ii) mitotic cell density to tumour growth and, under resistant conditions, (iii) retinoblastoma protein inhibition to cell survival. Results: The model successfully described vemurafenib/abemaciclib-mediated changes in MAPK pathway and cell cycle biomarkers. Initial tumour shrinkage by vemurafenib, acquisition of resistance and subsequent abemaciclib-mediated efficacy were successfully captured and externally validated. Model simulations illustrate the benefit of intermittent vemurafenib therapy over continuous treatment, and indicate that continuous abemaciclib in combination with intermittent vemurafenib offers the potential for considerable tumour regression. Conclusions: The quantitative pharmacology of the abemaciclib/vemurafenib combination was successfully characterised and an optimised, clinically-relevant dosing strategy was identified. PMID:26978007

High-resolution melting analysis for prenatal diagnosis of beta-thalassemia in northern Thailand.

PubMed

Charoenkwan, Pimlak; Sirichotiyakul, Supatra; Phusua, Arunee; Suanta, Sudjai; Fanhchaksai, Kanda; Sae-Tung, Rattika; Sanguansermsri, Torpong

2017-12-01

High-resolution melting (HRM) analysis is a rapid mutation analysis which assesses the pattern of reduction of fluorescence signal after subjecting the amplified PCR product with saturated fluorescence dye to an increasing temperature. We used HRM analysis for prenatal diagnosis of beta-thalassemia disease in northern Thailand. Five PCR-HRM protocols were used to detect point mutations in five different segments of the beta-globin gene, and one protocol to detect the 3.4 kb beta-globin deletion. We sought to characterize the mutations in carriers and to enable prenatal diagnosis in 126 couples at risk of having a fetus with beta-thalassemia disease. The protocols identified 18 common mutations causing beta-thalassemia, including the rare codon 132 (A-T) mutation. Each mutation showed a specific HRM pattern and all results were in concordance with those from direct DNA sequencing or gap-PCR methods. In cases of beta-thalassemia disease resulting from homozygosity for a mutation or compound heterozygosity for two mutations on the same amplified segment, the HRM patterns were different to those of a single mutation and were specific for each combination. HRM analysis is a simple and useful method for mutation identification in beta-thalassemia carriers and prenatal diagnosis of beta-thalassemia in northern Thailand.

Novel myosin mutations for hereditary hearing loss revealed by targeted genomic capture and massively parallel sequencing

PubMed Central

Brownstein, Zippora; Abu-Rayyan, Amal; Karfunkel-Doron, Daphne; Sirigu, Serena; Davidov, Bella; Shohat, Mordechai; Frydman, Moshe; Houdusse, Anne; Kanaan, Moien; Avraham, Karen B

2014-01-01

Hereditary hearing loss is genetically heterogeneous, with a large number of genes and mutations contributing to this sensory, often monogenic, disease. This number, as well as large size, precludes comprehensive genetic diagnosis of all known deafness genes. A combination of targeted genomic capture and massively parallel sequencing (MPS), also referred to as next-generation sequencing, was applied to determine the deafness-causing genes in hearing-impaired individuals from Israeli Jewish and Palestinian Arab families. Among the mutations detected, we identified nine novel mutations in the genes encoding myosin VI, myosin VIIA and myosin XVA, doubling the number of myosin mutations in the Middle East. Myosin VI mutations were identified in this population for the first time. Modeling of the mutations provided predicted mechanisms for the damage they inflict in the molecular motors, leading to impaired function and thus deafness. The myosin mutations span all regions of these molecular motors, leading to a wide range of hearing phenotypes, reinforcing the key role of this family of proteins in auditory function. This study demonstrates that multiple mutations responsible for hearing loss can be identified in a relatively straightforward manner by targeted-gene MPS technology and concludes that this is the optimal genetic diagnostic approach for identification of mutations responsible for hearing loss. PMID:24105371

Cri du Chat Syndrome and Primary Ciliary Dyskinesia: A Common Genetic Cause on Chromosome 5p

PubMed Central

Shapiro, Adam J.; Weck, Karen E.; Chao, Kay C.; Rosenfeld, Margaret; Nygren, Anders O. H.; Knowles, Michael R.; Leigh, Margaret W.; Zariwala, Maimoona A.

2014-01-01

Cri du chat syndrome (CdCS) and primary ciliary dyskinesia (PCD) are rare diseases that present with frequent respiratory symptoms. PCD can be caused by hemizygous DNAH5 mutation in combination with a 5p segmental deletion attributable to CdCS on the opposite chromosome. Chronic oto-sino-pulmonary symptoms or organ laterality defects in CdCS should prompt an evaluation for PCD. PMID:25066065

Mutation in WDR4 impairs tRNA m(7)G46 methylation and causes a distinct form of microcephalic primordial dwarfism.

PubMed

Shaheen, Ranad; Abdel-Salam, Ghada M H; Guy, Michael P; Alomar, Rana; Abdel-Hamid, Mohamed S; Afifi, Hanan H; Ismail, Samira I; Emam, Bayoumi A; Phizicky, Eric M; Alkuraya, Fowzan S

2015-09-28

Primordial dwarfism is a state of extreme prenatal and postnatal growth deficiency, and is characterized by marked clinical and genetic heterogeneity. Two presumably unrelated consanguineous families presented with an apparently novel form of primordial dwarfism in which severe growth deficiency is accompanied by distinct facial dysmorphism, brain malformation (microcephaly, agenesis of corpus callosum, and simplified gyration), and severe encephalopathy with seizures. Combined autozygome/exome analysis revealed a novel missense mutation in WDR4 as the likely causal variant. WDR4 is the human ortholog of the yeast Trm82, an essential component of the Trm8/Trm82 holoenzyme that effects a highly conserved and specific (m(7)G46) methylation of tRNA. The human mutation and the corresponding yeast mutation result in a significant reduction of m(7)G46 methylation of specific tRNA species, which provides a potential mechanism for primordial dwarfism associated with this lesion, since reduced m(7)G46 modification causes a growth deficiency phenotype in yeast. Our study expands the number of biological pathways underlying primordial dwarfism and adds to a growing list of human diseases linked to abnormal tRNA modification.

An engineered tale-transcription factor rescues transcription of factor VII impaired by promoter mutations and enhances its endogenous expression in hepatocytes.

PubMed

Barbon, Elena; Pignani, Silvia; Branchini, Alessio; Bernardi, Francesco; Pinotti, Mirko; Bovolenta, Matteo

2016-06-24

Tailored approaches to restore defective transcription responsible for severe diseases have been poorly explored. We tested transcription activator-like effectors fused to an activation domain (TALE-TFs) in a coagulation factor VII (FVII) deficiency model. In this model, the deficiency is caused by the -94C > G or -61T > G mutation, which abrogate the binding of Sp1 or HNF-4 transcription factors. Reporter assays in hepatoma HepG2 cells naturally expressing FVII identified a single TALE-TF (TF4) that, by targeting the region between mutations, specifically trans-activated both the variant (>100-fold) and wild-type (20-40-fold) F7 promoters. Importantly, in the genomic context of transfected HepG2 and transduced primary hepatocytes, TF4 increased F7 mRNA and protein levels (2- to 3-fold) without detectable off-target effects, even for the homologous F10 gene. The ectopic F7 expression in renal HEK293 cells was modestly affected by TF4 or by TALE-TF combinations. These results provide experimental evidence for TALE-TFs as gene-specific tools useful to counteract disease-causing promoter mutations.

A distal point mutation in the streptavidin-biotin complex preserves structure but diminishes binding affinity: experimental evidence of electronic polarization effects?

PubMed

Baugh, Loren; Le Trong, Isolde; Cerutti, David S; Gülich, Susanne; Stayton, Patrick S; Stenkamp, Ronald E; Lybrand, Terry P

2010-06-08

We have identified a distal point mutation in streptavidin that causes a 1000-fold reduction in biotin binding affinity without disrupting the equilibrium complex structure. The F130L mutation creates a small cavity occupied by a water molecule; however, all neighboring side chain positions are preserved, and protein-biotin hydrogen bonds are unperturbed. Molecular dynamics simulations reveal a reduced mobility of biotin binding residues but no observable destabilization of protein-ligand interactions. Our combined structural and computational studies suggest that the additional water molecule may affect binding affinity through an electronic polarization effect that impacts the highly cooperative hydrogen bonding network in the biotin binding pocket.

Single-nucleotide variants in two Hedgehog genes, SHH and HHIP, as genetic cause of combined pituitary hormone deficiency.

PubMed

Gorbenko del Blanco, Darya; de Graaff, Laura C G; Visser, Theo J; Hokken-Koelega, Anita C S

2013-03-01

Combined pituitary hormone deficiency (CPHD) is characterized by deficiencies of two or more anterior pituitary hormones. Its genetic cause is unknown in the majority of cases. The Hedgehog (Hh) signalling pathway has been implicated in disorders associated with pituitary development. Mutations in Sonic Hedgehog (SHH) have been described in patients with holoprosencephaly (with or without pituitary involvement). Hedgehog interacting protein (HHIP) has been associated with variations in adult height in genome wide association studies. We investigated whether mutations in these two genes of the Hh pathway, SHH and HHIP, could result in 'idiopathic' CPHD. We directly sequenced the coding regions and exon - intron boundaries of SHH and HHIP in 93 CPHD patients of the Dutch HYPOPIT study in whom mutations in the classical CPHD genes PROP1, POU1F1, HESX1, LHX3 and LHX4 had been ruled out. We compared the expression of Hh genes in Hep3B transfected cells between wild-type proteins and mutants. We identified three single-nucleotide variants (p.Ala226Thr, c.1078C>T and c.*8G>T) in SHH. The function of the latter was severely affected in our in vitro assay. In HHIP, we detected a new activating variant c.-1G>C, which increases HHIP's inhibiting function on the Hh pathway. Our results suggest involvement of the Hedgehog pathway in CPHD. We suggest that both SHH and HHIP are investigated as a second screening in CPHD, after mutations in the classical CPHD genes have been ruled out. © 2012 Blackwell Publishing Ltd.

The Genetic Inheritance of the Blue-eyed White Phenotype in Alpacas (Vicugna pacos)

PubMed Central

Johnson, Warren E.; Appleton, Belinda R.

2014-01-01

White-spotting patterns in mammals can be caused by mutations in the gene KIT, whose protein is necessary for the normal migration and survival of melanocytes from the neural crest. The alpaca (Vicugna pacos) blue-eyed white (BEW) phenotype is characterized by 2 blue eyes and a solid white coat over the whole body. Breeders hypothesize that the BEW phenotype in alpacas is caused by the combination of the gene causing gray fleece and a white-spotting gene. We performed an association study using KIT flanking and intragenic markers with 40 unrelated alpacas, of which 17 were BEW. Two microsatellite alleles at KIT-related markers were significantly associated (P < 0.0001) with the BEW phenotype (bew1 and bew2). In a larger cohort of 171 related individuals, we identify an abundance of an allele (bew1) in gray animals and the occurrence of bew2 homozygotes that are solid white with pigmented eyes. Association tests accounting for population structure and familial relatedness are consistent with a proposed model where these alleles are in linkage disequilibrium with a mutation or mutations that contribute to the BEW phenotype and to individual differences in fleece color. PMID:23144493

Mutation of Hip’s Carboxy-Terminal Region Inhibits a Transitional Stage of Progesterone Receptor Assembly

PubMed Central

Prapapanich, Viravan; Chen, Shiying; Smith, David F.

1998-01-01

Steroid receptor complexes are assembled through an ordered, multistep pathway involving multiple components of the cytoplasmic chaperone machinery. Two of these components are Hsp70-binding proteins, Hip and Hop, that have some limited homology in their C-terminal regions, outside the sequences mapped for Hsp70 binding. Within this region of Hip is a DPEV sequence that occurs twice; in Hop, one DPEV sequence plus a partial second sequence occurs. In an effort to better understand Hip function as it relates to assembly of progesterone receptor complexes, the DPEV region of Hip was targeted for mutations. Each DPEV sequence was mutated to an APAV sequence, singly or in combination. The combined mutation, APAV2, was further combined with a deletion of Hip’s tetratricopeptide repeat region that is required for Hsp70 binding or with a deletion of Hip’s GGMP repeat. An additional mutant was prepared by truncation of Hip’s DPEV-containing C terminus. By comparing interactions of various Hip forms with Hsp70, it was determined that mutation of the DPEV sequences created a dominant inhibitory form of Hip. The mutant Hip-Hsp70 complex was not prevented from interacting with progesterone receptor, but the mutant caused a dose-dependent inhibition of receptor assembly with Hsp90. The behavior of the Hip mutant is consistent with a model in which Hip and Hop are required to facilitate the transition from an early receptor complex with Hsp70 into later complexes containing Hsp90. PMID:9447991

A reversion of an IL2RG mutation in combined immunodeficiency providing competitive advantage to the majority of CD8+ T cells

PubMed Central

Kuijpers, Taco W.; van Leeuwen, Ester M.M.; Barendregt, Barbara H.; Klarenbeek, Paul; aan de Kerk, Daan J.; Baars, Paul A.; Jansen, Machiel H.; de Vries, Niek; van Lier, René A.W.; van der Burg, Mirjam

2013-01-01

Mutations in the common gamma chain (γc, CD132, encoded by the IL2RG gene) can lead to B+T−NK− X-linked severe combined immunodeficiency, as a consequence of unresponsiveness to γc-cytokines such as interleukins-2, -7 and -15. Hypomorphic mutations in CD132 may cause combined immunodeficiencies with a variety of clinical presentations. We analyzed peripheral blood mononuclear cells of a 6-year-old boy with normal lymphocyte counts, who suffered from recurrent pneumonia and disseminated mollusca contagiosa. Since proliferative responses of T cells and NK cells to γc -cytokines were severely impaired, we performed IL2RG gene analysis, showing a heterozygous mutation in the presence of a single X-chromosome. Interestingly, an IL2RG reversion to normal predominated in both naïve and antigen-primed CD8+ T cells and increased over time. Only the revertant CD8+ T cells showed normal expression of CD132 and the various CD8+ T cell populations had a different T-cell receptor repertoire. Finally, a fraction of γδ+ T cells and differentiated CD4+CD27− effector-memory T cells carried the reversion, whereas NK or B cells were repeatedly negative. In conclusion, in a patient with a novel IL2RG mutation, gene-reverted CD8+ T cells accumulated over time. Our data indicate that selective outgrowth of particular T-cell subsets may occur following reversion at the level of committed T progenitor cells. PMID:23403317

A reversion of an IL2RG mutation in combined immunodeficiency providing competitive advantage to the majority of CD8+ T cells.

PubMed

Kuijpers, Taco W; van Leeuwen, Ester M M; Barendregt, Barbara H; Klarenbeek, Paul; aan de Kerk, Daan J; Baars, Paul A; Jansen, Machiel H; de Vries, Niek; van Lier, René A W; van der Burg, Mirjam

2013-07-01

Mutations in the common gamma chain (γc, CD132, encoded by the IL2RG gene) can lead to B(+)T(-)NK(-) X-linked severe combined immunodeficiency, as a consequence of unresponsiveness to γc-cytokines such as interleukins-2, -7 and -15. Hypomorphic mutations in CD132 may cause combined immunodeficiencies with a variety of clinical presentations. We analyzed peripheral blood mononuclear cells of a 6-year-old boy with normal lymphocyte counts, who suffered from recurrent pneumonia and disseminated mollusca contagiosa. Since proliferative responses of T cells and NK cells to γc -cytokines were severely impaired, we performed IL2RG gene analysis, showing a heterozygous mutation in the presence of a single X-chromosome. Interestingly, an IL2RG reversion to normal predominated in both naïve and antigen-primed CD8(+) T cells and increased over time. Only the revertant CD8(+) T cells showed normal expression of CD132 and the various CD8(+) T cell populations had a different T-cell receptor repertoire. Finally, a fraction of γδ(+) T cells and differentiated CD4(+)CD27(-) effector-memory T cells carried the reversion, whereas NK or B cells were repeatedly negative. In conclusion, in a patient with a novel IL2RG mutation, gene-reverted CD8(+) T cells accumulated over time. Our data indicate that selective outgrowth of particular T-cell subsets may occur following reversion at the level of committed T progenitor cells.

Mitochondrial tRNALeu(UUR) C3275T, tRNAGln T4363C and tRNALys A8343G mutations may be associated with PCOS and metabolic syndrome.

PubMed

Ding, Yu; Xia, Bo-Hou; Zhang, Cai-Juan; Zhuo, Guang-Chao

2018-02-05

Polycystic ovary syndrome (PCOS) is a very prevalent endocrine disease affecting reproductive women. Clinically, patients with this disorder are more vulnerable to develop type 2 diabetes mellitus (T2DM), cardiovascular events, as well as metabolic syndrome (MetS). To date, the molecular mechanism underlying PCOS remains largely unknown. Previously, we showed that mitochondrial dysfunction caused by mitochondrial DNA (mtDNA) mutation was an important cause for PCOS. In the current study, we described the clinical and biochemical features of a three-generation pedigree with maternally transmitted MetS, combined with PCOS. A total of three matrilineal relatives exhibited MetS including obesity, high triglyceride (TG) and Hemoglobin A1c (HbA1c) levels, and hypertension. Whereas one patient from the third generation manifestated PCOS. Mutational analysis of the whole mitochondrial genes from the affected individuals identified a set of genetic variations belonging to East Asia haplogroup B4b1c. Among these variants, the homoplasmic C3275T mutation disrupted a highly evolutionary conserved base-pairing (28A-46C) on the variable region of tRNA Leu(UUR) , whereas the T4363C mutation created a new base-pairing (31T-37A) in the anticodon stem of tRNA Gln , furthermore, the A8343G mutation occurred at the very conserved position of tRNA Lys and may result the failure in mitochondrial tRNAs (mt-tRNAs) metabolism. Biochemical analysis revealed the deficiency in mitochondrial functions including lower levels of mitochondrial membrane potential (MMP), ATP production and mtDNA copy number, while a significantly increased reactive oxygen species (ROS) generation was observed in polymononuclear leukocytes (PMNs) from the individuals carrying these mt-tRNA mutations, suggesting that these mutations may cause mitochondrial dysfunction that was responsible for the clinical phenotypes. Taken together, our data indicated that mt-tRNA mutations were associated with MetS and PCOS in this family, which shaded additional light into the pathophysiology of PCOS that were manifestated by mitochondrial dysfunction. Copyright © 2017 Elsevier B.V. All rights reserved.

Explosive mutation accumulation triggered by heterozygous human Pol ε proofreading-deficiency is driven by suppression of mismatch repair

PubMed Central

Campbell, Brittany B; Ungerleider, Nathan; Light, Nicholas; Wu, Tong; LeCompte, Kimberly G; Goksenin, A Yasemin; Bunnell, Bruce A; Tabori, Uri; Shlien, Adam

2018-01-01

Tumors defective for DNA polymerase (Pol) ε proofreading have the highest tumor mutation burden identified. A major unanswered question is whether loss of Pol ε proofreading by itself is sufficient to drive this mutagenesis, or whether additional factors are necessary. To address this, we used a combination of next generation sequencing and in vitro biochemistry on human cell lines engineered to have defects in Pol ε proofreading and mismatch repair. Absent mismatch repair, monoallelic Pol ε proofreading deficiency caused a rapid increase in a unique mutation signature, similar to that observed in tumors from patients with biallelic mismatch repair deficiency and heterozygous Pol ε mutations. Restoring mismatch repair was sufficient to suppress the explosive mutation accumulation. These results strongly suggest that concomitant suppression of mismatch repair, a hallmark of colorectal and other aggressive cancers, is a critical force for driving the explosive mutagenesis seen in tumors expressing exonuclease-deficient Pol ε. PMID:29488881

Evolution of high-level resistance during low-level antibiotic exposure.

PubMed

Wistrand-Yuen, Erik; Knopp, Michael; Hjort, Karin; Koskiniemi, Sanna; Berg, Otto G; Andersson, Dan I

2018-04-23

It has become increasingly clear that low levels of antibiotics present in many environments can select for resistant bacteria, yet the evolutionary pathways for resistance development during exposure to low amounts of antibiotics remain poorly defined. Here we show that Salmonella enterica exposed to sub-MIC levels of streptomycin evolved high-level resistance via novel mechanisms that are different from those observed during lethal selections. During lethal selection only rpsL mutations are found, whereas at sub-MIC selection resistance is generated by several small-effect resistance mutations that combined confer high-level resistance via three different mechanisms: (i) alteration of the ribosomal RNA target (gidB mutations), (ii) reduction in aminoglycoside uptake (cyoB, nuoG, and trkH mutations), and (iii) induction of the aminoglycoside-modifying enzyme AadA (znuA mutations). These results demonstrate how the strength of the selective pressure influences evolutionary trajectories and that even weak selective pressures can cause evolution of high-level resistance.

Detection of c. -32T>G (IVS1-13T>G) mutation of Pompe disease by real-time PCR in dried blood spot specimen.

PubMed

Bobillo Lobato, Joaquin; Sánchez Peral, Blas A; Durán Parejo, Pilar; Jiménez Jiménez, Luis M

2013-03-15

Pompe disease, or acid maltase deficiency, is a genetic muscle disorder caused by mutations in the gene encoding the acid alpha-glucosidase (GAA) enzyme, which is essential for the degradation of glycogen to glucose in lysosomes. The wide clinical variability is resulted from genetic heterogeneity, and many different mutations of the GAA gene have been reported. Some of these mutations are associated with specific phenotypes, such as the c. -32T>G (IVS1-13T>G) mutation seen in late-onset Pompe disease. We used a real-time PCR, after genomic DNA extraction isolated from DBS (dried blood spots) and PCR amplification. Our results successfully detected in controls and patients have been 100% concordant with sequencing results. This assay combines simple sample processing and rapid analysis and it allows to detect the patients with a milder form and slower progression of this disease with a high reliability. Copyright © 2013 Elsevier B.V. All rights reserved.

Explosive mutation accumulation triggered by heterozygous human Pol ε proofreading-deficiency is driven by suppression of mismatch repair.

PubMed

Hodel, Karl P; de Borja, Richard; Henninger, Erin E; Campbell, Brittany B; Ungerleider, Nathan; Light, Nicholas; Wu, Tong; LeCompte, Kimberly G; Goksenin, A Yasemin; Bunnell, Bruce A; Tabori, Uri; Shlien, Adam; Pursell, Zachary F

2018-02-28

Tumors defective for DNA polymerase (Pol) ε proofreading have the highest tumor mutation burden identified. A major unanswered question is whether loss of Pol ε proofreading by itself is sufficient to drive this mutagenesis, or whether additional factors are necessary. To address this, we used a combination of next generation sequencing and in vitro biochemistry on human cell lines engineered to have defects in Pol ε proofreading and mismatch repair. Absent mismatch repair, monoallelic Pol ε proofreading deficiency caused a rapid increase in a unique mutation signature, similar to that observed in tumors from patients with biallelic mismatch repair deficiency and heterozygous Pol ε mutations. Restoring mismatch repair was sufficient to suppress the explosive mutation accumulation. These results strongly suggest that concomitant suppression of mismatch repair, a hallmark of colorectal and other aggressive cancers, is a critical force for driving the explosive mutagenesis seen in tumors expressing exonuclease-deficient Pol ε. © 2018, Hodel et al.

Analysis of Transcription Factors Key for Mouse Pancreatic Development Establishes NKX2-2 and MNX1 Mutations as Causes of Neonatal Diabetes in Man

PubMed Central

Flanagan, Sarah E.; De Franco, Elisa; Lango Allen, Hana; Zerah, Michele; Abdul-Rasoul, Majedah M.; Edge, Julie A.; Stewart, Helen; Alamiri, Elham; Hussain, Khalid; Wallis, Sam; de Vries, Liat; Rubio-Cabezas, Oscar; Houghton, Jayne A.L.; Edghill, Emma L.; Patch, Ann-Marie; Ellard, Sian; Hattersley, Andrew T.

2014-01-01

Summary Understanding transcriptional regulation of pancreatic development is required to advance current efforts in developing beta cell replacement therapies for patients with diabetes. Current knowledge of key transcriptional regulators has predominantly come from mouse studies, with rare, naturally occurring mutations establishing their relevance in man. This study used a combination of homozygosity analysis and Sanger sequencing in 37 consanguineous patients with permanent neonatal diabetes to search for homozygous mutations in 29 transcription factor genes important for murine pancreatic development. We identified homozygous mutations in 7 different genes in 11 unrelated patients and show that NKX2-2 and MNX1 are etiological genes for neonatal diabetes, thus confirming their key role in development of the human pancreas. The similar phenotype of the patients with recessive mutations and mice with inactivation of a transcription factor gene support there being common steps critical for pancreatic development and validate the use of rodent models for beta cell development. PMID:24411943

Structural Mechanism of the Pan-BCR-ABL Inhibitor Ponatinib (AP24534): Lessons for Overcoming Kinase Inhibitor Resistance

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhou, Tianjun; Commodore, Lois; Huang, Wei-Sheng

2012-01-20

The BCR-ABL inhibitor imatinib has revolutionized the treatment of chronic myeloid leukemia. However, drug resistance caused by kinase domain mutations has necessitated the development of new mutation-resistant inhibitors, most recently against the T315I gatekeeper residue mutation. Ponatinib (AP24534) inhibits both native and mutant BCR-ABL, including T315I, acting as a pan-BCR-ABL inhibitor. Here, we undertook a combined crystallographic and structure-activity relationship analysis on ponatinib to understand this unique profile. While the ethynyl linker is a key inhibitor functionality that interacts with the gatekeeper, virtually all other components of ponatinib play an essential role in its T315I inhibitory activity. The extensive networkmore » of optimized molecular contacts found in the DFG-out binding mode leads to high potency and renders binding less susceptible to disruption by single point mutations. The inhibitory mechanism exemplified by ponatinib may have broad relevance to designing inhibitors against other kinases with mutated gatekeeper residues.« less

Diminishing-returns epistasis decreases adaptability along an evolutionary trajectory.

PubMed

Wünsche, Andrea; Dinh, Duy M; Satterwhite, Rebecca S; Arenas, Carolina Diaz; Stoebel, Daniel M; Cooper, Tim F

2017-03-01

Populations evolving in constant environments exhibit declining adaptability. Understanding the basis of this pattern could reveal underlying processes determining the repeatability of evolutionary outcomes. In principle, declining adaptability can be due to a decrease in the effect size of beneficial mutations, a decrease in the rate at which they occur, or some combination of both. By evolving Escherichia coli populations started from different steps along a single evolutionary trajectory, we show that declining adaptability is best explained by a decrease in the size of available beneficial mutations. This pattern reflected the dominant influence of negative genetic interactions that caused new beneficial mutations to confer smaller benefits in fitter genotypes. Genome sequencing revealed that starting genotypes that were more similar to one another did not exhibit greater similarity in terms of new beneficial mutations, supporting the view that epistasis acts globally, having a greater influence on the effect than on the identity of available mutations along an adaptive trajectory. Our findings provide support for a general mechanism that leads to predictable phenotypic evolutionary trajectories.

Clinical problems of colorectal cancer and endometrial cancer cases with unknown cause of tumor mismatch repair deficiency (suspected Lynch syndrome).

PubMed

Buchanan, Daniel D; Rosty, Christophe; Clendenning, Mark; Spurdle, Amanda B; Win, Aung Ko

2014-01-01

Carriers of a germline mutation in one of the DNA mismatch repair (MMR) genes have a high risk of developing numerous different cancers, predominantly colorectal cancer and endometrial cancer (known as Lynch syndrome). MMR gene mutation carriers develop tumors with MMR deficiency identified by tumor microsatellite instability or immunohistochemical loss of MMR protein expression. Tumor MMR deficiency is used to identify individuals most likely to carry an MMR gene mutation. However, MMR deficiency can also result from somatic inactivation, most commonly methylation of the MLH1 gene promoter. As tumor MMR testing of all incident colorectal and endometrial cancers (universal screening) is becoming increasingly adopted, a growing clinical problem is emerging for individuals who have tumors that show MMR deficiency who are subsequently found not to carry an MMR gene mutation after genetic testing using the current diagnostic approaches (Sanger sequencing and multiplex ligation-dependent probe amplification) and who also show no evidence of MLH1 methylation. The inability to determine the underlying cause of tumor MMR deficiency in these "Lynch-like" or "suspected Lynch syndrome" cases has significant implications on the clinical management of these individuals and their relatives. When the data from published studies are combined, 59% (95% confidence interval [CI]: 55% to 64%) of colorectal cancers and 52% (95% CI: 41% to 62%) of endometrial cancers with MMR deficiency were identified as suspected Lynch syndrome. Recent studies estimated that colorectal cancer risk for relatives of suspected Lynch syndrome cases is lower than for relatives of those with MMR gene mutations, but higher than for relatives of those with tumor MMR deficiency resulting from methylation of the MLH1 gene promoter. The cause of tumor MMR deficiency in suspected Lynch syndrome cases is likely due to either unidentified germline MMR gene mutations, somatic cell mosaicism, or biallelic somatic inactivation. Determining the underlying cause of tumor MMR deficiency in suspected Lynch syndrome cases is likely to reshape the current triaging schemes used to identify germline MMR gene mutations in cancer-affected individuals and their relatives.

Mutation induction in haploid yeast after split-dose radiation exposure. II. Combination of UV-irradiation and X-rays.

PubMed

Keller, B; Zölzer, F; Kiefer, J

2004-01-01

Split-dose protocols can be used to investigate the kinetics of recovery from radiation damage and to elucidate the mechanisms of cell inactivation and mutation induction. In this study, a haploid strain of the yeast, Saccharomyces cerevisiae, wild-type with regard to radiation sensitivity, was irradiated with 254-nm ultraviolet (UV) light and then exposed to X-rays after incubation for 0-6 hr. The cells were incubated either on nutrient medium or salt agar between the treatments. Loss of reproductive ability and mutation to canavanine resistance were measured. When the X-ray exposure immediately followed UV-irradiation, the X-ray survival curves had the same slope irrespective of the pretreatment, while the X-ray mutation induction curves were changed from linear to linear quadratic with increasing UV fluence. Incubations up to about 3 hr on nutrient medium between the treatments led to synergism with respect to cell inactivation and antagonism with respect to mutation, but after 4-6 hr the two treatments acted independently. Incubation on salt agar did not cause any change in the survival curves, but there was a strong suppression of X-ray-induced mutation with increasing UV fluence. On the basis of these results, we suggest that mutation after combined UV and X-ray exposure is affected not only by the induction and suppression of DNA repair processes, but also by radiation-induced modifications of cell-cycle progression and changes in the expression of the mutant phenotype. Copyright 2004 Wiley-Liss, Inc.

Leaky RAG Deficiency in Adult Patients with Impaired Antibody Production against Bacterial Polysaccharide Antigens

PubMed Central

Geier, Christoph B.; Piller, Alexander; Linder, Angela; Sauerwein, Kai M. T.; Eibl, Martha M.; Wolf, Hermann M.

2015-01-01

Loss of function mutations in the recombination activating genes RAG1 and RAG2 have been reported to cause a T-B-NK+ type of severe combined immunodeficiency. In addition identification of hypomorphic mutations in RAG1 and RAG2 has led to an expansion of the spectrum of disease to include Omenn syndrome, early onset autoimmunity, granuloma, chronic cytomegalovirus- or EBV-infection with expansion of gamma/delta T-cells, idiophatic CD4 lymphopenia and a phenotype resembling common variable immunodeficiency. Herein we describe a novel presentation of leaky RAG1 and RAG2 deficiency in two unrelated adult patients with impaired antibody production against bacterial polysaccharide antigens. Clinical manifestation included recurrent pneumonia, sinusitis, otitis media and in one patient recurrent cutaneous vasculitis. Both patients harbored a combination of a null mutation on one allele with a novel hypomorphic RAG1/2 mutation on the other allele. One of these novel mutations affected the start codon of RAG1 and resulted in an aberrant gene and protein expression. The second novel RAG2 mutation leads to a truncated RAG2 protein, lacking the C-terminus with intact core RAG2 and reduced VDJ recombination capacity as previously described in a mouse model. Both patients presented with severely decreased numbers of naïve CD4+ T cells and defective T independent IgG responses to bacterial polysaccharide antigens, while T cell-dependent IgG antibody formation e.g. after tetanus or TBEV vaccination was intact. In conclusion, hypomorphic mutations in genes responsible for SCID should be considered in adults with predominantly antibody deficiency. PMID:26186701

Cosegregation of intragenic markers with a novel mutation that causes Crigler-Najjar syndrome type I: implication in carrier detection and prenatal diagnosis.

PubMed Central

Moghrabi, N; Clarke, D J; Burchell, B; Boxer, M

1993-01-01

Crigler-Najjar syndrome type 1 (CN-1) is a familial disorder characterized by severe unconjugated hyperbilirubinemia and jaundice and leads to kernicterus, neurological damage, and eventual death unless treated with liver transplantation. Previous reports identified mutations in the UGT1 gene complex to be the cause of the disease. The total absence of all phenol/bilirubin UGT proteins and their activities in liver homogenate of a CN-1 patient was determined by enzymological and immunochemical analysis. A novel homozygous nonsense mutation (CGA-->TGA) was identified in the patient by the combined techniques of PCR and direct sequencing. This mutation was located in exon 3 of the constant region in the gene complex which is common to all phenol and bilirubin UGTs. The segregation of the mutation in the patient's family was analyzed and confirmed the recessive nature of the disease. Newly developed intragenic polymorphic probes (UGT1* 4 and UGT-Const) were used on Southern blots of MspI-digested genomic DNA of the patient and his family. The segregation of individual alleles within the family was observed from haplotypes generated. Comparison of the segregation of haplotypes with the mutation for the patient and his family revealed the allele identified by the A1-B1-C2 haplotype to be carrying the mutation. The risk of recombination occurring is negligible, because of the intragenic nature of the probes. This study demonstrates the potential usefulness of these probes in carrier detection and prenatal/presymptomatic diagnosis. Images Figure 2 Figure 3 Figure 4 Figure 5 PMID:8102509

Clinical manifestation and molecular genetic characterization of MYH9 disorders.

PubMed

Provaznikova, Dana; Geierova, Vera; Kumstyrova, Tereza; Kotlin, Roman; Mikulenkova, Dana; Zurkova, Kamila; Matoska, Vaclav; Hrachovinova, Ingrid; Rittich, Simon

2009-08-01

Currently, the May-Hegglin anomaly (MHA), Sebastian (SBS), Fechtner (FTNS) and Epstein (EPS) syndrome are considered to be distinct clinical manifestations of a single disease caused by mutations of the MYH9 gene encoding the heavy chain of non-muscle myosin IIA (NMMHC-IIA). Manifestations of these disorders include giant platelets, thrombocytopenia and combinations of the presence of granulocyte inclusions, deafness, cataracts and renal failure. We examined 15 patients from 10 unrelated families on whom we performed immunostaining of NMMHC-IIA in blood samples. Polymerase chain reaction (PCR) analysis of selected exons of the MYH9 gene revealed mutations in nine samples with one novel mutation. Results of fluorescence and mutational analysis were compared with clinical manifestations of the MYH9 disorder. We also determined the number of glycoprotein sites on the surface of platelets. Most patients had an increased number of glycoproteins, which could be due to platelet size.

Timothy syndrome-like condition with syndactyly but without prolongation of the QT interval.

PubMed

Kosaki, Rika; Ono, Hiroshi; Terashima, Hiroshi; Kosaki, Kenjiro

2018-05-07

Timothy syndrome is characterized by a unique combination of a prolongation of the corrected QT interval of the electrocardiogram and bilateral cutaneous syndactyly of the fingers and the toes and is caused by heterozygous mutations in CACNA1C, a gene encoding a calcium channel. After the discovery of the CACNA1C gene as the causative gene for Timothy syndrome, patients with CACNA1C mutations with QT prolongation but without syndactyly were described. Here, we report a 5-year-old female patient with cutaneous syndactyly, developmental delay, and pulmonary hypertension. Exome analysis showed a previously undescribed de novo heterozygous mutation in the CACNA1C gene, p.Arg1024Gly. To our knowledge, this patient is the first to exhibit syndactyly and to carry a CACNA1C mutation but to not have QT prolongation, which has long been considered an obligatory feature of Timothy syndrome. © 2018 Wiley Periodicals, Inc.

Mutation of CDH23, encoding a new member of the cadherin gene family, causes Usher syndrome type 1D.

PubMed

Bolz, H; von Brederlow, B; Ramírez, A; Bryda, E C; Kutsche, K; Nothwang, H G; Seeliger, M; del C-Salcedó Cabrera, M; Vila, M C; Molina, O P; Gal, A; Kubisch, C

2001-01-01

Usher syndrome type I (USH1) is an autosomal recessive disorder characterized by congenital sensorineural hearing loss, vestibular dysfunction and visual impairment due to early onset retinitis pigmentosa (RP). So far, six loci (USH1A-USH1F) have been mapped, but only two USH1 genes have been identified: MYO7A for USH1B and the gene encoding harmonin for USH1C. We identified a Cuban pedigree linked to the locus for Usher syndrome type 1D (MIM 601067) within the q2 region of chromosome 10). Affected individuals present with congenital deafness and a highly variable degree of retinal degeneration. Using a positional candidate approach, we identified a new member of the cadherin gene superfamily, CDH23. It encodes a protein of 3,354 amino acids with a single transmembrane domain and 27 cadherin repeats. In the Cuban family, we detected two different mutations: a severe course of the retinal disease was observed in individuals homozygous for what is probably a truncating splice-site mutation (c.4488G-->C), whereas mild RP is present in individuals carrying the homozygous missense mutation R1746Q. A variable expression of the retinal phenotype was seen in patients with a combination of both mutations. In addition, we identified two mutations, Delta M1281 and IVS51+5G-->A, in a German USH1 patient. Our data show that different mutations in CDH23 result in USH1D with a variable retinal phenotype. In an accompanying paper, it is shown that mutations in the mouse ortholog cause disorganization of inner ear stereocilia and deafness in the waltzer mouse.

Structural and functional analysis of rare missense mutations in human chorionic gonadotrophin β-subunit

PubMed Central

Nagirnaja, Liina; Venclovas, Česlovas; Rull, Kristiina; Jonas, Kim C.; Peltoketo, Hellevi; Christiansen, Ole B.; Kairys, Visvaldas; Kivi, Gaily; Steffensen, Rudi; Huhtaniemi, Ilpo T.; Laan, Maris

2012-01-01

Heterodimeric hCG is one of the key hormones determining early pregnancy success. We have previously identified rare missense mutations in hCGβ genes with potential pathophysiological importance. The present study assessed the impact of these mutations on the structure and function of hCG by applying a combination of in silico (sequence and structure analysis, molecular dynamics) and in vitro (co-immunoprecipitation, immuno- and bioassays) approaches. The carrier status of each mutation was determined for 1086 North-Europeans [655 patients with recurrent miscarriage (RM)/431 healthy controls from Estonia, Finland and Denmark] using PCR-restriction fragment length polymorphism. The mutation CGB5 p.Val56Leu (rs72556325) was identified in a single heterozygous RM patient and caused a structural hindrance in the formation of the hCGα/β dimer. Although the amount of the mutant hCGβ assembled into secreted intact hCG was only 10% compared with the wild-type, a stronger signaling response was triggered upon binding to its receptor, thus compensating the effect of poor dimerization. The mutation CGB8 p.Pro73Arg (rs72556345) was found in five heterozygotes (three RM cases and two control individuals) and was inherited by two of seven studied live born children. The mutation caused ∼50% of secreted β-subunits to acquire an alternative conformation, but did not affect its biological activity. For the CGB8 p.Arg8Trp (rs72556341) substitution, the applied in vitro methods revealed no alterations in the assembly of intact hCG as also supported by an in silico analysis. In summary, the accumulated data indicate that only mutations with neutral or mild functional consequences might be tolerated in the major hCGβ genes CGB5 and CGB8. PMID:22554618

Exome capture sequencing identifies a novel mutation in BBS4

PubMed Central

Wang, Hui; Chen, Xianfeng; Dudinsky, Lynn; Patenia, Claire; Chen, Yiyun; Li, Yumei; Wei, Yue; Abboud, Emad B.; Al-Rajhi, Ali A.; Lewis, Richard Alan; Lupski, James R.; Mardon, Graeme; Gibbs, Richard A.; Perkins, Brian D.

2011-01-01

Purpose Leber congenital amaurosis (LCA) is one of the most severe eye dystrophies characterized by severe vision loss at an early stage and accounts for approximately 5% of all retinal dystrophies. The purpose of this study was to identify a novel LCA disease allele or gene and to develop an approach combining genetic mapping with whole exome sequencing. Methods Three patients from King Khaled Eye Specialist Hospital (KKESH205) underwent whole genome single nucleotide polymorphism genotyping, and a single candidate region was identified. Taking advantage of next-generation high-throughput DNA sequencing technologies, whole exome capture sequencing was performed on patient KKESH205#7. Sanger direct sequencing was used during the validation step. The zebrafish model was used to examine the function of the mutant allele. Results A novel missense mutation in Bardet-Biedl syndrome 4 protein (BBS4) was identified in a consanguineous family from Saudi Arabia. This missense mutation in the fifth exon (c.253G>C;p.E85Q) of BBS4 is likely a disease-causing mutation as it segregates with the disease. The mutation is not found in the single nucleotide polymorphism (SNP) database, the 1000 Genomes Project, or matching normal controls. Functional analysis of this mutation in zebrafish indicates that the G253C allele is pathogenic. Coinjection of the G253C allele cannot rescue the mislocalization of rhodopsin in the retina when BBS4 is knocked down by morpholino injection. Immunofluorescence analysis in cell culture shows that this missense mutation in BBS4 does not cause obvious defects in protein expression or pericentriolar localization. Conclusions This mutation likely mainly reduces or abolishes BBS4 function in the retina. Further studies of this allele will provide important insights concerning the pleiotropic nature of BBS4 function. PMID:22219648

Atlanto-axial malformation and instability in dogs with pituitary dwarfism due to an LHX3 mutation.

PubMed

Voorbij, A M W Y; Meij, B P; van Bruggen, L W L; Grinwis, G C M; Stassen, Q E M; Kooistra, H S

2015-01-01

Canine pituitary dwarfism or combined pituitary hormone deficiency (CPHD) in shepherd dogs is associated with an LHX3 mutation and can lead to a wide range of clinical manifestations. Some dogs with CPHD have neurological signs that are localized to the cervical spine. In human CPHD, caused by an LHX3 mutation, anatomical abnormalities in the atlanto-axial (C1-C2) joint have been described. To evaluate the presence of atlanto-axial malformations in dogs with pituitary dwarfism associated with an LHX3 mutation and to investigate the degree of similarity between the atlanto-axial anomalies found in canine and human CPHD patients with an LHX3 mutation. Three client-owned Czechoslovakian wolfdogs and 1 client-owned German shepherd dog, previously diagnosed with pituitary dwarfism caused by an LHX3 mutation, with neurological signs indicating a cervical spinal disorder. Radiography, computed tomography, and magnetic resonance imaging of the cranial neck and skull, necropsy, and histology. Diagnostic imaging identified abnormal positioning of the dens axis and incomplete ossification of the suture lines between the ossification centers of the atlas with concurrent atlanto-axial instability and dynamic compression of the spinal cord by the dens axis. The malformations and aberrant motion at C1-C2 were confirmed at necropsy and histology. The atlanto-axial abnormalities of the dwarf dogs resemble those encountered in human CPHD patients with an LHX3 mutation. These findings suggest an association between the LHX3 mutation in dogs with CPHD and atlanto-axial malformations. Consequently, pituitary dwarfs should be monitored closely for neurological signs. Copyright © 2015 The Authors. Journal of Veterinary Internal Medicine published by Wiley Periodicals, Inc. on behalf of American College of Veterinary Internal Medicine.

A novel rasopathy caused by recurrent de novo missense mutations in PPP1CB closely resembles Noonan syndrome with loose anagen hair.

PubMed

Gripp, Karen W; Aldinger, Kimberly A; Bennett, James T; Baker, Laura; Tusi, Jessica; Powell-Hamilton, Nina; Stabley, Deborah; Sol-Church, Katia; Timms, Andrew E; Dobyns, William B

2016-09-01

Noonan syndrome is a rasopathy caused by mutations in multiple genes encoding components of the RAS/MAPK pathway. Despite its variable phenotype, limited genotype-phenotype correlations exist. Noonan syndrome with loose anagen hair (NS-LAH) is characterized by its distinctive hair anomalies, developmental differences, and structural brain abnormalities and is caused by a single recurrent missense SHOC2 mutation. SHOC2 forms a complex with protein phosphatase 1 (PP1C). Protein phosphatases counterbalance kinases and control activation of signaling proteins, such as the mitogen-activated protein kinases of the RAS/MAPK pathway. Here we report four patients with de novo missense mutations in protein phosphatase one catalytic subunit beta (PPP1CB), sharing a recognizable phenotype. Three individuals had the recurrent PPP1CB c.146G>C, p.Pro49Arg mutation, the fourth had a c.166G>C, p.Ala56Pro change. All had relative or absolute macrocephaly, low-set and posteriorly angulated ears, and developmental delay. Slow growing and/or sparse hair and/or an unruly hair texture was present in all. Three individuals had feeding difficulties requiring feeding tubes. One of two males had cryptorchidism, another had pectus excavatum. Short stature was present in three. A female with the recurrent mutation had a Dandy-Walker malformation and optic nerve hypoplasia. Mild ventriculomegaly occurred in all, cerebellar tonsillar ectopia was seen in two and progressed to Chiari 1 malformation in one individual. Based on the combination of phenotypic findings and PPP1CB's effect on RAF dephosphorylation within the RAS/MAPK pathway, this novel condition can be considered a rasopathy, most similar to NS-LAH. Collectively, these mutations meet the standardized criteria for pathogenicity. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Novel compound heterozygous mutations in MYO7A Associated with Usher syndrome 1 in a Chinese family.

PubMed

Gao, Xue; Wang, Guo-Jian; Yuan, Yong-Yi; Xin, Feng; Han, Ming-Yu; Lu, Jing-Qiao; Zhao, Hui; Yu, Fei; Xu, Jin-Cao; Zhang, Mei-Guang; Dong, Jiang; Lin, Xi; Dai, Pu

2014-01-01

Usher syndrome is an autosomal recessive disease characterized by sensorineural hearing loss, age-dependent retinitis pigmentosa (RP), and occasionally vestibular dysfunction. The most severe form is Usher syndrome type 1 (USH1). Mutations in the MYO7A gene are responsible for USH1 and account for 29-55% of USH1 cases. Here, we characterized a Chinese family (no. 7162) with USH1. Combining the targeted capture of 131 known deafness genes, next-generation sequencing, and bioinformatic analysis, we identified two deleterious compound heterozygous mutations in the MYO7A gene: a reported missense mutation c.73G>A (p.G25R) and a novel nonsense mutation c.462C>A (p.C154X). The two compound variants are absent in 219 ethnicity-matched controls, co-segregates with the USH clinical phenotypes, including hearing loss, vestibular dysfunction, and age-dependent penetrance of progressive RP, in family 7162. Therefore, we concluded that the USH1 in this family was caused by compound heterozygous mutations in MYO7A.

PPM1D Mosaic Truncating Variants in Ovarian Cancer Cases May Be Treatment-Related Somatic Mutations

PubMed Central

Pharoah, Paul D. P.; Song, Honglin; Dicks, Ed; Intermaggio, Maria P.; Harrington, Patricia; Baynes, Caroline; Alsop, Kathryn; Bogdanova, Natalia; Cicek, Mine S.; Cunningham, Julie M.; Fridley, Brooke L.; Gentry-Maharaj, Aleksandra; Hillemanns, Peter; Lele, Shashi; Lester, Jenny; McGuire, Valerie; Moysich, Kirsten B.; Poblete, Samantha; Sieh, Weiva; Sucheston-Campbell, Lara; Widschwendter, Martin; Whittemore, Alice S.; Dörk, Thilo; Menon, Usha; Odunsi, Kunle; Goode, Ellen L.; Karlan, Beth Y.; Bowtell, David D.; Gayther, Simon A.; Ramus, Susan J.

2016-01-01

Mosaic truncating mutations in the protein phosphatase, Mg2+/Mn2+-dependent, 1D (PPM1D) gene have recently been reported with a statistically significantly greater frequency in lymphocyte DNA from ovarian cancer case patients compared with unaffected control patients. Using massively parallel sequencing (MPS) we identified truncating PPM1D mutations in 12 of 3236 epithelial ovarian cancer (EOC) case patients (0.37%) but in only one of 3431 unaffected control patients (0.03%) (P = .001). All statistical tests were two-sided. A combination of Sanger sequencing, pyrosequencing, and MPS data suggested that 12 of the 13 mutations were mosaic. All mutations were identified in post-chemotherapy treatment blood samples from case patients (n = 1827) (average 1234 days post-treatment in carriers) rather than from cases collected pretreatment (less than 14 days after diagnosis, n = 1384) (P = .002). These data suggest that PPM1D variants in EOC cases are primarily somatic mosaic mutations caused by treatment and are not associated with germline predisposition to EOC. PMID:26823519

Network and Atomistic Simulations Unveil the Structural Determinants of Mutations Linked to Retinal Diseases

PubMed Central

Mariani, Simona; Dell'Orco, Daniele; Felline, Angelo; Raimondi, Francesco; Fanelli, Francesca

2013-01-01

A number of incurable retinal diseases causing vision impairments derive from alterations in visual phototransduction. Unraveling the structural determinants of even monogenic retinal diseases would require network-centered approaches combined with atomistic simulations. The transducin G38D mutant associated with the Nougaret Congenital Night Blindness (NCNB) was thoroughly investigated by both mathematical modeling of visual phototransduction and atomistic simulations on the major targets of the mutational effect. Mathematical modeling, in line with electrophysiological recordings, indicates reduction of phosphodiesterase 6 (PDE) recognition and activation as the main determinants of the pathological phenotype. Sub-microsecond molecular dynamics (MD) simulations coupled with Functional Mode Analysis improve the resolution of information, showing that such impairment is likely due to disruption of the PDEγ binding cavity in transducin. Protein Structure Network analyses additionally suggest that the observed slight reduction of theRGS9-catalyzed GTPase activity of transducin depends on perturbed communication between RGS9 and GTP binding site. These findings provide insights into the structural fundamentals of abnormal functioning of visual phototransduction caused by a missense mutation in one component of the signaling network. This combination of network-centered modeling with atomistic simulations represents a paradigm for future studies aimed at thoroughly deciphering the structural determinants of genetic retinal diseases. Analogous approaches are suitable to unveil the mechanism of information transfer in any signaling network either in physiological or pathological conditions. PMID:24009494

Mutations in ARL2BP, Encoding ADP-Ribosylation-Factor-Like 2 Binding Protein, Cause Autosomal-Recessive Retinitis Pigmentosa

PubMed Central

Davidson, Alice E.; Schwarz, Nele; Zelinger, Lina; Stern-Schneider, Gabriele; Shoemark, Amelia; Spitzbarth, Benjamin; Gross, Menachem; Laxer, Uri; Sosna, Jacob; Sergouniotis, Panagiotis I.; Waseem, Naushin H.; Wilson, Robert; Kahn, Richard A.; Plagnol, Vincent; Wolfrum, Uwe; Banin, Eyal; Hardcastle, Alison J.; Cheetham, Michael E.; Sharon, Dror; Webster, Andrew R.

2013-01-01

Retinitis pigmentosa (RP) is a genetically heterogeneous retinal degeneration characterized by photoreceptor death, which results in visual failure. Here, we used a combination of homozygosity mapping and exome sequencing to identify mutations in ARL2BP, which encodes an effector protein of the small GTPases ARL2 and ARL3, as causative for autosomal-recessive RP (RP66). In a family affected by RP and situs inversus, a homozygous, splice-acceptor mutation, c.101−1G>C, which alters pre-mRNA splicing of ARLBP2 in blood RNA, was identified. In another family, a homozygous c.134T>G (p.Met45Arg) mutation was identified. In the mouse retina, ARL2BP localized to the basal body and cilium-associated centriole of photoreceptors and the periciliary extension of the inner segment. Depletion of ARL2BP caused cilia shortening. Moreover, depletion of ARL2, but not ARL3, caused displacement of ARL2BP from the basal body, suggesting that ARL2 is vital for recruiting or anchoring ARL2BP at the base of the cilium. This hypothesis is supported by the finding that the p.Met45Arg amino acid substitution reduced binding to ARL2 and caused the loss of ARL2BP localization at the basal body in ciliated nasal epithelial cells. These data demonstrate a role for ARL2BP and ARL2 in primary cilia function and that this role is essential for normal photoreceptor maintenance and function. PMID:23849777

Cri du chat syndrome and primary ciliary dyskinesia: a common genetic cause on chromosome 5p.

PubMed

Shapiro, Adam J; Weck, Karen E; Chao, Kay C; Rosenfeld, Margaret; Nygren, Anders O H; Knowles, Michael R; Leigh, Margaret W; Zariwala, Maimoona A

2014-10-01

Cri du chat syndrome (CdCS) and primary ciliary dyskinesia (PCD) are rare diseases that present with frequent respiratory symptoms. PCD can be caused by hemizygous DNAH5 mutation in combination with a 5p segmental deletion attributable to CdCS on the opposite chromosome. Chronic oto-sino-pulmonary symptoms or organ laterality defects in CdCS should prompt an evaluation for PCD. Copyright © 2014 Elsevier Inc. All rights reserved.

Cone opsins, colour blindness and cone dystrophy: Genotype-phenotype correlations.

PubMed

Gardner, J C; Michaelides, M; Hardcastle, A J

2016-05-25

X-linked cone photoreceptor disorders caused by mutations in the OPN1LW (L) and OPN1MW (M) cone opsin genes on chromosome Xq28 include a range of conditions from mild stable red-green colour vision deficiencies to severe cone dystrophies causing progressive loss of vision and blindness. Advances in molecular genotyping and functional analyses of causative variants, combined with deep retinal phenotyping, are unravelling genetic mechanisms underlying the variability of cone opsin disorders.

Two-Exon Skipping within MLPH Is Associated with Coat Color Dilution in Rabbits

PubMed Central

Lehner, Stefanie; Gähle, Marion; Dierks, Claudia; Stelter, Ricarda; Gerber, Jonathan; Brehm, Ralph; Distl, Ottmar

2013-01-01

Coat color dilution turns black coat color to blue and red color to cream and is a characteristic in many mammalian species. Matings among Netherland Dwarf, Loh, and Lionhead Dwarf rabbits over two generations gave evidence for a monogenic autosomal recessive inheritance of coat colour dilution. Histological analyses showed non-uniformly distributed, large, agglomerating melanin granules in the hair bulbs of coat color diluted rabbits. We sequenced the cDNA of MLPH in two dilute and one black rabbit for polymorphism detection. In both color diluted rabbits, skipping of exons 3 and 4 was present resulting in altered amino acids at p.QGL[37-39]QWA and a premature stop codon at p.K40*. Sequencing of genomic DNA revealed a c.111-5C>A splice acceptor mutation within the polypyrimidine tract of intron 2 within MLPH. This mutation presumably causes skipping of exons 3 and 4. In 14/15 dilute rabbits, the c.111-5C>A mutation was homozygous and in a further dilute rabbit, heterozygous and in combination with a homozygous frame shift mutation within exon 6 (c.585delG). In conclusion, our results demonstrated a colour dilution associated MLPH splice variant causing a strongly truncated protein (p.Q37QfsX4). An involvement of further MLPH-associated mutations needs further investigations. PMID:24376820

Adult cases of mitochondrial DNA depletion due to TK2 defect: an expanding spectrum.

PubMed

Béhin, A; Jardel, C; Claeys, K G; Fagart, J; Louha, M; Romero, N B; Laforêt, P; Eymard, B; Lombès, A

2012-02-28

In this study we aim to demonstrate the occurrence of adult forms of TK2 mutations causing progressive mitochondrial myopathy with significant muscle mitochondrial DNA (mtDNA) depletion. Patients' investigations included serum creatine kinase, blood lactate, electromyographic, echocardiographic, and functional respiratory analyses as well as TK2 gene sequencing and TK2 activity measurement. Mitochondrial activities and mtDNA were analyzed in the patients' muscle biopsy. The 3 adult patients with TK2 mutations presented with slowly progressive myopathy compatible with a fairly normal life during decades. Apart from its much slower progression, these patients' phenotype closely resembled that of pediatric cases including early onset, absence of CNS symptoms, generalized muscle weakness predominating on axial and proximal muscles but affecting facial, ocular, and respiratory muscles, typical mitochondrial myopathy with a mosaic pattern of COX-negative and ragged-red fibers, combined mtDNA-dependent respiratory complexes deficiency and mtDNA depletion. In accordance with the disease's relatively slow progression, the residual mtDNA content was higher than that observed in pediatric cases. That difference was not explained by the type of the TK2 mutations or by the residual TK2 activity. TK2 mutations can cause mitochondrial myopathy with a slow progression. Comparison of patients with similar mutations but different disease progression might address potential mechanisms of mtDNA maintenance modulation.

JAK and MPL mutations in myeloid malignancies.

PubMed

Tefferi, Ayalew

2008-03-01

The Janus family of non-receptor tyrosine kinases (JAK1, JAK2, JAK3 and tyrosine kinase 2) transduces signals downstream of type I and II cytokine receptors via signal transducers and activators of transcription (STATs). JAK3 is important in lymphoid and JAK2 in myeloid cell proliferation and differentiation. The thrombopoietin receptor MPL is one of several JAK2 cognate receptors and is essential for myelopoiesis in general and megakaryopoiesis in particular. Germline loss-of-function (LOF) JAK3 and MPL mutations cause severe combined immunodeficiency and congenital amegakaryocytic thrombocytopenia, respectively. Germline gain-of-function (GOF) MPL mutation (MPLS505N) causes familial thrombocytosis. Somatic JAK3 (e.g. JAK3A572V, JAK3V722I, JAK3P132T) and fusion JAK2 (e.g. ETV6-JAK2, PCM1-JAK2, BCR-JAK2) mutations have respectively been described in acute megakaryocytic leukemia and acute leukemia/chronic myeloid malignancies. However, current attention is focused on JAK2 (e.g. JAK2V617F, JAK2 exon 12 mutations) and MPL (e.g. MPLW515L/K/S, MPLS505N) mutations associated with myeloproliferative neoplasms (MPNs). A JAK2 mutation, primarily JAK2V617F, is invariably associated with polycythemia vera (PV). The latter mutation also occurs in the majority of patients with essential thrombocythemia (ET) or primary myelofibrosis (PMF). MPL mutational frequency in MPNs is substantially less (<10%). In general, despite a certain degree of genotype - phenotype correlations, the prognostic relevance of harbouring one of these mutations, or their allele burden when present, remains dubious. Regardless, based on the logical assumption that amplified JAK-STAT signalling is central to the pathogenesis of PV, ET and PMF, several anti-JAK2 tyrosine kinase inhibitors have been developed and are currently being tested in humans with these disorders.

Mechanistic Approaches to Improve Correction of the Most Common Disease-Causing Mutation in Cystic Fibrosis

PubMed Central

Bali, Vedrana; Lazrak, Ahmed; Guroji, Purushotham; Matalon, Sadis; Bebok, Zsuzsanna

2016-01-01

The most common mutation in the cystic fibrosis transmembrane conductance regulator (CFTR) gene leads to deletion of the phenylalanine at position 508 (ΔF508) in the CFTR protein and causes multiple folding and functional defects. Contrary to large-scale efforts by industry and academia, no significant therapeutic benefit has been achieved with a single “corrector”. Therefore, investigations concentrate on drug combinations. Orkambi (Vertex Pharmaceuticals), the first FDA-approved drug for treatment of cystic fibrosis (CF) caused by this mutation, is a combination of a corrector (VX-809) that facilitates ΔF508 CFTR biogenesis and a potentiator (VX-770), which improves its function. Yet, clinical trials utilizing this combination showed only modest therapeutic benefit. The low efficacy Orkambi has been attributed to VX-770-mediated destabilization of VX-809-rescued ΔF508 CFTR. Here we report that the negative effects of VX-770 can be reversed by increasing the half-life of the endoplasmic reticulum (ER) form (band B) of ΔF508 CFTR with another corrector (Corr-4a.) Although Corr-4a alone has only minimal effects on ΔF508 CFTR rescue, it increases the half-life of ΔF508 CFTR band B when it is present during half-life measurements. Our data shows that stabilization of band B ΔF508 CFTR with Corr-4a and simultaneous rescue with VX-809, leads to a >2-fold increase in cAMP-activated, CFTRinh-172-inhibited currents compared to VX-809 alone, or VX-809+VX-770. The negative effects of VX-770 and the Corr-4a protection are specific to the native I507-ATT ΔF508 CFTR without affecting the inherently more stable, synonymous variant I507-ATC ΔF508 CFTR. Our studies emphasize that stabilization of ΔF508 CFTR band B in the ER might improve its functional rescue by Orkambi. PMID:27214033

Eye Development Genes and Known Syndromes

PubMed Central

Slavotinek, Anne M.

