Mechanism of Substrate Recognition And PLP-Induced Conformational Changes in II-Diaminopimelate Aminotransferase From Arabidopsis Thaliana

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

Watanabe, N.; Clay, M.D.; Belkum, M.J.van

2009-05-26

LL-Diaminopimelate aminotransferase (LL-DAP-AT), a pyridoxal phosphate (PLP)-dependent enzyme in the lysine biosynthetic pathways of plants and Chlamydia, is a potential target for the development of herbicides or antibiotics. This homodimeric enzyme converts L-tetrahydrodipicolinic acid (THDP) directly to LL-DAP using L-glutamate as the source of the amino group. Earlier, we described the 3D structures of native and malate-bound LL-DAP-AT from Arabidopsis thaliana (AtDAP-AT). Seven additional crystal structures of AtDAP-AT and its variants are reported here as part of an investigation into the mechanism of substrate recognition and catalysis. Two structures are of AtDAP-AT with reduced external aldimine analogues: N-(5'-phosphopyridoxyl)-L-glutamate (PLP-Glu) andmore » N-(5'-phosphopyridoxyl)- LL-Diaminopimelate (PLP-DAP) bound in the active site. Surprisingly, they reveal that both L-glutamate and LL-DAP are recognized in a very similar fashion by the same sets of amino acid residues; both molecules adopt twisted V-shaped conformations. With both substrates, the {alpha}-carboxylates are bound in a salt bridge with Arg404, whereas the distal carboxylates are recognized via hydrogen bonds to the well-conserved side chains of Tyr37, Tyr125 and Lys129. The distal C{sup {var_epsilon}} amino group of LL-DAP is specifically recognized by several non-covalent interactions with residues from the other subunit (Asn309*, Tyr94*, Gly95*, and Glu97* (Amino acid designators followed by an asterisk (*) indicate that the residues originate in the other subunit of the dimer)) and by three bound water molecules. Two catalytically inactive variants of AtDAP-AT were created via site-directed mutagenesis of the active site lysine (K270N and K270Q). The structures of these variants permitted the observation of the unreduced external aldimines of PLP with L-glutamate and with LL-DAP in the active site, and revealed differences in the torsion angle about the PLP-substrate bond. Lastly

Role of the pyridine nitrogen in pyridoxal 5'-phosphate catalysis: activity of three classes of PLP enzymes reconstituted with deazapyridoxal 5'-phosphate.

PubMed

Griswold, Wait R; Toney, Michael D

2011-09-21

Pyridoxal 5'-phosphate (PLP; vitamin B(6))-catalyzed reactions have been well studied, both on enzymes and in solution, due to the variety of important reactions this cofactor catalyzes in nitrogen metabolism. Three functional groups are central to PLP catalysis: the C4' aldehyde, the O3' phenol, and the N1 pyridine nitrogen. In the literature, the pyridine nitrogen has traditionally been assumed to be protonated in enzyme active sites, with the protonated pyridine ring providing resonance stabilization of carbanionic intermediates. This assumption is certainly correct for some PLP enzymes, but the structures of other active sites are incompatible with protonation of N1, and, consequently, these enzymes are expected to use PLP in the N1-unprotonated form. For example, aspartate aminotransferase protonates the pyridine nitrogen for catalysis of transamination, while both alanine racemase and O-acetylserine sulfhydrylase are expected to maintain N1 in the unprotonated, formally neutral state for catalysis of racemization and β-elimination. Herein, kinetic results for these three enzymes reconstituted with 1-deazapyridoxal 5'-phosphate, an isosteric analogue of PLP lacking the pyridine nitrogen, are compared to those for the PLP enzyme forms. They demonstrate that the pyridine nitrogen is vital to the 1,3-prototropic shift central to transamination, but not to reactions catalyzed by alanine racemase or O-acetylserine sulfhydrylase. Not all PLP enzymes require the electrophilicity of a protonated pyridine ring to enable formation of carbanionic intermediates. It is proposed that modulation of cofactor electrophilicity plays a central role in controlling reaction specificity in PLP enzymes.

Biologically relevant conformational features of linear and cyclic proteolipid protein (PLP) peptide analogues obtained by high-resolution nuclear magnetic resonance and molecular dynamics

NASA Astrophysics Data System (ADS)

Kordopati, Golfo G.; Tzoupis, Haralambos; Troganis, Anastassios N.; Tsivgoulis, Gerasimos M.; Golic Grdadolnik, Simona; Simal, Carmen; Tselios, Theodore V.

2017-09-01

Proteolipid protein (PLP) is one of the main proteins of myelin sheath that are destroyed during the progress of multiple sclerosis (MS). The immunodominant PLP139-151 epitope is known to induce experimental autoimmune encephalomyelitis (EAE, animal model of MS), wherein residues 144 and 147 are recognized by T cell receptor (TCR) during the formation of trimolecular complex with peptide-antigen and major histocompability complex. The conformational behavior of linear and cyclic peptide analogues of PLP, namely PLP139-151 and cyclic (139-151) (L144, R147) PLP139-151, have been studied in solution by means of nuclear magnetic resonance (NMR) methods in combination with unrestrained molecular dynamics simulations. The results indicate that the side chains of mutated amino acids in the cyclic analogue have different spatial orientation compared with the corresponding side chains of the linear analogue, which can lead to reduced affinity to TCR. NMR experiments combined with theoretical calculations pave the way for the design and synthesis of potent restricted peptides of immunodominant PLP139-151 epitope as well as non peptide mimetics that rises as an ultimate goal.

Molecular dynamics simulations of apo, holo, and inactivator bound GABA-at reveal the role of active site residues in PLP dependent enzymes.

PubMed

Gökcan, Hatice; Monard, Gerald; Sungur Konuklar, F Aylin

2016-07-01

The pyridoxal 5-phosphate (PLP) cofactor is a significant organic molecule in medicinal chemistry. It is often found covalently bound to lysine residues in proteins to form PLP dependent enzymes. An example of this family of PLP dependent enzymes is γ-aminobutyric acid aminotransferase (GABA-AT) which is responsible for the degradation of the neurotransmitter GABA. Its inhibition or inactivation can be used to prevent the reduction of GABA concentration in brain which is the source of several neurological disorders. As a test case for PLP dependent enzymes, we have performed molecular dynamics simulations of GABA-AT to reveal the roles of the protein residues and its cofactor. Three different states have been considered: the apoenzyme, the holoenzyme, and the inactive state obtained after the suicide inhibition by vigabatrin. Different protonation states have also been considered for PLP and two key active site residues: Asp298 and His190. Together, 24 independent molecular dynamics trajectories have been simulated for a cumulative total of 2.88 µs. Our results indicate that, unlike in aqueous solution, the PLP pyridine moiety is protonated in GABA-AT. This is a consequence of a pKa shift triggered by a strong charge-charge interaction with an ionic "diad" formed by Asp298 and His190 that would help the activation of the first half-reaction of the catalytic mechanism in GABA-AT: the conversion of PLP to free pyridoxamine phosphate (PMP). In addition, our MD simulations exhibit additional strong hydrogen bond networks between the protein and PLP: the phosphate group is held in place by the donation of at least three hydrogen bonds while the carbonyl oxygen of the pyridine ring interacts with Gln301; Phe181 forms a π-π stacking interaction with the pyridine ring and works as a gate keeper with the assistance of Val300. All these interactions are hypothesized to help maintain free PMP in place inside the protein active site to facilitate the second half

Integrative proteomics, genomics, and translational immunology approaches reveal mutated forms of Proteolipid Protein 1 (PLP1) and mutant-specific immune response in multiple sclerosis.

PubMed

Qendro, Veneta; Bugos, Grace A; Lundgren, Debbie H; Glynn, John; Han, May H; Han, David K

2017-03-01

In order to gain mechanistic insights into multiple sclerosis (MS) pathogenesis, we utilized a multi-dimensional approach to test the hypothesis that mutations in myelin proteins lead to immune activation and central nervous system autoimmunity in MS. Mass spectrometry-based proteomic analysis of human MS brain lesions revealed seven unique mutations of PLP1; a key myelin protein that is known to be destroyed in MS. Surprisingly, in-depth genomic analysis of two MS patients at the genomic DNA and mRNA confirmed mutated PLP1 in RNA, but not in the genomic DNA. Quantification of wild type and mutant PLP RNA levels by qPCR further validated the presence of mutant PLP RNA in the MS patients. To seek evidence linking mutations in abundant myelin proteins and immune-mediated destruction of myelin, specific immune response against mutant PLP1 in MS patients was examined. Thus, we have designed paired, wild type and mutant peptide microarrays, and examined antibody response to multiple mutated PLP1 in sera from MS patients. Consistent with the idea of different patients exhibiting unique mutation profiles, we found that 13 out of 20 MS patients showed antibody responses against specific but not against all the mutant-PLP1 peptides. Interestingly, we found mutant PLP-directed antibody response against specific mutant peptides in the sera of pre-MS controls. The results from integrative proteomic, genomic, and immune analyses reveal a possible mechanism of mutation-driven pathogenesis in human MS. The study also highlights the need for integrative genomic and proteomic analyses for uncovering pathogenic mechanisms of human diseases. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Functional Evolution of PLP-dependent Enzymes based on Active-Site Structural Similarities

PubMed Central

Catazaro, Jonathan; Caprez, Adam; Guru, Ashu; Swanson, David; Powers, Robert

2014-01-01

Families of distantly related proteins typically have very low sequence identity, which hinders evolutionary analysis and functional annotation. Slowly evolving features of proteins, such as an active site, are therefore valuable for annotating putative and distantly related proteins. To date, a complete evolutionary analysis of the functional relationship of an entire enzyme family based on active-site structural similarities has not yet been undertaken. Pyridoxal-5’-phosphate (PLP) dependent enzymes are primordial enzymes that diversified in the last universal ancestor. Using the Comparison of Protein Active Site Structures (CPASS) software and database, we show that the active site structures of PLP-dependent enzymes can be used to infer evolutionary relationships based on functional similarity. The enzymes successfully clustered together based on substrate specificity, function, and three-dimensional fold. This study demonstrates the value of using active site structures for functional evolutionary analysis and the effectiveness of CPASS. PMID:24920327

Functional evolution of PLP-dependent enzymes based on active-site structural similarities.

PubMed

Catazaro, Jonathan; Caprez, Adam; Guru, Ashu; Swanson, David; Powers, Robert

2014-10-01

Families of distantly related proteins typically have very low sequence identity, which hinders evolutionary analysis and functional annotation. Slowly evolving features of proteins, such as an active site, are therefore valuable for annotating putative and distantly related proteins. To date, a complete evolutionary analysis of the functional relationship of an entire enzyme family based on active-site structural similarities has not yet been undertaken. Pyridoxal-5'-phosphate (PLP) dependent enzymes are primordial enzymes that diversified in the last universal ancestor. Using the comparison of protein active site structures (CPASS) software and database, we show that the active site structures of PLP-dependent enzymes can be used to infer evolutionary relationships based on functional similarity. The enzymes successfully clustered together based on substrate specificity, function, and three-dimensional-fold. This study demonstrates the value of using active site structures for functional evolutionary analysis and the effectiveness of CPASS. © 2014 Wiley Periodicals, Inc.

Interphase centrosome organization by the PLP-Cnn scaffold is required for centrosome function

PubMed Central

Lerit, Dorothy A.; Jordan, Holly A.; Poulton, John S.; Fagerstrom, Carey J.; Galletta, Brian J.; Peifer, Mark

2015-01-01

Pericentriolar material (PCM) mediates the microtubule (MT) nucleation and anchoring activity of centrosomes. A scaffold organized by Centrosomin (Cnn) serves to ensure proper PCM architecture and functional changes in centrosome activity with each cell cycle. Here, we investigate the mechanisms that spatially restrict and temporally coordinate centrosome scaffold formation. Focusing on the mitotic-to-interphase transition in Drosophila melanogaster embryos, we show that the elaboration of the interphase Cnn scaffold defines a major structural rearrangement of the centrosome. We identify an unprecedented role for Pericentrin-like protein (PLP), which localizes to the tips of extended Cnn flares, to maintain robust interphase centrosome activity and promote the formation of interphase MT asters required for normal nuclear spacing, centrosome segregation, and compartmentalization of the syncytial embryo. Our data reveal that Cnn and PLP directly interact at two defined sites to coordinate the cell cycle–dependent rearrangement and scaffolding activity of the centrosome to permit normal centrosome organization, cell division, and embryonic viability. PMID:26150390

Interphase centrosome organization by the PLP-Cnn scaffold is required for centrosome function.

PubMed

Lerit, Dorothy A; Jordan, Holly A; Poulton, John S; Fagerstrom, Carey J; Galletta, Brian J; Peifer, Mark; Rusan, Nasser M

2015-07-06

Pericentriolar material (PCM) mediates the microtubule (MT) nucleation and anchoring activity of centrosomes. A scaffold organized by Centrosomin (Cnn) serves to ensure proper PCM architecture and functional changes in centrosome activity with each cell cycle. Here, we investigate the mechanisms that spatially restrict and temporally coordinate centrosome scaffold formation. Focusing on the mitotic-to-interphase transition in Drosophila melanogaster embryos, we show that the elaboration of the interphase Cnn scaffold defines a major structural rearrangement of the centrosome. We identify an unprecedented role for Pericentrin-like protein (PLP), which localizes to the tips of extended Cnn flares, to maintain robust interphase centrosome activity and promote the formation of interphase MT asters required for normal nuclear spacing, centrosome segregation, and compartmentalization of the syncytial embryo. Our data reveal that Cnn and PLP directly interact at two defined sites to coordinate the cell cycle-dependent rearrangement and scaffolding activity of the centrosome to permit normal centrosome organization, cell division, and embryonic viability.

Evolutionary Trails of Plant Group II Pyridoxal Phosphate-Dependent Decarboxylase Genes.

PubMed

Kumar, Rahul

2016-01-01

Type II pyridoxal phosphate-dependent decarboxylase (PLP_deC) enzymes play important metabolic roles during nitrogen metabolism. Recent evolutionary profiling of these genes revealed a sharp expansion of histidine decarboxylase genes in the members of Solanaceae family. In spite of the high sequence homology shared by PLP_deC orthologs, these enzymes display remarkable differences in their substrate specificities. Currently, limited information is available on the gene repertoires and substrate specificities of PLP_deCs which renders their precise annotation challenging and offers technical challenges in the immediate identification and biochemical characterization of their full gene complements in plants. Herein, we explored their evolutionary trails in a comprehensive manner by taking advantage of high-throughput data accessibility and computational approaches. We discussed the premise that has enabled an improved reconstruction of their evolutionary lineage and evaluated the factors offering constraints in their rapid functional characterization, till date. We envisage that the synthesized information herein would act as a catalyst for the rapid exploration of their biochemical specificity and physiological roles in more plant species.

Allosteric communication between the pyridoxal 5'-phosphate (PLP) and heme sites in the H2S generator human cystathionine β-synthase.

PubMed

Yadav, Pramod Kumar; Xie, Peter; Banerjee, Ruma

2012-11-02

Human cystathionine β-synthase (CBS) is a unique pyridoxal 5'-phosphate (PLP)-dependent enzyme that has a regulatory heme cofactor. Previous studies have demonstrated the importance of Arg-266, a residue at the heme pocket end of α-helix 8, for communication between the heme and PLP sites. In this study, we have examined the role of the conserved Thr-257 and Thr-260 residues, located at the other end of α-helix 8 on the heme electronic environment and on activity. The mutations at the two positions destabilize PLP binding, leading to lower PLP content and ~2- to ~500-fold lower activity compared with the wild-type enzyme. Activity is unresponsive to PLP supplementation, consistent with the pyridoxine-nonresponsive phenotype of the T257M mutation in a homocystinuric patient. The H(2)S-producing activities, also impacted by the mutations, show a different pattern of inhibition compared with the canonical transsulfuration reaction. Interestingly, the mutants exhibit contrasting sensitivities to the allosteric effector, S-adenosylmethionine (AdoMet); whereas T257M and T257I are inhibited, the other mutants are hyperactivated by AdoMet. All mutants showed an increased propensity of the ferrous heme to form an inactive species with a 424 nm Soret peak and exhibited significantly reduced enzyme activity in the ferrous and ferrous-CO states. Our results provide the first evidence for bidirectional transmission of information between the cofactor binding sites, suggest the additional involvement of this region in allosteric communication with the regulatory AdoMet-binding domain, and reveal the potential for independent modulation of the canonical transsulfuration versus H(2)S-generating reactions catalyzed by CBS.

Therapy of Pelizaeus-Merzbacher disease in mice by feeding a cholesterol-enriched diet.

PubMed

Saher, Gesine; Rudolphi, Fabian; Corthals, Kristina; Ruhwedel, Torben; Schmidt, Karl-Friedrich; Löwel, Siegrid; Dibaj, Payam; Barrette, Benoit; Möbius, Wiebke; Nave, Klaus-Armin

2012-07-01

Duplication of PLP1 (proteolipid protein gene 1) and the subsequent overexpression of the myelin protein PLP (also known as DM20) in oligodendrocytes is the most frequent cause of Pelizaeus-Merzbacher disease (PMD), a fatal leukodystrophy without therapeutic options. PLP binds cholesterol and is contained within membrane lipid raft microdomains. Cholesterol availability is the rate-limiting factor of central nervous system myelin synthesis. Transgenic mice with extra copies of the Plp1 gene are accurate models of PMD. Dysmyelination followed by demyelination, secondary inflammation and axon damage contribute to the severe motor impairment in these mice. The finding that in Plp1-transgenic oligodendrocytes, PLP and cholesterol accumulate in late endosomes and lysosomes (endo/lysosomes), prompted us to further investigate the role of cholesterol in PMD. Here we show that cholesterol itself promotes normal PLP trafficking and that dietary cholesterol influences PMD pathology. In a preclinical trial, PMD mice were fed a cholesterol-enriched diet. This restored oligodendrocyte numbers and ameliorated intracellular PLP accumulation. Moreover, myelin content increased, inflammation and gliosis were reduced and motor defects improved. Even after onset of clinical symptoms, cholesterol treatment prevented disease progression. Dietary cholesterol did not reduce Plp1 overexpression but facilitated incorporation of PLP into myelin membranes. These findings may have implications for therapeutic interventions in patients with PMD.

Gemfibrozil, a lipid-lowering drug, increases myelin genes in human oligodendrocytes via peroxisome proliferator-activated receptor-β.

PubMed

Jana, Malabendu; Mondal, Susanta; Gonzalez, Frank J; Pahan, Kalipada

2012-10-05

An increase in CNS remyelination and a decrease in CNS inflammation are important steps to halt the progression of multiple sclerosis. Earlier studies have shown that gemfibrozil, a lipid-lowering drug, has anti-inflammatory properties. The current study identified another novel property of gemfibrozil in stimulating the expression of myelin-specific genes (myelin basic protein, myelin oligodendrocyte glycoprotein, 2',3'-cyclic-nucleotide 3'-phosphodiesterase, and proteolipid protein (PLP)) in primary human oligodendrocytes, mixed glial cells, and spinal cord organotypic cultures. Although gemfibrozil is a known activator of peroxisome proliferator-activated receptor-α (PPAR-α), we were unable to detect PPAR-α in either gemfibrozil-treated or untreated human oligodendrocytes, and gemfibrozil increased the expression of myelin genes in oligodendrocytes isolated from both wild type and PPAR-α(-/-) mice. On the other hand, gemfibrozil markedly increased the expression of PPAR-β but not PPAR-γ. Consistently, antisense knockdown of PPAR-β, but not PPAR-γ, abrogated the stimulatory effect of gemfibrozil on myelin genes in human oligodendrocytes. Gemfibrozil also did not up-regulate myelin genes in oligodendroglia isolated from PPAR-β(-/-) mice. Chromatin immunoprecipitation analysis showed that gemfibrozil induced the recruitment of PPAR-β to the promoter of PLP and myelin oligodendrocyte glycoprotein genes in human oligodendrocytes. Furthermore, gemfibrozil treatment also led to the recruitment of PPAR-β to the PLP promoter in vivo in the spinal cord of experimental autoimmune encephalomyelitis mice and suppression of experimental autoimmune encephalomyelitis symptoms in PLP-T cell receptor transgenic mice. These results suggest that gemfibrozil stimulates the expression of myelin genes via PPAR-β and that gemfibrozil, a prescribed drug for humans, may find further therapeutic use in demyelinating diseases.

Treatment of phantom limb pain (PLP) based on augmented reality and gaming controlled by myoelectric pattern recognition: a case study of a chronic PLP patient

PubMed Central

Ortiz-Catalan, Max; Sander, Nichlas; Kristoffersen, Morten B.; Håkansson, Bo; Brånemark, Rickard

2014-01-01

A variety of treatments have been historically used to alleviate phantom limb pain (PLP) with varying efficacy. Recently, virtual reality (VR) has been employed as a more sophisticated mirror therapy. Despite the advantages of VR over a conventional mirror, this approach has retained the use of the contralateral limb and is therefore restricted to unilateral amputees. Moreover, this strategy disregards the actual effort made by the patient to produce phantom motions. In this work, we investigate a treatment in which the virtual limb responds directly to myoelectric activity at the stump, while the illusion of a restored limb is enhanced through augmented reality (AR). Further, phantom motions are facilitated and encouraged through gaming. The proposed set of technologies was administered to a chronic PLP patient who has shown resistance to a variety of treatments (including mirror therapy) for 48 years. Individual and simultaneous phantom movements were predicted using myoelectric pattern recognition and were then used as input for VR and AR environments, as well as for a racing game. The sustained level of pain reported by the patient was gradually reduced to complete pain-free periods. The phantom posture initially reported as a strongly closed fist was gradually relaxed, interestingly resembling the neutral posture displayed by the virtual limb. The patient acquired the ability to freely move his phantom limb, and a telescopic effect was observed where the position of the phantom hand was restored to the anatomically correct distance. More importantly, the effect of the interventions was positively and noticeably perceived by the patient and his relatives. Despite the limitation of a single case study, the successful results of the proposed system in a patient for whom other medical and non-medical treatments have been ineffective justifies and motivates further investigation in a wider study. PMID:24616655

Treatment of phantom limb pain (PLP) based on augmented reality and gaming controlled by myoelectric pattern recognition: a case study of a chronic PLP patient.

PubMed

Ortiz-Catalan, Max; Sander, Nichlas; Kristoffersen, Morten B; Håkansson, Bo; Brånemark, Rickard

2014-01-01

A variety of treatments have been historically used to alleviate phantom limb pain (PLP) with varying efficacy. Recently, virtual reality (VR) has been employed as a more sophisticated mirror therapy. Despite the advantages of VR over a conventional mirror, this approach has retained the use of the contralateral limb and is therefore restricted to unilateral amputees. Moreover, this strategy disregards the actual effort made by the patient to produce phantom motions. In this work, we investigate a treatment in which the virtual limb responds directly to myoelectric activity at the stump, while the illusion of a restored limb is enhanced through augmented reality (AR). Further, phantom motions are facilitated and encouraged through gaming. The proposed set of technologies was administered to a chronic PLP patient who has shown resistance to a variety of treatments (including mirror therapy) for 48 years. Individual and simultaneous phantom movements were predicted using myoelectric pattern recognition and were then used as input for VR and AR environments, as well as for a racing game. The sustained level of pain reported by the patient was gradually reduced to complete pain-free periods. The phantom posture initially reported as a strongly closed fist was gradually relaxed, interestingly resembling the neutral posture displayed by the virtual limb. The patient acquired the ability to freely move his phantom limb, and a telescopic effect was observed where the position of the phantom hand was restored to the anatomically correct distance. More importantly, the effect of the interventions was positively and noticeably perceived by the patient and his relatives. Despite the limitation of a single case study, the successful results of the proposed system in a patient for whom other medical and non-medical treatments have been ineffective justifies and motivates further investigation in a wider study.

Gemfibrozil, a Lipid-lowering Drug, Increases Myelin Genes in Human Oligodendrocytes via Peroxisome Proliferator-activated Receptor-β*

PubMed Central

Jana, Malabendu; Mondal, Susanta; Gonzalez, Frank J.; Pahan, Kalipada

2012-01-01

An increase in CNS remyelination and a decrease in CNS inflammation are important steps to halt the progression of multiple sclerosis. Earlier studies have shown that gemfibrozil, a lipid-lowering drug, has anti-inflammatory properties. The current study identified another novel property of gemfibrozil in stimulating the expression of myelin-specific genes (myelin basic protein, myelin oligodendrocyte glycoprotein, 2′,3′-cyclic-nucleotide 3′-phosphodiesterase, and proteolipid protein (PLP)) in primary human oligodendrocytes, mixed glial cells, and spinal cord organotypic cultures. Although gemfibrozil is a known activator of peroxisome proliferator-activated receptor-α (PPAR-α), we were unable to detect PPAR-α in either gemfibrozil-treated or untreated human oligodendrocytes, and gemfibrozil increased the expression of myelin genes in oligodendrocytes isolated from both wild type and PPAR-α(−/−) mice. On the other hand, gemfibrozil markedly increased the expression of PPAR-β but not PPAR-γ. Consistently, antisense knockdown of PPAR-β, but not PPAR-γ, abrogated the stimulatory effect of gemfibrozil on myelin genes in human oligodendrocytes. Gemfibrozil also did not up-regulate myelin genes in oligodendroglia isolated from PPAR-β(−/−) mice. Chromatin immunoprecipitation analysis showed that gemfibrozil induced the recruitment of PPAR-β to the promoter of PLP and myelin oligodendrocyte glycoprotein genes in human oligodendrocytes. Furthermore, gemfibrozil treatment also led to the recruitment of PPAR-β to the PLP promoter in vivo in the spinal cord of experimental autoimmune encephalomyelitis mice and suppression of experimental autoimmune encephalomyelitis symptoms in PLP-T cell receptor transgenic mice. These results suggest that gemfibrozil stimulates the expression of myelin genes via PPAR-β and that gemfibrozil, a prescribed drug for humans, may find further therapeutic use in demyelinating diseases. PMID:22879602

Developmental Activation of the Proteolipid Protein Promoter Transgene in Neuronal and Oligodendroglial Cells of Neostriatum in Mice

PubMed Central

Fulton, Daniel; Paez, Pablo; Spreur, Vilma; Handley, Vance; Colwell, Christopher S.; Campagnoni, Anthony; Fisher, Robin

2011-01-01

Prior studies suggest that non-canonical proteolipid protein (PLP) gene expression occurs during development in non-myelinating neurons as well as myelinating oligodendroglia in mammalian brain. To assess this possibility in neostriatum, a region of uncertain PLP gene expression in neurons, morphological and electrophysiological tools were used to determine phenotypes of cells with activation of a PLP promoter transgene during the early postnatal period in mice. PLP gene expression is evident in both neuronal and oligodendroglial phenotypes in developing neostriatum, a conclusion based on three novel observations: (1) An enhanced green fluorescent protein (EGFP) reporter of PLP promoter activation was localized in two distinct populations of cells, which exhibit collective, developmental differences of morphological and electrophysiological characteristics in accord with neuronal and oligodendroglial phenotypes of neostriatal cells found during the early postnatal period in both transgenic and wild-type mice. (2) The EGFP reporter of PLP promoter activation was appropriately positioned to serve as a regulator of PLP gene expression. It colocalized with native PLP proteins in both neuronal and oligodendroglial phenotypes; however, only soma-restricted PLP protein isoforms were found in the neuronal phenotype, while classic and soma-restricted PLP protein isoforms were found in the oligodendroglial phenotype. (3) As shown by EGFP reporter, PLP promoter activation was placed to regulate PLP gene expression in only one neuronal phenotype among the several that constitute neostriatum. It was localized in medium spiny neurons, but not large aspiny neurons. These outcomes have significant implications for the non-canonical functional roles of PLP gene expression in addition to myelinogenesis in mammalian brain, and are consistent with potentially independent pathologic loci in neurons during the course of human mutational disorders of PLP gene expression. PMID:21912090

Gene expression in the developing cerebellum during perinatal hypo- and hyperthyroidism.

PubMed

Figueiredo, B C; Almazan, G; Ma, Y; Tetzlaff, W; Miller, F D; Cuello, A C

1993-03-01

The intensity of p75NGFR receptor-like immunoreactivity and the mRNAs encoding p75NGFR, T alpha 1 alpha-tubulin, GAP-43 and the myelin proteins MBP and PLP were measured in the developing cerebellum to study the effects of perinatal thyroid hormone imbalance in rats. Results compared to age-matched controls provide in vivo evidence for differential gene regulation by thyroid hormone in the developing cerebellum. We found that p75NGFR immunoreactivity was strikingly elevated in hypothyroid rats, whereas p75NGFR mRNA content remained only twice as high as that of control levels on postnatal day 15 (P15). When p75NGFR immunoreactivity was still elevated in hypothyroid rats, Purkinje cells exhibited proximal axonal varicosities, axonal twisting and differences in axonal caliber. The mRNAs encoding proteins involved with neurite growth-promoting elements, T alpha 1 alpha-tubulin and GAP-43, were also increased in hypothyroidism, possibly reflecting a neuronal response to a deficiency in, or damage to, cerebellar neurons, or a general delay in their down regulation. Similar increases were not observed for the myelin specific genes. MBP and PLP mRNAs were first detected on P2 of hyperthyroid rats, and they increased with age. Hypo- or hyperthyroidism did not affect the initial onset of MBP and PLP expression, however, hyperthyroidism increased levels of PLP and MBP mRNAs between P2 and P10. By contrast, the most consistent decrease in MBP and PLP mRNAs in rats with thyroid hormone deficiency was observed only on P10. At later times (P15 and P30), the two mRNA levels were similar to controls in all groups. These results are consistent with a role for thyroid hormone in the earlier stages of cerebellar myelination. Hypothryoidism led to specific increases in T alpha 1 alpha-tubulin and GAP-43 mRNAs, and in the immunoreactivity and mRNA levels of p75NGFR receptor--all changes that may play a role in the observed abnormal neuronal outgrowth.

4′-CyanoPLP presents better prospect for the experimental detection of elusive cyclic intermediate radical in the reaction of lysine 5,6-aminomutase

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

Maity, Amarendra Nath; Ke, Shyue-Chu, E-mail: ke@mail.ndhu.edu.tw

2015-02-06

Graphical abstract: The results of our calculations suggest that the reaction of 4′-cyanoPLP with lysine 5,6-aminomutase offers better prospect for the experimental detection of elusive cyclic azacyclopropylcarbinyl radical, which is proposed to be a key intermediate in the reaction of pyridoxal-5′-phosphate dependent radical aminomutases. - Highlights: • 4′-CyanoI{sup ·} is the lowest energy radical intermediate in the reaction of 5,6-LAM. • 4′-CyanoPLP offers good prospect for the experimental observation of elusive I{sup ·}. • The calculated HFCCs would help to characterize 4′-cyanoI{sup ·} by EPR. - Abstract: The results of our calculations suggest that the reaction of 4′-cyanoPLP with lysinemore » 5,6-aminomutase offers better prospect for the experimental detection of elusive cyclic azacyclopropylcarbinyl radical (I{sup ·}), which is proposed to be a key intermediate in the reaction of pyridoxal-5′-phosphate dependent radical aminomutases. We have calculated the corresponding hyperfine coupling constants (HFCCs) for {sup 14}N and {sup 13}C of cyano group using several basis sets to help the characterization of 4′-cyanoI{sup ·}.« less

Association of New Putative Epitopes of Myelin Proteolipid Protein (58-74) with Pathogenesis of Multiple Sclerosis.

PubMed

Zamanzadeh, Zahra; Ahangari, Ghasem; Ataei, Mitra; Pouragahi, Samie; Nabavi, Seyed Massood; Sadeghi, Mehdi; Sanati, Mohammad Hossein

2016-10-01

Multiple sclerosis (MS) is an autoimmune disease in which auto-reactive T cells react with self-antigens expressed in the central nervous system (CNS). The main cause of MS is unknown. Nonetheless, the most probable theory is based on molecular mimicry, which suggests that some infections can activate T cells against brain auto-antigens like myelin proteolipid protein (PLP) and initiate the disease cascade. This study is conducted to evaluate the activatory effects of PLP58-74 on T lymphocytes and humoral immunity. PLP58-74 was considered as an immunodominant epitope candidate of PLP using bioinformatics tools. Patients and healthy individuals' peripheral blood mononuclear cells (PBMCs) were treated with PLP58-74 and its proliferative effects were evaluated through assessing proliferating cell nuclear antigen (PCNA) gene expression changes by real time PCR and immunocytochemistry assay. Finally, the rate of CD4+ and CD8+ T cells were assessed by flowcytometry. ELISA was also performed to measure anti PLP58-74 antibody in patients' serum. PLP58-74 induced proliferation in patients' PBMCs while it did not influence PBMCs of healthy individuals. CD4+ T cells were the main activated cells in reaction to PLP58-74 which increased from 22% to 39.91%. In addition, immune assay showed threefold increase in specific anti PLP58-74 IgG in patients compared to healthy controls. Results showed that PLP58-74 can stimulate CD4+ T cells and humoral immunity. Therefore it seems that the epitopes of some microorganisms mimicking PLP such as PLP58-74 might have a potential role in the initiation of MS.

Identification and characterization of a pyridoxal 5'-phosphate phosphatase in the silkworm (Bombyx mori).

PubMed

Huang, ShuoHao; Han, CaiYun; Ma, ZhenQiao; Zhou, Jie; Zhang, JianYun; Huang, LongQuan

2017-03-01

Vitamin B 6 comprises six interconvertible pyridine compounds, among which pyridoxal 5'-phosphate (PLP) is a coenzyme for over 140 enzymes. PLP is also a very reactive aldehyde. The most well established mechanism for maintaining low levels of free PLP is its dephosphorylation by phosphatases. A human PLP-specific phosphatase has been identified and characterized. However, very little is known about the phosphatase in other living organisms. In this study, a cDNA clone of putative PLP phosphatase was identified from B. mori and characterized. The cDNA encodes a polypeptide of 343 amino acid residues, and the recombinant enzyme purified from E. coli exhibited properties similar to that of human PLP phosphatase. B. mori has a single copy of the PLPP gene, which is located on 11th chromosome, spans a 5.7kb region and contains five exons and four introns. PLP phosphatase transcript was detected in every larva tissue except hemolymph, and was most highly represented in Malpighian tube. We further down-regulated the gene expression of the PLP phosphatase in 5th instar larvae with the RNA interference. However, no significant changes in the gene expression of PLP biosynthetic enzymes and composition of B 6 vitamers were detected as compared with the control. Copyright © 2017 Elsevier Inc. All rights reserved.

Enhanced method of magnetic powder alignment for production of PLP Nd-Fe-B magnets

NASA Astrophysics Data System (ADS)

Popov, A. G.; Golovnia, O. A.; Protasov, A. V.

2017-04-01

It is demonstrated how the high degree of powder alignment in PLP magnets can be achieved by loading the powder into a container placed in a magnetic field of moderate strength. The strip-cast alloy with a composition of 30.00 Nd, 1.95 Dy, 66.42 Fe, 0.99 B, 0.54 Co, 0.1 Ga (wt%) was subjected to hydrogen decrepitation and then milled in a vibratory mill in toluene to an average particle size of 2.9 μm determined by the FSSS method. The powder was compacted in the magnetic field of 0.2 - 1.2 T to the filling density 2.6 - 3.2×103 kg/m3. It is shown that loading the powder into a container placed in a magnetic field enhances the degree of powder alignment in sintered Nd-Fe-B magnets produced from non-pressed powder. At the filling density less than 3.2×103 kg/m3, the density of magnets is high but insufficient, because of the formation of magnetostatic chains of particles, which impedes the powder compaction. The simulation by the discrete-element method qualitatively proves that the magnetostatic interaction of the chains of particles that are formed in the course of loading in the magnetic field stimulates a decrease in the density of the sintered magnets and its non-uniform distribution over the sample. As a result of the optimization of the parameters of the alignment and compaction of the powder loaded in a magnetic field, PLP magnets with Br ≥1.34 T, Hc ≥950 kA/m, (BH)max ≥340 kJ/m3, and the degree of alignment exceeding 96% were produced.

The chaperone role of the pyridoxal 5'-phosphate and its implications for rare diseases involving B6-dependent enzymes.

PubMed

Cellini, Barbara; Montioli, Riccardo; Oppici, Elisa; Astegno, Alessandra; Voltattorni, Carla Borri

2014-02-01

The biologically active form of the B6 vitamers is pyridoxal 5'-phosphate (PLP), which plays a coenzymatic role in several distinct enzymatic activities ranging from the synthesis, interconversion and degradation of amino acids to the replenishment of one-carbon units, synthesis and degradation of biogenic amines, synthesis of tetrapyrrolic compounds and metabolism of amino-sugars. In the catalytic process of PLP-dependent enzymes, the substrate amino acid forms a Schiff base with PLP and the electrophilicity of the PLP pyridine ring plays important roles in the subsequent catalytic steps. While the essential role of PLP in the acquisition of biological activity of many proteins is long recognized, the finding that some PLP-enzymes require the coenzyme for refolding in vitro points to an additional role of PLP as a chaperone in the folding process. Mutations in the genes encoding PLP-enzymes are causative of several rare inherited diseases. Patients affected by some of these diseases (AADC deficiency, cystathionuria, homocystinuria, gyrate atrophy, primary hyperoxaluria type 1, xanthurenic aciduria, X-linked sideroblastic anaemia) can benefit, although at different degrees, from the administration of pyridoxine, a PLP precursor. The effect of the coenzyme is not limited to mutations that affect the enzyme-coenzyme interaction, but also to those that cause folding defects, reinforcing the idea that PLP could play a chaperone role and improve the folding efficiency of misfolded variants. In this review, recent biochemical and cell biology studies highlighting the chaperoning activity of the coenzyme on folding-defective variants of PLP-enzymes associated with rare diseases are presented and discussed. Copyright © 2013 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

Girl with signs of Pelizaeus-Merzbacher disease heterozygous for a mutation in exon 2 of the proteolipid protein gene

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

Hodes, M.E.; DeMyer, W.E.; Pratt, V.M.

1995-02-13

We studied a female infant with clinical signs of Pelizaeus-Merzbacher disease (PMD), who has a familial mutation (C{sup 41}{r_arrow}T) in exon 2 of the proteolipid protein gene (PLP), and selected relatives. While the carrier mother and grandmother of the proposita currently are neurologically normal and show normal T2 magnetic resonance imaging (MRI) of the brain, the infant has a neurological picture, MRIs, and brain auditory evoked response (BAER) consistent with that diagnosis. The data here presented show that PMD can occur in females carrying a mutation in the PLP gene. Our experience with the MRIs of this patient, her mothermore » and grandmother, and those of a previously reported family show that molecular genetic analysis and not MRI is the appropriate means for carrier detection. 22 refs., 5 figs.« less

Genetic homogeneity of Pelizaeus-Merzbacher disease: Tight linkage to the proteolipoprotein locus in 16 affected families

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

Boespflug-Tanguy, O.; Mimault, C.; Cavagna, A.