2011-01-01

Anophthalmia and microphthalmia (A/M) are significant eye defects because they can have profound effects on visual acuity. A/M is associated with non-ocular abnormalities in an estimated 33–95% of cases and around 25% of patients have an underlying genetic syndrome that is diagnosable. Syndrome recognition is important for targeted molecular genetic testing, prognosis and for counseling regarding recurrence risks. This review provides clinical and molecular information for several of the commonest syndromes associated with A/M: Anophthalmia-Esophageal-Genital syndrome, caused by SOX2 mutations, Anophthalmia and pituitary abnormalities caused by OTX2 mutations, Matthew-Wood syndrome caused by STRA6 mutations, Oculocardiafaciodental syndrome and Lenz microphthalmia caused by BCOR mutations, Microphthalmia Linear Skin pigmentation syndrome caused by HCCS mutations, Anophthalmia, pituitary abnormalities, polysyndactyly caused by BMP4 mutations and Waardenburg anophthalmia caused by mutations in SMOC1. In addition, we briefly discuss the ocular and extraocular phenotypes associated with several other important eye developmental genes, including GDF6, VSX2, RAX, SHH, SIX6 and PAX6. PMID:22005280

PIK3CA-associated developmental disorders exhibit distinct classes of mutations with variable expression and tissue distribution

PubMed Central

Timms, Andrew E.; Conti, Valerio; Girisha, Katta M.; Martin, Beth; Olds, Carissa; Collins, Sarah; Park, Kaylee; Carter, Melissa; Krägeloh-Mann, Inge; Chitayat, David; Parikh, Aditi Shah; Bradshaw, Rachael; Torti, Erin; Braddock, Stephen; Burke, Leah; Ghedia, Sondhya; Stephan, Mark; Stewart, Fiona; Prasad, Chitra; Napier, Melanie; Saitta, Sulagna; Straussberg, Rachel; Gabbett, Michael; O’Connor, Bridget C.; Yin, Lim Jiin; Lai, Angeline Hwei Meeng; Martin, Nicole; McKinnon, Margaret; Addor, Marie-Claude; Schwartz, Charles E.; Lanoel, Agustina; Conway, Robert L.; Devriendt, Koenraad; Tatton-Brown, Katrina; Pierpont, Mary Ella; Painter, Michael; Worgan, Lisa; Reggin, James; Hennekam, Raoul; Pritchard, Colin C.; Aracena, Mariana; Gripp, Karen W.; Cordisco, Maria; Van Esch, Hilde; Garavelli, Livia; Curry, Cynthia; Goriely, Anne; Kayserilli, Hulya; Shendure, Jay; Graham, John; Guerrini, Renzo; Dobyns, William B.

2016-01-01

Mosaicism is increasingly recognized as a cause of developmental disorders with the advent of next-generation sequencing (NGS). Mosaic mutations of PIK3CA have been associated with the widest spectrum of phenotypes associated with overgrowth and vascular malformations. We performed targeted NGS using 2 independent deep-coverage methods that utilize molecular inversion probes and amplicon sequencing in a cohort of 241 samples from 181 individuals with brain and/or body overgrowth. We identified PIK3CA mutations in 60 individuals. Several other individuals (n = 12) were identified separately to have mutations in PIK3CA by clinical targeted-panel testing (n = 6), whole-exome sequencing (n = 5), or Sanger sequencing (n = 1). Based on the clinical and molecular features, this cohort segregated into three distinct groups: (a) severe focal overgrowth due to low-level but highly activating (hotspot) mutations, (b) predominantly brain overgrowth and less severe somatic overgrowth due to less-activating mutations, and (c) intermediate phenotypes (capillary malformations with overgrowth) with intermediately activating mutations. Sixteen of 29 PIK3CA mutations were novel. We also identified constitutional PIK3CA mutations in 10 patients. Our molecular data, combined with review of the literature, show that PIK3CA-related overgrowth disorders comprise a discontinuous spectrum of disorders that correlate with the severity and distribution of mutations. PMID:27631024

Screening for mutations in rhodopsin and peripherin/RDS in patients with autosomal dominant retinitis pigmentosa

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rodriguez, J.A.; Gannon, A.M.; Daiger, S.P.

1994-09-01

Mutations in rhodopsin account for approximately 30% of all cases of autosomal dominant retinits pigmentosa (adRP) and mutations in peripherin/RDS account for an additional 5% of cases. Also, mutations in rhodopsin can cause autosomal recessive retinitis pigmentosa and mutations in peripherin/RDS can cause dominant macular degeneration. Most disease-causing mutations in rhodopsin and peripherin/RDS are unique to one family or, at most, to a few families within a limited geographic region, though a few mutations are found in multiple, unrelated families. To further determine the spectrum of genetic variation in these genes, we screened DNA samples from 134 unrelated patients withmore » retinitis pigmentosa for mutations in both rhodopsin and peripherin/RDS using SSCP followed by genomic sequencing. Of the 134 patients, 86 were from families with apparent adRP and 48 were either isolated cases or were from families with an equivocal mode of inheritance. Among these patients we found 14 distinct rhodopsin mutations which are likely to cause retinal disease. Eleven of these mutations were found in one individual or one family only, whereas the Pro23His mutation was found in 14 {open_quotes}unrelated{close_quotes}individuals. The splice-site mutation produces dominant disease though with highly variable expression. Among the remaining patients were found 6 distinct peripherin/RDS mutations which are likely to cause retinal disease. These mutations were also found in one patient or family only, except the Gly266Asp mutation which was found in two unrelated patients. These results confirm the expected frequency and broad spectrum of mutations causing adRP.« less

Preclinical rationale for combination of crizotinib with mitomycin C for the treatment of advanced colorectal cancer.

PubMed

Lev, Avital; Deihimi, Safoora; Shagisultanova, Elena; Xiu, Joanne; Lulla, Amriti R; Dicker, David T; El-Deiry, Wafik S

2017-09-02

Colorectal cancer (CRC) is a leading cause of cancer-related deaths in the United States. We analyzed 26 MSI-High and 558 non-MSI-High CRC tumors. BRCA2 mutations were highly enriched (50%) in MSI-High CRC. Immunohistochemistry showed that BRCA2-mutated MSI-High CRC had high c-MET (64%) expression compared with BRCA-WT (17%). We hypothesized a mechanistic link between BRCA2-deficiency and c-MET overexpression and synergistic interaction between drugs that treat BRCA-deficient tumors (mitomycin C (MMC) or PARP inhibitors) and c-MET inhibitors (crizotinib). We tested CRC cell lines for sensitivity to MMC plus crizotinib or other drug combinations including PARP-inhibitors. Combined treatment of tumor cells with crizotinib and MMC led to increased apoptosis as compared with each drug alone. Additionally, combination treatment with increasing concentrations of both drugs demonstrated a synergistic anti-cancer effect (CI = 0.006-0.74). However, we found no evidence for c-MET upregulation upon effective BRCA2 knockdown in tumor cells -/+DNA damage. Although we found no mechanistic link between BRCA2 deficiency and c-MET overexpression, c-MET is frequently overexpressed in CRC and BRCA2 is mutated especially in MSI-H CRC. The combination of crizotinib with MMC appeared synergistic regardless of MSI or BRCA2 status. Using an in-vivo CRC xenograft model we found reduced tumor growth with combined crizotinib and MMC therapy (p = 0.0088). Our preclinical results support clinical testing of the combination of MMC and crizotinib in advanced CRC. Targeting cell survival mediated by c-MET in combination with targeting DNA repair may be a reasonable strategy for therapy development in CRC or other cancers.

Unpacking 'Artemisinin Resistance'.

PubMed

Wang, Jigang; Xu, Chengchao; Lun, Zhao-Rong; Meshnick, Steven R

2017-06-01

Artemisinin and its derivatives, in combination with partner drugs, are currently the most effective treatments for malaria parasite infection. Even though artemisinin has been widely used for decades, its mechanism of action had remained controversial until recently. Artemisinin combination therapies (ACTs) have recently been found to be losing efficacy in Southeast Asia. This 'artemisinin resistance', defined by a delayed parasite clearance time, has been associated with several genetic mutations. As with any other drug resistance phenotype, resistance can best be understood based on its mechanism of action. Recently, it was demonstrated that artemisinin attacks multiple parasitic targets, suggesting that mutations in drug targets are unlikely to cause high-level artemisinin resistance. These findings will help us to better understand the mechanisms of artemisinin resistance and suggest protocol modifications that may improve the efficacy of ACTs. Copyright © 2017 Elsevier Ltd. All rights reserved.

ORAI1 mutations abolishing store-operated Ca2+ entry cause anhidrotic ectodermal dysplasia with immunodeficiency.

PubMed

Lian, Jayson; Cuk, Mario; Kahlfuss, Sascha; Kozhaya, Lina; Vaeth, Martin; Rieux-Laucat, Frédéric; Picard, Capucine; Benson, Melina J; Jakovcevic, Antonia; Bilic, Karmen; Martinac, Iva; Stathopulos, Peter; Kacskovics, Imre; Vraetz, Thomas; Speckmann, Carsten; Ehl, Stephan; Issekutz, Thomas; Unutmaz, Derya; Feske, Stefan

2017-11-16

Store-operated Ca 2+ entry (SOCE) through Ca 2+ release-activated Ca 2+ channels is an essential signaling pathway in many cell types. Ca 2+ release-activated Ca 2+ channels are formed by ORAI1, ORAI2, and ORAI3 proteins and activated by stromal interaction molecule (STIM) 1 and STIM2. Mutations in the ORAI1 and STIM1 genes that abolish SOCE cause a combined immunodeficiency (CID) syndrome that is accompanied by autoimmunity and nonimmunologic symptoms. We performed molecular and immunologic analysis of patients with CID, anhidrosis, and ectodermal dysplasia of unknown etiology. We performed DNA sequencing of the ORAI1 gene, modeling of mutations on ORAI1 crystal structure, analysis of ORAI1 mRNA and protein expression, SOCE measurements, immunologic analysis of peripheral blood lymphocyte populations by using flow cytometry, and histologic and ultrastructural analysis of patient tissues. We identified 3 novel autosomal recessive mutations in ORAI1 in unrelated kindreds with CID, autoimmunity, ectodermal dysplasia with anhidrosis, and muscular dysplasia. The patients were homozygous for p.V181SfsX8, p.L194P, and p.G98R mutations in the ORAI1 gene that suppressed ORAI1 protein expression and SOCE in the patients' lymphocytes and fibroblasts. In addition to impaired T-cell cytokine production, ORAI1 mutations were associated with strongly reduced numbers of invariant natural killer T and regulatory T (Treg) cells and altered composition of γδ T-cell and natural killer cell subsets. ORAI1 null mutations are associated with reduced numbers of invariant natural killer T and Treg cells that likely contribute to the patients' immunodeficiency and autoimmunity. ORAI1-deficient patients have dental enamel defects and anhidrosis, representing a new form of anhidrotic ectodermal dysplasia with immunodeficiency that is distinct from previously reported patients with anhidrotic ectodermal dysplasia with immunodeficiency caused by mutations in the nuclear factor κB signaling pathway (IKBKG and NFKBIA). Copyright © 2017 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

C. elegans whole-genome sequencing reveals mutational signatures related to carcinogens and DNA repair deficiency

PubMed Central

Meier, Bettina; Cooke, Susanna L.; Weiss, Joerg; Bailly, Aymeric P.; Alexandrov, Ludmil B.; Marshall, John; Raine, Keiran; Maddison, Mark; Anderson, Elizabeth; Stratton, Michael R.; Campbell, Peter J.

2014-01-01

Mutation is associated with developmental and hereditary disorders, aging, and cancer. While we understand some mutational processes operative in human disease, most remain mysterious. We used Caenorhabditis elegans whole-genome sequencing to model mutational signatures, analyzing 183 worm populations across 17 DNA repair-deficient backgrounds propagated for 20 generations or exposed to carcinogens. The baseline mutation rate in C. elegans was approximately one per genome per generation, not overtly altered across several DNA repair deficiencies over 20 generations. Telomere erosion led to complex chromosomal rearrangements initiated by breakage–fusion–bridge cycles and completed by simultaneously acquired, localized clusters of breakpoints. Aflatoxin B1 induced substitutions of guanines in a GpC context, as observed in aflatoxin-induced liver cancers. Mutational burden increased with impaired nucleotide excision repair. Cisplatin and mechlorethamine, DNA crosslinking agents, caused dose- and genotype-dependent signatures among indels, substitutions, and rearrangements. Strikingly, both agents induced clustered rearrangements resembling “chromoanasynthesis,” a replication-based mutational signature seen in constitutional genomic disorders, suggesting that interstrand crosslinks may play a pathogenic role in such events. Cisplatin mutagenicity was most pronounced in xpf-1 mutants, suggesting that this gene critically protects cells against platinum chemotherapy. Thus, experimental model systems combined with genome sequencing can recapture and mechanistically explain mutational signatures associated with human disease. PMID:25030888

JAG1 mutations are found in approximately one third of patients presenting with only one or two clinical features of Alagille syndrome.

PubMed

Guegan, K; Stals, K; Day, M; Turnpenny, P; Ellard, S

2012-07-01

Alagille syndrome is a multisystem disorder characterized by highly variable expressivity, most frequently caused by heterozygous JAG1 gene mutations. Classic diagnostic criteria combine the presence of bile duct paucity on liver biopsy with three of five systems affected; liver, heart, skeleton, eye and dysmorphic facies. The aim of this study was to determine the prevalence and distribution of JAG1 mutations in patients referred for routine clinical diagnostic testing. Clinical data were available for 241 patients from 135 families. The index cases were grouped according to the number of systems affected (heart, liver, skeletal, eye and facies) and the mutation frequency calculated for each group. JAG1 mutations were identified in 59/135 (44%) probands. The highest mutation detection rates were observed in patients with the most frequent presenting features of Alagille syndrome; ranging from 20% (one system) to 86% (five systems). The overall mutation pick-up rate in a clinical diagnostic setting was lower than in previous research studies. Identification of a JAG1 gene mutation is particularly useful for those patients with atypical or mild Alagille syndrome who do not meet classic diagnostic criteria as it provides a definite molecular diagnosis and allows accurate genetic counselling for the family. © 2011 John Wiley & Sons A/S.

Adaptor protein complex 4 deficiency causes severe autosomal-recessive intellectual disability, progressive spastic paraplegia, shy character, and short stature.

PubMed

Abou Jamra, Rami; Philippe, Orianne; Raas-Rothschild, Annick; Eck, Sebastian H; Graf, Elisabeth; Buchert, Rebecca; Borck, Guntram; Ekici, Arif; Brockschmidt, Felix F; Nöthen, Markus M; Munnich, Arnold; Strom, Tim M; Reis, Andre; Colleaux, Laurence

2011-06-10

Intellectual disability inherited in an autosomal-recessive fashion represents an important fraction of severe cognitive-dysfunction disorders. Yet, the extreme heterogeneity of these conditions markedly hampers gene identification. Here, we report on eight affected individuals who were from three consanguineous families and presented with severe intellectual disability, absent speech, shy character, stereotypic laughter, muscular hypotonia that progressed to spastic paraplegia, microcephaly, foot deformity, decreased muscle mass of the lower limbs, inability to walk, and growth retardation. Using a combination of autozygosity mapping and either Sanger sequencing of candidate genes or next-generation exome sequencing, we identified one mutation in each of three genes encoding adaptor protein complex 4 (AP4) subunits: a nonsense mutation in AP4S1 (NM_007077.3: c.124C>T, p.Arg42(∗)), a frameshift mutation in AP4B1 (NM_006594.2: c.487_488insTAT, p.Glu163_Ser739delinsVal), and a splice mutation in AP4E1 (NM_007347.3: c.542+1_542+4delGTAA, r.421_542del, p.Glu181Glyfs(∗)20). Adaptor protein complexes (AP1-4) are ubiquitously expressed, evolutionarily conserved heterotetrameric complexes that mediate different types of vesicle formation and the selection of cargo molecules for inclusion into these vesicles. Interestingly, two mutations affecting AP4M1 and AP4E1 have recently been found to cause cerebral palsy associated with severe intellectual disability. Combined with previous observations, these results support the hypothesis that AP4-complex-mediated trafficking plays a crucial role in brain development and functioning and demonstrate the existence of a clinically recognizable syndrome due to deficiency of the AP4 complex. Copyright © 2011 The American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.

Mutation screening of patients with Alzheimer disease identifies APP locus duplication in a Swedish patient

PubMed Central

2011-01-01

Background Missense mutations in three different genes encoding amyloid-β precursor protein, presenilin 1 and presenilin 2 are recognized to cause familial early-onset Alzheimer disease. Also duplications of the amyloid precursor protein gene have been shown to cause the disease. At the Dept. of Geriatric Medicine, Karolinska University Hospital, Sweden, patients are referred for mutation screening for the identification of nucleotide variations and for determining copy-number of the APP locus. Methods We combined the method of microsatellite marker genotyping with a quantitative real-time PCR analysis to detect duplications in patients with Alzheimer disease. Results In 22 DNA samples from individuals diagnosed with clinical Alzheimer disease, we identified one patient carrying a duplication on chromosome 21 which included the APP locus. Further mapping of the chromosomal region by array-comparative genome hybridization showed that the duplication spanned a maximal region of 1.09 Mb. Conclusions This is the first report of an APP duplication in a Swedish Alzheimer patient and describes the use of quantitative real-time PCR as a tool for determining copy-number of the APP locus. PMID:22044463

Genetic Correction of SOD1 Mutant iPSCs Reveals ERK and JNK Activated AP1 as a Driver of Neurodegeneration in Amyotrophic Lateral Sclerosis.

PubMed

Bhinge, Akshay; Namboori, Seema C; Zhang, Xiaoyu; VanDongen, Antonius M J; Stanton, Lawrence W

2017-04-11

Although mutations in several genes with diverse functions have been known to cause amyotrophic lateral sclerosis (ALS), it is unknown to what extent causal mutations impinge on common pathways that drive motor neuron (MN)-specific neurodegeneration. In this study, we combined induced pluripotent stem cells-based disease modeling with genome engineering and deep RNA sequencing to identify pathways dysregulated by mutant SOD1 in human MNs. Gene expression profiling and pathway analysis followed by pharmacological screening identified activated ERK and JNK signaling as key drivers of neurodegeneration in mutant SOD1 MNs. The AP1 complex member JUN, an ERK/JNK downstream target, was observed to be highly expressed in MNs compared with non-MNs, providing a mechanistic insight into the specific degeneration of MNs. Importantly, investigations of mutant FUS MNs identified activated p38 and ERK, indicating that network perturbations induced by ALS-causing mutations converge partly on a few specific pathways that are drug responsive and provide immense therapeutic potential. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

Mutation screening of patients with Alzheimer disease identifies APP locus duplication in a Swedish patient.

PubMed

Thonberg, Håkan; Fallström, Marie; Björkström, Jenny; Schoumans, Jacqueline; Nennesmo, Inger; Graff, Caroline

2011-11-01

Missense mutations in three different genes encoding amyloid-β precursor protein, presenilin 1 and presenilin 2 are recognized to cause familial early-onset Alzheimer disease. Also duplications of the amyloid precursor protein gene have been shown to cause the disease. At the Dept. of Geriatric Medicine, Karolinska University Hospital, Sweden, patients are referred for mutation screening for the identification of nucleotide variations and for determining copy-number of the APP locus. We combined the method of microsatellite marker genotyping with a quantitative real-time PCR analysis to detect duplications in patients with Alzheimer disease. In 22 DNA samples from individuals diagnosed with clinical Alzheimer disease, we identified one patient carrying a duplication on chromosome 21 which included the APP locus. Further mapping of the chromosomal region by array-comparative genome hybridization showed that the duplication spanned a maximal region of 1.09 Mb. This is the first report of an APP duplication in a Swedish Alzheimer patient and describes the use of quantitative real-time PCR as a tool for determining copy-number of the APP locus.

CFTR rescue with VX-809 and VX-770 favors the repair of primary airway epithelial cell cultures from patients with class II mutations in the presence of Pseudomonas aeruginosa exoproducts.

PubMed

Adam, Damien; Bilodeau, Claudia; Sognigbé, Laura; Maillé, Émilie; Ruffin, Manon; Brochiero, Emmanuelle

2018-04-13

Progressive airway damage due to bacterial infections, especially with Pseudomonas aeruginosa remains the first cause of morbidity and mortality in CF patients. Our previous work revealed a repair delay in CF airway epithelia compared to non-CF. This delay was partially prevented after CFTR correction (with VRT-325) in the absence of infection. Our goals were now to evaluate the effect of the Orkambi combination (CFTR VX-809 corrector + VX-770 potentiator) on the repair of CF primary airway epithelia, in infectious conditions. Primary airway epithelial cell cultures from patients with class II mutations were mechanically injured and wound healing rates and transepithelial resistances were monitored after CFTR rescue, in the absence and presence of P. aeruginosa exoproducts. Our data revealed that combined treatment with VX-809 and VX-770 elicited a greater beneficial impact on airway epithelial repair than VX-809 alone, in the absence of infection. The treatment with Orkambi was effective not only in airway epithelial cell cultures from patients homozygous for the F508del mutation but also from heterozygous patients carrying F508del and another class II mutation (N1303 K, I507del). The stimulatory effect of the Orkambi treatment was prevented by CFTR inhibition with GlyH101. Finally, Orkambi combination elicited a slight but significant improvement in airway epithelial repair and transepithelial resistance, despite the presence of P. aeruginosa exoproducts. Our findings indicate that Orkambi may favor airway epithelial integrity in CF patients with class II mutations. Complementary approaches would however be needed to further improve CFTR rescue and airway epithelial repair. Copyright © 2018 European Cystic Fibrosis Society. Published by Elsevier B.V. All rights reserved.

Two novel LHX3 mutations in patients with combined pituitary hormone deficiency including cervical rigidity and sensorineural hearing loss.

PubMed

Ramzan, Khushnooda; Bin-Abbas, Bassam; Al-Jomaa, Lolwa; Allam, Rabab; Al-Owain, Mohammed; Imtiaz, Faiqa

2017-03-16

Congenital combined pituitary hormone deficiency (CPHD) is a rare heterogeneous group of conditions. CPHD-type 3 (CPHD3; MIM# 221750) is caused by recessive mutations in LHX3, a LIM-homeodomain transcription factor gene. The isoforms of LHX3 are critical for pituitary gland formation and specification of the anterior pituitary hormone-secreting cell types. They also play distinct roles in the development of neuroendocrine and auditory systems. Here, we summarize the clinical, endocrinological, radiological and molecular features of three patients from two unrelated families. Clinical evaluation revealed severe CPHD coupled with cervical vertebral malformations (rigid neck, scoliosis), mild developmental delay and moderate sensorineural hearing loss (SNHL). The patients were diagnosed with CPHD3 based on the array of hormone deficiencies and other associated syndromic symptoms, suggestive of targeted LHX3 gene sequencing. A novel missense mutation c.437G > T (p. Cys146Phe) and a novel nonsense mutation c.466C > T (p. Arg156Ter), both in homozygous forms, were found. The altered Cys146 resides in the LIM2 domain of the encoded protein and is a phylogenetically conserved residue, which mediates LHX3 transcription factor binding with a zinc cation. The p. Arg156Ter is predicted to result in a severely truncated protein, lacking the DNA binding homeodomain. Considering genotype/phenotype correlation, we suggest that the presence of SNHL and limited neck rotation should be considered in the differential diagnosis of CPHD3 to facilitate molecular diagnosis. This report describes the first LHX3 mutations from Saudi patients and highlights the importance of combining molecular diagnosis with the clinical findings. In addition, it also expands the knowledge of LHX3-related CPHD3 phenotype and the allelic spectrum for this gene.

Phase IB Study of Vemurafenib in Combination with Irinotecan and Cetuximab in Patients with Metastatic Colorectal Cancer with BRAFV600E Mutation.

PubMed

Hong, David S; Morris, Van K; El Osta, Badi; Sorokin, Alexey V; Janku, Filip; Fu, Siqing; Overman, Michael J; Piha-Paul, Sarina; Subbiah, Vivek; Kee, Bryan; Tsimberidou, Apostolia M; Fogelman, David; Bellido, Jorge; Shureiqi, Imad; Huang, Helen; Atkins, Johnique; Tarcic, Gabi; Sommer, Nicolas; Lanman, Richard; Meric-Bernstam, Funda; Kopetz, Scott

2016-12-01

In vitro, EGFR inhibition, combined with the BRAF inhibitor vemurafenib, causes synergistic cytotoxicity for BRAF V600E metastatic colorectal cancer, further augmented by irinotecan. The safety and efficacy of vemurafenib, irinotecan, and cetuximab in BRAF-mutated malignancies are not defined. In this 3+3 phase I study, patients with BRAF V600E -advanced solid cancers received cetuximab and irinotecan with escalating doses of vemurafenib. Nineteen patients (18 with metastatic colorectal cancer and 1 with appendiceal cancer) were enrolled. Three patients experienced dose-limiting toxicities. The MTD of vemurafenib was 960 mg twice daily. Six of 17 evaluable patients (35%) achieved a radiographic response by Response Evaluation Criteria in Solid Tumors 1.1 criteria, consistent with in vivo models demonstrating tumor regressions with the triplet regimen. Median progression-free survival was 7.7 months. BRAF V600E circulating cell-free DNA (cfDNA) trends correlated with radiographic changes, and acquired mutations from cfDNA in genes reactivating MAPK signaling were observed at progression. Vemurafenib, in combination with irinotecan and cetuximab, was well tolerated in patients with refractory, BRAF-mutated metastatic colorectal cancer, and both survival outcomes and response rates exceeded prior reports for vemurafenib and for irinotecan plus cetuximab in BRAF V600E metastatic colorectal cancer. In vivo models demonstrated regressions with the triplet, in contrast with vemurafenib and cetuximab alone. cfDNA predicted radiographic response and identified mutations reactivating the MAPK pathway upon progression. Cancer Discov; 6(12); 1352-65. ©2016 AACR.This article is highlighted in the In This Issue feature, p. 1293. ©2016 American Association for Cancer Research.

Potentiator Ivacaftor Abrogates Pharmacological Correction of ΔF508 CFTR in Cystic Fibrosis

PubMed Central

Cholon, Deborah M.; Quinney, Nancy L.; Fulcher, M. Leslie; Esther, Charles R.; Das, Jhuma; Dokholyan, Nikolay V.; Randell, Scott H.; Boucher, Richard C.; Gentzsch, Martina

2014-01-01

Cystic Fibrosis (CF) is caused by mutations in the CF transmembrane conductance regulator (CFTR). Newly developed “correctors” such as lumacaftor (VX-809) that improve CFTR maturation and trafficking and “potentiators” such as ivacaftor (VX-770) that enhance channel activity may provide important advances in CF therapy. Although VX-770 has demonstrated substantial clinical efficacy in the small subset of patients with a mutation (G551D) that affects only channel activity, a single compound is not sufficient to treat patients with the more common CFTR mutation, ΔF508. Thus, patients with ΔF508 will likely require treatment with both correctors and potentiators to achieve clinical benefit. However, whereas the effectiveness of acute treatment with this drug combination has been demonstrated in vitro, the impact of chronic therapy has not been established. In studies of human primary airway epithelial cells, we found that both acute and chronic treatment with VX-770 improved CFTR function in cells with the G551D mutation, consistent with clinical studies. In contrast, chronic VX-770 administration caused a dose-dependent reversal of VX-809-mediated CFTR correction in ΔF508 homozygous cultures. This result reflected the destabilization of corrected ΔF508 CFTR by VX-770, dramatically increasing its turnover rate. Chronic VX-770 treatment also reduced mature wild-type CFTR levels and function. These findings demonstrate that chronic treatment with CFTR potentiators and correctors may have unexpected effects that cannot be predicted from short-term studies. Combining of these drugs to maximize rescue of ΔF508 CFTR may require changes in dosing and/or development of new potentiator compounds that do not interfere with CFTR stability. PMID:25101886

A rare case of unprovoked venous thromboembolism manifestation in a young patient with antithrombin Type IIB deficiency combined with inferior vena cava anomaly from Lithuania.

PubMed

Saulytė-Trakymienė, Sonata; Adomaitienė, Irina; Unkrig, Susanne; Oldenburg, Johannes; Ivaškevičius, Vytautas

2017-01-01

Hereditary antithrombin (AT) deficiency is an autosomal-dominant disorder predisposing to venous and arterial thrombosis. Homozygosity resulting in severe AT deficiency is not compatible with life, apart from homozygous mutations affecting the heparin-binding site representing the most severe thrombophilia. A 12-year-old previously healthy boy of Romani origin presented with a swollen, painful left leg and fever. Imaging revealed signs of inferior vena cava (IVC) thrombosis, the presence of interrupted intrahepatic IVC with azygos continuation and bilateral iliofemoral thrombosis with enlargement of the azygous and hemiazygos venous system. In addition, right pleural effusion and signs of bilateral renal pericortical cysts, possibly caused by retroperitoneal lymphangiectasia, were disclosed. Thrombophilia screening involving protein C, Protein S, Antithrombin (chromogenic assays based on the inhibition of FIIa and FXa, antigen concentration), APC resistance, prothrombin mutation and Lupus anticoagulants was performed using standard laboratory methods. Genetic analysis of the SERPINC1 gene was done by direct sequencing. Thrombophilia screening showed isolated decreased AT activity at 21% (RR 80-120%). AT levels were retested and remained decreased (33-36%) on two consecutive occasions. SERPINC1 gene analysis revealed a previously described homozygous mutation (Budapest III) causing type IIB deficiency (c.391C>T; p.Leu131Phe). A family study confirmed the same mutation in both parents and three siblings. The patient improved significantly following warfarin therapy and over the past 2.5 years did not experience new thromboembolism. This case represents a rare combination of two risk factors provoking manifestation of spontaneous venous thromboembolism at a young age requiring permanent anticoagulant therapy. Schattauer GmbH.

Allele-specific Characterization of Alanine: Glyoxylate Aminotransferase Variants Associated with Primary Hyperoxaluria

PubMed Central

Lage, Melissa D.; Pittman, Adrianne M. C.; Roncador, Alessandro; Cellini, Barbara; Tucker, Chandra L.

2014-01-01

Primary Hyperoxaluria Type 1 (PH1) is a rare autosomal recessive kidney stone disease caused by deficiency of the peroxisomal enzyme alanine: glyoxylate aminotransferase (AGT), which is involved in glyoxylate detoxification. Over 75 different missense mutations in AGT have been found associated with PH1. While some of the mutations have been found to affect enzyme activity, stability, and/or localization, approximately half of these mutations are completely uncharacterized. In this study, we sought to systematically characterize AGT missense mutations associated with PH1. To facilitate analysis, we used two high-throughput yeast-based assays: one that assesses AGT specific activity, and one that assesses protein stability. Approximately 30% of PH1-associated missense mutations are found in conjunction with a minor allele polymorphic variant, which can interact to elicit complex effects on protein stability and trafficking. To better understand this allele interaction, we functionally characterized each of 34 mutants on both the major (wild-type) and minor allele backgrounds, identifying mutations that synergize with the minor allele. We classify these mutants into four distinct categories depending on activity/stability results in the different alleles. Twelve mutants were found to display reduced activity in combination with the minor allele, compared with the major allele background. When mapped on the AGT dimer structure, these mutants reveal localized regions of the protein that appear particularly sensitive to interactions with the minor allele variant. While the majority of the deleterious effects on activity in the minor allele can be attributed to synergistic interaction affecting protein stability, we identify one mutation, E274D, that appears to specifically affect activity when in combination with the minor allele. PMID:24718375

IDH1(R132H) mutation causes a less aggressive phenotype and radiosensitizes human malignant glioma cells independent of the oxygenation status.

PubMed

Kessler, Jacqueline; Güttler, Antje; Wichmann, Henri; Rot, Swetlana; Kappler, Matthias; Bache, Matthias; Vordermark, Dirk

2015-09-01

In malignant glioma the presence of the IDH1 mutation (IDH1(R132H)) is associated with better clinical outcome. However, it is unclear whether IDH1 mutation is associated with a less aggressive phenotype or directly linked to increased sensitivity to radiotherapy. We determined the influence of IDH1(R132H) mutant protein on proliferation and growth in 3D culture, migration, cell survival and radiosensitivity in vitro under normoxia (21% O2) and hypoxia (<1% O2) in a panel of human malignant glioma cell lines (U-251MG, U-343MG, LN-229) with stable overexpression of wild-type (IDH1(wt)) and mutated IDH1 (IDH1(R132H)). Overexpression of IDH1(R132H) in glioma cells resulted in slightly decreased cell proliferation, considerably reduced cell migration and caused differences in growth properties in 3D spheroid cultures. Furthermore, IDH1(R132H)-positive cells consistently demonstrated an increased radiosensitivity in human malignant glioma cells U-251MG (DMF10: 1.52, p<0.01 and 1.42, p<0.01), U-343MG (DMF10: 1.78, p<0.01 and 1.75, p<0.01) and LN-229 (DMF10: 1.41, p<0.05 and 1.68, p<0.01) under normoxia and hypoxia, respectively. Our data indicate that IDH1(R132H) mutation causes both a less aggressive biological behavior and direct radiosensitization of human malignant glioma cells. Targeting IDH1 appears to be an attractive approach in combination with radiotherapy. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

A FRMD7 variant in a Japanese family causes congenital nystagmus.

PubMed

Kohmoto, Tomohiro; Okamoto, Nana; Satomura, Shigeko; Naruto, Takuya; Komori, Takahide; Hashimoto, Toshiaki; Imoto, Issei

2015-01-01

Idiopathic congenital nystagmus (ICN) is a genetically heterogeneous eye movement disorder that causes a large proportion of childhood visual impairment. Here we describe a missense variant (p.L292P) within a mutation-rich region of FRMD7 detected in three affected male siblings in a Japanese family with X-linked ICN. Combining sequence analysis and results from structural and functional predictions, we report p.L292P as a variant potentially disrupting FRMD7 function associated with X-linked ICN.

A FRMD7 variant in a Japanese family causes congenital nystagmus

PubMed Central

Kohmoto, Tomohiro; Okamoto, Nana; Satomura, Shigeko; Naruto, Takuya; Komori, Takahide; Hashimoto, Toshiaki; Imoto, Issei

2015-01-01

Idiopathic congenital nystagmus (ICN) is a genetically heterogeneous eye movement disorder that causes a large proportion of childhood visual impairment. Here we describe a missense variant (p.L292P) within a mutation-rich region of FRMD7 detected in three affected male siblings in a Japanese family with X-linked ICN. Combining sequence analysis and results from structural and functional predictions, we report p.L292P as a variant potentially disrupting FRMD7 function associated with X-linked ICN. PMID:27081518

Genetic analyses of bone morphogenetic protein 2, 4 and 7 in congenital combined pituitary hormone deficiency.

PubMed

Breitfeld, Jana; Martens, Susanne; Klammt, Jürgen; Schlicke, Marina; Pfäffle, Roland; Krause, Kerstin; Weidle, Kerstin; Schleinitz, Dorit; Stumvoll, Michael; Führer, Dagmar; Kovacs, Peter; Tönjes, Anke

2013-12-01

The complex process of development of the pituitary gland is regulated by a number of signalling molecules and transcription factors. Mutations in these factors have been identified in rare cases of congenital hypopituitarism but for most subjects with combined pituitary hormone deficiency (CPHD) genetic causes are unknown. Bone morphogenetic proteins (BMPs) affect induction and growth of the pituitary primordium and thus represent plausible candidates for mutational screening of patients with CPHD. We sequenced BMP2, 4 and 7 in 19 subjects with CPHD. For validation purposes, novel genetic variants were genotyped in 1046 healthy subjects. Additionally, potential functional relevance for most promising variants has been assessed by phylogenetic analyses and prediction of effects on protein structure. Sequencing revealed two novel variants and confirmed 30 previously known polymorphisms and mutations in BMP2, 4 and 7. Although phylogenetic analyses indicated that these variants map within strongly conserved gene regions, there was no direct support for their impact on protein structure when applying predictive bioinformatics tools. A mutation in the BMP4 coding region resulting in an amino acid exchange (p.Arg300Pro) appeared most interesting among the identified variants. Further functional analyses are required to ultimately map the relevance of these novel variants in CPHD.

Genetic analyses of bone morphogenetic protein 2, 4 and 7 in congenital combined pituitary hormone deficiency

PubMed Central

2013-01-01

Background The complex process of development of the pituitary gland is regulated by a number of signalling molecules and transcription factors. Mutations in these factors have been identified in rare cases of congenital hypopituitarism but for most subjects with combined pituitary hormone deficiency (CPHD) genetic causes are unknown. Bone morphogenetic proteins (BMPs) affect induction and growth of the pituitary primordium and thus represent plausible candidates for mutational screening of patients with CPHD. Methods We sequenced BMP2, 4 and 7 in 19 subjects with CPHD. For validation purposes, novel genetic variants were genotyped in 1046 healthy subjects. Additionally, potential functional relevance for most promising variants has been assessed by phylogenetic analyses and prediction of effects on protein structure. Results Sequencing revealed two novel variants and confirmed 30 previously known polymorphisms and mutations in BMP2, 4 and 7. Although phylogenetic analyses indicated that these variants map within strongly conserved gene regions, there was no direct support for their impact on protein structure when applying predictive bioinformatics tools. Conclusions A mutation in the BMP4 coding region resulting in an amino acid exchange (p.Arg300Pro) appeared most interesting among the identified variants. Further functional analyses are required to ultimately map the relevance of these novel variants in CPHD. PMID:24289245

Genotypic and phenotypic analysis of 396 individuals with mutations in Sonic Hedgehog.

PubMed

Solomon, Benjamin D; Bear, Kelly A; Wyllie, Adrian; Keaton, Amelia A; Dubourg, Christele; David, Veronique; Mercier, Sandra; Odent, Sylvie; Hehr, Ute; Paulussen, Aimee; Clegg, Nancy J; Delgado, Mauricio R; Bale, Sherri J; Lacbawan, Felicitas; Ardinger, Holly H; Aylsworth, Arthur S; Bhengu, Ntombenhle Louisa; Braddock, Stephen; Brookhyser, Karen; Burton, Barbara; Gaspar, Harald; Grix, Art; Horovitz, Dafne; Kanetzke, Erin; Kayserili, Hulya; Lev, Dorit; Nikkel, Sarah M; Norton, Mary; Roberts, Richard; Saal, Howard; Schaefer, G B; Schneider, Adele; Smith, Erika K; Sowry, Ellen; Spence, M Anne; Shalev, Stavit A; Steiner, Carlos E; Thompson, Elizabeth M; Winder, Thomas L; Balog, Joan Z; Hadley, Donald W; Zhou, Nan; Pineda-Alvarez, Daniel E; Roessler, Erich; Muenke, Maximilian

2012-07-01

Holoprosencephaly (HPE), the most common malformation of the human forebrain, may result from mutations in over 12 genes. Sonic Hedgehog (SHH) was the first such gene discovered; mutations in SHH remain the most common cause of non-chromosomal HPE. The severity spectrum is wide, ranging from incompatibility with extrauterine life to isolated midline facial differences. To characterise genetic and clinical findings in individuals with SHH mutations. Through the National Institutes of Health and collaborating centres, DNA from approximately 2000 individuals with HPE spectrum disorders were analysed for SHH variations. Clinical details were examined and combined with published cases. This study describes 396 individuals, representing 157 unrelated kindreds, with SHH mutations; 141 (36%) have not been previously reported. SHH mutations more commonly resulted in non-HPE (64%) than frank HPE (36%), and non-HPE was significantly more common in patients with SHH than in those with mutations in the other common HPE related genes (p<0.0001 compared to ZIC2 or SIX3). Individuals with truncating mutations were significantly more likely to have frank HPE than those with non-truncating mutations (49% vs 35%, respectively; p=0.012). While mutations were significantly more common in the N-terminus than in the C-terminus (including accounting for the relative size of the coding regions, p=0.00010), no specific genotype-phenotype correlations could be established regarding mutation location. SHH mutations overall result in milder disease than mutations in other common HPE related genes. HPE is more frequent in individuals with truncating mutations, but clinical predictions at the individual level remain elusive.

The R245X mutation of PCDH15 in Ashkenazi Jewish children diagnosed with nonsyndromic hearing loss foreshadows retinitis pigmentosa.

PubMed

Brownstein, Zippora; Ben-Yosef, Tamar; Dagan, Orit; Frydman, Moshe; Abeliovich, Dvorah; Sagi, Michal; Abraham, Fabian A; Taitelbaum-Swead, Riki; Shohat, Mordechai; Hildesheimer, Minka; Friedman, Thomas B; Avraham, Karen B

2004-06-01

Usher syndrome is a frequent cause of the combination of deafness and blindness due to retinitis pigmentosa (RP). Five genes are known to underlie different forms of Usher syndrome type I (USH1). In the Ashkenazi Jewish population, the R245X mutation of the PCDH15 gene may be the most common cause of USH1 (Ben-Yosef T, Ness SL, Madeo AC, Bar-Lev A, Wolfman JH, Ahmed ZM, Desnick RK, Willner JP, Avraham KB, Ostrer H, Oddoux C, Griffith AJ, Friedman TB N Engl J Med 348: 1664-1670, 2003). To estimate what percentage of Ashkenazi Jewish children born with profound hearing loss will develop RP due to R245X, we examined the prevalence of the R245X PCDH15 mutation and its carrier rate among Ashkenazi Jews in Israel. Among probands diagnosed with nonsyndromic hearing loss not due to mutations of connexin 26 (GJB2) and/or connexin 30 (GJB6), and below the age of 10, 2 of 20 (10%) were homozygous for the R245X mutation. Among older nonsyndromic deaf individuals, no homozygotes were detected, although one individual was heterozygous for R245X. The carrier rate of the R245X mutation among the normal hearing Ashkenazi population in Israel was estimated at 1%. Ashkenazi Jewish children with profound prelingual hearing loss should be evaluated for the R245X PCDH15 mutation and undergo ophthalmologic evaluation to determine whether they will develop RP. Rehabilitation can then begin before loss of vision. Early use of cochlear implants in such cases may rescue these individuals from a dual neurosensory deficit.

Recessively inherited multiple epiphyseal dysplasia with normal stature, club foot, and double layered patella caused by a DTDST mutation

PubMed Central

Superti-Furga, A.; Neumann, L.; Riebel, T.; Eich, G.; Steinmann, B.; Spranger, J.; Kunze, J.

1999-01-01

We have observed over 25 different mutations in the diastrophic dysplasia sulphate transporter gene (DTDST) in association with the recessive disorders achondrogenesis 1B, atelosteogenesis 2, and diastrophic dysplasia. The c862t (R279W) transition is the most common mutation in non-Finnish patients, but in these disorders it is usually combined with other DTDST mutations. We had not seen a case of homozygosity for c862t (R279W) until we analysed DNA from a 36 year old male with tall-normal stature (180 cm) who asked for genetic counselling for suspected multiple epiphyseal dysplasia. He was treated for club foot and hip dysplasia at birth. Skeletal changes consistent with multiple epiphyseal dysplasia, with the peculiar finding of a double layered patella, were recognised during childhood. Cleft palate, swelling of the ear pinna, and hitch hiker thumb were absent. He was found to be homozygous, and both healthy parents heterozygous, for the R279W mutation in DTDST, and his fibroblasts showed a sulphate incorporation defect typical of DTDST disorders. Counselling was given for a recessive disorder, thereby considerably reducing the probability of affected offspring.
  Multiple epiphyseal dysplasia is more frequently caused by dominant mutations in the COMP (EDM1, McKusick 132400) and COL9A2 genes (EDM2, McKusick 600204). A few other patients and families with features similar to our proband have been described previously and considered to have autosomal recessive MED (EDM4, McKusick 226900). This observation confirms the existence of this entity and assigns it to the phenotypic spectrum associated with mutations at the DTDST locus.


Keywords: multiple epiphyseal dysplasia; DTDST; double layered patella PMID:10465113

Integrating in silico prediction methods, molecular docking, and molecular dynamics simulation to predict the impact of ALK missense mutations in structural perspective.

PubMed

Doss, C George Priya; Chakraborty, Chiranjib; Chen, Luonan; Zhu, Hailong

2014-01-01

Over the past decade, advancements in next generation sequencing technology have placed personalized genomic medicine upon horizon. Understanding the likelihood of disease causing mutations in complex diseases as pathogenic or neutral remains as a major task and even impossible in the structural context because of its time consuming and expensive experiments. Among the various diseases causing mutations, single nucleotide polymorphisms (SNPs) play a vital role in defining individual's susceptibility to disease and drug response. Understanding the genotype-phenotype relationship through SNPs is the first and most important step in drug research and development. Detailed understanding of the effect of SNPs on patient drug response is a key factor in the establishment of personalized medicine. In this paper, we represent a computational pipeline in anaplastic lymphoma kinase (ALK) for SNP-centred study by the application of in silico prediction methods, molecular docking, and molecular dynamics simulation approaches. Combination of computational methods provides a way in understanding the impact of deleterious mutations in altering the protein drug targets and eventually leading to variable patient's drug response. We hope this rapid and cost effective pipeline will also serve as a bridge to connect the clinicians and in silico resources in tailoring treatments to the patients' specific genotype.

Seamless correction of the sickle cell disease mutation of the HBB gene in human induced pluripotent stem cells using TALENs.

PubMed

Sun, Ning; Zhao, Huimin

2014-05-01

Sickle cell disease (SCD) is the most common human genetic disease which is caused by a single mutation of human β-globin (HBB) gene. The lack of long-term treatment makes the development of reliable cell and gene therapies highly desirable. Disease-specific patient-derived human induced pluripotent stem cells (hiPSCs) have great potential for developing novel cell and gene therapies. With the disease-causing mutations corrected in situ, patient-derived hiPSCs can restore normal cell functions and serve as a renewable autologous cell source for the treatment of genetic disorders. Here we successfully utilized transcription activator-like effector nucleases (TALENs), a recently emerged novel genome editing tool, to correct the SCD mutation in patient-derived hiPSCs. The TALENs we have engineered are highly specific and generate minimal off-target effects. In combination with piggyBac transposon, TALEN-mediated gene targeting leaves no residual ectopic sequences at the site of correction and the corrected hiPSCs retain full pluripotency and a normal karyotype. Our study demonstrates an important first step of using TALENs for the treatment of genetic diseases such as SCD, which represents a significant advance toward hiPSC-based cell and gene therapies. © 2013 Wiley Periodicals, Inc.

Mutations in FLVCR2 associated with Fowler syndrome and survival beyond infancy.

PubMed

Kvarnung, M; Taylan, F; Nilsson, D; Albåge, M; Nordenskjöld, M; Anderlid, B M; Nordgren, A; Syk Lundberg, E

2016-01-01

Proliferative vasculopathy and hydranencephaly-hydrocephaly syndrome (PVHH, OMIM 225790), also known as Fowler syndrome, is a rare autosomal recessive disorder, caused by mutations in FLVCR2. Hallmarks of the syndrome are glomerular vasculopathy in the central nervous system, severe hydrocephaly, hypokinesia and arthrogryphosis. The disorder is considered prenatally lethal. We report the first patients, a brother and a sister, with Fowler syndrome and survival beyond infancy. The patients present a phenotype of severe intellectual and neurologic disability with seizures, absence of functional movements, and no means of communication. Imaging of the brain showed calcifications, profound ventriculomegaly with only a thin edging of the cerebral cortex and hypoplastic cerebellum. Investigation with whole-exome sequencing (WES) revealed, in both patients, a homozygous pathogenic mutation in FLVCR2, c.1289C>T, compatible with a diagnosis of Fowler syndrome. The results highlight the power of combining WES with a thorough clinical examination in order to identify disease-causing mutations in patients whose clinical presentation differs from previously described cases. Specifically, the findings demonstrate that Fowler syndrome is a diagnosis to consider, not only prenatally but also in severely affected children with gross ventriculomegaly on brain imaging. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

cis-Regulatory Mutations Are a Genetic Cause of Human Limb Malformations

PubMed Central

VanderMeer, Julia E.; Ahituv, Nadav

2011-01-01

The underlying mutations that cause human limb malformations are often difficult to determine, particularly for limb malformations that occur as isolated traits. Evidence from a variety of studies shows that cis-regulatory mutations, specifically in enhancers, can lead to some of these isolated limb malformations. Here, we provide a review of human limb malformations that have been shown to be caused by enhancer mutations and propose that cis-regulatory mutations will continue to be identified as the cause of additional human malformations as our understanding of regulatory sequences improves. PMID:21509892

Functional analysis of Waardenburg syndrome-associated PAX3 and SOX10 mutations: report of a dominant-negative SOX10 mutation in Waardenburg syndrome type II.

PubMed

Zhang, Hua; Chen, Hongsheng; Luo, Hunjin; An, Jing; Sun, Lin; Mei, Lingyun; He, Chufeng; Jiang, Lu; Jiang, Wen; Xia, Kun; Li, Jia-Da; Feng, Yong

2012-03-01

Waardenburg syndrome (WS) is an auditory-pigmentary disorder resulting from melanocyte defects, with varying combinations of sensorineural hearing loss and abnormal pigmentation of the hair, skin, and inner ear. WS is classified into four subtypes (WS1-WS4) based on additional symptoms. PAX3 and SOX10 are two transcription factors that can activate the expression of microphthalmia-associated transcription factor (MITF), a critical transcription factor for melanocyte development. Mutations of PAX3 are associated with WS1 and WS3, while mutations of SOX10 cause WS2 and WS4. Recently, we identified some novel WS-associated mutations in PAX3 and SOX10 in a cohort of Chinese WS patients. Here, we further identified an E248fsX30 SOX10 mutation in a family of WS2. We analyzed the subcellular distribution, expression and in vitro activity of two PAX3 mutations (p.H80D, p.H186fsX5) and four SOX10 mutations (p.E248fsX30, p.G37fsX58, p.G38fsX69 and p.R43X). Except H80D PAX3, which retained partial activity, the other mutants were unable to activate MITF promoter. The H80D PAX3 and E248fsX30 SOX10 were localized in the nucleus as wild type (WT) proteins, whereas the other mutant proteins were distributed in both cytoplasm and nucleus. Furthermore, E248fsX30 SOX10 protein retained the DNA-binding activity and showed dominant-negative effect on WT SOX10. However, E248fsX30 SOX10 protein seems to decay faster than the WT one, which may underlie the mild WS2 phenotype caused by this mutation.

Amelogenesis Imperfecta; Genes, Proteins, and Pathways

PubMed Central

Smith, Claire E. L.; Poulter, James A.; Antanaviciute, Agne; Kirkham, Jennifer; Brookes, Steven J.; Inglehearn, Chris F.; Mighell, Alan J.

2017-01-01

Amelogenesis imperfecta (AI) is the name given to a heterogeneous group of conditions characterized by inherited developmental enamel defects. AI enamel is abnormally thin, soft, fragile, pitted and/or badly discolored, with poor function and aesthetics, causing patients problems such as early tooth loss, severe embarrassment, eating difficulties, and pain. It was first described separately from diseases of dentine nearly 80 years ago, but the underlying genetic and mechanistic basis of the condition is only now coming to light. Mutations in the gene AMELX, encoding an extracellular matrix protein secreted by ameloblasts during enamel formation, were first identified as a cause of AI in 1991. Since then, mutations in at least eighteen genes have been shown to cause AI presenting in isolation of other health problems, with many more implicated in syndromic AI. Some of the encoded proteins have well documented roles in amelogenesis, acting as enamel matrix proteins or the proteases that degrade them, cell adhesion molecules or regulators of calcium homeostasis. However, for others, function is less clear and further research is needed to understand the pathways and processes essential for the development of healthy enamel. Here, we review the genes and mutations underlying AI presenting in isolation of other health problems, the proteins they encode and knowledge of their roles in amelogenesis, combining evidence from human phenotypes, inheritance patterns, mouse models, and in vitro studies. An LOVD resource (http://dna2.leeds.ac.uk/LOVD/) containing all published gene mutations for AI presenting in isolation of other health problems is described. We use this resource to identify trends in the genes and mutations reported to cause AI in the 270 families for which molecular diagnoses have been reported by 23rd May 2017. Finally we discuss the potential value of the translation of AI genetics to clinical care with improved patient pathways and speculate on the possibility of novel treatments and prevention strategies for AI. PMID:28694781

Amelogenesis Imperfecta; Genes, Proteins, and Pathways.

PubMed

Smith, Claire E L; Poulter, James A; Antanaviciute, Agne; Kirkham, Jennifer; Brookes, Steven J; Inglehearn, Chris F; Mighell, Alan J

2017-01-01

Amelogenesis imperfecta (AI) is the name given to a heterogeneous group of conditions characterized by inherited developmental enamel defects. AI enamel is abnormally thin, soft, fragile, pitted and/or badly discolored, with poor function and aesthetics, causing patients problems such as early tooth loss, severe embarrassment, eating difficulties, and pain. It was first described separately from diseases of dentine nearly 80 years ago, but the underlying genetic and mechanistic basis of the condition is only now coming to light. Mutations in the gene AMELX , encoding an extracellular matrix protein secreted by ameloblasts during enamel formation, were first identified as a cause of AI in 1991. Since then, mutations in at least eighteen genes have been shown to cause AI presenting in isolation of other health problems, with many more implicated in syndromic AI. Some of the encoded proteins have well documented roles in amelogenesis, acting as enamel matrix proteins or the proteases that degrade them, cell adhesion molecules or regulators of calcium homeostasis. However, for others, function is less clear and further research is needed to understand the pathways and processes essential for the development of healthy enamel. Here, we review the genes and mutations underlying AI presenting in isolation of other health problems, the proteins they encode and knowledge of their roles in amelogenesis, combining evidence from human phenotypes, inheritance patterns, mouse models, and in vitro studies. An LOVD resource (http://dna2.leeds.ac.uk/LOVD/) containing all published gene mutations for AI presenting in isolation of other health problems is described. We use this resource to identify trends in the genes and mutations reported to cause AI in the 270 families for which molecular diagnoses have been reported by 23rd May 2017. Finally we discuss the potential value of the translation of AI genetics to clinical care with improved patient pathways and speculate on the possibility of novel treatments and prevention strategies for AI.

Genetic heterogeneity and consanguinity lead to a “double hit”: Homozygous mutations of MYO7A and PDE6B in a patient with retinitis pigmentosa

PubMed Central

Goldenberg-Cohen, Nitza; Banin, Eyal; Zalzstein, Yael; Cohen, Ben; Rotenstreich, Ygal; Rizel, Leah; Basel-Vanagaite, Lina

2013-01-01

Purpose Retinitis pigmentosa (RP), the most genetically heterogeneous disorder in humans, actually represents a group of pigmentary retinopathies characterized by night blindness followed by visual-field loss. RP can appear as either syndromic or nonsyndromic. One of the most common forms of syndromic RP is Usher syndrome, characterized by the combination of RP, hearing loss, and vestibular dysfunction. Methods The underlying cause of the appearance of syndromic and nonsyndromic RP in three siblings from a consanguineous Israeli Muslim Arab family was studied with whole-genome homozygosity mapping followed by whole exome sequencing. Results The family was found to segregate novel mutations of two different genes: myosin VIIA (MYO7A), which causes type 1 Usher syndrome, and phosphodiesterase 6B, cyclic guanosine monophosphate-specific, rod, beta (PDE6B), which causes nonsyndromic RP. One affected child was homozygous for both mutations. Since the retinal phenotype seen in this patient results from overlapping pathologies, one might expect to find severe retinal degeneration. Indeed, he was diagnosed with RP based on an abnormal electroretinogram (ERG) at a young age (9 months). However, this early diagnosis may be biased, as two of his older siblings had already been diagnosed, leading to increased awareness. At the age of 32 months, he had relatively good vision with normal visual fields. Further testing of visual function and structure at different ages in the three siblings is needed to determine whether the two RP-causing genes mutated in this youngest sibling confer increased disease severity. Conclusions This report further supports the genetic heterogeneity of RP, and demonstrates how consanguinity could increase intrafamilial clustering of multiple hereditary diseases. Moreover, this report provides a unique opportunity to study the clinical implications of the coexistence of pathogenic mutations in two RP-causative genes in a human patient. PMID:23882135

Genetic heterogeneity and consanguinity lead to a "double hit": homozygous mutations of MYO7A and PDE6B in a patient with retinitis pigmentosa.

PubMed

Goldenberg-Cohen, Nitza; Banin, Eyal; Zalzstein, Yael; Cohen, Ben; Rotenstreich, Ygal; Rizel, Leah; Basel-Vanagaite, Lina; Ben-Yosef, Tamar

2013-01-01

Retinitis pigmentosa (RP), the most genetically heterogeneous disorder in humans, actually represents a group of pigmentary retinopathies characterized by night blindness followed by visual-field loss. RP can appear as either syndromic or nonsyndromic. One of the most common forms of syndromic RP is Usher syndrome, characterized by the combination of RP, hearing loss, and vestibular dysfunction. The underlying cause of the appearance of syndromic and nonsyndromic RP in three siblings from a consanguineous Israeli Muslim Arab family was studied with whole-genome homozygosity mapping followed by whole exome sequencing. THE FAMILY WAS FOUND TO SEGREGATE NOVEL MUTATIONS OF TWO DIFFERENT GENES: myosin VIIA (MYO7A), which causes type 1 Usher syndrome, and phosphodiesterase 6B, cyclic guanosine monophosphate-specific, rod, beta (PDE6B), which causes nonsyndromic RP. One affected child was homozygous for both mutations. Since the retinal phenotype seen in this patient results from overlapping pathologies, one might expect to find severe retinal degeneration. Indeed, he was diagnosed with RP based on an abnormal electroretinogram (ERG) at a young age (9 months). However, this early diagnosis may be biased, as two of his older siblings had already been diagnosed, leading to increased awareness. At the age of 32 months, he had relatively good vision with normal visual fields. Further testing of visual function and structure at different ages in the three siblings is needed to determine whether the two RP-causing genes mutated in this youngest sibling confer increased disease severity. This report further supports the genetic heterogeneity of RP, and demonstrates how consanguinity could increase intrafamilial clustering of multiple hereditary diseases. Moreover, this report provides a unique opportunity to study the clinical implications of the coexistence of pathogenic mutations in two RP-causative genes in a human patient.

Molecular diagnosis of α-thalassemia in a multiethnic population.

PubMed

Gilad, Oded; Shemer, Orna Steinberg; Dgany, Orly; Krasnov, Tanya; Nevo, Michal; Noy-Lotan, Sharon; Rabinowicz, Ron; Amitai, Nofar; Ben-Dor, Shifra; Yaniv, Isaac; Yacobovich, Joanne; Tamary, Hannah

2017-06-01

α-Thalassemia, one of the most common genetic diseases, is caused by deletions or point mutations affecting one to four α-globin genes. Molecular diagnosis is important to prevent the most severe forms of the disease. However, the diagnosis of α-thalassemia is complex due to a high variability of the genetic defects involved, with over 250 described mutations. We summarize herein the findings of genetic analyses of DNA samples referred to our laboratory for the molecular diagnosis of α-thalassemia, along with a detailed clinical description. We utilized a diagnostic algorithm including Gap-PCR, to detect known deletions, followed by sequencing of the α-globin gene, to identify known and novel point mutations, and multiplex ligation-dependent probe amplification (MLPA) for the diagnosis of rare or novel deletions. α-Thalassemia was diagnosed in 662 of 975 samples referred to our laboratory. Most commonly found were deletions (75.3%, including two novel deletions previously described by us); point mutations comprised 25.4% of the cases, including five novel mutations. Our population included mostly Jews (of Ashkenazi and Sephardic origin) and Muslim Arabs, who presented with a higher rate of point mutations and hemoglobin H disease. Overall, we detected 53 different genotype combinations causing a spectrum of clinical phenotypes, from asymptomatic to severe anemia. Our work constitutes the largest group of patients with α-thalassemia originating in the Mediterranean whose clinical characteristics and molecular basis have been determined. We suggest a diagnostic algorithm that leads to an accurate molecular diagnosis in multiethnic populations. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Detection of Clinically Relevant Genetic Variants in Autism Spectrum Disorder by Whole-Genome Sequencing

PubMed Central

Jiang, Yong-hui; Yuen, Ryan K.C.; Jin, Xin; Wang, Mingbang; Chen, Nong; Wu, Xueli; Ju, Jia; Mei, Junpu; Shi, Yujian; He, Mingze; Wang, Guangbiao; Liang, Jieqin; Wang, Zhe; Cao, Dandan; Carter, Melissa T.; Chrysler, Christina; Drmic, Irene E.; Howe, Jennifer L.; Lau, Lynette; Marshall, Christian R.; Merico, Daniele; Nalpathamkalam, Thomas; Thiruvahindrapuram, Bhooma; Thompson, Ann; Uddin, Mohammed; Walker, Susan; Luo, Jun; Anagnostou, Evdokia; Zwaigenbaum, Lonnie; Ring, Robert H.; Wang, Jian; Lajonchere, Clara; Wang, Jun; Shih, Andy; Szatmari, Peter; Yang, Huanming; Dawson, Geraldine; Li, Yingrui; Scherer, Stephen W.

2013-01-01

Autism Spectrum Disorder (ASD) demonstrates high heritability and familial clustering, yet the genetic causes remain only partially understood as a result of extensive clinical and genomic heterogeneity. Whole-genome sequencing (WGS) shows promise as a tool for identifying ASD risk genes as well as unreported mutations in known loci, but an assessment of its full utility in an ASD group has not been performed. We used WGS to examine 32 families with ASD to detect de novo or rare inherited genetic variants predicted to be deleterious (loss-of-function and damaging missense mutations). Among ASD probands, we identified deleterious de novo mutations in six of 32 (19%) families and X-linked or autosomal inherited alterations in ten of 32 (31%) families (some had combinations of mutations). The proportion of families identified with such putative mutations was larger than has been previously reported; this yield was in part due to the comprehensive and uniform coverage afforded by WGS. Deleterious variants were found in four unrecognized, nine known, and eight candidate ASD risk genes. Examples include CAPRIN1 and AFF2 (both linked to FMR1, which is involved in fragile X syndrome), VIP (involved in social-cognitive deficits), and other genes such as SCN2A and KCNQ2 (linked to epilepsy), NRXN1, and CHD7, which causes ASD-associated CHARGE syndrome. Taken together, these results suggest that WGS and thorough bioinformatic analyses for de novo and rare inherited mutations will improve the detection of genetic variants likely to be associated with ASD or its accompanying clinical symptoms. PMID:23849776

Calmodulin Mutations Associated with Recurrent Cardiac Arrest in Infants

PubMed Central

Crotti, Lia; Johnson, Christopher N.; Graf, Elisabeth; De Ferrari, Gaetano M.; Cuneo, Bettina F.; Ovadia, Marc; Papagiannis, John; Feldkamp, Michael D.; Rathi, Subodh G.; Kunic, Jennifer D.; Pedrazzini, Matteo; Wieland, Thomas; Lichtner, Peter; Beckmann, Britt-Maria; Clark, Travis; Shaffer, Christian; Benson, D. Woodrow; Kääb, Stefan; Meitinger, Thomas; Strom, Tim M.; Chazin, Walter J.; Schwartz, Peter J.; George, Alfred L.

2013-01-01

Background Life-threatening disorders of heart rhythm may arise during infancy and can result in the sudden and tragic death of a child. We performed exome sequencing on two unrelated infants presenting with recurrent cardiac arrest to discover a genetic cause. Methods and Results We ascertained two unrelated infants (probands) with recurrent cardiac arrest and dramatically prolonged QTc interval who were both born to healthy parents. The two parent-child trios were investigated using exome sequencing to search for de novo genetic variants. We then performed follow-up candidate gene screening on an independent cohort of 82 subjects with congenital long-QT syndrome without an identified genetic cause. Biochemical studies were performed to determine the functional consequences of mutations discovered in two genes encoding calmodulin. We discovered three heterozygous de novo mutations in either CALM1 or CALM2, two of the three human genes encoding calmodulin, in the two probands and in two additional subjects with recurrent cardiac arrest. All mutation carriers were infants who exhibited life-threatening ventricular arrhythmias combined variably with epilepsy and delayed neurodevelopment. Mutations altered residues in or adjacent to critical calcium binding loops in the calmodulin carboxyl-terminal domain. Recombinant mutant calmodulins exhibited several fold reductions in calcium binding affinity. Conclusions Human calmodulin mutations disrupt calcium ion binding to the protein and are associated with a life-threatening condition in early infancy. Defects in calmodulin function will disrupt important calcium signaling events in heart affecting membrane ion channels, a plausible molecular mechanism for potentially deadly disturbances in heart rhythm during infancy. PMID:23388215

The identification of HESX1 mutations in Kallmann syndrome

PubMed Central

Newbern, Kayce; Natrajan, Nithya; Kim, Hyung-Goo; Chorich, Lynn .P.; Halvorson, Lisa; Cameron, Richard S.; Layman, Lawrence C.

2013-01-01

Objective To determine if HESX1 mutations are present in patients with idiopathic hypogonadotropic hypogonadism (IHH)/Kallmann syndrome (KS). HESX1 mutations have previously been characterized in patients with septo-optic dysplasia (SOD), isolated growth hormone deficiency (IGHD), and combined pituitary hormone deficiency (CPHD). We hypothesized that IHH/KS represents a milder phenotypic variant of SOD. Design PCR-based DNA sequencing was performed on 217 well-characterized IHH/KS patients. Putative missense mutations were analyzed by sorting intolerant from tolerant (SIFT) and Clustal Ω. Setting An academic medical center Patients 217 IHH/KS and 192 controls Interventions DNA was extracted from patients and controls; genotype/phenotype comparisons were made Main Outcome Measures DNA sequence of HESX1, SIFT analysis, and ortholog alignment Results Two novel heterozygous missense mutations (p.H42Y and p.V75L) and previously reported heterozygous missense mutation p.Q6H in HESX1 were identified in 3/217 (1.4%) patients. All were males with KS. Both p.Q6H and p.H42Y were predicted to be deleterious by SIFT, while p.V75L was conserved in 8/9 species. No other IHH/KS gene mutations were present. Conclusions HESX1 mutations may cause KS in addition to more severe phenotypes. Our findings expand the phenotypic spectrum of HESX1 mutations in humans, thereby broadening its role in development. PMID:23465708

Next-generation sequencing to solve complex inherited retinal dystrophy: A case series of multiple genes contributing to disease in extended families.

PubMed

Jones, Kaylie D; Wheaton, Dianna K; Bowne, Sara J; Sullivan, Lori S; Birch, David G; Chen, Rui; Daiger, Stephen P

2017-01-01

With recent availability of next-generation sequencing (NGS), it is becoming more common to pursue disease-targeted panel testing rather than traditional sequential gene-by-gene dideoxy sequencing. In this report, we describe using NGS to identify multiple disease-causing mutations that contribute concurrently or independently to retinal dystrophy in three relatively small families. Family members underwent comprehensive visual function evaluations, and genetic counseling including a detailed family history. A preliminary genetic inheritance pattern was assigned and updated as additional family members were tested. Family 1 (FAM1) and Family 2 (FAM2) were clinically diagnosed with retinitis pigmentosa (RP) and had a suspected autosomal dominant pedigree with non-penetrance (n.p.). Family 3 (FAM3) consisted of a large family with a diagnosis of RP and an overall dominant pedigree, but the proband had phenotypically cone-rod dystrophy. Initial genetic analysis was performed on one family member with traditional Sanger single gene sequencing and/or panel-based testing, and ultimately, retinal gene-targeted NGS was required to identify the underlying cause of disease for individuals within the three families. Results obtained in these families necessitated further genetic and clinical testing of additional family members to determine the complex genetic and phenotypic etiology of each family. Genetic testing of FAM1 (n = 4 affected; 1 n.p.) identified a dominant mutation in RP1 (p.Arg677Ter) that was present for two of the four affected individuals but absent in the proband and the presumed non-penetrant individual. Retinal gene-targeted NGS in the fourth affected family member revealed compound heterozygous mutations in USH2A (p. Cys419Phe, p.Glu767Serfs*21). Genetic testing of FAM2 (n = 3 affected; 1 n.p.) identified three retinal dystrophy genes ( PRPH2 , PRPF8 , and USH2A ) with disease-causing mutations in varying combinations among the affected family members. Genetic testing of FAM3 (n = 7 affected) identified a mutation in PRPH2 (p.Pro216Leu) tracking with disease in six of the seven affected individuals. Additional retinal gene-targeted NGS testing determined that the proband also harbored a multiple exon deletion in the CRX gene likely accounting for her cone-rod phenotype; her son harbored only the mutation in CRX , not the familial mutation in PRPH2 . Multiple genes contributing to the retinal dystrophy genotypes within a family were discovered using retinal gene-targeted NGS. Families with noted examples of phenotypic variation or apparent non-penetrant individuals may offer a clue to suspect complex inheritance. Furthermore, this finding underscores that caution should be taken when attributing a single gene disease-causing mutation (or inheritance pattern) to a family as a whole. Identification of a disease-causing mutation in a proband, even with a clear inheritance pattern in hand, may not be sufficient for targeted, known mutation analysis in other family members.

Unique presentation of cutis laxa with Leigh-like syndrome due to ECHS1 deficiency.

PubMed

Balasubramaniam, S; Riley, L G; Bratkovic, D; Ketteridge, D; Manton, N; Cowley, M J; Gayevskiy, V; Roscioli, T; Mohamed, M; Gardeitchik, T; Morava, E; Christodoulou, J

2017-09-01

Clinical finding of cutis laxa, characterized by wrinkled, redundant, sagging, nonelastic skin, is of growing significance due to its occurrence in several different inborn errors of metabolism (IEM). Metabolic cutis laxa results from Menkes syndrome, caused by a defect in the ATPase copper transporting alpha (ATP7A) gene; congenital disorders of glycosylation due to mutations in subunit 7 of the component of oligomeric Golgi (COG7)-congenital disorders of glycosylation (CDG) complex; combined disorder of N- and O-linked glycosylation, due to mutations in ATPase H+ transporting V0 subunit a2 (ATP6VOA2) gene; pyrroline-5-carboxylate reductase 1 deficiency; pyrroline-5-carboxylate synthase deficiency; macrocephaly, alopecia, cutis laxa, and scoliosis (MACS) syndrome, due to Ras and Rab interactor 2 (RIN2) mutations; transaldolase deficiency caused by mutations in the transaldolase 1 (TALDO1) gene; Gerodermia osteodysplastica due to mutations in the golgin, RAB6-interacting (GORAB or SCYL1BP1) gene; and mitogen-activated pathway (MAP) kinase defects, caused by mutations in several genes [protein tyrosine phosphatase, non-receptor-type 11 (PTPN11), RAF, NF, HRas proto-oncogene, GTPase (HRAS), B-Raf proto-oncogene, serine/threonine kinase (BRAF), MEK1/2, KRAS proto-oncogene, GTPase (KRAS), SOS Ras/Rho guanine nucleotide exchange factor 2 (SOS2), leucine rich repeat scaffold protein (SHOC2), NRAS proto-oncogene, GTPase (NRAS), and Raf-1 proto-oncogene, serine/threonine kinase (RAF1)], which regulate the Ras-MAPK cascade. Here, we further expand the list of inborn errors of metabolism associated with cutis laxa by describing the clinical presentation of a 17-month-old girl with Leigh-like syndrome due to enoyl coenzyme A hydratase, short chain, 1, mitochondria (ECHS1) deficiency, a mitochondrial matrix enzyme that catalyzes the second step of the beta-oxidation spiral of fatty acids and plays an important role in amino acid catabolism, particularly valine.

Analysis of transcription factors key for mouse pancreatic development establishes NKX2-2 and MNX1 mutations as causes of neonatal diabetes in man.

PubMed

Flanagan, Sarah E; De Franco, Elisa; Lango Allen, Hana; Zerah, Michele; Abdul-Rasoul, Majedah M; Edge, Julie A; Stewart, Helen; Alamiri, Elham; Hussain, Khalid; Wallis, Sam; de Vries, Liat; Rubio-Cabezas, Oscar; Houghton, Jayne A L; Edghill, Emma L; Patch, Ann-Marie; Ellard, Sian; Hattersley, Andrew T

2014-01-07

Understanding transcriptional regulation of pancreatic development is required to advance current efforts in developing beta cell replacement therapies for patients with diabetes. Current knowledge of key transcriptional regulators has predominantly come from mouse studies, with rare, naturally occurring mutations establishing their relevance in man. This study used a combination of homozygosity analysis and Sanger sequencing in 37 consanguineous patients with permanent neonatal diabetes to search for homozygous mutations in 29 transcription factor genes important for murine pancreatic development. We identified homozygous mutations in 7 different genes in 11 unrelated patients and show that NKX2-2 and MNX1 are etiological genes for neonatal diabetes, thus confirming their key role in development of the human pancreas. The similar phenotype of the patients with recessive mutations and mice with inactivation of a transcription factor gene support there being common steps critical for pancreatic development and validate the use of rodent models for beta cell development. Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.

Iris phenotypes and pigment dispersion caused by genes influencing pigmentation

PubMed Central

Hawes, Norman L.; Trantow, Colleen M.; Chang, Bo; John, Simon W.M.

2010-01-01

Summary Spontaneous mutations altering mouse coat colors have been a classic resource for discovery of numerous molecular pathways. Although often overlooked, the mouse iris is also densely pigmented and easily observed, thus representing a similarly powerful opportunity for studying pigment cell biology. Here, we present an analysis of iris phenotypes among sixteen mouse strains with mutations influencing melanosomes. Many of these strains exhibit biologically and medically relevant phenotypes, including pigment dispersion, a common feature of several human ocular diseases. Pigment dispersion was identified in several strains with mutant alleles known to influence melanosomes, including beige, light, and vitiligo. Pigment dispersion was also detected in the recently arising spontaneous coat color variant, nm2798. We have identified the nm2798 mutation as a missense mutation in the Dct gene, an identical re-occurrence of the slaty light mutation. These results suggest that dysregulated events of melanosomes can be potent contributors to the pigment dispersion phenotype. Combined, these findings illustrate the utility of studying iris phenotypes as a means of discovering new pathways, and re-linking old ones, to processes of pigmented cells in health and disease. PMID:18715234

Iris phenotypes and pigment dispersion caused by genes influencing pigmentation.

PubMed

Anderson, Michael G; Hawes, Norman L; Trantow, Colleen M; Chang, Bo; John, Simon W M

2008-10-01

Spontaneous mutations altering mouse coat colors have been a classic resource for discovery of numerous molecular pathways. Although often overlooked, the mouse iris is also densely pigmented and easily observed, thus representing a similarly powerful opportunity for studying pigment cell biology. Here, we present an analysis of iris phenotypes among 16 mouse strains with mutations influencing melanosomes. Many of these strains exhibit biologically and medically relevant phenotypes, including pigment dispersion, a common feature of several human ocular diseases. Pigment dispersion was identified in several strains with mutant alleles known to influence melanosomes, including beige, light, and vitiligo. Pigment dispersion was also detected in the recently arising spontaneous coat color variant, nm2798. We have identified the nm2798 mutation as a missense mutation in the Dct gene, an identical re-occurrence of the slaty light mutation. These results suggest that dysregulated events of melanosomes can be potent contributors to the pigment dispersion phenotype. Combined, these findings illustrate the utility of studying iris phenotypes as a means of discovering new pathways, and re-linking old ones, to processes of pigmented cells in health and disease.

Electron holes appear to trigger cancer-implicated mutations

NASA Astrophysics Data System (ADS)

Miller, John; Villagran, Martha

Malignant tumors are caused by mutations, which also affect their subsequent growth and evolution. We use a novel approach, computational DNA hole spectroscopy [M.Y. Suarez-Villagran & J.H. Miller, Sci. Rep. 5, 13571 (2015)], to compute spectra of enhanced hole probability based on actual sequence data. A hole is a mobile site of positive charge created when an electron is removed, for example by radiation or contact with a mutagenic agent. Peaks in the hole spectrum depict sites where holes tend to localize and potentially trigger a base pair mismatch during replication. Our studies of reveal a correlation between hole spectrum peaks and spikes in human mutation frequencies. Importantly, we also find that hole peak positions that do not coincide with large variant frequencies often coincide with cancer-implicated mutations and/or (for coding DNA) encoded conserved amino acids. This enables combining hole spectra with variant data to identify critical base pairs and potential cancer `driver' mutations. Such integration of DNA hole and variance spectra could also prove invaluable for pinpointing critical regions, and sites of driver mutations, in the vast non-protein-coding genome. Supported by the State of Texas through the Texas Ctr. for Superconductivity.

PPM1D Mosaic Truncating Variants in Ovarian Cancer Cases May Be Treatment-Related Somatic Mutations.

PubMed

Pharoah, Paul D P; Song, Honglin; Dicks, Ed; Intermaggio, Maria P; Harrington, Patricia; Baynes, Caroline; Alsop, Kathryn; Bogdanova, Natalia; Cicek, Mine S; Cunningham, Julie M; Fridley, Brooke L; Gentry-Maharaj, Aleksandra; Hillemanns, Peter; Lele, Shashi; Lester, Jenny; McGuire, Valerie; Moysich, Kirsten B; Poblete, Samantha; Sieh, Weiva; Sucheston-Campbell, Lara; Widschwendter, Martin; Whittemore, Alice S; Dörk, Thilo; Menon, Usha; Odunsi, Kunle; Goode, Ellen L; Karlan, Beth Y; Bowtell, David D; Gayther, Simon A; Ramus, Susan J

2016-03-01

Mosaic truncating mutations in the protein phosphatase, Mg(2+)/Mn(2+)-dependent, 1D (PPM1D) gene have recently been reported with a statistically significantly greater frequency in lymphocyte DNA from ovarian cancer case patients compared with unaffected control patients. Using massively parallel sequencing (MPS) we identified truncating PPM1D mutations in 12 of 3236 epithelial ovarian cancer (EOC) case patients (0.37%) but in only one of 3431 unaffected control patients (0.03%) (P = .001). All statistical tests were two-sided. A combination of Sanger sequencing, pyrosequencing, and MPS data suggested that 12 of the 13 mutations were mosaic. All mutations were identified in post-chemotherapy treatment blood samples from case patients (n = 1827) (average 1234 days post-treatment in carriers) rather than from cases collected pretreatment (less than 14 days after diagnosis, n = 1384) (P = .002). These data suggest that PPM1D variants in EOC cases are primarily somatic mosaic mutations caused by treatment and are not associated with germline predisposition to EOC. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Clinical and Molecular Aspects of an Informative Family with Neurofibromatosis Type 1 and Noonan Phenotype

PubMed Central

Stevenson, David A.; Viskochil, David H.; Rope, Alan F.; Carey, John C.

2011-01-01

NF-Noonan syndrome (NFNS) has been described as a unique phenotype, combining manifestations of neurofibromatosis type 1 (NF1) and Noonan syndromes, which are separate syndromes. Potential etiologies of NF-Noonan syndrome include a discrete syndrome of distinct etiology, co-segregation of two mutated common genes, variable clinical expressivity of NF1, and/or allelic heterogeneity. We present an informative family with an unusual NF1 mutation with variable features of NF1 and Noonan syndrome. We hypothesize that an NF1 mutant allele can lead to diagnostic manifestations of Noonan syndrome, supporting the hypothesis that NF1 allelic heterogeneity causes NFNS. PMID:16542390

Comprehensive analysis of Stargardt macular dystrophy patients reveals new genotype-phenotype correlations and unexpected diagnostic revisions

PubMed Central

Zaneveld, Jacques; Siddiqui, Sorath; Li, Huajin; Wang, Xia; Wang, Hui; Wang, Keqing; Li, Hui; Ren, Huanan; Lopez, Irma; Dorfman, Allison; Khan, Ayesha; Wang, Feng; Salvo, Jason; Gelowani, Violet; Li, Yumei; Sui, Ruifang; Koenekoop, Robert; Chen, Rui

2014-01-01

Purpose Stargardt macular dystrophy (STGD) results in early central vision loss. We sought to explain the genetic cause of STGD in a cohort of 88 patients from three different cultural backgrounds. Methods Next Generation Sequencing using a novel capture panel was used to search for disease causing mutations. Unsolved patients were clinically re-examined and tested for copy number variations (CNVs) as well as intronic mutations. Results We determined the cause of disease in 67% of our patients. Our analysis identified 35 novel ABCA4 alleles. Eleven patients had mutations in genes not previously reported to cause STGD. Finally, 45% of our unsolved patients had single deleterious mutations in ABCA4, a recessive disease gene. No likely pathogenic CNVs were identified. Conclusions This study expands our knowledge of STGD by identifying dozens of novel STGD causing alleles. The frequency of patients with single mutations in ABCA4 is higher than controls, indicating these mutations contribute to disease. Eleven patients were explained by mutations outside ABCA4 underlining the need to genotype all retinal disease genes to maximize genetic diagnostic rates. Few ABCA4 mutations were observed in our French Canadian patients. This population may contain an unidentified founder mutation. Our results indicate that CNVs are unlikely to be a major cause of STGD. PMID:25474345

Both TALENs and CRISPR/Cas9 directly target the HBB IVS2–654 (C > T) mutation in β-thalassemia-derived iPSCs

PubMed Central

Xu, Peng; Tong, Ying; Liu, Xiu-zhen; Wang, Ting-ting; Cheng, Li; Wang, Bo-yu; Lv, Xiang; Huang, Yue; Liu, De-pei

2015-01-01

β-Thalassemia is one of the most common genetic blood diseases and is caused by either point mutations or deletions in the β-globin (HBB) gene. The generation of patient-specific induced pluripotent stem cells (iPSCs) and subsequent correction of the disease-causing mutations may be a potential therapeutic strategy for this disease. Due to the low efficiency of typical homologous recombination, endonucleases, including TALENs and CRISPR/Cas9, have been widely used to enhance the gene correction efficiency in patient-derived iPSCs. Here, we designed TALENs and CRISPR/Cas9 to directly target the intron2 mutation site IVS2-654 in the globin gene. We observed different frequencies of double-strand breaks (DSBs) at IVS2-654 loci using TALENs and CRISPR/Cas9, and TALENs mediated a higher homologous gene targeting efficiency compared to CRISPR/Cas9 when combined with the piggyBac transposon donor. In addition, more obvious off-target events were observed for CRISPR/Cas9 compared to TALENs. Finally, TALENs-corrected iPSC clones were selected for erythroblast differentiation using the OP9 co-culture system and detected relatively higher transcription of HBB than the uncorrected cells. This comparison of using TALENs or CRISPR/Cas9 to correct specific HBB mutations in patient-derived iPSCs will guide future applications of TALENs- or CRISPR/Cas9-based gene therapies in monogenic diseases. PMID:26156589

Both TALENs and CRISPR/Cas9 directly target the HBB IVS2-654 (C > T) mutation in β-thalassemia-derived iPSCs.

PubMed

Xu, Peng; Tong, Ying; Liu, Xiu-zhen; Wang, Ting-ting; Cheng, Li; Wang, Bo-yu; Lv, Xiang; Huang, Yue; Liu, De-pei

2015-07-09

β-Thalassemia is one of the most common genetic blood diseases and is caused by either point mutations or deletions in the β-globin (HBB) gene. The generation of patient-specific induced pluripotent stem cells (iPSCs) and subsequent correction of the disease-causing mutations may be a potential therapeutic strategy for this disease. Due to the low efficiency of typical homologous recombination, endonucleases, including TALENs and CRISPR/Cas9, have been widely used to enhance the gene correction efficiency in patient-derived iPSCs. Here, we designed TALENs and CRISPR/Cas9 to directly target the intron2 mutation site IVS2-654 in the globin gene. We observed different frequencies of double-strand breaks (DSBs) at IVS2-654 loci using TALENs and CRISPR/Cas9, and TALENs mediated a higher homologous gene targeting efficiency compared to CRISPR/Cas9 when combined with the piggyBac transposon donor. In addition, more obvious off-target events were observed for CRISPR/Cas9 compared to TALENs. Finally, TALENs-corrected iPSC clones were selected for erythroblast differentiation using the OP9 co-culture system and detected relatively higher transcription of HBB than the uncorrected cells. This comparison of using TALENs or CRISPR/Cas9 to correct specific HBB mutations in patient-derived iPSCs will guide future applications of TALENs- or CRISPR/Cas9-based gene therapies in monogenic diseases.

Biochemical and genetic analysis of Leigh syndrome patients in Korea.

PubMed

Chae, Jong-Hee; Lee, Jin Sook; Kim, Ki Joong; Hwang, Yong Seung; Hirano, Michio

2008-06-01

Sixteen Korean patients with Leigh syndrome were identified at the Seoul National University Children's Hospital in 2001-2006. Biochemical or molecular defects were identified in 14 patients (87.5%). Thirteen patients had respiratory chain enzyme defects; 9 had complex I deficiency, and 4 had combined defects of complex I+III+IV. Based on the biochemical defects, targeted genetic studies in 4 patients with complex I deficiency revealed two heteroplasmic mitochondrial DNA mutations in ND genes. One patient had the mitochondrial DNA T8993G point mutation. No mitochondrial DNA defects were identified in 11 (68.7%) of our LS patients, who probably have mutations in nuclear DNA. Although a limited study based in a single tertiary medical center, our findings suggest that isolated complex I deficiency may be the most common cause of Leigh syndrome in Korea.

Congenital hypopituitarism due to POU1F1 gene mutation.

PubMed

Lee, Ni-Chung; Tsai, Wen-Yu; Peng, Shinn-Forng; Tung, Yi-Ching; Chien, Yin-Hsiu; Hwu, Wuh-Liang

2011-01-01

POU1F1 (Pit-1; Gene ID 5449) is an anterior pituitary transcriptional factor, and POU1F1 mutation is known to cause anterior pituitary hypoplasia, growth hormone and prolactin deficiency and various degree of hypothyroidism. We report here a patient who presented with growth failure and central hypothyroidism since early infancy. However, treatment with thyroxine gave no effect and he subsequently developed calf muscle pseudohypertrophy (Kocher-Debre-Semelaigne syndrome), elevation of creatinine kinase, dilated cardiomyopathy and pericardial effusion. Final diagnosis was made by combined pituitary function test and sequencing analysis that revealed POU1F1 gene C.698T > C (p.F233S) mutation. The rarity of the disease can result in delayed diagnosis and treatment. Copyright © 2011 Formosan Medical Association & Elsevier. Published by Elsevier B.V. All rights reserved.

Identification of Mutations in SLC24A4, Encoding a Potassium-Dependent Sodium/Calcium Exchanger, as a Cause of Amelogenesis Imperfecta

PubMed Central

Parry, David A.; Poulter, James A.; Logan, Clare V.; Brookes, Steven J.; Jafri, Hussain; Ferguson, Christopher H.; Anwari, Babra M.; Rashid, Yasmin; Zhao, Haiqing; Johnson, Colin A.; Inglehearn, Chris F.; Mighell, Alan J.

2013-01-01

A combination of autozygosity mapping and exome sequencing identified a null mutation in SLC24A4 in a family with hypomineralized amelogenesis imperfect a (AI), a condition in which tooth enamel formation fails. SLC24A4 encodes a calcium transporter upregulated in ameloblasts during the maturation stage of amelogenesis. Screening of further AI families identified a missense mutation in the ion-binding site of SLC24A4 expected to severely diminish or abolish the ion transport function of the protein. Furthermore, examination of previously generated Slc24a4 null mice identified a severe defect in tooth enamel that reflects impaired amelogenesis. These findings support a key role for SLC24A4 in calcium transport during enamel formation. PMID:23375655

Novel nonsense mutation of the endothelin-B receptor gene in a family with Waardenburg-Hirschsprung disease.

PubMed

Syrris, P; Carter, N D; Patton, M A

1999-11-05

Waardenburg syndrome (WS) comprises sensorineural hearing loss, hypopigmentation of skin and hair, and pigmentary disturbances of the irides. Four types of WS have been classified to date; in WS type IV (WS4), patients additionally have colonic aganglionosis (Hirschsprung disease, HSCR). Mutations in the endothelin-3 (EDN3), endothelin-B receptor (EDNRB), and Sox10 genes have been identified as causative for WS type IV. We screened a family with a combined WS-HSCR phenotype for mutations in the EDNRB locus using standard DNA mutation analysis and sequencing techniques. We have identified a novel nonsense mutation at codon 253 (CGA-->TGA, Arg-->STOP). This mutation leads to a premature end of the translation of EDNRB at exon 3, and it is predicted to produce a truncated and nonfunctional endothelin-B receptor. All affected relatives were heterozygous for the Arg(253)-->STOP mutation, whereas it was not observed in over 50 unrelated individuals used as controls. These data confirm the role of EDNRB in the cause of the Waardenburg-Hirschsprung syndrome and demonstrate that in WS-HSCR there is a lack of correlation between phenotype and genotype and a variable expression of disease even within the same family. Copyright 1999 Wiley-Liss, Inc.

High-resolution melting analysis for detection of MYH9 mutations.

PubMed

Provaznikova, Dana; Kumstyrova, Tereza; Kotlin, Roman; Salaj, Peter; Matoska, Vaclav; Hrachovinova, Ingrid; Rittich, Simon

2008-09-01

May-Hegglin anomaly (MHA), Sebastian (SBS), Fechtner (FTNS) and Epstein (EPS) syndromes are rare autosomal dominant disorders with giant platelets and thrombocytopenia. Other manifestations of these disorders are combinations of the presence of granulocyte inclusions and deafness, cataracts and renal failure. Currently, MHA, SBS, FTNS and EPS are considered to be distinct clinical manifestation of a single illness caused by mutations of the MYH9 gene encoding the heavy chain of non-muscle myosin IIA (NMMHC-IIA). As the MYH9 gene has a high number of exons, it takes much time and material to use this method for the detection of MYH9 mutations. Recently, a new method has been introduced for scanning DNA mutations without the need for direct sequencing: high-resolution melting analysis (HRMA). Mutation detection with HRMA relies on the intercalation of the specific dye (LC Green plus) in double-strand DNA and fluorescence monitoring of PCR product melting profiles. In our study, we optimized the conditions and used HRMA for rapid screening of mutations in all MYH9 exons in seven affected individuals from four unrelated families with suspected MYH9 disorders. Samples identified by HRMA as positive for the mutation were analysed by direct sequencing. HRMA saved us over 85% of redundant sequencing.

Adaptive Mutations in Influenza A/California/07/2009 Enhance Polymerase Activity and Infectious Virion Production.

PubMed

Slaine, Patrick D; MacRae, Cara; Kleer, Mariel; Lamoureux, Emily; McAlpine, Sarah; Warhuus, Michelle; Comeau, André M; McCormick, Craig; Hatchette, Todd; Khaperskyy, Denys A

2018-05-18

Mice are not natural hosts for influenza A viruses (IAVs), but they are useful models for studying antiviral immune responses and pathogenesis. Serial passage of IAV in mice invariably causes the emergence of adaptive mutations and increased virulence. Here, we report the adaptation of IAV reference strain A/California/07/2009(H1N1) (also known as CA/07) in outbred Swiss Webster mice. Serial passage led to increased virulence and lung titers, and dissemination of the virus to brains. We adapted a deep-sequencing protocol to identify and enumerate adaptive mutations across all genome segments. Among mutations that emerged during mouse-adaptation, we focused on amino acid substitutions in polymerase subunits: polymerase basic-1 (PB1) T156A and F740L and polymerase acidic (PA) E349G. These mutations were evaluated singly and in combination in minigenome replicon assays, which revealed that PA E349G increased polymerase activity. By selectively engineering three PB1 and PA mutations into the parental CA/07 strain, we demonstrated that these mutations in polymerase subunits decreased the production of defective viral genome segments with internal deletions and dramatically increased the release of infectious virions from mouse cells. Together, these findings increase our understanding of the contribution of polymerase subunits to successful host adaptation.

Utility of genetic testing for the detection of late-onset hearing loss in neonates.

PubMed

Lim, B Gail; Clark, Reese H; Kelleher, Amy S; Lin, Zhili; Spitzer, Alan R

2013-12-01

The purpose of this study was to demonstrate the utility of molecular testing in the detection of potentially important causes of delayed hearing loss missed by current audiometric screening at birth. We enrolled infants who had received a newborn audiometric hearing screen and a filter paper blood collection for state newborn screening. A central laboratory ran the SoundGene® panel. Of 3,681 infants studied, 35 (0.95%) had a positive SoundGene panel, 16 had mitochondrial mutations, 9 had Pendred mutations, 5 were cytomegalovirus (CMV) DNA positive, 2 had connexin mutations, and 3 had a combination of different mutations. Infants with an abnormal SoundGene panel were at increased risk for hearing loss compared to neonates without mutations. Three (8.6%) of the 35 subjects had persistent hearing loss compared to 5 (0.21%) of 2,398 subjects with no report of mutation (p < .01). Of 3,681 infants studied, 8 (0.22%) had persistent hearing loss: 5 (62.5%) had abnormal newborn audiometric screens, 2 (25%) had an abnormal SoundGene panel (1 was CMV positive, 1 had a mitochondrial mutation), and 1 (12.5%) had no identifiable risk factors. A positive SoundGene panel identifies infants who are not identified by audiometric testing and may be at risk for hearing loss.

Neutral lipid-storage disease with myopathy and extended phenotype with novel PNPLA2 mutation.

PubMed

Massa, Roberto; Pozzessere, Simone; Rastelli, Emanuele; Serra, Laura; Terracciano, Chiara; Gibellini, Manuela; Bozzali, Marco; Arca, Marcello

2016-04-01

Neutral lipid-storage disease with myopathy is caused by mutations in PNPLA2, which produce skeletal and cardiac myopathy. We report a man with multiorgan neutral lipid storage and unusual multisystem clinical involvement, including cognitive impairment. Quantitative brain MRI with voxel-based morphometry and extended neuropsychological assessment were performed. In parallel, the coding sequences and intron/exon boundaries of the PNPLA2 gene were screened by direct sequencing. Neuropsychological assessment revealed global cognitive impairment, and brain MRI showed reduced gray matter volume in the temporal lobes. Molecular characterization revealed a novel homozygous mutation in exon 5 of PNPLA2 (c.714C>A), resulting in a premature stop codon (p.Cys238*). Some PNPLA2 mutations, such as the one described here, may present with an extended phenotype, including brain involvement. In these cases, complete neuropsychological testing, combined with quantitative brain MRI, may help to characterize and quantify cognitive impairment. © 2016 Wiley Periodicals, Inc.

A mutation in canine CLN5 causes neuronal ceroid lipofuscinosis in Border collie dogs.

PubMed

Melville, Scott A; Wilson, Carmen L; Chiang, Chiu S; Studdert, Virginia P; Lingaas, Frode; Wilton, Alan N

2005-09-01

Neuronal ceroid lipofuscinosis (NCL) is a neurodegenerative disease found in Border collie dogs, humans, and other animals. Disease gene studies in humans and animals provided candidates for the NCL gene in Border collies. A combination of linkage analysis and comparative genomics localized the gene to CFA22 in an area syntenic to HSA13q that contains the CLN5 gene responsible for the Finnish variant of human late infantile NCL. Sequencing of CLN5 revealed a nonsense mutation (Q206X) within exon 4 that correlated with NCL in Border collies. This truncation mutation should result in a protein product of a size similar to that of some mutations identified in human CLN5 and therefore the Border collie may make a good model for human NCL. A simple test was developed to enable screening of the Border collie population for carriers so the disease can be eliminated as a problem in the breed.

Cp/Heph mutant mice have iron-induced neurodegeneration diminished by deferiprone

PubMed Central

Zhao, Liangliang; Hadziahmetovic, Majda; Wang, Chenguang; Xu, Xueying; Song, Ying; Jinnah, H.A.; Wodzinska, Jolanta; Iacovelli, Jared; Wolkow, Natalie; Krajacic, Predrag; Weissberger, Alyssa Cwanger; Connelly, John; Spino, Michael; Lee, Michael K.; Connor, James; Giasson, Benoit; Harris, Z. Leah; Dunaief, Joshua L.

2016-01-01

Brain iron accumulates in several neurodegenerative diseases and can cause oxidative damage, but mechanisms of brain iron homeostasis are incompletely understood. Patients with mutations in the cellular iron-exporting ferroxidase ceruloplasmin (Cp) have brain iron accumulation causing neurodegeneration. Here, we assessed the brains of mice with combined mutation of Cp and its homolog hephaestin. Compared to single mutants, brain iron accumulation was accelerated in double mutants in the cerebellum, substantia nigra, and hippocampus. Iron accumulated within glia, while neurons were iron deficient. There was loss of both neurons and glia. Mice developed ataxia and tremor, and most died by 9 months. Treatment with the oral iron chelator deferiprone diminished brain iron levels, protected against neuron loss, and extended lifespan. Ferroxidases play important, partially overlapping roles in brain iron homeostasis by facilitating iron export from glia, making iron available to neurons. PMID:26303407

8-oxoguanine causes spontaneous de novo germline mutations in mice.

PubMed

Ohno, Mizuki; Sakumi, Kunihiko; Fukumura, Ryutaro; Furuichi, Masato; Iwasaki, Yuki; Hokama, Masaaki; Ikemura, Toshimichi; Tsuzuki, Teruhisa; Gondo, Yoichi; Nakabeppu, Yusaku

2014-04-15

Spontaneous germline mutations generate genetic diversity in populations of sexually reproductive organisms, and are thus regarded as a driving force of evolution. However, the cause and mechanism remain unclear. 8-oxoguanine (8-oxoG) is a candidate molecule that causes germline mutations, because it makes DNA more prone to mutation and is constantly generated by reactive oxygen species in vivo. We show here that endogenous 8-oxoG caused de novo spontaneous and heritable G to T mutations in mice, which occurred at different stages in the germ cell lineage and were distributed throughout the chromosomes. Using exome analyses covering 40.9 Mb of mouse transcribed regions, we found increased frequencies of G to T mutations at a rate of 2 × 10(-7) mutations/base/generation in offspring of Mth1/Ogg1/Mutyh triple knockout (TOY-KO) mice, which accumulate 8-oxoG in the nuclear DNA of gonadal cells. The roles of MTH1, OGG1, and MUTYH are specific for the prevention of 8-oxoG-induced mutation, and 99% of the mutations observed in TOY-KO mice were G to T transversions caused by 8-oxoG; therefore, we concluded that 8-oxoG is a causative molecule for spontaneous and inheritable mutations of the germ lineage cells.

Fasting boosts sensitivity of human skin melanoma to cisplatin-induced cell death

DOE Office of Scientific and Technical Information (OSTI.GOV)

Antunes, Fernanda; Corazzari, Marco; National Institute for Infectious Diseases IRCCS “Lazzaro Spallanzani”

Melanoma is one of leading cause of tumor death worldwide. Anti-cancer strategy includes combination of different chemo-therapeutic agents as well as radiation; however these treatments have limited efficacy and induce significant toxic effects on healthy cells. One of most promising novel therapeutic approach to cancer therapy is the combination of anti-cancer drugs with calorie restriction. Here we investigated the effect Cisplatin (CDDP), one of the most potent chemotherapeutic agent used to treat tumors, in association with fasting in wild type and mutated BRAF{sup V600E} melanoma cell lines. Here we show that nutrient deprivation can consistently enhance the sensitivity of tumormore » cells to cell death induction by CDDP, also of those malignancies particularly resistant to any treatment, such as oncogenic BRAF melanomas. Mechanistic studies revealed that the combined therapy induced cell death is characterized by ROS accumulation and ATF4 in the absence of ER-stress. In addition, we show that autophagy is not involved in the enhanced sensitivity of melanoma cells to combined CDDP/EBSS-induced apoptosis. While, the exposure to 2-DG further enhanced the apoptotic rate observed in SK Mel 28 cells upon treatment with both CDDP and EBSS. - Highlights: • Calorie restriction associated to chemo-therapeutic drugs enhance cell death induction in many resistant malignancies • Cisplatin in association with starvation significantly increases cell death also in those high resistant melanoma cells bearing BRAF mutations • Combined treatment also including 2-DG results in similar cell death levels in both wild type and mutated BRAF cells.« less

Treacher Collins syndrome: clinical implications for the paediatrician--a new mutation in a severely affected newborn and comparison with three further patients with the same mutation, and review of the literature.

PubMed

Schlump, Jan-Ulrich; Stein, Anja; Hehr, Ute; Karen, Tanja; Möller-Hartmann, Claudia; Elcioglu, Nursel H; Bogdanova, Nadja; Woike, Hartmut Fritz; Lohmann, Dietmar R; Felderhoff-Mueser, Ursula; Linz, Annette; Wieczorek, Dagmar

2012-11-01

Treacher Collins syndrome (TCS) is the most common and well-known mandibulofacial dysostosis caused by mutations in at least three genes involved in pre-rRNA transcription, the TCOF1, POLR1D and POLR1C genes. We present a severely affected male individual with TCS with a heterozygous de novo frameshift mutation within the TCOF1 gene (c.790_791delAG,p.Ser264GlnfsX7) and compare the clinical findings with three previously unpublished, milder affected individuals from two families with the same mutation. We elucidate typical clinical features of TCS and its clinical implications for the paediatrician and mandibulofacial surgeon, especially in severely affected individuals and give a short review of the literature. The clinical data of these three families illustrate that the phenotype associated with this specific mutation has a wide intra- and interfamilial variability, which confirms that variable expressivity in carriers of TCOF1 mutations is not a simple consequence of the mutation but might be modified by the combination of genetic, environmental and stochastic factors. Being such a highly complex disease treatment of individuals with TCS should be tailored to the specific needs of each individual, preferably by a multidisciplinary team consisting of paediatricians, craniofacial surgeons and geneticists.

C. elegans whole-genome sequencing reveals mutational signatures related to carcinogens and DNA repair deficiency.

PubMed

Meier, Bettina; Cooke, Susanna L; Weiss, Joerg; Bailly, Aymeric P; Alexandrov, Ludmil B; Marshall, John; Raine, Keiran; Maddison, Mark; Anderson, Elizabeth; Stratton, Michael R; Gartner, Anton; Campbell, Peter J

2014-10-01

Mutation is associated with developmental and hereditary disorders, aging, and cancer. While we understand some mutational processes operative in human disease, most remain mysterious. We used Caenorhabditis elegans whole-genome sequencing to model mutational signatures, analyzing 183 worm populations across 17 DNA repair-deficient backgrounds propagated for 20 generations or exposed to carcinogens. The baseline mutation rate in C. elegans was approximately one per genome per generation, not overtly altered across several DNA repair deficiencies over 20 generations. Telomere erosion led to complex chromosomal rearrangements initiated by breakage-fusion-bridge cycles and completed by simultaneously acquired, localized clusters of breakpoints. Aflatoxin B1 induced substitutions of guanines in a GpC context, as observed in aflatoxin-induced liver cancers. Mutational burden increased with impaired nucleotide excision repair. Cisplatin and mechlorethamine, DNA crosslinking agents, caused dose- and genotype-dependent signatures among indels, substitutions, and rearrangements. Strikingly, both agents induced clustered rearrangements resembling "chromoanasynthesis," a replication-based mutational signature seen in constitutional genomic disorders, suggesting that interstrand crosslinks may play a pathogenic role in such events. Cisplatin mutagenicity was most pronounced in xpf-1 mutants, suggesting that this gene critically protects cells against platinum chemotherapy. Thus, experimental model systems combined with genome sequencing can recapture and mechanistically explain mutational signatures associated with human disease. © 2014 Meier et al.; Published by Cold Spring Harbor Laboratory Press.

Identification and Analysis of Mot3, a Zinc Finger Protein That Binds to the Retrotransposon Ty Long Terminal Repeat (δ) in Saccharomyces cerevisiae

PubMed Central

Madison, Jon M.; Dudley, Aimée M.; Winston, Fred

1998-01-01

Spt3 and Mot1 are two transcription factors of Saccharomyces cerevisiae that are thought to act in a related fashion to control the function of TATA-binding protein (TBP). Current models suggest that while Spt3 and Mot1 do not directly interact, they do function in a related fashion to stabilize the TBP-TATA interaction at particular promoters. Consistent with this model, certain combinations of spt3 and mot1 mutations are inviable. To identify additional proteins related to Spt3 and Mot1 functions, we screened for high-copy-number suppressors of the mot1 spt3 inviability. This screen identified a previously unstudied gene, MOT3, that encodes a zinc finger protein. We show that Mot3 binds in vitro to three sites within the retrotransposon Ty long terminal repeat (δ) sequence. One of these sites is immediately 5′ of the δ TATA region. Although a mot3 null mutation causes no strong phenotypes, it does cause some mild phenotypes, including a very modest increase in Ty mRNA levels, partial suppression of transcriptional defects caused by a mot1 mutation, and partial suppression of an spt3 mutation. These results, in conjunction with those of an independent study of Mot3 (A. Grishin, M. Rothenberg, M. A. Downs, and K. J. Blumer, Genetics, in press), suggest that this protein plays a varied role in gene expression that may be largely redundant with other factors. PMID:9528759

Bolivian kindred with combined spinocerebellar ataxia types 2 and 10.

PubMed

Baizabal-Carvallo, J F; Xia, G; Botros, P; Laguna, J; Ashizawa, T; Jankovic, J

2015-08-01

Spinocerebellar ataxias (SCA) are a group of rare hereditary neurodegenerative disorders. Rare cases of two SCA mutations in the same individual have been reported in the literature, however, family descriptions are lacking. To characterize a family with combined SCA2 and SCA10 mutations. Analysis of the clinical features and genetic findings of a Bolivian family expressing both SCA2 and SCA10 mutations. The index case and his mother had both SCA2 and SCA10 mutations with a combined clinical phenotype of both disorders, including slow saccades (SCA2) and seizures (SCA10). The uncle of the index case had only an SCA10 mutation. Although the presence of two SCA mutations in the same individuals may be coincidental, the low probability of having both mutations suggests that these mutations might be particularly prevalent in Bolivian population. This is the first description of a family with two SCA mutations with affected subjects having a combined SCA2 and SCA10 phenotype. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Target gene analyses of 39 amelogenesis imperfecta kindreds

PubMed Central

Chan, Hui-Chen; Estrella, Ninna M. R. P.; Milkovich, Rachel N.; Kim, Jung-Wook; Simmer, James P.; Hu, Jan C-C.

2012-01-01

Previously, mutational analyses identified six disease-causing mutations in 24 amelogenesis imperfecta (AI) kindreds. We have since expanded the number of AI kindreds to 39, and performed mutation analyses covering the coding exons and adjoining intron sequences for the six proven AI candidate genes [amelogenin (AMELX), enamelin (ENAM), family with sequence similarity 83, member H (FAM83H), WD repeat containing domain 72 (WDR72), enamelysin (MMP20), and kallikrein-related peptidase 4 (KLK4)] and for ameloblastin (AMBN) (a suspected candidate gene). All four of the X-linked AI families (100%) had disease-causing mutations in AMELX, suggesting that AMELX is the only gene involved in the aetiology of X-linked AI. Eighteen families showed an autosomal-dominant pattern of inheritance. Disease-causing mutations were identified in 12 (67%): eight in FAM83H, and four in ENAM. No FAM83H coding-region or splice-junction mutations were identified in three probands with autosomal-dominant hypocalcification AI (ADHCAI), suggesting that a second gene may contribute to the aetiology of ADHCAI. Six families showed an autosomal-recessive pattern of inheritance, and disease-causing mutations were identified in three (50%): two in MMP20, and one in WDR72. No disease-causing mutations were found in 11 families with only one affected member. We conclude that mutation analyses of the current candidate genes for AI have about a 50% chance of identifying the disease-causing mutation in a given kindred. PMID:22243262

Sarcomere protein gene mutations and inherited heart disease: a beta-cardiac myosin heavy chain mutation causing endocardial fibroelastosis and heart failure.

PubMed

Kamisago, Mitsuhiro; Schmitt, Joachim P; McNamara, Dennis; Seidman, Christine; Seidman, J G

2006-01-01

Inherited human cardiomyopathies often lead to heart failure. A common feature of these conditions is that affected individuals can express the disease causing mutations for many years without showing clinical signs of the disease. Previous studies have demonstrated that sarcomere protein gene mutations can cause either dilated cardiomyopathy or hypertrophic cardiomyopathy. Here we demonstrate that the Arg442His missense mutation in beta-cardiac myosin heavy chain (betaMHC) causes dilated cardiomyopathy, endocardial fibroelastosis and heart failure at a very early age. Using standard genetic engineering tools we and others have made murine models by introducing human disease causing mutations into mice. The central hypothesis of these studies has been that by identifying the pathophysiological pathways activated by these mutations we can define enzymatic activities that are modified during the disease process and which may be involved in pathways that involve more common forms of cardiac disease. Murine models bearing different mutant myosins are being used to address whether each disease causing mutant betaMHC activates the same or different cellular pathways. Dissecting the molecular pathways modulated by mutations in sarcomere protein genes as well as other genes has already demonstrated that there are multiple pathways leading to cardiac remodelling and heart failure. Defining the mechanisms by which mutations in the same genes activate different cellular pathways remains an important question.

A Japanese family with nonautoimmune hyperthyroidism caused by a novel heterozygous thyrotropin receptor gene mutation.

PubMed

Nakamura, Akie; Morikawa, Shuntaro; Aoyagi, Hayato; Ishizu, Katsura; Tajima, Toshihiro

2014-06-01

Hyperthyroidism caused by activating mutations of the thyrotropin receptor gene (TSHR) is rare in the pediatric population. We found a Japanese family with hyperthyroidism without autoantibody. DNA sequence analysis of TSHR was undertaken in this family. The functional consequences for the Gs-adenylyl cyclase and Gq/11-phospholipase C signaling pathways and cell surface expression of receptors were determined in vitro using transiently transfected human embryonic kidney 293 cells. We identified a heterozygous mutation (M453R) in exon 10 of TSHR. In this family, this mutation was found in all individuals who exhibited hyperthyroidism. The results showed that this mutation resulted in constitutive activation of the Gs-adenylyl cyclase system. However, this mutation also caused a reduction in the activation capacity of the Gq/11-phospholipase C pathway, compared with the wild type. We demonstrate that the M453R mutation is the cause of nonautoimmune hyperthyroidism.

Analysis of anti-malarial resistance markers in pfmdr1 and pfcrt across Southeast Asia in the Tracking Resistance to Artemisinin Collaboration.

PubMed

Srimuang, Krongkan; Miotto, Olivo; Lim, Pharath; Fairhurst, Rick M; Kwiatkowski, Dominic P; Woodrow, Charles J; Imwong, Mallika

2016-11-08

Declining anti-malarial efficacy of artemisinin-based combination therapy, and reduced Plasmodium falciparum susceptibility to individual anti-malarials are being documented across an expanding area of Southeast Asia (SEA). Genotypic markers complement phenotypic studies in assessing the efficacy of individual anti-malarials. The markers pfmdr1 and pfcrt were genotyped in parasite samples obtained in 2011-2014 at 14 TRAC (Tracking Resistance to Artemisinin Collaboration) sites in mainland Southeast Asia using a combination of PCR and next-generation sequencing methods. Pfmdr1 amplification, a marker of mefloquine and lumefantrine resistance, was highly prevalent at Mae Sot on the Thailand-Myanmar border (59.8% of isolates) and common (more than 10%) at sites in central Myanmar, eastern Thailand and western Cambodia; however, its prevalence was lower than previously documented in Pailin, western Cambodia. The pfmdr1 Y184F mutation was common, particularly in and around Cambodia, and the F1226Y mutation was found in about half of samples in Mae Sot. The functional significance of these two mutations remains unclear. Other previously documented pfmdr1 mutations were absent or very rare in the region. The pfcrt mutation K76T associated with chloroquine resistance was found in 98.2% of isolates. The CVIET haplotype made up 95% or more of isolates in western SEA while the CVIDT haplotype was common (30-40% of isolates) in north and northeastern Cambodia, southern Laos, and southern Vietnam. These findings generate cause for concern regarding the mid-term efficacy of artemether-lumefantrine in Myanmar, while the absence of resistance-conferring pfmdr1 mutations and SVMNT pfcrt haplotypes suggests that amodiaquine could be an efficacious component of anti-malarial regimens in SEA.

Identification of a novel BRCA2 and CHEK2 A-C-G-C haplotype in Turkish patients affected with breast cancer.

PubMed

Haytural, Hazal; Yalcinkaya, Nazli; Akan, Gokce; Arikan, Soykan; Ozkok, Elif; Cakmakoglu, Bedia; Yaylim, Ilhan; Aydin, Makbule; Atalar, Fatmahan

2013-01-01

Many breast cancers are caused by certain rare and familial mutations in the high or moderate penetrance genes BRCA1, BRCA2 and CHEK2. The aim of this study was to examine the allele and genotype frequencies of seven mutations in BRCA1, BRCA2 and CHEK2 genes in breast cancer patients and to investigate their isolated and combined associations with breast cancer risk. We genotyped seven mutations in BRCA1, BRCA2 and CHEK2 genes and then analyzed single variations and haplotype associations in 106 breast cancer patients and 80 healthy controls. We found significant associations in the analyses of CHEK2- 1100delC (p=0.001) and BRCA1-5382insC (p=0.021) mutations in breast cancer patients compared to controls. The highest risk was observed among breast cancer patients carrying both CHEK2-1100delC and BRCA2- Met784Val mutations (OR=0.093; 95%CI 0.021-0.423; p=0.001). We identified one previously undescribed BRCA2 and a CHEK2 four-marker haplotype of A-C-G-C which was overrepresented (?2=7.655; p=0.0057) in the patient group compared to controls. In this study, we identified a previously undescribed BRCA2 and CHEK2 A-C-G-C haplotype in association with the breast cancer in our population. Our results further suggest that the CHEK2-1100delC mutation in combination with BRCA2-Met784Val may lead to an unexpected high risk which needs to be confirmed in larger cohorts in order to better understand their role in the development and prognosis of breast cancer.

Rare Frequency of Mutations in Pituitary Transcription Factor Genes in Combined Pituitary Hormone or Isolated Growth Hormone Deficiencies in Korea.

PubMed

Choi, Jin Ho; Jung, Chang Woo; Kang, Eungu; Kim, Yoon Myung; Heo, Sun Hee; Lee, Beom Hee; Kim, Gu Hwan; Yoo, Han Wook

2017-05-01

Congenital hypopituitarism is caused by mutations in pituitary transcription factors involved in the development of the hypothalamic-pituitary axis. Mutation frequencies of genes involved in congenital hypopituitarism are extremely low and vary substantially between ethnicities. This study was undertaken to compare the clinical, endocrinological, and radiological features of patients with an isolated growth hormone deficiency (IGHD) or combined pituitary hormone deficiency (CPHD). This study included 27 patients with sporadic IGHD and CPHD. A mutation analysis of the POU1F1, PROP1, LHX3, LHX4, and HESX1 genes was performed using genomic DNA from peripheral blood leukocytes. IGHD and CPHD were observed in 4 and 23 patients, respectively. Mean age at diagnosis was 8.28±7.25 years for IGHD and 13.48±10.46 years for CPHD (p=0.37). Serum insulin-like growth factor-1 and peak growth hormone (GH) levels following GH stimulation tests were significantly lower in patients with CPHD than in those with IGHD (p<0.05). Sellar MRI findings revealed structural abnormalities in 3 patients with IGHD (75%) and 21 patients with CPHD (91.3%) (p=0.62). A mutation analysis identified homozygous p.R109Q mutations in HESX1 in a patient with CPHD. Patients with CPHD had more severe GHD than those with IGHD. The frequency of defects in the genes encoding pituitary transcription factors was extremely low in Korean patients with congenital hypopituitarism. Environmental factors and the impact of other causative genes may contribute to this clinical phenotype. © Copyright: Yonsei University College of Medicine 2017

Rare Frequency of Mutations in Pituitary Transcription Factor Genes in Combined Pituitary Hormone or Isolated Growth Hormone Deficiencies in Korea

PubMed Central

Choi, Jin-Ho; Jung, Chang-Woo; Kang, Eungu; Kim, Yoon-Myung; Heo, Sun Hee; Lee, Beom Hee; Kim, Gu-Hwan

2017-01-01

Purpose Congenital hypopituitarism is caused by mutations in pituitary transcription factors involved in the development of the hypothalamic-pituitary axis. Mutation frequencies of genes involved in congenital hypopituitarism are extremely low and vary substantially between ethnicities. This study was undertaken to compare the clinical, endocrinological, and radiological features of patients with an isolated growth hormone deficiency (IGHD) or combined pituitary hormone deficiency (CPHD). Materials and Methods This study included 27 patients with sporadic IGHD and CPHD. A mutation analysis of the POU1F1, PROP1, LHX3, LHX4, and HESX1 genes was performed using genomic DNA from peripheral blood leukocytes. Results IGHD and CPHD were observed in 4 and 23 patients, respectively. Mean age at diagnosis was 8.28±7.25 years for IGHD and 13.48±10.46 years for CPHD (p=0.37). Serum insulin-like growth factor-1 and peak growth hormone (GH) levels following GH stimulation tests were significantly lower in patients with CPHD than in those with IGHD (p<0.05). Sellar MRI findings revealed structural abnormalities in 3 patients with IGHD (75%) and 21 patients with CPHD (91.3%) (p=0.62). A mutation analysis identified homozygous p.R109Q mutations in HESX1 in a patient with CPHD. Patients with CPHD had more severe GHD than those with IGHD. Conclusion The frequency of defects in the genes encoding pituitary transcription factors was extremely low in Korean patients with congenital hypopituitarism. Environmental factors and the impact of other causative genes may contribute to this clinical phenotype. PMID:28332357

The CDC Hemophilia B mutation project mutation list: a new online resource.

PubMed

Li, Tengguo; Miller, Connie H; Payne, Amanda B; Craig Hooper, W

2013-11-01

Hemophilia B (HB) is caused by mutations in the human gene F9. The mutation type plays a pivotal role in genetic counseling and prediction of inhibitor development. To help the HB community understand the molecular etiology of HB, we have developed a listing of all F9 mutations that are reported to cause HB based on the literature and existing databases. The Centers for Disease Control and Prevention (CDC) Hemophilia B Mutation Project (CHBMP) mutation list is compiled in an easily accessible format of Microsoft Excel and contains 1083 unique mutations that are reported to cause HB. Each mutation is identified using Human Genome Variation Society (HGVS) nomenclature standards. The mutation types and the predicted changes in amino acids, if applicable, are also provided. Related information including the location of mutation, severity of HB, the presence of inhibitor, and original publication reference are listed as well. Therefore, our mutation list provides an easily accessible resource for genetic counselors and HB researchers to predict inhibitors. The CHBMP mutation list is freely accessible at http://www.cdc.gov/hemophiliamutations.

KRAS and the Reality of Personalized Medicine in Non-Small Cell Lung Cancer

PubMed Central

Kilgoz, Havva O; Bender, Guzide; Scandura, Joseph M; Viale, Agnes; Taneri, Bahar

2016-01-01

Lung cancer is the leading cause of mortality among all cancer types worldwide. The latest available global statistics of the World Health Organization report 1.59 million casualities in 2012. Worldwide, 1 in 5 cancer deaths are caused by lung cancer. In 2016, in the United States alone, there are an estimated 224,390 new cases of lung cancer, of which 158,080 are expected to result in death, as reported by the National Cancer Institute. Non-small cell lung cancer (NSCLC), a histological subtype, comprises about 85% of all cases, which is nearly 9 out of 10 lung cancer patients. Efforts are under way to develop and improve targeted therapy strategies. Certain mutations are being clinically targeted, such as those in EGFR and ALK genes. However, one of the most frequently mutated genes in NSCLC is the Kirsten rat sarcoma viral oncogene homolog (KRAS), which is currently not targetable. Approximately 25% of all types of NSCLC tumors contain KRAS mutations, which remain as an undruggable challenge. These mutations are indicative of poor prognosis and show negative response to standard chemotherapy. Furthermore, tumors harboring KRAS mutations are unlikely to respond to currently available targeted treatments such as tyrosine kinase inhibitors. Therefore, there is a definitive, urgent need to generate new targeted therapy approaches for KRAS mutations. Current strategies have major limitations and revolve around targeting molecules upstream and downstream of KRAS. Direct targeting is not available in the clinic. Combination therapies using multiple agents are being sought. Concentrated efforts are needed to accelerate basic research and consecutive clinical trials to achieve effective targeting of KRAS. PMID:27447490

Novel USH2A compound heterozygous mutations cause RP/USH2 in a Chinese family.

PubMed

Liu, Xiaowen; Tang, Zhaohui; Li, Chang; Yang, Kangjuan; Gan, Guanqi; Zhang, Zibo; Liu, Jingyu; Jiang, Fagang; Wang, Qing; Liu, Mugen

2010-03-17

To identify the disease-causing gene in a four-generation Chinese family affected with retinitis pigmentosa (RP). Linkage analysis was performed with a panel of microsatellite markers flanking the candidate genetic loci of RP. These loci included 38 known RP genes. The complete coding region and exon-intron boundaries of Usher syndrome 2A (USH2A) were sequenced with the proband DNA to screen the disease-causing gene mutation. Restriction fragment length polymorphism (RFLP) analysis and direct DNA sequence analysis were done to demonstrate co-segregation of the USH2A mutations with the family disease. One hundred normal controls were used without the mutations. The disease-causing gene in this Chinese family was linked to the USH2A locus on chromosome 1q41. Direct DNA sequence analysis of USH2A identified two novel mutations in the patients: one missense mutation p.G1734R in exon 26 and a splice site mutation, IVS32+1G>A, which was found in the donor site of intron 32 of USH2A. Neither the p.G1734R nor the IVS32+1G>A mutation was found in the unaffected family members or the 100 normal controls. One patient with a homozygous mutation displayed only RP symptoms until now, while three patients with compound heterozygous mutations in the family of study showed both RP and hearing impairment. This study identified two novel mutations: p.G1734R and IVS32+1G>A of USH2A in a four-generation Chinese RP family. In this study, the heterozygous mutation and the homozygous mutation in USH2A may cause Usher syndrome Type II or RP, respectively. These two mutations expand the mutant spectrum of USH2A.