1994-09-01

Among the numerous leukodystrophies that have an early onset and no biochemical markers, Pelizaeus-Merzbacher disease (PMD) is one that can be identified using strict clinical criteria and demonstrating an abnormal formation of myelin that is restricted to the CNS in electrophysiological studies and brain magnetic resonance imaging (MRI). In PMD, 12 different base substitutions and one total deletion of the genomic region containing the PLP gene have been reported, but, despite extensive analysis, PLP exon mutations have been found in only 10%-25% of the families analyzed. To test the genetic homogeneity of this disease, the authors have carried out linkagemore » analysis with polymorphic markers of the PLP genomic region in 16 families selected on strict diagnostic criteria of PMD. They observed a tight linkage of the PMD locus with markers of the PLP gene (cDNA PLP, exon IV polymorphism) and of the Xq22 region (DXS17, DXS94, and DXS287), whereas the markers located more proximally (DXYS1X and DXS3) or distally (DXS11) were not linked to the PMD locus. Multipoint analysis gave a maximal location score for the PMD locus (13.98) and the PLP gene (8.32) in the same interval between DXS94 and DXS287, suggesting that in all families PMD is linked to the PLP locus. Mutations of the extraexonic PLP gene sequences or of another unknown close gene could be involved in PMD. In an attempt to identify molecular defects of this genomic region that are responsible for PMD, these results meant that RFLP analysis could be used to improve genetic counseling for the numerous affected families in which a PLP exon mutation could not be demonstrated. 39 refs., 2 figs., 2 tabs.« less

Brain magnetic resonance imaging findings and auditory brainstem response in a child with spastic paraplegia 2 due to a PLP1 splice site mutation.

PubMed

Kubota, Kazuo; Saito, Yoshiaki; Ohba, Chihiro; Saitsu, Hirotomo; Fukuyama, Tetsuhiro; Ishiyama, Akihiko; Saito, Takashi; Komaki, Hirofumi; Nakagawa, Eiji; Sugai, Kenji; Sasaki, Masayuki; Matsumoto, Naomichi

2015-01-01

A boy with spastic paraplegia type 2 (SPG2) due to a novel splice site mutation of PLP1 presented with progressive spasticity of lower limbs, which was first observed during late infancy, when he gained the ability to walk with support. His speech was slow and he had dysarthria. The patient showed mildly delayed intellectual development. Subtotal dysmyelination in the central nervous system was revealed, which was especially prominent in structures known to be myelinated during earlier period, whereas structures that are myelinated later were better myelinated. These findings on the brain magnetic resonance imaging were unusual for subjects with PLP1 mutations. Peaks I and II of the auditory brainstem response (ABR) were normally provoked, but peaks III-V were not clearly demarcated, similarly to the findings in Pelizaeus-Merzbacher disease. These findings of brain MRI and ABR may be characteristic for a subtype of SPG2 patients. Copyright © 2014 The Japanese Society of Child Neurology. Published by Elsevier B.V. All rights reserved.

Altered GPM6A/M6 dosage impairs cognition and causes phenotypes responsive to cholesterol in human and Drosophila.

PubMed

Gregor, Anne; Kramer, Jamie M; van der Voet, Monique; Schanze, Ina; Uebe, Steffen; Donders, Rogier; Reis, André; Schenck, Annette; Zweier, Christiane

2014-12-01

Glycoprotein M6A (GPM6A) is a neuronal transmembrane protein of the PLP/DM20 (proteolipid protein) family that associates with cholesterol-rich lipid rafts and promotes filopodia formation. We identified a de novo duplication of the GPM6A gene in a patient with learning disability and behavioral anomalies. Expression analysis in blood lymphocytes showed increased GPM6A levels. An increase of patient-derived lymphoblastoid cells carrying membrane protrusions supports a functional effect of this duplication. To study the consequences of GPM6A dosage alterations in an intact nervous system, we employed Drosophila melanogaster as a model organism. We found that knockdown of Drosophila M6, the sole member of the PLP family in flies, in the wing, and whole organism causes malformation and lethality, respectively. These phenotypes as well as the protrusions of patient-derived lymphoblastoid cells with increased GPM6A levels can be alleviated by cholesterol supplementation. Notably, overexpression as well as loss of M6 in neurons specifically compromises long-term memory in the courtship conditioning paradigm. Our findings thus indicate a critical role of correct GPM6A/M6 levels for cognitive function and support a role of the GPM6A duplication for the patient's phenotype. Together with other recent findings, this study highlights compromised cholesterol homeostasis as a recurrent feature in cognitive phenotypes. © 2014 WILEY PERIODICALS, INC.

Pyridoxal phosphate synthases PdxS/PdxT are required for Actinobacillus pleuropneumoniae viability, stress tolerance and virulence.

PubMed

Xie, Fang; Li, Gang; Wang, Yalei; Zhang, Yanhe; Zhou, Long; Wang, Chengcheng; Liu, Shuanghong; Liu, Siguo; Wang, Chunlai

2017-01-01

Pyridoxal 5'-phosphate (PLP) is an essential cofactor for numerous enzymes involved in a diversity of cellular processes in living organisms. Previous analysis of the Actinobacillus pleuropneumoniae S-8 genome sequence revealed the presence of pdxS and pdxT genes, which are implicated in deoxyxylulose 5-phosphate (DXP)-independent pathway of PLP biosynthesis; however, little is known about their roles in A. pleuropneumoniae pathogenicity. Our data demonstrated that A. pleuropneumoniae could synthesize PLP by PdxS and PdxT enzymes. Disruption of the pdxS and pdxT genes rendered the pathogen auxotrophic for PLP, and the defective growth as a result of these mutants was chemically compensated by the addition of PLP, suggesting the importance of PLP production for A. pleuropneumoniae growth and viability. Additionally, the pdxS and pdxT deletion mutants displayed morphological defects as indicated by irregular and aberrant shapes in the absence of PLP. The reduced growth of the pdxS and pdxT deletion mutants under osmotic and oxidative stress conditions suggests that the PLP synthases PdxS/PdxT are associated with the stress tolerance of A. pleuropneumoniae. Furthermore, disruption of the PLP biosynthesis pathway led to reduced colonization and attenuated virulence of A. pleuropneumoniae in the BALB/c mouse model. The data presented in this study reveal the critical role of PLP synthases PdxS/PdxT in viability, stress tolerance, and virulence of A. pleuropneumoniae.

Decreased number and increased volume with mitochondrial enlargement of cerebellar synaptic terminals in a mouse model of chronic demyelination.

PubMed

Nguyen, Huy Bang; Sui, Yang; Thai, Truc Quynh; Ikenaka, Kazuhiro; Oda, Toshiyuki; Ohno, Nobuhiko

2018-05-23

Impaired nerve conduction, axonal degeneration, and synaptic alterations contribute to neurological disabilities in inflammatory demyelinating diseases. Cerebellar dysfunction is associated with demyelinating disorders, but the alterations of axon terminals in cerebellar gray matter during chronic demyelination are still unclear. We analyzed the morphological and ultrastructural changes of climbing fiber terminals in a mouse model of hereditary chronic demyelination. Three-dimensional ultrastructural analyses using serial block-face scanning electron microscopy and immunostaining for synaptic markers were performed in a demyelination mouse model caused by extra copies of myelin gene (PLP4e). At 1 month old, many myelinated axons were observed in PLP4e and wild-type mice, but demyelinated axons and axons with abnormally thin myelin were prominent in PLP4e mice at 5 months old. The density of climbing fiber terminals was significantly reduced in PLP4e mice at 5 months old. Reconstruction of climbing fiber terminals revealed that PLP4e climbing fibers had increased varicosity volume and enlarged mitochondria in the varicosities at 5-month-old mice. These results suggest that chronic demyelination is associated with alterations and loss of climbing fiber terminals in the cerebellar cortex, and that synaptic changes may contribute to cerebellar phenotypes observed in hereditary demyelinating disorders.

Linkage analysis and physical mapping near the gene for x-linked agammaglobulinemia at Xq22

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

Parolini, O.; Lassiter, G.L.; Henry, M.J.

The gene for x-linked agammaglobulinemia (XLA) has been mapped to Xq22. No recombinations have been reported between the gene and the prob p212 at DXS178; however, this probe is informative in only 30-40% of women and the reported flanking markers, DXS3 and DXS94, and 10-15 cM apart. To identify additional probes that might be useful in genetic counseling, we examined 11 polymorphisms that have been mapped to the Xq21.3-q22 region in 13 families with XLA. In addition, pulsed-field gel electrophoresis and yeast artificial chromosomes (YACs) were used to further characterize the segman of DNA within which the gene for SLAmore » must lie. The results demonstrated that DXS366 and DXS442, which share a 430-kb pulsed-field fragment, could replace DXS3 as proximal flanking markers. Probes at DXS178 and DXS265 identified the same 145-kb pulsed-field fragment, and both loci were contained within a 200-kb YAC identified with the probe p212. A highly polymorphic CA repeat (DCS178CA) was isolated from one end of this YAC and used in linkage analysis. Probes at DXS101 and DXS328 shared several pulsed-field fragments, the smallest of which was 250 kb. No recombinations were seen between XLA and the DXS178-DXS265-DXS178CA complex, DXS101, DXS328, DXS87, or the gene for proteolipid protein (PLP). Key crossovers, when combined with the linkage data from families with Alport syndrome, suggested the following order of loci: cen-DXS3-DXS366-DXS442-(PLP, DXS101, DXS328, DXS178-DXS265-DXS178CA complex, XL)-(DXS87, DXS94)-DXS327-(DXS350, DXS362)-tel. Our studies also limit the segment of DNA within which the XLA gene must lie to the 3- to 4-cM distance between DCS442 and DXS94 and they identify and orient polymorphisms that can be used in genetic counseling not only for XLA but also for Pelizaeus-Merzbacher disease (PLP deficiency), Alport syndrome (COL4A5 deficiency), and Fabry disease ([alpha]-galactosidase A difficiency). 31 refs., 5 figs., 2 tabs.« less

Mitochondrial Hsp60 Chaperonopathy Causes an Autosomal-Recessive Neurodegenerative Disorder Linked to Brain Hypomyelination and Leukodystrophy

PubMed Central

Magen, Daniella; Georgopoulos, Costa; Bross, Peter; Ang, Debbie; Segev, Yardena; Goldsher, Dorit; Nemirovski, Alexandra; Shahar, Eli; Ravid, Sarit; Luder, Anthony; Heno, Bayan; Gershoni-Baruch, Ruth; Skorecki, Karl; Mandel, Hanna

2008-01-01

Hypomyelinating leukodystrophies (HMLs) are disorders involving aberrant myelin formation. The prototype of primary HMLs is the X-linked Pelizaeus-Merzbacher disease (PMD) caused by mutations in PLP1. Recently, homozygous mutations in GJA12 encoding connexin 47 were found in patients with autosomal-recessive Pelizaeus-Merzbacher-like disease (PMLD). However, many patients of both genders with PMLD carry neither PLP1 nor GJA12 mutations. We report a consanguineous Israeli Bedouin kindred with clinical and radiological findings compatible with PMLD, in which linkage to PLP1 and GJA12 was excluded. Using homozygosity mapping and mutation analysis, we have identified a homozygous missense mutation (D29G) not previously described in HSPD1, encoding the mitochondrial heat-shock protein 60 (Hsp60) in all affected individuals. The D29G mutation completely segregates with the disease-associated phenotype. The pathogenic effect of D29G on Hsp60-chaperonin activity was verified by an in vivo E. coli complementation assay, which demonstrated compromised ability of the D29G-Hsp60 mutant protein to support E. coli survival, especially at high temperatures. The disorder, which we have termed MitCHAP-60 disease, can be distinguished from spastic paraplegia 13 (SPG13), another Hsp60-associated autosomal-dominant neurodegenerative disorder, by its autosomal-recessive inheritance pattern, as well as by its early-onset, profound cerebral involvement and lethality. Our findings suggest that Hsp60 defects can cause neurodegenerative pathologies of varying severity, not previously suspected on the basis of the SPG13 phenotype. These findings should help to clarify the important role of Hsp60 in myelinogenesis and neurodegeneration. PMID:18571143

Comprehensive Cancer-Predisposition Gene Testing in an Adult Multiple Primary Tumor Series Shows a Broad Range of Deleterious Variants and Atypical Tumor Phenotypes.

PubMed

Whitworth, James; Smith, Philip S; Martin, Jose-Ezequiel; West, Hannah; Luchetti, Andrea; Rodger, Faye; Clark, Graeme; Carss, Keren; Stephens, Jonathan; Stirrups, Kathleen; Penkett, Chris; Mapeta, Rutendo; Ashford, Sofie; Megy, Karyn; Shakeel, Hassan; Ahmed, Munaza; Adlard, Julian; Barwell, Julian; Brewer, Carole; Casey, Ruth T; Armstrong, Ruth; Cole, Trevor; Evans, Dafydd Gareth; Fostira, Florentia; Greenhalgh, Lynn; Hanson, Helen; Henderson, Alex; Hoffman, Jonathan; Izatt, Louise; Kumar, Ajith; Kwong, Ava; Lalloo, Fiona; Ong, Kai Ren; Paterson, Joan; Park, Soo-Mi; Chen-Shtoyerman, Rakefet; Searle, Claire; Side, Lucy; Skytte, Anne-Bine; Snape, Katie; Woodward, Emma R; Tischkowitz, Marc D; Maher, Eamonn R

2018-06-12

Multiple primary tumors (MPTs) affect a substantial proportion of cancer survivors and can result from various causes, including inherited predisposition. Currently, germline genetic testing of MPT-affected individuals for variants in cancer-predisposition genes (CPGs) is mostly targeted by tumor type. We ascertained pre-assessed MPT individuals (with at least two primary tumors by age 60 years or at least three by 70 years) from genetics centers and performed whole-genome sequencing (WGS) on 460 individuals from 440 families. Despite previous negative genetic assessment and molecular investigations, pathogenic variants in moderate- and high-risk CPGs were detected in 67/440 (15.2%) probands. WGS detected variants that would not be (or were not) detected by targeted resequencing strategies, including low-frequency structural variants (6/440 [1.4%] probands). In most individuals with a germline variant assessed as pathogenic or likely pathogenic (P/LP), at least one of their tumor types was characteristic of variants in the relevant CPG. However, in 29 probands (42.2% of those with a P/LP variant), the tumor phenotype appeared discordant. The frequency of individuals with truncating or splice-site CPG variants and at least one discordant tumor type was significantly higher than in a control population (χ 2 = 43.642; p ≤ 0.0001). 2/67 (3%) probands with P/LP variants had evidence of multiple inherited neoplasia allele syndrome (MINAS) with deleterious variants in two CPGs. Together with variant detection rates from a previous series of similarly ascertained MPT-affected individuals, the present results suggest that first-line comprehensive CPG analysis in an MPT cohort referred to clinical genetics services would detect a deleterious variant in about a third of individuals. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

Genes and Mutations Causing Autosomal Dominant Retinitis Pigmentosa

PubMed Central

Daiger, Stephen P.; Bowne, Sara J.; Sullivan, Lori S.

2015-01-01

Retinitis pigmentosa (RP) has a prevalence of approximately one in 4000; 25%–30% of these cases are autosomal dominant retinitis pigmentosa (adRP). Like other forms of inherited retinal disease, adRP is exceptionally heterogeneous. Mutations in more than 25 genes are known to cause adRP, more than 1000 mutations have been reported in these genes, clinical findings are highly variable, and there is considerable overlap with other types of inherited disease. Currently, it is possible to detect disease-causing mutations in 50%–75% of adRP families in select populations. Genetic diagnosis of adRP has advantages over other forms of RP because segregation of disease in families is a useful tool for identifying and confirming potentially pathogenic variants, but there are disadvantages too. In addition to identifying the cause of disease in the remaining 25% of adRP families, a central challenge is reconciling clinical diagnosis, family history, and molecular findings in patients and families. PMID:25304133

Neurotoxocarosis alters myelin protein gene transcription and expression.

PubMed

Heuer, Lea; Beyerbach, Martin; Lühder, Fred; Beineke, Andreas; Strube, Christina

2015-06-01

Neurotoxocarosis is an infection of the central nervous system caused by migrating larvae of the common dog and cat roundworms (Toxocara canis and Toxocara cati), which are zoonotic agents. As these parasites are prevalent worldwide and neuropathological and molecular investigations on neurotoxocarosis are scare, this study aims to characterise nerve fibre demyelination associated with neurotoxocarosis on a molecular level. Transcription of eight myelin-associated genes (Cnp, Mag, Mbp, Mog, Mrf-1, Nogo-A, Plp1, Olig2) was determined in the mouse model during six time points of the chronic phase of infection using qRT-PCR. Expression of selected proteins was analysed by Western blotting or immunohistochemistry. Additionally, demyelination and neuronal damage were investigated histologically. Significant differences (p ≤ 0.05) between transcription rates of T. canis-infected and uninfected control mice were detected for all analysed genes while T. cati affected five of eight investigated genes. Interestingly, 2', 3 ´-cyclic nucleotide 3'-phosphodiesterase (Cnp) and myelin oligodendrocyte glycoprotein (Mog) were upregulated in both T. canis- and T. cati-infected mice preceding demyelination. Later, CNPase expression was additionally enhanced. As expected, myelin basic protein (Mbp) was downregulated in cerebra and cerebella of T. canis-infected mice when severe demyelination was present 120 days post infectionem (dpi). The transcriptional pattern observed in the present study appears to reflect direct traumatic and hypoxic effects of larval migration as well as secondary processes including host immune reactions, demyelination and attempts to remyelinate damaged areas.

An essential cell cycle regulation gene causes hybrid inviability in Drosophila

PubMed Central

Phadnis, Nitin; Baker, EmilyClare P.; Cooper, Jacob C.; Frizzell, Kimberly A.; Hsieh, Emily; de la Cruz, Aida Flor A.; Shendure, Jay; Kitzman, Jacob O.; Malik, Harmit S.

2015-01-01

Speciation, the process by which new biological species arise, involves the evolution of reproductive barriers such as hybrid sterility or inviability between populations. However, identifying hybrid incompatibility genes remains a key obstacle in understanding the molecular basis of reproductive isolation. We devised a genomic screen, which identified a cell cycle regulation gene as the cause of male inviability in hybrids between Drosophila melanogaster and D. simulans. Ablation of the D. simulans allele of this gene is sufficient to rescue the adult viability of hybrid males. This dominantly acting cell cycle regulator causes mitotic arrest and, thereby, inviability of male hybrid larvae. Our genomic method provides a facile means to accelerate the identification of hybrid incompatibility genes in other model and non-model systems. PMID:26680200

Mutations in the pericentrin (PCNT) gene cause primordial dwarfism.

PubMed

Rauch, Anita; Thiel, Christian T; Schindler, Detlev; Wick, Ursula; Crow, Yanick J; Ekici, Arif B; van Essen, Anthonie J; Goecke, Timm O; Al-Gazali, Lihadh; Chrzanowska, Krystyna H; Zweier, Christiane; Brunner, Han G; Becker, Kristin; Curry, Cynthia J; Dallapiccola, Bruno; Devriendt, Koenraad; Dörfler, Arnd; Kinning, Esther; Megarbane, André; Meinecke, Peter; Semple, Robert K; Spranger, Stephanie; Toutain, Annick; Trembath, Richard C; Voss, Egbert; Wilson, Louise; Hennekam, Raoul; de Zegher, Francis; Dörr, Helmuth-Günther; Reis, André

2008-02-08

Fundamental processes influencing human growth can be revealed by studying extreme short stature. Using genetic linkage analysis, we find that biallelic loss-of-function mutations in the centrosomal pericentrin (PCNT) gene on chromosome 21q22.3 cause microcephalic osteodysplastic primordial dwarfism type II (MOPD II) in 25 patients. Adults with this rare inherited condition have an average height of 100 centimeters and a brain size comparable to that of a 3-month-old baby, but are of near-normal intelligence. Absence of PCNT results in disorganized mitotic spindles and missegregation of chromosomes. Mutations in related genes are known to cause primary microcephaly (MCPH1, CDK5RAP2, ASPM, and CENPJ).

Gene expression profiling of the hippocampal dentate gyrus in an adult toxicity study captures a variety of neurodevelopmental dysfunctions in rat models of hypothyroidism.

PubMed

Shiraki, Ayako; Saito, Fumiyo; Akane, Hirotoshi; Akahori, Yumi; Imatanaka, Nobuya; Itahashi, Megu; Yoshida, Toshinori; Shibutani, Makoto

2016-01-01

We previously found that developmental hypothyroidism changed the expression of genes in the rat hippocampal dentate gyrus, a brain region where adult neurogenesis is known to occur. In the present study, we performed brain region-specific global gene expression profiling in an adult rat hypothyroidism model to see if it reflected the developmental neurotoxicity we saw in the developmental hypothyroidism model. Starting when male rats were 5 weeks old, we administered 6-propyl-2-thiouracil at a doses of 0, 0.1 and 10 mg kg(-1) body weight by gavage for 28 days. We selected four brain regions to represent both cerebral and cerebellar tissues: hippocampal dentate gyrus, cerebral cortex, corpus callosum and cerebellar vermis. We observed significant alterations in the expression of genes related to neural development (Eph family genes and Robo3) in the cerebral cortex and hippocampal dentate gyrus and in the expression of genes related to myelination (Plp1 and Mbp) in the hippocampal dentate gyrus. We observed only minor changes in the expression of these genes in the corpus callosum and cerebellar vermis. We used real-time reverse-transcription polymerase chain reaction to confirm Chrdl1, Hes5, Mbp, Plp1, Slit1, Robo3 and the Eph family transcript expression changes. The most significant changes in gene expression were found in the dentate gyrus. Considering that the gene expression profile of the adult dentate gyrus closely related to neurogenesis, 28-day toxicity studies looking at gene expression changes in adult hippocampal dentate gyrus may also detect possible developmental neurotoxic effects. Copyright © 2015 John Wiley & Sons, Ltd.

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.

A single gene causes both male sterility and segregation distortion in Drosophila hybrids*

PubMed Central

Phadnis, Nitin; Orr, H. Allen

2008-01-01

A central goal of evolutionary biology is to identify the genes and evolutionary forces that cause speciation, the emergence of reproductive isolation between populations. Despite the identification of several genes that cause hybrid sterility or inviability— many of which have evolved rapidly under positive Darwinian selection— little is known about the ecological or genomic forces that drive the evolution of postzygotic isolation. Here we show that the same gene, Overdrive, causes both male sterility and segregation distortion in F1 hybrids between the Bogota and USA subspecies of Drosophila pseudoobscura. This segregation distorter gene is essential for hybrid sterility, a strong reproductive barrier between these young taxa. Our results suggest that genetic conflict may be an important evolutionary force in speciation. PMID:19074311

A single gene causes both male sterility and segregation distortion in Drosophila hybrids.

PubMed

Phadnis, Nitin; Orr, H Allen

2009-01-16

A central goal of evolutionary biology is to identify the genes and evolutionary forces that cause speciation, the emergence of reproductive isolation between populations. Despite the identification of several genes that cause hybrid sterility or inviability-many of which have evolved rapidly under positive Darwinian selection-little is known about the ecological or genomic forces that drive the evolution of postzygotic isolation. Here, we show that the same gene, Overdrive, causes both male sterility and segregation distortion in F1 hybrids between the Bogota and U.S. subspecies of Drosophila pseudoobscura. This segregation distorter gene is essential for hybrid sterility, a strong reproductive barrier between these young taxa. Our results suggest that genetic conflict may be an important evolutionary force in speciation.

An essential cell cycle regulation gene causes hybrid inviability in Drosophila.

PubMed

Phadnis, Nitin; Baker, EmilyClare P; Cooper, Jacob C; Frizzell, Kimberly A; Hsieh, Emily; de la Cruz, Aida Flor A; Shendure, Jay; Kitzman, Jacob O; Malik, Harmit S

2015-12-18

Speciation, the process by which new biological species arise, involves the evolution of reproductive barriers, such as hybrid sterility or inviability between populations. However, identifying hybrid incompatibility genes remains a key obstacle in understanding the molecular basis of reproductive isolation. We devised a genomic screen, which identified a cell cycle-regulation gene as the cause of male inviability in hybrids resulting from a cross between Drosophila melanogaster and D. simulans. Ablation of the D. simulans allele of this gene is sufficient to rescue the adult viability of hybrid males. This dominantly acting cell cycle regulator causes mitotic arrest and, thereby, inviability of male hybrid larvae. Our genomic method provides a facile means to accelerate the identification of hybrid incompatibility genes in other model and nonmodel systems. Copyright © 2015, American Association for the Advancement of Science.

Measurement of pyridoxal 5'-phosphate, pyridoxal, and 4-pyridoxic acid in the cerebrospinal fluid of children.

PubMed

Akiyama, Tomoyuki; Akiyama, Mari; Hayashi, Yumiko; Shibata, Takashi; Hanaoka, Yoshiyuki; Toda, Soichiro; Imai, Katsumi; Hamano, Shin-Ichiro; Okanishi, Tohru; Yoshinaga, Harumi; Kobayashi, Katsuhiro

2017-03-01

We quantified pyridoxal 5'-phosphate (PLP), pyridoxal (PL), and 4-pyridoxic acid (PA) in the cerebrospinal fluid (CSF) of children and to investigate the effect of age, sex, epilepsy, and anti-epileptic drug (AED) therapy on these vitamers. CSF samples prospectively collected from 116 pediatric patients were analyzed. PLP, PL, and PA were measured using high-performance liquid chromatography with fluorescence detection, using pre-column derivatization by semicarbazide. Effects of age, sex, epilepsy, and AEDs on these vitamers and the PLP/PL ratio were evaluated using multiple linear regression models. The PLP, PL, and PA concentrations were correlated negatively with age and the PLP/PL ratio was correlated positively with age. Multiple regression analysis revealed that the presence of epilepsy was associated with lower PLP concentrations and PLP/PL ratios but sex and AED therapy had no influence on these values. The observed ranges of these vitamers in epileptic and non-epileptic patients were demonstrated. We showed the age dependence of PLP and PL in CSF from pediatric patients. Epileptic patients had lower PLP concentrations and PLP/PL ratios than non-epileptic patients, but it is unknown whether this is the cause, or a result, of epilepsy. Copyright © 2016 Elsevier B.V. All rights reserved.

Loss of Sfpq Causes Long-Gene Transcriptopathy in the Brain.

PubMed

Takeuchi, Akihide; Iida, Kei; Tsubota, Toshiaki; Hosokawa, Motoyasu; Denawa, Masatsugu; Brown, J B; Ninomiya, Kensuke; Ito, Mikako; Kimura, Hiroshi; Abe, Takaya; Kiyonari, Hiroshi; Ohno, Kinji; Hagiwara, Masatoshi

2018-05-01

Genes specifically expressed in neurons contain members with extended long introns. Longer genes present a problem with respect to fulfilment of gene length transcription, and evidence suggests that dysregulation of long genes is a mechanism underlying neurodegenerative and psychiatric disorders. Here, we report the discovery that RNA-binding protein Sfpq is a critical factor for maintaining transcriptional elongation of long genes. We demonstrate that Sfpq co-transcriptionally binds to long introns and is required for sustaining long-gene transcription by RNA polymerase II through mediating the interaction of cyclin-dependent kinase 9 with the elongation complex. Phenotypically, Sfpq disruption caused neuronal apoptosis in developing mouse brains. Expression analysis of Sfpq-regulated genes revealed specific downregulation of developmentally essential neuronal genes longer than 100 kb in Sfpq-disrupted brains; those genes are enriched in associations with neurodegenerative and psychiatric diseases. The identified molecular machinery yields directions for targeted investigations of the association between long-gene transcriptopathy and neuronal diseases. Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.

NDRC: A Disease-Causing Genes Prioritized Method Based on Network Diffusion and Rank Concordance.

PubMed

Fang, Minghong; Hu, Xiaohua; Wang, Yan; Zhao, Junmin; Shen, Xianjun; He, Tingting

2015-07-01

Disease-causing genes prioritization is very important to understand disease mechanisms and biomedical applications, such as design of drugs. Previous studies have shown that promising candidate genes are mostly ranked according to their relatedness to known disease genes or closely related disease genes. Therefore, a dangling gene (isolated gene) with no edges in the network can not be effectively prioritized. These approaches tend to prioritize those genes that are highly connected in the PPI network while perform poorly when they are applied to loosely connected disease genes. To address these problems, we propose a new disease-causing genes prioritization method that based on network diffusion and rank concordance (NDRC). The method is evaluated by leave-one-out cross validation on 1931 diseases in which at least one gene is known to be involved, and it is able to rank the true causal gene first in 849 of all 2542 cases. The experimental results suggest that NDRC significantly outperforms other existing methods such as RWR, VAVIEN, DADA and PRINCE on identifying loosely connected disease genes and successfully put dangling genes as potential candidate disease genes. Furthermore, we apply NDRC method to study three representative diseases, Meckel syndrome 1, Protein C deficiency and Peroxisome biogenesis disorder 1A (Zellweger). Our study has also found that certain complex disease-causing genes can be divided into several modules that are closely associated with different disease phenotype.

Mutations in the deubiquitinase gene USP8 cause Cushing's disease.

PubMed

Reincke, Martin; Sbiera, Silviu; Hayakawa, Akira; Theodoropoulou, Marily; Osswald, Andrea; Beuschlein, Felix; Meitinger, Thomas; Mizuno-Yamasaki, Emi; Kawaguchi, Kohei; Saeki, Yasushi; Tanaka, Keiji; Wieland, Thomas; Graf, Elisabeth; Saeger, Wolfgang; Ronchi, Cristina L; Allolio, Bruno; Buchfelder, Michael; Strom, Tim M; Fassnacht, Martin; Komada, Masayuki

2015-01-01

Cushing's disease is caused by corticotroph adenomas of the pituitary. To explore the molecular mechanisms of endocrine autonomy in these tumors, we performed exome sequencing of 10 corticotroph adenomas. We found somatic mutations in the USP8 deubiquitinase gene in 4 of 10 adenomas. The mutations clustered in the 14-3-3 protein binding motif and enhanced the proteolytic cleavage and catalytic activity of USP8. Cleavage of USP8 led to increased deubiqutination of the EGF receptor, impairing its downregulation and sustaining EGF signaling. USP8 mutants enhanced promoter activity of the gene encoding proopiomelanocortin. In summary, our data show that dominant mutations in USP8 cause Cushing's disease via activation of EGF receptor signaling.

Genes of the de novo and Salvage Biosynthesis Pathways of Vitamin B6 are Regulated under Oxidative Stress in the Plant Pathogen Rhizoctonia solani

PubMed Central

Samsatly, Jamil; Chamoun, Rony; Gluck-Thaler, Emile; Jabaji, Suha

2016-01-01

Vitamin B6 is recognized as an important cofactor required for numerous metabolic enzymes, and has been shown to act as an antioxidant and play a role in stress responses. It can be synthesized through two different routes: salvage and de novo pathways. However, little is known about the possible function of the vitamin B6 pathways in the fungal plant pathogen Rhizoctonia solani. Using genome walking, the de novo biosynthetic pathway genes; RsolPDX1 and RsolPDX2 and the salvage biosynthetic pathway gene, RsolPLR were sequenced. The predicted amino acid sequences of the three genes had high degrees of similarity to other fungal PDX1, PDX2, and PLR proteins and are closely related to other R. solani anastomosis groups. We also examined their regulation when subjected to reactive oxygen species (ROS) stress inducers, the superoxide generator paraquat, or H2O2, and compared it to the well-known antioxidant genes, catalase and glutathione-S-transferase (GST). The genes were differentially regulated with transcript levels as high as 33 fold depending on the gene and type of stress reflecting differences in the type of damage induced by ROS. Exogenous addition of the vitamers PN or PLP in culture medium significantly induced the transcription of the vitamin B6 de novo encoding genes as early as 0.5 hour post treatment (HPT). On the other hand, transcription of RsolPLR was vitamer-specific; a down regulation upon supplementation of PN and upregulation with PLP. Our results suggest that accumulation of ROS in R. solani mycelia is linked to transcriptional regulation of the three genes and implicate the vitamin B6 biosynthesis machinery in R. solani, similar to catalases and GST, as an antioxidant stress protector against oxidative stress. PMID:26779127

A novel splicing mutation in GALT gene causing Galactosemia in Ecuadorian family.

PubMed

De Lucca, M; Barba, C; Casique, L

2017-07-01

Classic Galactosemia (OMIM 230400) is an autosomal recessive disorder of galactose metabolism caused by mutations in the galactose-1-phosphate uridyl transferase (GALT) gene. This disease caused by the inability to metabolize galactose is potentially life-threatening but its pathophysiology has not been clearly defined. GALT gene presents high allelic heterogeneity and around 336 variations have been identified. Here, we report the case of a patient with Classic Galactosemia who was detected during a neonatal screening in Ecuador. Molecular study revealed a mutation in GALT gene intron 1, c.82+3A>G in homozygous condition, this mutation has not been previously reported. This gene variation was not found in any of the 119 healthy Ecuadorian individuals used as control. Furthermore, the mutation was the only alteration detected in the propositus's GALT after sequencing all exons and introns of this gene. In silico modeling predicted that the mutation was pathogenic. Copyright © 2017. Published by Elsevier B.V.

Comprehensive detection of genes causing a phenotype using phenotype sequencing and pathway analysis.

PubMed

Harper, Marc; Gronenberg, Luisa; Liao, James; Lee, Christopher

2014-01-01

Discovering all the genetic causes of a phenotype is an important goal in functional genomics. We combine an experimental design for detecting independent genetic causes of a phenotype with a high-throughput sequencing analysis that maximizes sensitivity for comprehensively identifying them. Testing this approach on a set of 24 mutant strains generated for a metabolic phenotype with many known genetic causes, we show that this pathway-based phenotype sequencing analysis greatly improves sensitivity of detection compared with previous methods, and reveals a wide range of pathways that can cause this phenotype. We demonstrate our approach on a metabolic re-engineering phenotype, the PEP/OAA metabolic node in E. coli, which is crucial to a substantial number of metabolic pathways and under renewed interest for biofuel research. Out of 2157 mutations in these strains, pathway-phenoseq discriminated just five gene groups (12 genes) as statistically significant causes of the phenotype. Experimentally, these five gene groups, and the next two high-scoring pathway-phenoseq groups, either have a clear connection to the PEP metabolite level or offer an alternative path of producing oxaloacetate (OAA), and thus clearly explain the phenotype. These high-scoring gene groups also show strong evidence of positive selection pressure, compared with strictly neutral selection in the rest of the genome.

Staphylococcal food poisoning caused by Staphylococcus argenteus harboring staphylococcal enterotoxin genes.

PubMed

Wakabayashi, Yuki; Umeda, Kaoru; Yonogi, Shinya; Nakamura, Hiromi; Yamamoto, Kaori; Kumeda, Yuko; Kawatsu, Kentaro

2018-01-16

Staphylococcal food poisoning (SFP) is caused by staphylococcal enterotoxins (SEs) preformed in food materials. SE genes are encoded on mobile genetic elements and are widely found across Staphylococcus species including S. argenteus, although most SFP cases are caused by S. aureus. S. argenteus, recently discriminated from S. aureus as a novel species, are non-pigmented staphylococci phenotypically related to S. aureus. In 2014 and 2015, two independent food poisoning cases occurred in Osaka, Japan, in which non-pigmented staphylococci were predominantly isolated. Several enterotoxin genes (seb, seg, sei, sem, sen, seo, and selu2) were found in their genome and the production of SEB was confirmed by reverse passive agglutination tests. The non-pigmented isolates from patients, food handlers, food, and cooking utensils all produced the same pulsed-field gel electrophoresis pattern. These non-pigmented isolates were coagulase-positive and biochemically identical to S. aureus. We performed further genetic analysis using nucA sequencing and multi-locus sequence typing, and identified these isolates as S. argenteus. We also found that seb was encoded on the Staphylococcus aureus pathogenicity island, while seg, sei, sem, sen, seo, and selu2 were encoded on the enterotoxin gene cluster. From these results, we concluded that the two food poisoning outbreaks were SFP cases caused by S. argenteus harboring SE genes. Copyright © 2017 Elsevier B.V. All rights reserved.

Pelizaeus-Merzbacher disease in patients with molecularly confirmed diagnosis.

PubMed

Mierzewska, H; Jamroz, E; Mazurczak, T; Hoffman-Zacharska, D; Szczepanik, E

2016-01-01

Pelizaeus-Merzbacher disease (PMD) is X-linked hypomyelinating leukodystrophy caused by mutations of the PLP1 gene, which codes the proteolipid protein 1. The result of mutations is abnormal myelination - hypomyelination and dysmyelination of cerebral white matter, and in some form of the disease hypomyelinating peripheral neuropathy. DNA samples from 68 patients suspected of PMD due to the clinical course and hypomyelination at magnetic resonance imaging (MRI) were analyzed. Medical history and detailed clinical course of PMD patients were also analyzed. Different mutations of the PLP1 gene were detected in 14 boys from 11 families (~20%). Amongst the molecularly confirmed patients, 13 presented classical PMD forms but clinical phenotypes varied in the severity even amongst siblings. One patient presented a severe connatal form. One mother, obligate carrier, presented complicated SPG2 (spastic paraparesis). There was no phenotype-genotype correlation in our material. In many cases PMD was suspected with a delay of many years, sometimes only after birth of another affected child in the family. Pelizaeus-Merzbacher disease was most frequently misdiagnosed as cerebral palsy.

A Naturally Occurring Recombinant Enterovirus Expresses a Torovirus Deubiquitinase.