De novo mutations in ATP1A3 cause alternating hemiplegia of childhood

PubMed Central

Heinzen, Erin L.; Swoboda, Kathryn J.; Hitomi, Yuki; Gurrieri, Fiorella; Nicole, Sophie; de Vries, Boukje; Tiziano, F. Danilo; Fontaine, Bertrand; Walley, Nicole M.; Heavin, Sinéad; Panagiotakaki, Eleni; Fiori, Stefania; Abiusi, Emanuela; Di Pietro, Lorena; Sweney, Matthew T.; Newcomb, Tara M.; Viollet, Louis; Huff, Chad; Jorde, Lynn B.; Reyna, Sandra P.; Murphy, Kelley J.; Shianna, Kevin V.; Gumbs, Curtis E.; Little, Latasha; Silver, Kenneth; Ptác̆ek, Louis J.; Haan, Joost; Ferrari, Michel D.; Bye, Ann M.; Herkes, Geoffrey K.; Whitelaw, Charlotte M.; Webb, David; Lynch, Bryan J.; Uldall, Peter; King, Mary D.; Scheffer, Ingrid E.; Neri, Giovanni; Arzimanoglou, Alexis; van den Maagdenberg, Arn M.J.M.; Sisodiya, Sanjay M.; Mikati, Mohamad A.; Goldstein, David B.; Nicole, Sophie; Gurrieri, Fiorella; Neri, Giovanni; de Vries, Boukje; Koelewijn, Stephany; Kamphorst, Jessica; Geilenkirchen, Marije; Pelzer, Nadine; Laan, Laura; Haan, Joost; Ferrari, Michel; van den Maagdenberg, Arn; Zucca, Claudio; Bassi, Maria Teresa; Franchini, Filippo; Vavassori, Rosaria; Giannotta, Melania; Gobbi, Giuseppe; Granata, Tiziana; Nardocci, Nardo; De Grandis, Elisa; Veneselli, Edvige; Stagnaro, Michela; Gurrieri, Fiorella; Neri, Giovanni; Vigevano, Federico; Panagiotakaki, Eleni; Oechsler, Claudia; Arzimanoglou, Alexis; Nicole, Sophie; Giannotta, Melania; Gobbi, Giuseppe; Ninan, Miriam; Neville, Brian; Ebinger, Friedrich; Fons, Carmen; Campistol, Jaume; Kemlink, David; Nevsimalova, Sona; Laan, Laura; Peeters-Scholte, Cacha; van den Maagdenberg, Arn; Casaer, Paul; Casari, Giorgio; Sange, Guenter; Spiel, Georg; Boneschi, Filippo Martinelli; Zucca, Claudio; Bassi, Maria Teresa; Schyns, Tsveta; Crawley, Francis; Poncelin, Dominique; Vavassori, Rosaria

2012-01-01

Alternating hemiplegia of childhood (AHC) is a rare, severe neurodevelopmental syndrome characterized by recurrent hemiplegic episodes and distinct neurologic manifestations. AHC is usually a sporadic disorder with unknown etiology. Using exome sequencing of seven patients with AHC, and their unaffected parents, we identified de novo nonsynonymous mutations in ATP1A3 in all seven AHC patients. Subsequent sequence analysis of ATP1A3 in 98 additional patients revealed that 78% of AHC cases have a likely causal ATP1A3 mutation, including one inherited mutation in a familial case of AHC. Remarkably, six ATP1A3 mutations explain the majority of patients, including one observed in 36 patients. Unlike ATP1A3 mutations that cause rapid-onset-dystonia-parkinsonism, AHC-causing mutations revealed consistent reductions in ATPase activity without effects on protein expression. This work identifies de novo ATP1A3 mutations as the primary cause of AHC, and offers insight into disease pathophysiology by expanding the spectrum of phenotypes associated with mutations in this gene. PMID:22842232

Nitrative and oxidative DNA damage caused by K-ras mutation in mice

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ohnishi, Shiho; Saito, Hiromitsu; Suzuki, Noboru

2011-09-23

Highlights: {yields} Mutated K-ras in transgenic mice caused nitrative DNA damage, 8-nitroguanine. {yields} The mutagenic 8-nitroguanine seemed to be generated by iNOS via Ras-MAPK signal. {yields} Mutated K-ras produces additional mutagenic lesions, as a new oncogenic role. -- Abstract: Ras mutation is important for carcinogenesis. Carcinogenesis consists of multi-step process with mutations in several genes. We investigated the role of DNA damage in carcinogenesis initiated by K-ras mutation, using conditional transgenic mice. Immunohistochemical analysis revealed that mutagenic 8-nitroguanine and 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG) were apparently formed in adenocarcinoma caused by mutated K-ras. 8-Nitroguanine was co-localized with iNOS, eNOS, NF-{kappa}B, IKK, MAPK, MEK,more » and mutated K-ras, suggesting that oncogenic K-ras causes additional DNA damage via signaling pathway involving these molecules. It is noteworthy that K-ras mutation mediates not only cell over-proliferation but also the accumulation of mutagenic DNA lesions, leading to carcinogenesis.« less

The MEK1/2 Inhibitor AZD6244 Sensitizes BRAF-Mutant Thyroid Cancer to Vemurafenib.

PubMed

Song, Hao; Zhang, Jinna; Ning, Liang; Zhang, Honglai; Chen, Dong; Jiao, Xuelong; Zhang, Kejun

2018-05-08

BACKGROUND [i]BRAF[/i]V600E mutation occurs in approximately 45% of papillary thyroid cancer (PTC) cases, and 25% of anaplastic thyroid cancer (ATC) cases. Vemurafenib/PLX4032, a selective BRAF inhibitor, suppresses extracellular signal-regulated kinase kinase/extracellular signal-regulated kinase 1/2 (MEK/ERK1/2) signaling and shows beneficial effects in patients with metastatic melanoma harboring the [i]BRAFV600E[/i] mutation. However, the response to vemurafenib is limited in BRAF-mutant thyroid cancer. The present study evaluated the effect of vemurafenib in combination with the selective MEK1/2 inhibitor AZD6244 on cell survival and explored the mechanism underlying the combined effect of vemurafenib and AZD6244 on thyroid cancer cells harboring BRAFV600E. MATERIAL AND METHODS Thyroid cancer 8505C and BCPAP cells harboring the [i]BRAFV600E[/i] mutation were exposed to vemurafenib (0.01, 0.1, and 1 µM) and AZD6244 (0.01, 0.1, and 1 µM) alone or in the indicated combinations for the indicated times. Cell viability was detected by the MTT assay. Cell cycle distribution and induction of apoptosis were detected by flow cytometry. The expression of cyclin D1, P27, (P)-ERK1/2 was evaluated by Western blotting. The effect of vemurafenib or AZD6244 or their combination on the growth of 8505C cells was examined in orthotopic xenograft mouse models [i]in vivo[/i]. RESULTS Vemurafenib alone did not increase cell apoptosis, whereas it decreased cell viability by promoting cell cycle arrest in BCPAP and 8505C cells. AZD6244 alone increased cell apoptosis by inducing cell cycle arrest in BCPAP and 8505C cells. Combination treatment with AZD6244 and vemurafenib significantly decreased cell viability and increased apoptosis in both BCPAP and 8505C cells compared with the effects of each drug alone. AZD6244 alone abolished phospho-ERK1/2 (pERK1/2) expression at 48 h, whereas vemurafenib alone downregulated pERK1/2 at 4-6 h, with rapid recovery of expression, reaching the highest level at 24-48 h. Combined treatment for 48 h completely inhibited pERK1/2 expression. Combination treatment with vemurafenib and AZD6244 inhibited cell growth and induced apoptosis by causing cell-cycle arrest, with the corresponding changes in the expression of the cell cycle regulators p27Kip1 and cyclin D1. Co-administration of vemurafenib and AZD6244 [i]in vivo[/i] had a significant synergistic antitumor effect in a nude mouse model. CONCLUSIONS Vemurafenib activated pERK1/2 and induced vemurafenib resistance in thyroid cancer cells. Combination treatment with vemurafenib and AZD6244 inhibited ERK signaling and caused cell cycle arrest, resulting in cell growth inhibition. Combination treatment in patients with thyroid cancer harboring the [i]BRAFV600E[/i] mutation may overcome vemurafenib resistance and enhance the therapeutic effect.

Important Mutations Contributing to High-Level Penicillin Resistance in Taiwan19F-14, Taiwan23F-15, and Spain23F-1 of Streptococcus pneumoniae Isolated from Taiwan.

PubMed

Liu, Esther Yip-Mei; Chang, Jen-Chang; Lin, Jung-Chung; Chang, Feng-Yee; Fung, Chang-Phone

2016-12-01

Penicillin-resistant Streptococcus pneumoniae is a serious concern worldwide. In this study, we analyzed the cause of β-lactam resistance in pandemic multidrug-resistant clones. A total of 41 penicillin-nonsusceptible clinical isolates were collected from 1996 to 2012. Sero- and molecular typing confirmed that these isolates were clonal types of Taiwan 19F -14, Taiwan 23F -15, and Spain 23F -1. Sero-switching was found in four isolates. All isolates were multidrug resistant. Sequencing analysis of the penicillin binding proteins (PBPs) was performed on PBP1a, 2b, and 2x, and a large number of mutations were identified in comparing to clinical penicillin-susceptible isolates and the recipient strain R6 used for homologous recombination. The T 451 A substitution was the key amino acid in PBP2b that contributed to penicillin resistance. T 338 A in PBP2x played a role in resistance and reached the highest level of resistance when combined with other mutations in PBP2x. High-level penicillin resistance could not be obtained without the combination of mutations in PBP1a with PBP2b and 2x. The amino acid substitutions in PBP1a, 2b, and 2x were the crucial factors for β-lactam resistance.

Genetic heterogeneity of pseudoxanthoma elasticum: the Chinese signature profile of ABCC6 and ENPP1 mutations.

PubMed

Jin, Liang; Jiang, Qiujie; Wu, Zhengsheng; Shao, Changxia; Zhou, Yong; Yang, Luting; Uitto, Jouni; Wang, Gang

2015-05-01

Pseudoxanthoma elasticum (PXE), an autosomal recessive disorder characterized by ectopic mineralization, is caused by mutations in the ABCC6 gene. We examined clinically 29 Chinese PXE patients from unrelated families, so far the largest cohort of Asian PXE patients. In a subset of 22 patients, we sequenced ABCC6 and another candidate gene, ENPP1, and conducted pathogenicity analyses for each variant. We identified a total of 17 distinct mutations in ABCC6, 15 of them being, to our knowledge, previously unreported, including 5 frameshift and 10 missense variants. In addition, a missense mutation in combination with a recurrent nonsense mutation in ENPP1 was discovered in a pediatric PXE case. No cases with p.R1141X or del23-29 mutations, common in Caucasian patient populations, were identified. The 10 missense mutations in ABCC6 were expressed in the mouse liver via hydrodynamic tail-vein injections. One mutant protein showed cytoplasmic accumulation indicating abnormal subcellular trafficking, while the other nine mutants showed correct plasma membrane location. These nine mutations were further investigated for their pathogenicity using a recently developed zebrafish mRNA rescue assay. Minimal rescue of the morpholino-induced phenotype was achieved with eight of the nine mutant human ABCC6 mRNAs tested, implying pathogenicity. This study demonstrates that the Chinese PXE population harbors unique ABCC6 mutations. These genetic data have implications for allele-specific therapy currently being developed for PXE.

Reality of Single Circulating Tumor Cell Sequencing for Molecular Diagnostics in Pancreatic Cancer.

PubMed

Court, Colin M; Ankeny, Jacob S; Sho, Shonan; Hou, Shuang; Li, Qingyu; Hsieh, Carolyn; Song, Min; Liao, Xinfang; Rochefort, Matthew M; Wainberg, Zev A; Graeber, Thomas G; Tseng, Hsian-Rong; Tomlinson, James S

2016-09-01

To understand the potential and limitations of circulating tumor cell (CTC) sequencing for molecular diagnostics, we investigated the feasibility of identifying the ubiquitous KRAS mutation in single CTCs from pancreatic cancer (PC) patients. We used the NanoVelcro/laser capture microdissection CTC platform, combined with whole genome amplification and KRAS Sanger sequencing. We assessed both KRAS codon-12 coverage and the degree that allele dropout during whole genome amplification affected the detection of KRAS mutations from single CTCs. We isolated 385 single cells, 163 from PC cell lines and 222 from the blood of 12 PC patients, and obtained KRAS sequence coverage in 218 of 385 single cells (56.6%). For PC cell lines with known KRAS mutations, single mutations were detected in 67% of homozygous cells but only 37.4% of heterozygous single cells, demonstrating that both coverage and allele dropout are important causes of mutation detection failure from single cells. We could detect KRAS mutations in CTCs from 11 of 12 patients (92%) and 33 of 119 single CTCs sequenced, resulting in a KRAS mutation detection rate of 27.7%. Importantly, KRAS mutations were never found in the 103 white blood cells sequenced. Sequencing of groups of cells containing between 1 and 100 cells determined that at least 10 CTCs are likely required to reliably assess KRAS mutation status from CTCs. Copyright © 2016 American Society for Investigative Pathology and the Association for Molecular Pathology. Published by Elsevier Inc. All rights reserved.

Pathogenetic role of the deafness-related M34T mutation of Cx26

PubMed Central

Bicego, Massimiliano; Beltramello, Martina; Melchionda, Salvatore; Carella, Massimo; Piazza, Valeria; Zelante, Leopoldo; Bukauskas, Feliksas F.; Arslan, Edoardo; Cama, Elona; Pantano, Sergio; Bruzzone, Roberto; D’Andrea, Paola; Mammano, Fabio

2010-01-01

Mutations in the GJB2 gene, which encodes the gap junction protein connexin26 (Cx26), are the major cause of genetic non-syndromic hearing loss. The role of the allelic variant M34T in causing hereditary deafness remains controversial. By combining genetic, clinical, biochemical, electrophysiological and structural modeling studies, we have re-assessed the pathogenetic role of the M34T mutation. Genetic and audiological data indicate that the majority of heterozygous carriers and all five compound heterozygotes exhibited an impaired auditory function. Functional expression in transiently transfected HeLa cells showed that, although M34T was correctly synthesized and targeted to the plasma membrane, it inefficiently formed intercellular channels that displayed an abnormal electrical behavior and retained only 11% of the unitary conductance of the wild-type protein (HCx26wt). Moreover, M34T channels failed to support the intercellular diffusion of Lucifer Yellow and the spreading of mechanically induced intercellular Ca2+ waves. When co-expressed together with HCx26wt, M34T exerted dominant-negative effects on cell–cell coupling. Our findings are consistent with a structural model, predicting that the mutation leads to a constriction of the channel pore. These data support the view that M34T is a pathological variant of Cx26 associated with hearing impairment. PMID:16849369

Dominant mutations in KAT6A cause intellectual disability with recognizable syndromic features.

PubMed

Tham, Emma; Lindstrand, Anna; Santani, Avni; Malmgren, Helena; Nesbitt, Addie; Dubbs, Holly A; Zackai, Elaine H; Parker, Michael J; Millan, Francisca; Rosenbaum, Kenneth; Wilson, Golder N; Nordgren, Ann

2015-03-05

Through a multi-center collaboration study, we here report six individuals from five unrelated families, with mutations in KAT6A/MOZ detected by whole-exome sequencing. All five different de novo heterozygous truncating mutations were located in the C-terminal transactivation domain of KAT6A: NM_001099412.1: c.3116_3117 delCT, p.(Ser1039∗); c.3830_3831insTT, p.(Arg1278Serfs∗17); c.3879 dupA, p.(Glu1294Argfs∗19); c.4108G>T p.(Glu1370∗) and c.4292 dupT, p.(Leu1431Phefs∗8). An additional subject with a 0.23 MB microdeletion including the entire KAT6A reading frame was identified with genome-wide array comparative genomic hybridization. Finally, by detailed clinical characterization we provide evidence that heterozygous mutations in KAT6A cause a distinct intellectual disability syndrome. The common phenotype includes hypotonia, intellectual disability, early feeding and oromotor difficulties, microcephaly and/or craniosynostosis, and cardiac defects in combination with subtle facial features such as bitemporal narrowing, broad nasal tip, thin upper lip, posteriorly rotated or low-set ears, and microretrognathia. The identification of human subjects complements previous work from mice and zebrafish where knockouts of Kat6a/kat6a lead to developmental defects. Copyright © 2015 The American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.

Exome Sequencing Identifies a Novel CEACAM16 Mutation Associated with Autosomal Dominant Nonsyndromic Hearing Loss DFNA4B in a Chinese Family

PubMed Central

He, Chufeng; Li, Haibo; Qing, Jie; Grati, Mhamed; Hu, Zhengmao; Li, Jiada; Hu, Yiqiao; Xia, Kun; Mei, Lingyun; Wang, Xingwei; Yu, Jianjun; Chen, Hongsheng; Jiang, Lu; Liu, Yalan; Men, Meichao; Zhang, Hailin; Guan, Liping; Xiao, Jingjing; Zhang, Jianguo; Liu, Xuezhong; Feng, Yong

2014-01-01

Autosomal dominant nonsyndromic hearing loss (ADNSHL/DFNA) is a highly genetically heterogeneous disorder. Hitherto only about 30 ADNSHL-causing genes have been identified and many unknown genes remain to be discovered. In this research, genome-wide linkage analysis mapped the disease locus to a 4.3 Mb region on chromosome 19q13 in SY-026, a five-generation nonconsanguineous Chinese family affected by late-onset and progressive ADNSHL. This linkage region showed partial overlap with the previously reported DFNA4. Simultaneously, probands were analyzed using exome capture followed by next generation sequencing. Encouragingly, a heterozygous missense mutation, c.505G>A (p.G169R) in exon 3 of the CEACAM16 gene (carcinoembryonic antigen-related cell adhesion molecule 16), was identified via this combined strategy. Sanger sequencing verified that the mutation co-segregated with hearing loss in the family and that it was not present in 200 unrelated control subjects with matched ancestry. This is the second report in the literature of a family with ADNSHL caused by CEACAM16 mutation. Immunofluorescence staining and Western blots also prove CEACAM16 to be a secreted protein. Furthermore, our studies in transfected HEK293T cells show that the secretion efficacy of the mutant CEACAM16 is much lower than that of the wild-type, suggesting a deleterious effect of the sequence variant. PMID:25589040

Exome sequencing identifies a novel CEACAM16 mutation associated with autosomal dominant nonsyndromic hearing loss DFNA4B in a Chinese family.

PubMed

Wang, Honghan; Wang, Xinwei; He, Chufeng; Li, Haibo; Qing, Jie; Grati, Mhamed; Hu, Zhengmao; Li, Jiada; Hu, Yiqiao; Xia, Kun; Mei, Lingyun; Wang, Xingwei; Yu, Jianjun; Chen, Hongsheng; Jiang, Lu; Liu, Yalan; Men, Meichao; Zhang, Hailin; Guan, Liping; Xiao, Jingjing; Zhang, Jianguo; Liu, Xuezhong; Feng, Yong

2015-03-01

Autosomal dominant nonsyndromic hearing loss (ADNSHL/DFNA) is a highly genetically heterogeneous disorder. Hitherto only about 30 ADNSHL-causing genes have been identified and many unknown genes remain to be discovered. In this research, genome-wide linkage analysis mapped the disease locus to a 4.3 Mb region on chromosome 19q13 in SY-026, a five-generation nonconsanguineous Chinese family affected by late-onset and progressive ADNSHL. This linkage region showed partial overlap with the previously reported DFNA4. Simultaneously, probands were analyzed using exome capture followed by next-generation sequencing. Encouragingly, a heterozygous missense mutation, c.505G>A (p.G169R) in exon 3 of the CEACAM16 gene (carcinoembryonic antigen-related cell adhesion molecule 16), was identified via this combined strategy. Sanger sequencing verified that the mutation co-segregated with hearing loss in the family and that it was not present in 200 unrelated control subjects with matched ancestry. This is the second report in the literature of a family with ADNSHL caused by CEACAM16 mutation. Immunofluorescence staining and western blots also prove CEACAM16 to be a secreted protein. Furthermore, our studies in transfected HEK293T cells show that the secretion efficacy of the mutant CEACAM16 is much lower than that of the wild type, suggesting a deleterious effect of the sequence variant.

Molecular Genetics of the Usher Syndrome in Lebanon: Identification of 11 Novel Protein Truncating Mutations by Whole Exome Sequencing

PubMed Central

Reddy, Ramesh; Fahiminiya, Somayyeh; El Zir, Elie; Mansour, Ahmad; Megarbane, Andre; Majewski, Jacek; Slim, Rima

2014-01-01

Background Usher syndrome (USH) is a genetically heterogeneous condition with ten disease-causing genes. The spectrum of genes and mutations causing USH in the Lebanese and Middle Eastern populations has not been described. Consequently, diagnostic approaches designed to screen for previously reported mutations were unlikely to identify the mutations in 11 unrelated families, eight of Lebanese and three of Middle Eastern origins. In addition, six of the ten USH genes consist of more than 20 exons, each, which made mutational analysis by Sanger sequencing of PCR-amplified exons from genomic DNA tedious and costly. The study was aimed at the identification of USH causing genes and mutations in 11 unrelated families with USH type I or II. Methods Whole exome sequencing followed by expanded familial validation by Sanger sequencing. Results We identified disease-causing mutations in all the analyzed patients in four USH genes, MYO7A, USH2A, GPR98 and CDH23. Eleven of the mutations were novel and protein truncating, including a complex rearrangement in GPR98. Conclusion Our data highlight the genetic diversity of Usher syndrome in the Lebanese population and the time and cost-effectiveness of whole exome sequencing approach for mutation analysis of genetically heterogeneous conditions caused by large genes. PMID:25211151

A screen to identify Drosophila genes required for integrin-mediated adhesion.

PubMed Central

Walsh, E P; Brown, N H

1998-01-01

Drosophila integrins have essential adhesive roles during development, including adhesion between the two wing surfaces. Most position-specific integrin mutations cause lethality, and clones of homozygous mutant cells in the wing do not adhere to the apposing surface, causing blisters. We have used FLP-FRT induced mitotic recombination to generate clones of randomly induced mutations in the F1 generation and screened for mutations that cause wing blisters. This phenotype is highly selective, since only 14 lethal complementation groups were identified in screens of the five major chromosome arms. Of the loci identified, 3 are PS integrin genes, 2 are blistered and bloated, and the remaining 9 appear to be newly characterized loci. All 11 nonintegrin loci are required on both sides of the wing, in contrast to integrin alpha subunit genes. Mutations in 8 loci only disrupt adhesion in the wing, similar to integrin mutations, while mutations in the 3 other loci cause additional wing defects. Mutations in 4 loci, like the strongest integrin mutations, cause a "tail-up" embryonic lethal phenotype, and mutant alleles of 1 of these loci strongly enhance an integrin mutation. Thus several of these loci are good candidates for genes encoding cytoplasmic proteins required for integrin function. PMID:9755209

Molecular genetics of the Usher syndrome in Lebanon: identification of 11 novel protein truncating mutations by whole exome sequencing.

PubMed

Reddy, Ramesh; Fahiminiya, Somayyeh; El Zir, Elie; Mansour, Ahmad; Megarbane, Andre; Majewski, Jacek; Slim, Rima

2014-01-01

Usher syndrome (USH) is a genetically heterogeneous condition with ten disease-causing genes. The spectrum of genes and mutations causing USH in the Lebanese and Middle Eastern populations has not been described. Consequently, diagnostic approaches designed to screen for previously reported mutations were unlikely to identify the mutations in 11 unrelated families, eight of Lebanese and three of Middle Eastern origins. In addition, six of the ten USH genes consist of more than 20 exons, each, which made mutational analysis by Sanger sequencing of PCR-amplified exons from genomic DNA tedious and costly. The study was aimed at the identification of USH causing genes and mutations in 11 unrelated families with USH type I or II. Whole exome sequencing followed by expanded familial validation by Sanger sequencing. We identified disease-causing mutations in all the analyzed patients in four USH genes, MYO7A, USH2A, GPR98 and CDH23. Eleven of the mutations were novel and protein truncating, including a complex rearrangement in GPR98. Our data highlight the genetic diversity of Usher syndrome in the Lebanese population and the time and cost-effectiveness of whole exome sequencing approach for mutation analysis of genetically heterogeneous conditions caused by large genes.

Background sequence characteristics influence the occurrence and severity of disease-causing mtDNA mutations

PubMed Central

Wei, Wei; Hudson, Gavin

2017-01-01

Inherited mitochondrial DNA (mtDNA) mutations have emerged as a common cause of human disease, with mutations occurring multiple times in the world population. The clinical presentation of three pathogenic mtDNA mutations is strongly associated with a background mtDNA haplogroup, but it is not clear whether this is limited to a handful of examples or is a more general phenomenon. To address this, we determined the characteristics of 30,506 mtDNA sequences sampled globally. After performing several quality control steps, we ascribed an established pathogenicity score to the major alleles for each sequence. The mean pathogenicity score for known disease-causing mutations was significantly different between mtDNA macro-haplogroups. Several mutations were observed across all haplogroup backgrounds, whereas others were only observed on specific clades. In some instances this reflected a founder effect, but in others, the mutation recurred but only within the same phylogenetic cluster. Sequence diversity estimates showed that disease-causing mutations were more frequent on young sequences, and genomes with two or more disease-causing mutations were more common than expected by chance. These findings implicate the mtDNA background more generally in recurrent mutation events that have been purified through natural selection in older populations. This provides an explanation for the low frequency of mtDNA disease reported in specific ethnic groups. PMID:29253894

Cellular Reprogramming, Genome Editing, and Alternative CRISPR Cas9 Technologies for Precise Gene Therapy of Duchenne Muscular Dystrophy

PubMed Central

Xu, Huaigeng

2017-01-01

In the past decade, the development of two innovative technologies, namely, induced pluripotent stem cells (iPSCs) and the CRISPR Cas9 system, has enabled researchers to model diseases derived from patient cells and precisely edit DNA sequences of interest, respectively. In particular, Duchenne muscular dystrophy (DMD) has been an exemplary monogenic disease model for combining these technologies to demonstrate that genome editing can correct genetic mutations in DMD patient-derived iPSCs. DMD is an X-linked genetic disorder caused by mutations that disrupt the open reading frame of the dystrophin gene, which plays a critical role in stabilizing muscle cells during contraction and relaxation. The CRISPR Cas9 system has been shown to be capable of targeting the dystrophin gene and rescuing its expression in in vitro patient-derived iPSCs and in vivo DMD mouse models. In this review, we highlight recent advances made using the CRISPR Cas9 system to correct genetic mutations and discuss how emerging CRISPR technologies and iPSCs in a combined platform can play a role in bringing a therapy for DMD closer to the clinic. PMID:28607562

KMeyeDB: a graphical database of mutations in genes that cause eye diseases.

PubMed

Kawamura, Takashi; Ohtsubo, Masafumi; Mitsuyama, Susumu; Ohno-Nakamura, Saho; Shimizu, Nobuyoshi; Minoshima, Shinsei

2010-06-01

KMeyeDB (http://mutview.dmb.med.keio.ac.jp/) is a database of human gene mutations that cause eye diseases. We have substantially enriched the amount of data in the database, which now contains information about the mutations of 167 human genes causing eye-related diseases including retinitis pigmentosa, cone-rod dystrophy, night blindness, Oguchi disease, Stargardt disease, macular degeneration, Leber congenital amaurosis, corneal dystrophy, cataract, glaucoma, retinoblastoma, Bardet-Biedl syndrome, and Usher syndrome. KMeyeDB is operated using the database software MutationView, which deals with various characters of mutations, gene structure, protein functional domains, and polymerase chain reaction (PCR) primers, as well as clinical data for each case. Users can access the database using an ordinary Internet browser with smooth user-interface, without user registration. The results are displayed on the graphical windows together with statistical calculations. All mutations and associated data have been collected from published articles. Careful data analysis with KMeyeDB revealed many interesting features regarding the mutations in 167 genes that cause 326 different types of eye diseases. Some genes are involved in multiple types of eye diseases, whereas several eye diseases are caused by different mutations in one gene.

Genetic testing in steroid-resistant nephrotic syndrome: why, who, when and how?

PubMed

Preston, Rebecca; Stuart, Helen M; Lennon, Rachel

2017-11-27

Steroid-resistant nephrotic syndrome (SRNS) is a common cause of chronic kidney disease in childhood and has a significant risk of rapid progression to end-stage renal disease. The identification of over 50 monogenic causes of SRNS has revealed dysfunction in podocyte-associated proteins in the pathogenesis of proteinuria, highlighting their essential role in glomerular function. Recent technological advances in high-throughput sequencing have enabled indication-driven genetic panel testing for patients with SRNS. The availability of genetic testing, combined with the significant phenotypic variability of monogenic SRNS, poses unique challenges for clinicians when directing genetic testing. This highlights the need for clear clinical guidelines that provide a systematic approach for mutational screening in SRNS. The likelihood of identifying a causative mutation is inversely related to age at disease onset and is increased with a positive family history or the presence of extra-renal manifestations. An unequivocal molecular diagnosis could allow for a personalised treatment approach with weaning of immunosuppressive therapy, avoidance of renal biopsy and provision of accurate, well-informed genetic counselling. Identification of novel causative mutations will continue to unravel the pathogenic mechanisms of glomerular disease and provide new insights into podocyte biology and glomerular function.

Ectodermal Dysplasia: A Genetic Review

PubMed Central

Prashanth, S

2012-01-01

Abstract Ectodermal dysplasia is a rare hereditary disorder with a characteristic physiognomy. It is a genetic disorder affecting the development or function of the teeth, hair, nails and sweat glands. Depending on the particular syndrome ectodermal dysplasia can also affect the skin, the lens or retina of the eye, parts of the inner ear, the development of fingers and toes, the nerves and other parts of the body. Each syndrome usually involves a different combination of symptoms, which can range from mild to severe. The history and lessons learned from hypohidrotic ectodermal dysplasia (HED) may serve as an example for unraveling of the cause and pathogenesis of other ectodermal dysplasia syndromes by demonstrating that phenotypically identical syndromes can be caused by mutations in different genes (EDA, EDAR, EDARADD), that mutations in the same gene can lead to different phenotypes and that mutations in the genes further downstream in the same signaling pathway (NEMO) may modify the phenotype quite profoundly. The aim of this paper is to describe and discuss the etiology, genetic review, clinical manifestations and treatment options of this hereditary disorder. How to cite this article: Deshmukh S, Prashanth S. Ectodermal Dysplasia: A Genetic Review. Int J Clin Pediatr Dent 2012; 5(3):197-202. PMID:25206167

Ectodermal dysplasia: a genetic review.

PubMed

Deshmukh, Seema; Prashanth, S

2012-09-01

Ectodermal dysplasia is a rare hereditary disorder with a characteristic physiognomy. It is a genetic disorder affecting the development or function of the teeth, hair, nails and sweat glands. Depending on the particular syndrome ectodermal dysplasia can also affect the skin, the lens or retina of the eye, parts of the inner ear, the development of fingers and toes, the nerves and other parts of the body. Each syndrome usually involves a different combination of symptoms, which can range from mild to severe. The history and lessons learned from hypohidrotic ectodermal dysplasia (HED) may serve as an example for unraveling of the cause and pathogenesis of other ectodermal dysplasia syndromes by demonstrating that phenotypically identical syndromes can be caused by mutations in different genes (EDA, EDAR, EDARADD), that mutations in the same gene can lead to different phenotypes and that mutations in the genes further downstream in the same signaling pathway (NEMO) may modify the phenotype quite profoundly. The aim of this paper is to describe and discuss the etiology, genetic review, clinical manifestations and treatment options of this hereditary disorder. How to cite this article: Deshmukh S, Prashanth S. Ectodermal Dysplasia: A Genetic Review. Int J Clin Pediatr Dent 2012; 5(3):197-202.

Phelan-McDermid syndrome due to SHANK3 mutation in an intellectually disabled adult male: successful treatment with lithium.

PubMed

Egger, Jos I M; Verhoeven, Willem M A; Groenendijk-Reijenga, Renske; Kant, Sarina G

2017-09-28

For 30 years, Phelan and co-workers described a syndrome characterised by neonatal hypotonia, global developmental delay, strongly impaired speech, sleep disturbances and hyperreactivity to sensory stimuli. This Phelan-McDermid syndrome (PMS), also presenting with symptoms from the autism spectrum and a higher risk of developing seizure disorders, may be caused by a deletion of chromosome 22q13 or by a mutation in the SHANK3 gene. Its core psychopathological phenotype comprises symptoms from the bipolar spectrum for which generally treatment with a mood-stabilising anticonvulsant in combination with an atypical antipsychotic seems to be most effective. In addition to two elsewhere published adolescent patients, we here describe in detail the history of an adult male patient with PMS caused by a SHANK3 mutation in whom successive treatment regimens with antipsychotics and mood-stabilising anticonvulsants were all ineffective. Ultimately, addition of lithium to existing olanzapine therapy led to enduring stabilisation of mood and behaviour. © BMJ Publishing Group Ltd (unless otherwise stated in the text of the article) 2017. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

Current insights into the molecular genetic basis of dwarfism in livestock.

PubMed

Boegheim, Iris J M; Leegwater, Peter A J; van Lith, Hein A; Back, Willem

2017-06-01

Impairment of bone growth at a young age leads to dwarfism in adulthood. Dwarfism can be categorised as either proportionate, an overall size reduction without changes in body proportions, or disproportionate, a size reduction in one or more limbs, with changes in body proportions. Many forms of dwarfism are inherited and result from structural disruptions or disrupted signalling pathways. Hormonal disruptions are evident in Brooksville miniature Brahman cattle and Z-linked dwarfism in chickens, caused by mutations in GH1 and GHR. Furthermore, mutations in IHH are the underlying cause of creeper achondroplasia in chickens. Belgian blue cattle display proportionate dwarfism caused by a mutation in RNF11, while American Angus cattle dwarfism is caused by a mutation in PRKG2. Mutations in EVC2 are associated with dwarfism in Japanese brown cattle and Tyrolean grey cattle. Fleckvieh dwarfism is caused by mutations in the GON4L gene. Mutations in COL10A1 and COL2A1 cause dwarfism in pigs and Holstein cattle, both associated with structural disruptions, while several mutations in ACAN are associated with bulldog-type dwarfism in Dexter cattle and dwarfism in American miniature horses. In other equine breeds, such as Shetland ponies and Friesian horses, dwarfism is caused by mutations in SHOX and B4GALT7. In Texel sheep, chondrodysplasia is associated with a deletion in SLC13A1. This review discusses genes known to be involved in these and other forms of dwarfism in livestock. Copyright © 2017 Elsevier Ltd. All rights reserved.

Positive Selection during the Evolution of the Blood Coagulation Factors in the Context of Their Disease-Causing Mutations

PubMed Central

Rallapalli, Pavithra M.; Orengo, Christine A.; Studer, Romain A.; Perkins, Stephen J.

2014-01-01

Blood coagulation occurs through a cascade of enzymes and cofactors that produces a fibrin clot, while otherwise maintaining hemostasis. The 11 human coagulation factors (FG, FII–FXIII) have been identified across all vertebrates, suggesting that they emerged with the first vertebrates around 500 Ma. Human FVIII, FIX, and FXI are associated with thousands of disease-causing mutations. Here, we evaluated the strength of selective pressures on the 14 genes coding for the 11 factors during vertebrate evolution, and compared these with human mutations in FVIII, FIX, and FXI. Positive selection was identified for fibrinogen (FG), FIII, FVIII, FIX, and FX in the mammalian Primates and Laurasiatheria and the Sauropsida (reptiles and birds). This showed that the coagulation system in vertebrates was under strong selective pressures, perhaps to adapt against blood-invading pathogens. The comparison of these results with disease-causing mutations reported in FVIII, FIX, and FXI showed that the number of disease-causing mutations, and the probability of positive selection were inversely related to each other. It was concluded that when a site was under positive selection, it was less likely to be associated with disease-causing mutations. In contrast, sites under negative selection were more likely to be associated with disease-causing mutations and be destabilizing. A residue-by-residue comparison of the FVIII, FIX, and FXI sequence alignments confirmed this. This improved understanding of evolutionary changes in FVIII, FIX, and FXI provided greater insight into disease-causing mutations, and better assessments of the codon sites that may be mutated in applications of gene therapy. PMID:25158795

Coding Sequence Mutations Identified in MYH7, TNNT2, SCN5A, CSRP3, LBD3, and TCAP from 313 Patients with Familial or Idiopathic Dilated Cardiomyopathy

PubMed Central

Hershberger, Ray E.; Parks, Sharie B.; Kushner, Jessica D.; Li, Duanxiang; Ludwigsen, Susan; Jakobs, Petra; Nauman, Deirdre; Burgess, Donna; Partain, Julie; Litt, Michael

2008-01-01

Abstract Background: More than 20 genes have been reported to cause idiopathic and familial dilated cardiomyopathy (IDC/FDC), but the frequency of genetic causation remains poorly understood. Methods and Results: Blood samples were collected and DNA prepared from 313 patients, 183 with FDC and 130 with IDC. Genomic DNA underwent bidirectional sequencing of six genes, and mutation carriers were followed up by evaluation of additional family members. We identified in 36 probands, 31 unique protein‐altering variants (11.5% overall) that were not identified in 253 control subjects (506 chromosomes). These included 13 probands (4.2%) with 12 β‐myosin heavy chain (MYH7) mutations, nine probands (2.9%) with six different cardiac troponin T (TNNT2) mutations, eight probands (2.6%) carrying seven different cardiac sodium channel (SCN5A) mutations, three probands (1.0%) with three titin‐cap or telethonin (TCAP) mutations, three probands (1.0%) with two LIM domain binding 3 (LDB3) mutations, and one proband (0.3%) with a muscle LIM protein (CSRP3) mutation. Four nucleotide changes did not segregate with phentoype and/or did not alter a conserved amino acid and were therefore considered unlikely to be disease‐causing. Mutations in 11 probands were assessed as likely disease‐causing, and in 21 probands were considered possibly disease‐causing. These 32 probands included 14 of the 130 with IDC (10.8%) and 18 of the 183 with FDC (9.8%) Conclusions: Mutations of these six genes each account for a small fraction of the genetic cause of FDC/IDC. The frequency of possible or likely disease‐causing mutations in these genes is similar for IDC and FDC. PMID:19412328

Rhythm Defects Caused by Newly Engineered Null Mutations in Drosophila's cryptochrome Gene

PubMed Central

Dolezelova, Eva; Dolezel, David; Hall, Jeffrey C.

2007-01-01

Much of the knowledge about cryptochrome function in Drosophila stems from analyzing the cryb mutant. Several features of this variant's light responsiveness imply either that CRYb retains circadian-photoreceptive capacities or that additional CRY-independent light-input routes subserve these processes. Potentially to resolve these issues, we generated cry knock-out mutants (cry0's) by gene replacement. They behaved in an anomalously rhythmic manner in constant light (LL). However, cry0 flies frequently exhibited two separate circadian components in LL, not observed in most previous cryb analyses. Temperature-dependent circadian phenotypes exhibited by cry0 flies suggest that CRY is involved in core pacemaking. Further locomotor experiments combined cry0 with an externally blinding mutation (norpAP24), which caused the most severe decrements of circadian photoreception observed so far. cryb cultures were shown previously to exhibit either aperiodic or rhythmic eclosion in separate studies. We found cry0 to eclose in a solidly periodic manner in light:dark cycles or constant darkness. Furthermore, both cry0 and cryb eclosed rhythmically in LL. These findings indicate that the novel cry0 type causes more profound defects than does the cryb mutation, implying that CRYb retains residual activity. Because some norpAP24 cry0 individuals can resynchronize to novel photic regimes, an as-yet undetermined light-input route exists in Drosophila. PMID:17720919

Advancing epilepsy treatment through personalized genetic zebrafish models.

PubMed

Griffin, A; Krasniak, C; Baraban, S C

2016-01-01

With an increase in the number of disease causing genetic mutations identified from epilepsy cohorts, zebrafish are proving to be an attractive vertebrate model for functional analysis of these allele variants. Not only do zebrafish have conserved gene functions, but larvae harboring mutations in identified human epileptic genes show spontaneous seizure activity and mimic the convulsive behavioral movements observed in humans. With zebrafish being compatible with medium to high-throughput screening, they are also proving to be a unique and powerful system for early preclinical drug screening, including novel target identification, pharmacology, and toxicology. Additionally, with recent advances in genomic engineering technologies, it is now possible to study the precise pathophysiology of patient-specific gene mutations in zebrafish. The following sections highlight how the unique attributes of zebrafish, in combination with genetic modifications, are continuing to transform our understanding of epilepsy and help identify personalized therapeutics for specific patient cohorts. © 2016 Elsevier B.V. All rights reserved.

Structural study of the effects of mutations in proteins to identify the molecular basis of the loss of local structural fluidity leading to the onset of autoimmune diseases

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ali, Ananya; Ghosh, Semanti; Bagchi, Angshuman

Protein-Protein Interactions (PPIs) are crucial in most of the biological processes and PPI dysfunctions are known to be associated with the onsets of various diseases. One of such diseases is the auto-immune disease. Auto-immune diseases are one among the less studied group of diseases with very high mortality rates. Thus, we tried to correlate the appearances of mutations with their probable biochemical basis of the molecular mechanisms leading to the onset of the disease phenotypes. We compared the effects of the Single Amino Acid Variants (SAVs) in the wild type and mutated proteins to identify any structural deformities that mightmore » lead to altered PPIs leading ultimately to disease onset. For this we used Relative Solvent Accessibility (RSA) as a spatial parameter to compare the structural perturbation in mutated and wild type proteins. We observed that the mutations were capable to increase intra-chain PPIs whereas inter-chain PPIs would remain mostly unaltered. This might lead to more intra-molecular friction causing a deleterious alteration of protein's normal function. A Lyapunov exponent analysis, using the altered RSA values due to polymorphic and disease causing mutations, revealed polymorphic mutations have a positive mean value for the Lyapunov exponent while disease causing mutations have a negative mean value. Thus, local spatial stochasticity has been lost due to disease causing mutations, indicating a loss of structural fluidity. The amino acid conversion plot also showed a clear tendency of altered surface patch residue conversion propensity than polymorphic conversions. So far, this is the first report that compares the effects of different kinds of mutations (disease and non-disease causing polymorphic mutations) in the onset of autoimmune diseases. - Highlights: • Protein-Protein Interaction. • Changes in Relative Solvent Accessibility (RSA). • Amino acid conversion matrix. • Polymorphic mutations. • Disease causing mutations.« less

A KCNH2 branch point mutation causing aberrant splicing contributes to an explanation of genotype-negative long QT syndrome.

PubMed

Crotti, Lia; Lewandowska, Marzena A; Schwartz, Peter J; Insolia, Roberto; Pedrazzini, Matteo; Bussani, Erica; Dagradi, Federica; George, Alfred L; Pagani, Franco

2009-02-01

Genetic screening of long QT syndrome (LQTS) fails to identify disease-causing mutations in about 30% of patients. So far, molecular screening has focused mainly on coding sequence mutations or on substitutions at canonical splice sites. The purpose of this study was to explore the possibility that intronic variants not at canonical splice sites might affect splicing regulatory elements, lead to aberrant transcripts, and cause LQTS. Molecular screening was performed through DHPLC and sequence analysis. The role of the intronic mutation identified was assessed with a hybrid minigene splicing assay. A three-generation LQTS family was investigated. Molecular screening failed to identify an obvious disease-causing mutation in the coding sequences of the major LQTS genes but revealed an intronic A-to-G substitution in KCNH2 (IVS9-28A/G) cosegregating with the clinical phenotype in family members. In vitro analysis proved that the mutation disrupts the acceptor splice site definition by affecting the branch point (BP) sequence and promoting intron retention. We further demonstrated a tight functional relationship between the BP and the polypyrimidine tract, whose weakness is responsible for the pathological effect of the IVS9-28A/G mutation. We identified a novel BP mutation in KCNH2 that disrupts the intron 9 acceptor splice site definition and causes LQT2. The present finding demonstrates that intronic mutations affecting pre-mRNA processing may contribute to the failure of traditional molecular screening in identifying disease-causing mutations in LQTS subjects and offers a rationale strategy for the reduction of genotype-negative cases.

In silico investigation of molecular effects caused by missense mutations in creatine transporter protein

NASA Astrophysics Data System (ADS)

Zhang, Zhe; Schwatz, Charles; Alexov, Emil

2011-03-01

Creatine transporter (CT) protein, which is encoded by SLC6A8 gene, is essential for taking up the creatine in the cell, which in turn plays a key role in the spatial and temporal maintenance of energy in skeletal and cardiac muscle cells. It was shown that some missense mutations in CT cause mental retardation, while others are harmless non-synonymous single nucleoside polymorphism (nsSNP). Currently fifteen missense mutations in CT are known, among which twelve are disease-causing. Sequence analysis reveals that there is no clear trend distinguishing disease-causing from harmless missense mutations. Because of that, we built 3D model of the CT using highly homologous template and use the model to investigate the effects of mutations of CT stability and hydrogen bond network. It is demonstrated that disease-causing mutations affect the folding free energy and ionization states of titratable group in much greater extend as compared with harmless mutations. Supported by grants from NLM, NIH, grant numbers 1R03LM009748 and 1R03LM009748-S1.

CD40 agonist antibody mediated improvement of chronic Cryptosporidium infection in patients with X-linked hyper IgM syndrome

USDA-ARS?s Scientific Manuscript database

X-linked hyper-IgM syndrome (XHM) is a combined immune deficiency disorder caused by mutations in CD40 ligand. We tested CP-870,893, a human CD40 agonist monoclonal antibody, in the treatment of two XHM patients with biliary Cryptosporidiosis. CP-870,893 activated B cells and APCs in vitro, restori...

Clinical guideline seom: hereditary colorectal cancer.

PubMed

Guillén-Ponce, C; Serrano, R; Sánchez-Heras, A B; Teulé, A; Chirivella, I; Martín, T; Martínez, E; Morales, R; Robles, L

2015-12-01

Genetic mutations have been identified as the cause of inherited cancer risk in some colon cancer; these mutations are estimated to account for only 5-6 % of colorectal cancer (CRC) cases overall. Up to 25-30 % of patients have a family history of CRC that suggests a hereditary component, common exposures among family members, or a combination of both. Cancers in people with a hereditary predisposition typically occur at an earlier age than in sporadic cases. A predisposition to CRC may include a predisposition to other cancers, such as endometrial cancer. We describe genetics, current diagnosis and management of CRC hereditary syndromes pointing to a multidisciplinary approach to achieve the best results in patients and family outcomes.

Preimplantation diagnosis for neurofibromatosis.

PubMed

Verlinsky, Yury; Rechitsky, Svetlana; Verlinsky, Oleg; Chistokhina, Anna; Sharapova, Tatyana; Masciangelo, Christina; Levy, Michael; Kaplan, Brian; Lederer, Kevin; Kuliev, Anver

2002-01-01

Preimplantation genetic diagnosis (PGD) has recently been performed for inherited cancer predisposition determined by p53 tumour suppressor gene mutations, suggesting the usefulness of PGD for late onset disorders with genetic predisposition, including those caused by the germline mutations of other tumour suppressor genes. Here PGD was performed for two couples, one at risk for producing a child with maternally derived neurofibromatosis type I (NF1), and the other with paternally derived neurofibromatosis type II (NF2). The procedure involved a standard IVF protocol, combined with testing of oocytes or embryos prior to their transfer back to the patients. Maternal mutation Trp-->Ter (TGG-->TGA) in exon 29 of the NF1 gene was tested by sequential PCR analysis of the first and second polar bodies, and paternal L141P mutation in exon 4 of the NF2 gene by embryo biopsy at the cleavage stage. In both cases, multiplex nested PCR was applied, involving NF1 and NF2 mutation analysis simultaneously with the 3 and 2 linked markers, respectively. Of 57 oocytes tested in four PGD cycles for NF1 mutation, 26 mutation-free oocytes were detected, from which eight were preselected for transfer, two in each cycle. These produced two clinical pregnancies, one confirmed to be mutation free by chorionic villus sampling but ending in a stillbirth, and the other still ongoing. Of 18 embryos analysed in a cycle performed for NF2 mutation, eight mutation-free embryos were detected, three of which were transferred back to the patient, resulting in a singleton pregnancy and the birth of a mutation-free child. This suggests that PGD is a useful approach for avoiding the birth of children with inherited cancer predisposition, determined by NF1 and NF2 gene mutations.

Combination of retinitis pigmentosa and hearing loss caused by a novel mutation in PRPH2 and a known mutation in GJB2: importance for differential diagnosis of Usher syndrome.

PubMed

Fakin, Ana; Zupan, Andrej; Glavač, Damjan; Hawlina, Marko

2012-12-15

Purpose of this study was to molecularly characterize a family in which two brothers (46 and 36 years) presented with a combination of retinitis pigmentosa (RP) and severe sensorineural hearing loss while father and sister (71 and 41 years) presented with isolated RP. Retinal phenotype was compared with phenotype of 17 patients with Usher syndrome type 1. Ophthalmological examination included assessment of Snellen visual acuity, color vision with Ishihara tables, Goldmann perimetry (targets II/1-4) and microperimetry. Fundus autofluorescence imaging and optical coherence tomography were performed. Direct sequencing of all coding exons and flanking intronic sequences of GJB2 (gap junction protein, beta 2) and PRPH2 (peripherin 2) genes was performed in younger brother. Other family members were analyzed with sequencing (GJB2), high resolution melt analysis (GJB2) or restriction enzymes (PRPH2). Brothers with hearing loss were found to carry a homozygous c.35 delG mutation in GJB2, the most common mutation associated with recessive hearing loss. All patients were found to carry a novel heterozygous mutation c.389T>C (p.Leu130Pro) on PRPH2. Age of onset was higher in PRPH2 than USH1 patients, however with some overlap. Differentiation from retinal phenotype of USH1 could only be made in the oldest patient, who retained good central visual function after more than three decades of disease. Copyright © 2012 Elsevier Ltd. All rights reserved.

Prenatal diagnosis in a cystic fibrosis family: a combined molecular strategy for a precise diagnosis.

PubMed

Chávez-Saldaña, Margarita; García-Cavazos, Ricardo; Vigueras, Rosa María; Orozco, Lorena

2011-01-01

The high genetic heterogeneity in populations with a wide spectrum of mutations in the CF transmembrane conductance regulator gene (CFTR), makes the detection of mutations a very hard and difficult task, thereby limiting the accurate diagnosis of the disease, mainly in patients with uncharacterized mutations. Molecular strategies, like targeted identification of the most frequent CFTR mutations in Mexican population combined with linkage analysis using markers, is very useful for carrier detection and for prenatal diagnosis in affected families with CF. In this paper we show that the combination of methodologies was a crucial alternative to reach a precise prenatal CF diagnosis. We documented CF diagnosis in a 14th-week fetus combining the screening of the most common mutations in Mexican population with linkage analysis of two extragenic polymorphisms (XV2C/TaqI and KM19/PstI). We determined that the fetus inherited the PG542X mutation from its mother and an unknown mutation from its father through the chromosomal phases analysis.

New evidence for positive selection helps explain the paternal age effect observed in achondroplasia

PubMed Central

Shinde, Deepali N.; Elmer, Dominik P.; Calabrese, Peter; Boulanger, Jérôme; Arnheim, Norman; Tiemann-Boege, Irene

2013-01-01

There are certain de novo germline mutations associated with genetic disorders whose mutation rates per generation are orders of magnitude higher than the genome average. Moreover, these mutations occur exclusively in the male germ line and older men have a higher probability of having an affected child than younger ones, known as the paternal age effect (PAE). The classic example of a genetic disorder exhibiting a PAE is achondroplasia, caused predominantly by a single-nucleotide substitution (c.1138G>A) in FGFR3. To elucidate what mechanisms might be driving the high frequency of this mutation in the male germline, we examined the spatial distribution of the c.1138G>A substitution in a testis from an 80-year-old unaffected man. Using a technology based on bead-emulsion amplification, we were able to measure mutation frequencies in 192 individual pieces of the dissected testis with a false-positive rate lower than 2.7 × 10−6. We observed that most mutations are clustered in a few pieces with 95% of all mutations occurring in 27% of the total testis. Using computational simulations, we rejected the model proposing an elevated mutation rate per cell division at this nucleotide site. Instead, we determined that the observed mutation distribution fits a germline selection model, where mutant spermatogonial stem cells have a proliferative advantage over unmutated cells. Combined with data on several other PAE mutations, our results support the idea that the PAE, associated with a number of Mendelian disorders, may be explained primarily by a selective mechanism. PMID:23740942

Novel mutations in GALNT3 causing hyperphosphatemic familial tumoral calcinosis.

PubMed

Yancovitch, Alan; Hershkovitz, Dov; Indelman, Margareta; Galloway, Peter; Whiteford, Margo; Sprecher, Eli; Kılıç, Esra

2011-09-01

Hyperphosphatemic familial tumoral calcinosis (HFTC) is known to be caused by mutations in at least three genes: FGF23, GALNT3 and KL. Two families with two affected members suffering from HFTC were scrutinized for mutations in these candidate genes. We identified in both families homozygous missense mutations affecting highly conserved amino acids in GALNT3. One of the mutations is a novel mutation, whereas the second mutation was reported before in a compound heterozygous state. Our data expand the spectrum of known mutations in GALNT3 and contribute to a better understanding of the phenotypic manifestations of mutations in this gene.

Disruption of the APC gene by t(5;7) translocation in a Turcot family.

PubMed

Sahnane, Nora; Bernasconi, Barbara; Carnevali, Ileana; Furlan, Daniela; Viel, Alessandra; Sessa, Fausto; Tibiletti, Maria Grazia

2016-03-01

Turcot syndrome (TS) refers to the combination of colorectal polyps and primary tumours of the central nervous system. TS is a heterogeneous genetic condition due to APC and/or mismatch repair germline mutations. When APC is involved the vast majority of mutations are truncating, but in approximately 20%-30% of patients with familial polyposis no germline mutation can be found. A 30-year-old Caucasian woman with a positive pedigree for TS was referred to our Genetic Counselling Service. She was negative for APC and MUTYH but showed a reciprocal balanced translocation t(5;7)(q22;p15) at chromosome analysis. FISH analysis using specific BAC probes demonstrated that 5q22 breakpoint disrupted the APC gene. Transcript analysis by MLPA and digital PCR revealed that the cytogenetic rearrangement involving the 3' end of the APC gene caused a defective expression of a truncated transcript. This result allowed cytogenetic analysis to be offered to all the other family members and segregation analysis clearly demonstrated that all the carriers were affected, whereas non-carriers did not have the polyposis. A cytogenetic approach permitted the identification of the mutation-causing disease in this family, and the segregation analysis together with the transcript study supported the pathogenetic role of this mutation. Karyotype analysis was used as a predictive test in all members of this family. This family suggests that clinically positive TS and FAP cases, which test negative with standard molecular analysis, could be easily and cost-effectively resolved by a classical and molecular cytogenetic approach. Copyright © 2015 Elsevier Inc. All rights reserved.

Optimizing the molecular diagnosis of CDKL5 gene-related epileptic encephalopathy in boys.

PubMed

Mei, Davide; Darra, Francesca; Barba, Carmen; Marini, Carla; Fontana, Elena; Chiti, Laura; Parrini, Elena; Dalla Bernardina, Bernardo; Guerrini, Renzo

2014-11-01

Mutations involving the cyclin-dependent kinase-like 5 (CDKL5) gene cause an early onset epileptic encephalopathy (EE) with severe neurologic impairment and a skewed 12:1 female-to-male ratio. To date, 18 mutations have been described in boys. We analyzed our cohort of boys with early onset EE to assess the diagnostic yield of our molecular approach. We studied 74 boys who presented early onset severe seizures, including infantile spasms and developmental delay, in the setting of EE, using Sanger sequencing, next-generation sequencing (NGS) and multiplex ligation-dependent probe amplification (MLPA). We identified alterations involving CDKL5 in four boys (5.4%) using NGS in one and MLPA in three. Three of four mutations were indicative of somatic mosaicism. CDKL5 gene mutations accounted for 5.4% of boys with early onset EE. Somatic mosaic mutations might be even more represented than germline mutations, probably because their less deleterious effect enhances viability of the male embryo. The molecular approach used for CDKL5 screening remarkably influences the diagnostic yield in boys. Diagnosis is optimized by Sanger sequencing combined with array-based methods or MLPA; alternatively, NGS targeted resequencing designed to also detect copy number alterations, may be performed. Wiley Periodicals, Inc. © 2014 International League Against Epilepsy.

Screening of mutations affecting protein stability and dynamics of FGFR1—A simulation analysis

PubMed Central

Doss, C. George Priya; Rajith, B.; Garwasis, Nimisha; Mathew, Pretty Raju; Raju, Anand Solomon; Apoorva, K.; William, Denise; Sadhana, N.R.; Himani, Tanwar; Dike, IP.

2012-01-01

Single amino acid substitutions in Fibroblast Growth Factor Receptor 1 (FGFR1) destabilize protein and have been implicated in several genetic disorders like various forms of cancer, Kallamann syndrome, Pfeiffer syndrome, Jackson Weiss syndrome, etc. In order to gain functional insight into mutation caused by amino acid substitution to protein function and expression, special emphasis was laid on molecular dynamics simulation techniques in combination with in silico tools such as SIFT, PolyPhen 2.0, I-Mutant 3.0 and SNAP. It has been estimated that 68% nsSNPs were predicted to be deleterious by I-Mutant, slightly higher than SIFT (37%), PolyPhen 2.0 (61%) and SNAP (58%). From the observed results, P722S mutation was found to be most deleterious by comparing results of all in silico tools. By molecular dynamics approach, we have shown that P722S mutation leads to increase in flexibility, and deviated more from the native structure which was supported by the decrease in the number of hydrogen bonds. In addition, biophysical analysis revealed a clear insight of stability loss due to P722S mutation in FGFR1 protein. Majority of mutations predicted by these in silico tools were in good concordance with the experimental results. PMID:27896051

Screening of mutations affecting protein stability and dynamics of FGFR1-A simulation analysis.

PubMed

Doss, C George Priya; Rajith, B; Garwasis, Nimisha; Mathew, Pretty Raju; Raju, Anand Solomon; Apoorva, K; William, Denise; Sadhana, N R; Himani, Tanwar; Dike, I P

2012-12-01

Single amino acid substitutions in Fibroblast Growth Factor Receptor 1 ( FGFR1 ) destabilize protein and have been implicated in several genetic disorders like various forms of cancer, Kallamann syndrome, Pfeiffer syndrome, Jackson Weiss syndrome, etc. In order to gain functional insight into mutation caused by amino acid substitution to protein function and expression, special emphasis was laid on molecular dynamics simulation techniques in combination with in silico tools such as SIFT, PolyPhen 2.0, I-Mutant 3.0 and SNAP. It has been estimated that 68% nsSNPs were predicted to be deleterious by I-Mutant, slightly higher than SIFT (37%), PolyPhen 2.0 (61%) and SNAP (58%). From the observed results, P722S mutation was found to be most deleterious by comparing results of all in silico tools. By molecular dynamics approach, we have shown that P722S mutation leads to increase in flexibility, and deviated more from the native structure which was supported by the decrease in the number of hydrogen bonds. In addition, biophysical analysis revealed a clear insight of stability loss due to P722S mutation in FGFR1 protein. Majority of mutations predicted by these in silico tools were in good concordance with the experimental results.

Three rare diseases in one Sib pair: RAI1, PCK1, GRIN2B mutations associated with Smith-Magenis Syndrome, cytosolic PEPCK deficiency and NMDA receptor glutamate insensitivity.

PubMed

Adams, David R; Yuan, Hongjie; Holyoak, Todd; Arajs, Katrina H; Hakimi, Parvin; Markello, Thomas C; Wolfe, Lynne A; Vilboux, Thierry; Burton, Barbara K; Fajardo, Karin Fuentes; Grahame, George; Holloman, Conisha; Sincan, Murat; Smith, Ann C M; Wells, Gordon A; Huang, Yan; Vega, Hugo; Snyder, James P; Golas, Gretchen A; Tifft, Cynthia J; Boerkoel, Cornelius F; Hanson, Richard W; Traynelis, Stephen F; Kerr, Douglas S; Gahl, William A

2014-11-01

The National Institutes of Health Undiagnosed Diseases Program evaluates patients for whom no diagnosis has been discovered despite a comprehensive diagnostic workup. Failure to diagnose a condition may arise from the mutation of genes previously unassociated with disease. However, we hypothesized that this could also co-occur with multiple genetic disorders. Demonstrating a complex syndrome caused by multiple disorders, we report two siblings manifesting both similar and disparate signs and symptoms. They shared a history of episodes of hypoglycemia and lactic acidosis, but had differing exam findings and developmental courses. Clinical acumen and exome sequencing combined with biochemical and functional studies identified three genetic conditions. One sibling had Smith-Magenis Syndrome and a nonsense mutation in the RAI1 gene. The second sibling had a de novo mutation in GRIN2B, which resulted in markedly reduced glutamate potency of the encoded receptor. Both siblings had a protein-destabilizing homozygous mutation in PCK1, which encodes the cytosolic isoform of phosphoenolpyruvate carboxykinase (PEPCK-C). In summary, we present the first clinically-characterized mutation of PCK1 and demonstrate that complex medical disorders can represent the co-occurrence of multiple diseases. Copyright © 2014 Elsevier Inc. All rights reserved.

Rapid genotyping of common MeCP2 mutations with an electronic DNA microchip using serial differential hybridization.

PubMed

Thistlethwaite, William A; Moses, Linda M; Hoffbuhr, Kristen C; Devaney, Joseph M; Hoffman, Eric P

2003-05-01

Rett syndrome is a neurodevelopmental disorder that affects females almost exclusively, and in which eight common point mutations on the X-linked MeCP2 gene are knows to cause over 70% of mutation-positive cases. We explored the use of a novel platform to detect the eight common mutations in Rett syndrome patients to expedite and simplify the process of identification of known genotypes. The Nanogen workstation consists of a two-color assay based on electric hybridization and thermal discrimination, all performed on an electronically active NanoChip. This genotyping platform was tested on 362 samples of a pre-determined genotype, which had been previously identified by a combination of DHPLC (denaturing high performance liquid chromatography) and direct sequencing. This genotyping technique proved to be rapid, facile, and displayed a specificity of 100% with 3% ambiguity. In addition, we present consecutive testing of seven mutations on a single pad of the NanoChip. This was accomplished by tagging down two amplimers together and serially hybridizing for seven different loci, allowing us to genotype samples for seven of the eight common Rett mutations on a single pad. This novel method displayed the same level of specificity and accuracy as the single amplimer reactions, and proved to be faster and more economical.

Multiple levels of redundant processes inhibit Caenorhabditis elegans vulval cell fates.

PubMed

Andersen, Erik C; Saffer, Adam M; Horvitz, H Robert

2008-08-01

Many mutations cause obvious abnormalities only when combined with other mutations. Such synthetic interactions can be the result of redundant gene functions. In Caenorhabditis elegans, the synthetic multivulva (synMuv) genes have been grouped into multiple classes that redundantly inhibit vulval cell fates. Animals with one or more mutations of the same class undergo wild-type vulval development, whereas animals with mutations of any two classes have a multivulva phenotype. By varying temperature and genetic background, we determined that mutations in most synMuv genes within a single synMuv class enhance each other. However, in a few cases no enhancement was observed. For example, mutations that affect an Mi2 homolog and a histone methyltransferase are of the same class and do not show enhancement. We suggest that such sets of genes function together in vivo and in at least some cases encode proteins that interact physically. The approach of genetic enhancement can be applied more broadly to identify potential protein complexes as well as redundant processes or pathways. Many synMuv genes are evolutionarily conserved, and the genetic relationships we have identified might define the functions not only of synMuv genes in C. elegans but also of their homologs in other organisms.

Multiple Levels of Redundant Processes Inhibit Caenorhabditis elegans Vulval Cell Fates

PubMed Central

Andersen, Erik C.; Saffer, Adam M.; Horvitz, H. Robert

2008-01-01

Many mutations cause obvious abnormalities only when combined with other mutations. Such synthetic interactions can be the result of redundant gene functions. In Caenorhabditis elegans, the synthetic multivulva (synMuv) genes have been grouped into multiple classes that redundantly inhibit vulval cell fates. Animals with one or more mutations of the same class undergo wild-type vulval development, whereas animals with mutations of any two classes have a multivulva phenotype. By varying temperature and genetic background, we determined that mutations in most synMuv genes within a single synMuv class enhance each other. However, in a few cases no enhancement was observed. For example, mutations that affect an Mi2 homolog and a histone methyltransferase are of the same class and do not show enhancement. We suggest that such sets of genes function together in vivo and in at least some cases encode proteins that interact physically. The approach of genetic enhancement can be applied more broadly to identify potential protein complexes as well as redundant processes or pathways. Many synMuv genes are evolutionarily conserved, and the genetic relationships we have identified might define the functions not only of synMuv genes in C. elegans but also of their homologs in other organisms. PMID:18689876

Genotyping of K-ras codons 12 and 13 mutations in colorectal cancer by capillary electrophoresis.

PubMed

Chen, Yen-Ling; Chang, Ya-Sian; Chang, Jan-Gowth; Wu, Shou-Mei

2009-06-26

Point mutations of the K-ras gene located in codons 12 and 13 cause poor responses to the anti-epidermal growth factor receptor (anti-EGFR) therapy of colorectal cancer (CRC) patients. Besides, mutations of K-ras gene have also been proven to play an important role in human tumor progression. We established a simple and effective capillary electrophoresis (CE) method for simultaneous point mutation detection in codons 12 and 13 of K-ras gene. We combined one universal fluorescence-based nonhuman-sequence primer and two fragment-oriented primers in one tube, and performed this two-in-one polymerase chain reaction (PCR). PCR fragments included wild type and seven point mutations at codons 12 and 13 of K-ras gene. The amplicons were analyzed by single-strand conformation polymorphism (SSCP)-CE method. The CE analysis was performed by using a 1x Tris-borate-EDTA (TBE) buffer containing 1.5% (w/v) hydroxyethylcellulose (HEC) (MW 250,000) under reverse polarity with 15 degrees C and 30 degrees C. Ninety colorectal cancer patients were blindly genotyped using this developed method. The results showed good agreement with those of DNA sequencing method. The SSCP-CE was feasible for mutation screening of K-ras gene in populations.

Identification of a novel missense mutation of MAF in a Japanese family with congenital cataract by whole exome sequencing: a clinical report and review of literature.

PubMed

Narumi, Yoko; Nishina, Sachiko; Tokimitsu, Motoharu; Aoki, Yoko; Kosaki, Rika; Wakui, Keiko; Azuma, Noriyuki; Murata, Toshinori; Takada, Fumio; Fukushima, Yoshimitsu; Kosho, Tomoki

2014-05-01

Congenital cataracts are the most important cause of severe visual impairment in infants. Genetic factors contribute to the disease development and 29 genes are known to cause congenital cataracts. Identifying the genetic cause of congenital cataracts can be difficult because of genetic heterogeneity. V-maf avian musculoaponeurotic fibrosarcoma oncogene homolog (MAF) encodes a basic region/leucine zipper transcription factor that plays a key role as a regulator of embryonic lens fiber cell development. MAF mutations have been reported to cause juvenile-onset pulverulent cataract, microcornea, iris coloboma, and other anterior segment dysgenesis. We report on six patients in a family who have congenital cataracts were identified MAF mutation by whole exome sequencing (WES). The heterozygous MAF mutation Q303L detected in the present family occurs in a well conserved glutamine residue at the basic region of the DNA-binding domain. All affected members showed congenital cataracts. Three of the six members showed microcornea and one showed iris coloboma. Congenital cataracts with MAF mutation exhibited phenotypically variable cataracts within the family. Review of the patients with MAF mutations supports the notion that congenital cataracts caused by MAF mutations could be accompanied by microcornea and/or iris coloboma. WES is a useful tool for detecting disease-causing mutations in patients with genetically heterogeneous conditions. © 2014 Wiley Periodicals, Inc.

Targeted Resequencing of 29 Candidate Genes and Mouse Expression Studies Implicate ZIC3 and FOXF1 in Human VATER/VACTERL Association.

PubMed

Hilger, Alina C; Halbritter, Jan; Pennimpede, Tracie; van der Ven, Amelie; Sarma, Georgia; Braun, Daniela A; Porath, Jonathan D; Kohl, Stefan; Hwang, Daw-Yang; Dworschak, Gabriel C; Hermann, Bernhard G; Pavlova, Anna; El-Maarri, Osman; Nöthen, Markus M; Ludwig, Michael; Reutter, Heiko; Hildebrandt, Friedhelm

2015-12-01

The VATER/VACTERL association describes the combination of congenital anomalies including vertebral defects, anorectal malformations, cardiac defects, tracheoesophageal fistula with or without esophageal atresia, renal malformations, and limb defects. As mutations in ciliary genes were observed in diseases related to VATER/VACTERL, we performed targeted resequencing of 25 ciliary candidate genes as well as disease-associated genes (FOXF1, HOXD13, PTEN, ZIC3) in 123 patients with VATER/VACTERL or VATER/VACTERL-like phenotype. We detected no biallelic mutation in any of the 25 ciliary candidate genes; however, identified an identical, probably disease-causing ZIC3 missense mutation (p.Gly17Cys) in four patients and a FOXF1 de novo mutation (p.Gly220Cys) in a further patient. In situ hybridization analyses in mouse embryos between E9.5 and E14.5 revealed Zic3 expression in limb and prevertebral structures, and Foxf1 expression in esophageal, tracheal, vertebral, anal, and genital tubercle tissues, hence VATER/VACTERL organ systems. These data provide strong evidence that mutations in ZIC3 or FOXF1 contribute to VATER/VACTERL. © 2015 WILEY PERIODICALS, INC.

The DNA cytosine deaminase APOBEC3H haplotype I likely contributes to breast and lung cancer mutagenesis.

PubMed

Starrett, Gabriel J; Luengas, Elizabeth M; McCann, Jennifer L; Ebrahimi, Diako; Temiz, Nuri A; Love, Robin P; Feng, Yuqing; Adolph, Madison B; Chelico, Linda; Law, Emily K; Carpenter, Michael A; Harris, Reuben S

2016-09-21

Cytosine mutations within TCA/T motifs are common in cancer. A likely cause is the DNA cytosine deaminase APOBEC3B (A3B). However, A3B-null breast tumours still have this mutational bias. Here we show that APOBEC3H haplotype I (A3H-I) provides a likely solution to this paradox. A3B-null tumours with this mutational bias have at least one copy of A3H-I despite little genetic linkage between these genes. Although deemed inactive previously, A3H-I has robust activity in biochemical and cellular assays, similar to A3H-II after compensation for lower protein expression levels. Gly105 in A3H-I (versus Arg105 in A3H-II) results in lower protein expression levels and increased nuclear localization, providing a mechanism for accessing genomic DNA. A3H-I also associates with clonal TCA/T-biased mutations in lung adenocarcinoma suggesting this enzyme makes broader contributions to cancer mutagenesis. These studies combine to suggest that A3B and A3H-I, together, explain the bulk of 'APOBEC signature' mutations in cancer.

Novel deletions involving the USH2A gene in patients with Usher syndrome and retinitis pigmentosa.

PubMed

García-García, Gema; Aller, Elena; Jaijo, Teresa; Aparisi, Maria J; Larrieu, Lise; Faugère, Valérie; Blanco-Kelly, Fiona; Ayuso, Carmen; Roux, Anne-Francoise; Millán, José M

2014-01-01

The aim of the present work was to identify and characterize large rearrangements involving the USH2A gene in patients with Usher syndrome and nonsyndromic retinitis pigmentosa. The multiplex ligation-dependent probe amplification (MLPA) technique combined with a customized array-based comparative genomic hybridization (aCGH) analysis was applied to 40 unrelated patients previously screened for point mutations in the USH2A gene in which none or only one pathologic mutation was identified. We detected six large deletions involving USH2A in six out of the 40 cases studied. Three of the patients were homozygous for the deletion, and the remaining three were compound heterozygous with a previously identified USH2A point mutation. In five of these cases, the patients displayed Usher type 2, and the remaining case displayed nonsyndromic retinitis pigmentosa. The exact breakpoint junctions of the deletions found in USH2A in four of these cases were characterized. Our study highlights the need to develop improved efficient strategies of mutation screening based upon next generation sequencing (NGS) that reduce cost, time, and complexity and allow simultaneous identification of all types of disease-causing mutations in diagnostic procedures.

Presenilin 1 mutations influence processing and trafficking of the ApoE receptor apoER2.

PubMed

Wang, Wei; Moerman-Herzog, Andrea M; Slaton, Arthur; Barger, Steven W

2017-01-01

Presenilin (PS)-1 is an intramembrane protease serving as the catalytic component of γ-secretase. Mutations in the PS1 gene are the most common cause of familial Alzheimer's disease (FAD). The low-density lipoprotein (LDL)-receptor family member apoER2 is a γ-secretase substrate that has been associated with AD in several ways, including acting as a receptor for apolipoprotein E (ApoE). ApoER2 is processed by γ-secretase into a C-terminal fragment (γ-CTF) that appears to regulate gene expression. FAD PS1 mutations were tested for effects on apoER2. PS1 mutation R278I showed impaired γ-secretase activity for apoER2 in the basal state or after exposure to Reelin. PS1 M146V mutation permitted accumulation of apoER2 CTFs after Reelin treatment, whereas no difference was seen between wild-type (WT) and M146V in the basal state. PS1 L282V mutation, combined with the γ-secretase inhibitor N-(N-[3,5-Difluorophenacetyl]-L-alanyl)-S-phenylglycine t-butyl ester, greatly reduced the cell-surface levels of apoER2 without affecting total apoER2 levels, suggesting a defect in receptor trafficking. These findings indicate that impaired processing or localization of apoER2 may contribute to the pathogenic effects of FAD mutations in PS1. Published by Elsevier Inc.

Adaptive Mutations in Influenza A/California/07/2009 Enhance Polymerase Activity and Infectious Virion Production

PubMed Central

Slaine, Patrick D.; MacRae, Cara; Kleer, Mariel; Lamoureux, Emily; McAlpine, Sarah; Warhuus, Michelle; Comeau, André M.; Hatchette, Todd

2018-01-01

Mice are not natural hosts for influenza A viruses (IAVs), but they are useful models for studying antiviral immune responses and pathogenesis. Serial passage of IAV in mice invariably causes the emergence of adaptive mutations and increased virulence. Here, we report the adaptation of IAV reference strain A/California/07/2009(H1N1) (also known as CA/07) in outbred Swiss Webster mice. Serial passage led to increased virulence and lung titers, and dissemination of the virus to brains. We adapted a deep-sequencing protocol to identify and enumerate adaptive mutations across all genome segments. Among mutations that emerged during mouse-adaptation, we focused on amino acid substitutions in polymerase subunits: polymerase basic-1 (PB1) T156A and F740L and polymerase acidic (PA) E349G. These mutations were evaluated singly and in combination in minigenome replicon assays, which revealed that PA E349G increased polymerase activity. By selectively engineering three PB1 and PA mutations into the parental CA/07 strain, we demonstrated that these mutations in polymerase subunits decreased the production of defective viral genome segments with internal deletions and dramatically increased the release of infectious virions from mouse cells. Together, these findings increase our understanding of the contribution of polymerase subunits to successful host adaptation. PMID:29783694

Amplicon Resequencing Identified Parental Mosaicism for Approximately 10% of “de novo” SCN1A Mutations in Children with Dravet Syndrome

PubMed Central

Xu, Xiaojing; Yang, Xiaoxu; Wu, Qixi; Liu, Aijie; Yang, Xiaoling; Ye, Adam Yongxin; Huang, August Yue; Li, Jiarui; Wang, Meng; Yu, Zhe; Wang, Sheng; Zhang, Zhichao; Wu, Xiru

2015-01-01

ABSTRACT The majority of children with Dravet syndrome (DS) are caused by de novo SCN1A mutations. To investigate the origin of the mutations, we developed and applied a new method that combined deep amplicon resequencing with a Bayesian model to detect and quantify allelic fractions with improved sensitivity. Of 174 SCN1A mutations in DS probands which were considered “de novo” by Sanger sequencing, we identified 15 cases (8.6%) of parental mosaicism. We identified another five cases of parental mosaicism that were also detectable by Sanger sequencing. Fraction of mutant alleles in the 20 cases of parental mosaicism ranged from 1.1% to 32.6%. Thirteen (65% of 20) mutations originated paternally and seven (35% of 20) maternally. Twelve (60% of 20) mosaic parents did not have any epileptic symptoms. Their mutant allelic fractions were significantly lower than those in mosaic parents with epileptic symptoms (P = 0.016). We identified mosaicism with varied allelic fractions in blood, saliva, urine, hair follicle, oral epithelium, and semen, demonstrating that postzygotic mutations could affect multiple somatic cells as well as germ cells. Our results suggest that more sensitive tools for detecting low‐level mosaicism in parents of families with seemingly “de novo” mutations will allow for better informed genetic counseling. PMID:26096185

Four USH2A founder mutations underlie the majority of Usher syndrome type 2 cases among non-Ashkenazi Jews.

PubMed

Auslender, Noa; Bandah, Dikla; Rizel, Leah; Behar, Doron M; Shohat, Mordechai; Banin, Eyal; Allon-Shalev, Stavit; Sharony, Reuven; Sharon, Dror; Ben-Yosef, Tamar

2008-06-01

Type 2 Usher syndrome (USH2) is a recessively inherited disorder, characterized by the combination of early onset, moderate-to-severe, sensorineural hearing loss, and vision impairment due to retinitis pigmentosa. From 74% to 90% of USH2 cases are caused by mutations of the USH2A gene. USH2A is composed of 72 exons, encoding for usherin, an extracellular matrix protein, which plays an important role in the development and maintenance of neurosensory cells in both retina and cochlea. To date, over 70 pathogenic mutations of USH2A have been reported in individuals of various ethnicities. Many of these mutations are rare private mutations segregating in single families. The aim of the current work was to investigate the genetic basis for USH2 among Jews of various origins. We found that four USH2A mutations (c.239-240insGTAC, c.1000C>T, c.2209C>T, and c.12067-2A>G) account for 64% of mutant alleles underlying USH2 in Jewish families of non-Ashkenazi descent. Considering the very large size of the USH2A gene and the high number of mutations detected in USH2 patients worldwide, our findings have significant implications for genetic counseling and carrier screening in various Jewish populations.

Autosomal recessive POLR1D mutation with decrease of TCOF1 mRNA is responsible for Treacher Collins syndrome.

PubMed

Schaefer, Elise; Collet, Corinne; Genevieve, David; Vincent, Marie; Lohmann, Dietmar R; Sanchez, Elodie; Bolender, Chantal; Eliot, Marie-Madeleine; Nürnberg, Gudrun; Passos-Bueno, Maria-Rita; Wieczorek, Dagmar; van Maldergem, Lionel; Doray, Bérénice

2014-09-01

Treacher Collins syndrome is a mandibulofacial dysostosis caused by mutations in genes involved in ribosome biogenesis and synthesis. TCOF1 mutations are observed in ~80% of the patients and are inherited in an autosomal dominant manner. Recently, two other genes have been reported in <2% of patients--POLR1D in patients with autosomal dominant inheritance, and POLR1C in patients with autosomal recessive inheritance. We performed direct sequencing of TCOF1, POLR1C, and POLR1D in two unrelated consanguineous families. The four affected children shared the same homozygous mutation in POLR1D (c.163C>G, p.Leu55Val). This mutation is localized in a region encoding the dimerization domain of the RNA polymerase. It is supposed that this mutation impairs RNA polymerase, resulting in a lower amount of mature dimeric ribosomes. A functional analysis of the transcripts of TCOF1 by real-time quantitative reverse transcription-polymerase chain reaction was performed in the first family, demonstrating a 50% reduction in the index case, compatible with this hypothesis. This is the first report of POLR1D mutation being responsible for an autosomal recessive inherited Treacher Collins syndrome. These results reinforce the concept of genetic heterogeneity of Treacher Collins syndrome and underline the importance of combining clinical expertise and familial molecular analyses for appropriate genetic counseling.

Disease-Causing Mutations in the G Protein Gαs Subvert the Roles of GDP and GTP.

PubMed

Hu, Qi; Shokat, Kevan M

2018-05-17

The single most frequent cancer-causing mutation across all heterotrimeric G proteins is R201C in Gαs. The current model explaining the gain-of-function activity of the R201 mutations is through the loss of GTPase activity and resulting inability to switch off to the GDP state. Here, we find that the R201C mutation can bypass the need for GTP binding by directly activating GDP-bound Gαs through stabilization of an intramolecular hydrogen bond network. Having found that a gain-of-function mutation can convert GDP into an activator, we postulated that a reciprocal mutation might disrupt the normal role of GTP. Indeed, we found R228C, a loss-of-function mutation in Gαs that causes pseudohypoparathyroidism type 1a (PHP-Ia), compromised the adenylyl cyclase-activating activity of Gαs bound to a non-hydrolyzable GTP analog. These findings show that disease-causing mutations in Gαs can subvert the canonical roles of GDP and GTP, providing new insights into the regulation mechanism of G proteins. Copyright © 2018 Elsevier Inc. All rights reserved.

Nuclear factors involved in mitochondrial translation cause a subgroup of combined respiratory chain deficiency.

PubMed

Kemp, John P; Smith, Paul M; Pyle, Angela; Neeve, Vivienne C M; Tuppen, Helen A L; Schara, Ulrike; Talim, Beril; Topaloglu, Haluk; Holinski-Feder, Elke; Abicht, Angela; Czermin, Birgit; Lochmüller, Hanns; McFarland, Robert; Chinnery, Patrick F; Chrzanowska-Lightowlers, Zofia M A; Lightowlers, Robert N; Taylor, Robert W; Horvath, Rita

2011-01-01

Mutations in several mitochondrial DNA and nuclear genes involved in mitochondrial protein synthesis have recently been reported in combined respiratory chain deficiency, indicating a generalized defect in mitochondrial translation. However, the number of patients with pathogenic mutations is small, implying that nuclear defects of mitochondrial translation are either underdiagnosed or intrauterine lethal. No comprehensive studies have been reported on large cohorts of patients with combined respiratory chain deficiency addressing the role of nuclear genes affecting mitochondrial protein synthesis to date. We investigated a cohort of 52 patients with combined respiratory chain deficiency without causative mitochondrial DNA mutations, rearrangements or depletion, to determine whether a defect in mitochondrial translation defines the pathomechanism of their clinical disease. We followed a combined approach of sequencing known nuclear genes involved in mitochondrial protein synthesis (EFG1, EFTu, EFTs, MRPS16, TRMU), as well as performing in vitro functional studies in 22 patient cell lines. The majority of our patients were children (<15 years), with an early onset of symptoms <1 year of age (65%). The most frequent clinical presentation was mitochondrial encephalomyopathy (63%); however, a number of patients showed cardiomyopathy (33%), isolated myopathy (15%) or hepatopathy (13%). Genomic sequencing revealed compound heterozygous mutations in the mitochondrial transfer ribonucleic acid modifying factor (TRMU) in a single patient only, presenting with early onset, reversible liver disease. No pathogenic mutation was detected in any of the remaining 51 patients in the other genes analysed. In vivo labelling of mitochondrial polypeptides in 22 patient cell lines showed overall (three patients) or selective (four patients) defects of mitochondrial translation. Immunoblotting for mitochondrial proteins revealed decreased steady state levels of proteins in some patients, but normal or increased levels in others, indicating a possible compensatory mechanism. In summary, candidate gene sequencing in this group of patients has a very low detection rate (1/52), although in vivo labelling of mitochondrial translation in 22 patient cell lines indicate that a nuclear defect affecting mitochondrial protein synthesis is responsible for about one-third of combined respiratory chain deficiencies (7/22). In the remaining patients, the impaired respiratory chain activity is most likely the consequence of several different events downstream of mitochondrial translation. Clinical classification of patients with biochemical analysis, genetic testing and, more importantly, in vivo labelling and immunoblotting of mitochondrial proteins show incoherent results, but a systematic review of these data in more patients may reveal underlying mechanisms, and facilitate the identification of novel factors involved in combined respiratory chain deficiency.

In vitro and in vivo evidence of a potential A(H1N1)pdm09 antigenic drift mediated by escape mutations in the haemagglutinin Sa antigenic site.

PubMed

Retamal, Miguel; Abed, Yacine; Rhéaume, Chantal; Baz, Mariana; Boivin, Guy

2017-06-01

Influenza A(H1N1)pdm09 virus continues to circulate worldwide without evidence of significant antigenic drift between 2009 and 2016. By using escape mutants, we previously identified six haemagglutinin (HA) changes (T80R, G143E, G158E, N159D, K166E and A198E) that were located within antigenic sites. Combinations of these mutations were introduced into the A(H1N1)pdm09 HA plasmid by mutagenesis. Reassortant 6 : 2 viruses containing both the HA and NA genes of the A(H1N1)pdm09 and the six internal gene segments of A/PR/8/34 were rescued by reverse genetics. In vitro, HA inhibition and microneutralization assays showed that the HA hexa-mutant reassortant virus (RG1) escaped A(H1N1)pdm09 hyper-immune ferret antiserum recognition. C57Black/6 mice that received the vaccine formulated with A/California/07/09 were challenged with 2×104 p.f.u. of either the 6 : 2 wild-type (WT) or RG1 viruses. Reductions in body weight loss, mortality rate and lung viral titre were observed in immunized animals challenged with the 6 : 2 WT virus compared to non-immunized mice. However, immunization did not protect mice challenged with RG1 virus. To further characterize the mutations causing this antigenic change, 11 additional RG viruses whose HA gene contained single or combinations of mutations were evaluated in vitro. Although the RG1 virus was still the least reactive against hyper-immune serum by HAI testing, mutations G158E and N159D within the Sa antigenic site appeared to play the major role in the altered antigenicity of the A(H1N1)pdm09 virus. These results show that the Sa antigenic site contains the most prominent epitopes susceptible to cause an antigenic drift, escaping actual vaccine protection.

[Hereditary hypomelanocytoses: the role of PAX3, SOX10, MITF, SNAI2, KIT, EDN3 and EDNRB genes].

PubMed

Otręba, Michał; Miliński, Maciej; Buszman, Ewa; Wrześniok, Dorota; Beberok, Artur

2013-11-26

Hypo- and hyperpigmentation disorders are the most severe dermatological diseases observed in patients from all over the world. These disorders can be divided into melanoses connected with disorders of melanocyte function and melanocytoses connected with melanocyte development. The article presents some hereditary hypomelanocytoses, which are caused by abnormal melanoblast development, migration and proliferation as well as by abnormal melanocyte viability and proliferation. These disorders are represented by Waardenburg syndrome, piebaldism and Tietz syndrome, and are caused by different mutations of various or the same genes. The types of mutations comprise missense and nonsense mutations, frameshifts (in-frame insertions or deletions), truncating variations, splice alterations and non-stop mutations. It has been demonstrated that mutations of the same gene may cause different hypopigmentation syndromes that may have similar phenotypes. For example, mutations of the MITF gene cause Waardenburg syndrome type 2A as well as Tietz syndrome. It has also been demonstrated that mutations of different genes may cause an identical syndrome. For example, mutations of MITF, SNAI2 and SOX10 genes are observed in Waardenburg syndrome type II and mutations of EDNRB, EDN3 and SOX10 genes are responsible for Waardenburg syndrome type IV. In turn, mutation of the KIT gene and/or heterozygous deletion of the SNAI2 gene result in piebaldism disease. The knowledge of the exact mechanisms of pigmentary disorders may be useful in the development of new therapeutic approaches to their treatment.

Genetic Screening of Selected Disease-Causing Mutations in Glutaryl-CoA Dehydrogenase Gene among Indian Patients with Glutaric Aciduria Type I.

PubMed

Tp, Kruthika-Vinod; Muntaj, Shaik; Devaraju, K S; Kamate, M; Vedamurthy, A B

2017-09-01

Glutaric aciduria type I (GA-I) is an organic aciduria caused by glutaryl-CoA dehydrogenase (GCDH) deficiency. There are limited studies on GA-I from India. A total of 48 Indian GA-I patients were screened for selected disease-causing mutations such as R402W, A421V, A293T, R227P, and V400M using polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP). Among these patients, 9 (18.8%) had R402W mutation, and none had A421V, A293T, R227P, or V400M mutation. One low excretor mutation (P286S) and several novel mutations (I152M, Q144P, and E414X) were also found in this study. We conclude that among selected mutations, R402W is the most common mutation found among Indian GA-I patients.

Novel HSF4 mutation causes congenital total white cataract in a Chinese family.

PubMed

Ke, Tie; Wang, Qing K; Ji, Binchu; Wang, Xu; Liu, Ping; Zhang, Xianqin; Tang, Zhaohui; Ren, Xiang; Liu, Mugen

2006-08-01

To identify the disease-causing gene (mutation) in a Chinese family affected with autosomal dominant congenital total white cataract. Observational case series. Genotyping and linkage analyses were used to identify the linkage of the disease-causing gene in the Chinese family to the HSF4 gene encoding a member of the family of heat shock transcription factors (HSFs). Direct DNA sequence analysis was used to identify the disease-causing mutation. Polymerase chain reaction/restriction fragment length polymorphism analysis was used to demonstrate cosegregation of the HSF4 mutation with the cataract and the absence of the mutation in the normal controls. The cataract gene in the Chinese family was linked to marker D16S3043, and further haplotype analysis defined the causative gene between D16S515 and D16S415 within which HSF4 is located. A novel mutation c.221G>A was identified in HSF4, which results in substitution of a highly conserved arginine residue by histidine at codon 74 (p.R74H). The R74H mutation cosegregated with the affected individuals in the family and did not exist in unaffected family members and 150 unrelated normal controls. These results identified a novel missense mutation R74H in the transcription factor gene HSF4 in a Chinese cataract family and expand the spectrum of HSF4 mutations causing cataract.

Genetic stability of Rift Valley fever virus MP-12 vaccine during serial passages in culture cells.

PubMed

Lokugamage, Nandadeva; Ikegami, Tetsuro

2017-01-01

Rift Valley fever (RVF) is a mosquito-borne zoonotic disease endemic to Africa which affects both ruminants and humans. RVF causes serious damage to the livestock industry and is also a threat to public health. The Rift Valley fever virus has a segmented negative-stranded RNA genome consisting of Large (L)-, Medium (M)-, and Small (S)-segments. The live-attenuated MP-12 vaccine is immunogenic in livestock and humans, and is conditionally licensed for veterinary use in the U.S. The MP-12 strain encodes 23 mutations (nine amino acid substitutions) and is attenuated through a combination of mutations in the L-, M-, and S-segments. Among them, the M-U795C, M-A3564G, and L-G3104A mutations contribute to viral attenuation through the L- and M-segments. The M-U795C, M-A3564G, L-U533C, and L-G3750A mutations are also independently responsible for temperature-sensitive (ts) phenotype. We hypothesized that a serial passage of the MP-12 vaccine in culture cells causes reversions of the MP-12 genome. The MP-12 vaccine and recombinant rMP12-ΔNSs16/198 were serially passaged 25 times. Droplet digital PCR analysis revealed that the reversion occurred at L-G3750A during passages of MP-12 in Vero or MRC-5 cells. The reversion also occurred at M-A3564G and L-U533C of rMP12-ΔNSs16/198 in Vero cells. Reversion mutations were not found in MP-12 or the variant, rMP12-TOSNSs, in the brains of mice with encephalitis. This study characterized genetic stability of the MP-12 vaccine and the potential risk of reversion mutation at the L-G3750A ts mutation after excessive viral passages in culture cells.

Identification of a novel COL1A1 frameshift mutation, c.700delG, in a Chinese osteogenesis imperfecta family

PubMed Central

Wang, Xiran; Pei, Yu; Dou, Jingtao; Lu, Juming; Li, Jian; Lv, Zhaohui

2015-01-01

Osteogenesis imperfecta (OI) is a family of genetic disorders associated with bone loss and fragility. Mutations associated with OI have been found in genes encoding the type I collagen chains. People with OI type I often produce insufficient α1-chain type I collagen because of frameshift, nonsense, or splice site mutations in COL1A1 or COL1A2. This report is of a Chinese daughter and mother who had both experienced two bone fractures. Because skeletal fragility is predominantly inherited, we focused on identifying mutations in COL1A1 and COL1A2 genes. A novel mutation in COL1A1, c.700delG, was detected by genomic DNA sequencing in the mother and daughter, but not in their relatives. The identification of this mutation led to the conclusion that they were affected by mild OI type I. Open reading frame analysis indicated that this frameshift mutation would truncate α1-chain type I collagen at residue p263 (p.E234KfsX264), while the wild-type protein would contain 1,464 residues. The clinical data were consistent with the patients’ diagnosis of mild OI type I caused by haploinsufficiency of α1-chain type I collagen. Combined with previous reports, identification of the novel mutation COL1A1-c.700delG in these patients suggests that additional genetic and environmental factors may influence the severity of OI. PMID:25983617

Homozygosity mapping in autosomal recessive retinitis pigmentosa families detects novel mutations

PubMed Central

Marzouka, Nour al Dain; Hebrard, Maxime; Manes, Gaël; Sénéchal, Audrey; Meunier, Isabelle; Hamel, Christian P.

2013-01-01

Purpose Autosomal recessive retinitis pigmentosa (arRP) is a genetically heterogeneous disease resulting in progressive loss of photoreceptors that leads to blindness. To date, 36 genes are known to cause arRP, rendering the molecular diagnosis a challenge. The aim of this study was to use homozygosity mapping to identify the causative mutation in a series of inbred families with arRP. Methods arRP patients underwent standard ophthalmic examination, Goldman perimetry, fundus examination, retinal OCT, autofluorescence measurement, and full-field electroretinogram. Fifteen consanguineous families with arRP excluded for USH2A and EYS were genotyped on 250 K SNP arrays. Homozygous regions were listed, and known genes within these regions were PCR sequenced. Familial segregation and mutation analyzes were performed. Results We found ten mutations, seven of which were novel mutations in eight known genes, including RP1, IMPG2, NR2E3, PDE6A, PDE6B, RLBP1, CNGB1, and C2ORF71, in ten out of 15 families. The patients carrying RP1, C2ORF71, and IMPG2 mutations presented with severe RP, while those with PDE6A, PDE6B, and CNGB1 mutations were less severely affected. The five families without mutations in known genes could be a source of identification of novel genes. Conclusions Homozygosity mapping combined with systematic screening of known genes results in a positive molecular diagnosis in 66.7% of families. PMID:24339724

Germline mutations in RYR1 are associated with foetal akinesia deformation sequence/lethal multiple pterygium syndrome.

PubMed

McKie, Arthur B; Alsaedi, Atif; Vogt, Julie; Stuurman, Kyra E; Weiss, Marjan M; Shakeel, Hassan; Tee, Louise; Morgan, Neil V; Nikkels, Peter G J; van Haaften, Gijs; Park, Soo-Mi; van der Smagt, Jasper J; Bugiani, Marianna; Maher, Eamonn R

2014-12-05

Foetal akinesia deformation sequence syndrome (FADS) is a genetically heterogeneous disorder characterised by the combination of foetal akinesia and developmental defects which may include pterygia (joint webbing). Traditionally multiple pterygium syndrome (MPS) has been divided into two forms: prenatally lethal (LMPS) and non-lethal Escobar type (EVMPS) types. Interestingly, FADS, LMPS and EVMPS may be allelic e.g. each of these phenotypes may result from mutations in the foetal acetylcholine receptor gamma subunit gene (CHRNG). Many cases of FADS and MPS do not have a mutation in a known FADS/MPS gene and we undertook molecular genetic studies to identify novel causes of these phenotypes. After mapping a novel locus for FADS/LMPS to chromosome 19, we identified a homozygous null mutation in the RYR1 gene in a consanguineous kindred with recurrent LMPS pregnancies. Resequencing of RYR1 in a cohort of 66 unrelated probands with FADS/LMPS/EVMPS (36 with FADS/LMPS and 30 with EVMPS) revealed two additional homozygous mutations (in frame deletions). The overall frequency of RYR1 mutations in probands with FADS/LMPS was 8.3%. Our findings report, for the first time, a homozygous RYR1 null mutation and expand the range of RYR1-related phenotypes to include early lethal FADS/LMPS. We suggest that RYR1 mutation analysis should be performed in cases of severe FADS/LMPS even in the absence of specific histopathological indicators of RYR1-related disease.

[Clinical and molecular study in a family with autosomal dominant hypohidrotic ectodermal dysplasia].

PubMed

Callea, Michele; Cammarata-Scalisi, Francisco; Willoughby, Colin E; Giglio, Sabrina R; Sani, Ilaria; Bargiacchi, Sara; Traficante, Giovanna; Bellacchio, Emanuele; Tadini, Gianluca; Yavuz, Izzet; Galeotti, Angela; Clarich, Gabriella

2017-02-01

Hypohidrotic ectodermal dysplasia (HED) is a rare disease characterized by deficiency in development of structure derived from the ectoderm and is caused by mutations in the genes EDA, EDAR, or EDARADD. Phenotypes caused by mutations in these three may exhibit similar clinical features, explained by a common signaling pathway. Mutations in EDA gene cause X linked HED, which is the most common form. Mutations in EDAR and EDARADD genes cause autosomal dominant and recessive form of HED. The most striking clinical findings in HED are hypodontia, hypotrichosis and hypohidrosis that can lead to episodes of hyperthermia. We report on clinical findings in a child with HED with autosomal dominant inheritance pattern with a heterozygous mutation c.1072C>T (p.Arg358X) in the EDAR gene. A review of the literature with regard to other cases presenting the same mutation has been carried out and discussed. Sociedad Argentina de Pediatría.

Targeting Nonsense Mutations in Diseases with Translational Read-Through-Inducing Drugs (TRIDs).

PubMed

Nagel-Wolfrum, Kerstin; Möller, Fabian; Penner, Inessa; Baasov, Timor; Wolfrum, Uwe

2016-04-01

In recent years, remarkable advances in the ability to diagnose genetic disorders have been made. The identification of disease-causing genes allows the development of gene-specific therapies with the ultimate goal to develop personalized medicines for each patient according to their own specific genetic defect. In-depth genotyping of many different genes has revealed that ~12% of inherited genetic disorders are caused by in-frame nonsense mutations. Nonsense (non-coding) mutations are caused by point mutations, which generate premature termination codons (PTCs) that cause premature translational termination of the mRNA, and subsequently inhibit normal full-length protein expression. Recently, a gene-based therapeutic approach for genetic diseases caused by nonsense mutations has emerged, namely the so-called translational read-through (TR) therapy. Read-through therapy is based on the discovery that small molecules, known as TR-inducing drugs (TRIDs), allow the translation machinery to suppress a nonsense codon, elongate the nascent peptide chain, and consequently result in the synthesis of full-length protein. Several TRIDs are currently under investigation and research has been performed on several genetic disorders caused by nonsense mutations over the years. These findings have raised hope for the usage of TR therapy as a gene-based pharmacogenetic therapy for nonsense mutations in various genes responsible for a variety of genetic diseases.

A Presenilin-1 Mutation Identified in Familial Alzheimer Disease with Cotton Wool Plaques Causes a Nearly Complete Loss of γ-Secretase Activity*

PubMed Central

Heilig, Elizabeth A.; Xia, Weiming; Shen, Jie; Kelleher, Raymond J.

2010-01-01

Mutations in presenilin-1 and presenilin-2 (PS1 and PS2) are the most common cause of familial Alzheimer disease. PS1 and PS2 are the presumptive catalytic components of the multisubunit γ-secretase complex, which proteolyzes a number of type I transmembrane proteins, including the amyloid precursor protein (APP) and Notch. APP processing by γ-secretase produces β-amyloid peptides (Aβ40 and Aβ42) that accumulate in the Alzheimer disease brain. Here we identify a pathogenic L435F mutation in PS1 in two affected siblings with early-onset familial Alzheimer disease characterized by deposition of cerebral cotton wool plaques. The L435F mutation resides in a conserved C-terminal PAL sequence implicated in active site conformation and catalytic activity. The impact of PS1 mutations in and around the PAL motif on γ-secretase activity was assessed by expression of mutant PS1 in mouse embryo fibroblasts lacking endogenous PS1 and PS2. Surprisingly, the L435F mutation caused a nearly complete loss of γ-secretase activity, including >90% reductions in the generation of Aβ40, Aβ42, and the APP and Notch intracellular domains. Two nonpathogenic PS1 mutations, P433L and L435R, caused essentially complete loss of γ-secretase activity, whereas two previously identified pathogenic PS1 mutations, P436Q and P436S, caused partial loss of function with substantial reductions in production of Aβ40, Aβ42, and the APP and Notch intracellular domains. These results argue against overproduction of Aβ42 as an essential property of presenilin proteins bearing pathogenic mutations. Rather, our findings provide support for the hypothesis that pathogenic mutations cause a general loss of presenilin function. PMID:20460383

Dual HER2\\PIK3CA targeting overcomes single-agent acquired resistance in HER2 amplified uterine serous carcinoma cell lines in vitro and in vivo

PubMed Central

Lopez, Salvatore; Cocco, Emiliano; Black, Jonathan; Bellone, Stefania; Bonazzoli, Elena; Predolini, Federica; Ferrari, Francesca; Schwab, Carlton L.; English, Diana P.; Ratner, Elena; Silasi, Dan-Arin; Azodi, Masoud; Schwartz, Peter E.; Terranova, Corrado; Angioli, Roberto; Santin, Alessandro D.

2015-01-01

HER2/neu gene amplification and PIK3CA driver mutations are common in uterine serous carcinoma (USC), and may represent ideal therapeutic targets against this aggressive variant of endometrial cancer. We examined the sensitivity to neratinib, taselisib and the combination of the two compounds in in vitro and in vivo experiments using PIK3CA mutated and PIK3CA-wild type HER2/neu amplified USC cell lines. Cell viability and cell cycle distribution were assessed using flow-cytometry assays. Downstream signaling was assessed by immunoblotting. Preclinical efficacy of single versus dual inhibition was evaluated in vivo using two USC-xenografts. We found both single agent neratinib and taselisib to be active but only transiently effective in controlling the in vivo growth of USC xenografts harboring HER2/neu gene amplification with or without oncogenic PIK3CA mutations. In contrast, the combination of the two inhibitors caused a stronger and long lasting growth inhibition in both USC xenografts when compared to single agent therapy. Combined targeting of HER2 and PIK3CA was associated with a significant and dose-dependent increase in the percentage of cells in the G0/G1 phase of the cell cycle and a dose-dependent decline in the phosphorylation of S6. Importantly, dual inhibition therapy initiated after tumor progression in single agent-treated mice was still remarkably effective at inducing tumor regression in both large PIK3CA or pan-ErbB inhibitor-resistant USC xenografts. Dual HER2/PIK3CA blockade may represent a novel therapeutic option for USC patients harboring tumors with HER2/neu gene amplification and mutated or wild type PIK3CA resistant to chemotherapy. PMID:26333383

A Population Genetic Model for the Initial Spread of Partially Resistant Malaria Parasites under Anti-Malarial Combination Therapy and Weak Intrahost Competition

PubMed Central

Kim, Yuseob; Escalante, Ananias A.; Schneider, Kristan A.

2014-01-01

To develop public-health policies that extend the lifespan of affordable anti-malarial drugs as effective treatment options, it is necessary to understand the evolutionary processes leading to the origin and spread of mutations conferring drug resistance in malarial parasites. We built a population-genetic model for the emergence of resistance under combination drug therapy. Reproductive cycles of parasites are specified by their absolute fitness determined by clinical parameters, thus coupling the evolutionary-genetic with population-dynamic processes. Initial mutations confer only partial drug-resistance. Therefore, mutant parasites rarely survive combination therapy and within-host competition is very weak among parasites. The model focuses on the early phase of such unsuccessful recurrent mutations. This ends in the rare event of mutants enriching in an infected individual from which the successful spread of resistance over the entire population is initiated. By computer simulations, the waiting time until the establishment of resistant parasites is analysed. Resistance spreads quickly following the first appearance of a host infected predominantly by mutant parasites. This occurs either through a rare transmission of a resistant parasite to an uninfected host or through a rare failure of drugs in removing “transient” mutant alleles. The emergence of resistance is delayed with lower mutation rate, earlier treatment, higher metabolic cost of resistance, longer duration of high drug dose, and higher drug efficacy causing a stronger reduction in the sensitive and resistant parasites’ fitnesses. Overall, contrary to other studies’ proposition, the current model based on absolute fitness suggests that aggressive drug treatment delays the emergence of drug resistance. PMID:25007207

Heterozygous Mutations Causing Partial Prohormone Convertase 1 Deficiency Contribute to Human Obesity

PubMed Central

Creemers, John W.M.; Choquet, Hélène; Stijnen, Pieter; Vatin, Vincent; Pigeyre, Marie; Beckers, Sigri; Meulemans, Sandra; Than, Manuel E.; Yengo, Loïc; Tauber, Maithé; Balkau, Beverley; Elliott, Paul; Jarvelin, Marjo-Riitta; Van Hul, Wim; Van Gaal, Luc; Horber, Fritz; Pattou, François; Froguel, Philippe; Meyre, David

2012-01-01

Null mutations in the PCSK1 gene, encoding the proprotein convertase 1/3 (PC1/3), cause recessive monogenic early onset obesity. Frequent coding variants that modestly impair PC1/3 function mildly increase the risk for common obesity. The aim of this study was to determine the contribution of rare functional PCSK1 mutations to obesity. PCSK1 exons were sequenced in 845 nonconsanguineous extremely obese Europeans. Eight novel nonsynonymous PCSK1 mutations were identified, all heterozygous. Seven mutations had a deleterious effect on either the maturation or the enzymatic activity of PC1/3 in cell lines. Of interest, five of these novel mutations, one of the previously described frequent variants (N221D), and the mutation found in an obese mouse model (N222D), affect residues at or near the structural calcium binding site Ca-1. The prevalence of the newly identified mutations was assessed in 6,233 obese and 6,274 lean European adults and children, which showed that carriers of any of these mutations causing partial PCSK1 deficiency had an 8.7-fold higher risk to be obese than wild-type carriers. These results provide the first evidence of an increased risk of obesity in heterozygous carriers of mutations in the PCSK1 gene. Furthermore, mutations causing partial PCSK1 deficiency are present in 0.83% of extreme obesity phenotypes. PMID:22210313

Heterozygous mutations causing partial prohormone convertase 1 deficiency contribute to human obesity.

PubMed

Creemers, John W M; Choquet, Hélène; Stijnen, Pieter; Vatin, Vincent; Pigeyre, Marie; Beckers, Sigri; Meulemans, Sandra; Than, Manuel E; Yengo, Loïc; Tauber, Maithé; Balkau, Beverley; Elliott, Paul; Jarvelin, Marjo-Riitta; Van Hul, Wim; Van Gaal, Luc; Horber, Fritz; Pattou, François; Froguel, Philippe; Meyre, David

2012-02-01

Null mutations in the PCSK1 gene, encoding the proprotein convertase 1/3 (PC1/3), cause recessive monogenic early onset obesity. Frequent coding variants that modestly impair PC1/3 function mildly increase the risk for common obesity. The aim of this study was to determine the contribution of rare functional PCSK1 mutations to obesity. PCSK1 exons were sequenced in 845 nonconsanguineous extremely obese Europeans. Eight novel nonsynonymous PCSK1 mutations were identified, all heterozygous. Seven mutations had a deleterious effect on either the maturation or the enzymatic activity of PC1/3 in cell lines. Of interest, five of these novel mutations, one of the previously described frequent variants (N221D), and the mutation found in an obese mouse model (N222D), affect residues at or near the structural calcium binding site Ca-1. The prevalence of the newly identified mutations was assessed in 6,233 obese and 6,274 lean European adults and children, which showed that carriers of any of these mutations causing partial PCSK1 deficiency had an 8.7-fold higher risk to be obese than wild-type carriers. These results provide the first evidence of an increased risk of obesity in heterozygous carriers of mutations in the PCSK1 gene. Furthermore, mutations causing partial PCSK1 deficiency are present in 0.83% of extreme obesity phenotypes.

Mutations affecting gyrase in Haemophilus influenzae.

PubMed Central

Setlow, J K; Cabrera-Juárez, E; Albritton, W L; Spikes, D; Mutschler, A

1985-01-01

Mutants separately resistant to novobiocin, coumermycin, nalidixic acid, and oxolinic acid contained gyrase activity as measured in vitro that was resistant to the antibiotics, indicating that the mutations represented structural alterations of the enzyme. One Novr mutant contained an altered B subunit of the enzyme, as judged by the ability of a plasmid, pNov1, containing the mutation to complement a temperature-sensitive gyrase B mutation in Escherichia coli and to cause novobiocin resistance in that strain. Three other Novr mutations did not confer antibiotic resistance to the gyrase but appeared to increase the amount of active enzyme in the cell. One of these, novB1, could only act in cis, whereas a new mutation, novC, could act in trans. An RNA polymerase mutation partially substituted for the novB1 mutation, suggesting that novB1 may be a mutation in a promoter region for the B subunit gene. Growth responses of strains containing various combinations of mutations on plasmids or on the chromosome indicated that low-level resistance to novobiocin or coumermycin may have resulted from multiple copies of wild-type genes coding for the gyrase B subunit, whereas high-level resistance required a structural change in the gyrase B gene and was also dependent on alteration in a regulatory region. When there was mismatch at the novB locus, with the novB1 mutation either on a plasmid or the chromosome, and the corresponding wild-type gene present in trans, chromosome to plasmid recombination during transformation was much higher than when the genes matched, probably because plasmid to chromosome recombination, eliminating the plasmid, was inhibited by the mismatch. PMID:2997115

[Two novel pathogenic mutations of GAN gene identified in a patient with giant axonal neuropathy].

PubMed

Wang, Juan; Ma, Qingwen; Cai, Qin; Liu, Yanna; Wang, Wei; Ren, Zhaorui

2016-06-01

To explore the disease-causing mutations in a patient suspected for giant axonal neuropathy(GAN). Target sequence capture sequencing was used to screen potential mutations in genomic DNA extracted from peripheral blood sample of the patient. Sanger sequencing was applied to confirm the detected mutation. The mutation was verified among 400 GAN alleles from 200 healthy individuals by Sanger sequencing. The function of the mutations was predicted by bioinformatics analysis. The patient was identified as a compound heterozygote carrying two novel pathogenic GAN mutations, i.e., c.778G>T (p.Glu260Ter) and c.277G>A (p.Gly93Arg). Sanger sequencing confirmed that the c.778G>T (p.Glu260Ter) mutation was inherited from his father, while c.277G>A (p.Gly93Arg) was inherited from his mother. The same mutations was not found in the 200 healthy individuals. Bioinformatics analysis predicted that the two mutations probably caused functional abnormality of gigaxonin. Two novel GAN mutations were detected in a patient with GAN. Both mutations are pathogenic and can cause abnormalities of gigaxonin structure and function, leading to pathogenesis of GAN. The results may also offer valuable information for similar diseases.

Partial uniparental isodisomy of chromosome 16 unmasks a deleterious biallelic mutation in IFT140 that causes Mainzer-Saldino syndrome.

PubMed

Helm, Benjamin M; Willer, Jason R; Sadeghpour, Azita; Golzio, Christelle; Crouch, Eric; Vergano, Samantha Schrier; Katsanis, Nicholas; Davis, Erica E

2017-07-19

The ciliopathies represent an umbrella group of >50 clinical entities that share both clinical features and molecular etiology underscored by structural and functional defects of the primary cilium. Despite the advances in gene discovery, this group of entities continues to pose a diagnostic challenge, in part due to significant genetic and phenotypic heterogeneity and variability. We consulted a pediatric case from asymptomatic, non-consanguineous parents who presented as a suspected ciliopathy due to a constellation of retinal, renal, and skeletal findings. Although clinical panel sequencing of genes implicated in nephrotic syndromes yielded no likely causal mutation, an oligo-SNP microarray identified a ~20-Mb region of homozygosity, with no altered gene dosage, on chromosome 16p13. Intersection of the proband's phenotypes with known disease genes within the homozygous region yielded a single candidate, IFT140, encoding a retrograde intraflagellar transport protein implicated previously in several ciliopathies, including the phenotypically overlapping Mainzer-Saldino syndrome (MZSDS). Sanger sequencing yielded a maternally inherited homozygous c.634G>A; p.Gly212Arg mutation altering the exon 6 splice donor site. Functional studies in cells from the proband showed that the locus produced two transcripts: a majority message containing a mis-splicing event that caused a premature termination codon and a minority message homozygous for the p.Gly212Arg allele. Zebrafish in vivo complementation studies of the latter transcript demonstrated a loss of function effect. Finally, we conducted post-hoc trio-based whole exome sequencing studies to (a) test the possibility of other causal loci in the proband and (b) explain the Mendelian error of segregation for the IFT140 mutation. We show that the proband harbors a chromosome 16 maternal heterodisomy, with segmental isodisomy at 16p13, likely due to a meiosis I error in the maternal gamete. Using clinical phenotyping combined with research-based genetic and functional studies, we have characterized a recurrent IFT140 mutation in the proband; together, these data are consistent with MZSDS. Additionally, we report a rare instance of a uniparental isodisomy unmasking a deleterious mutation to cause a ciliary disorder.

ALDH1A3 Mutations Cause Recessive Anophthalmia and Microphthalmia

PubMed Central

Fares-Taie, Lucas; Gerber, Sylvie; Chassaing, Nicolas; Clayton-Smith, Jill; Hanein, Sylvain; Silva, Eduardo; Serey, Margaux; Serre, Valérie; Gérard, Xavier; Baumann, Clarisse; Plessis, Ghislaine; Demeer, Bénédicte; Brétillon, Lionel; Bole, Christine; Nitschke, Patrick; Munnich, Arnold; Lyonnet, Stanislas; Calvas, Patrick; Kaplan, Josseline; Ragge, Nicola; Rozet, Jean-Michel

2013-01-01

Anophthalmia and microphthalmia (A/M) are early-eye-development anomalies resulting in absent or small ocular globes, respectively. A/M anomalies occur in syndromic or nonsyndromic forms. They are genetically heterogeneous, some mutations in some genes being responsible for both anophthalmia and microphthalmia. Using a combination of homozygosity mapping, exome sequencing, and Sanger sequencing, we identified homozygosity for one splice-site and two missense mutations in the gene encoding the A3 isoform of the aldehyde dehydrogenase 1 (ALDH1A3) in three consanguineous families segregating A/M with occasional orbital cystic, neurological, and cardiac anomalies. ALDH1A3 is a key enzyme in the formation of a retinoic acid gradient along the dorso-ventral axis during early eye development. Transitory expression of mutant ALDH1A3 open reading frames showed that both missense mutations reduce the accumulation of the enzyme, potentially leading to altered retinoic acid synthesis. Although the role of retinoic acid signaling in eye development is well established, our findings provide genetic evidence of a direct link between retinoic-acid-synthesis dysfunction and early-eye-development anomalies in humans. PMID:23312594

Integrative Analysis of PRKAG2 Cardiomyopathy iPS and Microtissue Models Identifies AMPK as a Regulator of Metabolism, Survival, and Fibrosis.

PubMed

Hinson, J Travis; Chopra, Anant; Lowe, Andre; Sheng, Calvin C; Gupta, Rajat M; Kuppusamy, Rajarajan; O'Sullivan, John; Rowe, Glenn; Wakimoto, Hiroko; Gorham, Joshua; Burke, Michael A; Zhang, Kehan; Musunuru, Kiran; Gerszten, Robert E; Wu, Sean M; Chen, Christopher S; Seidman, Jonathan G; Seidman, Christine E

2016-12-20

AMP-activated protein kinase (AMPK) is a metabolic enzyme that can be activated by nutrient stress or genetic mutations. Missense mutations in the regulatory subunit, PRKAG2, activate AMPK and cause left ventricular hypertrophy, glycogen accumulation, and ventricular pre-excitation. Using human iPS cell models combined with three-dimensional cardiac microtissues, we show that activating PRKAG2 mutations increase microtissue twitch force by enhancing myocyte survival. Integrating RNA sequencing with metabolomics, PRKAG2 mutations that activate AMPK remodeled global metabolism by regulating RNA transcripts to favor glycogen storage and oxidative metabolism instead of glycolysis. As in patients with PRKAG2 cardiomyopathy, iPS cell and mouse models are protected from cardiac fibrosis, and we define a crosstalk between AMPK and post-transcriptional regulation of TGFβ isoform signaling that has implications in fibrotic forms of cardiomyopathy. Our results establish critical connections among metabolic sensing, myocyte survival, and TGFβ signaling. Copyright © 2016 The Author(s). Published by Elsevier Inc. All rights reserved.

KRAS oncogene in non-small cell lung cancer: clinical perspectives on the treatment of an old target.

PubMed

Román, Marta; Baraibar, Iosune; López, Inés; Nadal, Ernest; Rolfo, Christian; Vicent, Silvestre; Gil-Bazo, Ignacio

2018-02-19

Lung neoplasms are the leading cause of death by cancer worldwide. Non-small cell lung cancer (NSCLC) constitutes more than 80% of all lung malignancies and the majority of patients present advanced disease at onset. However, in the last decade, multiple oncogenic driver alterations have been discovered and each of them represents a potential therapeutic target. Although KRAS mutations are the most frequently oncogene aberrations in lung adenocarcinoma patients, effective therapies targeting KRAS have yet to be developed. Moreover, the role of KRAS oncogene in NSCLC remains unclear and its predictive and prognostic impact remains controversial. The study of the underlying biology of KRAS in NSCLC patients could help to determine potential candidates to evaluate novel targeted agents and combinations that may allow a tailored treatment for these patients. The aim of this review is to update the current knowledge about KRAS-mutated lung adenocarcinoma, including a historical overview, the biology of the molecular pathways involved, the clinical relevance of KRAS mutations as a prognostic and predictive marker and the potential therapeutic approaches for a personalized treatment of KRAS-mutated NSCLC patients.

Screening of MITF and SOX10 regulatory regions in Waardenburg syndrome type 2.

PubMed

Baral, Viviane; Chaoui, Asma; Watanabe, Yuli; Goossens, Michel; Attie-Bitach, Tania; Marlin, Sandrine; Pingault, Veronique; Bondurand, Nadege

2012-01-01

Waardenburg syndrome (WS) is a rare auditory-pigmentary disorder that exhibits varying combinations of sensorineural hearing loss and pigmentation defects. Four subtypes are clinically defined based on the presence or absence of additional symptoms. WS type 2 (WS2) can result from mutations within the MITF or SOX10 genes; however, 70% of WS2 cases remain unexplained at the molecular level, suggesting that other genes might be involved and/or that mutations within the known genes escaped previous screenings. The recent identification of a deletion encompassing three of the SOX10 regulatory elements in a patient presenting with another WS subtype, WS4, defined by its association with Hirschsprung disease, led us to search for deletions and point mutations within the MITF and SOX10 regulatory elements in 28 yet unexplained WS2 cases. Two nucleotide variations were identified: one in close proximity to the MITF distal enhancer (MDE) and one within the U1 SOX10 enhancer. Functional analyses argued against a pathogenic effect of these variations, suggesting that mutations within regulatory elements of WS genes are not a major cause of this neurocristopathy.

DFG-out Mode of Inhibition by an Irreversible Type-1 Inhibitor Capable of Overcoming Gate-Keeper Mutations in FGF Receptors

PubMed Central

2015-01-01

Drug-resistance acquisition through kinase gate-keeper mutations is a major hurdle in the clinic. Here, we determined the first crystal structures of the human FGFR4 kinase domain (FGFR4K) alone and complexed with ponatinib, a promiscuous type-2 (DFG-out) kinase inhibitor, and an oncogenic FGFR4K harboring the V550L gate-keeper mutation bound to FIIN-2, a new type-1 irreversible inhibitor. Remarkably, like ponatinib, FIIN-2 also binds in the DFG-out mode despite lacking a functional group necessary to occupy the pocket vacated upon the DFG-out flip. Structural analysis reveals that the covalent bond between FIIN-2 and a cysteine, uniquely present in the glycine-rich loop of FGFR kinases, facilitates the DFG-out conformation, which together with the internal flexibility of FIIN-2 enables FIIN-2 to avoid the steric clash with the gate-keeper mutation that causes the ponatinib resistance. The structural data provide a blueprint for the development of next generation anticancer inhibitors through combining the salient inhibitory mechanisms of ponatinib and FIIN-2. PMID:25317566

Mutations in ZMYND10, a Gene Essential for Proper Axonemal Assembly of Inner and Outer Dynein Arms in Humans and Flies, Cause Primary Ciliary Dyskinesia

PubMed Central

Moore, Daniel J.; Onoufriadis, Alexandros; Shoemark, Amelia; Simpson, Michael A.; zur Lage, Petra I.; de Castro, Sandra C.; Bartoloni, Lucia; Gallone, Giuseppe; Petridi, Stavroula; Woollard, Wesley J.; Antony, Dinu; Schmidts, Miriam; Didonna, Teresa; Makrythanasis, Periklis; Bevillard, Jeremy; Mongan, Nigel P.; Djakow, Jana; Pals, Gerard; Lucas, Jane S.; Marthin, June K.; Nielsen, Kim G.; Santoni, Federico; Guipponi, Michel; Hogg, Claire; Antonarakis, Stylianos E.; Emes, Richard D.; Chung, Eddie M.K.; Greene, Nicholas D.E.; Blouin, Jean-Louis; Jarman, Andrew P.; Mitchison, Hannah M.