PubMed

Shang, Pengcheng; Misra, Saurav; Hause, Ben; Fang, Ying

2017-07-15

Enteroviruses (EVs) are implicated in a wide range of diseases in humans and animals. In this study, a novel enterovirus (enterovirus species G [EVG]) (EVG 08/NC_USA/2015) was isolated from a diagnostic sample from a neonatal pig diarrhea case and identified by using metagenomics and complete genome sequencing. The viral genome shares 75.4% nucleotide identity with a prototypic EVG strain (PEV9 UKG/410/73). Remarkably, a 582-nucleotide insertion, flanked by 3C pro cleavage sites at the 5' and 3' ends, was found in the 2C/3A junction region of the viral genome. This insertion encodes a predicted protease with 54 to 68% amino acid identity to torovirus (ToV) papain-like protease (PLP) (ToV-PLP). Structural homology modeling predicts that this protease adopts a fold and a catalytic site characteristic of minimal PLP catalytic domains. This structure is similar to those of core catalytic domains of the foot-and-mouth disease virus leader protease and coronavirus PLPs, which act as deubiquitinating and deISGylating (interferon [IFN]-stimulated gene 15 [ISG15]-removing) enzymes on host cell substrates. Importantly, the recombinant ToV-PLP protein derived from this novel enterovirus also showed strong deubiquitination and deISGylation activities and demonstrated the ability to suppress IFN-β expression. Using reverse genetics, we generated a ToV-PLP knockout recombinant virus. Compared to the wild-type virus, the ToV-PLP knockout mutant virus showed impaired growth and induced higher expression levels of innate immune genes in infected cells. These results suggest that ToV-PLP functions as an innate immune antagonist; enterovirus G may therefore gain fitness through the acquisition of ToV-PLP from a recombination event. IMPORTANCE Enteroviruses comprise a highly diversified group of viruses. Genetic recombination has been considered a driving force for viral evolution; however, recombination between viruses from two different orders is a rare event. In this study, we

A Naturally Occurring Recombinant Enterovirus Expresses a Torovirus Deubiquitinase

PubMed Central

Shang, Pengcheng; Misra, Saurav

2017-01-01

ABSTRACT Enteroviruses (EVs) are implicated in a wide range of diseases in humans and animals. In this study, a novel enterovirus (enterovirus species G [EVG]) (EVG 08/NC_USA/2015) was isolated from a diagnostic sample from a neonatal pig diarrhea case and identified by using metagenomics and complete genome sequencing. The viral genome shares 75.4% nucleotide identity with a prototypic EVG strain (PEV9 UKG/410/73). Remarkably, a 582-nucleotide insertion, flanked by 3Cpro cleavage sites at the 5′ and 3′ ends, was found in the 2C/3A junction region of the viral genome. This insertion encodes a predicted protease with 54 to 68% amino acid identity to torovirus (ToV) papain-like protease (PLP) (ToV-PLP). Structural homology modeling predicts that this protease adopts a fold and a catalytic site characteristic of minimal PLP catalytic domains. This structure is similar to those of core catalytic domains of the foot-and-mouth disease virus leader protease and coronavirus PLPs, which act as deubiquitinating and deISGylating (interferon [IFN]-stimulated gene 15 [ISG15]-removing) enzymes on host cell substrates. Importantly, the recombinant ToV-PLP protein derived from this novel enterovirus also showed strong deubiquitination and deISGylation activities and demonstrated the ability to suppress IFN-β expression. Using reverse genetics, we generated a ToV-PLP knockout recombinant virus. Compared to the wild-type virus, the ToV-PLP knockout mutant virus showed impaired growth and induced higher expression levels of innate immune genes in infected cells. These results suggest that ToV-PLP functions as an innate immune antagonist; enterovirus G may therefore gain fitness through the acquisition of ToV-PLP from a recombination event. IMPORTANCE Enteroviruses comprise a highly diversified group of viruses. Genetic recombination has been considered a driving force for viral evolution; however, recombination between viruses from two different orders is a rare event. In this

Gene repair of an Usher syndrome causing mutation by zinc-finger nuclease mediated homologous recombination.

PubMed

Overlack, Nora; Goldmann, Tobias; Wolfrum, Uwe; Nagel-Wolfrum, Kerstin

2012-06-26

Human Usher syndrome (USH) is the most frequent cause of inherited deaf-blindness. It is clinically and genetically heterogeneous, assigned to three clinical types of which the most severe type is USH1. No effective treatment for the ophthalmic component of USH exists. Gene augmentation is an attractive strategy for hereditary retinal diseases. However, several USH genes, like USH1C, are expressed in various isoforms, hampering gene augmentation. As an alternative treatment strategy, we applied the zinc-finger nuclease (ZFN) technology for targeted gene repair of an USH1C, causing mutation by homologous recombination. We designed ZFNs customized for the p.R31X nonsense mutation in Ush1c. We evaluated ZFNs for DNA cleavage capability and analyzed ZFNs biocompatibilities by XTT assays. We demonstrated ZFNs mediated gene repair on genomic level by digestion assays and DNA sequencing, and on protein level by indirect immunofluorescence and Western blot analyses. The specifically designed ZFNs did not show cytotoxic effects in a p.R31X cell line. We demonstrated that ZFN induced cleavage of their target sequence. We showed that simultaneous application of ZFN and rescue DNA induced gene repair of the disease-causing mutation on the genomic level, resulting in recovery of protein expression. In our present study, we analyzed for the first time ZFN-activated gene repair of an USH gene. The data highlight the ability of ZFNs to induce targeted homologous recombination and mediate gene repair in USH. We provide further evidence that the ZFN technology holds great potential to recover disease-causing mutations in inherited retinal disorders.

Prioritization of orphan disease-causing genes using topological feature and GO similarity between proteins in interaction networks.

PubMed

Li, Min; Li, Qi; Ganegoda, Gamage Upeksha; Wang, JianXin; Wu, FangXiang; Pan, Yi

2014-11-01

Identification of disease-causing genes among a large number of candidates is a fundamental challenge in human disease studies. However, it is still time-consuming and laborious to determine the real disease-causing genes by biological experiments. With the advances of the high-throughput techniques, a large number of protein-protein interactions have been produced. Therefore, to address this issue, several methods based on protein interaction network have been proposed. In this paper, we propose a shortest path-based algorithm, named SPranker, to prioritize disease-causing genes in protein interaction networks. Considering the fact that diseases with similar phenotypes are generally caused by functionally related genes, we further propose an improved algorithm SPGOranker by integrating the semantic similarity of GO annotations. SPGOranker not only considers the topological similarity between protein pairs in a protein interaction network but also takes their functional similarity into account. The proposed algorithms SPranker and SPGOranker were applied to 1598 known orphan disease-causing genes from 172 orphan diseases and compared with three state-of-the-art approaches, ICN, VS and RWR. The experimental results show that SPranker and SPGOranker outperform ICN, VS, and RWR for the prioritization of orphan disease-causing genes. Importantly, for the case study of severe combined immunodeficiency, SPranker and SPGOranker predict several novel causal genes.

[From gene to disease; primary hyperoxaluria type I caused by mutations in the AGXT gene].

PubMed

van Woerden, C S; Groothof, J W; Wanders, R J A; Waterham, H R; Wijburg, F R

2006-07-29

Primary hyperoxaluria type I (PH1) is a congenital defect in glyoxylate metabolism caused by a deficiency in the liver-specific peroxisomal enzyme known as alanine glyoxylate aminotransferase (AGT). The deficiency is due to mutations in the AGXT gene, located on chromosome 2q37.3, and results in the conversion of glyoxylate to oxalate. The crystallisation of oxalate with calcium results in symptoms varying from a solitary kidney stone to end-stage renal disease with systemic oxalosis. The diagnosis is based on increased oxalate and glycolate excretion in the urine, reduced AGT activity in liver tissue, and confirmed mutations in the AGXT gene. Over 50 disease-causing mutations have been identified in PH1, which are associated with a wide range of effects on the AGT enzyme. Homozygous Gly170Arg or Phei52Ile mutations are associated with a reduction in urinary oxalate excretion upon pyridoxine administration and long-term preservation of renal function when treatment is initiated in a timely manner. Homozygous 33insC and Gly82Arg mutations result in a much poorer prognosis. Mutational analysis of the AGXT gene in PH1 patients can be a useful tool for establishing the diagnosis and choosing an appropriate therapeutic strategy.

Differential gene expression patterns between smokers and non‐smokers: cause or consequence?

PubMed Central

Jansen, Rick; Brooks, Andy; Willemsen, Gonneke; van Grootheest, Gerard; de Geus, Eco; Smit, Jan H.; Penninx, Brenda W.; Boomsma, Dorret I.

2015-01-01

Abstract The molecular mechanisms causing smoking‐induced health decline are largely unknown. To elucidate the molecular pathways involved in cause and consequences of smoking behavior, we conducted a genome‐wide gene expression study in peripheral blood samples targeting 18 238 genes. Data of 743 smokers, 1686 never smokers and 890 ex‐smokers were available from two population‐based cohorts from the Netherlands. In addition, data of 56 monozygotic twin pairs discordant for ever smoking were used. One hundred thirty‐two genes were differentially expressed between current smokers and never smokers (P < 1.2 × 10−6, Bonferroni correction). The most significant genes were G protein‐coupled receptor 15 (P < 1 × 10−150) and leucine‐rich repeat neuronal 3 (P < 1 × 10−44). The smoking‐related genes were enriched for immune system, blood coagulation, natural killer cell and cancer pathways. By taking the data of ex‐smokers into account, expression of these 132 genes was classified into reversible (94 genes), slowly reversible (31 genes), irreversible (6 genes) or inconclusive (1 gene). Expression of 6 of the 132 genes (three reversible and three slowly reversible) was confirmed to be reactive to smoking as they were differentially expressed in monozygotic pairs discordant for smoking. Cis‐expression quantitative trait loci for GPR56 and RARRES3 (downregulated in smokers) were associated with increased number of cigarettes smoked per day in a large genome‐wide association meta‐analysis, suggesting a causative effect of GPR56 and RARRES3 expression on smoking behavior. In conclusion, differential gene expression patterns in smokers are extensive and cluster in several underlying disease pathways. Gene expression differences seem mainly direct consequences of smoking, and largely reversible after smoking cessation. However, we also identified DNA variants that may influence smoking behavior via the mediating gene

Characterization of Wnt and Notch-Responsive Lgr5+ Hair Cell Progenitors in the Striolar Region of the Neonatal Mouse Utricle

PubMed Central

You, Dan; Guo, Luo; Li, Wenyan; Sun, Shan; Chen, Yan; Chai, Renjie; Li, Huawei

2018-01-01

Dysfunctions in hearing and balance are largely connected with hair cell (HC) loss. Although regeneration of HCs in the adult cochlea does not occur, there is still limited capacity for HC regeneration in the mammalian utricle from a distinct population of supporting cells (SCs). In response to HC damage, these Lgr5+ SCs, especially those in the striolar region, can regenerate HCs. In this study, we isolated Lgr5+ SCs and Plp1+ SCs (which originate from the striolar and extrastriolar regions, respectively) from transgenic mice by flow cytometry so as to compare the properties of these two subsets of SCs. We found that the Lgr5+ progenitors had greater proliferation and HC regeneration ability than the Plp1+ SCs and that the Lgr5+ progenitors responded more strongly to Wnt and Notch signaling than Plp1+ SCs. We then compared the gene expression profiles of the two populations by RNA-Seq and identified several genes that were significantly differentially expressed between the two populations, including genes involved in the cell cycle, transcription and cell signaling pathways. Targeting these genes and pathways might be a potential way to activate HC regeneration. PMID:29760650

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.

Retinoschisislike alterations in the mouse eye caused by gene targeting of the Norrie disease gene.

PubMed

Ruether, K; van de Pol, D; Jaissle, G; Berger, W; Tornow, R P; Zrenner, E

1997-03-01

To investigate the retinal function and morphology of mice carrying a replacement mutation in exon 2 of the Norrie disease gene. Recently, Norrie disease mutant mice have been generated using gene targeting technology. The mutation removes the 56 N-terminal amino acids of the Norrie gene product. Ganzfeld electroretinograms (ERGs) were obtained in five animals hemizygous or homozygous for the mutant gene and in three female animals heterozygous for the mutant gene. As controls, three males carrying the wild-type gene were examined. Electroretinogram testing included rod a- and b-wave V-log I functions, oscillatory potentials, and cone responses. The fundus morphology has been visualized by scanning laser ophthalmoscopy. Rod and cone ERG responses and fundus morphology were not significantly different among female heterozygotes and wild-type mice. In contrast, the hemizygous mice displayed a severe loss of ERG b-wave, leading to a negatively shaped scotopic ERG and a marked reduction of oscillatory potentials. The a-wave was normal at low intensities, and only with brighter flashes was there a moderate amplitude loss. Cone amplitudes were barely recordable in the gene-targeted males. Ophthalmoscopy revealed snowflakelike vitreal changes, retinoschisis, and pigment epithelium irregularities in hemizygotes and homozygotes, but no changes in female heterozygotes. The negatively shaped scotopic ERG in male mice with a Norrie disease gene mutation probably was caused by retinoschisis. Pigment epithelial changes and degenerations of the outer retina are relatively mild. These findings may be a clue to the embryonal retinoschisislike pathogenesis of Norrie disease in humans or it may indicate a different expression of the Norrie disease gene defect in mice compared to that in humans.

Distinct distal myopathy phenotype caused by VCP gene mutation in a Finnish family.

PubMed

Palmio, Johanna; Sandell, Satu; Suominen, Tiina; Penttilä, Sini; Raheem, Olayinka; Hackman, Peter; Huovinen, Sanna; Haapasalo, Hannu; Udd, Bjarne

2011-08-01

Inclusion body myopathy with Paget disease and frontotemporal dementia (IBMPFD) is caused by mutations in the valosin-containing protein (VCP) gene. We report a new distal phenotype caused by VCP gene mutation in a Finnish family with nine affected members in three generations. Patients had onset of distal leg muscle weakness and atrophy in the anterior compartment muscles after age 35, which caused a foot drop at age 50. None of the siblings had scapular winging, proximal myopathy, cardiomyopathy or respiratory problems during long-term follow-up. Three distal myopathy patients developed rapidly progressive dementia, became bedridden and died of cachexia and pneumonia and VCP gene mutation P137L (c.410C>T) was then identified in the family. Late onset autosomal dominant distal myopathy with rimmed vacuolar muscle pathology was not sufficient for exact diagnosis in this family until late-occurring dementia provided the clue for molecular diagnosis. VCP needs to be considered in the differential diagnostic work-up in patients with distal myopathy phenotype. Copyright © 2011 Elsevier B.V. All rights reserved.

pSW2, a Novel Low-Temperature-Inducible Gene Expression Vector Based on a Filamentous Phage of the Deep-Sea Bacterium Shewanella piezotolerans WP3.

PubMed

Yang, Xin-Wei; Jian, Hua-Hua; Wang, Feng-Ping

2015-08-15

A low-temperature-inducible protein expression vector (pSW2) based on a filamentous phage (SW1) of the deep-sea bacterium Shewanella piezotolerans WP3 was constructed. This vector replicated stably in Escherichia coli and Shewanella species, and its copy number increased at low temperatures. The pSW2 vector can be utilized as a complementation plasmid in WP3, and it can also be used for the production of complex cytochromes with multiple heme groups, which has the potential for application for metal ion recovery or bioremediation. Promoters of low-temperature-inducible genes in WP3 were fused into the vector to construct a series of vectors for enhancing protein expression at low temperature. The maximum green fluorescent protein intensity was obtained when the promoter for the hfq gene was used. The WP3/pSW2 system can efficiently produce a patatin-like protein (PLP) from a metagenomic library that tends to form inclusion bodies in E. coli. The yields of PLP in the soluble fraction were 8.3 mg/liter and 4.7 mg/liter of culture at 4°C and 20°C, respectively. Moreover, the pSW2 vector can be broadly utilized in other Shewanella species, such as S. oneidensis and S. psychrophila. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Comparative genomics of pyridoxal 5′-phosphate-dependent transcription factor regulons in Bacteria

PubMed Central

Suvorova, Inna A.

2016-01-01

The MocR-subfamily transcription factors (MocR-TFs) characterized by the GntR-family DNA-binding domain and aminotransferase-like sensory domain are broadly distributed among certain lineages of Bacteria. Characterized MocR-TFs bind pyridoxal 5′-phosphate (PLP) and control transcription of genes involved in PLP, gamma aminobutyric acid (GABA) and taurine metabolism via binding specific DNA operator sites. To identify putative target genes and DNA binding motifs of MocR-TFs, we performed comparative genomics analysis of over 250 bacterial genomes. The reconstructed regulons for 825 MocR-TFs comprise structural genes from over 200 protein families involved in diverse biological processes. Using the genome context and metabolic subsystem analysis we tentatively assigned functional roles for 38 out of 86 orthologous groups of studied regulators. Most of these MocR-TF regulons are involved in PLP metabolism, as well as utilization of GABA, taurine and ectoine. The remaining studied MocR-TF regulators presumably control genes encoding enzymes involved in reduction/oxidation processes, various transporters and PLP-dependent enzymes, for example aminotransferases. Predicted DNA binding motifs of MocR-TFs are generally similar in each orthologous group and are characterized by two to four repeated sequences. Identified motifs were classified according to their structures. Motifs with direct and/or inverted repeat symmetry constitute the majority of inferred DNA motifs, suggesting preferable TF dimerization in head-to-tail or head-to-head configuration. The obtained genomic collection of in silico reconstructed MocR-TF motifs and regulons in Bacteria provides a basis for future experimental characterization of molecular mechanisms for various regulators in this family. PMID:28348826

Novel homozygous missense mutation in ALDH7A1 causes neonatal pyridoxine dependent epilepsy.

PubMed

Coci, Emanuele G; Codutti, Luca; Fink, Christian; Bartsch, Sophie; Grüning, Gunnar; Lücke, Thomas; Kurth, Ingo; Riedel, Joachim

2017-04-01

Pyridoxine dependent epilepsy (PDE) (OMIM#266100) is a neonatal form of epilepsy, caused by dysfunction of the enzyme α-aminoadipic semialdehyde dehydrogenase (ALDH7A1 or Antiquitin). This enzyme converts α-aminoadipic semialdehyde (α-AASA) into α-aminoadipate (AAA), a critical step in the lysine metabolism of the brain. ALDH7A1 dysfunction causes an accumulation of α-AASA and δ 1 -piperideine-6-carboxylic acid (P6C), which are in equilibrium with each other. P6C binds and inactivates pyridoxal 5'-phosphate (PLP), the active form of pyridoxine. Individuals affected by ALDH7A1 deficiency show pre-natal and post-natal seizures, which respond to oral pyridoxine but not to other pediatric anti-epileptic drugs. We discovered a novel missense mutation (c.566G > A, p.Gly189Glu) in homozygous state residing in the NAD+ binding domain coding region of exon 6 and affecting an highly conserved amino acid residue. The seizures stopped under post-natal pyridoxine therapy, nevertheless a longer follow-up is needed to evaluate the intellectual development of the child, who is additionally treated with oral l-arginine since the 13th month of life. Developmental delay with or without structural cortex abnormalities were reported in several patients. A brain MRI scan revealed hyperintense white matter in the right cerebellum compatible with cerebellar gliosis. Taken together, our studies enlarge the group of missense pathogenic mutations of ALDH7A1 gene and reveal a novel cerebellar finding within the PDE patients cohort. Copyright © 2016 Elsevier Ltd. All rights reserved.

Mutation Update of ARSA and PSAP Genes Causing Metachromatic Leukodystrophy.

PubMed

Cesani, Martina; Lorioli, Laura; Grossi, Serena; Amico, Giulia; Fumagalli, Francesca; Spiga, Ivana; Filocamo, Mirella; Biffi, Alessandra

2016-01-01

Metachromatic leukodystrophy is a neurodegenerative disorder characterized by progressive demyelination. The disease is caused by variants in the ARSA gene, which codes for the lysosomal enzyme arylsulfatase A, or, more rarely, in the PSAP gene, which codes for the activator protein saposin B. In this Mutation Update, an extensive review of all the ARSA- and PSAP-causative variants published in the literature to date, accounting for a total of 200 ARSA and 10 PSAP allele types, is presented. The detailed ARSA and PSAP variant lists are freely available on the Leiden Online Variation Database (LOVD) platform at http://www.LOVD.nl/ARSA and http://www.LOVD.nl/PSAP, respectively. © 2015 WILEY PERIODICALS, INC.

Vitamin B6 Prevents IL-1β Protein Production by Inhibiting NLRP3 Inflammasome Activation.

PubMed

Zhang, Peipei; Tsuchiya, Kohsuke; Kinoshita, Takeshi; Kushiyama, Hiroko; Suidasari, Sofya; Hatakeyama, Mizuki; Imura, Hisanori; Kato, Norihisa; Suda, Takashi

2016-11-18

Vitamin B6 includes six water-soluble vitamers: pyridoxal (PL), pyridoxamine (PM), pyridoxine (PN), and their phosphorylated forms. Pyridoxal 5'-phosphate (PLP) is an important cofactor for many metabolic enzymes. Several lines of evidence demonstrate that blood levels of PLP are significantly lower in patients with inflammation than in control subjects and that vitamin B6 has anti-inflammatory effects, with therapeutic potential for a variety of inflammatory diseases. Although one of our group previously demonstrated that PL inhibits the NF-κB pathway, the molecular mechanism by which vitamin B6 suppresses inflammation is not well understood. Here, we showed that both PL and PLP suppressed the expression of cytokine genes in macrophages by inhibiting Toll-like receptor (TLR)-mediated TAK1 phosphorylation and the subsequent NF-κB and JNK activation. Furthermore, PL and PLP abolished NLRP3-dependent caspase-1 processing and the subsequent secretion of mature IL-1β and IL-18 in LPS-primed macrophages. In contrast, PM and PN had little effect on IL-1β production. PLP, but not PL, markedly reduced the production of mitochondrial reactive oxygen species (ROS) in peritoneal macrophages. Importantly, PL and PLP reduced IL-1β production induced by LPS and ATP, or by LPS alone, in mice. Moreover, PL and PLP protected mice from lethal endotoxic shock. Collectively, these findings reveal novel anti-inflammatory activities for vitamin B6 and suggest its potential for preventing inflammatory diseases driven by the NLRP3 inflammasome. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

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.

Prediction of cardioembolic, arterial and lacunar causes of cryptogenic stroke by gene expression and infarct location

PubMed Central

Jickling, Glen C; Stamova, Boryana; Ander, Bradley P; Zhan, Xinhua; Liu, Dazhi; Sison, Shara-Mae; Verro, Piero; Sharp, Frank R

2012-01-01

Background and Purpose The cause of ischemic stroke remains unclear, or cryptogenic, in as many as 35% of stroke patients. Not knowing the cause of stroke restricts optimal implementation of prevention therapy and limits stroke research. We demonstrate how gene expression profiles in blood can be used in conjunction with a measure of infarct location on neuroimaging to predict a probable cause in cryptogenic stroke. Methods The cause of cryptogenic stroke was predicted using previously described profiles of differentially expressed genes characteristic of patients with cardioembolic, arterial and lacunar stroke. RNA was isolated from peripheral blood of 131 cryptogenic strokes and compared to profiles derived from 149 strokes of known cause. Each sample was run on Affymetrix U133 Plus2.0 microarrays. Cause of cryptogenic stroke was predicted using gene expression in blood and infarct location. Results Cryptogenic strokes were predicted to be 58% cardioembolic, 18% arterial, 12% lacunar and 12% unclear etiology. Cryptogenic stroke of predicted cardioembolic etiology had more prior myocardial infarction and higher CHA2DS2-VASc scores compared to stroke of predicted arterial etiology. Predicted lacunar strokes had higher systolic and diastolic blood pressures and lower NIHSS compared to predicted arterial and cardioembolic strokes. Cryptogenic strokes of unclear predicted etiology were less likely to have a prior TIA or ischemic stroke. Conclusions Gene expression in conjunction with a measure of infarct location can predict a probable cause in cryptogenic strokes. Predicted groups require further evaluation to determine whether relevant clinical, imaging, or therapeutic differences exist for each group. PMID:22627989

Novel biallelic mutations in MSH6 and PMS2 genes: gene conversion as a likely cause of PMS2 gene inactivation.

PubMed

Auclair, Jessie; Leroux, Dominique; Desseigne, Françoise; Lasset, Christine; Saurin, Jean Christophe; Joly, Marie Odile; Pinson, Stéphane; Xu, Xiao Li; Montmain, Gilles; Ruano, Eric; Navarro, Claudine; Puisieux, Alain; Wang, Qing

2007-11-01

Since the first report by our group in 1999, more than 20 unrelated biallelic mutations in DNA mismatch repair genes (MMR) have been identified. In the present report, we describe two novel cases: one carrying compound heterozygous mutations in the MSH6 gene; and the other, compound heterozygous mutations in the PMS2 gene. Interestingly, the inactivation of one PMS2 allele was likely caused by gene conversion. Although gene conversion has been suggested to be a mutation mechanism underlying PMS2 inactivation, this is the first report of its involvement in a pathogenic mutation. The clinical features of biallelic mutation carriers were similar to other previously described patients, with the presence of café-au-lait spots (CALS), early onset of brain tumors, and colorectal neoplasia. Our data provide further evidence of the existence, although rare, of a distinct recessively inherited syndrome on the basis of MMR constitutional inactivation. The identification of this syndrome should be useful for genetic counseling, especially in families with atypical hereditary nonpolyposis colon cancer (HNPCC) associated with childhood cancers, and for the clinical surveillance of these mutation carriers. 2007 Wiley-Liss, Inc.

Folate-mediated one-carbon metabolism genes and interactions with nutritional factors on colorectal cancer risk: Women's Health Initiative Observational Study.

PubMed

Cheng, Ting-Yuan David; Makar, Karen W; Neuhouser, Marian L; Miller, Joshua W; Song, Xiaoling; Brown, Elissa C; Beresford, Shirley A A; Zheng, Yingye; Poole, Elizabeth M; Galbraith, Rachel L; Duggan, David J; Habermann, Nina; Bailey, Lynn B; Maneval, David R; Caudill, Marie A; Toriola, Adetunji T; Green, Ralph; Ulrich, Cornelia M

2015-10-15

Investigations of folate-mediated one-carbon metabolism (FOCM) genes and gene-nutrient interactions with respect to colorectal cancer (CRC) risk are limited to candidate polymorphisms and dietary folate. This study comprehensively investigated associations between genetic variants in FOCM and CRC risk and whether the FOCM nutrient status modified these associations. Two hundred eighty-eight candidate and tagging single-nucleotide polymorphisms (SNPs) in 30 FOCM genes were genotyped for 821 incident CRC case-control matched pairs in the Women's Health Initiative Observational Study cohort. FOCM biomarkers (red blood cell [RBC] folate, plasma folate, pyridoxal-5'-phosphate [PLP], vitamin B12, and homocysteine) and self-reported alcohol consumption were measured at the baseline. Conditional logistic regression was implemented; effect modification was examined on the basis of known enzyme-nutrient relations. Statistically significant associations were observed between CRC risk and functionally defined candidate SNPs of methylenetetrahydrofolate dehydrogenase 1 (MTHFD1; K134R), 5-methyltetrahydrofolate-homocysteine methyltransferase reductase (MTRR; P450R), and PR domain containing 2 with ZNF domain (PRDM2; S450N) and a literature candidate SNP of thymidylate synthase (TYMS; g.676789A>T; nominal P < .05). In addition, suggestive associations were noted for tagging SNPs in cystathionine-β-synthase (CBS), dihydrofolate reductase (DHFR), DNA (cytosine-5-)-methyltransferase 3β (DNMT3B), methionine adenosyltransferase I α (MAT1A), MTHFD1, and MTRR (nominal P < .05; adjusted P, not significant). Significant interactions between nutrient biomarkers and candidate polymorphisms were observed for 1) plasma/RBC folate and folate hydrolase 1 (FOLH1), paraoxonase 1 (PON1), transcobalamin II (TCN2), DNMT1, and DNMT3B; 2) plasma PLP and TYMS TS3; 3) plasma B12 and betaine-homocysteine S-methyltransferase 2 (BHMT2); and 4) homocysteine and methylenetetrahydrofolate reductase

Deletion of the SHOX gene in patients with short stature of unknown cause.

PubMed

Morizio, E; Stuppia, L; Gatta, V; Fantasia, D; Guanciali Franchi, P; Rinaldi, M M; Scarano, G; Concolino, D; Giannotti, A; Verrotti, A; Chiarelli, F; Calabrese, G; Palka, G

2003-06-15

A fluorescence in situ hybridization (FISH) study was performed in 56 patients with short stature of unknown cause in order to establish the role of deletion of the SHOX gene in this population. FISH analysis was carried out on metaphase spreads and interphase lymphocytes from blood smears using a probe specific for the SHOX gene. Deletion of SHOX was found in four patients (7.1%). No skeletal abnormalities were detected in these patients either at the physical examination or at X-rays of the upper and lower limbs. Present results indicate that SHOX plays an important role also in short stature of unknown cause, and FISH analysis appears as an easy, appropriate, and inexpensive method for the detection of SHOX deletion. Copyright 2003 Wiley-Liss, Inc.

Racemization of alanine by the alanine racemases from Salmonella typhimurium and Bacillus stearothermophilus: energetic reaction profiles

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

Faraci, W.S.; Walsh, C.T.

1988-05-03

Alanine racemases are bacterial pyridoxal 5'-phosphate (PLP) dependent enzymes providing D-alanine as an essential building block for biosynthesis of the peptidoglycan layer of the cell wall. Two isozymic alanine racemases, encoded by the dadB gene and the alr gene, from the Gram-negative mesophilic Salmonella typhimurium and one from the Gram-positive thermophilic Bacillus stearothermophilus have been examined for the racemization mechanism. Substrate deuterium isotope effects and solvent deuterium isotope effects have been measured in both L ..-->.. D and D..-->.. L directions for all three enzymes to assess the degree to which abstraction of the ..cap alpha..-proton or protonation of substratemore » PLP carbanion is limiting in catalysis. Additionally, experiments measuring internal return of ..cap alpha..-/sup 3/H from substrate to product and solvent exchange/substrate conversion experiments in /sup 3/H/sub 2/O have been used with each enzyme to examine the partitioning of substrate PLP carbanion intermediates and to obtain the relative heights of kinetically significant energy barriers in alanine racemase catalysis.« less

Saccharomyces cerevisiae Differential Functionalization of Presumed ScALT1 and ScALT2 Alanine Transaminases Has Been Driven by Diversification of Pyridoxal Phosphate Interactions

PubMed Central

Rojas-Ortega, Erendira; Aguirre-López, Beatriz; Reyes-Vivas, Horacio; González-Andrade, Martín; Campero-Basaldúa, Jose C.; Pardo, Juan P.; González, Alicia

2018-01-01

Saccharomyces cerevisiae arose from an interspecies hybridization (allopolyploidiza-tion), followed by Whole Genome Duplication. Diversification analysis of ScAlt1/ScAlt2 indicated that while ScAlt1 is an alanine transaminase, ScAlt2 lost this activity, constituting an example in which one of the members of the gene pair lacks the apparent ancestral physiological role. This paper analyzes structural organization and pyridoxal phosphate (PLP) binding properties of ScAlt1 and ScAlt2 indicating functional diversification could have determined loss of ScAlt2 alanine transaminase activity and thus its role in alanine metabolism. It was found that ScAlt1 and ScAlt2 are dimeric enzymes harboring 67% identity and intact conservation of the catalytic residues, with very similar structures. However, tertiary structure analysis indicated that ScAlt2 has a more open conformation than that of ScAlt1 so that under physiological conditions, while PLP interaction with ScAlt1 allows the formation of two tautomeric PLP isomers (enolimine and ketoenamine) ScAlt2 preferentially forms the ketoenamine PLP tautomer, indicating a modified polarity of the active sites which affect the interaction of PLP with these proteins, that could result in lack of alanine transaminase activity in ScAlt2. The fact that ScAlt2 forms a catalytically active Schiff base with PLP and its position in an independent clade in “sensu strictu” yeasts suggests this protein has a yet undiscovered physiological function. PMID:29867852

Mutations in PRPF31 Inhibit Pre-mRNA Splicing of Rhodopsin Gene and Cause Apoptosis of Retinal Cells

PubMed Central

Yuan, Liya; Kawada, Mariko; Havlioglu, Necat; Tang, Hao; Wu, Jane Y.

2007-01-01

Mutations in human PRPF31 gene have been identified in patients with autosomal dominant retinitis pigmentosa (adRP). To begin to understand mechanisms by which defects in this general splicing factor cause retinal degeneration, we examined the relationship between PRPF31 and pre-mRNA splicing of photoreceptor-specific genes. We used a specific anti-PRPF31 antibody to immunoprecipitate splicing complexes from retinal cells and identified the transcript of rhodopsin gene (RHO) among RNA species associated with PRPF31-containing complexes. Mutant PRPF31 proteins significantly inhibited pre-mRNA splicing of intron 3 in RHO gene. In primary retinal cell cultures, expression of the mutant PRPF31 proteins reduced rhodopsin expression and caused apoptosis of rhodopsin-positive retinal cells. This primary retinal culture assay provides an in vitro model to study photoreceptor cell death caused by PRPF31 mutations. Our results demonstrate that mutations in PRPF31 gene affect RHO pre-mRNA splicing and reveal a link between PRPF31 and RHO, two major adRP genes. PMID:15659613

Absence of IFNγ Increases Brain Pathology in EAE-susceptible DRB1*0301.DQ8 HLA Transgenic Mice Through Secretion of Pro-inflammatory Cytokine IL-17 and Induction of Pathogenic Monocytes/Microglia into the CNS

PubMed Central

Mangalam, Ashutosh; Luo, Ningling; Luckey, David; Papke, Louisa; Hubbard, Alyssa; Wussow, Arika; Smart, Michele; Giri, Shailendra; Rodriguez, Moses; David, Chella

2014-01-01

Multiple sclerosis (MS) is an inflammatory, demyelinating disease of the central nervous system (CNS) of presumed autoimmune origin. Of all the genetic factors linked with MS, MHC class-II molecules have the strongest association. Generation of HLA class-II transgenic mice has helped to elucidate the role of HLA class-II genes in chronic inflammatory and demyelinating diseases. We have shown that the human HLA-DRB1*0301 gene predisposes to proteolipid protein (PLP)-induced EAE, whereas HLA-DQβ1*0601 (DQ6) was resistant. We also showed that the DQ6 molecule protects from EAE in DRB1*0301.DQ6 double transgenic mice by producing anti-inflammatory interferon gamma (IFNγ). HLA-DQβ1*0302 (DQ8) transgenic mice were also resistant to PLP91-110-induced EAE, but production of pro-inflammatory IL-17 exacerbated disease in DRB1*0301.DQ8 mice. To further confirm the role of IFNγ in protection, we generated DRB1*0301.DQ8 mice lacking IFNγ (DRB1*0301.DQ8.IFNγ−/−). Immunization with PLP91-110 peptide caused atypical EAE in DRB1*0301.DQ8.IFNγ−/− mice characterized by ataxia, spasticity and dystonia, hallmarks of brain-specific disease. Severe brain specific inflammation and demyelination in DRB1*0301.DQ8.IFNγ−/− mice with minimal spinal cord pathology further confirmed brain-specific pathology. Atypical EAE in DRB1*0301.DQ8.IFNγ−/− mice was associated with increased encephalitogenicity of CD4 T cells and their ability to produce higher levels of IL-17 and GM-CSF compared to DRB1*0301.DQ8 mice. Further, areas with demyelination showed increased presence of CD68+ inflammatory cells, suggesting an important role for monocytes/microglia in causing brain pathology. Thus, our study supports a protective role for IFNγ in the demyelination of brain through down regulation of IL-17/GM-CSF and induction of neuro-protective factors in the brain by monocytes/microglial cells. PMID:25339670

Direct evidence that an extended hydrogen-bonding network influences activation of pyridoxal 5'-phosphate in aspartate aminotransferase

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

Dajnowicz, Steven; Parks, Jerry M.; Hu, Xiche

We used pyridoxal 5'-phosphate (PLP) is a fundamental, multifunctional enzyme cofactor to catalyze a wide variety of chemical reactions involved in amino acid metabolism. PLP-dependent enzymes optimize specific chemical reactions by modulating the electronic states of PLP through distinct active site environments. In aspartate aminotransferase (AAT), an extended hydrogen bond network is coupled to the pyridinyl nitrogen of the PLP, influencing the electrophilicity of the cofactor. This network, which involves residues Asp-222, His-143, Thr-139, His-189, and structural waters, is located at the edge of PLP opposite the reactive Schiff base. We demonstrate that this hydrogen bond network directly influences themore » protonation state of the pyridine nitrogen of PLP, which affects the rates of catalysis. We analyzed perturbations caused by single- and double-mutant variants using steady-state kinetics, high resolution X-ray crystallography, and quantum chemical calculations. Protonation of the pyridinyl nitrogen to form a pyridinium cation induces electronic delocalization in the PLP, which correlates with the enhancement in catalytic rate in AAT. Therefore, PLP activation is controlled by the proximity of the pyridinyl nitrogen to the hydrogen bond microenvironment. Quantum chemical calculations indicate that Asp-222, which is directly coupled to the pyridinyl nitrogen, increases the pKa of the pyridine nitrogen and stabilizes the pyridinium cation. His-143 and His-189 also increase the pKa of the pyridine nitrogen but, more significantly, influence the position of the proton that resides between Asp-222 and the pyridinyl nitrogen. Our findings indicate that the second shell residues directly enhance the rate of catalysis in AAT.« less

Direct evidence that an extended hydrogen-bonding network influences activation of pyridoxal 5'-phosphate in aspartate aminotransferase

DOE PAGES

Dajnowicz, Steven; Parks, Jerry M.; Hu, Xiche; ...

2017-02-23

We used pyridoxal 5'-phosphate (PLP) is a fundamental, multifunctional enzyme cofactor to catalyze a wide variety of chemical reactions involved in amino acid metabolism. PLP-dependent enzymes optimize specific chemical reactions by modulating the electronic states of PLP through distinct active site environments. In aspartate aminotransferase (AAT), an extended hydrogen bond network is coupled to the pyridinyl nitrogen of the PLP, influencing the electrophilicity of the cofactor. This network, which involves residues Asp-222, His-143, Thr-139, His-189, and structural waters, is located at the edge of PLP opposite the reactive Schiff base. We demonstrate that this hydrogen bond network directly influences themore » protonation state of the pyridine nitrogen of PLP, which affects the rates of catalysis. We analyzed perturbations caused by single- and double-mutant variants using steady-state kinetics, high resolution X-ray crystallography, and quantum chemical calculations. Protonation of the pyridinyl nitrogen to form a pyridinium cation induces electronic delocalization in the PLP, which correlates with the enhancement in catalytic rate in AAT. Therefore, PLP activation is controlled by the proximity of the pyridinyl nitrogen to the hydrogen bond microenvironment. Quantum chemical calculations indicate that Asp-222, which is directly coupled to the pyridinyl nitrogen, increases the pKa of the pyridine nitrogen and stabilizes the pyridinium cation. His-143 and His-189 also increase the pKa of the pyridine nitrogen but, more significantly, influence the position of the proton that resides between Asp-222 and the pyridinyl nitrogen. Our findings indicate that the second shell residues directly enhance the rate of catalysis in AAT.« less

De novo mutations in genes of mediator complex causing syndromic intellectual disability: mediatorpathy or transcriptomopathy?

PubMed

Caro-Llopis, Alfonso; Rosello, Monica; Orellana, Carmen; Oltra, Silvestre; Monfort, Sandra; Mayo, Sonia; Martinez, Francisco

2016-12-01

Mutations in the X-linked gene MED12 cause at least three different, but closely related, entities of syndromic intellectual disability. Recently, a new syndrome caused by MED13L deleterious variants has been described, which shows similar clinical manifestations including intellectual disability, hypotonia, and other congenital anomalies. Genotyping of 1,256 genes related with neurodevelopment was performed by next-generation sequencing in three unrelated patients and their healthy parents. Clinically relevant findings were confirmed by conventional sequencing. Each patient showed one de novo variant not previously reported in the literature or databases. Two different missense variants were found in the MED12 or MED13L genes and one nonsense mutation was found in the MED13L gene. The phenotypic consequences of these mutations are closely related and/or have been previously reported in one or other gene. Additionally, MED12 and MED13L code for two closely related partners of the mediator kinase module. Consequently, we propose the concept of a common MED12/MED13L clinical spectrum, encompassing Opitz-Kaveggia syndrome, Lujan-Fryns syndrome, Ohdo syndrome, MED13L haploinsufficiency syndrome, and others.

Apolipoprotein gene polymorphisms as cause of cholesterol QTLs in mice.

PubMed

Suto, Jun-ichi

2005-06-01

Quantitative trait locus (QTL) analyses of plasma cholesterol levels were carried out in three sets of F(2) mice that were formed in a 'round-robin' manner from C57BL/6J, KK (-A(y)), and RR strains. Six QTLs were identified on chromosomes 1 (Cq1, Cq2, and Cq6), 3 (Cq3), and 9 (Cq4 and Cq5); of these, Cq2 colocalized with Cq6, and Cq4 colocalized with Cq5. The major candidate gene for Cq2 and Cq6 is Apoa2, and that for Cq4 and Cq5 is Apoa4. The adequacy of polymorphisms in candidate genes as cause of QTLs was investigated in this study. For Apoa2, three different alleles (Apoa2(a), Apoa2(b), and Apoa2(c)) are known. Since there was no significant physiologic difference between Apoa2(a) and Apoa2(c) alleles, previous hypothesis that Apoa2(b) was different from Apoa2(a) and Apoa2(c) in the ability to increase cholesterol levels was further supported. Presumably, G-to-A substitution at nucleotide 84 and/or C-to-T substitution at nucleotide 182 are crucial to make the Apoa2(b) unique. On the other hand, for Apoa4, the most striking polymorphism was the number of Glu-Gln-Ala/Val-Gln repeats in carboxyl end; however, this might not be responsible for QTLs. Instead, a silent mutation, C-to-T substitution at nucleotide 771, was shown to be completely correlated with the occurrence of QTLs in a total of six F(2) intercrosses. Provisionally, but reasonably, these base substitutions are qualified as primary causes that constitute QTL effect. The potential strategy for identifying genes and base substitutions underlying QTLs is discussed.

[Short stature caused by SHOX gene haploinsufficiency: from diagnosis to treatment].

PubMed

Jorge, Alexander A L; Nishi, Mirian Y; Funari, Mariana F A; Souza, Silvia C; Arnhold, Ivo J P; Mendonça, Berenice B

2008-07-01

Studies involving patients with short stature and partial deletion of sex chromosomes identified SHOX gene in the pseudoautosomal region of the X and Y chromosomes. SHOX haploinsufficiency is an important cause of short stature in a diversity of clinical conditions. It explains 2/3 of short stature observed in Turner syndrome (TS) patients. Heterozygous mutations in SHOX are observed in 77% of patients with Leri-Weill dyschondrosteosis, a common dominant inherited skeletal dysplasia and in 3% of children with idiopathic short stature, indicating that SHOX defects are the most frequent monogenetic cause of short stature. The sitting height/height ratio (SH/H) standard deviation score is a simple way to assess body proportions and together with a careful exam of other family members, effectively selected a group of patients that presented a high frequency of SHOX mutations. Growth hormone treatment of short stature due to TS is well established and considering the common etiology of short stature in patients with isolated defects of SHOX gene, this treatment is also proposed for these patients. Here, we review clinical, molecular and therapeutic aspects of SHOX haploinsufficiency.

Mutations in the ABCA4 (ABCR) gene are the major cause of autosomal recessive cone-rod dystrophy.

PubMed

Maugeri, A; Klevering, B J; Rohrschneider, K; Blankenagel, A; Brunner, H G; Deutman, A F; Hoyng, C B; Cremers, F P

2000-10-01

The photoreceptor cell-specific ATP-binding cassette transporter gene (ABCA4; previously denoted "ABCR") is mutated, in most patients, with autosomal recessive (AR) Stargardt disease (STGD1) or fundus flavimaculatus (FFM). In addition, a few cases with AR retinitis pigmentosa (RP) and AR cone-rod dystrophy (CRD) have been found to have ABCA4 mutations. To evaluate the importance of the ABCA4 gene as a cause of AR CRD, we selected 5 patients with AR CRD and 15 patients from Germany and The Netherlands with isolated CRD. Single-strand conformation-polymorphism analysis and sequencing revealed 19 ABCA4 mutations in 13 (65%) of 20 patients. In six patients, mutations were identified in both ABCA4 alleles; in seven patients, mutations were detected in one allele. One complex ABCA4 allele (L541P;A1038V) was found exclusively in German patients with CRD; one patient carried this complex allele homozygously, and five others were compound heterozygous. These findings suggest that mutations in the ABCA4 gene are the major cause of AR CRD. A primary role of the ABCA4 gene in STGD1/FFM and AR CRD, together with the gene's involvement in an as-yet-unknown proportion of cases with AR RP, strengthens the idea that mutations in the ABCA4 gene could be the most frequent cause of inherited retinal dystrophy in humans.

HlyU Is a Positive Regulator of Hemolysin Expression in Vibrio anguillarum ▿

PubMed Central

Li, Ling; Mou, Xiangyu; Nelson, David R.

2011-01-01

The two hemolysin gene clusters previously identified in Vibrio anguillarum, the vah1 cluster and the rtxACHBDE cluster, are responsible for the hemolytic and cytotoxic activities of V. anguillarum in fish. In this study, we used degenerate PCR to identify a positive hemolysin regulatory gene, hlyU, from the unsequenced V. anguillarum genome. The hlyU gene of V. anguillarum encodes a 92-amino-acid protein and is highly homologous to other bacterial HlyU proteins. An hlyU mutant was constructed, which exhibited an ∼5-fold decrease in hemolytic activity on sheep blood agar with no statistically significant decrease in cytotoxicity of the wild-type strain. Complementation of the hlyU mutation restored both hemolytic activity and cytotoxic activity. Both semiquantitative reverse transcription-PCR (RT-PCR) and quantitative real-time RT-PCR (qRT-PCR) were used to examine expression of the hemolysin genes under exponential and stationary-phase conditions in wild-type, hlyU mutant, and hlyU complemented strains. Compared to the wild-type strain, expression of rtx genes decreased in the hlyU mutant, while expression of vah1 and plp was not affected in the hlyU mutant. Complementation of the hlyU mutation restored expression of the rtx genes and increased vah1 and plp expression to levels higher than those in the wild type. The transcriptional start sites in both the vah1-plp and rtxH-rtxB genes' intergenic regions were determined using 5′ random amplification of cDNA ends (5′-RACE), and the binding sites for purified HlyU were discovered using DNA gel mobility shift experiments and DNase protection assays. PMID:21764937

Identification of HIBCH gene mutations causing autosomal recessive Leigh syndrome: a gene involved in valine metabolism.

PubMed

Soler-Alfonso, Claudia; Enns, Gregory M; Koenig, Mary Kay; Saavedra, Heather; Bonfante-Mejia, Eliana; Northrup, Hope

2015-03-01

Leigh syndrome is a progressive neurodegenerative disorder with usual onset of symptoms during the first year of life. The disorder has been associated with mutations in over 30 genes. This difficulty with genetic heterogeneity makes whole exome sequencing a more cost-effective approach for investigation of etiology. We describe an individual with typical Leigh syndrome who was found to have compound heterozygous mutations in the gene HIBCH (3-hydroxyisobutyryl coenzyme A hydrolase), an enzyme involved in the catabolism of valine. She exhibited significant clinical improvement after a valine-restricted diet. A subset of patients with uncharacterized Leigh syndrome present with specific biochemical abnormalities. This report highpoints the challenges and restrictions of routine metabolic testing and features the recognition of inborn errors of metabolism as potential treatable causes of Leigh syndrome. Copyright © 2015 Elsevier Inc. All rights reserved.

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.

Schizophrenia: A Pathogenetic Autoimmune Disease Caused by Viruses and Pathogens and Dependent on Genes

PubMed Central

Carter, C. J.

2011-01-01

Many genes have been implicated in schizophrenia as have viral prenatal or adult infections and toxoplasmosis or Lyme disease. Several autoantigens also target key pathology-related proteins. These factors are interrelated. Susceptibility genes encode for proteins homologous to those of the pathogens while the autoantigens are homologous to pathogens' proteins, suggesting that the risk-promoting effects of genes and risk factors are conditional upon each other, and dependent upon protein matching between pathogen and susceptibility gene products. Pathogens' proteins may act as dummy ligands, decoy receptors, or via interactome interference. Many such proteins are immunogenic suggesting that antibody mediated knockdown of multiple schizophrenia gene products could contribute to the disease, explaining the immune activation in the brain and lymphocytes in schizophrenia, and the preponderance of immune-related gene variants in the schizophrenia genome. Schizophrenia may thus be a “pathogenetic” autoimmune disorder, caused by pathogens, genes, and the immune system acting together, and perhaps preventable by pathogen elimination, or curable by the removal of culpable antibodies and antigens. PMID:22567321

A novel synonymous variant in the AVP gene associated with adFNDI causes partial RNA missplicing.

PubMed

Kvistgaard, Helene; Christensen, Jane H; Johansson, Jan-Ove; Gregersen, Niels; Rittig, Charlotte; Rittig, Soeren; Corydon, Thomas Juhl

2018-06-27

Objective: Autosomal dominant familial neurohypophyseal diabetes insipidus (adFNDI) is characterized by severe polyuria and polydipsia and is caused by variations in the gene encoding the AVP prohormone. The study aimed to ascertain a correct diagnosis, to identify the underlying genetic cause of adFNDI in a Swedish kindred, and to test the hypothesis that the identified synonymous exonic variant in the AVP gene (c.324G>A), causes missplicing, and endoplasmic reticulum (ER) retention of the prohormone. Three affected family members were admitted for fluid deprivation test and dDAVP challenge test. Direct sequencing of the AVP gene was performed in affected subjects, and genotyping of the identified variant was performed in family members. The variant was examined by expression of AVP minigenes containing the entire coding regions as well as intron 2 of AVP. Clinical tests revealed significant phenotypical variation with both complete and partial adFNDI phenotype. DNA analysis revealed a synonymous c.324G>A substitution in one allele of the AVP gene in affected family members only. Cellular studies revealed both normally spliced and misspliced pre-mRNA in cells transfected with the AVP c.324G>A minigene. Confocal laser scanning microscopy showed collective localization of the variant prohormone to ER and vesicular structures at the tip of cellular processes. We have identified a synonymous variant affecting the second nucleotide of exon 3 in the AVP gene (c.324G>A) in a kindred in which adFNDI segregates. Notably, we showed that this variant causes partial missplicing of pre-mRNA resulting in accumulation of variant prohormone in ER. Our study suggests that even a small amount of aberrant mRNA might be sufficient to disturb cellular function resulting in adFNDI.
. ©2018S. Karger AG, Basel.

Mutations in the ABCA4 (ABCR) Gene Are the Major Cause of Autosomal Recessive Cone-Rod Dystrophy

PubMed Central

Maugeri, Alessandra; Klevering, B. Jeroen; Rohrschneider, Klaus; Blankenagel, Anita; Brunner, Han G.; Deutman, August F.; Hoyng, Carel B.; Cremers, Frans P. M.

2000-01-01

The photoreceptor cell–specific ATP-binding cassette transporter gene (ABCA4; previously denoted “ABCR”) is mutated in most patients with autosomal recessive (AR) Stargardt disease (STGD1) or fundus flavimaculatus (FFM). In addition, a few cases with AR retinitis pigmentosa (RP) and AR cone-rod dystrophy (CRD) have been found to have ABCA4 mutations. To evaluate the importance of the ABCA4 gene as a cause of AR CRD, we selected 5 patients with AR CRD and 15 patients with isolated CRD, all from Germany and The Netherlands . Single-strand conformation–polymorphism analysis and sequencing revealed 19 ABCA4 mutations in 13 (65%) of 20 patients. In six patients, mutations were identified in both ABCA4 alleles; in seven patients, mutations were detected in one allele. One complex ABCA4 allele (L541P;A1038V) was found exclusively in German patients with CRD; one patient carried this complex allele homozygously, and five others were compound heterozygous. These findings suggest that mutations in the ABCA4 gene are the major cause of AR CRD. A primary role of the ABCA4 gene in STGD1/FFM and AR CRD, together with the gene's involvement in an as-yet-unknown proportion of cases with AR RP, strengthens the idea that mutations in the ABCA4 gene could be the most frequent cause of inherited retinal dystrophy in humans. PMID:10958761

Autosomal recessive retinitis pigmentosa caused by mutations in the MAK gene.

PubMed

Stone, Edwin M; Luo, Xunda; Héon, Elise; Lam, Byron L; Weleber, Richard G; Halder, Jennifer A; Affatigato, Louisa M; Goldberg, Jacqueline B; Sumaroka, Alexander; Schwartz, Sharon B; Cideciyan, Artur V; Jacobson, Samuel G

2011-12-28

To determine the disease expression in autosomal recessive (ar) retinitis pigmentosa (RP) caused by mutations in the MAK (male germ cell-associated kinase) gene. Patients with RP and MAK gene mutations (n = 24; age, 32-77 years at first visit) were studied by ocular examination, perimetry, and optical coherence tomography (OCT). All but one MAK patient were homozygous for an identical truncating mutation in exon 9 and had Ashkenazi Jewish heritage. The carrier frequency of this mutation among 1207 unrelated Ashkenazi control subjects was 1 in 55, making it the most common cause of heritable retinal disease in this population and MAK-associated RP the sixth most common Mendelian disease overall in this group. Visual acuities could be normal into the eighth decade of life. Kinetic fields showed early loss in the superior-temporal quadrant. With more advanced disease, superior and midperipheral function was lost, but the nasal field remained. Only a central island was present at late stages. Pigmentary retinopathy was less prominent in the superior nasal quadrant. Rod-mediated vision was abnormal but detectable in the residual field; all patients had rod>cone dysfunction. Photoreceptor layer thickness was normal centrally but decreased with eccentricity. At the stages studied, there was no evidence of photoreceptor ciliary elongation. The patterns of disease expression in the MAK form of arRP showed some resemblance to patterns described in autosomal dominant RP, especially the form caused by RP1 mutations. The similarity in phenotypes is of interest, considering that there is experimental evidence of interaction between Mak and RP1 in the photoreceptor cilium.

[From gene to disease; genetic causes of hearing loss and visual impairment sometimes accompanied by vestibular problems (Usher syndrome)].

PubMed

Pennings, R J E; Kremer, H; Deutman, A F; Kimberling, W J; Cremers, C W R J

2002-12-07

Usher syndrome is an autosomal recessively inherited disease, characterised by sensorineural hearing loss, tapetoretinal degeneration and in some cases vestibular problems. Based on the clinical heterogeneity, the disease can be classified into three clinical types (I, II and III), which have their own genetic subtypes (Usher 1A-Usher IG, Usher 2A-Usher 2C and Usher 3). The majority of the Usher type I cases are caused by mutations in the MYO7A gene (Usher 1B) while mutations in the USH2A gene (Usher 2A) are the cause of most cases of type II. Usher syndrome type III, caused by mutations in the USH3 gene, is frequently seen only in Finland.

Contiguous gene deletion of chromosome 2p16.3-p21 as a cause of Lynch syndrome.

PubMed

Salo-Mullen, Erin E; Lynn, Patricio B; Wang, Lu; Walsh, Michael; Gopalan, Anuradha; Shia, Jinru; Tran, Christina; Man, Fung Ying; McBride, Sean; Schattner, Mark; Zhang, Liying; Weiser, Martin R; Stadler, Zsofia K

2018-01-01

Lynch syndrome is an autosomal dominant condition caused by pathogenic mutations in the DNA mismatch repair (MMR) genes. Although commonly associated with clinical features such as intellectual disability and congenital anomalies, contiguous gene deletions may also result in cancer predisposition syndromes. We report on a 52-year-old male with Lynch syndrome caused by deletion of chromosome 2p16.3-p21. The patient had intellectual disability and presented with a prostatic adenocarcinoma with an incidentally identified synchronous sigmoid adenocarcinoma that exhibited deficient MMR with an absence of MSH2 and MSH6 protein expression. Family history was unrevealing. Physical exam revealed short stature, brachycephaly with a narrow forehead and short philtrum, brachydactyly of the hands, palmar transverse crease, broad and small feet with hyperpigmentation of the soles. The patient underwent total colectomy with ileorectal anastomosis for a pT3N1 sigmoid adenocarcinoma. Germline genetic testing of the MSH2, MSH6, and EPCAM genes revealed full gene deletions. SNP-array based DNA copy number analysis identified a deletion of 4.8 Mb at 2p16.3-p21. In addition to the three Lynch syndrome associated genes, the deleted chromosomal section encompassed genes including NRXN1, CRIPT, CALM2, FBXO11, LHCGR, MCFD2, TTC7A, EPAS1, PRKCE, and 15 others. Contiguous gene deletions have been described in other inherited cancer predisposition syndromes, such as Familial Adenomatous Polyposis. Our report and review of the literature suggests that contiguous gene deletion within the 2p16-p21 chromosomal region is a rare cause of Lynch syndrome, but presents with distinct phenotypic features, highlighting the need for recognition and awareness of this syndromic entity.

Hypothyroidism coordinately and transiently affects myelin protein gene expression in most rat brain regions during postnatal development.

PubMed

Ibarrola, N; Rodríguez-Peña, A

1997-03-28

To assess the role of thyroid hormone on myelin gene expression, we have studied the effect of hypothyroidism on the mRNA steady state levels for the major myelin protein genes: myelin basic protein (MBP), proteolipid protein (PLP), myelin-associated glycoprotein (MAG) and 2':3'-cyclic nucleotide 3'-phosphodiesterase (CNP) in different rat brain regions, during the first postnatal month. We found that hypothyroidism reduces the levels of every myelin protein transcript, with striking differences between the different brain regions. Thus, in the more caudal regions, the effect of hypothyroidism was extremely modest, being only evident at the earlier stages of myelination. In contrast, in the striatum and the cerebral cortex the important decrease in the myelin protein transcripts is maintained beyond the first postnatal month. Therefore, thyroid hormone modulates in a synchronous fashion the expression of the myelin genes and the length of its effect depends on the brain region. On the other hand, hyperthyroidism leads to an increase of the major myelin protein transcripts above control values. Finally, lack of thyroid hormone does not change the expression of the oligodendrocyte progenitor-specific gene, the platelet derived growth factor receptor alpha.

Generic HPLC platform for automated enzyme reaction monitoring: Advancing the assay toolbox for transaminases and other PLP-dependent enzymes.

PubMed

Börner, Tim; Grey, Carl; Adlercreutz, Patrick

2016-08-01

Methods for rapid and direct quantification of enzyme kinetics independent of the substrate stand in high demand for both fundamental research and bioprocess development. This study addresses the need for a generic method by developing an automated, standardizable HPLC platform monitoring reaction progress in near real-time. The method was applied to amine transaminase (ATA) catalyzed reactions intensifying process development for chiral amine synthesis. Autosampler-assisted pipetting facilitates integrated mixing and sampling under controlled temperature. Crude enzyme formulations in high and low substrate concentrations can be employed. Sequential, small (1 µL) sample injections and immediate detection after separation permits fast reaction monitoring with excellent sensitivity, accuracy and reproducibility. Due to its modular design, different chromatographic techniques, e.g. reverse phase and size exclusion chromatography (SEC) can be employed. A novel assay for pyridoxal 5'-phosphate-dependent enzymes is presented using SEC for direct monitoring of enzyme-bound and free reaction intermediates. Time-resolved changes of the different cofactor states, e.g. pyridoxal 5'-phosphate, pyridoxamine 5'-phosphate and the internal aldimine were traced in both half reactions. The combination of the automated HPLC platform with SEC offers a method for substrate-independent screening, which renders a missing piece in the assay and screening toolbox for ATAs and other PLP-dependent enzymes. Copyright © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Coenzyme Q10 and Pyridoxal Phosphate Deficiency Is a Common Feature in Mucopolysaccharidosis Type III.

PubMed

Yubero, Dèlia; Montero, Raquel; O'Callaghan, Mar; Pineda, Mercè; Meavilla, Silvia; Delgadillo, Veronica; Sierra, Cristina; Altimira, Laura; Navas, Plácido; Pope, Simon; Oppenheim, Marcus; Neergheen, Viruna; Ghosh, Arunabha; Mills, Phillipa; Clayton, Peter; Footitt, Emma; Cleary, Maureen; Hargreaves, Iain; Jones, Simon A; Heales, Simon; Artuch, Rafael

2016-01-01

Mucopolysaccharidoses (MPS) are a group of lysosomal storage disorders caused by deficiencies of lysosomal enzymes catalyzing degradation of glycosaminoglycans (GAGs). Previously, we reported a secondary plasma coenzyme Q 10 (CoQ) deficiency in MPS patients. For this study, nine MPS patients were recruited in the Hospital Sant Joan de Déu (HSJD, Barcelona) and two patients in the Neurometabolic Unit, National Hospital (NMU, London), to explore the nutritional status of MPS type III patients by analyzing several vitamins and micronutrients in blood and in cerebrospinal fluid. Plasma CoQ and plasma and cerebrospinal fluid pyridoxal phosphate (PLP) content were analyzed by high-pressure liquid chromatography (HPLC) with electrochemical and fluorescence detection, respectively. We found that most MPS-III patients disclosed low plasma pyridoxal phosphate (PLP) values (seven out of nine) and also low plasma CoQ concentrations (eight out of nine). We observed significantly lower median values of PLP, tocopherol, and CoQ (Mann-Whitney U test, p = 0.006, p = 0.004, and p = 0.001, respectively) in MPS patients when compared with age-matched controls. Chi-square test showed a significant association between the fact of having low plasma PLP and CoQ values in the whole cohort of patients. Cerebrospinal fluid PLP values were clearly deficient in the two patients studied. In conclusion, we report a combined CoQ and PLP deficiency in MPS-III patients. These observations could be related to the complexity of the physiopathology of the disease. If our results are confirmed in larger series of patients, CoQ and PLP therapy could be trialed as coadjuvant therapy with the current MPS treatments.

Autosomal Recessive Retinitis Pigmentosa Caused by Mutations in the MAK Gene

PubMed Central

Luo, Xunda; Héon, Elise; Lam, Byron L.; Weleber, Richard G.; Halder, Jennifer A.; Affatigato, Louisa M.; Goldberg, Jacqueline B.; Sumaroka, Alexander; Schwartz, Sharon B.; Cideciyan, Artur V.; Jacobson, Samuel G.

2011-01-01

Purpose. To determine the disease expression in autosomal recessive (ar) retinitis pigmentosa (RP) caused by mutations in the MAK (male germ cell-associated kinase) gene. Methods. Patients with RP and MAK gene mutations (n = 24; age, 32–77 years at first visit) were studied by ocular examination, perimetry, and optical coherence tomography (OCT). Results. All but one MAK patient were homozygous for an identical truncating mutation in exon 9 and had Ashkenazi Jewish heritage. The carrier frequency of this mutation among 1207 unrelated Ashkenazi control subjects was 1 in 55, making it the most common cause of heritable retinal disease in this population and MAK-associated RP the sixth most common Mendelian disease overall in this group. Visual acuities could be normal into the eighth decade of life. Kinetic fields showed early loss in the superior–temporal quadrant. With more advanced disease, superior and midperipheral function was lost, but the nasal field remained. Only a central island was present at late stages. Pigmentary retinopathy was less prominent in the superior nasal quadrant. Rod-mediated vision was abnormal but detectable in the residual field; all patients had rod>cone dysfunction. Photoreceptor layer thickness was normal centrally but decreased with eccentricity. At the stages studied, there was no evidence of photoreceptor ciliary elongation. Conclusions. The patterns of disease expression in the MAK form of arRP showed some resemblance to patterns described in autosomal dominant RP, especially the form caused by RP1 mutations. The similarity in phenotypes is of interest, considering that there is experimental evidence of interaction between Mak and RP1 in the photoreceptor cilium. PMID:22110072

Two novel mutations in the BCKDHB gene that cause maple syrup urine disease.

PubMed

Han, Bingjuan; Han, Bingchao; Guo, Bin; Liu, Yingxia; Cao, Zhiyang

2018-01-06

Maple syrup urine disease (MSUD) is a rare metabolic disorder of autosomal recessive inheritance caused by decreased activity of branched-chain α-ketoacid dehydrogenase complex (BCKD). Mutations in the three genes (BCKDHA, BCKDHB and DBT) are associated with MSUD. Here, we describe the presenting symptoms, clinical course and gene mutation analysis of a Chinese boy with MSUD. Plasma amino acid analysis was performed by tandem mass spectrometry and the levels of organic acids in urine were measured with gas chromatography-mass spectrometry. The BCKDHB gene was sequenced by Sanger method. Furthermore, the significance of the novel mutations was predicted by Polyphen and Mutationtaster. After diagnosis, the patient was fed with protein-restricted diet to reduce intake of BCAA and was treated with l -carnitine. Metabolic parameters, clinical presentation and mental development were followed up. The patient was diagnosed as MSUD. Two novel BCKDHB mutations (c.523 T > C and c.478-25_552del100) were identified. In silico analysis predicted that the two mutations were "disease causing". The boy tolerated the treatment well and had symptomatic improvement. He presented with mild hypotonia and had nearly normal DQ scores at the age of 10 months. The two novel mutations resulted in the clinical manifestations of MSUD. Our results may reflect the heterogeneity of the pathogenic variants found in patients with MSUD. Copyright © 2018. Published by Elsevier B.V.

[Using exon combined target region capture sequencing chip to detect the disease-causing genes of retinitis pigmentosa].

PubMed

Rong, Weining; Chen, Xuejuan; Li, Huiping; Liu, Yani; Sheng, Xunlun

2014-06-01

To detect the disease-causing genes of 10 retinitis pigmentosa pedigrees by using exon combined target region capture sequencing chip. Pedigree investigation study. From October 2010 to December 2013, 10 RP pedigrees were recruited for this study in Ningxia Eye Hospital. All the patients and family members received complete ophthalmic examinations. DNA was abstracted from patients, family members and controls. Using exon combined target region capture sequencing chip to screen the candidate disease-causing mutations. Polymerase chain reaction (PCR) and direct sequencing were used to confirm the disease-causing mutations. Seventy patients and 23 normal family members were recruited from 10 pedigrees. Among 10 RP pedigrees, 1 was autosomal dominant pedigrees and 9 were autosomal recessive pedigrees. 7 mutations related to 5 genes of 5 pedigrees were detected. A frameshift mutation on BBS7 gene was detected in No.2 pedigree, the patients of this pedigree combined with central obesity, polydactyly and mental handicap. No.2 pedigree was diagnosed as Bardet-Biedl syndrome finally. A missense mutation was detected in No.7 and No.10 pedigrees respectively. Because the patients suffered deafness meanwhile, the final diagnosis was Usher syndrome. A missense mutation on C3 gene related to age-related macular degeneration was also detected in No. 7 pedigrees. A nonsense mutation and a missense mutation on CRB1 gene were detected in No. 1 pedigree and a splicesite mutation on PROM1 gene was detected in No. 5 pedigree. Retinitis pigmentosa is a kind of genetic eye disease with diversity clinical phenotypes. Rapid and effective genetic diagnosis technology combined with clinical characteristics analysis is helpful to improve the level of clinical diagnosis of RP.

Two novel mutations in the GAN gene causing giant axonal neuropathy.

PubMed

Normendez-Martínez, Monica Irad; Monterde-Cruz, Lucero; Martínez, Roberto; Marquez-Harper, Magdalena; Esquitin-Garduño, Nayelli; Valdes-Flores, Margarita; Casas-Avila, Leonora; de Leon-Suarez, Valeria Ponce; Romero-Díaz, Viktor Javier; Hidalgo-Bravo, Alberto

2018-06-06

Giant axonal neuropathy (GAN) is a rare neurodegenerative disease transmitted in an autosomal recessive mode. This disorder presents motor and sensitive symptoms with an onset in early childhood. Progressive neurodegeneration makes the patients wheelchair dependent by the end of the second decade of life. Affected individuals do not survive beyond the third decade of life. Molecular analysis has identified mutations in the gene GAN in patients with this disorder. This gene produces a protein called gigaxonin which is presumably involved in protein degradation via the ubiquitin-proteasome system. However, the underlying molecular mechanism is not clearly understood yet. Here we present the first patient from Mexico with clinical data suggesting GAN. Sequencing of the GAN gene was carried out. Changes in the nucleotide sequence were investigated for their possible impact on protein function and structure using the publicly available prediction tools PolyPhen-2 and PANTHER. The patient is a compound heterozygous carrying two novel mutations in the GAN gene. The sequence analysis revealed two missense mutations in the Kelch repeats domain. In one allele, a C>T transition was found in exon 9 at the nucleotide position 55393 (g.55393C>T). In the other allele, a transversion G>T in exon 11 at the nucleotide position 67471 (g.67471G>T) was observed. Both of the bioinformatic tools predicted that these amino acid substitutions would have a negative impact on gigaxonin's function. This work provides useful information for health professionals and expands the spectrum of disease-causing mutations in the GAN gene and it is the first documented case in Mexican population.

Splicing defect in FKBP10 gene causes autosomal recessive osteogenesis imperfecta disease: a case report.

PubMed

Maghami, Fatemeh; Tabei, Seyed Mohammad Bagher; Moravej, Hossein; Dastsooz, Hassan; Modarresi, Farzaneh; Silawi, Mohammad; Faghihi, Mohammad Ali

2018-05-25

Osteogenesis imperfecta (OI) is a group of connective tissue disorder caused by mutations of genes involved in the production of collagen and its supporting proteins. Although the majority of reported OI variants are in COL1A1 and COL1A2 genes, recent reports have shown problems in other non-collagenous genes involved in the post translational modifications, folding and transport, transcription and proliferation of osteoblasts, bone mineralization, and cell signaling. Up to now, 17 types of OI have been reported in which types I to IV are the most frequent cases with autosomal dominant pattern of inheritance. Here we report an 8- year- old boy with OI who has had multiple fractures since birth and now he is wheelchair-dependent. To identify genetic cause of OI in our patient, whole exome sequencing (WES) was carried out and it revealed a novel deleterious homozygote splice acceptor site mutation (c.1257-2A > G, IVS7-2A > G) in FKBP10 gene in the patient. Then, the identified mutation was confirmed using Sanger sequencing in the proband as homozygous and in his parents as heterozygous, indicating its autosomal recessive pattern of inheritance. In addition, we performed RT-PCR on RNA transcripts originated from skin fibroblast of the proband to analyze the functional effect of the mutation on splicing pattern of FKBP10 gene and it showed skipping of the exon 8 of this gene. Moreover, Real-Time PCR was carried out to quantify the expression level of FKBP10 in the proband and his family members in which it revealed nearly the full decrease in the level of FKBP10 expression in the proband and around 75% decrease in its level in the carriers of the mutation, strongly suggesting the pathogenicity of the mutation. Our study identified, for the first time, a private pathogenic splice site mutation in FKBP10 gene and further prove the involvement of this gene in the rare cases of autosomal recessive OI type XI with distinguished clinical manifestations.

Molecular identification of Neofabraea species associated with bull's-eye rot on apple using rolling-circle amplification of partial EF-1α sequence.

PubMed

Lin, Huijiao; Jiang, Xiang; Yi, Jianping; Wang, Xinguo; Zuo, Ranling; Jiang, Zide; Wang, Weifang; Zhou, Erxun

2018-01-01

A rolling-circle amplification (RCA) method with padlock probes targeted on EF-1α regions was developed for rapid detection of apple bull's-eye rot pathogens, including Neofabraea malicorticis, N. perennans, N. kienholzii, and N. vagabunda (synonym: N. alba). Four padlock probes (PLP-Nm, PLP-Np, PLP-Nk, and PLP-Nv) were designed and tested against 28 samples, including 22 BER pathogen cultures, 4 closely related species, and 2 unrelated species that may cause serious apple decays. The assay successfully identified all the bull's-eye rot pathogenic fungi at the level of species, while no cross-reaction was observed in all target species and no false-positive reaction was observed with all strains used for reference. This study showed that the use of padlock probes and the combination of probe signal amplification by RCA provided an effective and sensitive method for the rapid identification of Neofabraea spp. The method could therefore be a useful tool for monitoring bull's-eye rot pathogens in port quarantine and orchard epidemiological studies.

Deficiency of liver Comparative Gene Identification-58 causes steatohepatitis and fibrosis in mice

PubMed Central

Guo, Feng; Ma, Yinyan; Kadegowda, Anil K. G.; Betters, Jenna L.; Xie, Ping; Liu, George; Liu, Xiuli; Miao, Hongming; Ou, Juanjuan; Su, Xiong; Zheng, Zhenlin; Xue, Bingzhong; Shi, Hang; Yu, Liqing

2013-01-01

Triglyceride (TG) accumulation in hepatocytes (hepatic steatosis) preludes the development of advanced nonalcoholic fatty liver diseases (NAFLDs) such as steatohepatitis, fibrosis, and cirrhosis. Mutations in human Comparative Gene Identification-58 (CGI-58) cause cytosolic TG-rich lipid droplets to accumulate in almost all cell types including hepatocytes. However, it is unclear if CGI-58 mutation causes hepatic steatosis locally or via altering lipid metabolism in other tissues. To directly address this question, we created liver-specific CGI-58 knockout (LivKO) mice. LivKO mice on standard chow diet displayed microvesicular and macrovesicular panlobular steatosis, and progressed to advanced NAFLD stages over time, including lobular inflammation and centrilobular fibrosis. Compared with CGI-58 floxed control littermates, LivKO mice showed 8-fold and 52-fold increases in hepatic TG content, which was associated with 40% and 58% decreases in hepatic TG hydrolase activity at 16 and 42 weeks, respectively. Hepatic cholesterol also increased significantly in LivKO mice. At 42 weeks, LivKO mice showed increased hepatic oxidative stress, plasma aminotransferases, and hepatic mRNAs for genes involved in fibrosis and inflammation, such as α-smooth muscle actin, collagen type 1 α1, tumor necrosis factor α, and interleukin-1β. In conclusion, CGI-58 deficiency in the liver directly causes not only hepatic steatosis but also steatohepatitis and fibrosis. PMID:23733885

Gene expression profiling: cell cycle deregulation and aneuploidy do not cause breast cancer formation in WAP-SVT/t transgenic animals.

PubMed

Klein, Andreas; Guhl, Eva; Zollinger, Raphael; Tzeng, Yin-Jeh; Wessel, Ralf; Hummel, Michael; Graessmann, Monika; Graessmann, Adolf

2005-05-01

Microarray studies revealed that as a first hit the SV40 T/t antigen causes deregulation of 462 genes in mammary gland cells (ME cells) of WAP-SVT/t transgenic animals. The majority of deregulated genes are cell proliferation specific and Rb-E2F dependent, causing ME cell proliferation and gland hyperplasia but not breast cancer formation. In the breast tumor cells a further 207 genes are differentially expressed, most of them belonging to the cell communication category. In tissue culture breast tumor cells frequently switch off WAP-SVT/t transgene expression and regain the morphology and growth characteristics of normal ME cells, although the tumor-revertant cells are aneuploid and only 114 genes regain the expression level of normal ME cells. The profile of retransformants shows that only 38 deregulated genes are tumor-specific, and that none of them is considered to be a typical breast cancer gene.

Methamphetamine Causes Differential Alterations in Gene Expression and Patterns of Histone Acetylation/Hypoacetylation in the Rat Nucleus Accumbens

PubMed Central

Martin, Tracey A.; Jayanthi, Subramaniam; McCoy, Michael T.; Brannock, Christie; Ladenheim, Bruce; Garrett, Tiffany; Lehrmann, Elin; Becker, Kevin G.; Cadet, Jean Lud

2012-01-01

Methamphetamine (METH) addiction is associated with several neuropsychiatric symptoms. Little is known about the effects of METH on gene expression and epigenetic modifications in the rat nucleus accumbens (NAC). Our study investigated the effects of a non-toxic METH injection (20 mg/kg) on gene expression, histone acetylation, and the expression of the histone acetyltransferase (HAT), ATF2, and of the histone deacetylases (HDACs), HDAC1 and HDAC2, in that structure. Microarray analyses done at 1, 8, 16 and 24 hrs after the METH injection identified METH-induced changes in the expression of genes previously implicated in the acute and longterm effects of psychostimulants, including immediate early genes and corticotropin-releasing factor (Crf). In contrast, the METH injection caused time-dependent decreases in the expression of other genes including Npas4 and cholecystokinin (Cck). Pathway analyses showed that genes with altered expression participated in behavioral performance, cell-to-cell signaling, and regulation of gene expression. PCR analyses confirmed the changes in the expression of c-fos, fosB, Crf, Cck, and Npas4 transcripts. To determine if the METH injection caused post-translational changes in histone markers, we used western blot analyses and identified METH-mediated decreases in histone H3 acetylated at lysine 9 (H3K9ac) and lysine 18 (H3K18ac) in nuclear sub-fractions. In contrast, the METH injection caused time-dependent increases in acetylated H4K5 and H4K8. The changes in histone acetylation were accompanied by decreased expression of HDAC1 but increased expression of HDAC2 protein levels. The histone acetyltransferase, ATF2, showed significant METH-induced increased in protein expression. These results suggest that METH-induced alterations in global gene expression seen in rat NAC might be related, in part, to METH-induced changes in histone acetylation secondary to changes in HAT and HDAC expression. The causal role that HATs and HDACs might

Mutations in the profilin 1 gene cause familial amyotrophic lateral sclerosis.

PubMed

Wu, Chi-Hong; Fallini, Claudia; Ticozzi, Nicola; Keagle, Pamela J; Sapp, Peter C; Piotrowska, Katarzyna; Lowe, Patrick; Koppers, Max; McKenna-Yasek, Diane; Baron, Desiree M; Kost, Jason E; Gonzalez-Perez, Paloma; Fox, Andrew D; Adams, Jenni; Taroni, Franco; Tiloca, Cinzia; Leclerc, Ashley Lyn; Chafe, Shawn C; Mangroo, Dev; Moore, Melissa J; Zitzewitz, Jill A; Xu, Zuo-Shang; van den Berg, Leonard H; Glass, Jonathan D; Siciliano, Gabriele; Cirulli, Elizabeth T; Goldstein, David B; Salachas, Francois; Meininger, Vincent; Rossoll, Wilfried; Ratti, Antonia; Gellera, Cinzia; Bosco, Daryl A; Bassell, Gary J; Silani, Vincenzo; Drory, Vivian E; Brown, Robert H; Landers, John E

2012-08-23

Amyotrophic lateral sclerosis (ALS) is a late-onset neurodegenerative disorder resulting from motor neuron death. Approximately 10% of cases are familial (FALS), typically with a dominant inheritance mode. Despite numerous advances in recent years, nearly 50% of FALS cases have unknown genetic aetiology. Here we show that mutations within the profilin 1 (PFN1) gene can cause FALS. PFN1 is crucial for the conversion of monomeric (G)-actin to filamentous (F)-actin. Exome sequencing of two large ALS families showed different mutations within the PFN1 gene. Further sequence analysis identified 4 mutations in 7 out of 274 FALS cases. Cells expressing PFN1 mutants contain ubiquitinated, insoluble aggregates that in many cases contain the ALS-associated protein TDP-43. PFN1 mutants also display decreased bound actin levels and can inhibit axon outgrowth. Furthermore, primary motor neurons expressing mutant PFN1 display smaller growth cones with a reduced F/G-actin ratio. These observations further document that cytoskeletal pathway alterations contribute to ALS pathogenesis.