2013-01-01

Primary ciliary dyskinesia (PCD) is a ciliopathy characterized by airway disease, infertility, and laterality defects, often caused by dual loss of the inner dynein arms (IDAs) and outer dynein arms (ODAs), which power cilia and flagella beating. Using whole-exome and candidate-gene Sanger resequencing in PCD-affected families afflicted with combined IDA and ODA defects, we found that 6/38 (16%) carried biallelic mutations in the conserved zinc-finger gene BLU (ZMYND10). ZMYND10 mutations conferred dynein-arm loss seen at the ultrastructural and immunofluorescence level and complete cilia immotility, except in hypomorphic p.Val16Gly (c.47T>G) homozygote individuals, whose cilia retained a stiff and slowed beat. In mice, Zmynd10 mRNA is restricted to regions containing motile cilia. In a Drosophila model of PCD, Zmynd10 is exclusively expressed in cells with motile cilia: chordotonal sensory neurons and sperm. In these cells, P-element-mediated gene silencing caused IDA and ODA defects, proprioception deficits, and sterility due to immotile sperm. Drosophila Zmynd10 with an equivalent c.47T>G (p.Val16Gly) missense change rescued mutant male sterility less than the wild-type did. Tagged Drosophila ZMYND10 is localized primarily to the cytoplasm, and human ZMYND10 interacts with LRRC6, another cytoplasmically localized protein altered in PCD. Using a fly model of PCD, we conclude that ZMYND10 is a cytoplasmic protein required for IDA and ODA assembly and that its variants cause ciliary dysmotility and PCD with laterality defects. PMID:23891471

HIBCH mutations can cause Leigh-like disease with combined deficiency of multiple mitochondrial respiratory chain enzymes and pyruvate dehydrogenase.

PubMed

Ferdinandusse, Sacha; Waterham, Hans R; Heales, Simon J R; Brown, Garry K; Hargreaves, Iain P; Taanman, Jan-Willem; Gunny, Roxana; Abulhoul, Lara; Wanders, Ronald J A; Clayton, Peter T; Leonard, James V; Rahman, Shamima

2013-12-04

Deficiency of 3-hydroxy-isobutyryl-CoA hydrolase (HIBCH) caused by HIBCH mutations is a rare cerebral organic aciduria caused by disturbance of valine catabolism. Multiple mitochondrial respiratory chain (RC) enzyme deficiencies can arise from a number of mechanisms, including defective maintenance or expression of mitochondrial DNA. Impaired biosynthesis of iron-sulphur clusters and lipoic acid can lead to pyruvate dehydrogenase complex (PDHc) deficiency in addition to multiple RC deficiencies, known as the multiple mitochondrial dysfunctions syndrome. Two brothers born to distantly related Pakistani parents presenting in early infancy with a progressive neurodegenerative disorder, associated with basal ganglia changes on brain magnetic resonance imaging, were investigated for suspected Leigh-like mitochondrial disease. The index case had deficiencies of multiple RC enzymes and PDHc in skeletal muscle and fibroblasts respectively, but these were normal in his younger brother. The observation of persistently elevated hydroxy-C4-carnitine levels in the younger brother led to suspicion of HIBCH deficiency, which was investigated by biochemical assay in cultured skin fibroblasts and molecular genetic analysis. Specific spectrophotometric enzyme assay revealed HIBCH activity to be below detectable limits in cultured skin fibroblasts from both brothers. Direct Sanger sequence analysis demonstrated a novel homozygous pathogenic missense mutation c.950G

The role of p53 in combination radioimmunotherapy with 64Cu-DOTA-cetuximab and cisplatin in a mouse model of colorectal cancer.

PubMed

Guo, Yunjun; Parry, Jesse J; Laforest, Richard; Rogers, Buck E; Anderson, Carolyn J

2013-09-01

Radioimmunotherapy has been successfully used in the treatment of lymphoma but thus far has not demonstrated significant efficacy in humans beyond disease stabilization in solid tumors. Radioimmunotherapy with (64)Cu was highly effective in a hamster model of colorectal cancer, but targeted radiotherapies with this radionuclide have since not shown as much success. It is widely known that mutations in key proteins play a role in the success or failure of cancer therapies. For example, the KRAS mutation is predictive of poor response to anti-epidermal growth factor receptor therapies in colorectal cancer, whereas p53 is frequently mutated in tumors, causing resistance to multiple therapeutic regimens. We previously showed that nuclear localization of (64)Cu-labeled DOTA-cetuximab was enhanced in p53 wild-type tumor cells. Here, we examine the role of p53 in the response to radioimmunotherapy with (64)Cu-DOTA-cetuximab in KRAS-mutated HCT116 tumor-bearing mice, with and without cisplatin, which upregulates wild-type p53. Experiments with HCT116 cells that are p53 +/+ (p53 wild-type) and -/- (p53 null) grown in cell culture demonstrated that preincubation with cisplatin increased expression of p53 and subsequently enhanced localization of (64)Cu from (64)Cu-acetate and (64)Cu-DOTA-cetuximab to the tumor cell nuclei. Radioimmunotherapy studies in p53-positive HCT116 tumor-bearing mice, receiving either radioimmunotherapy alone or in combination with cisplatin, showed significantly longer survival in mice receiving unlabeled cetuximab or cisplatin alone or in combination (all, P < 0.01). In contrast, the p53-negative tumor-bearing mice treated with radioimmunotherapy alone or combined with cisplatin showed no survival advantage, compared with control groups (all, P > 0.05). Together, these data suggest that (64)Cu specifically delivered to epidermal growth factor receptor-positive tumors by cetuximab can suppress tumor growth despite the KRAS status and present opportunities for personalized clinical treatment strategies in colorectal cancer.

A new common mutation in the cardiac beta-myosin heavy chain gene in Finnish patients with hypertrophic cardiomyopathy.

PubMed

Jääskeläinen, Pertti; Heliö, Tiina; Aalto-Setälä, Katriina; Kaartinen, Maija; Ilveskoski, Erkki; Hämäläinen, Liisa; Melin, John; Kärkkäinen, Satu; Peuhkurinen, Keijo; Nieminen, Markku S; Laakso, Markku; Kuusisto, Johanna

2014-09-01

In the nationwide FinHCM Study including 306 Finnish patients with hypertrophic cardiomyopathy (HCM), we have previously identified two founder mutations in the alpha-tropomyosin (TPM1-D175N) and myosin-binding protein C (MYBPC3-Q1061X) genes, accounting for 18% of all cases. Objective. To screen additional mutations, previously identified in eastern Finnish cohorts with HCM, in the FinHCM Study population. Ten mutations in the beta-myosin heavy chain gene (MYH7), TPM1, and MYBPC3 were screened. MYH7-R1053Q was found in 17 of 306 patients (5.6%). No carriers of MYH7-R719W or N696S were found. A novel TPM1-D175G mutation was found in a single patient. MYBPC3 mutations were found in 14 patients: IVS5-2A-C in two, IVS14-13G-A in two, K811del in six, and A851insT in four patients. Altogether, a HCM-causing mutation was identified in 32 patients, accounting for 10.5% of all cases. In addition, two MYBPC3 variants R326Q and V896M with uncertain pathogenicity were found in eight and in 10 patients, respectively. Combining the present findings with our previous results, a causative mutation was identified in 28% of the FinHCM cohort. MYH7-R1053Q was the third most common mutation, and should be screened in all new cases of HCM in Finland.

Structural study of the effects of mutations in proteins to identify the molecular basis of the loss of local structural fluidity leading to the onset of autoimmune diseases.

PubMed

Ali, Ananya; Ghosh, Semanti; Bagchi, Angshuman

2017-02-26

Protein-Protein Interactions (PPIs) are crucial in most of the biological processes and PPI dysfunctions are known to be associated with the onsets of various diseases. One of such diseases is the auto-immune disease. Auto-immune diseases are one among the less studied group of diseases with very high mortality rates. Thus, we tried to correlate the appearances of mutations with their probable biochemical basis of the molecular mechanisms leading to the onset of the disease phenotypes. We compared the effects of the Single Amino Acid Variants (SAVs) in the wild type and mutated proteins to identify any structural deformities that might lead to altered PPIs leading ultimately to disease onset. For this we used Relative Solvent Accessibility (RSA) as a spatial parameter to compare the structural perturbation in mutated and wild type proteins. We observed that the mutations were capable to increase intra-chain PPIs whereas inter-chain PPIs would remain mostly unaltered. This might lead to more intra-molecular friction causing a deleterious alteration of protein's normal function. A Lyapunov exponent analysis, using the altered RSA values due to polymorphic and disease causing mutations, revealed polymorphic mutations have a positive mean value for the Lyapunov exponent while disease causing mutations have a negative mean value. Thus, local spatial stochasticity has been lost due to disease causing mutations, indicating a loss of structural fluidity. The amino acid conversion plot also showed a clear tendency of altered surface patch residue conversion propensity than polymorphic conversions. So far, this is the first report that compares the effects of different kinds of mutations (disease and non-disease causing polymorphic mutations) in the onset of autoimmune diseases. Copyright © 2017 Elsevier Inc. All rights reserved.

Heterozygous ABCC8 mutations are a cause of MODY.

PubMed

Bowman, P; Flanagan, S E; Edghill, E L; Damhuis, A; Shepherd, M H; Paisey, R; Hattersley, A T; Ellard, S

2012-01-01

The ABCC8 gene encodes the sulfonylurea receptor 1 (SUR1) subunit of the pancreatic beta cell ATP-sensitive potassium (K(ATP)) channel. Inactivating mutations cause congenital hyperinsulinism (CHI) and activating mutations cause transient neonatal diabetes (TNDM) or permanent neonatal diabetes (PNDM) that can usually be treated with sulfonylureas. Sulfonylurea sensitivity is also a feature of HNF1A and HNF4A MODY, but patients referred for genetic testing with clinical features of these types of diabetes do not always have mutations in the HNF1A/4A genes. Our aim was to establish whether mutations in the ABCC8 gene cause MODY that is responsive to sulfonylurea therapy. We sequenced the ABCC8 gene in 85 patients with a BMI <30 kg/m², no family history of neonatal diabetes and who were deemed sensitive to sulfonylureas by the referring clinician or were sulfonylurea-treated. All had tested negative for mutations in the HNF1A and HNF4A genes. ABCC8 mutations were found in seven of the 85 (8%) probands. Four patients were heterozygous for previously reported mutations and four novel mutations, E100K, G214R, Q485R and N1245D, were identified. Only four probands fulfilled MODY criteria, with two diagnosed after 25 years and one patient, who had no family history of diabetes, as a result of a proven de novo mutation. ABCC8 mutations can cause MODY in patients whose clinical features are similar to those with HNF1A/4A MODY. Therefore, sequencing of ABCC8 in addition to the known MODY genes should be considered if such features are present, to facilitate optimal clinical management of these patients.

Observation of c.260A > G mutation in superoxide dismutase 1 that causes p.Asn86Ser in Iranian amyotrophic lateral sclerosis patient and absence of genotype/phenotype correlation.

PubMed

Khani, Marzieh; Alavi, Afagh; Nafissi, Shahriar; Elahi, Elahe

2015-07-06

Amyotrophic lateral sclerosis (ALS) is the most common motor neuron disorder in European populations. ALS can be sporadic ALS (SALS) or familial ALS (FALS). Among 20 known ALS genes, mutations in C9orf72 and superoxide dismutase 1 (SOD1) are the most common genetic causes of the disease. Whereas C9orf72 mutations are more common in Western populations, the contribution of SOD1 to ALS in Iran is more than C9orf72. At present, a clear genotype/phenotype correlation for ALS has not been identified. We aimed to perform mutation screening of SOD1 in a newly identified Iranian FALS patient and to assess whether a genotype/phenotype correlation for the identified mutation exists. The five exons of SOD1 and flanking intronic sequences of a FALS proband were screened for mutations by direct sequencing. The clinical features of the proband were assessed by a neuromuscular specialist (SN). The phenotypic presentations were compared to previously reported patients with the same mutation. Heterozygous c.260A > G mutation in SOD1 that causes Asn86Ser was identified in the proband. Age at onset was 34 years and site of the first presentation was in the lower extremities. Comparisons of clinical features of different ALS patients with the same mutation evidenced variable presentations. The c.260A > G mutation in SOD1 that causes Asn86Ser appears to cause ALS with variable clinical presentations.

Dosage Mutator Genes in Saccharomyces cerevisiae: A Novel Mutator Mode-of-Action of the Mph1 DNA Helicase.

PubMed

Ang, J Sidney; Duffy, Supipi; Segovia, Romulo; Stirling, Peter C; Hieter, Philip

2016-11-01

Mutations that cause genome instability are considered important predisposing events that contribute to initiation and progression of cancer. Genome instability arises either due to defects in genes that cause an increased mutation rate (mutator phenotype), or defects in genes that cause chromosome instability (CIN). To extend the catalog of genome instability genes, we systematically explored the effects of gene overexpression on mutation rate, using a forward-mutation screen in budding yeast. We screened ∼5100 plasmids, each overexpressing a unique single gene, and characterized the five strongest mutators, MPH1 (mutator phenotype 1), RRM3, UBP12, PIF1, and DNA2 We show that, for MPH1, the yeast homolog of Fanconi Anemia complementation group M (FANCM), the overexpression mutator phenotype is distinct from that of mph1Δ. Moreover, while four of our top hits encode DNA helicases, the overexpression of 48 other DNA helicases did not cause a mutator phenotype, suggesting this is not a general property of helicases. For Mph1 overexpression, helicase activity was not required for the mutator phenotype; in contrast Mph1 DEAH-box function was required for hypermutation. Mutagenesis by MPH1 overexpression was independent of translesion synthesis (TLS), but was suppressed by overexpression of RAD27, a conserved flap endonuclease. We propose that binding of DNA flap structures by excess Mph1 may block Rad27 action, creating a mutator phenotype that phenocopies rad27Δ. We believe this represents a novel mutator mode-of-action and opens up new prospects to understand how upregulation of DNA repair proteins may contribute to mutagenesis. Copyright © 2016 by the Genetics Society of America.

In the Thick of It: HCM-Causing Mutations in Myosin Binding Proteins of the Thick Filament

PubMed Central

Harris, Samantha P.; Lyons, Ross G.; Bezold, Kristina L.

2010-01-01

In the 20 yrs since the discovery of the first mutation linked to familial hypertrophic cardiomyopathy (HCM) an astonishing number of mutations affecting numerous sarcomeric proteins have been described. Among the most prevalent of these are mutations that affect thick filament binding proteins including the myosin essential and regulatory light chains and cardiac myosin binding protein-C (cMyBP-C). However, despite the frequency with which myosin binding proteins, especially cMyBP-C, have been linked to inherited cardiomyopathies, the functional consequences of mutations in these proteins and the mechanisms by which they cause disease are still only partly understood. The purpose of this review is to summarize the known disease-causing mutations that affect the major thick filament binding proteins and to relate these mutations to protein function. Conclusions emphasize the impact that discovery of HCM causing mutations has had on fueling insights into the basic biology of thick filament proteins and reinforce the idea that myosin binding proteins are dynamic regulators of the activation state of the thick filament that contribute to the speed and force of myosin driven muscle contraction. Additional work is still needed to determine the mechanisms by which individual mutations induce hypertrophic phenotypes. PMID:21415409

Induced mutations in mice and genetic risk assessment in humans

DOE Office of Scientific and Technical Information (OSTI.GOV)

Selby, P.B.

1980-01-01

In studies on mice, in contrast to studies on humans, it is possible to perform carefully controlled experiments with the exposures one desires. The necessity for having separate mammalian tests for looking at the induction of gene mutations and small deficiencies, and at the induction of chromosomal aberrations, is obvious. Mutagens can differ as to which of these types of damage they are more likely to cause. The reason for focusing attention on the mouse in a discussion of hazard from induced gene mutations and small deficiencies is the existence of techniques in this mammal for readily studying the inductionmore » of such genetic effects. Many mutations at the molecular level cause no apparent changes at the gene-product level and many mutations that cause changes at the gene-product level cause no detectable phenotypic changes in heterozygotes. Many dominant mutations that change the phenotype cause no serious handicap. For these reasons, risk estimation for important chemicals must rely heavily on studies on the induction of those germinal mutations in mammals that are easily related to human dominant disorders, such as skeletal and cataract mutations. Molecular or enzyme studies cannot provide definitive answers about risk. The specific-locus method should help greatly in assessing the genetic risks to humans from chemicals. The new sensitive-indicator method should complement it in providing a tool for attacking the question of what treatments induce gene mutations and small deficiencies and for approximating first-generation damage to the skeleton. (ERB)« less

Isocitrate dehydrogenase 1 R132C mutation occurs exclusively in microsatellite stable colorectal cancers with the CpG island methylator phenotype.

PubMed

Whitehall, V L J; Dumenil, T D; McKeone, D M; Bond, C E; Bettington, M L; Buttenshaw, R L; Bowdler, L; Montgomery, G W; Wockner, L F; Leggett, B A

2014-11-01

The CpG Island Methylator Phenotype (CIMP) is fundamental to an important subset of colorectal cancer; however, its cause is unknown. CIMP is associated with microsatellite instability but is also found in BRAF mutant microsatellite stable cancers that are associated with poor prognosis. The isocitrate dehydrogenase 1 (IDH1) gene causes CIMP in glioma due to an activating mutation that produces the 2-hydroxyglutarate oncometabolite. We therefore examined IDH1 alteration as a potential cause of CIMP in colorectal cancer. The IDH1 mutational hotspot was screened in 86 CIMP-positive and 80 CIMP-negative cancers. The entire coding sequence was examined in 81 CIMP-positive colorectal cancers. Forty-seven cancers varying by CIMP-status and IDH1 mutation status were examined using Illumina 450K DNA methylation microarrays. The R132C IDH1 mutation was detected in 4/166 cancers. All IDH1 mutations were in CIMP cancers that were BRAF mutant and microsatellite stable (4/45, 8.9%). Unsupervised hierarchical cluster analysis identified an IDH1 mutation-like methylation signature in approximately half of the CIMP-positive cancers. IDH1 mutation appears to cause CIMP in a small proportion of BRAF mutant, microsatellite stable colorectal cancers. This study provides a precedent that a single gene mutation may cause CIMP in colorectal cancer, and that this will be associated with a specific epigenetic signature and clinicopathological features.

Two new mutations in the 3' coding region of the glycogen debranching enzyme in a glycogen storage disease type IIIa Ashkenazi Jewish patient.

PubMed

Parvari, R; Shen, J; Hershkovitz, E; Chen, Y T; Moses, S W

1998-04-01

Glycogen storage disease type III (GSD III) is an autosomal recessive disease caused by the deficiency of glycogen debranching enzyme (AGL). We report the finding of two new mutations in a GSD IIIa Ashkenazi Jewish patient. Both mutations are insertion of an adenine into a stretch of 8 adenines towards the 3' end of the coding region, one at position 3904 (3904insA) in exon 30, the second at position 4214 (4214insA) in exon 32. The mutations cause frameshifts and premature terminations of the glycogen debranching enzyme, the first causing a frameshift at amino acid 1304, the second causing a frameshift at amino acid 1408 of the total of 1532. These mutations demonstrate the importance of the 125 amino acids at the carboxy-terminus of the debrancher enzyme for its activity and support the suggestion that the putative glycogen binding domain is located in the carboxy-terminus of the AGL. The mutations cause distinctive single-strand conformation polymorphism (SSCP) patterns enabling easy detection.

Cutaneous squamous and neuroendocrine carcinoma: genetically and immunohistochemically different from Merkel cell carcinoma

PubMed Central

Pulitzer, Melissa P; Brannon, A Rose; Berger, Michael F; Louis, Peter; Scott, Sasinya N; Jungbluth, Achim A; Coit, Daniel G; Brownell, Isaac; Busam, Klaus J

2016-01-01

Cutaneous neuroendocrine (Merkel cell) carcinoma most often arises de novo in the background of a clonally integrated virus, the Merkel cell polyomavirus, and is notable for positive expression of retinoblastoma 1 (RB1) protein and low expression of p53 compared with the rare Merkel cell polyomavirus-negative Merkel cell carcinomas. Combined squamous and Merkel cell tumors are consistently negative for Merkel cell polyomavirus. Little is known about their immunophenotypic or molecular profile. Herein, we studied 10 combined cutaneous squamous cell and neuroendocrine carcinomas for immunohistochemical expression of p53, retinoblastoma 1 protein, neurofilament, p63, and cytokeratin 20 (CK20). We compared mutation profiles of five combined Merkel cell carcinomas and seven ‘pure’ Merkel cell carcinomas using targeted next-generation sequencing. Combined tumors were from the head, trunk, and leg of Caucasian males and one female aged 52–89. All cases were highly p53- and p63-positive and neurofilament-negative in the squamous component, whereas RB1-negative in both components. Eight out of 10 were p53-positive, 3/10 p63-positive, and 3/10 focally neurofilament-positive in the neuroendocrine component. Six out of 10 were CK20-positive in any part. By next-generation sequencing, combined tumors were highly mutated, with an average of 48 mutations per megabase compared with pure tumors, which showed 1.25 mutations per megabase. RB1 and p53 mutations were identified in all five combined tumors. Combined tumors represent an immunophenotypically and genetically distinct variant of primary cutaneous neuroendocrine carcinomas, notable for a highly mutated genetic profile, significant p53 expression and/or mutation, absent RB1 expression in the context of increased RB1 mutation, and minimal neurofilament expression. PMID:26022453

Cutaneous squamous and neuroendocrine carcinoma: genetically and immunohistochemically different from Merkel cell carcinoma.

PubMed

Pulitzer, Melissa P; Brannon, A Rose; Berger, Michael F; Louis, Peter; Scott, Sasinya N; Jungbluth, Achim A; Coit, Daniel G; Brownell, Isaac; Busam, Klaus J

2015-08-01

Cutaneous neuroendocrine (Merkel cell) carcinoma most often arises de novo in the background of a clonally integrated virus, the Merkel cell polyomavirus, and is notable for positive expression of retinoblastoma 1 (RB1) protein and low expression of p53 compared with the rare Merkel cell polyomavirus-negative Merkel cell carcinomas. Combined squamous and Merkel cell tumors are consistently negative for Merkel cell polyomavirus. Little is known about their immunophenotypic or molecular profile. Herein, we studied 10 combined cutaneous squamous cell and neuroendocrine carcinomas for immunohistochemical expression of p53, retinoblastoma 1 protein, neurofilament, p63, and cytokeratin 20 (CK20). We compared mutation profiles of five combined Merkel cell carcinomas and seven 'pure' Merkel cell carcinomas using targeted next-generation sequencing. Combined tumors were from the head, trunk, and leg of Caucasian males and one female aged 52-89. All cases were highly p53- and p63-positive and neurofilament-negative in the squamous component, whereas RB1-negative in both components. Eight out of 10 were p53-positive, 3/10 p63-positive, and 3/10 focally neurofilament-positive in the neuroendocrine component. Six out of 10 were CK20-positive in any part. By next-generation sequencing, combined tumors were highly mutated, with an average of 48 mutations per megabase compared with pure tumors, which showed 1.25 mutations per megabase. RB1 and p53 mutations were identified in all five combined tumors. Combined tumors represent an immunophenotypically and genetically distinct variant of primary cutaneous neuroendocrine carcinomas, notable for a highly mutated genetic profile, significant p53 expression and/or mutation, absent RB1 expression in the context of increased RB1 mutation, and minimal neurofilament expression.

Hearing loss caused by a P2RX2 mutation identified in a MELAS family with a coexisting mitochondrial 3243AG mutation

PubMed Central

Moteki, Hideaki; Azaiez, Hela; Booth, Kevin T; Hattori, Mitsuru; Sato, Ai; Sato, Yoshihiko; Motobayashi, Mitsuo; Sloan, Christina M; Kolbe, Diana L; Shearer, A Eliot; Smith, Richard J H; Usami, Shin-ichi

2015-01-01

Objective We present a family with a mitochondrial DNA 3243A>G mutation resulting in MELAS, of which some members have hearing loss where a novel mutation in the P2RX2 gene was identified. Methods One hundred ninety-four (194) Japanese subjects from unrelated families were enrolled in the study. Targeted genomic enrichment and massively parallel sequencing of all known non-syndromic hearing loss genes were performed to identify the genetic causes of hearing loss. Results A novel mutation in the P2RX2 gene, that corresponded to c.601G>A (p.Asp201Tyr) was identified. Two patients carried the mutation, and had severe SNHL, while other members with MELAS (who did not carry the P2RX2 mutation) had normal hearing. Conclusion This is the first case report of a diagnosis of hearing loss caused by P2RX2 mutation in patients with MELAS. A potential explanation is that decreasing ATP production due to MELAS with mitochondrial 3243A>G mutation might suppress activation of P2X2 receptors. We also suggest that hearing loss caused by the P2RX2 mutation might be influenced by the decrease in ATP production due to MELAS, and that nuclear genetic factors may play a modifying role in mitochondrial dysfunction. PMID:25788561

Modification of mutagenesis initiated by ultraviolet light through posttreatment of bacteria with basic dyes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Witkin, E M

1961-12-01

Post treatment with basic dyes, in concentrations that retard cell division, was found to influence the tnduction of mutations to prototrophy by UV light in a tyrosine-requirtng strain of E.Coli. Pyronin, which is unique among the dyes in tts selective affinity for RNA, was found to duplicate the effects of chloramphenicol or amino acfd deprtvatfon in causfng the rapid and irreversible loss of potential prototrophs (mutation frequency decline, or MFD). Acriflavtne, methyl green. crystal violet, methylene blue, and toluidine blue, all of which are known to combine with DNA, delay or retard the occurrence of MFD under conditions of aminomore » acid deprivation. When acriflavine is removed from its combination with cellular components by the addition of an excess of sodium deoxyrtbonucleate, MFD begins promptly. The same basic dyes that delay MFD were also found to interfere with the fixation of mutations (MF) in an amino acid- enriched medium, and to cause marked enhancement of the mutagenic potency of low doses of UV light. While showing no independent mutagenic activity for unirradiated bacteria, all the dyes except pyronin increased the yield of induced mutations signtficantly when added to the enriched medium upon which trradiated bacteria were incubated.These results were interpreted as evidence that UV light initiates mutagenesis by producing unstable changas directly in genic DNA. MFD is interpreted as a repair process, blocked by the machinery of RNA and protein synthesis and by the presence of certain basic dyes.« less

ETFDH mutations as a major cause of riboflavin-responsive multiple acyl-CoA dehydrogenation deficiency.

PubMed

Olsen, Rikke K J; Olpin, Simon E; Andresen, Brage S; Miedzybrodzka, Zofia H; Pourfarzam, Morteza; Merinero, Begoña; Frerman, Frank E; Beresford, Michael W; Dean, John C S; Cornelius, Nanna; Andersen, Oluf; Oldfors, Anders; Holme, Elisabeth; Gregersen, Niels; Turnbull, Douglass M; Morris, Andrew A M

2007-08-01

Multiple acyl-CoA dehydrogenation deficiency (MADD) is a disorder of fatty acid, amino acid and choline metabolism that can result from defects in two flavoproteins, electron transfer flavoprotein (ETF) or ETF: ubiquinone oxidoreductase (ETF:QO). Some patients respond to pharmacological doses of riboflavin. It is unknown whether these patients have defects in the flavoproteins themselves or defects in the formation of the cofactor, FAD, from riboflavin. We report 15 patients from 11 pedigrees. All the index cases presented with encephalopathy or muscle weakness or a combination of these symptoms; several had previously suffered cyclical vomiting. Urine organic acid and plasma acyl-carnitine profiles indicated MADD. Clinical and biochemical parameters were either totally or partly corrected after riboflavin treatment. All patients had mutations in the gene for ETF:QO. In one patient, we show that the ETF:QO mutations are associated with a riboflavin-sensitive impairment of ETF:QO activity. This patient also had partial deficiencies of flavin-dependent acyl-CoA dehydrogenases and respiratory chain complexes, most of which were restored to control levels after riboflavin treatment. Low activities of mitochondrial flavoproteins or respiratory chain complexes have been reported previously in two of our patients with ETF:QO mutations. We postulate that riboflavin-responsive MADD may result from defects of ETF:QO combined with general mitochondrial dysfunction. This is the largest collection of riboflavin-responsive MADD patients ever reported, and the first demonstration of the molecular genetic basis for the disorder.

Loss of circadian clock accelerates aging in neurodegeneration-prone mutants

PubMed Central

Krishnan, Natraj; Rakshit, Kuntol; Chow, Eileen S.; Wentzell, Jill S.; Kretzschmar, Doris; Giebultowicz, Jadwiga M.

2012-01-01

Circadian clocks generate rhythms in molecular, cellular, physiological, and behavioral processes. Recent studies suggest that disruption of the clock mechanism accelerates organismal senescence and age-related pathologies in mammals. Impaired circadian rhythms are observed in many neurological diseases; however, it is not clear whether loss of rhythms is the cause or result of neurodegeneration, or both. To address this important question, we examined the effects of circadian disruption in Drosophila melanogaster mutants that display clock-unrelated neurodegenerative phenotypes. We combined a null mutation in the clock gene period (per01) that abolishes circadian rhythms, with a hypomorphic mutation in the carbonyl reductase gene sniffer (sni1), which displays oxidative stress induced neurodegeneration. We report that disruption of circadian rhythms in sni1 mutants significantly reduces their lifespan compared to single mutants. Shortened lifespan in double mutants was coupled with accelerated neuronal degeneration evidenced by vacuolization in the adult brain. In addition, per01 sni1 flies showed drastically impaired vertical mobility and increased accumulation of carbonylated proteins compared to age-matched single mutant flies. Loss of per function does not affect sni mRNA expression, suggesting that these genes act via independent pathways producing additive effects. Finally, we show that per01 mutation accelerates the onset of brain pathologies when combined with neurodegeneration-prone mutation in another gene, swiss cheese (sws1), which does not operate through the oxidative stress pathway. Taken together, our data suggest that the period gene may be causally involved in neuroprotective pathways in aging Drosophila. PMID:22227001

Long-term follow-up of combined pituitary hormone deficiency in two siblings with a Prophet of Pit-1 gene mutation.

PubMed

Georgopoulos, Neoklis A; Katsikis, Ilias; Giamalis, Petros; Koika, Vasiliki; Adonakis, George; Kourtis, Anargyros; Kourounis, George; Panidis, Dimitrios

2006-12-01

Combined pituitary hormone deficiency (CPHD) is a rare disorder resulting from an impaired pituitary function due to different causes, characterized by impaired secretion of growth hormone (GH) and one or more of the other anterior pituitary hormones. To date, 16 distinct human Prophet of Pit-1 (Prop1) gene mutations have been identified in patients with CPHD, inducing a phenotype involving GH, follicle-stimulating hormone (FSH), luteinizing hormone (LH), prolactin and thyroid-stimulating hormone (TSH), and rarely adrenocorticotropic hormone, deficiency. Herein we present two siblings of different sexes from a family with parental consanguinity presenting the 301-302delAG mutation in the Prop1 gene. The female presented failure of growth from the age of 6 years and was treated for 10 years with GH, ending in a final height (standard deviation score) of -0.28. TSH deficiency was manifested after the initiation of GH and was treated with thyroxine while puberty was initiated with conjugated estrogens. The male presented TSH deficiency since childhood, treated with thyroxine, and growth failure at the age of 14 years, treated for a period of 2 years with GH. Puberty was initiated with increasing doses of testosterone, while human chorionic gonadotropin was added in order to achieve increased testicular volume. In conclusion, these two siblings of different sexes with CPHD carrying the 301-302delAG mutation in the Prop1 gene presented a variable phenotype characterized by GH, TSH, LH and FSH deficiency.

A novel missense NDP mutation [p.(Cys93Arg)] with a manifesting carrier in an austrian family with Norrie disease.

PubMed

Parzefall, Thomas; Lucas, Trevor; Ritter, Markus; Ludwig, Martin; Ramsebner, Reinhard; Frohne, Alexandra; Schöfer, Christian; Hengstschläger, Markus; Frei, Klemens

2014-01-01

Norrie disease is a rare, X-linked genetic syndrome characterized by combined congenital blindness and progressive hearing impairment. Norrie disease is caused by alterations in the NDP gene encoding the growth factor norrin that plays a key role in vascular development and stabilization of the eye, inner ear and brain. We identified a family with 3 affected deafblind males and a single female carrier presenting with a serous retinal detachment but normal hearing. Genetic analysis revealed a novel c.277T>C missense mutation causing the substitution of a hydrophobic cysteine to a hydrophilic arginine [p.(Cys93Arg)] within the highly conserved cysteine knot domain of the norrin protein. These results should expand the scope for amniocentesis and genetic testing for Norrie disease which is gaining in importance due to novel postnatal therapeutic concepts to alleviate the devastating retinal symptoms of Norrie disease. © 2014 S. Karger AG, Basel.

Novel compound heterozygous mutations in MYO7A in a Chinese family with Usher syndrome type 1.

PubMed

Liu, Fei; Li, Pengcheng; Liu, Ying; Li, Weirong; Wong, Fulton; Du, Rong; Wang, Lei; Li, Chang; Jiang, Fagang; Tang, Zhaohui; Liu, Mugen

2013-01-01

To identify the disease-causing mutation(s) in a Chinese family with autosomal recessive Usher syndrome type 1 (USH1). An ophthalmic examination and an audiometric test were conducted to ascertain the phenotype of two affected siblings. The microsatellite marker D11S937, which is close to the candidate gene MYO7A (USH1B locus), was selected for genotyping. From the DNA of the proband, all coding exons and exon-intron boundaries of MYO7A were sequenced to identify the disease-causing mutation(s). Restriction fragment length polymorphism (RFLP) analysis was performed to exclude the alternative conclusion that the mutations are non-pathogenic rare polymorphisms. Based on severe hearing impairment, unintelligible speech, and retinitis pigmentosa, a clinical diagnosis of Usher syndrome type 1 was made. The genotyping results did not exclude the USH1B locus, which suggested that the MYO7A gene was likely the gene associated with the disease-causing mutation(s) in the family. With direct DNA sequencing of MYO7A, two novel compound heterozygous mutations (c.3742G>A and c.6051+1G>A) of MYO7A were identified in the proband. DNA sequence analysis and RFLP analysis of other family members showed that the mutations cosegregated with the disease. Unaffected members, including the parents, uncle, and sister of the proband, carry only one of the two mutations. The mutations were not present in the controls (100 normal Chinese subjects=200 chromosomes) according to the RFLP analysis. In this study, we identified two novel mutations, c.3742G>A (p.E1248K) and c.6051+1G>A (donor splice site mutation in intron 44), of MYO7A in a Chinese non-consanguineous family with USH1. The mutations cosegregated with the disease and most likely cause the phenotype in the two affected siblings who carry these mutations compound heterozygously. Our finding expands the mutational spectrum of MYO7A.

RTTN Mutations Cause Primary Microcephaly and Primordial Dwarfism in Humans

PubMed Central

Shamseldin, Hanan; Alazami, Anas M.; Manning, Melanie; Hashem, Amal; Caluseiu, Oana; Tabarki, Brahim; Esplin, Edward; Schelley, Susan; Innes, A. Micheil; Parboosingh, Jillian S.; Lamont, Ryan; Majewski, Jacek; Bernier, Francois P.; Alkuraya, Fowzan S.

2015-01-01

Primary microcephaly is a developmental brain anomaly that results from defective proliferation of neuroprogenitors in the germinal periventricular zone. More than a dozen genes are known to be mutated in autosomal-recessive primary microcephaly in isolation or in association with a more generalized growth deficiency (microcephalic primordial dwarfism), but the genetic heterogeneity is probably more extensive. In a research protocol involving autozygome mapping and exome sequencing, we recruited a multiplex consanguineous family who is affected by severe microcephalic primordial dwarfism and tested negative on clinical exome sequencing. Two candidate autozygous intervals were identified, and the second round of exome sequencing revealed a single intronic variant therein (c.2885+8A>G [p.Ser963∗] in RTTN exon 23). RT-PCR confirmed that this change creates a cryptic splice donor and thus causes retention of the intervening 7 bp of the intron and leads to premature truncation. On the basis of this finding, we reanalyzed the exome file of a second consanguineous family affected by a similar phenotype and identified another homozygous change in RTTN as the likely causal mutation. Combined linkage analysis of the two families confirmed that RTTN maps to the only significant linkage peak. Finally, through international collaboration, a Canadian multiplex family affected by microcephalic primordial dwarfism and biallelic mutation of RTTN was identified. Our results expand the phenotype of RTTN-related disorders, hitherto limited to polymicrogyria, to include microcephalic primordial dwarfism with a complex brain phenotype involving simplified gyration. PMID:26608784

The p.Cys169Tyr variant of connexin 26 is not a polymorphism

PubMed Central

Zonta, Francesco; Girotto, Giorgia; Buratto, Damiano; Crispino, Giulia; Morgan, Anna; Abdulhadi, Khalid; Alkowari, Moza; Badii, Ramin; Gasparini, Paolo; Mammano, Fabio

2015-01-01

Mutations in the GJB2 gene, which encodes the gap junction protein connexin 26 (Cx26), are the primary cause of hereditary prelingual hearing impairment. Here, the p.Cys169Tyr missense mutation of Cx26 (Cx26C169Y), previously classified as a polymorphism, has been identified as causative of severe hearing loss in two Qatari families. We have analyzed the effect of this mutation using a combination of confocal immunofluorescence microscopy and molecular dynamics simulations. At the cellular level, our results show that the mutant protein fails to form junctional channels in HeLa transfectants despite being correctly targeted to the plasma membrane. At the molecular level, this effect can be accounted for by disruption of the disulfide bridge that Cys169 forms with Cys64 in the wild-type structure (Cx26WT). The lack of the disulfide bridge in the Cx26C169Y protein causes a spatial rearrangement of two important residues, Asn176 and Thr177. In the Cx26WT protein, these residues play a crucial role in the intra-molecular interactions that permit the formation of an intercellular channel by the head-to-head docking of two opposing hemichannels resident in the plasma membrane of adjacent cells. Our results elucidate the molecular pathogenesis of hereditary hearing loss due to the connexin mutation and facilitate the understanding of its role in both healthy and affected individuals. PMID:25628337

Molecular analysis of the AGXT gene in patients suspected with hyperoxaluria type 1 and three novel mutations from Turkey.

PubMed

Isiyel, Emel; Ezgu, Sevcan A Bakkaloglu; Caliskan, Salim; Akman, Sema; Akil, Ipek; Tabel, Yilmaz; Akinci, Nurver; Ozdogan, Elif Bahat; Ozel, Ahmet; Eroglu, Fehime Kara; Ezgu, Fatih S

2016-12-01

Primary hyperoxaluria type 1 (PH1) is a rare, autosomal recessive disease, caused by the defect of AGXT gene encoding hepatic peroxisomal alanine glyoxylateaminotransferase (AGT). This enzyme is responsible for the conversion of glyoxylate to glycine. The diagnosis of PH1 should be suspected in infants and children with nephrocalcinosis or nephrolithiasis. Early diagnosis and treatment is crucial in preventing disease progression to end stage kidney disease (ESKD). In this study, AGXT gene sequence analyses were performed in 82 patients who were clinically suspected (hyperoxaluria and nephrolithiasis or nephrocalcinosis with or without renal impairment) to have PH1. Disease causing mutations have been found in fifteen patients from thirteen families (18%). Novel mutations have been found (c.458T>A (p.L153X), c.733_734delAA (p.Lys245Valfs*11), c.52 C>T (p.L18F)) in three of 13 families. There were 3-year lag time between initial symptoms and the time of PH1 is suspected; additionally, 5.5-year lag time between initial symptoms and definitive diagnosis. Consanguinity was detected in 77% of the patients with mutation. After genetic diagnosis, one patient received combined kidney and liver transplantation. AGXT gene sequencing is now the choice of diagnosis of PH1 due to its non-invasive nature compared to liver enzyme assay. Early diagnosis and accurate treatment in PH1 is important for better patient outcomes. Copyright © 2016. Published by Elsevier Inc.

Historical contingency and its biophysical basis in glucocorticoid receptor evolution.

PubMed

Harms, Michael J; Thornton, Joseph W

2014-08-14

Understanding how chance historical events shape evolutionary processes is a central goal of evolutionary biology. Direct insights into the extent and causes of evolutionary contingency have been limited to experimental systems, because it is difficult to know what happened in the deep past and to characterize other paths that evolution could have followed. Here we combine ancestral protein reconstruction, directed evolution and biophysical analysis to explore alternative 'might-have-been' trajectories during the ancient evolution of a novel protein function. We previously found that the evolution of cortisol specificity in the ancestral glucocorticoid receptor (GR) was contingent on permissive substitutions, which had no apparent effect on receptor function but were necessary for GR to tolerate the large-effect mutations that caused the shift in specificity. Here we show that alternative mutations that could have permitted the historical function-switching substitutions are extremely rare in the ensemble of genotypes accessible to the ancestral GR. In a library of thousands of variants of the ancestral protein, we recovered historical permissive substitutions but no alternative permissive genotypes. Using biophysical analysis, we found that permissive mutations must satisfy at least three physical requirements--they must stabilize specific local elements of the protein structure, maintain the correct energetic balance between functional conformations, and be compatible with the ancestral and derived structures--thus revealing why permissive mutations are rare. These findings demonstrate that GR evolution depended strongly on improbable, non-deterministic events, and this contingency arose from intrinsic biophysical properties of the protein.

R213W mutation in the retinoschisis 1 gene causes X-linked juvenile retinoschisis in a large Chinese family.

PubMed

Xu, Jun; Gu, Hong; Ma, Kai; Liu, Xipu; Snellingen, Torkel; Sun, Erdan; Wang, Ningli; Liu, Ningpu

2010-08-12

We identified a large Chinese family with X-linked juvenile retinoschisis. The purpose of this study was to report the clinical findings of the family and to identify the genetic mutation by screening the retinoschisis 1 (RS1) gene. Family history was collected and all family members underwent routine ophthalmic examination. Venous blood was collected from family members and genomic DNA was extracted. The exons of RS1 were screened by PCR followed by direct sequencing and/or restriction enzyme digestion. The pedigree of interest was a four-generation family with 52 family members, including seven affected individuals. The proband was a 5-year-old boy showing highly elevated bullous retinoschisis with moderate vitreous hemorrhage in both eyes. Vitrectomy was performed in the left eye of the proband. Five affected males showed large peripheral retinoschisis in both eyes, either involving the macula or combined with foveal stellate cystic change. One of the affected family members showed only a foveal stellate cystic change in both eyes without periphery retinoschisis. Visual acuity of affected individuals ranged from hand motion to 0.4. The R213W mutation in exon 6 of RS1 was identified in all affected individuals, predicting an amino acid substitution of arginine to tryptophan at codon 213. Our data show that the R213W mutation in RS1 causes various severities of retinoschisis in a large Chinese family, providing further evidence for X-linked juvenile retinoschisis phenotypic variability.

A novel autosomal partially dominant mutation designated G476D in the keratin 5 gene causing epidermolysis bullosa simplex Weber-Cockayne type: a family study with a genetic twist.

PubMed

Kowalewski, Cezary; Hamada, Takahiro; Wozniak, Katarzyna; Kawano, Yuko; Szczecinska, Weronika; Yasumoto, Shinichiro; Schwartz, Robert A; Hashimoto, Takashi

2007-07-01

Epidermolysis bullosa simplex Weber-Cockayne type (EBS-WC) is a genetically inherited skin disease characterized by blistering restricted to the palms and soles. Its inheritance in nearly all kindreds is caused by a dominant-negative mutation in either KRT5 or KRT14, the genes encoding keratin 5 and keratin 14 proteins, respectively. Rarely, recessive mutations have also been found. We described a family with EBS-WC caused by a novel autosomal dominant mutation (G476D) in the keratin 5 gene. One family member was first seen with mucosal erosions and generalized blisters localized on the anogenital area, trunk, face and sites of mechanical trauma. Molecular analysis in this patient showed the presence of an additional mutation, an autosomal recessive (G183E) one, in the same gene. This observation suggests an additional effect of a recessively inherited mutation modulating the phenotypic expression of EBS caused by a partially dominant mutation and is important for accurate genetic counseling.

Novel cases of Tunisian patients with mutations in the gene encoding 17β-hydroxysteroid dehydrogenase type 3 and a founder effect.

PubMed

Ben Rhouma, Bochra; Kallabi, Fakhri; Mahfoudh, Nadia; Ben Mahmoud, Afif; Engeli, Roger T; Kamoun, Hassen; Keskes, Leila; Odermatt, Alex; Belguith, Neila

2017-01-01

17β-Hydroxysteroid dehydrogenase type 3 (17β-HSD3) is expressed almost exclusively in the testis and converts Δ4-androstene-3,17-dione to testosterone. Mutations in the HSD17B3 gene causing 17β-HSD3 deficiency are responsible for a rare recessive form of 46, XY Disorders of Sex Development (46, XY DSD). We report novel cases of Tunisian patients with 17β-HSD3 deficiency due to previously reported mutations, i.e. p.C206X and p.G133R, as well as a case with the novel compound heterozygous mutations p.C206X and p.Q176P. Moreover, the previously reported polymorphism p.G289S was identified in a heterozygous state in combination with a novel non-coding variant c.54G>T, also in a heterozygous state, in a male patient presenting with micropenis and low testosterone levels. The identification of four different mutations in a cohort of eight patients confirms the generally observed genetic heterogeneity of 17β-HSD3 deficiency. Nevertheless, analysis of DNA from 272 randomly selected healthy controls from the same geographic area (region of Sfax) revealed a high carrier frequency for the p.C206X mutation of approximately 1 in 40. Genotype reconstruction of the affected pedigree members revealed that all p.C206X mutation carriers harbored the same haplotype, indicating inheritance of the mutation from a common ancestor. Thus, the identification of a founder effect and the elevated carrier frequency of the p.C206X mutation emphasize the importance to consider this mutation in the diagnosis and genetic counseling of affected 17β-HSD3 deficiency pedigrees in Tunisia. Copyright © 2016 Elsevier Ltd. All rights reserved.

Mutations in Plasmodium falciparum K13 propeller gene from Bangladesh (2009-2013).

PubMed

Mohon, Abu Naser; Alam, Mohammad Shafiul; Bayih, Abebe Genetu; Folefoc, Asongna; Shahinas, Dea; Haque, Rashidul; Pillai, Dylan R

2014-11-18

Bangladesh is a malaria hypo-endemic country sharing borders with India and Myanmar. Artemisinin combination therapy (ACT) remains successful in Bangladesh. An increase of artemisinin-resistant malaria parasites on the Thai-Cambodia and Thai-Myanmar borders is worrisome. K13 propeller gene (PF3D7_1343700 or PF13_0238) mutations have been linked to both in vitro artemisinin resistance and in vivo slow parasite clearance rates. This group undertook to evaluate if mutations seen in Cambodia have emerged in Bangladesh where ACT use is now standard for a decade. Samples were obtained from Plasmodium falciparum-infected malaria patients from Upazila health complexes (UHC) between 2009 and 2013 in seven endemic districts of Bangladesh. These districts included Khagrachari (Matiranga UHC), Rangamati (Rajasthali UHC), Cox's Bazar (Ramu and Ukhia UHC), Bandarban (Lama UHC), Mymensingh (Haluaghat UHC), Netrokona (Durgapur and Kalmakanda UHC), and Moulvibazar (Sreemangal and Kamalganj UHC). Out of 296 microscopically positive P. falciparum samples, 271 (91.6%) were confirmed as mono-infections by both real-time PCR and nested PCR. The K13 propeller gene from 253 (93.4%) samples was sequenced bi-directionally. One non-synonymous mutation (A578S) was found in Bangladeshi clinical isolates. The A578S mutation was confirmed and lies adjacent to the C580Y mutation, the major mutation causing delayed parasite clearance in Cambodia. Based on computational modeling A578S should have a significant effect on tertiary structure of the protein. The data suggest that P. falciparum in Bangladesh remains free of the C580Y mutation linked to delayed parasite clearance. However, the mutation A578S is present and based on structural analysis could affect K13 gene function. Further in vivo clinical studies are required to validate the effect of this mutation.

Mutations in Kelch-like 3 and Cullin 3 cause hypertension and electrolyte abnormalities

PubMed Central

Boyden, Lynn M.; Choi, Murim; Choate, Keith A.; Nelson-Williams, Carol J.; Farhi, Anita; Toka, Hakan R.; Tikhonova, Irina R.; Bjornson, Robert; Mane, Shrikant M.; Colussi, Giacomo; Lebel, Marcel; Gordon, Richard D.; Semmekrot, Ben A.; Poujol, Alain; Välimäki, Matti J.; De Ferrari, Maria E.; Sanjad, Sami A.; Gutkin, Michael; Karet, Fiona E.; Tucci, Joseph R.; Stockigt, Jim R.; Keppler-Noreuil, Kim M.; Porter, Craig C.; Anand, Sudhir K.; Whiteford, Margo L.; Davis, Ira D.; Dewar, Stephanie B.; Bettinelli, Alberto; Fadrowski, Jeffrey J.; Belsha, Craig W.; Hunley, Tracy E.; Nelson, Raoul D.; Trachtman, Howard; Cole, Trevor R. P.; Pinsk, Maury; Bockenhauer, Detlef; Shenoy, Mohan; Vaidyanathan, Priya; Foreman, John W.; Rasoulpour, Majid; Thameem, Farook; Al-Shahrouri, Hania Z.; Radhakrishnan, Jai; Gharavi, Ali G.; Goilav, Beatrice; Lifton, Richard P.

2012-01-01

Hypertension affects one billion people and is a principal reversible risk factor for cardiovascular disease. A rare Mendelian syndrome, pseudohypoaldosteronism type II (PHAII), featuring hypertension, hyperkalemia, and metabolic acidosis, has revealed previously unrecognized physiology orchestrating the balance between renal salt reabsorption versus K+ and H+ excretion1. We used exome sequencing to identify mutations in Kelch-like 3 (KLHL3) or Cullin 3 (CUL3) in 41 PHAII kindreds. KLHL3 mutations are either recessive or dominant, while CUL3 mutations are dominant and predominantly de novo. CUL3 and BTB-Kelch proteins such as KLHL3 are components of Cullin/RING E3 ligase complexes (CRLs) that ubiquitinate substrates bound to Kelch propeller domains2–8. Dominant KLHL3 mutations are clustered in short segments within the Kelch propeller and BTB domains implicated in substrate9 and Cullin5 binding, respectively. Diverse CUL3 mutations all result in skipping of exon 9, producing an in-frame deletion. Because dominant KLHL3 and CUL3 mutations both phenocopy recessive loss-of-function KLHL3 mutations, they may abrogate ubiquitination of KLHL3 substrates. Disease features are reversed by thiazide diuretics, which inhibit the Na-Cl cotransporter (NCC) in the distal nephron of the kidney; KLHL3 and CUL3 are expressed in this location, suggesting a mechanistic link between KLHL3/CUL3 mutations, increased Na-Cl reabsorption, and disease pathogenesis. These findings demonstrate the utility of exome sequencing in disease gene identification despite combined complexities of locus heterogeneity, mixed models of transmission, and frequent de novo mutation, and establish a fundamental role for KLHL3/CUL3 in blood pressure, K+, and pH homeostasis. PMID:22266938

Reengineering of a Corynebacterium glutamicum L-arginine and L-citrulline producer.

PubMed

Ikeda, Masato; Mitsuhashi, Satoshi; Tanaka, Kenji; Hayashi, Mikiro

2009-03-01

Toward the creation of a robust and efficient producer of L-arginine and L-citrulline (arginine/citrulline), we have performed reengineering of a Corynebacterium glutamicum strain by using genetic information of three classical producers. Sequence analysis of their arg operons identified three point mutations (argR123, argG92(up), and argG45) in one producer and one point mutation (argB26 or argB31) in each of the other two producers. Reconstitution of the former three mutations or of each argB mutation on a wild-type genome led to no production. Combined introduction of argB26 or argB31 with argR123 into a wild type gave rise to arginine/citrulline production. When argR123 was replaced by an argR-deleted mutation (Delta argR), the production was further increased. The best mutation set, Delta argR and argB26, was used to screen for the highest productivity in the backgrounds of different wild-type strains of C. glutamicum. This yielded a robust producer, RB, but the production was still one-third of that of the best classical producer. Transcriptome analysis revealed that the arg operon of the classical producer was much more highly upregulated than that of strain RB. Introduction of leuC456, a mutation derived from a classical L-lysine producer and provoking global induction of the amino acid biosynthesis genes, including the arg operon, into strain RB led to increased production but incurred retarded fermentation. On the other hand, replacement of the chromosomal argB by heterologous Escherichia coli argB, natively insensitive to arginine, caused a threefold-increased production without retardation, revealing that the limitation in strain RB was the activity of the argB product. To overcome this, in addition to argB26, the argB31 mutation was introduced into strain RB, which caused higher deregulation of the enzyme and resulted in dramatically increased production, like the strain with E. coli argB. This reconstructed strain displayed an enhanced performance, thus allowing significantly higher productivity of arginine/citrulline even at the suboptimal 38 degrees C.

Advanced cell-based modeling of the royal disease: characterization of the mutated F9 mRNA.

PubMed

Martorell, L; Luce, E; Vazquez, J L; Richaud-Patin, Y; Jimenez-Delgado, S; Corrales, I; Borras, N; Casacuberta-Serra, S; Weber, A; Parra, R; Altisent, C; Follenzi, A; Dubart-Kupperschmitt, A; Raya, A; Vidal, F; Barquinero, J

2017-11-01

Essentials The Royal disease (RD) is a form of hemophilia B predicted to be caused by a splicing mutation. We generated an iPSC-based model of the disease allowing mechanistic studies at the RNA level. F9 mRNA analysis in iPSC-derived hepatocyte-like cells showed the predicted abnormal splicing. Mutated F9 mRNA level was very low but we also found traces of wild type transcripts. Background The royal disease is a form of hemophilia B (HB) that affected many descendants of Queen Victoria in the 19th and 20th centuries. It was found to be caused by the mutation F9 c.278-3A>G. Objective To generate a physiological cell model of the disease and to study F9 expression at the RNA level. Methods Using fibroblasts from skin biopsies of a previously identified hemophilic patient bearing the F9 c.278-3A>G mutation and his mother, we generated induced pluripotent stem cells (iPSCs). Both the patient's and mother's iPSCs were differentiated into hepatocyte-like cells (HLCs) and their F9 mRNA was analyzed using next-generation sequencing (NGS). Results and Conclusion We demonstrated the previously predicted aberrant splicing of the F9 transcript as a result of an intronic nucleotide substitution leading to a frameshift and the generation of a premature termination codon (PTC). The F9 mRNA level in the patient's HLCs was significantly reduced compared with that of his mother, suggesting that mutated transcripts undergo nonsense-mediated decay (NMD), a cellular mechanism that degrades PTC-containing mRNAs. We also detected small proportions of correctly spliced transcripts in the patient's HLCs, which, combined with genetic variability in splicing and NMD machineries, could partially explain some clinical variability among affected members of the European royal families who had lifespans above the average. This work allowed the demonstration of the pathologic consequences of an intronic mutation in the F9 gene and represents the first bona fide cellular model of HB allowing the study of rare mutations at the RNA level. © 2017 International Society on Thrombosis and Haemostasis.

Two coexisting heterozygous frameshift mutations in PROP1 are responsible for a different phenotype of combined pituitary hormone deficiency.

PubMed

Ziemnicka, K; Budny, B; Drobnik, K; Baszko-Błaszyk, D; Stajgis, M; Katulska, K; Waśko, R; Wrotkowska, E; Słomski, R; Ruchała, M

2016-08-01

The role of genetic background in childhood-onset combined pituitary hormone deficiency (CPHD) has been extensively studied. The major contributors are the PROP1, POU1F1, LHX3, LHX4 and HESX1 genes coding transcription factors implicated in pituitary organogenesis. The clinical consequences of mutations encompass impaired synthesis of a growth hormone (GH) and one or more concurrent pituitary hormones (i.e. LH, FSH, TSH, PRL). Manifestation of the disorder may vary due to various mutation impacts on the final gene products or an influence of environmental factors during pituitary organogenesis. We describe the clinical and molecular characteristics of two brothers aged 47 and 39 years presenting an uncommon manifestation of congenital hypopituitarism. Sequencing of the PROP1, POU1F1, LHX3, LHX4 and HESX1 genes was performed to confirm the genetic origin of the disorder. A compound heterozygosity in the PROP1 gene has been identified for both probands. The first change represents a mutational hot spot (c.150delA, p.R53fsX164), whereas the second is a novel alteration (p.R112X) that leads to protein disruption. Based on precise genetic diagnosis, an in silico prediction of a p.R112X mutation on protein architecture was performed. The resulting clinical phenotype was surprisingly distinct compared to most patients with genetic alterations in PROP1 reported in the current literature. This may be caused by a residual activity of a newly identified p.R112X protein that preserves over 70 % of the homeodomain structure. This examination may confirm a key role of a DNA-binding homeodomain in maintaining PROP1 functionality and suggests a conceivable explanation of an unusual phenotype.

Consensus on management of metastatic colorectal cancer in Central America and the Caribbean: San José, Costa Rica, August 2016

PubMed Central

López, Roberto Ivan; Castro, Jenny Lissette; Cedeño, Heidy; Cisneros, Dagoberto; Corrales, Luis; González-Herrera, Ileana; Lima-Pérez, Mayté; Prestol, Rogelio; Salinas, Roberto; Soriano-García, Jorge Luis; T Zavala, Alejandra; Zetina, Luis Miguel; Zúñiga-Orlich, Carlos Eduardo

2018-01-01

Colorectal cancer (CRC) is the third most common cancer in men and the second most common in women worldwide. In Latin America and the Caribbean, it has a mortality of 56%. The median overall survival for patients with metastatic colorectal cancer (mCRC) is currently estimated as ~30 months, which has substantially improved through strategic changes in treatment and in the management of patients. As opposed to other metastatic cancers where first-line regimens are often determined, mCRC requires special attention because there is controversy in the possible combinations of the available drugs and the different periods of duration for each patient. Each combination must seek to be effective and to generate the minimum adverse effects as possible. Instead of giving the first-line regimen until the tumour progresses, treatment is often individualised. Furthermore, up to 60% of colorectal tumours are considered non-mutated or wild-type CRC. Not harbouring mutations in the RAS family of genes or mutations in the signalling pathways of the epidermal growth factor receptor causes a null response to anti-epidermal growth factor receptor antibody therapy, which implies even more complex considerations regarding its management. The primary objective of this consensus is to address the main scenarios of mCRC in order to warrant the most appropriate therapeutic intervention for these patients in the Central American and the Caribbean (CAC) region. This can lead to better clinical outcomes as well as quality of life for palliative patients. This document includes the formal expert consensus recommendations for scenarios of mutated and non-mutated mCRC, including synchronous or metachronous disease, management of mCRC with liver and lung metastasis, resectable, potentially resectable or non-resectable tumours and local in the CAC context. PMID:29636987

Non-HFE haemochromatosis

PubMed Central

Wallace, Daniel F; Subramaniam, V Nathan

2007-01-01

Non-HFE hereditary haemochromatosis (HH) refers to a genetically heterogeneous group of iron overload disorders that are unlinked to mutations in the HFE gene. The four main types of non-HFE HH are caused by mutations in the hemojuvelin, hepcidin, transferrin receptor 2 and ferroportin genes. Juvenile haemochromatosis is an autosomal recessive disorder and can be caused by mutations in either hemojuvelin or hepcidin. An adult onset form of HH similar to HFE-HH is caused by homozygosity for mutations in transferrin receptor 2. The autosomal dominant iron overload disorder ferroportin disease is caused by mutations in the iron exporter ferroportin. The clinical characteristics and molecular basis of the various types of non-HFE haemochromatosis are reviewed. The study of these disorders and the molecules involved has been invaluable in improving our understanding of the mechanisms involved in the regulation of iron metabolism. PMID:17729390

Variation, "evolution", immortality and genetic instabilities in tumour cells.

PubMed

Bignold, L P

2007-08-18

The pathological characteristics of tumour cells often include variation of their histopathological features (i.e. "degrees of de-differentiation") between cases of the same tumour type and between different foci within individual tumours. Usually, only a few cell lines from tumours are immortal. Currently, somatic mutation, replicative infidelity of DNA and aneuploidy are suggested as alternative mechanisms of genomic disturbance underlying tumours. Nevertheless, apart from Hansemann's ideas of "anaplasia" and "de-differentiation" (proposed in the 1890s), and supposed "evolutionary themes" in cancer cell biology, little has been published concerning how histopathologic variation and immortality in tumour cells might arise. This paper reviews applications of the concepts of "variation" to tumours, including concepts of "evolution" and "cellular Darwinism". It is proposed that combinations of somatic mutation, DNA replicative infidelity and aneuploidy may explain the variabilities in tumours, and provide immortality in occasional tumour cells. A possible model involves (i) an initial somatic mutation causing reduced replicative fidelity of DNA, which could be variable in intensity, and thus give rise to variations between cases; (ii) a phase of replicative infidelity of DNA causing daughter cells lines to develop various abnormalities to different degrees, and hence provide for variation between areas of the same tumour. As a last event (iii) occasional asymmetric chromosomal distributions (aneuploidy) might "refresh" the ability of a daughter cell to replicate DNA faithfully causing them to become immortal. Thus extensively mutant and variable, hyperploid, and occasionally immortal cells might arise.