Mechanical Strain Promotes Oligodendrocyte Differentiation by Global Changes of Gene Expression

PubMed Central

Jagielska, Anna; Lowe, Alexis L.; Makhija, Ekta; Wroblewska, Liliana; Guck, Jochen; Franklin, Robin J. M.; Shivashankar, G. V.; Van Vliet, Krystyn J.

2017-01-01

Differentiation of oligodendrocyte progenitor cells (OPC) to oligodendrocytes and subsequent axon myelination are critical steps in vertebrate central nervous system (CNS) development and regeneration. Growing evidence supports the significance of mechanical factors in oligodendrocyte biology. Here, we explore the effect of mechanical strains within physiological range on OPC proliferation and differentiation, and strain-associated changes in chromatin structure, epigenetics, and gene expression. Sustained tensile strain of 10–15% inhibited OPC proliferation and promoted differentiation into oligodendrocytes. This response to strain required specific interactions of OPCs with extracellular matrix ligands. Applied strain induced changes in nuclear shape, chromatin organization, and resulted in enhanced histone deacetylation, consistent with increased oligodendrocyte differentiation. This response was concurrent with increased mRNA levels of the epigenetic modifier histone deacetylase Hdac11. Inhibition of HDAC proteins eliminated the strain-mediated increase of OPC differentiation, demonstrating a role of HDACs in mechanotransduction of strain to chromatin. RNA sequencing revealed global changes in gene expression associated with strain. Specifically, expression of multiple genes associated with oligodendrocyte differentiation and axon-oligodendrocyte interactions was increased, including cell surface ligands (Ncam, ephrins), cyto- and nucleo-skeleton genes (Fyn, actinins, myosin, nesprin, Sun1), transcription factors (Sox10, Zfp191, Nkx2.2), and myelin genes (Cnp, Plp, Mag). These findings show how mechanical strain can be transmitted to the nucleus to promote oligodendrocyte differentiation, and identify the global landscape of signaling pathways involved in mechanotransduction. These data provide a source of potential new therapeutic avenues to enhance OPC differentiation in vivo. PMID:28473753

Rare Variants in the Gene ALPL That Cause Hypophosphatasia Are Strongly Associated With Ovarian and Uterine Disorders.

PubMed

Dahir, Kathryn M; Tilden, Daniel R; Warner, Jeremy L; Bastarache, Lisa; Smith, Derek K; Gifford, Aliya; Ramirez, Andrea H; Simmons, Jill S; Black, Margo M; Newman, John H; Denny, Josh C

2018-06-01

Mutations in alkaline phosphatase (AlkP), liver/bone/kidney (ALPL), which encodes tissue-nonspecific isozyme AlkP, cause hypophosphatasia (HPP). HPP is suspected by a low-serum AlkP. We hypothesized that some patients with bone or dental disease have undiagnosed HPP, caused by ALPL variants. Our objective was to discover the prevalence of these gene variants in the Vanderbilt University DNA Biobank (BioVU) and to assess phenotypic associations. We identified subjects in BioVU, a repository of DNA, that had at least one of three known, rare HPP disease-causing variants in ALPL: rs199669988, rs121918007, and/or rs121918002. To evaluate for phenotypic associations, we conducted a sequential phenome-wide association study of ALPL variants and then performed a de-identified manual record review to refine the phenotype. Out of 25,822 genotyped individuals, we identified 52 women and 53 men with HPP disease-causing variants in ALPL, 7/1000. None had a clinical diagnosis of HPP. For patients with ALPL variants, the average serum AlkP levels were in the lower range of normal or lower. Forty percent of men and 62% of women had documented bone and/or dental disease, compatible with the diagnosis of HPP. Forty percent of the female patients had ovarian pathology or other gynecological abnormalities compared with 15% seen in controls. Variants in the ALPL gene cause bone and dental disease in patients with and without the standard biomarker, low plasma AlkP. ALPL gene variants are more prevalent than currently reported and underdiagnosed. Gynecologic disease appears to be associated with HPP-causing variants in ALPL.

S81L and G170R mutations causing Primary Hyperoxaluria type I in homozygosis and heterozygosis: an example of positive interallelic complementation

PubMed Central

Montioli, Riccardo; Roncador, Alessandro; Oppici, Elisa; Mandrile, Giorgia; Giachino, Daniela Francesca; Cellini, Barbara; Borri Voltattorni, Carla

2014-01-01

Primary Hyperoxaluria type I (PH1) is a rare disease due to the deficit of peroxisomal alanine:glyoxylate aminotransferase (AGT), a homodimeric pyridoxal-5′-phosphate (PLP) enzyme present in humans as major (Ma) and minor (Mi) allele. PH1-causing mutations are mostly missense identified in both homozygous and compound heterozygous patients. Until now, the pathogenesis of PH1 has been only studied by approaches mimicking homozygous patients, whereas the molecular aspects of the genotype-enzymatic-clinical phenotype relationship in compound heterozygous patients are completely unknown. Here, for the first time, we elucidate the enzymatic phenotype linked to the S81L mutation on AGT-Ma, relative to a PLP-binding residue, and how it changes when the most common mutation G170R on AGT-Mi, known to cause AGT mistargeting without affecting the enzyme functionality, is present in the second allele. By using a bicistronic eukaryotic expression vector, we demonstrate that (i) S81L-Ma is mainly in its apo-form and has a significant peroxisomal localization and (ii) S81L and G170R monomers interact giving rise to the G170R-Mi/S81L-Ma holo-form, which is imported into peroxisomes and exhibits an enhanced functionality with respect to the parental enzymes. These data, integrated with the biochemical features of the heterodimer and the homodimeric counterparts in their purified recombinant form, (i) highlight the molecular basis of the pathogenicity of S81L-Ma and (ii) provide evidence for a positive interallelic complementation between the S81L and G170R monomers. Our study represents a valid approach to investigate the molecular pathogenesis of PH1 in compound heterozygous patients. PMID:24990153

Genes and Gut Bacteria Involved in Luminal Butyrate Reduction Caused by Diet and Loperamide.

PubMed

Hwang, Nakwon; Eom, Taekil; Gupta, Sachin K; Jeong, Seong-Yeop; Jeong, Do-Youn; Kim, Yong Sung; Lee, Ji-Hoon; Sadowsky, Michael J; Unno, Tatsuya

2017-11-28

Unbalanced dietary habits and gut dysmotility are causative factors in metabolic and functional gut disorders, including obesity, diabetes, and constipation. Reduction in luminal butyrate synthesis is known to be associated with gut dysbioses, and studies have suggested that restoring butyrate formation in the colon may improve gut health. In contrast, shifts in different types of gut microbiota may inhibit luminal butyrate synthesis, requiring different treatments to restore colonic bacterial butyrate synthesis. We investigated the influence of high-fat diets (HFD) and low-fiber diets (LFD), and loperamide (LPM) administration, on key bacteria and genes involved in reduction of butyrate synthesis in mice. MiSeq-based microbiota analysis and HiSeq-based differential gene analysis indicated that different types of bacteria and genes were involved in butyrate metabolism in each treatment. Dietary modulation depleted butyrate kinase and phosphate butyryl transferase by decreasing members of the Bacteroidales and Parabacteroides . The HFD also depleted genes involved in succinate synthesis by decreasing Lactobacillus . The LFD and LPM treatments depleted genes involved in crotonoyl-CoA synthesis by decreasing Roseburia and Oscilllibacter . Taken together, our results suggest that different types of bacteria and genes were involved in gut dysbiosis, and that selected treatments may be needed depending on the cause of gut dysfunction.

Novel mutations of the AGXT gene causing primary hyperoxaluria type 1.

PubMed

Yuen, Yuet-Ping; Lai, Chi-Kong; Tong, Gensy Mei-Wah; Wong, Ping-Nam; Wong, Francis Kim-Ming; Mak, Siu-Ka; Lo, Kin-Yee; Wong, Andrew Kui-Man; Tong, Sui-Fan; Chan, Yan-Wo; Lam, Ching-Wan

2004-01-01

Primary hyperoxaluria type 1 (PH1), an inherited cause of nephrolithiasis, is due to a functional defect of the liver-specific peroxisomal enzyme alanine:glyoxylate aminotransferase (AGT). A definitive PH1 diagnosis can be established by analyzing AGT activity in liver tissue or mutation analysis of the AGXT gene. The molecular basis of PH1 in three Chinese patients, two with adult-onset and one with childhood-onset recurrent nephrolithiasis, was established by analyzing the entire AGXT gene. Three novel mutations (c2T>C, c817insAG and c844C>T) and two previously reported mutations (c33insC and 679-IVS6+2delAAgt) were identified. c2T>C converts the initiation codon from ATG to ACG, which predicts significant reduction, if not complete abolition, of protein translation. c817insAG leads to a frameshift and changes the amino acid sequence after codon 274. c844C>T changes glutamine at codon 282 to a termination codon, resulting in protein truncation. This is the first report describing AGXT gene mutations in Chinese patients with PH1. AGXT genotypes cannot fully explain the clinical heterogeneity of PH1, and other factors involved in disease pathogenesis remain to be identified. Our experience emphasizes the importance of excluding PH1 in patients with recurrent nephrolithiasis to avoid delay or inappropriate management.

Exclusion of the APC gene as the cause of a variant form of familial adenomatous polyposis (FAP)

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

Stella, A.; Resta, N.; Susca, F.

Familial adenomatous polyposis (FAP) is a premalignant disease inherited as an autosomal dominant trait, characterized by hundreds to thousands of polyps in the colorectal tract. Recently, the syndrome has been shown to be caused by mutations in the APC (adenomatous polyposis coli) gene located on chromosome 5q21. The authors studied two families that both presented a phenotype different from that of the classical form of FAP. The most important findings observed in these two kindreds are (a) low and variable number of colonic polyps (from 5 to 100) and (b) a slower evolution of the disease, with colon cancer occurringmore » at a more advanced age than in FAP in spite of the early onset of intestinal manifestations. To determine whether mutations of the APC gene are also responsible for this variant syndrome, linkage studies were performed by using a series of markers both intragenic and tightly linked to the APC gene. The results provide evidence for exclusion of the APC gene as the cause of the variant form of polyposis present in the two families described. 30 refs., 1 fig., 1 tab.« less

Hypotrichosis and juvenile macular dystrophy caused by CDH3 mutation: A candidate disease for retinal gene therapy.

PubMed

Singh, Mandeep S; Broadgate, Suzanne; Mathur, Ranjana; Holt, Richard; Halford, Stephanie; MacLaren, Robert E

2016-05-09

Hypotrichosis with juvenile macular dystrophy (HJMD) is an autosomal recessive disorder that causes childhood visual impairment. HJMD is caused by mutations in CDH3 which encodes cadherin-3, a protein expressed in retinal pigment epithelium (RPE) cells that may have a key role in intercellular adhesion. We present a case of HJMD and analyse its phenotypic and molecular characteristics to assess the potential for retinal gene therapy as a means of preventing severe visual loss in this condition. Longitudinal in vivo imaging of the retina showed the relative anatomical preservation of the macula, which suggested the presence of a therapeutic window for gene augmentation therapy to preserve visual acuity. The coding sequence of CDH3 fits within the packaging limit of recombinant adeno-associated virus vectors that have been shown to be safe in clinical trials and can efficiently target RPE cells. This report expands the number of reported cases of HJMD and highlights the phenotypic characteristics to consider when selecting candidates for retinal gene therapy.

Establishing the precise evolutionary history of a gene improves prediction of disease-causing missense mutations

DOE PAGES

Adebali, Ogun; Reznik, Alexander O.; Ory, Daniel S.; ...

2016-02-18

Here, predicting the phenotypic effects of mutations has become an important application in clinical genetic diagnostics. Computational tools evaluate the behavior of the variant over evolutionary time and assume that variations seen during the course of evolution are probably benign in humans. However, current tools do not take into account orthologous/paralogous relationships. Paralogs have dramatically different roles in Mendelian diseases. For example, whereas inactivating mutations in the NPC1 gene cause the neurodegenerative disorder Niemann-Pick C, inactivating mutations in its paralog NPC1L1 are not disease-causing and, moreover, are implicated in protection from coronary heart disease. Methods: We identified major events inmore » NPC1 evolution and revealed and compared orthologs and paralogs of the human NPC1 gene through phylogenetic and protein sequence analyses. We predicted whether an amino acid substitution affects protein function by reducing the organism s fitness. As a result, removing the paralogs and distant homologs improved the overall performance of categorizing disease-causing and benign amino acid substitutions. In conclusion, the results show that a thorough evolutionary analysis followed by identification of orthologs improves the accuracy in predicting disease-causing missense mutations. We anticipate that this approach will be used as a reference in the interpretation of variants in other genetic diseases as well.« less

Establishing the precise evolutionary history of a gene improves prediction of disease-causing missense mutations

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

Adebali, Ogun; Reznik, Alexander O.; Ory, Daniel S.

Here, predicting the phenotypic effects of mutations has become an important application in clinical genetic diagnostics. Computational tools evaluate the behavior of the variant over evolutionary time and assume that variations seen during the course of evolution are probably benign in humans. However, current tools do not take into account orthologous/paralogous relationships. Paralogs have dramatically different roles in Mendelian diseases. For example, whereas inactivating mutations in the NPC1 gene cause the neurodegenerative disorder Niemann-Pick C, inactivating mutations in its paralog NPC1L1 are not disease-causing and, moreover, are implicated in protection from coronary heart disease. Methods: We identified major events inmore » NPC1 evolution and revealed and compared orthologs and paralogs of the human NPC1 gene through phylogenetic and protein sequence analyses. We predicted whether an amino acid substitution affects protein function by reducing the organism s fitness. As a result, removing the paralogs and distant homologs improved the overall performance of categorizing disease-causing and benign amino acid substitutions. In conclusion, the results show that a thorough evolutionary analysis followed by identification of orthologs improves the accuracy in predicting disease-causing missense mutations. We anticipate that this approach will be used as a reference in the interpretation of variants in other genetic diseases as well.« less

Myelin Proteolipid Protein Complexes with αv Integrin and AMPA Receptors In Vivo and Regulates AMPA-Dependent Oligodendrocyte Progenitor Cell Migration through the Modulation of Cell-Surface GluR2 Expression

PubMed Central

Harlow, Danielle E.; Saul, Katherine E.; Komuro, Hitoshi

2015-01-01

In previous studies, stimulation of ionotropic AMPA/kainate glutamate receptors on cultured oligodendrocyte cells induced the formation of a signaling complex that includes the AMPA receptor, integrins, calcium-binding proteins, and, surprisingly, the myelin proteolipid protein (PLP). AMPA stimulation of cultured oligodendrocyte progenitor cells (OPCs) also caused an increase in OPC migration. The current studies focused primarily on the formation of the PLP–αv integrin–AMPA receptor complex in vivo and whether complex formation impacts OPC migration in the brain. We found that in wild-type cerebellum, PLP associates with αv integrin and the calcium-impermeable GluR2 subunit of the AMPA receptor, but in mice lacking PLP, αv integrin did not associate with GluR2. Live imaging studies of OPC migration in ex vivo cerebellar slices demonstrated altered OPC migratory responses to neurotransmitter stimulation in the absence of PLP and GluR2 or when αv integrin levels were reduced. Chemotaxis assays of purified OPCs revealed that AMPA stimulation was neither attractive nor repulsive but clearly increased the migration rate of wild-type but not PLP null OPCs. AMPA receptor stimulation of wild-type OPCs caused decreased cell-surface expression of the GluR2 AMPA receptor subunit and increased intracellular Ca2+ signaling, whereas PLP null OPCs did not reduce GluR2 at the cell surface or increase Ca2+ signaling in response to AMPA treatment. Together, these studies demonstrate that PLP is critical for OPC responses to glutamate signaling and has important implications for OPC responses when levels of glutamate are high in the extracellular space, such as following demyelination. SIGNIFICANCE STATEMENT After demyelination, such as occurs in multiple sclerosis, remyelination of axons is often incomplete, leading to loss of neuronal function and clinical disability. Remyelination may fail because oligodendrocyte precursor cells (OPCs) do not completely migrate into

Novel XLRS1 gene mutations cause X-linked juvenile retinoschisis in Chinese families.

PubMed

Ma, Xiang; Li, Xiaoxin; Wang, Lihua

2008-01-01

To investigate various XLRS1 (RS1) gene mutations in Chinese families with X-linked juvenile retinoschisis (XLRS or RS). Genomic DNA was isolated from leukocytes of 29 male patients with X-linked juvenile retinoschisis, 38 female carriers, and 100 normal controls. All 6 exons of the RS1 gene were amplified by polymerase chain reaction, and the RS1 gene mutations were determined by direct sequencing. Eleven different RS1 mutations in 12 families were identified in the 29 male patients. The mutations comprised eight missense, two frameshift, and one splice donor site mutation. Four of these mutations, one frameshift mutation (26 del T) in exon 1, one frameshift mutation (488 del G) in exon 5, Asp145His and Arg156Gly in exon 5, have not been previously described. One novel non-disease-related polymorphism, 576C to T (Pro192Pro) in exon 6, was also found. Six recurrent mutations, Ser73Pro and Arg102Gln mutations in exon 4 and Arg200Cys, Arg209His, Arg213Gln, and Cys223Arg mutations in exon 6, were also identified in this study. RS1 gene mutations caused X-linked juvenile retinoschisis in these Chinese families.

Two common low density lipoprotein receptor gene mutations cause familial hypercholesterolemia in Afrikaners.

PubMed

Leitersdorf, E; Van der Westhuyzen, D R; Coetzee, G A; Hobbs, H H

1989-09-01

Familial hypercholesterolemia (FH), an autosomal dominant disease caused by mutations in the LDL receptor gene, is five times more frequent in the Afrikaner population of South Africa than it is in the population of the United States and Europe. It has been proposed that the high frequency is due to a founder effect. In this paper, we characterized 24 mutant LDL receptor alleles from 12 Afrikaner individuals homozygous for FH. We identified two mutations that together makeup greater than 95% of the mutant LDL receptor genes represented in our sample. Both mutations were basepair substitutions that result in single-amino acid changes. Each mutation can be detected readily with the polymerase chain reaction and restriction analysis. The finding of two common LDL receptor mutations in the Afrikaner FH homozygotes predicts that these mutations will predominate in the Afrikaner population and that the high frequency of FH is due to a founder effect. The increased incidence of ischemic heart disease in the Afrikaner population may in part be due to the high frequency of these two mutations in the LDL receptor gene.

Alu-mediated large deletion of the CDSN gene as a cause of peeling skin disease.

PubMed

Wada, T; Matsuda, Y; Muraoka, M; Toma, T; Takehara, K; Fujimoto, M; Yachie, A

2014-10-01

Peeling skin disease (PSD) is an autosomal recessive skin disorder caused by mutations in CDSN and is characterized by superficial peeling of the upper epidermis. Corneodesmosin (CDSN) is a major component of corneodesmosomes that plays an important role in maintaining epidermis integrity. Herein, we report a patient with PSD caused by a novel homozygous large deletion in the 6p21.3 region encompassing the CDSN gene, which abrogates CDSN expression. Several genes including C6orf15, PSORS1C1, PSORS1C2, CCHCR1, and TCF19 were also deleted, however, the patient showed only clinical features typical of PSD. The deletion size was 59.1 kb. Analysis of the sequence surrounding the breakpoint showed that both telomeric and centromeric breakpoints existed within Alu-S sequences that were oriented in opposite directions. These results suggest an Alu-mediated recombination event as the mechanism underlying the deletion in our patient. © 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Hypomorphic mutation in mouse Nppc gene causes retarded bone growth due to impaired endochondral ossification

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

Tsuji, Takehito; Kondo, Eri; Yasoda, Akihiro

2008-11-07

Long bone abnormality (lbab/lbab) is a spontaneous mutant mouse characterized by dwarfism with shorter long bones. A missense mutation was reported in the Nppc gene, which encodes C-type natriuretic peptide (CNP), but it has not been confirmed whether this mutation is responsible for the dwarf phenotype. To verify that the mutation causes the dwarfism of lbab/lbab mice, we first investigated the effect of CNP in lbab/lbab mice. By transgenic rescue with chondrocyte-specific expression of CNP, the dwarf phenotype in lbab/lbab mice was completely compensated. Next, we revealed that CNP derived from the lbab allele retained only slight activity to inducemore » cGMP production through its receptor. Histological analysis showed that both proliferative and hypertrophic zones of chondrocytes in the growth plate of lbab/lbab mice were markedly reduced. Our results demonstrate that lbab/lbab mice have a hypomorphic mutation in the Nppc gene that is responsible for dwarfism caused by impaired endochondral ossification.« less

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.

Gene Therapy for the Retinal Degeneration of Usher Syndrome Caused by Mutations in MYO7A.

PubMed

Lopes, Vanda S; Williams, David S

2015-01-20

Usher syndrome is a deaf-blindness disorder. One of the subtypes, Usher 1B, is caused by loss of function of the gene encoding the unconventional myosin, MYO7A. A variety of different viral-based delivery approaches have been tested for retinal gene therapy to prevent the blindness of Usher 1B, and a clinical trial based on one of these approaches has begun. This review evaluates the different approaches. Copyright © 2015 Cold Spring Harbor Laboratory Press; all rights reserved.

Investigation of genetic diversity and epidemiological characteristics of Pasteurella multocida isolates from poultry in southwest China by population structure, multi-locus sequence typing and virulence-associated gene profile analysis.

PubMed

Li, Zhangcheng; Cheng, Fangjun; Lan, Shimei; Guo, Jianhua; Liu, Wei; Li, Xiaoyan; Luo, Zeli; Zhang, Manli; Wu, Juan; Shi, Yang

2018-04-25

Fowl cholera caused by Pasteurella multocida has always been a disease of global importance for poultry production. The aim of this study was to obtain more information about the epidemiology of avian P. multocida infection in southwest China and the genetic characteristics of clinical isolates. P. multocida isolates were characterized by biochemical and molecular-biological methods. The distributions of the capsular serogroups, the phenotypic antimicrobial resistance profiles, lipopolysaccharide (LPS) genotyping and the presence of 19 virulence genes were investigated in 45 isolates of P. multocida that were associated with clinical disease in poultry. The genetic diversity of P. multocida strains was performed by 16S rRNA and rpoB gene sequence analysis as well as multilocus sequence typing (MLST). The results showed that most (80.0%) of the P. multocida isolates in this study represented special P. multocida subspecies, and 71.1% of the isolates showed multiple-drug resistance. 45 isolates belonged to capsular types: A (100%) and two LPS genotypes: L1 (95.6%) and L3 (4.4%). MLST revealed two new alleles (pmi77 and gdh57) and one new sequence type (ST342). ST129 types dominated in 45 P. multocida isolates. Isolates belonging to ST129 were with the genes ompH+plpB+ptfA+tonB, whereas ST342 included isolates with fur+hgbA+tonB genes. Population genetic analysis and the MLST results revealed that at least one new ST genotype was present in the avian P. multocida in China. These findings provide novel insights into the epidemiological characteristics of avian P. multocida isolates in southwest China.

Waardenburg syndrome type II in a Chinese patient caused by a novel nonsense mutation in the SOX10 gene.

PubMed

Ma, Jing; Zhang, Tie-Song; Lin, Ken; Sun, Hao; Jiang, Hong-Chao; Yang, Yan-Li; Low, Fan; Gao, Ying-Qin; Ruan, Biao

2016-06-01

Waardenburg syndrome is a congenital genetic disorder. It is the most common type of syndromic hearing impairment with highly genetic heterogeneity and proved to be related by 6 genes as follows: PAX3, MITF, SNAI2, EDN3, EDNRB and SOX10. This article aims to identify the genetic causes of a Chinese WS child patient. A Chinese WS child was collected for clinical data collection by questionnaire survey. DNA samples of proband and his parents were extracted from peripheral blood samples. Six candidate genes were sequenced by the Trusight One sequencing panel on the illumina NextSeq 500 platform. A novel nonsense heterozygous mutation was found in the coding region of exon 2 in the SOX10 gene of proband. The novel nonsense heterozygous mutation could cause the replacement of the 55th lysine codon by stop codon (484T > C, C142R) and further more possibly cause terminating the protein translation in advance. However, both proband's parents had no mutation of genes above mentioned. The gene mutation of SOX10 [NM_006941.3 c.163A > T] is a novel nonsense mutation. No record of this mutation has been found in dbSNP, HGMD, 1000 Genomes Project, ClinVar and ESP6500 databases. It meets the condition of PS2 of strong evidence in 2015 ACMG Standards and Guidelines. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Multiple organ gigantism caused by mutation in VmPPD gene in blackgram (Vigna mungo)

PubMed Central

Naito, Ken; Takahashi, Yu; Chaitieng, Bubpa; Hirano, Kumi; Kaga, Akito; Takagi, Kyoko; Ogiso-Tanaka, Eri; Thavarasook, Charaspon; Ishimoto, Masao; Tomooka, Norihiko

2017-01-01

Seed size is one of the most important traits in leguminous crops. We obtained a recessive mutant of blackgram that had greatly enlarged leaves, stems and seeds. The mutant produced 100% bigger leaves, 50% more biomass and 70% larger seeds though it produced 40% less number of seeds. We designated the mutant as multiple-organ-gigantism (mog) and found the mog phenotype was due to increase in cell numbers but not in cell size. We also found the mog mutant showed a rippled leaf (rl) phenotype, which was probably caused by a pleiotropic effect of the mutation. We performed a map-based cloning and successfully identified an 8 bp deletion in the coding sequence of VmPPD gene, an orthologue of Arabidopsis PEAPOD (PPD) that regulates arrest of cell divisions in meristematic cells. We found no other mutations in the neighboring genes between the mutant and the wild type. We also knocked down GmPPD genes and reproduced both the mog and rl phenotypes in soybean. Controlling PPD genes to produce the mog phenotype is highly valuable for breeding since larger seed size could directly increase the commercial values of grain legumes. PMID:28588392

Multiple organ gigantism caused by mutation in VmPPD gene in blackgram (Vigna mungo).

PubMed

Naito, Ken; Takahashi, Yu; Chaitieng, Bubpa; Hirano, Kumi; Kaga, Akito; Takagi, Kyoko; Ogiso-Tanaka, Eri; Thavarasook, Charaspon; Ishimoto, Masao; Tomooka, Norihiko

2017-03-01

Seed size is one of the most important traits in leguminous crops. We obtained a recessive mutant of blackgram that had greatly enlarged leaves, stems and seeds. The mutant produced 100% bigger leaves, 50% more biomass and 70% larger seeds though it produced 40% less number of seeds. We designated the mutant as multiple-organ-gigantism ( mog ) and found the mog phenotype was due to increase in cell numbers but not in cell size. We also found the mog mutant showed a rippled leaf ( rl ) phenotype, which was probably caused by a pleiotropic effect of the mutation. We performed a map-based cloning and successfully identified an 8 bp deletion in the coding sequence of VmPPD gene, an orthologue of Arabidopsis PEAPOD ( PPD ) that regulates arrest of cell divisions in meristematic cells . We found no other mutations in the neighboring genes between the mutant and the wild type. We also knocked down GmPPD genes and reproduced both the mog and rl phenotypes in soybean. Controlling PPD genes to produce the mog phenotype is highly valuable for breeding since larger seed size could directly increase the commercial values of grain legumes.

Functional analyses of the three simian hemorrhagic fever virus nonstructural protein 1 papain-like proteases.

PubMed

Vatter, Heather A; Di, Han; Donaldson, Eric F; Radu, Gertrud U; Maines, Taronna R; Brinton, Margo A

2014-08-01

The N-terminal region of simian hemorrhagic fever virus (SHFV) nonstructural polyprotein 1a is predicted to encode three papain-like proteases (PLP1α, PLP1β, and PLP1γ). Catalytic residues and cleavage sites for each of the SHFV PLP1s were predicted by alignment of the SHFV PLP1 region sequences with each other as well as with those of other arteriviruses, and the predicted catalytic residues were shown to be proximal by homology modeling of the SHFV nsp1s on porcine respiratory and reproductive syndrome virus (PRRSV) nsp1 crystal structures. The functionality of the predicted catalytic Cys residues and cleavage sites was tested by analysis of the autoproteolytic products generated in in vitro transcription/translation reactions done with wild-type or mutant SHFV nsp1 constructs. Cleavage sites were also analyzed by mass spectroscopy analysis of selected immunoprecipitated cleavage products. The data showed that each of the three SHFV PLP1s is an active protease. Cys63 was identified as the catalytic Cys of SHFV PLP1α and is adjacent to an Ala instead of the canonical Tyr observed in other arterivirus PLP1s. SHFV PLP1γ is able to cleave at both downstream and upstream nsp1 junction sites. Although intermediate precursor polyproteins as well as alternative products generated by each of the SHFV PLP1s cleaving at sites within the N-terminal region of nsp1β were produced in the in vitro reactions, Western blotting of SHFV-infected, MA104 cell lysates with SHFV nsp1 protein-specific antibodies detected only the three mature nsp1 proteins. SHFV is unique among arteriviruses in having three N-terminal papain-like protease 1 (PLP1) domains. Other arteriviruses encode one or two active PLP1s. This is the first functional study of the SHFV PLP1s. Analysis of the products of in vitro autoprocessing of an N-terminal SHFV nonstructural 1a polypeptide fragment showed that each of the three SHFV PLP1s is active, and the predicted catalytic Cys residues and cleavage sites

An atypical case of fragile X syndrome caused by a deletion that includes the FMR1 gene.

PubMed Central

Quan, F; Zonana, J; Gunter, K; Peterson, K L; Magenis, R E; Popovich, B W

1995-01-01

Fragile X syndrome is the most common form of inherited mental retardation and results from the transcriptional inactivation of the FMR1 gene. In the vast majority of cases, this is caused by the expansion of an unstable CGG repeat in the first exon of the FMR1 gene. We describe here a phenotypically atypical case of fragile X syndrome, caused by a deletion that includes the entire FMR1 gene and > or = 9.0 Mb of flanking DNA. The proband, RK, was a 6-year-old mentally retarded male with obesity and anal atresia. A diagnosis of fragile X syndrome was established by the failure of RK's DNA to hybridize to a 558-bp PstI-XhoI fragment (pfxa3) specific for the 5'-end of the FMR1 gene. The analysis of flanking markers in the interval from Xq26.3-q28 indicated a deletion extending from between 160-500 kb distal and 9.0 Mb proximal to the FMR1 gene. High-resolution chromosome banding confirmed a deletion with breakpoints in Xq26.3 and Xq27.3. This deletion was maternally transmitted and arose as a new mutation on the grandpaternal X chromosome. The maternal transmission of the deletion was confirmed by FISH using a 34-kb cosmid (c31.4) containing most of the FMR1 gene. These results indicated that RK carried a deletion of the FMR1 region with the most proximal breakpoint described to date. This patient's unusual clinical presentation may indicate the presence of genes located in the deleted interval proximal to the FMR1 locus that are able to modify the fragile X syndrome phenotype. Images Figure 5 Figure 2 Figure 4 Figure 6 Figure 7 PMID:7726157

A new missense mutation in the BCKDHB gene causes the classic form of maple syrup urine disease (MSUD).

PubMed

Miryounesi, Mohammad; Ghafouri-Fard, Soudeh; Goodarzi, Hamedreza; Fardaei, Majid

2015-05-01

Maple syrup urine disease (MSUD) is an autosomal recessive metabolic disease caused by mutations in the BCKDHA, BCKDHB, DBT and DLD genes, which encode the E1α, E1β, E2 and E3 subunits of the branched chain α ketoacid dehydrogenase (BCKD) complex, respectively. This complex is involved in the metabolism of branched-chain amino acids. In this study, we analyzed the DNA sequences of BCKDHA and BCKDHB genes in an infant who suffered from MSUD and died at the age of 6 months. We found a new missense mutation in exon 5 of BCKDHB gene (c.508C>T). The heterozygosity of the parents for the mentioned nucleotide change was confirmed by direct sequence analysis of the corresponding segment. Another missense mutation has been found in the same codon previously and shown by in silico analyses to be deleterious. This report provides further evidence that this amino acid change can cause classic MSUD.

Trajectory of phantom limb pain relief using mirror therapy: Retrospective analysis of two studies.

PubMed

Griffin, Sarah C; Curran, Sean; Chan, Annie W Y; Finn, Sacha B; Baker, Chris I; Pasquina, Paul F; Tsao, Jack W

2017-04-01

Research indicates that mirror therapy reduces phantom limb pain (PLP). Objectives were to determine when mirror therapy works in those who respond to treatment, the relevance of baseline PLP to when pain relief occurs, and what pain symptoms respond to mirror therapy. Data from two independent cohorts with unilateral lower limb amputation were analyzed for this study (n=33). Mirror therapy consisted of 15-min sessions in which amputees performed synchronous movements of the phantom and intact legs/feet. PLP was measured using a visual analogue scale and the Short-Form McGill Pain Questionnaire. The severity of PLP at the beginning of treatment predicted when pain relief occurred. Those with low baseline PLP experienced a reduction (p<0.05) in PLP by session 7 of treatment, those with medium baseline PLP experienced pain relief by session 14 of treatment, and those with high baseline PLP experienced pain relief by session 21 of treatment. Mirror therapy reduced throbbing, shooting, stabbing, sharp, cramping, aching, tender, splitting, tiring/exhausting, and punishing-cruel pain symptoms. The degree of PLP at baseline predicts when mirror therapy relieves pain. This article indicates that the degree of baseline PLP affects when mirror therapy relieves pain: relief occurs by session 7 in patients with low PLP but by session 21 in patients with high PLP. Clinicians should anticipate slower pain relief in patients who begin treatment with high levels of pain. ClinicalTrials.gov numbers:NCT00623818 and NCT00662415. Copyright © 2017 Scandinavian Association for the Study of Pain. All rights reserved.

Multi-modal antigen specific therapy for autoimmunity.

PubMed

Legge, K L; Bell, J J; Li, L; Gregg, R; Caprio, J C; Zaghouani, H

2001-10-01

Peripheral tolerance, represents an attractive strategy to down-regulate previously activated T cells and suppress an ongoing disease. Herein, immunoglobulins (Igs) were used to deliver self and altered self peptides for efficient peptide presentation without costimulation to test for modulation of experimental allergic encephalomyelitis (EAE). Accordingly, the encephalitogenic proteolipid protein (PLP) sequence 139-151 (referred to as PLP1) and an altered form of PLP1 known as PLP-LR were genetically expressed on Igs and the resulting Ig-PLP1 and Ig-PLP-LR were tested for efficient presentation of the peptides and for amelioration of ongoing EAE. Evidence is presented indicating that Ig-PLP1 as well as Ig-PLP-LR given in saline to mice with ongoing clinical EAE suppresses subsequent relapses. However, aggregation of both chimeras allows crosslinking of Fcgamma receptors (FcgammaRs) and induction of IL-10 production by APCs but does not promote the up-regulation of costimulatory molecules. Consequently, IL-10 displays bystander suppression and synergizes with presentation without costimulation to drive effective modulation of EAE. As Ig-PLP1 is more potent than Ig-PLP-LR in the down-regulation of T cells, we conclude that peptide affinity plays a critical role in this multi-modal approach of T cell modulation.

Two common low density lipoprotein receptor gene mutations cause familial hypercholesterolemia in Afrikaners.

PubMed Central

Leitersdorf, E; Van der Westhuyzen, D R; Coetzee, G A; Hobbs, H H

1989-01-01

Familial hypercholesterolemia (FH), an autosomal dominant disease caused by mutations in the LDL receptor gene, is five times more frequent in the Afrikaner population of South Africa than it is in the population of the United States and Europe. It has been proposed that the high frequency is due to a founder effect. In this paper, we characterized 24 mutant LDL receptor alleles from 12 Afrikaner individuals homozygous for FH. We identified two mutations that together makeup greater than 95% of the mutant LDL receptor genes represented in our sample. Both mutations were basepair substitutions that result in single-amino acid changes. Each mutation can be detected readily with the polymerase chain reaction and restriction analysis. The finding of two common LDL receptor mutations in the Afrikaner FH homozygotes predicts that these mutations will predominate in the Afrikaner population and that the high frequency of FH is due to a founder effect. The increased incidence of ischemic heart disease in the Afrikaner population may in part be due to the high frequency of these two mutations in the LDL receptor gene. Images PMID:2569482

131I-Traced PLGA-Lipid Nanoparticles as Drug Delivery Carriers for the Targeted Chemotherapeutic Treatment of Melanoma

NASA Astrophysics Data System (ADS)

Wang, Haiyan; Sheng, Weizhong

2017-05-01

Herein, folic acid (FA) conjugated Poly(d,l-lactide-co-glycolide) (PLGA)-lipid composites (FA-PL) were developed as nanocarriers for the targeted delivery of insoluble anti-cancer drug paclitaxel (PTX), resulting FA-PLP nanoparticles. Furthermore, 131I, as a radioactive tracer, was used to label FA-PLP nanoparticles (FA-PLP-131I) to evaluate their cell uptake activity, in vivo blood circulation, and biodistribution. The FA-PLP-131I nanoparticles had a spherical morphology with great stability, a narrow size distribution (165.6 and 181.2 nm), and -22.1 mV in average zeta potential. Confocal laser scanning microscopy indicated that the targeting molecule FA promotes PLP-131I uptake by melanoma B16F10 cells, which was further confirmed by the cell incorporation rate via 131I activity detection as measured by a gamma counter. FA-PLP-131I without PTX (FA-PL-131I) shows minor cytotoxicity, good biocompatibility, while FA-PLP-131I was demonstrated to have efficient cell viability suppression compared to free PTX and PLP-131I. Following intravenous injection, the blood circulation half-life of free PTX ( t 1/2 = 5.4 ± 0.23 h) was prolonged to 18.5 ± 0.5 h by FA-PLP-131I. Through FA targeting, the tumor uptake of FA-PLP-131I was approximately 4.41- and 12.8-fold higher compared to that of PLP-131I and free PTX-131I, respectively. Moreover, following 40 days of treatment, FA-PLP-131I showed an improved tumor inhibition effect compared to free PTX and PLP-131I, with no relapse and no remarkable systemic in vivo toxicity. The results demonstrate that the 131I-labeled PLGA-lipid nanoparticle can be simultaneously applied for targeted drug delivery and reliable tracking of drugs in vivo.

Ogura-CMS in Chinese cabbage (Brassica rapa ssp. pekinensis) causes delayed expression of many nuclear genes.