Toward the Mutational Landscape of Autosomal Dominant Retinitis Pigmentosa: A Comprehensive Analysis of 258 Spanish Families.

PubMed

Martin-Merida, Inmaculada; Aguilera-Garcia, Domingo; Jose, Patricia Fernandez-San; Blanco-Kelly, Fiona; Zurita, Olga; Almoguera, Berta; Garcia-Sandoval, Blanca; Avila-Fernandez, Almudena; Arteche, Ana; Minguez, Pablo; Carballo, Miguel; Corton, Marta; Ayuso, Carmen

2018-05-01

To provide a comprehensive overview of the molecular basis of autosomal dominant retinitis pigmentosa (adRP) in Spanish families. Thus, we established the molecular characterization rate, gene prevalence, and mutational spectrum in the largest European cohort reported to date. A total of 258 unrelated Spanish families with a clinical diagnosis of RP and suspected autosomal dominant inheritance were included. Clinical diagnosis was based on complete ophthalmologic examination and family history. Retrospective and prospective analysis of Spanish adRP families was carried out using a combined strategy consisting of classic genetic techniques and next-generation sequencing (NGS) for single-nucleotide variants and copy number variation (CNV) screening. Overall, 60% of our families were genetically solved. Interestingly, 3.1% of the cohort carried pathogenic CNVs. Disease-causing variants were found in an autosomal dominant gene in 55% of the families; however, X-linked and autosomal recessive forms were also identified in 3% and 2%, respectively. Four genes (RHO, PRPF31, RP1, and PRPH2) explained up to 62% of the solved families. Missense changes were most frequently found in adRP-associated genes; however, CNVs represented a relevant disease cause in PRPF31- and CRX-associated forms. Implementation of NGS technologies in the adRP study clearly increased the diagnostic yield compared with classic approaches. Our study outcome expands the spectrum of disease-causing variants, provides accurate data on mutation gene prevalence, and highlights the implication of CNVs as important contributors to adRP etiology.

Clinical variability of Waardenburg-Shah syndrome in patients with proximal 13q deletion syndrome including the endothelin-B receptor locus.

PubMed

Tüysüz, Beyhan; Collin, Anna; Arapoğlu, Müjde; Suyugül, Nezir

2009-10-01

Waardenburg-Shah syndrome (Waardenburg syndrome type IV-WS4) is an auditory-pigmentary disorder that combines clinical features of pigmentary abnormalities of the skin, hair and irides, sensorineural hearing loss, and Hirschsprung disease (HSCR). Mutations in the endothelin-B receptor (EDNRB) gene on 13q22 have been found to cause this syndrome. Mutations in both alleles cause the full phenotype, while heterozygous mutations cause isolated HSCR or HSCR with minor pigmentary anomalies and/or sensorineural deafness. We investigated the status of the EDNRB gene, by FISH analysis, in three patients with de novo proximal 13q deletions detected at cytogenetic analysis and examined the clinical variability of WS4 among these patients. Chromosome 13q was screened with locus specific FISH probes and breakpoints were determined at 13q22.1q31.3 in Patients 1 and 3, and at 13q21.1q31.3 in Patient 2. An EDNRB specific FISH probe was deleted in all three patients. All patients had common facial features seen in proximal 13q deletion syndrome and mild mental retardation. However, findings related to WS4 were variable; Patient 1 had hypopigmentation of the irides and HSCR, Patient 2 had prominent bicolored irides and mild bilateral hearing loss, and Patient 3 had only mild unilateral hearing loss. These data contribute new insights into the pathogenesis of WS4.

The genetic landscape of familial congenital hydrocephalus.

PubMed

Shaheen, Ranad; Sebai, Mohammed Adeeb; Patel, Nisha; Ewida, Nour; Kurdi, Wesam; Altweijri, Ikhlass; Sogaty, Sameera; Almardawi, Elham; Seidahmed, Mohammed Zain; Alnemri, Abdulrahman; Madirevula, Sateesh; Ibrahim, Niema; Abdulwahab, Firdous; Hashem, Mais; Al-Sheddi, Tarfa; Alomar, Rana; Alobeid, Eman; Sallout, Bahauddin; AlBaqawi, Badi; AlAali, Wajeih; Ajaji, Nouf; Lesmana, Harry; Hopkin, Robert J; Dupuis, Lucie; Mendoza-Londono, Roberto; Al Rukban, Hadeel; Yoon, Grace; Faqeih, Eissa; Alkuraya, Fowzan S

2017-06-01

Congenital hydrocephalus is an important birth defect, the genetics of which remains incompletely understood. To date, only 4 genes are known to cause Mendelian diseases in which congenital hydrocephalus is the main or sole clinical feature, 2 X-linked (L1CAM and AP1S2) and 2 autosomal recessive (CCDC88C and MPDZ). In this study, we aimed to determine the genetic etiology of familial congenital hydrocephalus with the assumption that these cases represent Mendelian forms of the disease. Exome sequencing combined, where applicable, with positional mapping. We identified a likely causal mutation in the majority of these families (21 of 27, 78%), spanning 16 genes, none of which is X-linked. Ciliopathies and dystroglycanopathies were the most common etiologies of congenital hydrocephalus in our cohort (19% and 26%, respectively). In 1 family with 4 affected members, we identified a homozygous truncating variant in EML1, which we propose as a novel cause of congenital hydrocephalus in addition to its suggested role in cortical malformation. Similarly, we show that recessive mutations in WDR81, previously linked to cerebellar ataxia, mental retardation, and disequilibrium syndrome 2, cause severe congenital hydrocephalus. Furthermore, we confirm the previously reported candidacy of MPDZ by presenting a phenotypic spectrum of congenital hydrocephalus associated with 5 recessive alleles. Our study highlights the importance of recessive mutations in familial congenital hydrocephalus and expands the locus heterogeneity of this condition. Ann Neurol 2017;81:890-897. © 2017 American Neurological Association.

Molecular characterisation of congenital glaucoma in a consanguineous Canadian community: a step towards preventing glaucoma related blindness

PubMed Central

Martin, S; Sutherland, J.; Levin, A.; Klose, R.; Priston, M.; Heon, E.

2000-01-01

Glaucoma is a leading cause of irreversible blindness in Canada. Congenital glaucoma usually manifests during the first years of life and is characterised by severe visual loss and autosomal recessive inheritance. Two disease loci, on chromosomes 1p36 and 2p21, have been associated with various forms of congenital glaucoma. A branch of a large six generation family from a consanguineous Amish community in south western Ontario was affected with congenital glaucoma and was studied by linkage and mutational analysis to identify the glaucoma related genetic defects. Linkage analysis using the MLINK component of the LINKAGE package (v 5.1) showed evidence of linkage to the 2p21 region (Zmax=3.34, θ=0, D2S1348 and D2S1346). Mutational analysis of the primary candidate gene, CYP1B1, was done by direct cycle sequencing, dideoxy fingerprinting analysis, and fragment analysis. Two different disease causing mutations in exon 3, 1410del13 and 1505G→A, both segregated with the disease phenotype. The two different combinations of these alleles appeared to result in a variable expressivity of the phenotype. The compound heterozygote appeared to have a milder phenotype when compared to the homozygotes for the 13 bp deletion. The congenital glaucoma phenotype for this large inbred Amish family is the result of mutations in CYP1B1 (2p21). The molecular information derived from this study will be used to help identify carriers of the CYP1B1 mutation in this community and optimise the management of those at risk of developing glaucoma.


Keywords: congenital glaucoma; CYP1B1; gene; genetic counselling PMID:10851252

Novel mutation detection of fibroblast growth factor receptor 1 (FGFR1) gene, FGFR2IIIa, FGFR2IIIb, FGFR2IIIc, FGFR3, FGFR4 gene for craniosynostosis: A prospective study in Asian Indian patient.

PubMed

Barik, Mayadhar; Bajpai, Minu; Malhotra, Arun; Samantaray, Jyotish Chandra; Dwivedi, Sadananda; Das, Sambhunath

2015-01-01

Craniosynostosis (CS) syndrome is an autosomal dominant condition classically combining craniosynostosis and non-syndromic craniosynostosis with digital anomalies of the hands and feet. The majority of cases are caused by heterozygous mutations in the third immunoglobulin-like domain (IgIII) of FGFR2, whilst a larger number of cases can be attributed to mutations outside this region of the protein. To find out the FGFR1, FGFR2, FGFR3 and FGFR4 gene in craniosynostosis syndrome. A hospital based prospective study. Prospective analysis of clinical records of patients registered in CS clinic from December 2007 to January 2015 was done in patients between 4 months to 13 years of age. We have performed genetic findings in a three generation Indian family with Craniosynostosis syndrome. We report for the first time the clinical and genetic findings in a three generation Indian family with Craniosynostosis syndrome caused by a heterozygous missense mutation, Thr 392 Thr and ser 311 try, located in the IgII domain of FGFR2. FGFR 3 and 4 gene basis syndrome was eponymously named. Genetic analysis demonstrated that 51/56 families to be unrelated. In FGFR3 gene 10/TM location of 1172 the nucleotide changes C>A, Ala 391 Glu 19/56 and Exon-19, 5q35.2 at conserved linker region the changes occurred pro 246 Arg in 25/56 families. Independent genetic origins, but phenotypic similarities in the 51 families add to the evidence supporting the theory of selfish spermatogonial selective advantage for this rare gain-of-function FGFR2 mutation.

Novel mutation detection of fibroblast growth factor receptor 1 (FGFR1) gene, FGFR2IIIa, FGFR2IIIb, FGFR2IIIc, FGFR3, FGFR4 gene for craniosynostosis: A prospective study in Asian Indian patient

PubMed Central

Barik, Mayadhar; Bajpai, Minu; Malhotra, Arun; Samantaray, Jyotish Chandra; Dwivedi, Sadananda; Das, Sambhunath

2015-01-01

Background: Craniosynostosis (CS) syndrome is an autosomal dominant condition classically combining craniosynostosis and non-syndromic craniosynostosis with digital anomalies of the hands and feet. The majority of cases are caused by heterozygous mutations in the third immunoglobulin-like domain (IgIII) of FGFR2, whilst a larger number of cases can be attributed to mutations outside this region of the protein. Aims: To find out the FGFR1, FGFR2, FGFR3 and FGFR4 gene in craniosynostosis syndrome. Settings and Design: A hospital based prospective study. Materials and Methods: Prospective analysis of clinical records of patients registered in CS clinic from December 2007 to January 2015 was done in patients between 4 months to 13 years of age. We have performed genetic findings in a three generation Indian family with Craniosynostosis syndrome. Results: We report for the first time the clinical and genetic findings in a three generation Indian family with Craniosynostosis syndrome caused by a heterozygous missense mutation, Thr 392 Thr and ser 311 try, located in the IgII domain of FGFR2. FGFR 3 and 4 gene basis syndrome was eponymously named. Genetic analysis demonstrated that 51/56 families to be unrelated. In FGFR3 gene 10/TM location of 1172 the nucleotide changes C>A, Ala 391 Glu 19/56 and Exon-19, 5q35.2 at conserved linker region the changes occurred pro 246 Arg in 25/56 families. Conclusions: Independent genetic origins, but phenotypic similarities in the 51 families add to the evidence supporting the theory of selfish spermatogonial selective advantage for this rare gain-of-function FGFR2 mutation. PMID:26557159

The Combination of CRISPR/Cas9 and iPSC Technologies in the Gene Therapy of Human β-thalassemia in Mice.

PubMed

Ou, Zhanhui; Niu, Xiaohua; He, Wenyin; Chen, Yuchang; Song, Bing; Xian, Yexing; Fan, Di; Tang, Daolin; Sun, Xiaofang

2016-09-01

β-thalassemia results from point mutations or small deletions in the β-globin (HBB) gene that ultimately cause anemia. The generation of induced pluripotent stem cells (iPSCs) from the somatic cells of patients in combination with subsequent homologous recombination-based gene correction provides new approaches to cure this disease. CRISPR/Cas9 is a genome editing tool that is creating a buzz in the scientific community for treating human diseases, especially genetic disorders. Here, we reported that correction of β-thalassemia mutations in patient-specific iPSCs using the CRISPR/Cas9 tool promotes hematopoietic differentiation in vivo. CRISPR/Cas9-corrected iPSC-derived hematopoietic stem cells (HSCs) were injected into sublethally-irradiated NOD-scid-IL2Rg-/- (NSI) mice. HBB expression was observed in these HSCs after hematopoietic differentiation in the NSI mice. Importantly, no tumor was found in the livers, lungs, kidneys, or bone marrow at 10 weeks in the NSI mice after implantation with these HSCs. Collectively, our findings demonstrated that CRISPR/Cas9 successfully corrects β-thalassemia mutations in patient-specific iPSCs. These CRISPR/Cas9-corrected iPSC-derived HSCs express normal HBB in mice without tumorigenic potential, suggesting a safe strategy for personalized treatment of β-thalassemia.

Genome Editing in Induced Pluripotent Stem Cells using CRISPR/Cas9.

PubMed

Ben Jehuda, Ronen; Shemer, Yuval; Binah, Ofer

2018-06-01

The development of the reprogramming technology led to generation of induced Pluripotent Stem Cells (iPSC) from a variety of somatic cells. Ever since, fast growing knowledge of different efficient protocols enabled the differentiation of these iPSCs into different cells types utilized for disease modeling. Indeed, iPSC-derived cells have been increasingly used for investigating molecular and cellular pathophysiological mechanisms underlying inherited diseases. However, a major barrier in the field of iPSC-based disease modeling relies on discriminating between the effects of the causative mutation and the genetic background of these cells. In the past decade, researchers have made great improvement in genome editing techniques, with one of the latest being CRISPR/Cas9. Using a single non-sequence specific protein combined with a small guiding RNA molecule, this state-of-the-art approach enables modifications of genes with high efficiency and accuracy. By so doing, this technique enables the generation of isogenic controls or isogenic mutated cell lines in order to focus on the pathologies caused by a specific mutation. In this article, we review the latest studies combining iPSC and CRISPR/Cas9 technologies for the investigation of the molecular and cellular mechanisms underlying inherited diseases including immunological, metabolic, hematological, neurodegenerative and cardiac diseases.

Revisiting sweat chloride test results based on recent guidelines for diagnosis of cystic fibrosis.

PubMed

Pagaduan, Jayson V; Ali, Mahesheema; Dowlin, Michael; Suo, Liye; Ward, Tabitha; Ruiz, Fadel; Devaraj, Sridevi

2018-03-01

Recent sweat chloride guidelines published by the Cystic Fibrosis Foundation changed the intermediate sweat chloride concentration range from 40-59 mmol/L to 30-59 mmol/L for age > 6 months. We wanted to know how this new guideline would impact detection of cystic fibrosis among patients who previously had sweat tests done at Texas Children's Hospital. We revisited sweat chloride test results (n = 3012) in the last 5 years at Texas Children's Hospital based on the new guidelines on diagnosis of cystic fibrosis from the Cystic Fibrosis Foundation. We identified 125 patients that would be reclassified in the intermediate sweat chloride value with the new guidelines that were classified as "unlikely to have CF" in the previous guidelines. 8 (32%) patients with CFTR gene testing were positive for CFTR gene mutation(s). 4 (50%) of these patients were identified to have 2 CFTR mutations. One had variant combination that was reported to cause CF but all were diagnosed with CFTR-related metabolic syndrome. Our findings concur with the new CF diagnosis guidelines that changing the intermediate cut-off to 30-59 mmol/L sweat chloride concentration in combination with CFTR genetic analysis enhances the probability of identifying individuals that have risk of developing CF or have CF and enables for earlier therapeutic intervention.

Nocturnal frontal lobe epilepsy caused by a mutation in the GATOR1 complex gene NPRL3.

PubMed

Korenke, Georg-Christoph; Eggert, Marlene; Thiele, Holger; Nürnberg, Peter; Sander, Thomas; Steinlein, Ortrud K

2016-03-01

Mutations in NPRL3, one of three genes that encode proteins of the mTORC1-regulating GATOR1 complex, have recently been reported to cause cortical dysplasia with focal epilepsy. We have now analyzed a multiplex epilepsy family by whole exome sequencing and identified a frameshift mutation (NM_001077350.2; c.1522delG; p.E508Rfs*46) within exon 13 of NPRL3. This truncating mutation causes an epilepsy phenotype characterized by early childhood onset of mainly nocturnal frontal lobe epilepsy. The penetrance in our family was low (three affected out of six mutation carriers), compared to families with either ion channel- or DEPDC5-associated familial nocturnal frontal lobe epilepsy. The absence of apparent structural brain abnormalities suggests that mutations in NPRL3 are not necessarily associated with focal cortical dysplasia but might be able to cause epilepsy by different, yet unknown pathomechanisms. Wiley Periodicals, Inc. © 2016 International League Against Epilepsy.

A novel STXBP1 mutation causes typical Rett syndrome in a Japanese girl.

PubMed

Yuge, Kotaro; Iwama, Kazuhiro; Yonee, Chihiro; Matsufuji, Mayumi; Sano, Nozomi; Saikusa, Tomoko; Yae, Yukako; Yamashita, Yushiro; Mizuguchi, Takeshi; Matsumoto, Naomichi; Matsuishi, Toyojiro

2018-06-01

Rett syndrome (RTT) is a neurodevelopmental disorder mostly caused by mutations in Methyl-CpG-binding protein 2 (MECP2); however, mutations in various other genes may lead to RTT-like phenotypes. Here, we report the first case of a Japanese girl with RTT caused by a novel syntaxin-binding protein 1 (STXBP1) frameshift mutation (c.60delG, p.Lys21Argfs*16). She showed epilepsy at one year of age, regression of acquired psychomotor abilities thereafter, and exhibited stereotypic hand and limb movements at 3 years of age. Her epilepsy onset was earlier than is typical for RTT patients. However, she fully met the 2010 diagnostic criteria of typical RTT. STXBP1 mutations cause early infantile epileptic encephalopathy (EIEE), various intractable epilepsies, and neurodevelopmental disorders. However, the case described here presented a unique clinical presentation of typical RTT without EIEE and a novel STXBP1 mutation. Copyright © 2018 The Japanese Society of Child Neurology. Published by Elsevier B.V. All rights reserved.

Mitochondrial DNA sequence context in the penetrance of mitochondrial t-RNA mutations: A study across multiple lineages with diagnostic implications

PubMed Central

Queen, Rachel A.; Steyn, Jannetta S.; Lord, Phillip

2017-01-01

Mitochondrial DNA (mtDNA) mutations are well recognized as an important cause of inherited disease. Diseases caused by mtDNA mutations exhibit a high degree of clinical heterogeneity with a complex genotype-phenotype relationship, with many such mutations exhibiting incomplete penetrance. There is evidence that the spectrum of mutations causing mitochondrial disease might differ between different mitochondrial lineages (haplogroups) seen in different global populations. This would point to the importance of sequence context in the expression of mutations. To explore this possibility, we looked for mutations which are known to cause disease in humans, in animals of other species unaffected by mtDNA disease. The mt-tRNA genes are the location of many pathogenic mutations, with the m.3243A>G mutation on the mt-tRNA-Leu(UUR) being the most frequently seen mutation in humans. This study looked for the presence of m.3243A>G in 2784 sequences from 33 species, as well as any of the other mutations reported in association with disease located on mt-tRNA-Leu(UUR). We report a number of disease associated variations found on mt-tRNA-Leu(UUR) in other chordates, as the major population variant, with m.3243A>G being seen in 6 species. In these, we also found a number of mutations which appear compensatory and which could prevent the pathogenicity associated with this change in humans. This work has important implications for the discovery and diagnosis of mtDNA mutations in non-European populations. In addition, it might provide a partial explanation for the conflicting results in the literature that examines the role of mtDNA variants in complex traits. PMID:29161289

Mutations in the small nuclear riboprotein 200 kDa gene (SNRNP200) cause 1.6% of autosomal dominant retinitis pigmentosa

PubMed Central

Sullivan, Lori S.; Avery, Cheryl E.; Sasser, Elizabeth M.; Roorda, Austin; Duncan, Jacque L.; Wheaton, Dianna H.; Birch, David G.; Branham, Kari E.; Heckenlively, John R.; Sieving, Paul A.; Daiger, Stephen P.

2013-01-01

Purpose The purpose of this project was to determine the spectrum and frequency of mutations in the small nuclear riboprotein 200 kDa gene (SNRNP200) that cause autosomal dominant retinitis pigmentosa (adRP). Methods A well-characterized adRP cohort of 251 families was tested for mutations in the exons and intron/exon junctions of SNRNP200 using fluorescent dideoxy sequencing. An additional 21 adRP families from the eyeGENE® Network were tested for possible mutations. Bioinformatic and segregation analysis was performed on novel variants. Results SNRNP200 mutations were identified in seven of the families tested. Two previously reported mutations, p.Arg681Cys and p.Ser1087Leu, were found in two families each. One family had the previously reported p.Arg681His mutation. Two novel SNRNP200 variants, p.Pro682Ser and p.Ala542Val, were also identified in one family each. Bioinformatic and segregation analyses suggested that these novel variants are likely to be pathogenic. Clinical examination of patients with SNRNP200 mutations showed a wide range of clinical symptoms and severity, including one instance of non-penetrance. Conclusions Mutations in SNRNP200 caused 1.6% of disease in our adRP cohort. Pathogenic mutations were found primarily in exons 16 and 25, but the novel p.Ala542Val mutation in exon 13 suggests that variation in other genetic regions is also responsible for causing dominant disease. SNRNP200 mutations were associated with a wide range of clinical symptoms similar to those of individuals with other splice-factor gene mutations. PMID:24319334

Defective pigment granule biogenesis and aberrant behavior caused by mutations in the Drosophila AP-3beta adaptin gene ruby.

PubMed Central

Kretzschmar, D; Poeck, B; Roth, H; Ernst, R; Keller, A; Porsch, M; Strauss, R; Pflugfelder, G O

2000-01-01

Lysosomal protein trafficking is a fundamental process conserved from yeast to humans. This conservation extends to lysosome-like organelles such as mammalian melanosomes and insect eye pigment granules. Recently, eye and coat color mutations in mouse (mocha and pearl) and Drosophila (garnet and carmine) were shown to affect subunits of the heterotetrameric adaptor protein complex AP-3 involved in vesicle trafficking. Here we demonstrate that the Drosophila eye color mutant ruby is defective in the AP-3beta subunit gene. ruby expression was found in retinal pigment and photoreceptor cells and in the developing central nervous system. ruby mutations lead to a decreased number and altered size of pigment granules in various cell types in and adjacent to the retina. Humans with lesions in the related AP-3betaA gene suffer from Hermansky-Pudlak syndrome, which is caused by defects in a number of lysosome-related organelles. Hermansky-Pudlak patients have a reduced skin pigmentation and suffer from internal bleeding, pulmonary fibrosis, and visual system malfunction. The Drosophila AP-3beta adaptin also appears to be involved in processes other than eye pigment granule biogenesis because all ruby allele combinations tested exhibited defective behavior in a visual fixation paradigm. PMID:10790396

Evolving ecological networks and the emergence of biodiversity patterns across temperature gradients.

PubMed

Stegen, James C; Ferriere, Regis; Enquist, Brian J

2012-03-22

In ectothermic organisms, it is hypothesized that metabolic rates mediate influences of temperature on the ecological and evolutionary processes governing biodiversity. However, it is unclear how and to what extent the influence of temperature on metabolism scales up to shape large-scale diversity patterns. In order to clarify the roles of temperature and metabolism, new theory is needed. Here, we establish such theory and model eco-evolutionary dynamics of trophic networks along a broad temperature gradient. In the model temperature can influence, via metabolism, resource supply, consumers' vital rates and mutation rate. Mutation causes heritable variation in consumer body size, which diversifies and governs consumer function in the ecological network. The model predicts diversity to increase with temperature if resource supply is temperature-dependent, whereas temperature-dependent consumer vital rates cause diversity to decrease with increasing temperature. When combining both thermal dependencies, a unimodal temperature-diversity pattern evolves, which is reinforced by temperature-dependent mutation rate. Studying coexistence criteria for two consumers showed that these outcomes are owing to temperature effects on mutual invasibility and facilitation. Our theory shows how and why metabolism can influence diversity, generates predictions useful for understanding biodiversity gradients and represents an extendable framework that could include factors such as colonization history and niche conservatism.

Deactivating germline mutations in LEMD3 cause osteopoikilosis and Buschke-Ollendorff syndrome, but not sporadic melorheostosis.

PubMed

Mumm, Steven; Wenkert, Deborah; Zhang, Xiafang; McAlister, William H; Mier, Richard J; Whyte, Michael P

2007-02-01

Autosomal dominant OPK and BOS feature widespread foci of osteosclerotic trabeculae without or with skin lesions, respectively. Occasionally, a larger area of dense bone in OPK or BOS resembles MEL, a sporadic sclerosing disorder primarily involving cortical bone. Others, finding deactivating germline LEMD3 mutations in OPK or BOS, concluded such defects explain all three conditions. We found germline LEMD3 mutations in OPK and BOS but not in sporadic MEL. In 2004, others discovered that heterozygous, loss-of-function, germline mutations in the LEMD3 gene (LEMD3 or MAN1) cause both osteopoikilosis (OPK) and Buschke-Ollendorff syndrome (BOS). OPK is an autosomal dominant, usually benign, skeletal dysplasia featuring multiple, small, especially metaphyseal, oval or round, dense trabecular foci distributed symmetrically throughout the skeleton. BOS combines OPK with connective tissue nevi comprised of collagen and elastin. In some OPK and BOS families, an individual may have relatively large, asymmetric areas of dense cortical bone interpreted as melorheostosis (MEL). MEL, however, classically refers to a sporadic, troublesome skeletal dysostosis featuring large, asymmetric, "flowing hyperostosis" of long bone cortices often with overlying, constricting soft tissue abnormalities. However, a heterozygous germline mutation in LEMD3 was offered to explain MEL. We studied 11 unrelated individuals with sclerosing bone disorders where LEMD3 mutation was a potential etiology: familial OPK (1), familial BOS (2), previously reported familial OPK with MEL (1), sporadic MEL (3), sporadic MEL with mixed-sclerosing-bone dystrophy (1), and patients with other unusual sclerosing bone disorders (3). All coding exons and adjacent mRNA splice sites for LEMD3 were amplified by PCR and sequenced using genomic DNA from leukocytes. We did not study lesional tissue from bone or skin. In the OPK family, a heterozygous nonsense mutation (c.1433T>A, p.L478X) was discovered in exon 1. In the two BOS families, a heterozygous nonsense mutation (exon 1, c.1323C>A, p.Y441X) and a heterozygous frame-shift mutation (exon 1, c.332_333insTC) were identified. In the individual with MEL and familial OPK, a heterozygous nonsense mutation (c.1963C>T, p.R655X) was detected in exon 7. However, no LEMD3 mutation was found for any other patient, including all four with sporadic MEL. We confirm that OPK and BOS individuals, including those with MEL-like lesions, have heterozygous, deactivating, germline LEMD3 mutations. However, MEL remains of unknown etiology.

Prevailing PA Mutation K356R in Avian Influenza H9N2 Virus Increases Mammalian Replication and Pathogenicity.

PubMed

Xu, Guanlong; Zhang, Xuxiao; Gao, Weihua; Wang, Chenxi; Wang, Jinliang; Sun, Honglei; Sun, Yipeng; Guo, Lu; Zhang, Rui; Chang, Kin-Chow; Liu, Jinhua; Pu, Juan

2016-09-15

Adaptation of the viral polymerase complex comprising PB1, PB2, and PA is necessary for efficient influenza A virus replication in new host species. We found that PA mutation K356R (PA-K356R) has become predominant since 2014 in avian H9N2 viruses in China as with seasonal human H1N1 viruses. The same mutation is also found in most human isolates of emergent avian H7N9 and H10N8 viruses whose six internal gene segments are derived from the H9N2 virus. We further demonstrated the mammalian adaptive functionality of the PA-K356R mutation. Avian H9N2 virus with the PA-K356R mutation in human A549 cells showed increased nuclear accumulation of PA and increased viral polymerase activity that resulted in elevated levels of viral transcription and virus output. The same mutant virus in mice also enhanced virus replication and caused lethal infection. In addition, combined mutation of PA-K356R and PB2-E627K, a well-known mammalian adaptive marker, in the H9N2 virus showed further cooperative increases in virus production and severity of infection in vitro and in vivo In summary, PA-K356R behaves as a novel mammalian tropism mutation, which, along with other mutations such as PB2-E627K, might render avian H9N2 viruses adapted for human infection. Mutations of the polymerase complex (PB1, PB2, and PA) of influenza A virus are necessary for viral adaptation to new hosts. This study reports a novel and predominant mammalian adaptive mutation, PA-K356R, in avian H9N2 viruses and human isolates of emergent H7N9 and H10N8 viruses. We found that PA-356R in H9N2 viruses causes significant increases in virus replication and severity of infection in human cells and mice and that PA-K356R cooperates with the PB2-E627K mutation, a well-characterized human adaptive marker, to exacerbate mammalian infection in vitro and in vivo Therefore, the PA-K356R mutation is a significant adaptation in H9N2 viruses and related H7N9 and H10N8 reassortants toward human infectivity. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Prevailing PA Mutation K356R in Avian Influenza H9N2 Virus Increases Mammalian Replication and Pathogenicity

PubMed Central

Xu, Guanlong; Zhang, Xuxiao; Gao, Weihua; Wang, Chenxi; Wang, Jinliang; Sun, Honglei; Sun, Yipeng; Guo, Lu; Zhang, Rui; Chang, Kin-Chow; Liu, Jinhua

2016-01-01

ABSTRACT Adaptation of the viral polymerase complex comprising PB1, PB2, and PA is necessary for efficient influenza A virus replication in new host species. We found that PA mutation K356R (PA-K356R) has become predominant since 2014 in avian H9N2 viruses in China as with seasonal human H1N1 viruses. The same mutation is also found in most human isolates of emergent avian H7N9 and H10N8 viruses whose six internal gene segments are derived from the H9N2 virus. We further demonstrated the mammalian adaptive functionality of the PA-K356R mutation. Avian H9N2 virus with the PA-K356R mutation in human A549 cells showed increased nuclear accumulation of PA and increased viral polymerase activity that resulted in elevated levels of viral transcription and virus output. The same mutant virus in mice also enhanced virus replication and caused lethal infection. In addition, combined mutation of PA-K356R and PB2-E627K, a well-known mammalian adaptive marker, in the H9N2 virus showed further cooperative increases in virus production and severity of infection in vitro and in vivo. In summary, PA-K356R behaves as a novel mammalian tropism mutation, which, along with other mutations such as PB2-E627K, might render avian H9N2 viruses adapted for human infection. IMPORTANCE Mutations of the polymerase complex (PB1, PB2, and PA) of influenza A virus are necessary for viral adaptation to new hosts. This study reports a novel and predominant mammalian adaptive mutation, PA-K356R, in avian H9N2 viruses and human isolates of emergent H7N9 and H10N8 viruses. We found that PA-356R in H9N2 viruses causes significant increases in virus replication and severity of infection in human cells and mice and that PA-K356R cooperates with the PB2-E627K mutation, a well-characterized human adaptive marker, to exacerbate mammalian infection in vitro and in vivo. Therefore, the PA-K356R mutation is a significant adaptation in H9N2 viruses and related H7N9 and H10N8 reassortants toward human infectivity. PMID:27384648

EGFR mutations predict a favorable outcome for malignant pleural effusion of lung adenocarcinoma with Tarceva therapy.

PubMed

Guo, Haisheng; Wan, Yunyan; Tian, Guangyan; Liu, Qinghua; Kang, Yanmeng; Li, Yuye; Yao, Zhouhong; Lin, Dianjie

2012-03-01

The aim of the present study was to evaluate the therapeutic effects and adverse reactions of Tarceva treatment for malignant pleural effusion (MPE) caused by metastatic lung adenocarcinomas. One hundred and twenty-eight patients who failed first-line chemotherapy drug treatment were divided into a mutation and a non-mutation group according to the presence or absence of epidermal growth factor receptor (EGFR) mutations. Each patient received closed drainage combined with simple negative pressure suction after thoracoscopic talc poudrage pleurodesis and oral Tarceva treatment. Short-term and long-term clinical therapeutic effects of Tarceva were evaluated. The EGFR mutation rate in pleural metastatic tissues of lung adenocarcinoma acquired through video-assisted thoracoscopic surgery was higher compared to that in surgical resection specimens, plasma specimens and pleural effusion specimens compared to previously reported results. There were significant statistical differences in the average extubation time (p<0.01), drainage volume of pleural effusion (p<0.05), Karnofsky score and formation of encapsulated pleural effusion 4 weeks after surgery (p<0.05) between these two groups. The number of patients with mild pleural hypertrophy in the mutation group was significantly higher compared to the non-mutation group (p<0.01), while the number of patients with severe pleural hypertrophy was significantly reduced (p<0.05). There was significant statistical discrepancy between these two groups in terms of improvement of peripheral blood carcinoembryonic antigen and tissue polypeptide antigen after 4 weeks of therapy. The complete remission rate and the efficacy rate were higher in the mutation group compared to that in the non-mutation group (p<0.05). There was a longer overall survival time after Tarceva treatment in patients with EGFR mutations than those without EGFR mutation. EGFR mutations predict a favorable outcome for malignant pleural effusion of lung adenocarcinoma with Tarceva therapy. Detection of EGFR mutations may determine the responsiveness of malignant pleural effusion to Tarceva treatment.

Molecular-genetic diagnostics of von Hippel-Lindau syndrome (VHL) in Bulgaria: first complex mutation event in the VHL gene.

PubMed

Glushkova, Maria; Dimova, Petia; Yordanova, Iglika; Todorov, Tihomir; Tourtourikov, Ivan; Mitev, Vanyo; Todorova, Albena

2018-02-01

Von Hippel-Lindau syndrome is an autosomal-dominant disease characterized by the formation of various tumours and cysts in many different parts of the body. Von Hippel-Lindau syndrome is caused by VHL gene mutations leading to production of impaired tumor suppressor Von Hippel-Lindau syndrome protein or its complete absence. To study five patients with clinically suspected Von Hippel-Lindau syndrome, who were referred for molecular genetic testing. Sanger sequencing of the coding regions of the VHL gene. Five clinically relevant germline mutations were detected. One of the pathogenic variants has not been previously reported. This novel mutation is a complex mutation event combining a duplication and an indel, rearranging exon 3 of the VHL gene - c. [516_517dupGTCAAGCCT; 532_542delCTGGACATCGTinsATTA], p. (Glu173Serfs*4). Overall, our results showed that the diagnosis of Von Hippel-Lindau syndrome in our country is difficult most probably because of its heterogeneous clinical manifestation and insufficient knowledge on the diagnostic criteria for the disease. From genetic point of view our results add some novel data on the mutation profile of the VHL gene. In order to prove or revise the diagnosis, early genetic testing is strongly recommended in affected patients and their family members to ensure appropriate follow-up and treatment of the malignancies.

A short insertion mutation disrupts genesis of miR-16 and causes increased body weight in domesticated chicken.

PubMed

Jia, Xinzheng; Lin, Huiran; Nie, Qinghua; Zhang, Xiquan; Lamont, Susan J

2016-11-03

Body weight is one of the most important quantitative traits with high heritability in chicken. We previously mapped a quantitative trait locus (QTL) for body weight by genome-wide association study (GWAS) in an F2 chicken resource population. To identify the causal mutations linked to this QTL, expression profiles were determined on livers of high-weight and low-weight chicken lines by microarray. Combining the expression pattern with SNP effects by GWAS, miR-16 was identified as the most likely potential candidate with a 3.8-fold decrease in high-weight lines. Re-sequencing revealed that a 54-bp insertion mutation in the upstream region of miR-15a-16 displayed high allele frequencies in high-weight commercial broiler line. This mutation resulted in lower miR-16 expression by introducing three novel splicing sites instead of the missing 5' terminal splicing of mature miR-16. Elevating miR-16 significantly inhibited DF-1 chicken embryo cell proliferation, consistent with a role in suppression of cellular growth. The 54-bp insertion was significantly associated with increased body weight, bone size and muscle mass. Also, the insertion mutation tended towards fixation in commercial broilers (Fst > 0.4). Our findings revealed a novel causative mutation for body weight regulation that aids our basic understanding of growth regulation in birds.

A short insertion mutation disrupts genesis of miR-16 and causes increased body weight in domesticated chicken

PubMed Central

Jia, Xinzheng; Lin, Huiran; Nie, Qinghua; Zhang, Xiquan; Lamont, Susan J.

2016-01-01

Body weight is one of the most important quantitative traits with high heritability in chicken. We previously mapped a quantitative trait locus (QTL) for body weight by genome-wide association study (GWAS) in an F2 chicken resource population. To identify the causal mutations linked to this QTL, expression profiles were determined on livers of high-weight and low-weight chicken lines by microarray. Combining the expression pattern with SNP effects by GWAS, miR-16 was identified as the most likely potential candidate with a 3.8-fold decrease in high-weight lines. Re-sequencing revealed that a 54-bp insertion mutation in the upstream region of miR-15a-16 displayed high allele frequencies in high-weight commercial broiler line. This mutation resulted in lower miR-16 expression by introducing three novel splicing sites instead of the missing 5′ terminal splicing of mature miR-16. Elevating miR-16 significantly inhibited DF-1 chicken embryo cell proliferation, consistent with a role in suppression of cellular growth. The 54-bp insertion was significantly associated with increased body weight, bone size and muscle mass. Also, the insertion mutation tended towards fixation in commercial broilers (Fst > 0.4). Our findings revealed a novel causative mutation for body weight regulation that aids our basic understanding of growth regulation in birds. PMID:27808177

Combined Growth Hormone and Thyroid-Stimulating Hormone Deficiency in a Japanese Patient with a Novel Frameshift Mutation in IGSF1.

PubMed

Asakura, Yumi; Abe, Kiyomi; Muroya, Koji; Hanakawa, Junko; Oto, Yuji; Narumi, Satoshi; Hasegawa, Tomonobu; Adachi, Masanori

2015-01-01

Recent reports have indicated that loss-of-function mutations in the immunoglobulin superfamily member 1 gene (IGSF1, OMIM 300888) cause congenital central hypothyroidism with macroorchidism. We conducted a next-generation sequencing-based comprehensive mutation screening for pituitary hormone deficiencies to elucidate molecular mechanisms other than anatomical abnormalities of the pituitary that might be responsible for multiple anterior hormone deficiency in a male patient who originally visited our institute complaining of short stature. He was born large for gestational age (4,370 g, +3.0 SD) after an obstructed labour. Endocrinological evaluation revealed growth hormone and thyroid-stimulating hormone deficiency. Magnetic resonance imaging showed a discontinuity of the pituitary stalk with an ectopic posterior lobe and a hypoplastic anterior lobe, likely explaining multiple anterior pituitary hormone deficiency. We identified a novel hemizygous IGSF1 mutation (c.1137_1138delCA, p.Asn380Glnfs*6) in the patient. In reviewing the literature, we noticed that all reported Japanese male IGSF1 mutation carriers were born larger than mean standards for gestational age (mean birth weight SD score of +2.0, 95% confidence interval 1.0-3.0). This case suggests that more attention should be paid to intrauterine growth and birth history when patients are suspected of having an IGSF1 mutation. © 2015 S. Karger AG, Basel.

Variable expressivity of the phenotype in two families with brachydactyly type E, craniofacial dysmorphism, short stature and delayed bone age caused by novel heterozygous mutations in the PTHLH gene.

PubMed

Jamsheer, Aleksander; Sowińska-Seidler, Anna; Olech, Ewelina M; Socha, Magdalena; Kozłowski, Kazimierz; Pyrkosz, Antoni; Trzeciak, Tomasz; Materna-Kiryluk, Anna; Latos-Bieleńska, Anna

2016-05-01

Brachydactyly refers to shortening of digits due to hypoplasia or aplasia of bones forming the hands and/or feet. Isolated brachydactyly type E (BDE), which is characterized by shortened metacarpals and/or metatarsals, results in a small proportion of patients from HOXD13 or PTHLH mutations, although in the majority of cases molecular lesion remains unknown. BDE, like other brachydactylies, shows clinical heterogeneity with highly variable intrafamilial and interindividual expressivity. In this study, we investigated two Polish cases (one familial and one sporadic) presenting with BDE and additional symptoms due to novel PTHLH mutations. Apart from BDE, the affected family showed short stature, mild craniofacial dysmorphism and delayed bone age. Sanger sequencing of PTHLH revealed a novel heterozygous frameshift mutation c.258delC(p.N87Tfs*18) in two affected individuals and one relative manifesting mild brachydactyly. The sporadic patient, in addition to BDE, presented with craniofacial dysmorphism, normal stature and bone age, and was demonstrated to carry a de novo heterozygous c.166C>T(p.R56*) mutation. Our paper reports on the two novel truncating PTHLH variants, resulting in variable combination of BDE and other symptoms. Data shown here expand the knowledge on the phenotypic presentation of PTHLH mutations, highlighting significant clinical variability and incomplete penetrance of the PTHLH-related symptoms.

Preexisting MEK1 Exon 3 Mutations in V600E/KBRAF Melanomas Do Not Confer Resistance to BRAF Inhibitors

PubMed Central

Shi, Hubing; Moriceau, Gatien; Kong, Xiangju; Koya, Richard C.; Nazarian, Ramin; Pupo, Gulietta M.; Bacchiocchi, Antonella; Dahlman, Kimberly B.; Chmielowski, Bartosz; Sosman, Jeffrey A.; Halaban, Ruth; Kefford, Richard F.; Long, Georgina V.; Ribas, Antoni; Lo, Roger S.

2012-01-01

BRAF inhibitors (BRAFi) induce antitumor responses in nearly 60% of patients with advanced V600E/KBRAF melanomas. Somatic activating MEK1 mutations are thought to be rare in melanomas, but their potential concurrence with V600E/KBRAF may be selected for by BRAFi. We sequenced MEK1/2 exon 3 in melanomas at baseline and upon disease progression. Of 31 baseline V600E/KBRAF melanomas, 5 (16%) carried concurrent somatic BRAF/MEK1 activating mutations. Three of 5 patients with BRAF/MEK1 double-mutant baseline melanomas showed objective tumor responses, consistent with the overall 60% frequency. No MEK1 mutation was found in disease progression melanomas, except when it was already identified at baseline. MEK1-mutant expression in V600E/KBRAF melanoma cell lines resulted in no significant alterations in p-ERK1/2 levels or growth-inhibitory sensitivities to BRAFi, MEK1/2 inhibitor (MEKi), or their combination. Thus, activating MEK1 exon 3 mutations identified herein and concurrent with V600E/KBRAF do not cause BRAFi resistance in melanoma. SIGNIFICANCE As BRAF inhibitors gain widespread use for treatment of advanced melanoma, bio-markers for drug sensitivity or resistance are urgently needed. We identify here concurrent activating mutations in BRAF and MEK1 in melanomas and show that the presence of a downstream mutation in MEK1 does not necessarily make BRAF–mutant melanomas resistant to BRAF inhibitors. PMID:22588879

Mitochondrial DNA disease—molecular insights and potential routes to a cure

DOE Office of Scientific and Technical Information (OSTI.GOV)

Russell, Oliver; Turnbull, Doug, E-mail: doug.turnbull@newcastle.ac.uk

2014-07-01

Mitochondrial DNA diseases are common neurological conditions caused by mutations in the mitochondrial genome or nuclear genes responsible for its maintenance. Current treatments for these disorders are focussed on the management of the symptoms, rather than the correction of biochemical defects caused by the mutation. This review focuses on the molecular effects of mutations, the symptoms they cause and current work focusing on the development of targeted treatments for mitochondrial DNA disease. - Highlights: • We discuss several common disease causing mtDNA mutations. • We highlight recent work linking pathogenicity to deletion size and heteroplasmy. • We discuss recent advancesmore » in the development of targeted mtDNA disease treatments.« less

[Kenny-Caffey syndrome and its related syndromes].

PubMed

Isojima, Tsuyoshi; Kitanaka, Sachiko

2015-11-01

Kenny-Caffey syndrome (KCS) is a very rare dysmorphologic syndrome characterized by proportionate short stature, cortical thickening and medullary stenosis of tubular bones, delayed closure of anterior fontanelle, eye abnormalities, and hypoparathyroidism. Two types of KCS were known: the autosomal recessive form (KCS type 1), which is caused by mutations of the TBCE gene, and the autosomal dominant form (KCS type 2), which is caused by mutations of the FAM111A gene. TBCE mutation also causes hypoparathyroidism-retardation-dysmorphism syndrome, and FAM111A mutation also causes gracile bone dysplasia. These two diseases can be called as KCS-related syndromes. In this article, we review the clinical manifestations of KCS and discuss its related syndromes.

Combined Occurrence of Autosomal Dominant Aniridia and Autosomal Recessive Albinism in Several Members of a Family.

PubMed

Yahalom, Claudia; Sharon, Dror; Dalia, Eli; Simhon, Shiran Ben; Shemesh, Efrat; Blumenfeld, Anat

2015-06-01

To characterize clinical and genetic aspects of a family with a unique combination of two hereditary blinding eye diseases. Comprehensive eye examination of proband and family members. Molecular analyses of the TYR and PAX6 genes. A young couple, both legally blind, requested genetic counselling regarding their ocular condition. The female was previously diagnosed with oculocutaneous albinism (OCA1A) and her spouse was diagnosed with Peters anomaly. A comprehensive clinical examination revealed that the female had OCA1A combined with signs of another ocular disease, showing some similarity to aniridia. A complete ocular examination of her family members revealed that her brother also suffered from the same combined phenotype, her father had typical OCA1A signs, and her mother and sister had aniridia-like phenotype, without clinical diagnosis until the time of presentation. Molecular analysis identified two compound heterozygous TYR mutations known to cause OCAIA and cosegregate with oculocutaneous albinism. In addition, we identified a novel heterozygous PAX6 mutation confirming the atypical aniridia phenotype. We report here a unique and rare clinical phenotype that is explained by the segregation of two severe inherited eye diseases. The clinical and genetic analysis in this family allowed them to receive accurate genetic counseling.

Beneficial effects of combining nilotinib and imatinib in preclinical models of BCR-ABL+ leukemias

PubMed Central

Weisberg, Ellen; Catley, Laurie; Wright, Renee D.; Moreno, Daisy; Banerji, Lolita; Ray, Arghya; Manley, Paul W.; Mestan, Juergen; Fabbro, Doriano; Jiang, Jingrui; Hall-Meyers, Elizabeth; Callahan, Linda; DellaGatta, Jamie L.; Kung, Andrew L.

2007-01-01

Drug resistance resulting from emergence of imatinib-resistant BCR-ABL point mutations is a significant problem in advanced-stage chronic myelogenous leukemia (CML). The BCR-ABL inhibitor, nilotinib (AMN107), is significantly more potent against BCR-ABL than imatinib, and is active against many imatinib-resistant BCR-ABL mutants. Phase 1/2 clinical trials show that nilotinib can induce remissions in patients who have previously failed imatinib, indicating that sequential therapy with these 2 agents has clinical value. However, simultaneous, rather than sequential, administration of 2 BCR-ABL kinase inhibitors is attractive for many reasons, including the theoretical possibility that this could reduce emergence of drug-resistant clones. Here, we show that exposure of a variety of BCR-ABL+ cell lines to imatinib and nilotinib results in additive or synergistic cytotoxicity, including testing of a large panel of cells expressing BCR-ABL point mutations causing resistance to imatinib in patients. Further, using a highly quantifiable bioluminescent in vivo model, drug combinations were at least additive in antileukemic activity, compared with each drug alone. These results suggest that despite binding to the same site in the same target kinase, the combination of imatinib and nilotinib is highly efficacious in these models, indicating that clinical testing of combinations of BCR-ABL kinase inhibitors is warranted. PMID:17068153

Pancreatic Agenesis due to Compound Heterozygosity for a Novel Enhancer and Truncating Mutation in the PTF1A Gene.

PubMed

Gabbay, Monica; Ellard, Sian; De Franco, Elisa; Moisés, Regina S

2017-09-01

Neonatal diabetes, defined as the onset of diabetes within the first six months of life, is very rarely caused by pancreatic agenesis. Homozygous truncating mutations in the PTF1A gene, which encodes a transcriptional factor, have been reported in patients with pancreatic and cerebellar agenesis, whilst mutations located in a distal pancreatic-specific enhancer cause isolated pancreatic agenesis. We report an infant, born to healthy non-consanguineous parents, with neonatal diabetes due to pancreatic agenesis. Initial genetic investigation included sequencing of KCNJ11, ABCC8 and INS genes, but no mutations were found. Following this, 22 neonatal diabetes associated genes were analyzed by a next generation sequencing assay. We found compound heterozygous mutations in the PTF1A gene: A frameshift mutation in exon 1 (c.437_462 del, p.Ala146Glyfs*116) and a mutation affecting a highly conserved nucleotide within the distal pancreatic enhancer (g.23508442A>G). Both mutations were confirmed by Sanger sequencing. Isolated pancreatic agenesis resulting from compound heterozygosity for truncating and enhancer mutations in the PTF1A gene has not been previously reported. This report broadens the spectrum of mutations causing pancreatic agenesis.

Pancreatic Agenesis due to Compound Heterozygosity for a Novel Enhancer and Truncating Mutation in the PTF1A Gene

PubMed Central

Gabbay, Monica; Ellard, Sian; De Franco, Elisa; Moisés, Regina S.

2017-01-01

Neonatal diabetes, defined as the onset of diabetes within the first six months of life, is very rarely caused by pancreatic agenesis. Homozygous truncating mutations in the PTF1A gene, which encodes a transcriptional factor, have been reported in patients with pancreatic and cerebellar agenesis, whilst mutations located in a distal pancreatic-specific enhancer cause isolated pancreatic agenesis. We report an infant, born to healthy non-consanguineous parents, with neonatal diabetes due to pancreatic agenesis. Initial genetic investigation included sequencing of KCNJ11, ABCC8 and INS genes, but no mutations were found. Following this, 22 neonatal diabetes associated genes were analyzed by a next generation sequencing assay. We found compound heterozygous mutations in the PTF1A gene: A frameshift mutation in exon 1 (c.437_462 del, p.Ala146Glyfs*116) and a mutation affecting a highly conserved nucleotide within the distal pancreatic enhancer (g.23508442A>G). Both mutations were confirmed by Sanger sequencing. Isolated pancreatic agenesis resulting from compound heterozygosity for truncating and enhancer mutations in the PTF1A gene has not been previously reported. This report broadens the spectrum of mutations causing pancreatic agenesis. PMID:28663161

May 2006 update in porphobilinogen deaminase gene polymorphisms and mutations causing acute intermittent porphyria: comparison with the situation in Slavic population.

PubMed

Hrdinka, M; Puy, H; Martasek, P

2006-01-01

Acute intermittent porphyria (AIP) is an autosomal dominant disorder of heme biosynthesis caused by molecular defects in the porphobilinogen deaminase (PBGD) gene. This paper reviews published mutations, their types, and polymorphisms within the PBGD gene. To date, 301 different mutations and 21 polymorphisms have been identified in the PBGD gene in AIP patients and individuals from various countries and ethnic groups. During the search for mutations identified among Slavic AIP patients we found 65 such mutations and concluded that there is not a distinct predominance of certain mutations in Slavs.

Low-level APC mutational mosaicism is the underlying cause in a substantial fraction of unexplained colorectal adenomatous polyposis cases.

PubMed

Spier, Isabel; Drichel, Dmitriy; Kerick, Martin; Kirfel, Jutta; Horpaopan, Sukanya; Laner, Andreas; Holzapfel, Stefanie; Peters, Sophia; Adam, Ronja; Zhao, Bixiao; Becker, Tim; Lifton, Richard P; Perner, Sven; Hoffmann, Per; Kristiansen, Glen; Timmermann, Bernd; Nöthen, Markus M; Holinski-Feder, Elke; Schweiger, Michal R; Aretz, Stefan

2016-03-01

In 30-50% of patients with colorectal adenomatous polyposis, no germline mutation in the known genes APC, causing familial adenomatous polyposis, MUTYH, causing MUTYH-associated polyposis, or POLE or POLD1, causing polymerase-proofreading-associated polyposis can be identified, although a hereditary aetiology is likely. This study aimed to explore the impact of APC mutational mosaicism in unexplained polyposis. To comprehensively screen for somatic low-level APC mosaicism, high-coverage next-generation sequencing of the APC gene was performed using DNA from leucocytes and a total of 53 colorectal tumours from 20 unrelated patients with unexplained sporadic adenomatous polyposis. APC mosaicism was assumed if the same loss-of-function APC mutation was present in ≥ 2 anatomically separated colorectal adenomas/carcinomas per patient. All mutations were validated using diverse methods. In 25% (5/20) of patients, somatic mosaicism of a pathogenic APC mutation was identified as underlying cause of the disease. In 2/5 cases, the mosaic level in leucocyte DNA was slightly below the sensitivity threshold of Sanger sequencing; while in 3/5 cases, the allelic fraction was either very low (0.1-1%) or no mutations were detectable. The majority of mosaic mutations were located outside the somatic mutation cluster region of the gene. The present data indicate a high prevalence of pathogenic mosaic APC mutations below the detection thresholds of routine diagnostics in adenomatous polyposis, even if high-coverage sequencing of leucocyte DNA alone is taken into account. This has important implications for both routine work-up and strategies to identify new causative genes in this patient group. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/

GLUT1 deficiency syndrome as a cause of encephalopathy that includes cognitive disability, treatment-resistant infantile epilepsy and a complex movement disorder.

PubMed

Graham, John M

2012-05-01

Glucose transporter-1 (GLUT1) deficiency syndrome is caused by heterozygous mutations in the SLC2A1 gene, resulting in impaired glucose transport into the brain. It is characterized by a low glucose concentration in the cerebrospinal fluid (hypoglycorrhachia) in the absence of hypoglycemia, in combination with low to normal lactate in the cerebrospinal fluid (CSF). It often results in treatment-resistant infantile epilepsy with progressive developmental disabilities and a complex movement disorder. Recognizing GLUT1 deficiency syndrome is important, since initiation of a ketogenic diet can reduce the frequency of seizures and the severity of the movement disorder. There can be a considerable delay in diagnosing GLUT1 deficiency syndrome, and this point is illustrated by the natural history of this disorder in a 21-year-old woman with severe, progressive neurological disabilities. Her encephalopathy consisted of treatment-resistant seizures, a complex movement disorder, progressive intellectual disability, and deceleration of her head growth after late infancy. Focused evaluation at age 21 revealed GLUT1 deficiency caused by a novel heterozygous missence mutation in exon 7 (c.938C > A; p.Ser313Try) in SLC2A1 as the cause for her disabilities. Copyright © 2011 Elsevier Masson SAS. All rights reserved.

Heterogeneity of Genetic Pathways toward Daptomycin Nonsusceptibility in Staphylococcus aureus Determined by Adjunctive Antibiotics

PubMed Central

Berti, Andrew D.; Baines, Sarah L.; Howden, Benjamin P.; Sakoulas, George; Nizet, Victor; Proctor, Richard A.

2015-01-01

Daptomycin is increasingly used in combination with other antibiotics to enhance antimicrobial efficacy and/or to mitigate the emergence of daptomycin nonsusceptibility (DNS). This study used a clinical methicillin-resistant Staphylococcus aureus (MRSA) strain in which DNS emerged upon therapy to examine the influence of antibiotic combinations on the development of mutations in specific genes (mprF, rpoBC, dltA, cls2, and yycFG) previously associated with DNS. Whole genomes of bacteria obtained following 28 days of in vitro exposure to daptomycin with or without adjunctive clarithromycin, linezolid, oxacillin, or trimethoprim-sulfamethoxazole were sequenced, and the sequences were compared to that of the progenitor isolate. The addition of oxacillin to medium containing daptomycin prevented the emergence of mprF mutation but did not prevent rpoBC mutation (P < 0.01). These isolates maintained susceptibility to daptomycin during the combined exposure (median MIC, 1 mg/liter). Daptomycin plus clarithromycin or linezolid resulted in low-level (1.5 to 8 mg/liter) and high-level (12 to 96 mg/liter) DNS, respectively, and did not prevent mprF mutation. However, these same combinations prevented rpoBC mutation. Daptomycin alone or combined with linezolid or trimethoprim-sulfamethoxazole resulted in high-level DNS and mutations in mprF plus rpoBC, cls2, and yycFG. Combining daptomycin with different antimicrobials alters the mutational space available for DNS development, thereby favoring the development of predictable collateral susceptibilities. PMID:25733508

Heterogeneity of genetic pathways toward daptomycin nonsusceptibility in Staphylococcus aureus determined by adjunctive antibiotics.

PubMed

Berti, Andrew D; Baines, Sarah L; Howden, Benjamin P; Sakoulas, George; Nizet, Victor; Proctor, Richard A; Rose, Warren E

2015-05-01

Daptomycin is increasingly used in combination with other antibiotics to enhance antimicrobial efficacy and/or to mitigate the emergence of daptomycin nonsusceptibility (DNS). This study used a clinical methicillin-resistant Staphylococcus aureus (MRSA) strain in which DNS emerged upon therapy to examine the influence of antibiotic combinations on the development of mutations in specific genes (mprF, rpoBC, dltA, cls2, and yycFG) previously associated with DNS. Whole genomes of bacteria obtained following 28 days of in vitro exposure to daptomycin with or without adjunctive clarithromycin, linezolid, oxacillin, or trimethoprim-sulfamethoxazole were sequenced, and the sequences were compared to that of the progenitor isolate. The addition of oxacillin to medium containing daptomycin prevented the emergence of mprF mutation but did not prevent rpoBC mutation (P < 0.01). These isolates maintained susceptibility to daptomycin during the combined exposure (median MIC, 1 mg/liter). Daptomycin plus clarithromycin or linezolid resulted in low-level (1.5 to 8 mg/liter) and high-level (12 to 96 mg/liter) DNS, respectively, and did not prevent mprF mutation. However, these same combinations prevented rpoBC mutation. Daptomycin alone or combined with linezolid or trimethoprim-sulfamethoxazole resulted in high-level DNS and mutations in mprF plus rpoBC, cls2, and yycFG. Combining daptomycin with different antimicrobials alters the mutational space available for DNS development, thereby favoring the development of predictable collateral susceptibilities. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Brief report: EGFR L858M/L861Q cis mutations confer selective sensitivity to afatinib

PubMed Central

Saxon, Jamie A.; Sholl, Lynette M.; Jänne, Pasi A.

2017-01-01

Introduction Tyrosine kinase inhibitors (TKIs) have been developed to treat patients with epidermal growth factor receptor (EGFR)-mutant lung cancers. However, the therapeutic efficacy of TKIs in patients with uncommon EGFR mutations remains unclear. Methods Next-generation sequencing was performed on a patient’s lung adenocarcinoma tumor sample, revealing rare combined in cis (on the same allele) EGFR mutations. Stable Ba/F3 and NIH-3T3 cell lines harboring the mutations were established to investigate the effect of first, second, and third generation EGFR TKIs on cell proliferation by MTS assay and EGFR phosphorylation by Western blotting. Results EGFR L858M/L861Q mutations in cis were detected in a non-small cell lung cancer patient’s tumor. The patient demonstrated primary resistance to erlotinib and was subsequently treated with afatinib, which caused tumor regression. In in vitro studies, first and third generation TKIs exhibited a decreased capacity to prevent EGFR phosphorylation and inhibit cell proliferation in EGFR L858M/L861Q cells compared to cells harboring the common EGFR L858R point mutation. In contrast, afatinib treatment reduced proliferation and inhibited EGFR phosphorylation in L858M/L861Q and L858R mutant cells at similar concentrations. Conclusions Afatinib may be a beneficial therapeutic option for a subset of lung cancer patients with rare EGFR mutations in their tumors. Understanding how uncommon mutations affect protein structure and TKI binding will be important for identifying effective targeted therapies for these patients. PMID:28088511

Gene mutation analysis in non-small cell lung cancer patients using bronchoalveolar lavage fluid and tumor tissue as diagnostic markers.