PubMed

Dong, Xiangshu; Kim, Wan Kyu; Lim, Yong-Pyo; Kim, Yeon-Ki; Hur, Yoonkang

2013-02-01

We investigated the mechanism regulating cytoplasmic male sterility (CMS) in Brassica rapa ssp. pekinensis using floral bud transcriptome analyses of Ogura-CMS Chinese cabbage and its maintainer line in B. rapa 300-K oligomeric probe (Br300K) microarrays. Ogura-CMS Chinese cabbage produced few and infertile pollen grains on indehiscent anthers. Compared to the maintainer line, CMS plants had shorter filaments and plant growth, and delayed flowering and pollen development. In microarray analysis, 4646 genes showed different expression, depending on floral bud size, between Ogura-CMS and its maintainer line. We found 108 and 62 genes specifically expressed in Ogura-CMS and its maintainer line, respectively. Ogura-CMS line-specific genes included stress-related, redox-related, and B. rapa novel genes. In the maintainer line, genes related to pollen coat and germination were specifically expressed in floral buds longer than 3mm, suggesting insufficient expression of these genes in Ogura-CMS is directly related to dysfunctional pollen. In addition, many nuclear genes associated with auxin response, ATP synthesis, pollen development and stress response had delayed expression in Ogura-CMS plants compared to the maintainer line, which is consistent with the delay in growth and development of Ogura-CMS plants. Delayed expression may reduce pollen grain production and/or cause sterility, implying that mitochondrial, retrograde signaling delays nuclear gene expression. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

New splicing mutation in the choline kinase beta (CHKB) gene causing a muscular dystrophy detected by whole-exome sequencing.

PubMed

Oliveira, Jorge; Negrão, Luís; Fineza, Isabel; Taipa, Ricardo; Melo-Pires, Manuel; Fortuna, Ana Maria; Gonçalves, Ana Rita; Froufe, Hugo; Egas, Conceição; Santos, Rosário; Sousa, Mário

2015-06-01

Muscular dystrophies (MDs) are a group of hereditary muscle disorders that include two particularly heterogeneous subgroups: limb-girdle MD and congenital MD, linked to 52 different genes (seven common to both subgroups). Massive parallel sequencing technology may avoid the usual stepwise gene-by-gene analysis. We report the whole-exome sequencing (WES) analysis of a patient with childhood-onset progressive MD, also presenting mental retardation and dilated cardiomyopathy. Conventional sequencing had excluded eight candidate genes. WES of the trio (patient and parents) was performed using the ion proton sequencing system. Data analysis resorted to filtering steps using the GEMINI software revealed a novel silent variant in the choline kinase beta (CHKB) gene. Inspection of sequence alignments ultimately identified the causal variant (CHKB:c.1031+3G>C). This splice site mutation was confirmed using Sanger sequencing and its effect was further evaluated with gene expression analysis. On reassessment of the muscle biopsy, typical abnormal mitochondrial oxidative changes were observed. Mutations in CHKB have been shown to cause phosphatidylcholine deficiency in myofibers, causing a rare form of CMD (only 21 patients reported). Notwithstanding interpretative difficulties that need to be overcome before the integration of WES in the diagnostic workflow, this work corroborates its utility in solving cases from highly heterogeneous groups of diseases, in which conventional diagnostic approaches fail to provide a definitive diagnosis.

Disrupted auto-regulation of the spliceosomal gene SNRPB causes cerebro–costo–mandibular syndrome

PubMed Central

Lynch, Danielle C.; Revil, Timothée; Schwartzentruber, Jeremy; Bhoj, Elizabeth J.; Innes, A. Micheil; Lamont, Ryan E.; Lemire, Edmond G.; Chodirker, Bernard N.; Taylor, Juliet P.; Zackai, Elaine H.; McLeod, D. Ross; Kirk, Edwin P.; Hoover-Fong, Julie; Fleming, Leah; Savarirayan, Ravi; Boycott, Kym; MacKenzie, Alex; Brudno, Michael; Bulman, Dennis; Dyment, David; Majewski, Jacek; Jerome-Majewska, Loydie A.; Parboosingh, Jillian S.; Bernier, Francois P.

2014-01-01

Elucidating the function of highly conserved regulatory sequences is a significant challenge in genomics today. Certain intragenic highly conserved elements have been associated with regulating levels of core components of the spliceosome and alternative splicing of downstream genes. Here we identify mutations in one such element, a regulatory alternative exon of SNRPB as the cause of cerebro–costo–mandibular syndrome. This exon contains a premature termination codon that triggers nonsense-mediated mRNA decay when included in the transcript. These mutations cause increased inclusion of the alternative exon and decreased overall expression of SNRPB. We provide evidence for the functional importance of this conserved intragenic element in the regulation of alternative splicing and development, and suggest that the evolution of such a regulatory mechanism has contributed to the complexity of mammalian development. PMID:25047197

Disrupted auto-regulation of the spliceosomal gene SNRPB causes cerebro-costo-mandibular syndrome.

PubMed

Lynch, Danielle C; Revil, Timothée; Schwartzentruber, Jeremy; Bhoj, Elizabeth J; Innes, A Micheil; Lamont, Ryan E; Lemire, Edmond G; Chodirker, Bernard N; Taylor, Juliet P; Zackai, Elaine H; McLeod, D Ross; Kirk, Edwin P; Hoover-Fong, Julie; Fleming, Leah; Savarirayan, Ravi; Majewski, Jacek; Jerome-Majewska, Loydie A; Parboosingh, Jillian S; Bernier, Francois P

2014-07-22

Elucidating the function of highly conserved regulatory sequences is a significant challenge in genomics today. Certain intragenic highly conserved elements have been associated with regulating levels of core components of the spliceosome and alternative splicing of downstream genes. Here we identify mutations in one such element, a regulatory alternative exon of SNRPB as the cause of cerebro-costo-mandibular syndrome. This exon contains a premature termination codon that triggers nonsense-mediated mRNA decay when included in the transcript. These mutations cause increased inclusion of the alternative exon and decreased overall expression of SNRPB. We provide evidence for the functional importance of this conserved intragenic element in the regulation of alternative splicing and development, and suggest that the evolution of such a regulatory mechanism has contributed to the complexity of mammalian development.

Alu element insertion in PKLR gene as a novel cause of pyruvate kinase deficiency in Middle Eastern patients.

PubMed

Lesmana, Harry; Dyer, Lisa; Li, Xia; Denton, James; Griffiths, Jenna; Chonat, Satheesh; Seu, Katie G; Heeney, Matthew M; Zhang, Kejian; Hopkin, Robert J; Kalfa, Theodosia A

2018-03-01

Pyruvate kinase deficiency (PKD) is the most frequent red blood cell enzyme abnormality of the glycolytic pathway and the most common cause of hereditary nonspherocytic hemolytic anemia. Over 250 PKLR-gene mutations have been described, including missense/nonsense, splicing and regulatory mutations, small insertions, small and gross deletions, causing PKD and hemolytic anemia of variable severity. Alu retrotransposons are the most abundant mobile DNA sequences in the human genome, contributing to almost 11% of its mass. Alu insertions have been associated with a number of human diseases either by disrupting a coding region or a splice signal. Here, we report on two unrelated Middle Eastern patients, both born from consanguineous parents, with transfusion-dependent hemolytic anemia, where sequence analysis revealed a homozygous insertion of AluYb9 within exon 6 of the PKLR gene, causing precipitous decrease of PKLR RNA levels. This Alu element insertion consists a previously unrecognized mechanism underlying pathogenesis of PKD. © 2017 Wiley Periodicals, Inc.

Novel mutations of the carbohydrate sulfotransferase-6 (CHST6) gene causing macular corneal dystrophy in India.

PubMed

Sultana, Afia; Sridhar, Mittanamalli S; Jagannathan, Aparna; Balasubramanian, Dorairajan; Kannabiran, Chitra; Klintworth, Gordon K

2003-12-22

Macular corneal dystrophy (MCD) is an autosomal recessive disorder characterized by progressive central haze, confluent punctate opacities and abnormal deposits in the cornea. It is caused by mutations in the carbohydrate sulfotransferase-6 (CHST6) gene, encoding corneal N-acetyl glucosamine-6-O-sulfotransferase (C-GlcNAc-6-ST). We screened the CHST6 gene for mutations in Indian families with MCD, in order to determine the range of pathogenic mutations. Genomic DNA was isolated from peripheral blood leukocytes of patients with MCD and normal controls. The coding regions of the CHST6 gene were amplified using three pairs of primers and amplified products were directly sequenced. We identified 22 (5 nonsense, 5 frameshift, 2 insertion, and 10 missense) mutations in 36 patients from 31 families with MCD, supporting the conclusion that loss of function of this gene is responsible for this corneal disease. Seventeen of these mutations are novel. These data highlight the allelic heterogeneity of macular corneal dystrophy in Indian patients.

GeneCOST: a novel scoring-based prioritization framework for identifying disease causing genes.

PubMed

Ozer, Bugra; Sağıroğlu, Mahmut; Demirci, Hüseyin

2015-11-15

Due to the big data produced by next-generation sequencing studies, there is an evident need for methods to extract the valuable information gathered from these experiments. In this work, we propose GeneCOST, a novel scoring-based method to evaluate every gene for their disease association. Without any prior filtering and any prior knowledge, we assign a disease likelihood score to each gene in correspondence with their variations. Then, we rank all genes based on frequency, conservation, pedigree and detailed variation information to find out the causative reason of the disease state. We demonstrate the usage of GeneCOST with public and real life Mendelian disease cases including recessive, dominant, compound heterozygous and sporadic models. As a result, we were able to identify causative reason behind the disease state in top rankings of our list, proving that this novel prioritization framework provides a powerful environment for the analysis in genetic disease studies alternative to filtering-based approaches. GeneCOST software is freely available at www.igbam.bilgem.tubitak.gov.tr/en/softwares/genecost-en/index.html. buozer@gmail.com Supplementary data are available at Bioinformatics online. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Increased Grik4 Gene Dosage Causes Imbalanced Circuit Output and Human Disease-Related Behaviors.

PubMed

Arora, Vineet; Pecoraro, Valeria; Aller, M Isabel; Román, Celia; Paternain, Ana V; Lerma, Juan

2018-06-26

Altered glutamatergic neurotransmission is thought to contribute to mental disorders and neurodegenerative diseases. Copy-number variation in genes associated with glutamatergic synapses represents a source of genetic variability, possibly underlying neurological and mental disease susceptibility. The GRIK4 gene encodes a high-affinity kainate receptor subunit of essentially unknown function, although de novo duplication of the 11q23.3-q24.1 locus to which it maps has been detected in autism and other disorders. To determine how changes in the dose of Grik4 affect synaptic activity, we studied mice overexpressing this gene in the forebrain. A mild gain in Grik4 enhances synaptic transmission, causing a persistent imbalance in inhibitory and excitatory activity and disturbing the circuits responsible for the main amygdala outputs. These changes in glutamatergic activity reverse when Grik4 levels are normalized; thus, they may account for the behavioral abnormalities in disorders like autism or schizophrenia. Copyright © 2018 Agencia Estatal Consejo Superior de Investigaciones Científicas. Published by Elsevier Inc. All rights reserved.

Phenotypic spectrum of autosomal recessive cone-rod dystrophies caused by mutations in the ABCA4 (ABCR) gene.

PubMed

Klevering, B Jeroen; Blankenagel, Anita; Maugeri, Alessandra; Cremers, Frans P M; Hoyng, Carel B; Rohrschneider, Klaus

2002-06-01

To describe the phenotype of 12 patients with autosomal recessive or isolated cone-rod types of progressive retinal degeneration (CRD) caused by mutations in the ABCA4 gene. The charts of patients who had originally received a diagnosis of isolated or autosomal recessive CRD were reviewed after molecular analysis revealed mutations in the ABCA4 gene. In two of the patients both the photopic and scotopic electroretinogram were nonrecordable. In the remainder, the photopic cone b-wave amplitudes appeared to be more seriously affected than the scotopic rod b-wave amplitudes. Although the clinical presentation was heterogeneous, all patients experienced visual loss early in life, impaired color vision, and a central scotoma. Fundoscopy revealed evidence of early-onset maculopathy, sometimes accompanied by involvement of the retinal periphery in the later stages of the disease. Mutations in the ABCA4 gene are the pathologic cause of the CRD-like dystrophy in these patients, and the resultant clinical pictures are complex and heterogeneous. Given this wide clinical spectrum of CRD-like phenotypes associated with ABCA4 mutations, detailed clinical subclassifications are difficult and may not be very useful.

Is There Convincing Evidence that Intermediate Repeats in the HTT Gene Cause Huntington's Disease?

PubMed

Oosterloo, Mayke; Van Belzen, Martine J; Bijlsma, Emilia K; Roos, Raymund A C

2015-01-01

Huntington's disease (HD) is a neurodegenerative disease associated with a CAG repeat expansion in the Huntingtin (HTT) gene. A trinucleotide size between 27 and 35 is considered 'intermediate' and not to cause symptoms and signs of HD. There are articles claiming otherwise, however publishing only the cases that have a HD phenotype introduces a significant publication bias. Our objective is to determine if there is convincing evidence that intermediate repeats (IA) cause HD. Previously published case reports on HTT intermediate repeat sizes and all cases from the Netherlands with an IA were reviewed for clinical symptoms and signs. Four patients had a clinical presentation of Huntington's disease and an IA out of ten reported cases in literature. Between 2001 and 2012, 1,690 patients were tested for HD in the Netherlands. One case out of 60 with an IA had a phenotype resembling HD, but had already been published in a case report. Given the high background frequency of intermediate alleles in several populations, the possibility of developing HD would have huge implications for 1-7% of the normal population. It is possible that IAs present as an endophenotype with the potential of subsequent clinical manifestations. However, given the scarcity of convincing cases, the lack of convincing biological evidence for pathogenicity of intermediate alleles, and many genes still to be discovered for HD mimics, we find that it is premature to claim that IAs can cause HD. We recommend systematic follow up of this group of individuals and if possible brain pathology for confirmation or exclusion of HD.

Pyridoxal 5'-phosphate, pyridoxal, and 4-pyridoxic acid in the paired serum and cerebrospinal fluid of children.

PubMed

Akiyama, Tomoyuki; Hayashi, Yumiko; Hanaoka, Yoshiyuki; Shibata, Takashi; Akiyama, Mari; Tsuchiya, Hiroki; Yamaguchi, Tokito; Kobayashi, Katsuhiro

2017-09-01

We quantified pyridoxal 5'-phosphate (PLP), pyridoxal (PL), and 4-pyridoxic acid (PA) in paired serum and cerebrospinal fluid (CSF) samples from children and investigated the effect of age on the concentrations and CSF-to-serum ratios of these vitamers. Serum and CSF samples prospectively collected from 49 pediatric patients were analyzed. PLP, PL, and PA were measured using high-performance liquid chromatography with fluorescence detection, using pre-column derivatization by semicarbazide. Effects of age on these vitamers, the PLP-to-PL ratio, CSF-to-serum PLP ratio, and CSF-to-serum PL ratio were evaluated using correlation analysis. The PLP, PL, and PA concentrations in the serum and CSF were higher at younger ages, except for CSF PA concentrations that were mostly below the limit of detection (<1.2nmol/l). The PLP-to-PL ratios in the serum and CSF correlated positively with age. The CSF-to-serum PLP ratio and CSF-to-serum PL ratio were independent of age. Age-related changes in PLP, PL, and PA in serum and in CSF from pediatric patients and CSF-to-serum ratios of PLP and PL demonstrated in this study will provide valuable information for evaluating PLP supply to the central nervous system from the peripheral blood. Copyright © 2017 Elsevier B.V. All rights reserved.

Quality and stability of extemporaneous pyridoxal phosphate preparations used in the treatment of paediatric epilepsy.

PubMed

Mohamed-Ahmed, Abeer H A; Wilson, Matthew P; Albuera, Maedelyn; Chen, Ting; Mills, Philippa B; Footitt, Emma J; Clayton, Peter T; Tuleu, Catherine

2017-04-01

To assess the pyridoxal 5'-phosphate (PLP) content and stability of extemporaneous PLP liquids prepared from dietary supplements used for the treatment of vitamin B 6 -dependent epilepsy. Pyridoxal 5'-phosphate liquids were prepared in accordance with the guidelines given to patients from marketed 50 mg PLP dietary capsules and tablets. The PLP content and its stability were evaluated under conditions resembling the clinical setting using reverse phase HPLC and mass spectrometry. Pyridoxal 5'-phosphate content in most of the extemporaneously prepared liquids from dietary supplements was found to be different from the expected amount (~16-60 mg). Most of these PLP extemporaneous liquids were stable at room temperature (protected from light) after 24 h but unstable after 4 h when exposed to light. A key photodegradation product of PLP in water was confirmed as 4-pyridoxic acid 5'-phosphate (PAP). Pyridoxal 5'-phosphate tablets from Solgar ® were found to be the most reliable product for the preparation of extemporaneous PLP liquids. This work highlighted the difference between the marketed PLP dietary supplements quality and the importance of proper storage of aqueous PLP. There is a need to develop pharmaceutical forms of PLP that ensure dose accuracy and avoid potentially unsafe impurities with the aim of enhancing safety and compliance. © 2017 Royal Pharmaceutical Society.

[Maple syrup urine disease caused by two novel BCKDHB gene mutations in a Chinese neonate].

PubMed

Shen, Yunlin; Gong, Xiaohui; Yan, Jingbin; Qin, Li; Qiu, Gang

2015-01-01

Maple syrup urine disease (MSUD) is an autosomal recessive metabolic disorder that is caused by mutations in the subunits of the branched chain α-ketoacid dehydrogenase (BCKD) complex. This report presents a Han ethnic Chinese newborn infant with the severe classic form of MSUD caused by two novel missense mutations in the BCKDHB gene. The clinical and biochemical data of a Chinese neonate with classic form of MSUD were analyzed, and the DNA sequences of BCKDHA, BCKDHB, DBT and DLD genes were investigated for mutations. Then the DNA samples of the proband and the patient's parents were tested with Sanger sequencing. The manifestations of this patient were poor feeding, low reaction, and compensatory metabolic acidosis. Tandem mass spectrometry (MS/MS) showed that leucine and valine were significantly higher than normal. Urine gas chromatography-mass spectrometry (GC/MS) showed significant abnormality. Brain CT scan showed white matter changes. We identified two previously unreported mutations in the BCKDHB gene, p.Leu194Phe (c.580 C>T) and p.Ser199Arg (c.597 T>G) in exon 5. Segregation analysis showed that the novel mutation p.Ser199Arg was maternally inherited and the novel mutation p.Leu194Phe was paternally inherited. Neither mutation was found in the 186 alleles of 93 normal Han ethnic Chinese individuals. In human BCKDHB protein crystal structure, the 194th and 199th amino acids changes are likely to affect the spatial structure of the protein. The 194th and 199th amino acid of human BCKDHB protein was conserved among species. PolyPhen protein function prediction indicated that the 194th and 199th amino acid changes were likely to affect protein function. Two novel missense mutations were identified in the BCKDHB gene in the Chinese patient with MSUD.

Potential for Cell-Mediated Immune Responses in Mouse Models of Pelizaeus-Merzbacher Disease

PubMed Central

Southwood, Cherie M.; Fykkolodziej, Bozena; Dachet, Fabien; Gow, Alexander

2013-01-01

Although activation of the innate and adaptive arms of the immune system are undoubtedly involved in the pathophysiology of neurodegenerative diseases, it is unclear whether immune system activation is a primary or secondary event. Increasingly, published studies link primary metabolic stress to secondary inflammatory responses inside and outside of the nervous system. In this study, we show that the metabolic stress pathway known as the unfolded protein response (UPR) leads to secondary activation of the immune system. First, we observe innate immune system activation in autopsy specimens from Pelizaeus-Merzbacher disease (PMD) patients and mouse models stemming from PLP1 gene mutations. Second, missense mutations in mildly- and severely-affected Plp1-mutant mice exhibit immune-associated expression profiles with greater disease severity causing an increasingly proinflammatory environment. Third, and unexpectedly, we find little evidence for dysregulated expression of major antioxidant pathways, suggesting that the unfolded protein and oxidative stress responses are separable. Together, these data show that UPR activation can precede innate and/or adaptive immune system activation and that neuroinflammation can be titrated by metabolic stress in oligodendrocytes. Whether or not such activation leads to autoimmune disease in humans is unclear, but the case report of steroid-mitigated symptoms in a PMD patient initially diagnosed with multiple sclerosis lends support. PMID:24575297

Novel Partitivirus Enhances Virulence of and Causes Aberrant Gene Expression in Talaromyces marneffei.

PubMed

Lau, Susanna K P; Lo, George C S; Chow, Franklin W N; Fan, Rachel Y Y; Cai, James J; Yuen, Kwok-Yung; Woo, Patrick C Y

2018-06-12

Talaromyces marneffei is the most important thermal dimorphic fungus causing systemic mycosis in Southeast Asia. We report the discovery of a novel partitivirus, Talaromyces marneffei partitivirus -1 (TmPV1). TmPV1 was detected in 7 (12.7%) of 55 clinical T. marneffei isolates. Complete genome sequencing of the seven TmPV1 isolates revealed two double-stranded RNA (dsRNA) segments encoding RNA-dependent RNA polymerase (RdRp) and capsid protein, respectively. Phylogenetic analysis showed that TmPV1 occupied a distinct clade among the members of the genus Gammapartitivirus Transmission electron microscopy confirmed the presence of isometric, nonenveloped viral particles of 30 to 45 nm in diameter, compatible with partitiviruses, in TmPV1-infected T. marneffei Quantitative reverse transcription-PCR (qRT-PCR) demonstrated higher viral load of TmPV1 in the yeast phase than in the mycelial phase of T. marneffei Two virus-free isolates, PM1 and PM41, were successfully infected by purified TmPV1 using protoplast transfection. Mice challenged with TmPV1-infected T. marneffei isolates showed significantly shortened survival time ( P < 0.0001) and higher fungal burden in organs than mice challenged with isogenic TmPV1-free isolates. Transcriptomic analysis showed that TmPV1 causes aberrant expression of various genes in T. marneffei , with upregulation of potential virulence factors and suppression of RNA interference (RNAi)-related genes. This is the first report of a mycovirus in a thermally dimorphic fungus. Further studies are required to ascertain the mechanism whereby TmPV1 enhances the virulence of T. marneffei in mice and the potential role of RNAi-related genes in antiviral defense in T. marneffei IMPORTANCE Talaromyces marneffei (formerly Penicillium marneffei ) is the most important thermal dimorphic fungus in Southeast Asia, causing highly fatal systemic penicilliosis in HIV-infected and immunocompromised patients. We discovered a novel mycovirus, TmPV1

Venezuelan equine encephalitis virus infection causes modulation of inflammatory and immune response genes in mouse brain

PubMed Central

Sharma, Anuj; Bhattacharya, Bhaskar; Puri, Raj K; Maheshwari, Radha K

2008-01-01

Background Neurovirulent Venezuelan equine encephalitis virus (VEEV) causes lethal encephalitis in equines and is transmitted to humans by mosquitoes. VEEV is highly infectious when transmitted by aerosol and has been developed as a bio-warfare agent, making it an important pathogen to study from a military and civilian standpoint. Molecular mechanisms of VEE pathogenesis are poorly understood. To study these, the gene expression profile of VEEV infected mouse brains was investigated. Changes in gene expression were correlated with histological changes in the brain. In addition, a molecular framework of changes in gene expression associated with progression of the disease was studied. Results Our results demonstrate that genes related to important immune pathways such as antigen presentation, inflammation, apoptosis and response to virus (Cxcl10, CxCl11, Ccl5, Ifr7, Ifi27 Oas1b, Fcerg1,Mif, Clusterin and MHC class II) were upregulated as a result of virus infection. The number of over-expressed genes (>1.5-fold level) increased as the disease progressed (from 197, 296, 400, to 1086 at 24, 48, 72 and 96 hours post infection, respectively). Conclusion Identification of differentially expressed genes in brain will help in the understanding of VEEV-induced pathogenesis and selection of biomarkers for diagnosis and targeted therapy of VEEV-induced neurodegeneration. PMID:18558011

Statins cause profound effects on gene expression in human cancer cells in vitro: the role of membrane microdomains.

PubMed

Garnett, David John; Greenhough, Trevor James

2012-01-01

There is increasing evidence that statin treatment can be beneficial in certain cancer patients. To determine if these benefits are a direct result of the cholesterol-lowering effects of statins or a result of secondary, protein transcription effects, the impacts of pravastatin and a cholesterol sequestrating agent methyl-beta-cyclodextrin (MbetaCD) on mRNA expression in the breast cancer cell MDA-MB-231 and the lung carcinoma cell Calu-1 have been compared by microarray techniques. The effects of these agents on cholesterol-rich rafts and caveolae, which have significance in cancer signaling, have also been examined. Both treatments caused a general downregulation of not only signal transduction including cancer pathway proteins, but also apoptosis and chemokine pathways, with statins impacting 35 genes by twofold or greater in MDA-MB-231 and > 300 genes in Calu-1. These manifold dysregulations could also explain the various side effects reportedly caused by statins. MbetaCD produced far fewer statistical events than pravastatin in the breast cancer line but many more in the lung cell line. Pravastatin increased expression of CAV1 but caveolae density decreased and overall raft density was unaffected. MbetaCD also caused an increase in CAV1 expression and reduced the prevalence of both rafts and caveolae. It is proposed that sequestration of cholesterol from the membrane by MbetaCD is not equivalent to blockade of the cholesterol pathway and causes different effects on microdomain-mediated signal transduction dependant on the cell line. The profound effects of statins on mRNA expression can be explained by the failure of caveolin-1 to properly complex with cholesterol in an altered sterol environment, with caveolae acting as the main loci for signaling directed towards those transcription processes unaffected by MbetaCD. Targeted inhibition of the postmevalonate pathway could offer an opportunity to specifically reduce caveolae-based signaling in cancer cells. The

Screening of potential genes contributing to the macrocycle drug resistance of C. albicans via microarray analysis

PubMed Central

Yang, Jing; Zhang, Wei; Sun, Jian; Xi, Zhiqin; Qiao, Zusha; Zhang, Jinyu; Wang, Yan; Ji, Ying; Feng, Wenli

2017-01-01

The aim of the present study was to investigate the potential genes involved in drug resistance of Candida albicans (C. albicans) by performing microarray analysis. The gene expression profile of GSE65396 was downloaded from the Gene Expression Omnibus, including a control, 15-min and 45-min macrocyclic compound RF59-treated group with three repeats for each. Following preprocessing using RAM, the differentially expressed genes (DEGs) were screened using the Limma package. Subsequently, the Kyoto Encyclopedia of Genes and Genomes pathways of these genes were analyzed using the Database for Annotation, Visualization and Integrated Discovery. Based on interactions estimated by the Search Tool for Retrieval of Interacting Gene, the protein-protein interaction (PPI) network was visualized using Cytoscape. Subnetwork analysis was performed using ReactomeFI. A total of 154 upregulated and 27 downregulated DEGs were identified in the 15-min treated group, compared with the control, and 235 upregulated and 233 downregulated DEGs were identified in the 45-min treated group, compared with the control. The upregulated DEGs were significantly enriched in the ribosome pathway. Based on the PPI network, PRP5, RCL1, NOP13, NOP4 and MRT4 were the top five nodes in the 15-min treated comparison. GIS2, URA3, NOP58, ELP3 and PLP7 were the top five nodes in the 45-min treated comparison, and its subnetwork was significantly enriched in the ribosome pathway. The macrocyclic compound RF59 had a notable effect on the ribosome and its associated pathways of C. albicans. RCL1, NOP4, MRT4, GIS2 and NOP58 may be important in RF59-resistance. PMID:28944888

A novel mutation in the AGXT gene causing primary hyperoxaluria type I: genotype-phenotype correlation.

PubMed

M'Dimegh, Saoussen; Aquaviva-Bourdain, Cécile; Omezzine, Asma; M'Barek, Ibtihel; Souche, Geneviéve; Zellama, Dorsaf; Abidi, Kamel; Achour, Abdelattif; Gargah, Tahar; Abroug, Saoussen; Bouslama, Ali

2016-09-01

Primary hyperoxaluria type I (PH1) is an autosomal recessive metabolic disorder caused by inherited mutations in the AGXT gene encoding liver peroxisomal alanine : glyoxylate aminotransferase (AGT) which is deficient or mistargeted to mitochondria. PH1 shows considerable phenotypic and genotypic heterogeneity. The incidence and severity of PH1 varies in different geographic regions. DNA samples of the affected members from two unrelated Tunisian families were tested by amplifying and sequencing each of the AGXT exons and intron-exon junctions. We identified a novel frameshift mutation in the AGXT gene, the c.406_410dupACTGC resulting in a truncated protein (p.Gln137Hisfs*19). It is found in homozygous state in two nonconsanguineous unrelated families from Tunisia. These molecular findings provide genotype/phenotype correlations in the intrafamilial phenotypic and permit accurate carrier detection, and prenatal diagnosis. The novel p.Gln137Hisfs*19 mutation detected in our study extend the spectrum of known AGXT gene mutations in Tunisia.

Gene Therapy

MedlinePlus

... material into the cells' genes. Researchers remove the original disease-causing genes from the viruses, replacing them ... into the body, the viruses may recover their original ability to cause disease. Possibility of causing a ...

Gene switching in Amoeba proteus caused by endosymbiotic bacteria.

PubMed

Jeon, Taeck J; Jeon, Kwang W

2004-02-01

The expression of genes for S-adenosylmethionine synthetase (SAMS), which catalyzes the synthesis of S-adenosylmethionine (AdoMet), a major methyl donor in cells, was studied in symbiont-free (D) and symbiont-bearing (xD) amoeba strains to determine the effect of bacterial endosymbionts. The symbionts suppressed the expression of the gene in host xD amoebae, but amoebae still exhibited about half the enzyme activity found in symbiont-free D amoebae. The study was aimed at elucidating mechanisms of the suppression of the amoeba's gene and determining the alternative source for the gene product. Unexpectedly, we found a second sams (sams2) gene in amoebae, which encoded 390 amino acids. Results of experiments measuring SAMS activities and amounts of AdoMet in D and xD amoebae showed that the half SAMS activity found in xD amoebae came from the amoeba's SAMS2 and not from their endosymbionts. The expression of amoeba sams genes was switched from sams1 to sams2 as a result of infection with X-bacteria, raising the possibility that the switch in the expression of sams genes by bacteria plays a role in the development of symbiosis and the host-pathogen interactions. This is the first report showing such a switch in the expression of host sams genes by infecting bacteria.

A cancer-causing gene is positively correlated with male aggression in Xiphophorus cortezi

PubMed Central

Fernandez, André A.

2010-01-01

The persistence of seemingly maladaptive genes in organisms challenges evolutionary biological thought. In Xiphophorus fishes, certain melanin patterns form malignant melanomas due to a cancer-causing gene (Xiphophorus melanoma receptor kinase; Xmrk), which arose several millions years ago from unequal meiotic recombination. Xiphophorus melanomas are male biased and induced by androgens however male behavior and Xmrk genotype has not been investigated. This study found that male X. cortezi with the spotted caudal (Sc) pattern, from which melanomas originate, displayed increased aggression in mirror image trials. Furthermore, Xmrk males (regardless of Sc phenotype) bit and performed more agonistic displays than Xmrk deficient males. Male aggressive response decreased when males viewed their Sc image as compared to their non-Sc image. Collectively, these results indicate that Xmrk males experience a competitive advantage over wild-type males and that intrasexual selection could be an important component in the evolutionary maintenance of this oncogene within Xiphophorus. PMID:20021547

Not all SCID pigs are created equally: Two independent mutations in the Artemis gene cause Severe Combined Immunodeficiency (SCID) in pigs

PubMed Central

Waide, Emily H; Dekkers, Jack CM; Ross, Jason W; Rowland, Raymond RR; Wyatt, Carol R; Ewen, Catherine L; Evans, Alyssa B; Thekkoot, Dinesh M; Boddicker, Nicholas J; Serão, Nick VL; Ellinwood, N Matthew; Tuggle, Christopher K

2017-01-01

Mutations in over 30 genes are known to result in impairment of the adaptive immune system, causing a group of disorders collectively known as severe combined immunodeficiency (SCID). SCID disorders are split into groups based on their presence and/or functionality of B, T, and NK cells. Piglets from a line of Yorkshire pigs at Iowa State University were shown to be affected by T− B− NK+ SCID, representing the first example of naturally occurring SCID in pigs. Here, we present evidence for two spontaneous mutations as the molecular basis for this SCID phenotype. Flow cytometry analysis of thymocytes showed an increased frequency of immature T cells in SCID pigs. Fibroblasts from these pigs were more sensitive to ionizing radiation than non-SCID piglets, eliminating the RAG1 and RAG2 genes. Genetic and molecular analyses showed two mutations were present in the Artemis gene, which in homozygous or compound heterozygous state cause the immunodeficient phenotype. Rescue of SCID fibroblast radiosensitivity by human Artemis protein demonstrated that the identified Artemis mutations are the direct cause of this cellular phenotype. The work presented here reveals two mutations in the Artemis gene that cause T− B− NK+ SCID in pigs. The SCID pig can be an important biomedical model, but these mutations would be undesirable in commercial pig populations. The identified mutations and associated genetic tests can be used to address both of these issues. PMID:26320255

Do Structural Missense Variants in the ATM Gene Found in Women With Breast Cancer Cause Breast Cancer in Knock-in Mouse Strains?

DTIC Science & Technology

2006-04-01

W81XWH-05-1-0282 TITLE: Do Structural Missense Variants in the ATM Gene Found in Women with Breast Cancer Cause Breast Cancer in "Knock-in...5a. CONTRACT NUMBER Do Structural Missense Variants in the ATM Gene Found in Women with Breast Cancer Cause Breast Cancer in "Knock-in" Mouse...human cohort-specific missense mutations will develop breast cancer with dominant inheritance in a subset of animals. It also is hypothesized that

Human immunodeficiency virus type 1 pol gene mutations which cause decreased susceptibility to 2',3'-dideoxycytidine.

PubMed Central

Fitzgibbon, J E; Howell, R M; Haberzettl, C A; Sperber, S J; Gocke, D J; Dubin, D T

1992-01-01

To investigate whether human immunodeficiency virus type 1 pol gene mutations are selected during prolonged 2',3'-dideoxycytidine (ddC) therapy, we used the polymerase chain reaction to amplify a portion of the reverse transcriptase segment of the pol gene from the peripheral blood mononuclear cell DNA of a patient with AIDS before and after an 80-week course of ddC therapy. The consensus sequence from the second sample contained a unique double mutation (ACT to GAT) in the codon for reverse transcriptase amino acid 69, causing substitution of aspartic acid (Asp) for the wild-type threonine (Thr). A mutation (ACA to ATA) also occurred in the codon for position 165, causing substitution of isoleucine (Ile) for Thr. The GAT (Asp) codon was introduced into the pol gene of a molecular clone of human immunodeficiency virus via site-directed mutagenesis. Following transfection, mutant and wild-type viruses were tested for susceptibility to ddC by a plaque reduction assay. The mutant virus was fivefold less susceptible to ddC than the wild type; cross-resistance to 3'-azido-3'-deoxythymidine or 2'3'-dideoxyinosine was not found. The Ile-165 mutation did not confer additional ddC resistance. The Asp-69 substitution may have contributed to the generation of resistant virus in this patient. Images PMID:1317143

Quantitative HRMAS proton total correlation spectroscopy applied to cultured melanoma cells treated by chloroethyl nitrosourea: demonstration of phospholipid metabolism alterations.

PubMed

Morvan, Daniel; Demidem, Aicha; Papon, Janine; Madelmont, Jean Claude

2003-02-01

Recent NMR spectroscopy developments, such as high-resolution magic angle spinning (HRMAS) probes and correlation-enhanced 2D sequences, now allow improved investigations of phospholipid (Plp) metabolism. Using these modalities we previously demonstrated that a mouse-bearing melanoma tumor responded to chloroethyl nitrosourea (CENU) treatment in vivo by altering its Plp metabolism. The aims of the present study were to investigate whether HRMAS proton total correlation spectroscopy (TOCSY) could be used as a quantitative technique to probe Plp metabolism, and to determine the Plp metabolism response of cultured B16 melanoma cells to CENU treatment in vitro. The exploited TOCSY signals of Plp derivatives arose from scalar coupling among the protons of neighbor methylene groups within base headgroups (choline and ethanolamine). For strongly expressed Plp derivatives, TOCSY signals were compared to saturation recovery signals and demonstrated a linear relationship. HRMAS proton TOCSY was thus used to provide concentrations of Plp derivatives during long-term follow-up of CENU-treated cell cultures. Strong Plp metabolism alteration was observed in treated cultured cells in vitro involving a down-regulation of phosphocholine, and a dramatic and irreversible increase of phosphoethanolamine. These findings are discussed in relation to previous in vivo data, and to Plp metabolism enzymatic involvement. Copyright 2003 Wiley-Liss, Inc.

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.

Applications of vitamin B6 cofactor pyridoxal 5‧-phosphate and pyridoxal 5‧-phosphate crowned gold nanoparticles for optical sensing of metal ions

NASA Astrophysics Data System (ADS)

Bothra, Shilpa; Upadhyay, Yachana; Kumar, Rajender; Sahoo, Suban K.

2017-03-01

Vitamin B6 cofactor pyridoxal 5‧-phosphate (PLP) and PLP crowned gold nanoparticles (PLP-AuNPs) was applied for the optical chemosensing of metal ions in aqueous medium. PLP showed a visually detectable colour change from colourless to yellow and 'turn-off' fluorescence in the presence of Fe3 +. The fluorescence intensity of PLP at 433 nm was also blue-shifted and enhanced at 395 nm upon addition of Al3 +. When the PLP was functionalized over AuNPs surface, the wine red colour of PLP-AuNPs was turned to purplish-blue and the SPR band at 525 nm was red-shifted upon addition of Al3 +, Cd2 + and Pb2 + due to the complexation-induced aggregation of nanoparticles. The developed sensing systems exhibited good selectivity and specificity for the detected analytes (Fe3 +, Al3 +, Cd2 + and Pb2 +).

A conserved 19-kDa Eimeria tenella antigen is a profilin-like protein.

PubMed

Fetterer, R H; Miska, K B; Jenkins, M C; Barfield, R C

2004-12-01

A wide range of recombinant proteins from Eimeria species have been reported to offer some degree of protection against infection and disease, but the specific biological function of these proteins is largely unknown. Previous studies have demonstrated a 19-kDa protein of unknown function designated SZ-1 in sporozoites and merozoites of Eimeria acervulina that can be used to confer partial protection against coccidiosis. Reverse transcriptase-polymerase chain reaction indicated that the gene for SZ-1 is expressed by all the asexual stages of Eimeria tenella. Rabbit antisera to recombinant SZ-1 recognized an approximately 19-kDa protein from extracts of E. tenella sporozoites, merozoites, sporulated oocysts, and oocysts in various stages of sporulation. Immunofluorescence antibody staining indicated specific staining of E. tenella sporozoites and merozoites. Staining was most intense in the cytoplasm of the posterior end of the parasite. The primary amino acid sequence of the gene for E. tenella SZ-1 deduced from the E. tenella genome indicated a conserved domain for the actin-regulatory protein profilin. A conserved binding site for poly-L-proline (PLP), characteristic of profilin was also observed. SZ-1 was separated from soluble extract of E. tenella proteins by affinity chromatography using a PLP ligand, confirming the ability of SZ-1 to bind PLP. SZ-1 also partially inhibited the polymerization of actin. The current results are consistent with the classification of SZ-1 as a profilin-related protein.

Early-Onset Severe Encephalopathy with Epilepsy: The BRAT1 Gene Should Be Added to the List of Causes.

PubMed

van de Pol, Laura A; Wolf, Nicole I; van Weissenbruch, Mirjam M; Stam, Cornelie J; Weiss, Janneke M; Waisfisz, Quinten; Kevelam, Sietske H; Bugiani, Mariana; van de Kamp, Jiddeke M; van der Knaap, Marjo S

2015-12-01

A variety of pathologies can underlie early-onset severe encephalopathy with epilepsy. To aid the diagnostic process in such patients we present an overview of causes, including the rapidly expanding list of genes involved. When no explanation is found, whole-exome sequencing (WES) can be used in an attempt to identify gene defects in patients suspected to suffer from a genetic form. We describe three siblings, born to consanguineous parents, with a lethal severe epileptic encephalopathy with early-infantile onset, including their magnetic resonance imaging, electroencephalography and, in one case, neuropathological findings. Using WES a homozygous frameshift mutation in the BRAT1 gene, c.638dup p.(Val214Glyfs*189), was identified. We present our cases in the context of all published cases with mutations in the BRAT1 gene and conclude that BRAT1 should be added to the growing list of genes related to early-onset severe encephalopathy with epilepsy. Georg Thieme Verlag KG Stuttgart · New York.

Familial Dysalbuminemic Hyperthyroxinemia in a Japanese Man Caused by a Point Albumin Gene Mutation (R218P)

PubMed Central

Osaki, Yoshinori; Hayashi, Yoshitaka; Nakagawa, Yoshinori; Yoshida, Katsumi; Ozaki, Hiroshi; Fukazawa, Hiroshi

2016-01-01

Familial dysalbuminemic hyperthyroxinemia (FDH) is a familial autosomal dominant disease caused by mutation in the albumin gene that produces a condition of euthyroid hyperthyroxinemia. In patients with FDH, serum-free thyroxine (FT4) and free triiodothyronine (FT3) concentrations as measured by several commercial methods are often falsely increased with normal thyrotropin (TSH). Therefore, several diagnostic steps are needed to differentiate TSH-secreting tumor or generalized resistance to thyroid hormone from FDH. We herein report a case of a Japanese man born in Aomori prefecture, with FDH caused by a mutant albumin gene (R218P). We found that a large number of FDH patients reported in Japan to date might have been born in Aomori prefecture and have shown the R218P mutation. In conclusion, FDH needs to be considered among the differential diagnoses in Japanese patients born in Aomori prefecture and showing normal TSH levels and elevated FT4 levels. PMID:27081329

XLID-Causing Mutations and Associated Genes Challenged in Light of Data From Large-Scale Human Exome Sequencing

PubMed Central

Piton, Amélie; Redin, Claire; Mandel, Jean-Louis

2013-01-01

Because of the unbalanced sex ratio (1.3–1.4 to 1) observed in intellectual disability (ID) and the identification of large ID-affected families showing X-linked segregation, much attention has been focused on the genetics of X-linked ID (XLID). Mutations causing monogenic XLID have now been reported in over 100 genes, most of which are commonly included in XLID diagnostic gene panels. Nonetheless, the boundary between true mutations and rare non-disease-causing variants often remains elusive. The sequencing of a large number of control X chromosomes, required for avoiding false-positive results, was not systematically possible in the past. Such information is now available thanks to large-scale sequencing projects such as the National Heart, Lung, and Blood (NHLBI) Exome Sequencing Project, which provides variation information on 10,563 X chromosomes from the general population. We used this NHLBI cohort to systematically reassess the implication of 106 genes proposed to be involved in monogenic forms of XLID. We particularly question the implication in XLID of ten of them (AGTR2, MAGT1, ZNF674, SRPX2, ATP6AP2, ARHGEF6, NXF5, ZCCHC12, ZNF41, and ZNF81), in which truncating variants or previously published mutations are observed at a relatively high frequency within this cohort. We also highlight 15 other genes (CCDC22, CLIC2, CNKSR2, FRMPD4, HCFC1, IGBP1, KIAA2022, KLF8, MAOA, NAA10, NLGN3, RPL10, SHROOM4, ZDHHC15, and ZNF261) for which replication studies are warranted. We propose that similar reassessment of reported mutations (and genes) with the use of data from large-scale human exome sequencing would be relevant for a wide range of other genetic diseases. PMID:23871722

XLID-causing mutations and associated genes challenged in light of data from large-scale human exome sequencing.

PubMed

Piton, Amélie; Redin, Claire; Mandel, Jean-Louis

2013-08-08

Because of the unbalanced sex ratio (1.3-1.4 to 1) observed in intellectual disability (ID) and the identification of large ID-affected families showing X-linked segregation, much attention has been focused on the genetics of X-linked ID (XLID). Mutations causing monogenic XLID have now been reported in over 100 genes, most of which are commonly included in XLID diagnostic gene panels. Nonetheless, the boundary between true mutations and rare non-disease-causing variants often remains elusive. The sequencing of a large number of control X chromosomes, required for avoiding false-positive results, was not systematically possible in the past. Such information is now available thanks to large-scale sequencing projects such as the National Heart, Lung, and Blood (NHLBI) Exome Sequencing Project, which provides variation information on 10,563 X chromosomes from the general population. We used this NHLBI cohort to systematically reassess the implication of 106 genes proposed to be involved in monogenic forms of XLID. We particularly question the implication in XLID of ten of them (AGTR2, MAGT1, ZNF674, SRPX2, ATP6AP2, ARHGEF6, NXF5, ZCCHC12, ZNF41, and ZNF81), in which truncating variants or previously published mutations are observed at a relatively high frequency within this cohort. We also highlight 15 other genes (CCDC22, CLIC2, CNKSR2, FRMPD4, HCFC1, IGBP1, KIAA2022, KLF8, MAOA, NAA10, NLGN3, RPL10, SHROOM4, ZDHHC15, and ZNF261) for which replication studies are warranted. We propose that similar reassessment of reported mutations (and genes) with the use of data from large-scale human exome sequencing would be relevant for a wide range of other genetic diseases. Copyright © 2013 The American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.

Lysine Decarboxylase with an Enhanced Affinity for Pyridoxal 5-Phosphate by Disulfide Bond-Mediated Spatial Reconstitution.

PubMed

Sagong, Hye-Young; Kim, Kyung-Jin

2017-01-01

Lysine decarboxylase (LDC) catalyzes the decarboxylation of l-lysine to produce cadaverine, an important industrial platform chemical for bio-based polyamides. However, due to high flexibility at the pyridoxal 5-phosphate (PLP) binding site, use of the enzyme for cadaverine production requires continuous supplement of large amounts of PLP. In order to develop an LDC enzyme from Selenomonas ruminantium (SrLDC) with an enhanced affinity for PLP, we introduced an internal disulfide bond between Ala225 and Thr302 residues with a desire to retain the PLP binding site in a closed conformation. The SrLDCA225C/T302C mutant showed a yellow color and the characteristic UV/Vis absorption peaks for enzymes with bound PLP, and exhibited three-fold enhanced PLP affinity compared with the wild-type SrLDC. The mutant also exhibited a dramatically enhanced LDC activity and cadaverine conversion particularly under no or low PLP concentrations. Moreover, introduction of the disulfide bond rendered SrLDC more resistant to high pH and temperature. The formation of the introduced disulfide bond and the maintenance of the PLP binding site in the closed conformation were confirmed by determination of the crystal structure of the mutant. This study shows that disulfide bond-mediated spatial reconstitution can be a platform technology for development of enzymes with enhanced PLP affinity.

MASA syndrome is caused by mutations in the neural cell adhesion gene, L1CAM

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

Schwartz, C.E.; Wang, Y.; Schroer, R.J.

1994-09-01

The MASA syndrome is a recessive X-linked disorder characterized by Mental retardation, Adducted thumbs, Shuffling gait and Aphasia. Recently we found that MASA in one family was likely caused by a point mutation in exon 6 of the L1CAM gene. This gene has also been shown to be involved in X-linked hydrocephalus (HSAS). We have screened 60 patients with either sporadic HSAS or MASA as well as two additional families with MASA. For the screening, we initially utilized 3 cDNA probes for the L1CAM gene. In one of the MASA families, K8310, two affected males were found to have anmore » altered BglII band. The band was present in their carrier mother but not in their normal brothers. This band was detected by the entire cDNA probe as well as the cDNA probe for 3{prime} end of the gene. Analysis of the L1CAM sequence indicated the altered BglII site is distal to the exon 28 but proximal to the punative poly A signal site. It is hypothesized that this point mutation alters the stability of the L1CAM mRNA. This is being tested using cell lines established from the two affected males.« less

Continuous variation caused by genes with graduated effects.

PubMed Central

Matthysse, S; Lange, K; Wagener, D K

1979-01-01

The classical polygenic theory of inheritance postulates a large number of genes with small, and essentially similar, effects. We propose instead a model with genes of gradually decreasing effects. The resulting phenotypic distribution is not normal; if the gene effects are geometrically decreasing, it can be triangular. The joint distribution of parent and offspring genic value is calculated. The most readily testable difference between the two models is that, in the decreasing-effect model, the variance of the offspring distribution from given parents depends on the parents' genic values. The more the parents deviate from the mean, the smaller the variance of the offspring should be. In the equal-effect model the offspring variance is independent of the parents' genic values. PMID:288073

Deletion of ETS-1, a gene in the Jacobsen syndrome critical region, causes ventricular septal defects and abnormal ventricular morphology in mice

PubMed Central

Ye, Maoqing; Coldren, Chris; Liang, Xingqun; Mattina, Teresa; Goldmuntz, Elizabeth; Benson, D. Woodrow; Ivy, Dunbar; Perryman, M.B.; Garrett-Sinha, Lee Ann; Grossfeld, Paul

2010-01-01

Congenital heart defects comprise the most common form of major birth defects, affecting 0.7% of all newborn infants. Jacobsen syndrome (11q-) is a rare chromosomal disorder caused by deletions in distal 11q. We have previously determined that a wide spectrum of the most common congenital heart defects occur in 11q-, including an unprecedented high frequency of hypoplastic left heart syndrome (HLHS). We identified an ∼7 Mb ‘cardiac critical region’ in distal 11q that contains a putative causative gene(s) for congenital heart disease. In this study, we utilized chromosomal microarray mapping to characterize three patients with 11q- and congenital heart defects that carry interstitial deletions overlapping the 7 Mb cardiac critical region. We propose that this 1.2 Mb region of overlap harbors a gene(s) that causes at least a subset of the congenital heart defects that occur in 11q-. We demonstrate that one gene in this region, ETS-1 (a member of the ETS family of transcription factors), is expressed in the endocardium and neural crest during early mouse heart development. Gene-targeted deletion of ETS-1 in mice in a C57/B6 background causes, with high penetrance, large membranous ventricular septal defects and a bifid cardiac apex, and less frequently a non-apex-forming left ventricle (one of the hallmarks of HLHS). Our results implicate an important role for the ETS-1 transcription factor in mammalian heart development and should provide important insights into some of the most common forms of congenital heart disease. PMID:19942620

Duplication in the Microtubule-Actin Cross-linking Factor 1 gene causes a novel neuromuscular condition

PubMed Central

Jørgensen, Louise H.; Mosbech, Mai-Britt; Færgeman, Nils J.; Graakjaer, Jesper; Jacobsen, Søren V.; Schrøder, Henrik D.

2014-01-01

Spectrins and plakins are important communicators linking cytoskeletal components to each other and to cellular junctions. Microtubule-actin cross-linking factor 1 (MACF1) belongs to the spectraplakin family and is involved in control of microtubule dynamics. Complete knock out of MACF1 in mice is associated with developmental retardation and embryonic lethality. Here we present a family with a novel neuromuscular condition. Genetic analyses show a heterozygous duplication resulting in reduced MACF1 gene product. The functional consequence is affected motility observed as periodic hypotonia, lax muscles and diminished motor skills, with heterogeneous presentation among the affected family members. To corroborate these findings we used RNA interference to knock down the VAB-10 locus containing the MACF1 homologue in C. elegans, and we could show that this also causes movement disturbances. These findings suggest that changes in the MACF1 gene is implicated in this neuromuscular condition, which is an important observation since MACF1 has not previously been associated with any human disease and thus presents a key to understanding the essential nature of this gene. PMID:24899269

Duplication in the microtubule-actin cross-linking factor 1 gene causes a novel neuromuscular condition.

PubMed

Jørgensen, Louise H; Mosbech, Mai-Britt; Færgeman, Nils J; Graakjaer, Jesper; Jacobsen, Søren V; Schrøder, Henrik D

2014-06-05

Spectrins and plakins are important communicators linking cytoskeletal components to each other and to cellular junctions. Microtubule-actin cross-linking factor 1 (MACF1) belongs to the spectraplakin family and is involved in control of microtubule dynamics. Complete knock out of MACF1 in mice is associated with developmental retardation and embryonic lethality. Here we present a family with a novel neuromuscular condition. Genetic analyses show a heterozygous duplication resulting in reduced MACF1 gene product. The functional consequence is affected motility observed as periodic hypotonia, lax muscles and diminished motor skills, with heterogeneous presentation among the affected family members. To corroborate these findings we used RNA interference to knock down the VAB-10 locus containing the MACF1 homologue in C. elegans, and we could show that this also causes movement disturbances. These findings suggest that changes in the MACF1 gene is implicated in this neuromuscular condition, which is an important observation since MACF1 has not previously been associated with any human disease and thus presents a key to understanding the essential nature of this gene.

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.

Somatic USP8 Gene Mutations Are a Common Cause of Pediatric Cushing Disease.

PubMed

Faucz, Fabio R; Tirosh, Amit; Tatsi, Christina; Berthon, Annabel; Hernández-Ramírez, Laura C; Settas, Nikolaos; Angelousi, Anna; Correa, Ricardo; Papadakis, Georgios Z; Chittiboina, Prashant; Quezado, Martha; Pankratz, Nathan; Lane, John; Dimopoulos, Aggeliki; Mills, James L; Lodish, Maya; Stratakis, Constantine A

2017-08-01

Somatic mutations in the ubiquitin-specific protease 8 (USP8) gene have been recently identified as the most common genetic alteration in patients with Cushing disease (CD). However, the frequency of these mutations in the pediatric population has not been extensively assessed. We investigated the status of the USP8 gene at the somatic level in a cohort of pediatric patients with corticotroph adenomas. The USP8 gene was fully sequenced in both germline and tumor DNA samples from 42 pediatric patients with CD. Clinical, biochemical, and imaging data were compared between patients with and without somatic USP8 mutations. Five different USP8 mutations (three missense, one frameshift, and one in-frame deletion) were identified in 13 patients (31%), all of them located in exon 14 at the previously described mutational hotspot, affecting the 14-3-3 binding motif of the protein. Patients with somatic mutations were older at disease presentation [mean 5.1 ± 2.1 standard deviation (SD) vs 13.1 ± 3.6 years, P = 0.03]. Levels of urinary free cortisol, midnight serum cortisol, and adrenocorticotropic hormone, as well as tumor size and frequency of invasion of the cavernous sinus, were not significantly different between the two groups. However, patients harboring somatic USP8 mutations had a higher likelihood of recurrence compared with patients without mutations (46.2% vs 10.3%, P = 0.009). Somatic USP8 gene mutations are a common cause of pediatric CD. Patients harboring a somatic mutation had a higher likelihood of tumor recurrence, highlighting the potential importance of this molecular defect for the disease prognosis and the development of targeted therapeutic options. Copyright © 2017 Endocrine Society

DUX4, a candidate gene for facioscapulohumeral muscular dystrophy, causes p53-dependent myopathy in vivo.

PubMed

Wallace, Lindsay M; Garwick, Sara E; Mei, Wenyan; Belayew, Alexandra; Coppee, Frederique; Ladner, Katherine J; Guttridge, Denis; Yang, Jing; Harper, Scott Q

2011-03-01

Facioscapulohumeral muscular dystrophy (FSHD) is associated with D4Z4 repeat contraction on human chromosome 4q35. This genetic lesion does not result in complete loss or mutation of any gene. Consequently, the pathogenic mechanisms underlying FSHD have been difficult to discern. In leading FSHD pathogenesis models, D4Z4 contractions are proposed to cause epigenetic changes, which ultimately increase expression of genes with myopathic potential. Although no gene has been conclusively linked to FSHD development, recent evidence supports a role for the D4Z4-encoded DUX4 gene in FSHD. In this study, our objective was to test the in vivo myopathic potential of DUX4. We delivered DUX4 to zebrafish and mouse muscle by transposon-mediated transgenesis and adeno-associated viral vectors, respectively. Overexpression of DUX4, which encodes a transcription factor, caused abnormalities associated with muscular dystrophy in zebrafish and mice. This toxicity required DNA binding, because a DUX4 DNA binding domain mutant produced no abnormalities. Importantly, we found the myopathic effects of DUX4 were p53 dependent, as p53 inhibition mitigated DUX4 toxicity in vitro, and muscles from p53 null mice were resistant to DUX4-induced damage. Our work demonstrates the myopathic potential of DUX4 in animal muscle. Considering previous studies showed DUX4 was elevated in FSHD patient muscles, our data support the hypothesis that DUX4 overexpression contributes to FSHD development. Moreover, we provide a p53-dependent mechanism for DUX4 toxicity that is consistent with previous studies showing p53 pathway activation in FSHD muscles. Our work justifies further investigation of DUX4 and the p53 pathway in FSHD pathogenesis. Copyright © 2010 American Neurological Association.

DUX4, a Candidate Gene for Facioscapulohumeral Muscular Dystrophy, Causes p53-Dependent Myopathy In Vivo

PubMed Central

Wallace, Lindsay M.; Garwick, Sara E.; Mei, Wenyan; Belayew, Alexandra; Coppee, Frederique; Ladner, Katherine J.; Guttridge, Denis; Yang, Jing; Harper, Scott Q.

2014-01-01

Objective Facioscapulohumeral muscular dystrophy (FSHD) is associated with D4Z4 repeat contraction on human chromosome 4q35. This genetic lesion does not result in complete loss or mutation of any gene. Consequently, the pathogenic mechanisms underlying FSHD have been difficult to discern. In leading FSHD pathogenesis models, D4Z4 contractions are proposed to cause epigenetic changes, which ultimately increase expression of genes with myopathic potential. Although no gene has been conclusively linked to FSHD development, recent evidence supports a role for the D4Z4-encoded DUX4 gene in FSHD. In this study, our objective was to test the in vivo myopathic potential of DUX4. Methods We delivered DUX4 to zebrafish and mouse muscle by transposon-mediated transgenesis and adeno-associated viral vectors, respectively. Results Overexpression of DUX4, which encodes a transcription factor, caused abnormalities associated with muscular dystrophy in zebrafish and mice. This toxicity required DNA binding, because a DUX4 DNA binding domain mutant produced no abnormalities. Importantly, we found the myopathic effects of DUX4 were p53 dependent, as p53 inhibition mitigated DUX4 toxicity in vitro, and muscles from p53 null mice were resistant to DUX4-induced damage. Interpretation Our work demonstrates the myopathic potential of DUX4 in animal muscle. Considering previous studies showed DUX4 was elevated in FSHD patient muscles, our data support the hypothesis that DUX4 overexpression contributes to FSHD development. Moreover, we provide a p53-dependent mechanism for DUX4 toxicity that is consistent with previous studies showing p53 pathway activation in FSHD muscles. Our work justifies further investigation of DUX4 and the p53 pathway in FSHD pathogenesis. PMID:21446026

An innovative strategy to clone positive modifier genes of defects caused by mtDNA mutations: MRPS18C as suppressor gene of m.3946G>A mutation in MT-ND1 gene.

PubMed

Rodríguez-García, María Elena; Cotrina-Vinagre, Francisco Javier; Carnicero-Rodríguez, Patricia; Martínez-Azorín, Francisco

2017-07-01

We have developed a new functional complementation approach to clone modifier genes which overexpression is able to suppress the biochemical defects caused by mtDNA mutations (suppressor genes). This strategy consists in transferring human genes into respiratory chain-deficient fibroblasts, followed by a metabolic selection in a highly selective medium. We used a normalized expression cDNA library in an episomal vector (pREP4) to transfect the fibroblasts, and a medium with glutamine and devoid of any carbohydrate source to select metabolically. Growing the patient's fibroblasts in this selective medium, the deficient cells rapidly disappear unless they are rescued by the cDNA of a suppressor gene. The use of an episomal vector allows us to carry out several rounds of transfection/selection (cyclical phenotypic rescue) to enrich the rescue with true clones of suppressor genes. Using fibroblasts from a patient with epileptic encephalopathy with the m.3946G>A (p.E214K) mutation in the MT-ND1 gene, several candidate genes were identified and one of them was characterized functionally. Thus, overexpression of MRPS18C gene (that encode for bS18m protein) suppressed the molecular defects produced by this mtDNA mutation, recovering the complex I activity and reducing the ROS produced by this complex to normal levels. We suggest that modulation of bS18m expression may be an effective therapeutic strategy for the patients with this mutation.

North Carolina macular dystrophy (MCDR1) caused by a novel tandem duplication of the PRDM13 gene

PubMed Central

Sullivan, Lori S.; Wheaton, Dianna K.; Locke, Kirsten G.; Jones, Kaylie D.; Koboldt, Daniel C.; Fulton, Robert S.; Wilson, Richard K.; Blanton, Susan H.; Birch, David G.; Daiger, Stephen P.

2016-01-01

Purpose To identify the underlying cause of disease in a large family with North Carolina macular dystrophy (NCMD). Methods A large four-generation family (RFS355) with an autosomal dominant form of NCMD was ascertained. Family members underwent comprehensive visual function evaluations. Blood or saliva from six affected family members and three unaffected spouses was collected and DNA tested for linkage to the MCDR1 locus on chromosome 6q12. Three affected family members and two unaffected spouses underwent whole exome sequencing (WES) and subsequently, custom capture of the linkage region followed by next-generation sequencing (NGS). Standard PCR and dideoxy sequencing were used to further characterize the mutation. Results Of the 12 eyes examined in six affected individuals, all but two had Gass grade 3 macular degeneration features. Large central excavation of the retinal and choroid layers, referred to as a macular caldera, was seen in an age-independent manner in the grade 3 eyes. The calderas are unique to affected individuals with MCDR1. Genome-wide linkage mapping and haplotype analysis of markers from the chromosome 6q region were consistent with linkage to the MCDR1 locus. Whole exome sequencing and custom-capture NGS failed to reveal any rare coding variants segregating with the phenotype. Analysis of the custom-capture NGS sequencing data for copy number variants uncovered a tandem duplication of approximately 60 kb on chromosome 6q. This region contains two genes, CCNC and PRDM13. The duplication creates a partial copy of CCNC and a complete copy of PRDM13. The duplication was found in all affected members of the family and is not present in any unaffected members. The duplication was not seen in 200 ethnically matched normal chromosomes. Conclusions The cause of disease in the original family with MCDR1 and several others has been recently reported to be dysregulation of the PRDM13 gene, caused by either single base substitutions in a DNase 1

North Carolina macular dystrophy (MCDR1) caused by a novel tandem duplication of the PRDM13 gene.

PubMed

Bowne, Sara J; Sullivan, Lori S; Wheaton, Dianna K; Locke, Kirsten G; Jones, Kaylie D; Koboldt, Daniel C; Fulton, Robert S; Wilson, Richard K; Blanton, Susan H; Birch, David G; Daiger, Stephen P

2016-01-01

To identify the underlying cause of disease in a large family with North Carolina macular dystrophy (NCMD). A large four-generation family (RFS355) with an autosomal dominant form of NCMD was ascertained. Family members underwent comprehensive visual function evaluations. Blood or saliva from six affected family members and three unaffected spouses was collected and DNA tested for linkage to the MCDR1 locus on chromosome 6q12. Three affected family members and two unaffected spouses underwent whole exome sequencing (WES) and subsequently, custom capture of the linkage region followed by next-generation sequencing (NGS). Standard PCR and dideoxy sequencing were used to further characterize the mutation. Of the 12 eyes examined in six affected individuals, all but two had Gass grade 3 macular degeneration features. Large central excavation of the retinal and choroid layers, referred to as a macular caldera, was seen in an age-independent manner in the grade 3 eyes. The calderas are unique to affected individuals with MCDR1. Genome-wide linkage mapping and haplotype analysis of markers from the chromosome 6q region were consistent with linkage to the MCDR1 locus. Whole exome sequencing and custom-capture NGS failed to reveal any rare coding variants segregating with the phenotype. Analysis of the custom-capture NGS sequencing data for copy number variants uncovered a tandem duplication of approximately 60 kb on chromosome 6q. This region contains two genes, CCNC and PRDM13 . The duplication creates a partial copy of CCNC and a complete copy of PRDM13 . The duplication was found in all affected members of the family and is not present in any unaffected members. The duplication was not seen in 200 ethnically matched normal chromosomes. The cause of disease in the original family with MCDR1 and several others has been recently reported to be dysregulation of the PRDM13 gene, caused by either single base substitutions in a DNase 1 hypersensitive site upstream of the CCNC

Telomeric repeat-containing RNA/G-quadruplex-forming sequences cause genome-wide alteration of gene expression in human cancer cells in vivo.

PubMed

Hirashima, Kyotaro; Seimiya, Hiroyuki

2015-02-27

Telomere erosion causes cell mortality, suggesting that longer telomeres enable more cell divisions. In telomerase-positive human cancer cells, however, telomeres are often kept shorter than those of surrounding normal tissues. Recently, we showed that cancer cell telomere elongation represses innate immune genes and promotes their differentiation in vivo. This implies that short telomeres contribute to cancer malignancy, but it is unclear how such genetic repression is caused by elongated telomeres. Here, we report that telomeric repeat-containing RNA (TERRA) induces a genome-wide alteration of gene expression in telomere-elongated cancer cells. Using three different cell lines, we found that telomere elongation up-regulates TERRA signal and down-regulates innate immune genes such as STAT1, ISG15 and OAS3 in vivo. Ectopic TERRA oligonucleotides repressed these genes even in cells with short telomeres under three-dimensional culture conditions. This appeared to occur from the action of G-quadruplexes (G4) in TERRA, because control oligonucleotides had no effect and a nontelomeric G4-forming oligonucleotide phenocopied the TERRA oligonucleotide. Telomere elongation and G4-forming oligonucleotides showed similar gene expression signatures. Most of the commonly suppressed genes were involved in the innate immune system and were up-regulated in various cancers. We propose that TERRA G4 counteracts cancer malignancy by suppressing innate immune genes. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

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.

NMR studies of the protonation states of pyridoxal-5‧-phosphate in water

NASA Astrophysics Data System (ADS)

Chan-Huot, Monique; Niether, Christiane; Sharif, Shasad; Tolstoy, Peter M.; Toney, Michael D.; Limbach, Hans-Heinrich

2010-07-01

We have measured the 13C NMR spectra of the cofactor pyridoxal-5'-phosphate (vitamin B 6, PLP) at 278 K in aqueous solution as a function of pH. By 13C enrichment of PLP in the C-4' and C-5' positions we were able to measure spectra down to pH 1. From the dependence of the 13C chemical shifts on pH, the p Ka values of PLP could be determined. In particular, the heretofore uncharacterized protonation state of PLP, in which the phosphate group as well as the pyridine ring and the phenolic groups are fully protonated, has been analyzed. The corresponding p Ka value of 2.4 indicates that the phosphate group is solely involved in the first deprotonation step. The 15N chemical shifts of the pyridine ring of PLP published previously are in good agreement with the new results. These shifts contain information about the tautomerism of the different protonation states of PLP. The implications of these findings for the biological function of PLP are discussed.

A large deletion of the AVPR2 gene causing severe nephrogenic diabetes insipidus in a Turkish family.

PubMed

Saglar, Emel; Deniz, Ferhat; Erdem, Beril; Karaduman, Tugce; Yönem, Arif; Cagiltay, Eylem; Mergen, Hatice

2014-05-01

X-linked nephrogenic diabetes insipidus (NDI) is a rare hereditary disease caused by mutations in arginine vasopressin type 2 receptor (AVPR2) and characterized by the production of large amounts of urine and an inability to concentrate urine in response to the antidiuretic hormone vasopressin. We have identified a novel 388 bp deletion starting in intron 1 and ending in exon 2 in the AVPR2 gene in a patient with NDI and in his family. We have revealed that this mutation is a de novo mutation for the mother of the proband patient. Prospective clinical data were collected for all family members. The water deprivation test confirmed the diagnosis of diabetes insipidus. The patient has severe symptoms like deep polyuria nocturia, polydipsia, and fatigue. He was given arginine vasopressin treatment while he was a child. However, he could not get well due to his nephrogenic type of illness. Both of his nephews have the same complains in addition to failure to grow. We have sequenced all exons and intron-exon boundaries of the AVPR2 gene of all family members. The analyses of bioinformatics and comparative genomics of the deletion were done via considering the DNA level damage. AVPR2 gene mutation results in the absence of the three transmembrane domains, two extracellular domains, and one cytoplasmic domain. Three-dimensional protein structure prediction was shown. We concluded that X-linked NDI and severity of illness in this family is caused by a novel 388 bp deletion in the AVPR2 gene that is predicted to truncate the receptor protein, and also this deletion may lead to dysfunctioning in protein activity and inefficient or inadequate binding abilities.

Brief review on pulse laser propulsion

NASA Astrophysics Data System (ADS)

Yu, Haichao; Li, Hanyang; Wang, Yan; Cui, Lugui; Liu, Shuangqiang; Yang, Jun

2018-03-01

Pulse laser propulsion (PLP) is an advanced propulsion concept can be used across a variety of fields with a wide range of applications. PLP reflects superior payload as well as decreased launch costs in comparison with other conventional methods of producing thrust, such as chemical propulsion or electric propulsion. Numerous researchers have attempted to exploit the potential applications of PLP. This paper first reviews concepts relevant to PLP, including the propulsion modes, breakdown regimes, and propulsion efficiency; the propulsion targets for different materials with the pulse laser are then discussed in detail, including the propulsion of solid and liquid microspheres. PLP applications such as the driven microsatellite, target surface particle removal, and orbital debris removal are also discussed. Although the PLP has been applied to a variety of fields, further research is yet warranted to establish its application in the aerospace field.

Familial retinoblastoma due to intronic LINE-1 insertion causes aberrant and noncanonical mRNA splicing of the RB1 gene.

PubMed

Rodríguez-Martín, Carlos; Cidre, Florencia; Fernández-Teijeiro, Ana; Gómez-Mariano, Gema; de la Vega, Leticia; Ramos, Patricia; Zaballos, Ángel; Monzón, Sara; Alonso, Javier

2016-05-01

Retinoblastoma (RB, MIM 180200) is the paradigm of hereditary cancer. Individuals harboring a constitutional mutation in one allele of the RB1 gene have a high predisposition to develop RB. Here, we present the first case of familial RB caused by a de novo insertion of a full-length long interspersed element-1 (LINE-1) into intron 14 of the RB1 gene that caused a highly heterogeneous splicing pattern of RB1 mRNA. LINE-1 insertion was inferred by mRNA studies and full-length sequenced by massive parallel sequencing. Some of the aberrant mRNAs were produced by noncanonical acceptor splice sites, a new finding that up to date has not been described to occur upon LINE-1 retrotransposition. Our results clearly show that RNA-based strategies have the potential to detect disease-causing transposon insertions. It also confirms that the incorporation of new genetic approaches, such as massive parallel sequencing, contributes to characterize at the sequence level these unique and exceptional genetic alterations.

Mutation of the Erwinia amylovora argD Gene Causes Arginine Auxotrophy, Nonpathogenicity in Apples, and Reduced Virulence in Pears

PubMed Central

Ramos, Laura S.; Lehman, Brian L.; Peter, Kari A.

2014-01-01

Fire blight is caused by Erwinia amylovora and is the most destructive bacterial disease of apples and pears worldwide. In this study, we found that E. amylovora argD(1000)::Tn5, an argD Tn5 transposon mutant that has the Tn5 transposon inserted after nucleotide 999 in the argD gene-coding region, was an arginine auxotroph that did not cause fire blight in apple and had reduced virulence in immature pear fruits. The E. amylovora argD gene encodes a predicted N-acetylornithine aminotransferase enzyme, which is involved in the production of the amino acid arginine. A plasmid-borne copy of the wild-type argD gene complemented both the nonpathogenic and the arginine auxotrophic phenotypes of the argD(1000)::Tn5 mutant. However, even when mixed with virulent E. amylovora cells and inoculated onto immature apple fruit, the argD(1000)::Tn5 mutant still failed to grow, while the virulent strain grew and caused disease. Furthermore, the pCR2.1-argD complementation plasmid was stably maintained in the argD(1000)::Tn5 mutant growing in host tissues without any antibiotic selection. Therefore, the pCR2.1-argD complementation plasmid could be useful for the expression of genes, markers, and reporters in E. amylovora growing in planta, without concern about losing the plasmid over time. The ArgD protein cannot be considered an E. amylovora virulence factor because the argD(1000)::Tn5 mutant was auxotrophic and had a primary metabolism defect. Nevertheless, these results are informative about the parasitic nature of the fire blight disease interaction, since they indicate that E. amylovora cannot obtain sufficient arginine from apple and pear fruit tissues or from apple vegetative tissues, either at the beginning of the infection process or after the infection has progressed to an advanced state. PMID:25172854

Induced pluripotent stem cells derived from a patient with autosomal dominant familial neurohypophyseal diabetes insipidus caused by a variant in the AVP gene.

PubMed

Toustrup, Lise Bols; Zhou, Yan; Kvistgaard, Helene; Gregersen, Niels; Rittig, Søren; Aagaard, Lars; Corydon, Thomas Juhl; Luo, Yonglun; Christensen, Jane H

2017-03-01

Autosomal dominant familial neurohypophyseal diabetes insipidus (adFNDI) is caused by variants in the arginine vasopressin (AVP) gene. Here we report the generation of induced pluripotent stem cells (iPSCs) from a 42-year-old man carrying an adFNDI causing variant in exon 1 of the AVP gene using lentivirus-mediated nuclear reprogramming. The iPSCs carried the expected variant in the AVP gene. Furthermore, the iPSCs expressed pluripotency markers; displayed in vitro differentiation potential to the three germ layers and had a normal karyotype consistent with the original fibroblasts. This iPSC line is useful in future studies focusing on the pathogenesis of adFNDI. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

Conditional ablation of the choroideremia gene causes age-related changes in mouse retinal pigment epithelium.

PubMed

Wavre-Shapton, Silène T; Tolmachova, Tanya; Lopes da Silva, Mafalda; da Silva, Mafalda Lopes; Futter, Clare E; Seabra, Miguel C

2013-01-01

The retinal pigment epithelium (RPE) is a pigmented monolayer of cells lying between the photoreceptors and a layer of fenestrated capillaries, the choriocapillaris. Choroideremia (CHM) is an X-linked progressive degeneration of these three layers caused by the loss of function of Rab Escort protein-1 (REP1). REP1 is involved in the prenylation of Rab proteins, key regulators of membrane trafficking. To study the pathological consequences of chronic disruption of membrane traffic in the RPE we used a cell type-specific knock-out mouse model of the disease, where the Chm/Rep1 gene is deleted only in pigmented cells (Chm(Flox), Tyr-Cre+). Transmission electron microscopy (TEM) was used to quantitate the melanosome distribution in the RPE and immunofluorescent staining of rhodopsin was used to quantitate phagocytosed rod outer segments in retinal sections. The ultrastructure of the RPE and Bruch's membrane at different ages was characterised by TEM to analyse age-related changes occurring as a result of defects in membrane traffic pathways. Chm/Rep1 gene knockout in RPE cells resulted in reduced numbers of melanosomes in the apical processes and delayed phagosome degradation. In addition, the RPE accumulated pathological changes at 5-6 months of age similar to those observed in 2-year old controls. These included the intracellular accumulation of lipofuscin-containing deposits, disorganised basal infoldings and the extracellular accumulation of basal laminar and basal linear deposits. The phenotype of the Chm(Flox), Tyr-Cre+ mice suggests that loss of the Chm/Rep1 gene causes premature accumulation of features of aging in the RPE. Furthermore, the striking similarities between the present observations and some of the phenotypes reported in age-related macular degeneration (AMD) suggest that membrane traffic defects may contribute to the pathogenesis of AMD.

Prediagnostic plasma vitamin B6 (pyridoxal 50-phosphate) and survival in patients with colorectal cancer

USDA-ARS?s Scientific Manuscript database

Higher plasma pyridoxal 5'-phosphate (PLP) levels are associated with a decreased incidence of colorectal cancer, but the influence of plasma PLP on survival of patients with colorectal cancer is unknown. We prospectively examined whether prediagnostic plasma PLP levels are associated with mortality...

A novel mutation of α-galactosidase A gene causes Fabry disease mimicking primary erythromelalgia in a Chinese family.

PubMed

Ge, Wei; Wei, Bin; Zhu, Hao; Miao, Zhigang; Zhang, Weimin; Leng, Cuihua; Li, Jizhen; Zhang, Dan; Sun, Miao; Xu, Xingshun

2017-05-01

Fabry disease is an X-linked genetic disorder caused by the mutations of α-galactosidase A (GLA, MIM 300644) gene presenting with various clinical symptoms including small-fiber peripheral neuropathy and limb burning pain. Here, we reported a Chinese pedigree with the initial diagnosis of primary erythromelalgia in an autosomal dominant (AD)-inherited pattern. Mutation analysis of SCN9A and GLA genes by direct sequencing and functional analysis of a novel mutation of GLA in cells were performed. Our data did not show any pathological mutations in SCN9A gene; however, a novel missense mutation c.139T>C (p.W47R) of GLA was identified in a male proband as well as two female carriers in this family. Enzyme assay of α-galactosidase A activity showed deficient enzyme activity in male patients and female carriers, further confirming the diagnosis of Fabry disease. Finally, a functional analysis indicated that the replacement of the 47th amino acid tryptophan (W47) with arginine (W47R) or glycine (W47G) led to reduced activity of α-galactosidase A in 293T cells. Therefore, these findings demonstrated that the novel mutation p.W47R of GLA is the cause of Fabry disease. Because Fabry disease and primary erythromelalgia share similar symptoms, it is a good strategy for clinical physicians to perform genetic mutation screenings on both SCN9A and GLA genes in those patients with limb burning pain but without a clear inheritant pattern.

Identification of a novel amino acid racemase from a hyperthermophilic archaeon Pyrococcus horikoshii OT-3 induced by D-amino acids.