PubMed

Li, Jian; Hu, Yi-Ming; Wang, Yi; Tang, Xing-Ping; Shi, Wei-Lin; Du, Yong-Jie

2014-12-09

Non-small cell lung cancer (NSCLC) is one of the main causes of cancer death in the world. Early detection of NSCLC can improve its outcome. The aim of this study was to identify the mutations of the KRAS and p53 genes in bronchoalveoar lavage (BAL) fluid for the early detection of peripheral NSCLC. We examined the DNA obtained from the tumor, nearby normal lung tissue, and matched BAL fluid for mutations in the KRAS and p53 genes; the material was obtained from 48 patients with peripheral NSCLC, and was analyzed by PCR-single strand conformation polymorphism and DNA sequencing. BAL fluids from 26 patients with benign lung disease were used as controls. Positive rates of KRAS and p53 mutations were distributed as follows: in NSCLC tissue, 52% and 58%; in BAL fluid of NSCLC patients, 38% and 44%; in normal lung tissue, 6% and 4%; and in BAL fluid of patients with benign lung disease, 8% and 4%. The combined detection of both KRAS and p53 mutations yielded a sensitivity of 66% for the diagnosis of peripheral NSCLC, which is markedly higher than that of cytology plus histology by first bronchoscopy (38%, p=0.008). In each patient with the 2 gene mutations in BAL fluid, mutation type and location were the same as those of the primary tumor. Our study indicates that the detection of the KRAS and p53 mutations in BAL fluids could be a helpful addition to cytology and histology examination for the diagnosis of peripheral NSCLC.

Single strand conformation polymorphism analysis of androgen receptor gene mutations in patients with androgen insensitivity syndromes: Application for diagnosis, genetic counseling, and therapy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hiort, O.; Huang, Q.; Sinnecker, G.H.G.

Recent studies indicate that mutations in the androgen receptor gene are associated with androgen insensitivity syndromes, a heterogeneous group of related disorders involving defective sexual differentiation in karyotypic males. In this report, the authors address the possibility of rapid mutational analysis of the androgen receptor gene for initial diagnosis, genetic counseling, and molecular subclassification of affected patients and their families. DNA from peripheral blood leukocytes of six patients from five families with various degrees of androgen insensitivity was studied. Exons 2 to 8 of the androgen receptor gene were analyzed using a combination of single strand conformation polymorphism analysis andmore » direct DNA sequencing. Female family members were also studied to identify heterozygote carriers. Point mutations in the AR gene were identified in all six patients, and all mutations caused amino acid substitutions. One patient with incomplete androgen insensitivity was a mosaic for the mutation. Four of the five mothers, as well as a young sister of one patient, were carriers of the mutation present in the affected child. The data show that new mutations may occur in the androgen receptor gene leading to sporadic androgen insensitivity syndrome. Molecular genetic characterization of the variant allele can serve as a primary tool for diagnosis and subsequent therapy, and can provide a basis for distinguishing heterozygous carriers in familial androgen resistance. The identification of carriers is of substantial clinical importance for genetic counseling. 29 refs., 2 figs., 1 tab.« less

Postzygotic HRAS mutation causing both keratinocytic epidermal nevus and thymoma and associated with bone dysplasia and hypophosphatemia due to elevated FGF23.

PubMed

Avitan-Hersh, Emily; Tatur, Sameh; Indelman, Margarita; Gepstein, Vardit; Shreter, Roni; Hershkovitz, Dov; Brick, Riva; Bergman, Reuven; Tiosano, Dov

2014-01-01

Epidermal nevus syndrome is a rare group of disorders characterized by the combination of congenital epidermal nevi and extracutaneous features, including skeletal, neurological, ocular, and other systemic findings. We report a case of keratinocytic epidermal nevus syndrome that includes a thymoma, bone dysplasia, and hypophosphatemia with elevated fibroblast growth factor 23 (FGF23) levels associated with postzygotic HRAS mutation. A 14-year-old boy was admitted due to recent limping. The physical examination revealed multiple right-sided linear epidermal nevi along Blaschko's lines. Magnetic resonance imaging showed cystic lesions in cervical bones and thymoma, and x-ray examination showed cystic lesions in the hands. Biochemical studies demonstrated severe hypophosphatemia, normocalcemia, high normal PTH, low 25-hydroxyvitamin D and low 1,25-dihydroxyvitamin D levels. The serum FGF23 C-terminal level was normal, but the intact FGF23 level was found to be elevated. Genetic evaluation revealed a heterozygote mutation in the HRAS gene in both the keratinocytic epidermal nevus and thymoma but not in DNA extracted from blood lymphocytes, thus establishing the mutation as postzygotic. Postzygotic mutations in HRAS lead to elevation of FGF23 levels, as found in mutated PHEX, FGF23, DMP1, and ENPP1 genes, which lead to hypophosphatemia. An identical postzygotic HRAS mutation was shown to be present in both keratinocytic epidermal nevus and thymoma and to be associated with bone lesions and hypophosphatemia due to elevated FGF23 levels. These may all be related to the HRAS mutation.

Mutation Frequency of the Major Frontotemporal Dementia Genes, MAPT, GRN and C9ORF72 in a Turkish Cohort of Dementia Patients.

PubMed

Guven, Gamze; Lohmann, Ebba; Bras, Jose; Gibbs, J Raphael; Gurvit, Hakan; Bilgic, Basar; Hanagasi, Hasmet; Rizzu, Patrizia; Heutink, Peter; Emre, Murat; Erginel-Unaltuna, Nihan; Just, Walter; Hardy, John; Singleton, Andrew; Guerreiro, Rita

2016-01-01

'Microtubule-associated protein tau' (MAPT), 'granulin' (GRN) and 'chromosome 9 open reading frame72' (C9ORF72) gene mutations are the major known genetic causes of frontotemporal dementia (FTD). Recent studies suggest that mutations in these genes may also be associated with other forms of dementia. Therefore we investigated whether MAPT, GRN and C9ORF72 gene mutations are major contributors to dementia in a random, unselected Turkish cohort of dementia patients. A combination of whole-exome sequencing, Sanger sequencing and fragment analysis/Southern blot was performed in order to identify pathogenic mutations and novel variants in these genes as well as other FTD-related genes such as the 'charged multivesicular body protein 2B' (CHMP2B), the 'FUS RNA binding protein' (FUS), the 'TAR DNA binding protein' (TARDBP), the 'sequestosome1' (SQSTM1), and the 'valosin containing protein' (VCP). We determined one pathogenic MAPT mutation (c.1906C>T, p.P636L) and one novel missense variant (c.38A>G, p.D13G). In GRN we identified a probably pathogenic TGAG deletion in the splice donor site of exon 6. Three patients were found to carry the GGGGCC expansions in the non-coding region of the C9ORF72 gene. In summary, a complete screening for mutations in MAPT, GRN and C9ORF72 genes revealed a frequency of 5.4% of pathogenic mutations in a random cohort of 93 Turkish index patients with dementia.

Pitfalls in molecular analysis for mismatch repair deficiency in a family with biallelic pms2 germline mutations.

PubMed

Leenen, C H M; Geurts-Giele, W R R; Dubbink, H J; Reddingius, R; van den Ouweland, A M; Tops, C M J; van de Klift, H M; Kuipers, E J; van Leerdam, M E; Dinjens, W N M; Wagner, A

2011-12-01

Heterozygous germline mutations in the mismatch repair (MMR) genes MLH1, MSH2, MSH6 and PMS2 cause Lynch syndrome. Biallelic mutations in the MMR genes are associated with a childhood cancer syndrome [constitutional mismatch repair deficiency (CMMR-D)]. This is predominantly characterized by hematological malignancies and tumors of the bowel and brain, often associated with signs of neurofibromatosis type 1 (NF1). Diagnostic strategies for selection of patients for MMR gene analysis include analysis of microsatellite instability (MSI) and immunohistochemical (IHC) analysis of MMR proteins in tumor tissue. We report the clinical characterization and molecular analyses of tumor specimens from a family with biallelic PMS2 germline mutations. This illustrates the pitfalls of present molecular screening strategies. Tumor tissues of five family members were analyzed for MSI and IHC. MSI was observed in only one of the analyzed tissues. However, IHC analysis of brain tumor tissue of the index patient and his sister showed absence of PMS2 expression, and germline mutation analyses showed biallelic mutations in PMS2: p.Ser46IIe and p.Pro246fs. The same heterozygous mutations were confirmed in the father and mother, respectively. These data support the conclusion that in case of a clinical phenotype of CMMR-D, it is advisable to routinely combine MSI analysis with IHC analysis for the expression of MMR proteins. With inconclusive or conflicting results, germline mutation analysis of the MMR genes should be considered after thorough counselling of the patients and/or their relatives. © 2011 John Wiley & Sons A/S.

The A31P missense mutation in cardiac myosin binding protein C alters protein structure but does not cause haploinsufficiency.

PubMed

van Dijk, Sabine J; Bezold Kooiker, Kristina; Mazzalupo, Stacy; Yang, Yuanzhang; Kostyukova, Alla S; Mustacich, Debbie J; Hoye, Elaine R; Stern, Joshua A; Kittleson, Mark D; Harris, Samantha P

2016-07-01

Mutations in MYBPC3, the gene encoding cardiac myosin binding protein C (cMyBP-C), are a major cause of hypertrophic cardiomyopathy (HCM). While most mutations encode premature stop codons, missense mutations causing single amino acid substitutions are also common. Here we investigated effects of a single proline for alanine substitution at amino acid 31 (A31P) in the C0 domain of cMyBP-C, which was identified as a natural cause of HCM in cats. Results using recombinant proteins showed that the mutation disrupted C0 structure, altered sensitivity to trypsin digestion, and reduced recognition by an antibody that preferentially recognizes N-terminal domains of cMyBP-C. Western blots detecting A31P cMyBP-C in myocardium of cats heterozygous for the mutation showed a reduced amount of A31P mutant protein relative to wild-type cMyBP-C, but the total amount of cMyBP-C was not different in myocardium from cats with or without the A31P mutation indicating altered rates of synthesis/degradation of A31P cMyBP-C. Also, the mutant A31P cMyBP-C was properly localized in cardiac sarcomeres. These results indicate that reduced protein expression (haploinsufficiency) cannot account for effects of the A31P cMyBP-C mutation and instead suggest that the A31P mutation causes HCM through a poison polypeptide mechanism that disrupts cMyBP-C or myocyte function. Copyright © 2016 Elsevier Inc. All rights reserved.

Determining the Location of DNA Modification and Mutation Caused by UVB Light in Skin Cancer

DTIC Science & Technology

2013-09-01

we obtain cleavage patterns consistent with the administered UV dosage and that sequencing libraries generated for both yeast and human cells show...understanding the mutations they cause. 15. SUBJECT TERMS UV DNA modification, HeLa cells, Skin Cancer 16. SECURITY CLASSIFICATION OF: 17...of mutations that are caused by UV light in cells and correlate them to modification frequencies. Understanding the initial chemical changes

MITOCHONDRIAL DNA DEPLETION SYNDROME DUE TO MUTATIONS IN THE RRM2B GENE

PubMed Central

Bornstein, Belén; Area, Estela; Flanigan, Kevin M.; Ganesh, Jaya; Jayakar, Parul; Swoboda, Kathryn J.; Coku, Jorida; Naini, Ali; Shanske, Sara; Tanji, Kurenai; Hirano, Michio; DiMauro, Salvatore

2014-01-01

Mitochondrial DNA depletion syndrome (MDS) is characterized by a reduction in mtDNA copy number and has been associated with mutations in eight nuclear genes, including enzymes involved in mitochondrial nucleotide metabolism (POLG, TK2, DGUOK, SUCLA2, SUCLG1, PEO1) and MPV17. Recently, mutations in The RRM2B gene, encoding the p53-controlled ribonucleotide reductase subunit, have been described in 7 infants from 4 families, who presented with various combinations of hypotonia, tubulopathy, seizures, respiratory distress, diarrhea, and lactic acidosis. All children died before 4 months of age. We sequenced the RRM2B gene in three unrelated cases with unexplained severe mtDNA depletion. The first patient developed intractable diarrhea, profound weakness, respiratory distress, and died at three months. The other two unrelated patients had a much milder phenotype and are still alive at ages 27 and 36 months. All three patients had lactic acidosis and severe depletion of mtDNA in muscle. Muscle histochemistry showed RRF and COX deficiency. Sequencing the RRM2B gene revealed three missense mutations and two single nucleotide deletions in exon 6, 8 and 9, confirming that RRM2B mutations are important causes of MDS and that the clinical phenotype is heterogeneous and not invariably fatal in infancy. PMID:18504129

Mitochondrial DNA depletion syndrome due to mutations in the RRM2B gene.

PubMed

Bornstein, Belén; Area, Estela; Flanigan, Kevin M; Ganesh, Jaya; Jayakar, Parul; Swoboda, Kathryn J; Coku, Jorida; Naini, Ali; Shanske, Sara; Tanji, Kurenai; Hirano, Michio; DiMauro, Salvatore

2008-06-01

Mitochondrial DNA depletion syndrome (MDS) is characterized by a reduction in mtDNA copy number and has been associated with mutations in eight nuclear genes, including enzymes involved in mitochondrial nucleotide metabolism (POLG, TK2, DGUOK, SUCLA2, SUCLG1, PEO1) and MPV17. Recently, mutations in the RRM2B gene, encoding the p53-controlled ribonucleotide reductase subunit, have been described in seven infants from four families, who presented with various combinations of hypotonia, tubulopathy, seizures, respiratory distress, diarrhea, and lactic acidosis. All children died before 4 months of age. We sequenced the RRM2B gene in three unrelated cases with unexplained severe mtDNA depletion. The first patient developed intractable diarrhea, profound weakness, respiratory distress, and died at 3 months. The other two unrelated patients had a much milder phenotype and are still alive at ages 27 and 36 months. All three patients had lactic acidosis and severe depletion of mtDNA in muscle. Muscle histochemistry showed RRF and COX deficiency. Sequencing the RRM2B gene revealed three missense mutations and two single nucleotide deletions in exons 6, 8, and 9, confirming that RRM2B mutations are important causes of MDS and that the clinical phenotype is heterogeneous and not invariably fatal in infancy.

Studying phenotypic evolution in domestic animals: a walk in the footsteps of Charles Darwin.

PubMed

Andersson, L

2009-01-01

Charles Darwin used domesticated plants and animals as proof of principle for his theory on phenotypic evolution by means of natural selection. Inspired by Darwin's work, we developed an intercross between the wild boar and domestic pigs to study the genetic basis for phenotypic changes during domestication. The difference in coat color is controlled by two major loci. Dominant white color is due to two consecutive mutations in the KIT gene: a 450-kb duplication and a splice mutation. Black spotting is caused by the combined effect of two mutations in MC1R: a missense mutation for dominant black color and a 2-bp insertion leading to a frameshift. A major discovery made using this pedigree is the identification of a single-nucleotide substitution in intron 3 of the gene for insulin-like growth factor 2 (IGF2) that is underlying a quantitative trait locus affecting muscle growth, size of the heart, and fat deposition. The mutation disrupts the interaction with a repressor and leads to threefold increased IGF2 expression in postnatal muscle. In a recent study, we have identified the IGF2 repressor, and this previously unknown protein, named ZBED6, is specific for placental mammals and derived from a domesticated DNA transposon.

[Familial male-limited precocious puberty due to Asp578His mutations in the LHCGR gene: clinical characteristics and gene analysis in an infant].

PubMed

Wang, Min; Li, Min; Liu, Yue-Sheng; Lei, Si-Min; Xiao, Yan-Feng

2017-11-01

The aim of the study was to provide a descriptive analysis of familial male-limited precocious puberty (FMPP), which is a rare inherited disease caused by heterozygous constitutively activating mutations of the luteinizing hormone/choriogonadotropin receptor gene (LHCGR). The patient was a ten-month-old boy, presenting with penile enlargement, pubic hair formation, and spontaneous erections. Based on the clinical manifestations and laboratory data, including sexual characteristics, serum testosterone levels, GnRH stimulation test, and bone age, this boy was diagnosed with peripheral precocious puberty. Subsequently the precocious puberty-related genes were analyzed by direct DNA sequencing of amplified PCR products from the patient and his parents. Genetic analysis revealed a novel heterozygous missense mutation c.1732G>C (Asp578His) of the LHCGR gene exon11 in the patient, which had never been reported. His parents had no mutations. After combined treatment with aromatase inhibitor letrozole and anti-androgen spironolactone for six months, the patient's symptoms were controlled. The findings in this study expand the mutation spectrum of the LHCGR gene, and provide molecular evidence for the etiologic diagnosis as well as for the genetic counseling and prenatal diagnosis in the family.

Novel deletions involving the USH2A gene in patients with Usher syndrome and retinitis pigmentosa

PubMed Central

García-García, Gema; Jaijo, Teresa; Aparisi, Maria J.; Larrieu, Lise; Faugère, Valérie; Blanco-Kelly, Fiona; Ayuso, Carmen; Roux, Anne-Francoise; Millán, José M.

2014-01-01

Purpose The aim of the present work was to identify and characterize large rearrangements involving the USH2A gene in patients with Usher syndrome and nonsyndromic retinitis pigmentosa. Methods The multiplex ligation-dependent probe amplification (MLPA) technique combined with a customized array-based comparative genomic hybridization (aCGH) analysis was applied to 40 unrelated patients previously screened for point mutations in the USH2A gene in which none or only one pathologic mutation was identified. Results We detected six large deletions involving USH2A in six out of the 40 cases studied. Three of the patients were homozygous for the deletion, and the remaining three were compound heterozygous with a previously identified USH2A point mutation. In five of these cases, the patients displayed Usher type 2, and the remaining case displayed nonsyndromic retinitis pigmentosa. The exact breakpoint junctions of the deletions found in USH2A in four of these cases were characterized. Conclusions Our study highlights the need to develop improved efficient strategies of mutation screening based upon next generation sequencing (NGS) that reduce cost, time, and complexity and allow simultaneous identification of all types of disease-causing mutations in diagnostic procedures. PMID:25352746

Effect of the G375C and G346E achondroplasia mutations on FGFR3 activation.

PubMed

He, Lijuan; Serrano, Christopher; Niphadkar, Nitish; Shobnam, Nadia; Hristova, Kalina

2012-01-01

Two mutations in FGFR3, G380R and G375C are known to cause achondroplasia, the most common form of human dwarfism. The G380R mutation accounts for 98% of the achondroplasia cases, and thus has been studied extensively. Here we study the effect of the G375C mutation on the phosphorylation and the cross-linking propensity of full-length FGFR3 in HEK 293 cells, and we compare the results to previously published results for the G380R mutant. We observe identical behavior of the two achondroplasia mutants in these experiments, a finding which supports a direct link between the severity of dwarfism phenotypes and the level and mechanism of FGFR3 over-activation. The mutations do not increase the cross-linking propensity of FGFR3, contrary to previous expectations that the achondroplasia mutations stabilize the FGFR3 dimers. Instead, the phosphorylation efficiency within un-liganded FGFR3 dimers is increased, and this increase is likely the underlying cause for pathogenesis in achondroplasia. We further investigate the G346E mutation, which has been reported to cause achondroplasia in one case. We find that this mutation does not increase FGFR3 phosphorylation and decreases FGFR3 cross-linking propensity, a finding which raises questions whether this mutation is indeed a genetic cause for human dwarfism.

Effect of the G375C and G346E Achondroplasia Mutations on FGFR3 Activation

PubMed Central

He, Lijuan; Serrano, Christopher; Niphadkar, Nitish; Shobnam, Nadia; Hristova, Kalina

2012-01-01

Two mutations in FGFR3, G380R and G375C are known to cause achondroplasia, the most common form of human dwarfism. The G380R mutation accounts for 98% of the achondroplasia cases, and thus has been studied extensively. Here we study the effect of the G375C mutation on the phosphorylation and the cross-linking propensity of full-length FGFR3 in HEK 293 cells, and we compare the results to previously published results for the G380R mutant. We observe identical behavior of the two achondroplasia mutants in these experiments, a finding which supports a direct link between the severity of dwarfism phenotypes and the level and mechanism of FGFR3 over-activation. The mutations do not increase the cross-linking propensity of FGFR3, contrary to previous expectations that the achondroplasia mutations stabilize the FGFR3 dimers. Instead, the phosphorylation efficiency within un-liganded FGFR3 dimers is increased, and this increase is likely the underlying cause for pathogenesis in achondroplasia. We further investigate the G346E mutation, which has been reported to cause achondroplasia in one case. We find that this mutation does not increase FGFR3 phosphorylation and decreases FGFR3 cross-linking propensity, a finding which raises questions whether this mutation is indeed a genetic cause for human dwarfism. PMID:22529939

Combining Protein and Strain Engineering for the Production of Glyco-Engineered Horseradish Peroxidase C1A in Pichia pastoris

PubMed Central

Capone, Simona; Ćorajević, Lejla; Bonifert, Günther; Murth, Patrick; Maresch, Daniel; Altmann, Friedrich; Herwig, Christoph; Spadiut, Oliver

2015-01-01

Horseradish peroxidase (HRP), conjugated to antibodies and lectins, is widely used in medical diagnostics. Since recombinant production of the enzyme is difficult, HRP isolated from plant is used for these applications. Production in the yeast Pichia pastoris (P. pastoris), the most promising recombinant production platform to date, causes hyperglycosylation of HRP, which in turn complicates conjugation to antibodies and lectins. In this study we combined protein and strain engineering to obtain an active and stable HRP variant with reduced surface glycosylation. We combined four mutations, each being beneficial for either catalytic activity or thermal stability, and expressed this enzyme variant as well as the unmutated wildtype enzyme in both a P. pastoris benchmark strain and a strain where the native α-1,6-mannosyltransferase (OCH1) was knocked out. Considering productivity in the bioreactor as well as enzyme activity and thermal stability, the mutated HRP variant produced in the P. pastoris benchmark strain turned out to be interesting for medical diagnostics. This variant shows considerable catalytic activity and thermal stability and is less glycosylated, which might allow more controlled and efficient conjugation to antibodies and lectins. PMID:26404235

Intronic splicing mutations in PTCH1 cause Gorlin syndrome.

PubMed

Bholah, Zaynab; Smith, Miriam J; Byers, Helen J; Miles, Emma K; Evans, D Gareth; Newman, William G

2014-09-01

Gorlin syndrome is an autosomal dominant disorder characterized by multiple early-onset basal cell carcinoma, odontogenic keratocysts and skeletal abnormalities. It is caused by heterozygous mutations in the tumour suppressor PTCH1. Routine clinical genetic testing, by Sanger sequencing and multiplex ligation-dependent probe amplification (MLPA) to confirm a clinical diagnosis of Gorlin syndrome, identifies a mutation in 60-90 % of cases. We undertook RNA analysis on lymphocytes from ten individuals diagnosed with Gorlin syndrome, but without known PTCH1 mutations by exonic sequencing or MLPA. Two altered PTCH1 transcripts were identified. Genomic DNA sequence analysis identified an intron 7 mutation c.1068-10T>A, which created a strong cryptic splice acceptor site, leading to an intronic insertion of eight bases; this is predicted to create a frameshift p.(His358Alafs*12). Secondly, a deep intronic mutation c.2561-2057A>G caused an inframe insertion of 78 intronic bases in the cDNA transcript, leading to a premature stop codon p.(Gly854fs*3). The mutations are predicted to cause loss of function of PTCH1, consistent with its tumour suppressor function. The findings indicate the importance of RNA analysis to detect intronic mutations in PTCH1 not identified by routine screening techniques.

FOXC2 disease-mutations identified in lymphedema-distichiasis patients cause both loss and gain of protein function

PubMed Central

Tavian, Daniela; Missaglia, Sara; Maltese, Paolo E.; Michelini, Sandro; Fiorentino, Alessandro; Ricci, Maurizio; Serrani, Roberta; Walter, Michael A.; Bertelli, Matteo

2016-01-01

Dominant mutations in the FOXC2 gene cause a form of lymphedema primarily of the limbs that usually develops at or after puberty. In 90-95% of patients, lymphedema is accompanied by distichiasis. FOXC2 is a member of the forkhead/winged-helix family of transcription factors and plays essential roles in different developmental pathways and physiological processes. We previously described six unrelated families with primary lymphedema-distichiasis in which patients showed different FOXC2 mutations located outside of the forkhead domain. Of those, four were missense mutations, one a frameshift mutation, and the last a stop mutation. To assess their pathogenic potential, we have now examined the subcellular localization and the transactivation activity of the mutated FOXC2 proteins. All six FOXC2 mutant proteins were able to localize into the nucleus; however, the frameshift truncated protein appeared to be sequestered into nuclear aggregates. A reduction in the ability to activate FOXC1/FOXC2 response elements was detected in 50% of mutations, while the remaining ones caused an increase of protein transactivation activity. Our data reveal that either a complete loss or a significant gain of FOXC2 function can cause a perturbation of lymphatic vessel formation leading to lymphedema. PMID:27276711

Primary Ciliary Dyskinesia Caused by Homozygous Mutation in DNAL1, Encoding Dynein Light Chain 1

PubMed Central

Mazor, Masha; Alkrinawi, Soliman; Chalifa-Caspi, Vered; Manor, Esther; Sheffield, Val C.; Aviram, Micha; Parvari, Ruti

2011-01-01

In primary ciliary dyskinesia (PCD), genetic defects affecting motility of cilia and flagella cause chronic destructive airway disease, randomization of left-right body asymmetry, and, frequently, male infertility. The most frequent defects involve outer and inner dynein arms (ODAs and IDAs) that are large multiprotein complexes responsible for cilia-beat generation and regulation, respectively. Although it has long been suspected that mutations in DNAL1 encoding the ODA light chain1 might cause PCD such mutations were not found. We demonstrate here that a homozygous point mutation in this gene is associated with PCD with absent or markedly shortened ODA. The mutation (NM_031427.3: c.449A>G; p.Asn150Ser) changes the Asn at position150, which is critical for the proper tight turn between the β strand and the α helix of the leucine-rich repeat in the hydrophobic face that connects to the dynein heavy chain. The mutation reduces the stability of the axonemal dynein light chain 1 and damages its interactions with dynein heavy chain and with tubulin. This study adds another important component to understanding the types of mutations that cause PCD and provides clinical information regarding a specific mutation in a gene not yet known to be associated with PCD. PMID:21496787

Distinct effects of tubulin isotype mutations on neurite growth in Caenorhabditis elegans

PubMed Central

Zheng, Chaogu; Diaz-Cuadros, Margarete; Nguyen, Ken C. Q.; Hall, David H.; Chalfie, Martin

2017-01-01

Tubulins, the building block of microtubules (MTs), play a critical role in both supporting and regulating neurite growth. Eukaryotic genomes contain multiple tubulin isotypes, and their missense mutations cause a range of neurodevelopmental defects. Using the Caenorhabditis elegans touch receptor neurons, we analyzed the effects of 67 tubulin missense mutations on neurite growth. Three types of mutations emerged: 1) loss-of-function mutations, which cause mild defects in neurite growth; 2) antimorphic mutations, which map to the GTP binding site and intradimer and interdimer interfaces, significantly reduce MT stability, and cause severe neurite growth defects; and 3) neomorphic mutations, which map to the exterior surface, increase MT stability, and cause ectopic neurite growth. Structure-function analysis reveals a causal relationship between tubulin structure and MT stability. This stability affects neuronal morphogenesis. As part of this analysis, we engineered several disease-associated human tubulin mutations into C. elegans genes and examined their impact on neuronal development at the cellular level. We also discovered an α-tubulin (TBA-7) that appears to destabilize MTs. Loss of TBA-7 led to the formation of hyperstable MTs and the generation of ectopic neurites; the lack of potential sites for polyamination and polyglutamination on TBA-7 may be responsible for this destabilization. PMID:28835377

A novel AMELX mutation causes hypoplastic amelogenesis imperfecta.

PubMed

Kim, Young-Jae; Kim, Youn Jung; Kang, Jenny; Shin, Teo Jeon; Hyun, Hong-Keun; Lee, Sang-Hoon; Lee, Zang Hee; Kim, Jung-Wook

2017-04-01

Amelogenesis imperfecta (AI) is a hereditary genetic defect affecting tooth enamel. AI is heterogeneous in clinical phenotype as well as in genetic etiology. To date, more than 10 genes have been associated with the etiology of AI. Amelogenin is the most abundant enamel matrix protein, most of which is encoded by the amelogenin gene in the X-chromosome (AMELX). More than 16 alternative splicing transcripts have been identified in the murine Amelx gene. The purpose of this study was to identify the genetic cause of an AI family. We recruited a family with hypoplastic AI and performed mutational analysis on the candidate gene based on the clinical phenotype. Mutational analysis revealed a missense mutation in exon 6 (NM_182680.1; c.242C > T), which changes a sequence in a highly conserved amino acid (NP_872621.1; p.Pro81Leu). Furthermore, a splicing assay using a minigene displayed that the mutation changed the mRNA splicing repertory. In this study, we identified a novel AMELX missense mutation causing hypoplastic AI, and this mutation also resulted in altered mRNA splicing. These results will not only expand the mutation spectrum causing AI but also broaden our understanding of the biological mechanism of enamel formation. Copyright © 2017 Elsevier Ltd. All rights reserved.

Activating mutations affecting the Dbl homology domain of SOS2 cause Noonan syndrome

PubMed Central

Cordeddu, Viviana; Yin, Jiani C.; Gunnarsson, Cecilia; Virtanen, Carl; Drunat, Séverine; Lepri, Francesca; De Luca, Alessandro; Rossi, Cesare; Ciolfi, Andrea; Pugh, Trevor J.; Bruselles, Alessandro; Priest, James R.; Pennacchio, Len A.; Lu, Zhibin; Danesh, Arnavaz; Quevedo, Rene; Hamid, Alaa; Martinelli, Simone; Pantaleoni, Francesca; Gnazzo, Maria; Daniele, Paola; Lissewski, Christina; Bocchinfuso, Gianfranco; Stella, Lorenzo; Odent, Sylvie; Philip, Nicole; Faivre, Laurence; Vlckova, Marketa; Seemanova, Eva; Digilio, Cristina; Zenker, Martin; Zampino, Giuseppe; Verloes, Alain; Dallapiccola, Bruno; Roberts, Amy E.; Cavé, Hélène; Gelb, Bruce D.; Neel, Benjamin G.; Tartaglia, Marco

2015-01-01

The RASopathies constitute a family of autosomal dominant disorders whose major features include facial dysmorphism, cardiac defects, reduced postnatal growth, variable cognitive deficits, ectodermal and skeletal anomalies, and susceptibility to certain malignancies. Noonan syndrome (NS), the commonest RASopathy, is genetically heterogeneous and caused by functional dysregulation of signal transducers and regulatory proteins with roles in the RAS/extracellular signal-regulated kinase (ERK) signal transduction pathway. Mutations in known disease genes account for approximately 80% of affected individuals. Here, we report that missense mutations altering son of sevenless, Drosophila, homolog 2 (SOS2), which encodes a RAS guanine nucleotide exchange factor, occur in a small percentage of subjects with NS. Four missense mutations were identified in five unrelated sporadic cases and families transmitting NS. Disease-causing mutations affected three conserved residues located in the Dbl homology domain, of which two are directly involved in the intramolecular binding network maintaining SOS2 in its auto-inhibited conformation. All mutations were found to promote enhanced signaling from RAS to ERK. Similar to NS-causing SOS1 mutations, the phenotype associated with SOS2 defects is characterized by normal development and growth, as well as marked ectodermal involvement. Unlike SOS1 mutations, however, those in SOS2 are restricted to the Dbl homology domain. PMID:26173643

Generation and analysis of knock-in mice carrying pseudohypoaldosteronism type II-causing mutations in the cullin 3 gene.

PubMed

Araki, Yuya; Rai, Tatemitsu; Sohara, Eisei; Mori, Takayasu; Inoue, Yuichi; Isobe, Kiyoshi; Kikuchi, Eriko; Ohta, Akihito; Sasaki, Sei; Uchida, Shinichi

2015-10-21

Pseudohypoaldosteronism type II (PHAII) is a hereditary hypertensive disease caused by mutations in four different genes: with-no-lysine kinases (WNK) 1 and 4, Kelch-like family member 3 (KLHL3), and cullin 3 (Cul3). Cul3 and KLHL3 form an E3 ligase complex that ubiquitinates and reduces the expression level of WNK4. PHAII-causing mutations in WNK4 and KLHL3 impair WNK4 ubiquitination. However, the molecular pathogenesis of PHAII caused by Cul3 mutations is unclear. In cultured cells and human leukocytes, PHAII-causing Cul3 mutations result in the skipping of exon 9, producing mutant Cul3 protein lacking 57 amino acids. However, whether this phenomenon occurs in the kidneys and is responsible for the pathogenesis of PHAII in vivo is unknown. We generated knock-in mice carrying a mutation in the C-terminus of intron 8 of Cul3, c.1207-1G>A, which corresponds to a PHAII-causing mutation in the human Cul3 gene. Heterozygous Cul3(G(-1)A/+) knock-in mice did not exhibit PHAII phenotypes, and the skipping of exon 9 was not evident in their kidneys. However, the level of Cul3 mRNA expression in the kidneys of heterozygous knock-in mice was approximately half that of wild-type mice. Furthermore, homozygous knock-in mice were nonviable. It suggested that the mutant allele behaved like a knockout allele and did not produce Cul3 mRNA lacking exon 9. A reduction in Cul3 expression alone was not sufficient to develop PHAII in the knock-in mice. Our findings highlighted the pathogenic role of mutant Cul3 protein and provided insight to explain why PHAII-causing mutations in Cul3 cause kidney-predominant PHAII phenotypes. © 2015. Published by The Company of Biologists Ltd.

Mutations in the G6PC3 gene cause Dursun syndrome.

PubMed

Banka, Siddharth; Newman, William G; Ozgül, R Koksal; Dursun, Ali

2010-10-01

Dursun syndrome is a triad of familial primary pulmonary hypertension, leucopenia, and atrial septal defect. Here we demonstrate that mutations in G6PC3 cause Dursun syndrome. Mutations in G6PC3 are known to also cause severe congenital neutropenia type 4. Identification of the genetic basis of Dursun syndrome expands the pre-existing knowledge about the phenotypic effects of mutations in G6PC3. We propose that Dursun syndrome should now be considered as a subset of severe congenital neutropenia type 4 with pulmonary hypertension as an important clinical feature. Copyright © 2010 Wiley-Liss, Inc.

Prevalence and penetrance of germline BRCA1 and BRCA2 mutations in a population series of 649 women with ovarian cancer.

PubMed

Risch, H A; McLaughlin, J R; Cole, D E; Rosen, B; Bradley, L; Kwan, E; Jack, E; Vesprini, D J; Kuperstein, G; Abrahamson, J L; Fan, I; Wong, B; Narod, S A

2001-03-01

A population-based series of 649 unselected incident cases of ovarian cancer diagnosed in Ontario, Canada, during 1995-96 was screened for germline mutations in BRCA1 and BRCA2. We specifically tested for 11 of the most commonly reported mutations in the two genes. Then, cases were assessed with the protein-truncation test (PTT) for exon 11 of BRCA1, with denaturing gradient gel electrophoresis for the remainder of BRCA1, and with PTT for exons 10 and 11 of BRCA2. No mutations were found in all 134 women with tumors of borderline histology. Among the 515 women with invasive cancers, we identified 60 mutations, 39 in BRCA1 and 21 in BRCA2. The total mutation frequency among women with invasive cancers, 11.7% (95% confidence interval [95%CI] 9.2%-14.8%), is higher than previous estimates. Hereditary ovarian cancers diagnosed at age <50 years were mostly (83%) due to BRCA1, whereas the majority (60%) of those diagnosed at age >60 years were due to BRCA2. Mutations were found in 19% of women reporting first-degree relatives with breast or ovarian cancer and in 6.5% of women with no affected first-degree relatives. Risks of ovarian, breast, and stomach cancers and leukemias/lymphomas were increased nine-, five-, six- and threefold, respectively, among first-degree relatives of cases carrying BRCA1 mutations, compared with relatives of noncarriers, and risk of colorectal cancer was increased threefold for relatives of cases carrying BRCA2 mutations. For carriers of BRCA1 mutations, the estimated penetrance by age 80 years was 36% for ovarian cancer and 68% for breast cancer. In breast-cancer risk for first-degree relatives, there was a strong trend according to mutation location along the coding sequence of BRCA1, with little evidence of increased risk for mutations in the 5' fifth, but 8.8-fold increased risk for mutations in the 3' fifth (95%CI 3.6-22.0), corresponding to a carrier penetrance of essentially 100%. Ovarian, colorectal, stomach, pancreatic, and prostate cancer occurred among first-degree relatives of carriers of BRCA2 mutations only when mutations were in the ovarian cancer-cluster region (OCCR) of exon 11, whereas an excess of breast cancer was seen when mutations were outside the OCCR. For cancers of all sites combined, the estimated penetrance of BRCA2 mutations was greater for males than for females, 53% versus 38%. Past studies may have underestimated the contribution of BRCA2 to ovarian cancer, because mutations in this gene cause predominantly late-onset cancer, and previous work has focused more on early-onset disease. If confirmed in future studies, the trend in breast-cancer penetrance, according to mutation location along the BRCA1 coding sequence, may have significant impact on treatment decisions for carriers of BRCA1-mutations. As well, BRCA2 mutations may prove to be a greater cause of cancer in male carriers than previously has been thought.

Loss of GATA-1 Full Length as a Cause of Diamond–Blackfan Anemia Phenotype

PubMed Central

Parrella, Sara; Aspesi, Anna; Quarello, Paola; Garelli, Emanuela; Pavesi, Elisa; Carando, Adriana; Nardi, Margherita; Ellis, Steven R.; Ramenghi, Ugo; Dianzani, Irma

2015-01-01

Mutations in the hematopoietic transcription factor GATA-1 alter the proliferation/differentiation of hemopoietic progenitors. Mutations in exon 2 interfere with the synthesis of the full-length isoform of GATA-1 and lead to the production of a shortened isoform, GATA-1s. These mutations have been found in patients with Diamond–Blackfan anemia (DBA), a congenital erythroid aplasia typically caused by mutations in genes encoding ribosomal proteins. We sequenced GATA-1 in 23 patients that were negative for mutations in the most frequently mutated DBA genes. One patient showed a c.2T > C mutation in the initiation codon leading to the loss of the full-length GATA-1 isoform. PMID:24453067

Frequency of rare mutations and common genetic variations in severe hypertriglyceridemia in the general population of Spain.

PubMed

Lamiquiz-Moneo, Itziar; Blanco-Torrecilla, Cristian; Bea, Ana M; Mateo-Gallego, Rocío; Pérez-Calahorra, Sofía; Baila-Rueda, Lucía; Cenarro, Ana; Civeira, Fernando; de Castro-Orós, Isabel

2016-04-23

Hypertriglyceridemia (HTG) is a common complex metabolic trait that results of the accumulation of relatively common genetic variants in combination with other modifier genes and environmental factors resulting in increased plasma triglyceride (TG) levels. The majority of severe primary hypertriglyceridemias is diagnosed in adulthood and their molecular bases have not been fully defined yet. The prevalence of HTG is highly variable among populations, possibly caused by differences in environmental factors and genetic background. However, the prevalence of very high TG and the frequency of rare mutations causing HTG in a whole non-selected population have not been previously studied. The total of 23,310 subjects over 18 years from a primary care-district in a middle-class area of Zaragoza (Spain) with TG >500 mg/dL were selected to establish HTG prevalence. Those affected of primary HTG were considered for further genetic analysis. The promoters, coding regions and exon-intron boundaries of LPL, LMF1, APOC2, APOA5, APOE and GPIHBP1 genes were sequenced. The frequency of rare variants identified was studied in 90 controls. One hundred ninety-four subjects (1.04%) had HTG and 90 subjects (46.4%) met the inclusion criteria for primary HTG. In this subgroup, nine patients (12.3%) were carriers of 7 rare variants in LPL, LMF1, APOA5, GPIHBP1 or APOE genes. Three of these mutations are described for the first time in this work. The presence of a rare pathogenic mutation did not confer a differential phenotype or a higher family history of HTG. The prevalence of rare mutations in candidate genes in subjects with primary HTG is low. The low frequency of rare mutations, the absence of a more severe phenotype or the dominant transmission of the HTG would not suggest the use of genetic analysis in the clinical practice in this population.

Tracking the origins and drivers of subclonal metastatic expansion in prostate cancer

DOE PAGES

Hong, Matthew K. H.; Macintyre, Geoff; Wedge, David C.; ...

2015-04-01

Tumour heterogeneity in primary prostate cancer is a well-established phenomenon. However, how the subclonal diversity of tumours changes during metastasis and progression to lethality is poorly understood. Here we reveal the precise direction of metastatic spread across four lethal prostate cancer patients using whole-genome and ultra-deep targeted sequencing of longitudinally collected primary and metastatic tumours. We find one case of metastatic spread to the surgical bed causing local recurrence, and another case of cross-metastatic site seeding combining with dynamic remoulding of subclonal mixtures in response to therapy. By ultra-deep sequencing end-stage blood, we detect both metastatic and primary tumour clones,more » even years after removal of the prostate. As a result, analysis of mutations associated with metastasis reveals an enrichment of TP53 mutations, and additional sequencing of metastases from 19 patients demonstrates that acquisition of TP53 mutations is linked with the expansion of subclones with metastatic potential which we can detect in the blood.« less

Generation of the SCN1A epilepsy mutation in hiPS cells using the TALEN technique

NASA Astrophysics Data System (ADS)

Chen, Wanjuan; Liu, Jingxin; Zhang, Longmei; Xu, Huijuan; Guo, Xiaogang; Deng, Sihao; Liu, Lipeng; Yu, Daiguan; Chen, Yonglong; Li, Zhiyuan

2014-06-01

Human induced pluripotent stem cells (iPSC) can be used to understand the pathological mechanisms of human disease. These cells are a promising source for cell-replacement therapy. However, such studies require genetically defined conditions. Such genetic manipulations can be performed using the novel Transcription Activator-Like Effector Nucleases (TALENs), which generate site-specific double-strand DNA breaks (DSBs) with high efficiency and precision. Combining the TALEN and iPSC methods, we developed two iPS cell lines by generating the point mutation A5768G in the SCN1A gene, which encodes the voltage-gated sodium channel Nav1.1 α subunit. The engineered iPSC maintained pluripotency and successfully differentiated into neurons with normal functional characteristics. The two cell lines differ exclusively at the epilepsy-susceptibility variant. The ability to robustly introduce disease-causing point mutations in normal hiPS cell lines can be used to generate a human cell model for studying epileptic mechanisms and for drug screening.

Tracking the origins and drivers of subclonal metastatic expansion in prostate cancer.

PubMed

Hong, Matthew K H; Macintyre, Geoff; Wedge, David C; Van Loo, Peter; Patel, Keval; Lunke, Sebastian; Alexandrov, Ludmil B; Sloggett, Clare; Cmero, Marek; Marass, Francesco; Tsui, Dana; Mangiola, Stefano; Lonie, Andrew; Naeem, Haroon; Sapre, Nikhil; Phal, Pramit M; Kurganovs, Natalie; Chin, Xiaowen; Kerger, Michael; Warren, Anne Y; Neal, David; Gnanapragasam, Vincent; Rosenfeld, Nitzan; Pedersen, John S; Ryan, Andrew; Haviv, Izhak; Costello, Anthony J; Corcoran, Niall M; Hovens, Christopher M

2015-04-01

Tumour heterogeneity in primary prostate cancer is a well-established phenomenon. However, how the subclonal diversity of tumours changes during metastasis and progression to lethality is poorly understood. Here we reveal the precise direction of metastatic spread across four lethal prostate cancer patients using whole-genome and ultra-deep targeted sequencing of longitudinally collected primary and metastatic tumours. We find one case of metastatic spread to the surgical bed causing local recurrence, and another case of cross-metastatic site seeding combining with dynamic remoulding of subclonal mixtures in response to therapy. By ultra-deep sequencing end-stage blood, we detect both metastatic and primary tumour clones, even years after removal of the prostate. Analysis of mutations associated with metastasis reveals an enrichment of TP53 mutations, and additional sequencing of metastases from 19 patients demonstrates that acquisition of TP53 mutations is linked with the expansion of subclones with metastatic potential which we can detect in the blood.

Missense variants in plakophilin-2 in arrhythmogenic right ventricular cardiomyopathy patients--disease-causing or innocent bystanders?

PubMed

Christensen, Alex Hørby; Benn, Marianne; Tybjaerg-Hansen, Anne; Haunso, Stig; Svendsen, Jesper Hastrup

2010-01-01

Mutations in genes encoding desmosomal proteins have been linked to arrhythmogenic right ventricular cardiomyopathy/dysplasia (ARVC/D). We hypothesized that a Scandinavian ARVC/D population would have a different spectrum of plakophilin-2 (PKP2) mutations and that some of the reported missense mutations may not be pathogenic. We screened 53 unrelated patients fulfilling Task Force criteria for ARVC/D for mutations in PKP2 by direct sequencing. Seven different mutations were identified: two insertion/deletions (E329fsX352, P401fsX406), 1 splice site (2146-2A>T), 1 non-sense (R79X) and 4 missense mutations (Q62K in 2 patients, G489R, G673V) of undeterminable pathogeneity. None of these mutations was present in 650 controls. Five of the mutations were novel. Seven patients carried reported missense mutations (D26N, S140F, V587I); however, these mutations were identified in our healthy controls, although at a lower frequency. Evaluation of all reported missense mutations in PKP2 showed unclear pathogeneity of several reported mutations. Fifteen percent of Danish ARVC/D patients carried PKP2 mutations. Our finding of reported disease-causing mutations at a low frequency among healthy controls suggests that these variants are disease modifying but not directly disease causing. We recommend conservative interpretation of missense variants in PKP2, functional characterization and large-scale sequencing to clarify normal variation in the gene.

Survival analysis of cancer risk reduction strategies for BRCA1/2 mutation carriers.

PubMed

Kurian, Allison W; Sigal, Bronislava M; Plevritis, Sylvia K

2010-01-10

Women with BRCA1/2 mutations inherit high risks of breast and ovarian cancer; options to reduce cancer mortality include prophylactic surgery or breast screening, but their efficacy has never been empirically compared. We used decision analysis to simulate risk-reducing strategies in BRCA1/2 mutation carriers and to compare resulting survival probability and causes of death. We developed a Monte Carlo model of breast screening with annual mammography plus magnetic resonance imaging (MRI) from ages 25 to 69 years, prophylactic mastectomy (PM) at various ages, and/or prophylactic oophorectomy (PO) at ages 40 or 50 years in 25-year-old BRCA1/2 mutation carriers. With no intervention, survival probability by age 70 is 53% for BRCA1 and 71% for BRCA2 mutation carriers. The most effective single intervention for BRCA1 mutation carriers is PO at age 40, yielding a 15% absolute survival gain; for BRCA2 mutation carriers, the most effective single intervention is PM, yielding a 7% survival gain if performed at age 40 years. The combination of PM and PO at age 40 improves survival more than any single intervention, yielding 24% survival gain for BRCA1 and 11% for BRCA2 mutation carriers. PM at age 25 instead of age 40 offers minimal incremental benefit (1% to 2%); substituting screening for PM yields a similarly minimal decrement in survival (2% to 3%). Although PM at age 25 plus PO at age 40 years maximizes survival probability, substituting mammography plus MRI screening for PM seems to offer comparable survival. These results may guide women with BRCA1/2 mutations in their choices between prophylactic surgery and breast screening.

Joint effects of pleiotropic selection and stabilizing selection on the maintenance of quantitative genetic variation at mutation-selection balance.

PubMed Central

Zhang, Xu-Sheng; Hill, William G

2002-01-01

In quantitative genetics, there are two basic "conflicting" observations: abundant polygenic variation and strong stabilizing selection that should rapidly deplete that variation. This conflict, although having attracted much theoretical attention, still stands open. Two classes of model have been proposed: real stabilizing selection directly on the metric trait under study and apparent stabilizing selection caused solely by the deleterious pleiotropic side effects of mutations on fitness. Here these models are combined and the total stabilizing selection observed is assumed to derive simultaneously through these two different mechanisms. Mutations have effects on a metric trait and on fitness, and both effects vary continuously. The genetic variance (V(G)) and the observed strength of total stabilizing selection (V(s,t)) are analyzed with a rare-alleles model. Both kinds of selection reduce V(G) but their roles in depleting it are not independent: The magnitude of pleiotropic selection depends on real stabilizing selection and such dependence is subject to the shape of the distributions of mutational effects. The genetic variation maintained thus depends on the kurtosis as well as the variance of mutational effects: All else being equal, V(G) increases with increasing leptokurtosis of mutational effects on fitness, while for a given distribution of mutational effects on fitness, V(G) decreases with increasing leptokurtosis of mutational effects on the trait. The V(G) and V(s,t) are determined primarily by real stabilizing selection while pleiotropic effects, which can be large, have only a limited impact. This finding provides some promise that a high heritability can be explained under strong total stabilizing selection for what are regarded as typical values of mutation and selection parameters. PMID:12242254

Expanding the SHOC2 mutation associated phenotype of Noonan syndrome with loose anagen hair: structural brain anomalies and myelofibrosis.

PubMed

Gripp, Karen W; Zand, Dina J; Demmer, Laurie; Anderson, Carol E; Dobyns, William B; Zackai, Elaine H; Denenberg, Elizabeth; Jenny, Kim; Stabley, Deborah L; Sol-Church, Katia

2013-10-01

Noonan syndrome is a heterogenous rasopathy typically presenting with short stature, characteristic facial features, cardiac abnormalities including pulmonic valve stenosis, ASD and hypertrophic cardiomyopathy (HCM), cryptorchidism, ectodermal abnormalities, and learning differences. The phenotype is variable, and limited genotype phenotype correlation exists with SOS1 mutations often associated with normal cognition and stature, RAF1 mutations entailing a high HCM risk, and certain PTPN11 mutations predisposing to juvenile myelomonocytic leukemia. The recently identified SHOC2 mutation (p.Ser2Gly) causes Noonan syndrome with loose anagen hair. We report five patients with this mutation. All had skin hyperpigmentation, sparse light colored hair, increased fine wrinkles, ligamentous laxity, developmental delay, and 4/4 had a structural cardiac anomaly. Hypotonia and macrocephaly occurred in 4/5 (80%); 3/5 (60%) had polyhydramnios, increased birth weight or required use of a feeding tube. Distinctive brain abnormalities included relative megalencephaly and enlarged subarachnoid spaces suggestive of benign external hydrocephalus, and a relatively small posterior fossa as indicated by a vertical tentorium. The combination of a large brain with a small posterior fossa likely resulted in the high rate of cerebellar tonsillar ectopia (3/4; 75%). Periventricular nodular heterotopia was seen in one patient with a thick and dysplastic corpus callosum. We report on the first hematologic neoplasm, myelofibrosis, in a 2-year-old patient with SHOC2 mutation. Myelofibrosis is exceedingly rare in children and young adults. The absence of a somatic JAK2 mutation, seen in the majority of patients with myelofibrosis, is noteworthy as it suggests that germline or somatic SHOC2 mutations are causally involved in myelofibrosis. Copyright © 2013 Wiley Periodicals, Inc.

Germline PMS2 and somatic POLE exonuclease mutations cause hypermutability of the leading DNA strand in biallelic mismatch repair deficiency syndrome brain tumours.

PubMed

Andrianova, Maria A; Chetan, Ghati Kasturirangan; Sibin, Madathan Kandi; Mckee, Thomas; Merkler, Doron; Narasinga, Rao Kvl; Ribaux, Pascale; Blouin, Jean-Louis; Makrythanasis, Periklis; Seplyarskiy, Vladimir B; Antonarakis, Stylianos E; Nikolaev, Sergey I

2017-11-01

Biallelic mismatch repair deficiency (bMMRD) in tumours is frequently associated with somatic mutations in the exonuclease domains of DNA polymerases POLE or POLD1, and results in a characteristic mutational profile. In this article, we describe the genetic basis of ultramutated high-grade brain tumours in the context of bMMRD. We performed exome sequencing of two second-cousin patients from a large consanguineous family of Indian origin with early onset of high-grade glioblastoma and astrocytoma. We identified a germline homozygous nonsense variant, p.R802*, in the PMS2 gene. Additionally, by genome sequencing of these tumours, we found extremely high somatic mutation rates (237/Mb and 123/Mb), as well as somatic mutations in the proofreading domain of POLE polymerase (p.P436H and p.L424V), which replicates the leading DNA strand. Most interestingly, we found, in both cancers, that the vast majority of mutations were consistent with the signature of POLE exo - , i.e. an abundance of C>A and C>T mutations, particularly in special contexts, on the leading strand. We showed that the fraction of mutations under positive selection among mutations in tumour suppressor genes is more than two-fold lower in ultramutated tumours than in other glioblastomas. Genetic analyses enabled the diagnosis of the two consanguineous childhood brain tumours as being due to a combination of PMS2 germline and POLE somatic variants, and confirmed them as bMMRD/POLE exo - disorders. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Novel mutations and mutation combinations of ryanodine receptor in a chlorantraniliprole resistant population of Plutella xylostella (L.)

PubMed Central

Guo, Lei; Liang, Pei; Zhou, Xuguo; Gao, Xiwu

2014-01-01

A previous study documented a glycine to glutamic acid mutation (G4946E) in ryanodine receptor (RyR) was highly correlated to diamide insecticide resistance in field populations of Plutella xylostella (Lepidoptera: Plutellidae). In this study, a field population collected in Yunnan province, China, exhibited a 2128-fold resistance to chlorantraniliprole. Sequence comparison between resistant and susceptible P. xylostella revealed three novel mutations including a glutamic acid to valine substitution (E1338D), a glutamine to leucine substitution (Q4594L) and an isoleucine to methionine substitution (I4790M) in highly conserved regions of RyR. Frequency analysis of all four mutations in this field population showed that the three new mutations showed a high frequency of 100%, while the G4946E had a frequency of 20%. Furthermore, the florescent ligand binding assay revealed that the RyR containing multiple mutations displayed a significantly lower affinity to the chlorantraniliprole. The combined results suggested that the co-existence of different combinations of the four mutations was involved in the chlorantraniliprole resistance. An allele-specific PCR based method was developed for the diagnosis of the four mutations in the field populations of P. xylostella. PMID:25377064

A de novo mutation in the AGXT gene causing primary hyperoxaluria type 1.

PubMed

Williams, Emma L; Kemper, Markus J; Rumsby, Gill

2006-09-01

Primary hyperoxaluria type 1 is caused by mutations in the alanine-glyoxylate aminotransferase (AGXT) gene. In cases in which no mutation was identified, linkage analysis can be used to confirm or exclude the diagnosis in other siblings. We present a family in which a sibling of the index case predicted to have primary hyperoxaluria type 1 by means of linkage analysis failed to show hyperoxaluria during the following 7 years, putting the diagnosis into question. Whole-gene sequence analysis identified 2 causative mutations in the index case, of which only 1, c.646A (Gly216Arg), was inherited. The other sequence change, c.33_34insC, was a de novo mutation occurring on the paternal allele. This particular mutation is a relatively common cause of primary hyperoxaluria type 1. It occurs in a run of 8 cytosines and therefore potentially is susceptible to polymerase slippage. This case illustrates 2 important points. First, biochemical confirmation of a genetic diagnosis should always be made in siblings diagnosed by using genetic tests. Second, de novo mutations should be considered as a potential, albeit rare, cause of primary hyperoxaluria type 1.

CFTR Mutations Spectrum and the Efficiency of Molecular Diagnostics in Polish Cystic Fibrosis Patients

PubMed Central

Ziętkiewicz, Ewa; Rutkiewicz, Ewa; Pogorzelski, Andrzej; Klimek, Barbara; Voelkel, Katarzyna; Witt, Michał

2014-01-01

Cystic fibrosis (CF) is caused by mutations in the cystic fibrosis transmembrane regulator gene (CFTR). In light of the strong allelic heterogeneity and regional specificity of the mutation spectrum, the strategy of molecular diagnostics and counseling in CF requires genetic tests to reflect the frequency profile characteristic for a given population. The goal of the study was to provide an updated comprehensive estimation of the distribution of CFTR mutations in Polish CF patients and to assess the effectiveness of INNOLiPA_CFTR tests in Polish population. The analyzed cohort consisted of 738 patients with the clinically confirmed CF diagnosis, prescreened for molecular defects using INNOLiPA_CFTR panels from Innogenetics. A combined efficiency of INNOLiPA CFTR_19 and CFTR_17_TnUpdate tests was 75.5%; both mutations were detected in 68.2%, and one mutation in 14.8% of the affected individuals. The group composed of all the patients with only one or with no mutation detected (109 and 126 individuals, respectively) was analyzed further using a mutation screening approach, i.e. SSCP/HD (single strand conformational polymorphism/heteroduplex) analysis of PCR products followed by sequencing of the coding sequence. As a result, 53 more mutations were found in 97 patients. The overall efficiency of the CF allele detection was 82.5% (7.0% increase compared to INNOLiPA tests alone). The distribution of the most frequent mutations in Poland was assessed. Most of the mutations repetitively found in Polish patients had been previously described in other European populations. The most frequent mutated allele, F508del, represented 54.5% of Polish CF chromosomes. Another eight mutations had frequencies over 1%, 24 had frequencies between 1 and 0.1%; c.2052-2053insA and c.3468+2_3468+3insT were the most frequent non-INNOLiPA mutations. Mutation distribution described herein is also relevant to the Polish diaspora. Our study also demonstrates that the reported efficiency of mutation detection strongly depends on the diagnostic experience of referring health centers. PMID:24586523

CFTR mutations spectrum and the efficiency of molecular diagnostics in Polish cystic fibrosis patients.

PubMed

Ziętkiewicz, Ewa; Rutkiewicz, Ewa; Pogorzelski, Andrzej; Klimek, Barbara; Voelkel, Katarzyna; Witt, Michał

2014-01-01

Cystic fibrosis (CF) is caused by mutations in the cystic fibrosis transmembrane regulator gene (CFTR). In light of the strong allelic heterogeneity and regional specificity of the mutation spectrum, the strategy of molecular diagnostics and counseling in CF requires genetic tests to reflect the frequency profile characteristic for a given population. The goal of the study was to provide an updated comprehensive estimation of the distribution of CFTR mutations in Polish CF patients and to assess the effectiveness of INNOLiPA_CFTR tests in Polish population. The analyzed cohort consisted of 738 patients with the clinically confirmed CF diagnosis, prescreened for molecular defects using INNOLiPA_CFTR panels from Innogenetics. A combined efficiency of INNOLiPA CFTR_19 and CFTR_17_TnUpdate tests was 75.5%; both mutations were detected in 68.2%, and one mutation in 14.8% of the affected individuals. The group composed of all the patients with only one or with no mutation detected (109 and 126 individuals, respectively) was analyzed further using a mutation screening approach, i.e. SSCP/HD (single strand conformational polymorphism/heteroduplex) analysis of PCR products followed by sequencing of the coding sequence. As a result, 53 more mutations were found in 97 patients. The overall efficiency of the CF allele detection was 82.5% (7.0% increase compared to INNOLiPA tests alone). The distribution of the most frequent mutations in Poland was assessed. Most of the mutations repetitively found in Polish patients had been previously described in other European populations. The most frequent mutated allele, F508del, represented 54.5% of Polish CF chromosomes. Another eight mutations had frequencies over 1%, 24 had frequencies between 1 and 0.1%; c.2052-2053insA and c.3468+2_3468+3insT were the most frequent non-INNOLiPA mutations. Mutation distribution described herein is also relevant to the Polish diaspora. Our study also demonstrates that the reported efficiency of mutation detection strongly depends on the diagnostic experience of referring health centers.

Epilepsy caused by an abnormal alternative splicing with dosage effect of the SV2A gene in a chicken model.

PubMed

Douaud, Marine; Feve, Katia; Pituello, Fabienne; Gourichon, David; Boitard, Simon; Leguern, Eric; Coquerelle, Gérard; Vieaud, Agathe; Batini, Cesira; Naquet, Robert; Vignal, Alain; Tixier-Boichard, Michèle; Pitel, Frédérique

2011-01-01

Photosensitive reflex epilepsy is caused by the combination of an individual's enhanced sensitivity with relevant light stimuli, such as stroboscopic lights or video games. This is the most common reflex epilepsy in humans; it is characterized by the photoparoxysmal response, which is an abnormal electroencephalographic reaction, and seizures triggered by intermittent light stimulation. Here, by using genetic mapping, sequencing and functional analyses, we report that a mutation in the acceptor site of the second intron of SV2A (the gene encoding synaptic vesicle glycoprotein 2A) is causing photosensitive reflex epilepsy in a unique vertebrate model, the Fepi chicken strain, a spontaneous model where the neurological disorder is inherited as an autosomal recessive mutation. This mutation causes an aberrant splicing event and significantly reduces the level of SV2A mRNA in homozygous carriers. Levetiracetam, a second generation antiepileptic drug, is known to bind SV2A, and SV2A knock-out mice develop seizures soon after birth and usually die within three weeks. The Fepi chicken survives to adulthood and responds to levetiracetam, suggesting that the low-level expression of SV2A in these animals is sufficient to allow survival, but does not protect against seizures. Thus, the Fepi chicken model shows that the role of the SV2A pathway in the brain is conserved between birds and mammals, in spite of a large phylogenetic distance. The Fepi model appears particularly useful for further studies of physiopathology of reflex epilepsy, in comparison with induced models of epilepsy in rodents. Consequently, SV2A is a very attractive candidate gene for analysis in the context of both mono- and polygenic generalized epilepsies in humans.