PubMed

Kawakami, Ryushi; Ohmori, Taketo; Sakuraba, Haruhiko; Ohshima, Toshihisa

2015-08-01

To date, there have been few reports analyzing the amino acid requirement for growth of hyperthermophilic archaea. We here found that the hyperthermophilic archaeon Pyrococcus horikoshii OT-3 requires Thr, Leu, Val, Phe, Tyr, Trp, His and Arg in the medium for growth, and shows slow growth in medium lacking Met or Ile. This largely corresponds to the presence, or absence, of genes related to amino acid biosynthesis in its genome, though there are exceptions. The amino acid requirements were dramatically lost by addition of D-isomers of Met, Leu, Val, allo-Ile, Phe, Tyr, Trp and Arg. Tracer analysis using (14)C-labeled D-Trp showed that D-Trp in the medium was used as a protein component in the cells, suggesting the presence of D-amino acid metabolic enzymes. Pyridoxal 5'-phosphate (PLP)-dependent racemase activity toward Met, Leu and Phe was detected in crude extract of P. horikoshii and was enhanced in cells grown in the medium supplemented with D-amino acids, especially D-allo-Ile. The gene encoding the racemase was narrowed down to one open reading frame on the basis of enzyme purification from P. horikoshii cells, and the recombinant enzyme exhibited PLP-dependent racemase activity toward several amino acids, including Met, Leu and Phe, but not Pro, Asp or Glu. This is the first report showing the presence in a hyperthermophilic archaeon of a PLP-dependent amino acid racemase with broad substrate specificity that is likely responsible for utilization of D-amino acids for growth.

Heart failure and sudden cardiac death in heritable thoracic aortic disease caused by pathogenic variants in the SMAD3 gene.

PubMed

Backer, Julie De; Braverman, Alan C

2018-05-01

Predominant cardiovascular manifestations in the spectrum of Heritable Thoracic Aortic Disease include by default aortic root aneurysms- and dissections, which may be associated with aortic valve disease. Mitral- and tricuspid valve prolapse are other commonly recognized features. Myocardial disease, characterized by heart failure and/or malignant arrhythmias has been reported in humans and in animal models harboring pathogenic variants in the Fibrillin1 gene. Description of clinical history of three cases from one family in Ghent (Belgium) and one family in St. Louis (US). We report on three cases from two families presenting end-stage heart failure (in two) and lethal arrhythmias associated with moderate left ventricular dilatation (in one). All three cases harbor a pathogenic variant in the SMAD3 gene, known to cause aneurysm osteoarthritis syndrome, Loeys-Dietz syndrome type 3 or isolated Heritable Thoracic Aortic Disease. These unusual presentations warrant awareness for myocardial disease in patients harboring pathogenic variants in genes causing Heritable Thoracic Aortic Disease and indicate the need for prospective studies in larger cohorts. © 2018 The Authors. Molecular Genetics & Genomic Medicine published by Wiley Periodicals, Inc.

The Relationship between the Expression of Ethylene-Related Genes and Papaya Fruit Ripening Disorder Caused by Chilling Injury

PubMed Central

Zou, Yuan; Zhang, Lin; Rao, Shen; Zhu, Xiaoyang; Ye, Lanlan; Chen, Weixin; Li, Xueping

2014-01-01

Papaya (Carica papaya L.) is sensitive to low temperature and easy to be subjected to chilling injury, which causes fruit ripening disorder. This study aimed to investigate the relationship between the expression of genes related to ethylene and fruit ripening disorder caused by chilling injury. Papaya fruits were firstly stored at 7°C and 12°C for 25 and 30 days, respectively, then treated with exogenous ethylene and followed by ripening at 25°C for 5 days. Chilling injury symptoms such as pulp water soaking were observed in fruit stored at 7°C on 20 days, whereas the coloration and softening were completely blocked after 25 days, Large differences in the changes in the expression levels of twenty two genes involved in ethylene were seen during 7°C-storage with chilling injury. Those genes with altered expression could be divided into three groups: the group of genes that were up-regulated, including ACS1/2/3, EIN2, EIN3s/EIL1, CTR1/2/3, and ERF1/3/4; the group of genes that were down-regulated, including ACO3, ETR1, CTR4, EBF2, and ERF2; and the group of genes that were un-regulated, including ACO1/2, ERS, and EBF1. The results also showed that pulp firmness had a significantly positive correlation with the expression of ACS2, ACO1, CTR1/4, EIN3a/b, and EBF1/2 in fruit without chilling injury. This positive correlation was changed to negative one in fruit after storage at 7°C for 25 days with chilling injury. The coloring index displayed significantly negative correlations with the expression levels of ACS2, ACO1/2, CTR4, EIN3a/b, ERF3 in fruit without chilling injury, but these correlations were changed into the positive ones in fruit after storage at 7°C for 25 days with chilling injury. All together, these results indicate that these genes may play important roles in the abnormal softening and coloration with chilling injury in papaya. PMID:25542021

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.

A simple high-performance liquid chromatography (HPLC) method for the measurement of pyridoxal-5-phosphate and 4-pyridoxic acid in human plasma.

PubMed

Cabo, Rona; Kozik, Karolina; Milanowski, Maciej; Hernes, Sigrunn; Slettan, Audun; Haugen, Margaretha; Ye, Shu; Blomhoff, Rune; Mansoor, M Azam

2014-06-10

Low concentration of plasma pyridoxal-5-phosphate (PLP) is associated with hyperhomocysteinemia and inflammation. Most methods for the measurement of plasma PLP require large specimen volume and involve the use of toxic reagents. We have developed a HPLC method for the measurement of PLP and 4-pyridoxic acid (4-PA) in plasma, which requires small specimen volume. The samples are prepared without adding any toxic reagents. Furthermore, we have examined whether intake of vitamin B6 affects the concentration of plasma PLP and 4-PA. The coefficient of variation of the method was 6% and the recovery of the added vitamin in plasma was about 100%. The concentrations of plasma PLP and 4-PA in 168 healthy subjects were 40.6 (8.4-165.0) nmol/L, median and (range) and 17.5 (3.7-114.79) nmol/L, median and (range) respectively. In the multiple regression analyses, the concentration of plasma PLP was associated with the concentration of plasma 4-PA (p<0.0001), BMI, (p=0.02) and sex, (p=0.0008). The concentration of plasma 4-PA was associated with plasma PLP (p<0.0001), serum folate (p=0.004), smoking (p=0.03) and vitamin B6 intake (p=0.01). The present method is suitable for large clinical studies for the measurement of plasma PLP and 4-PA. Our findings demonstrate that plasma 4-PA, BMI and sex are the major determinants of plasma PLP in healthy individuals. Copyright © 2014 Elsevier B.V. All rights reserved.

Mutation of a chitinase-like gene causes ectopic deposition of lignin, aberrant cell shapes, and overproduction of ethylene.

PubMed

Zhong, Ruiqin; Kays, Stanley J; Schroeder, Betty P; Ye, Zheng-Hua

2002-01-01

Chitinase-like proteins have long been proposed to play roles in normal plant growth and development, but no mutations in chitinase-like genes have been obtained previously to support this hypothesis. In this study, we have shown that the gene responsible for the elp1 mutation in Arabidopsis encodes a chitinase-like protein (AtCTL1). Mutation of this chitinase-like gene caused ectopic deposition of lignin and aberrant shapes of cells with incomplete cell walls in the pith of inflorescence stems. The AtCTL1 gene was expressed in all organs during normal plant growth and development, but it was not induced by wounding, salicylic acid, pectin fragments, or ethylene. Consistent with its ubiquitous expression pattern, mutation of the AtCTL1 gene affected many aspects of plant growth and development, including exaggerated hook curvature, reduced length and increased diameter of hypocotyls in dark-grown seedlings, and reduced root length and increased number of root hairs in light-grown seedlings. The mutant phenotypes could be rescued partially by ethylene inhibitors, and ethylene production in the mutant was significantly greater than in the wild type. Together, these results suggest that AtCTL1, a chitinase-like gene, is essential for normal plant growth and development in Arabidopsis.

VIP Gene Deletion in Mice Causes Cardiomyopathy Associated with Upregulation of Heart Failure Genes

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

Szema, Anthony M.; Hamidi, Sayyed A.; Smith, S. David

2013-05-20

Vasoactive Intestinal Peptide (VIP), a pulmonary vasodilator and inhibitor of vascular smooth muscle proliferation, is absent in pulmonary arteries of patients with idiopathic pulmonary arterial hypertension (PAH). We previously determined that targeted deletion of the VIP gene in mice leads to PAH with pulmonary vascular remodeling and right ventricular (RV) dilatation. Whether the left ventricle is also affected by VIP gene deletion is unknown. In the current study, we examined if VIP knockout mice (VIP-/-) develop both right (RV) and left ventricular (LV) cardiomyopathy, manifested by LV dilatation and systolic dysfunction, as well as overexpression of genes conducive to heartmore » failure.« less

The myelin proteolipid DMα in fishes.

PubMed

Brösamle, Christian

2010-05-01

Vertebrate myelin membranes are compacted and held in close apposition by three structural proteins of myelin, myelin basic protein, myelin protein zero (MPZ) and myelin proteolipid protein (PLP1/DMalpha). PLP1/DMalpha is considered to function as a scaffolding protein and play a role in intracellular trafficking in oligodendrocytes. In humans, point mutations, duplications or deletions of PLP1 are associated with Pelizaeus-Merzbacher disease and spastic paraplegia Type 2. PLP1 is highly conserved between mammals, but less so in lower vertebrates. This has led some researchers to question whether certain fish species express PLP1 orthologues at all, and to suggest that the function of PLP1/DMalpha in the central nervous system (CNS) may have been taken over by MPZ. Here, we review the evidence for the conservation of orthologues of PLP1/DMalpha in actinopterygian fishes and provide a comparison of currently available sequence data across 17 fish species. Our analysis demonstrates that orthologues of PLP1/DMalpha have been retained and are functionally expressed in many, if not all, extant species of bony fish. Many of the amino acids that, when mutated, are associated with severe CNS pathology are conserved in teleosts, demonstrating conservation of essential functions and justifying the development of novel disease models in species such as the zebrafish.

Atelosteogenesis type II is caused by mutations in the diastrophic dysplasia sulfate-transporter gene (DTDST): Evidence for a phenotypic series involving three chondrodysplasias

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

Haestbacka, J.; Lander, E.S.; Superti-Furga, A.

1996-02-01

Atelosteogenesis type II (AO II) is a neonatally lethal chondrodysplasia whose clinical and histological characteristics resemble those of another chondrodysplasia, the much less severe diastrophic dysplasia (DTD). The similarity suggests a shared pathogenesis involving lesions in the same biochemical pathway and perhaps the same gene. DTD is caused by mutations in the recently identified diastrophic dysplasia sulfate-transporter gene (DTDST). Here, we report that AOII patients also have DTDST mutations, which lead to defective uptake of inorganic sulfate and insufficient sulfation of macromolecules by patient mesenchymal cells in vitro. Together with our recent observation that a third even more severe chondrodysplasia,more » achondrogenesis type IB, is also caused by mutations in DTDST, these results demonstrate a phenotypic series of three chondrodysplasias of increasing severity caused by lesions in a single sulfate-transporter gene. The severity of the phenotype appears to be correlated with the predicted effect of the mutations on the residual activity of the DTDST protein. 24 refs., 6 figs., 1 tab.« less

Human male infertility and its genetic causes.

PubMed

Miyamoto, Toshinobu; Minase, Gaku; Shin, Takeshi; Ueda, Hiroto; Okada, Hiroshi; Sengoku, Kazuo

2017-04-01

Infertility affects about 15% of couples who wish to have children and half of these cases are associated with male factors. Genetic causes of azoospermia include chromosomal abnormalities, Y chromosome microdeletions, and specific mutations/deletions of several Y chromosome genes. Many researchers have analyzed genes in the AZF region on the Y chromosome; however, in 2003 the SYCP3 gene on chromosome 12 (12q23) was identified as causing azoospermia by meiotic arrest through a point mutation. We mainly describe the SYCP3 and PLK4 genes that we have studied in our laboratory, and add comments on other genes associated with human male infertility. Up to now, The 17 genes causing male infertility by their mutation have been reported in human. Infertility caused by nonobstructive azoospermia (NOA) is very important in the field of assisted reproductive technology. Even with the aid of chromosomal analysis, ultrasonography of the testis, and detailed endocrinology, only MD-TESE can confirm the presence of immature spermatozoa in the testes. We strongly hope that these studies help clinics avoid ineffective MD-TESE procedures.

A novel recessive mutation in the gene ELOVL4 causes a neuro-ichthyotic disorder with variable expressivity

PubMed Central

2014-01-01

Background A rare neuro-ichthyotic disorder characterized by ichthyosis, spastic quadriplegia and intellectual disability and caused by recessive mutations in ELOVL4, encoding elongase-4 protein has recently been described. The objective of the study was to search for sequence variants in the gene ELOVL4 in three affected individuals of a consanguineous Pakistani family exhibiting features of neuro-ichthyotic disorder. Methods Linkage in the family was searched by genotyping microsatellite markers linked to the gene ELOVL4, mapped at chromosome 6p14.1. Exons and splice junction sites of the gene ELOVL4 were polymerase chain reaction amplified and sequenced in an automated DNA sequencer. Results DNA sequence analysis revealed a novel homozygous nonsense mutation (c.78C > G; p.Tyr26*). Conclusions Our report further confirms the recently described ELOVL4-related neuro-ichthyosis and shows that the neurological phenotype can be absent in some individuals. PMID:24571530

Clinical characteristics of severe congenital neutropenia caused by novel ELANE gene mutations.

PubMed

Shu, Zhou; Li, Xiao-Hui; Bai, Xiao-Ming; Zhang, Zhi-Yong; Jiang, Li-ping; Tang, Xue-Mei; Zhao, Xiao-dong

2015-02-01

Mutations within the ELANE gene, which encodes human neutrophil elastase, are the most common genetic causes of severe congenital neutropenia (SCN). No cases of SCN have been previously described from a Chinese population. Herein, we describe the clinical, hematologic and molecular characteristics of 7 Chinese SCN cases with novel ELANE mutations. Seven Chinese pediatric patients (4 males and 3 females) with suspected SCN were enrolled in this study. Clinical data, peripheral blood, bone marrow and immune function were evaluated for SCN. ELANE genomic DNA and cDNA sequences from patients and potential carriers were analyzed using polymerase chain reaction (PCR) and direct sequencing. All the7 patients experienced recurrent infection (soft tissue, lung, oral cavity) during a period of 120 days. Noninfectious conditions such as anemia and osteopenia were found in most patients, and absolute peripheral neutrophil counts varied. DNA and cDNA sequencing demonstrated that the patients harbored a range of heterozygous ELANE gene mutations, including substitution, deletion, insertion and frame shift alterations. All the mutations had not been reported previously; however, no mutation carriers were identified among the parents or siblings, even in a family with 2 affected offspring. SCN cases were identified for the first time in China, and all patients carried novel ELANE mutations. Granulocyte-colony stimulating factor (G-CSF) was an effective treatment for most of the SCN patients and prevented life-threatening bacterial infections.

Reversal of gene dysregulation in cultured cytotrophoblasts reveals possible causes of preeclampsia

PubMed Central

Zhou, Yan; Gormley, Matthew J.; Hunkapiller, Nathan M.; Kapidzic, Mirhan; Stolyarov, Yana; Feng, Victoria; Nishida, Masakazu; Drake, Penelope M.; Bianco, Katherine; Wang, Fei; McMaster, Michael T.; Fisher, Susan J.

2013-01-01

During human pregnancy, a subset of placental cytotrophoblasts (CTBs) differentiates into cells that aggressively invade the uterus and its vasculature, anchoring the progeny and rerouting maternal blood to the placenta. In preeclampsia (PE), CTB invasion is limited, reducing placental perfusion and/or creating intermittent flow. This syndrome, affecting 4%–8% of pregnancies, entails maternal vascular alterations (e.g., high blood pressure, proteinuria, and edema) and, in some patients, fetal growth restriction. The only cure is removal of the faulty placenta, i.e., delivery. Previously, we showed that defective CTB differentiation contributes to the placental component of PE, but the causes were unknown. Here, we cultured CTBs isolated from PE and control placentas for 48 hours, enabling differentiation and invasion. In various severe forms of PE, transcriptomics revealed common aberrations in CTB gene expression immediately after isolation, including upregulation of SEMA3B, which resolved in culture. The addition of SEMA3B to normal CTBs inhibited invasion and recreated aspects of the PE phenotype. Additionally, SEMA3B downregulated VEGF signaling through the PI3K/AKT and GSK3 pathways, effects that were observed in PE CTBs. We propose that, in severe PE, the in vivo environment dysregulates CTB gene expression; the autocrine actions of the upregulated molecules (including SEMA3B) impair CTB differentiation, invasion and signaling; and patient-specific factors determine the signs. PMID:23934129

Imaging of Skeletal Disorders Caused by Fibroblast Growth Factor Receptor Gene Mutations.

PubMed

Sargar, Kiran M; Singh, Achint K; Kao, Simon C

2017-10-01

Fibroblast growth factors and fibroblast growth factor receptors (FGFRs) play important roles in human axial and craniofacial skeletal development. FGFR1, FGFR2, and FGFR3 are crucial for both chondrogenesis and osteogenesis. Mutations in the genes encoding FGFRs, types 1-3, are responsible for various skeletal dysplasias and craniosynostosis syndromes. Many of these disorders are relatively common in the pediatric population, and diagnosis is often challenging. These skeletal disorders can be classified based on which FGFR is affected. Skeletal disorders caused by type 1 mutations include Pfeiffer syndrome (PS) and osteoglophonic dysplasia, and disorders caused by type 2 mutations include Crouzon syndrome (CS), Apert syndrome (AS), and PS. Disorders caused by type 3 mutations include achondroplasia, hypochondroplasia, thanatophoric dysplasia (TD), severe achondroplasia with developmental delay and acanthosis nigricans, Crouzonodermoskeletal syndrome, and Muenke syndrome. Most of these mutations are inherited in an autosomal dominant fashion and are gain-of-function-type mutations. Imaging plays a key role in the evaluation of these skeletal disorders. Knowledge of the characteristic imaging and clinical findings can help confirm the correct diagnosis and guide the appropriate molecular genetic tests. Some characteristics and clinical findings include premature fusion of cranial sutures and deviated broad thumbs and toes in PS; premature fusion of cranial sutures and syndactyly of the hands and feet in AS; craniosynostosis, ocular proptosis, and absence of hand and foot abnormalities in CS; rhizomelic limb shortening, caudal narrowing of the lumbar interpediculate distance, small and square iliac wings, and trident hands in achondroplasia; and micromelia, bowing of the femora, and platyspondyly in TD. © RSNA, 2017.

A novel mutation in PGAP2 gene causes developmental delay, intellectual disability, epilepsy and microcephaly in consanguineous Saudi family.

PubMed

Naseer, Muhammad Imran; Rasool, Mahmood; Jan, Mohammed M; Chaudhary, Adeel G; Pushparaj, Peter Natesan; Abuzenadah, Adel M; Al-Qahtani, Mohammad H

2016-12-15

PGAP2 (Post-GPI Attachment to Proteins 2) gene is involved in lipid remodeling steps of Glycosylphosphatidylinositol (GPI)-anchor maturation. At the surface of the cell this gene is required for proper expression of GPI-anchored proteins. Hyperphosphatasia with mental retardation syndrome-3 is an autosomal recessive disorder usually characterized by severe mental retardation. Mutations in the PGAP2 gene cause hyperphosphatasia mental retardation syndrome-3. We have identified a large consanguineous family from Saudi origin segregating developmental delay, intellectual disability, epilepsy and microcephaly. Whole exome sequencing with 100× coverage was performed on two affected siblings of the family. Data analysis in the patient revealed a novel missense mutation c.191C>T in PGAP2 gene resulting in Alanine to Valine substitution (Ala64Val). The mutation was reconfirmed and validated by subsequent Sanger sequencing method. The mutation was ruled out in 100 unrelated healthy controls. We suggest that this pathogenic mutation disrupts the proper function of the gene proteins resulting in the disease state. Copyright © 2016 Elsevier B.V. All rights reserved.

Point mutation in the MITF gene causing Waardenburg syndrome type II in a three-generation Indian family.

PubMed

Lalwani, A K; Attaie, A; Randolph, F T; Deshmukh, D; Wang, C; Mhatre, A; Wilcox, E

1998-12-04

Waardenburg syndrome (WS) is an autosomal-dominant neural crest cell disorder phenotypically characterized by hearing impairment and disturbance of pigmentation. A presence of dystopia canthorum is indicative of WS type 1, caused by loss of function mutation in the PAX3 gene. In contrast, type 2 WS (WS2) is characterized by normally placed medial canthi and is genetically heterogeneous; mutations in MITF (microphthalmia associated transcription factor) associated with WS2 have been identified in some but not all affected families. Here, we report on a three-generation Indian family with a point mutation in the MITF gene causing WS2. This mutation, initially reported in a Northern European family, creates a stop codon in exon 7 and is predicted to result in a truncated protein lacking the HLH-Zip or Zip structure necessary for normal interaction with its target DNA motif. Comparison of the phenotype between the two families demonstrates a significant difference in pigmentary disturbance of the eye. This family, with the first documented case of two unrelated WS2 families harboring identical mutations, provides additional evidence for the importance of genetic background on the clinical phenotype.

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 frameshift deletion in the albumin gene causes analbuminemia in a young Turkish woman.

PubMed

Dagnino, Monica; Caridi, Gianluca; Aydin, Zeki; Ozturk, Savas; Karaali, Zeynep; Kazancioglu, Rumeyza; Cefle, Kivanc; Gursu, Meltem; Campagnoli, Monica; Galliano, Monica; Minchiotti, Lorenzo

2010-11-11

Analbuminemia is a rare autosomal recessive disorder manifested by the absence, or severe reduction, of circulating serum albumin. The analbuminemic trait was diagnosed in a young Turkish woman on the basis of her clinical symptoms (bilateral lower limb edema) and biochemical findings (minimal albumin amount and variable increases in other protein fractions). Total DNA from the analbuminemic proband and her parents was PCR-amplified using oligonucleotide primers designed to amplify the 14 exons of the albumin gene (ALB) and the flanking intron regions. The products were screened for mutations by single-strand conformation polymorphism (SSCP) and heteroduplex analyses (HA). HA allowed the identification of the mutation site in exon 12. Direct DNA sequencing of this abnormal fragment revealed that the analbuminemic trait was caused by a homozygous CA deletion at nucleotide positions c. 1614-1615 in the codons for Cys538 and Thr539. The subsequent frameshift should give rise to a putative truncated albumin variant in which the sequence Cys(538)-Thr-Leu-Ser has been changed to Cys(538)-Thr-Phe-Stop. The parents were heterozygous for the same mutation. Gel-based mutation detection and DNA sequencing substantiate the clinical diagnosis of congenital analbuminemia in our patient and show that the condition is caused by a novel mutation within the ALB gene. These results contribute to shed light on the molecular basis of this rare condition. 2010 Elsevier B.V. All rights reserved.

The effect of mirror therapy on the management of phantom limb pain.

PubMed

Yıldırım, Meltem; Kanan, Nevin

2016-07-01

In the last two decades, mirror therapy has become a frequently used method of managing phantom limb pain (PLP). However, the role of nurses in mirror therapy has not yet been well defined. This study examined the effect of mirror therapy on the management of PLP, and discusses the importance of mirror therapy in the nursing care of amputee patients. This quasi-experimental study was conducted in the pain management department of a university hospital and a prosthesis clinic in İstanbul, Turkey, with 15 amputee patients who had PLP. Forty minutes of practical mirror therapy training was given to the patients and they were asked to practice at home for 4 weeks. Patients were asked to record the severity of their PLP before and after the therapy each day using 0-10 Numeric Pain Intensity Scale. Mirror therapy practiced for 4 weeks provided a significant decrease in severity of PLP. There was no significant relationship between the effect of mirror therapy and demographic, amputation or PLP-related characteristics. Patients who were not using prosthesis had greater benefit from mirror therapy. Mirror therapy can be used as an adjunct to medical and surgical treatment of PLP. It is a method that patients can practice independently, enhancing self-control over phantom pain. As mirror therapy is a safe, economical, and easy-to-use treatment method, it should be considered in the nursing care plan for patients with PLP.

Case reports of juvenile GM1 gangliosidosisis type II caused by mutation in GLB1 gene.

PubMed

Karimzadeh, Parvaneh; Naderi, Samaneh; Modarresi, Farzaneh; Dastsooz, Hassan; Nemati, Hamid; Farokhashtiani, Tayebeh; Shamsian, Bibi Shahin; Inaloo, Soroor; Faghihi, Mohammad Ali

2017-07-17

Type II or juvenile GM1-gangliosidosis is an autosomal recessive lysosomal storage disorder, which is clinically distinct from infantile form of the disease by the lack of characteristic cherry-red spot and hepatosplenomegaly. The disease is characterized by slowly progressive neurodegeneration and mild skeletal changes. Due to the later age of onset and uncharacteristic presentation, diagnosis is frequently puzzled with other ataxic and purely neurological disorders. Up to now, 3-4 types of GM1-gangliosidosis have been reported and among them type I is the most common phenotype with the age of onset around 6 months. Various forms of GM1-gangliosidosis are caused by GLB1 gene mutations but severity of the disease and age of onset are directly related to the position and the nature of deleterious mutations. However, due to its unique genetic cause and overlapping clinical features, some researchers believe that GM1 gangliosidosis represents an overlapped disease spectrum instead of four distinct types. Here, we report a less frequent type of autosomal recessive GM1 gangliosidosis with perplexing clinical presentation in three families in the southwest part of Iran, who are unrelated but all from "Lurs" ethnic background. To identify disease-causing mutations, Whole Exome Sequencing (WES) utilizing next generation sequencing was performed. Four patients from three families were investigated with the age of onset around 3 years old. Clinical presentations were ataxia, gate disturbances and dystonia leading to wheelchair-dependent disability, regression of intellectual abilities, and general developmental regression. They all were born in consanguineous families with no previous documented similar disease in their parents. A homozygote missense mutation in GLB1 gene (c. 601 G > A, p.R201C) was found in all patients. Using Sanger sequencing this identified mutation was confirmed in the proband, their parents, grandparents, and extended family members, confirming

Mutation of the Erwinia amylovora argD gene causes arginine auxotrophy, nonpathogenicity in apples, and reduced virulence in pears.

PubMed

Ramos, Laura S; Lehman, Brian L; Peter, Kari A; McNellis, Timothy W

2014-11-01

Fire blight is caused by Erwinia amylovora and is the most destructive bacterial disease of apples and pears worldwide. In this study, we found that E. amylovora argD(1000)::Tn5, an argD Tn5 transposon mutant that has the Tn5 transposon inserted after nucleotide 999 in the argD gene-coding region, was an arginine auxotroph that did not cause fire blight in apple and had reduced virulence in immature pear fruits. The E. amylovora argD gene encodes a predicted N-acetylornithine aminotransferase enzyme, which is involved in the production of the amino acid arginine. A plasmid-borne copy of the wild-type argD gene complemented both the nonpathogenic and the arginine auxotrophic phenotypes of the argD(1000)::Tn5 mutant. However, even when mixed with virulent E. amylovora cells and inoculated onto immature apple fruit, the argD(1000)::Tn5 mutant still failed to grow, while the virulent strain grew and caused disease. Furthermore, the pCR2.1-argD complementation plasmid was stably maintained in the argD(1000)::Tn5 mutant growing in host tissues without any antibiotic selection. Therefore, the pCR2.1-argD complementation plasmid could be useful for the expression of genes, markers, and reporters in E. amylovora growing in planta, without concern about losing the plasmid over time. The ArgD protein cannot be considered an E. amylovora virulence factor because the argD(1000)::Tn5 mutant was auxotrophic and had a primary metabolism defect. Nevertheless, these results are informative about the parasitic nature of the fire blight disease interaction, since they indicate that E. amylovora cannot obtain sufficient arginine from apple and pear fruit tissues or from apple vegetative tissues, either at the beginning of the infection process or after the infection has progressed to an advanced state. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Genotype–phenotype characteristics and baseline natural history of heritable neuropathies caused by mutations in the MPZ gene

PubMed Central

Feely, Shawna; Scherer, Steven S.; Herrmann, David N.; Burns, Joshua; Muntoni, Francesco; Li, Jun; Siskind, Carly E.; Day, John W.; Laura, Matilde; Sumner, Charlotte J.; Lloyd, Thomas E.; Ramchandren, Sindhu; Shy, Rosemary R.; Grider, Tiffany; Bacon, Chelsea; Finkel, Richard S.; Yum, Sabrina W.; Moroni, Isabella; Piscosquito, Giuseppe; Pareyson, Davide; Reilly, Mary M.; Shy, Michael E.

2015-01-01

We aimed to characterize genotype–phenotype correlations and establish baseline clinical data for peripheral neuropathies caused by mutations in the myelin protein zero (MPZ) gene. MPZ mutations are the second leading cause of Charcot–Marie–Tooth disease type 1. Recent research makes clinical trials for patients with MPZ mutations a realistic possibility. However, the clinical severity varies with different mutations and natural history data on progression is sparse. We present cross-sectional data to begin to define the phenotypic spectrum and clinical baseline of patients with these mutations. A cohort of patients with MPZ gene mutations was identified in 13 centres of the Inherited Neuropathies Consortium - Rare Disease Clinical Research Consortium (INC-RDCRC) between 2009 and 2012 and at Wayne State University between 1996 and 2009. Patient phenotypes were quantified by the Charcot–Marie–Tooth disease neuropathy score version 1 or 2 and the Charcot–Marie–Tooth disease paediatric scale outcome instruments. Genetic testing was performed in all patients and/or in first- or second-degree relatives to document mutation in MPZ gene indicating diagnosis of Charcot–Marie–Tooth disease type 1B. There were 103 patients from 71 families with 47 different MPZ mutations with a mean age of 40 years (range 3–84 years). Patients and mutations were separated into infantile, childhood and adult-onset groups. The infantile onset group had higher Charcot–Marie–Tooth disease neuropathy score version 1 or 2 and slower nerve conductions than the other groups, and severity increased with age. Twenty-three patients had no family history of Charcot–Marie–Tooth disease. Sixty-one patients wore foot/ankle orthoses, 19 required walking assistance or support, and 10 required wheelchairs. There was hearing loss in 21 and scoliosis in 17. Forty-two patients did not begin walking until after 15 months of age. Half of the infantile onset patients then required

A second mutation in the type II procollagen gene (COL2AI) causing stickler syndrome (arthro-ophthalmopathy) is also a premature termination codon.

PubMed Central

Ahmad, N N; McDonald-McGinn, D M; Zackai, E H; Knowlton, R G; LaRossa, D; DiMascio, J; Prockop, D J

1993-01-01

Genetic linkage analyses suggest that mutations in type II collagen may be responsible for Stickler syndrome, or arthro-ophthalmopathy (AO), in many families. In the present study oligonucleotide primers were developed to amplify and directly sequence eight of the first nine exons of the gene for type II procollagen (COL2A1). Analysis of the eight exons in 10 unrelated probands with AO revealed that one had a single-base mutation in one allele that changed the codon of -CGA- for arginine at amino acid position alpha 1-9 in exon 7 to a premature termination signal for translation. The second mutation found to cause AO was, therefore, similar to the first in that both created premature termination signals in the COL2A1 gene. Since mutations producing premature termination signals have not previously been detected in genes for fibrillar collagens, the results raise the possibility that such mutations in the COL2A1 gene are a common cause of AO. Images Figure 2 Figure 3 PMID:8434604

Digital gene expression analysis of corky split vein caused by boron deficiency in 'Newhall' Navel Orange (Citrus sinensis Osbeck) for selecting differentially expressed genes related to vascular hypertrophy.

PubMed

Yang, Cheng-Quan; Liu, Yong-Zhong; An, Ji-Cui; Li, Shuang; Jin, Long-Fei; Zhou, Gao-Feng; Wei, Qing-Jiang; Yan, Hui-Qing; Wang, Nan-Nan; Fu, Li-Na; Liu, Xiao; Hu, Xiao-Mei; Yan, Ting-Shuai; Peng, Shu-Ang

2013-01-01

Corky split vein caused by boron (B) deficiency in 'Newhall' Navel Orange was studied in the present research. The boron-deficient citrus exhibited a symptom of corky split vein in mature leaves. Morphologic and anatomical surveys at four representative phases of corky split veins showed that the symptom was the result of vascular hypertrophy. Digital gene expression (DGE) analysis was performed based on the Illumina HiSeq™ 2000 platform, which was applied to analyze the gene expression profilings of corky split veins at four morphologic phases. Over 5.3 million clean reads per library were successfully mapped to the reference database and more than 22897 mapped genes per library were simultaneously obtained. Analysis of the differentially expressed genes (DEGs) revealed that the expressions of genes associated with cytokinin signal transduction, cell division, vascular development, lignin biosynthesis and photosynthesis in corky split veins were all affected. The expressions of WOL and ARR12 involved in the cytokinin signal transduction pathway were up-regulated at 1(st) phase of corky split vein development. Furthermore, the expressions of some cell cycle genes, CYCs and CDKB, and vascular development genes, WOX4 and VND7, were up-regulated at the following 2(nd) and 3(rd) phases. These findings indicated that the cytokinin signal transduction pathway may play a role in initiating symptom observed in our study.

A Partial Gene Deletion of SLC45A2 Causes Oculocutaneous Albinism in Doberman Pinscher Dogs

PubMed Central

Winkler, Paige A.; Gornik, Kara R.; Ramsey, David T.; Dubielzig, Richard R.; Venta, Patrick J.; Petersen-Jones, Simon M.; Bartoe, Joshua T.

2014-01-01

The first white Doberman pinscher (WDP) dog was registered by the American Kennel Club in 1976. The novelty of the white coat color resulted in extensive line breeding of this dog and her offspring. The WDP phenotype closely resembles human oculocutaneous albinism (OCA) and clinicians noticed a seemingly high prevalence of pigmented masses on these dogs. This study had three specific aims: (1) produce a detailed description of the ocular phenotype of WDPs, (2) objectively determine if an increased prevalence of ocular and cutaneous melanocytic tumors was present in WDPs, and (3) determine if a genetic mutation in any of the genes known to cause human OCA is causal for the WDP phenotype. WDPs have a consistent ocular phenotype of photophobia, hypopigmented adnexal structures, blue irides with a tan periphery and hypopigmented retinal pigment epithelium and choroid. WDPs have a higher prevalence of cutaneous melanocytic neoplasms compared with control standard color Doberman pinschers (SDPs); cutaneous tumors were noted in 12/20 WDP (<5 years of age: 4/12; >5 years of age: 8/8) and 1/20 SDPs (p<0.00001). Using exclusion analysis, four OCA causative genes were investigated for their association with WDP phenotype; TYR, OCA2, TYRP1 and SLC45A2. SLC45A2 was found to be linked to the phenotype and gene sequencing revealed a 4,081 base pair deletion resulting in loss of the terminus of exon seven of SLC45A2 (chr4∶77,062,968–77,067,051). This mutation is highly likely to be the cause of the WDP phenotype and is supported by a lack of detectable SLC45A2 transcript levels by reverse transcriptase PCR. The WDP provides a valuable model for studying OCA4 visual disturbances and melanocytic neoplasms in a large animal model. PMID:24647637

The p.Leu167del Mutation in APOE Gene Causes Autosomal Dominant Hypercholesterolemia by Down-regulation of LDL Receptor Expression in Hepatocytes.

PubMed

Cenarro, Ana; Etxebarria, Aitor; de Castro-Orós, Isabel; Stef, Marianne; Bea, Ana M; Palacios, Lourdes; Mateo-Gallego, Rocío; Benito-Vicente, Asier; Ostolaza, Helena; Tejedor, Teresa; Martín, César; Civeira, Fernando

2016-05-01

The p.Leu167del mutation in the APOE gene has been associated with hyperlipidemia. Our objective was to determine the frequency of p.Leu167del mutation in APOE gene in subjects with autosomal dominant hypercholesterolemia (ADH) in whom LDLR, APOB, and PCSK9 mutations had been excluded and to identify the mechanisms by which this mutant apo E causes hypercholesterolemia. The APOE gene was analyzed in a case-control study. The study was conducted at a University Hospital Lipid Clinic. Two groups (ADH, 288 patients; control, 220 normolipidemic subjects) were included. We performed sequencing of APOE gene and proteomic and cellular experiments. To determine the frequency of the p.Leu167del mutation and the mechanism by which it causes hypercholesterolemia. In the ADH group, nine subjects (3.1%) were carriers of the APOE c.500_502delTCC, p.Leu167del mutation, cosegregating with hypercholesterolemia in studied families. Proteomic quantification of wild-type and mutant apo E in very low-density lipoprotein (VLDL) from carrier subjects revealed that apo E3 is almost a 5-fold increase compared to mutant apo E. Cultured cell studies revealed that VLDL from mutation carriers had a significantly higher uptake by HepG2 and THP-1 cells compared to VLDL from subjects with E3/E3 or E2/E2 genotypes. Transcriptional down-regulation of LDLR was also confirmed. p.Leu167del mutation in APOE gene is the cause of hypercholesterolemia in the 3.1% of our ADH subjects without LDLR, APOB, and PCSK9 mutations. The mechanism by which this mutation is associated to ADH is that VLDL carrying the mutant apo E produces LDLR down-regulation, thereby raising plasma low-density lipoprotein cholesterol levels.

Exercise training attenuates experimental autoimmune encephalomyelitis by peripheral immunomodulation rather than direct neuroprotection.

PubMed

Einstein, Ofira; Fainstein, Nina; Touloumi, Olga; Lagoudaki, Roza; Hanya, Ester; Grigoriadis, Nikolaos; Katz, Abram; Ben-Hur, Tamir

2018-01-01

Conflicting results exist on the effects of exercise training (ET) on Experimental Autoimmune Encephalomyelitis (EAE), nor is it known how exercise impacts on disease progression. We examined whether ET ameliorates the development of EAE by modulating the systemic immune system or exerting direct neuroprotective effects on the CNS. Healthy mice were subjected to 6weeks of motorized treadmill running. The Proteolipid protein (PLP)-induced transfer EAE model in mice was utilized. To assess effects of ET on systemic autoimmunity, lymph-node (LN)-T cells from trained- vs. sedentary donor mice were transferred to naïve recipients. To assess direct neuroprotective effects of ET, PLP-reactive LN-T cells were transferred into recipient mice that were trained prior to EAE transfer or to sedentary mice. EAE severity was assessed in vivo and the characteristics of encephalitogenic LN-T cells derived from PLP-immunized mice were evaluated in vitro. LN-T cells obtained from trained mice induced an attenuated clinical and pathological EAE in recipient mice vs. cells derived from sedentary animals. Training inhibited the activation, proliferation and cytokine gene expression of PLP-reactive T cells in response to CNS-derived autoantigen, but strongly enhanced their proliferation in response to Concanavalin A, a non-specific stimulus. However, there was no difference in EAE severity when autoreactive encephalitogenic T cells were transferred to trained vs. sedentary recipient mice. ET inhibits immune system responses to an auto-antigen to attenuate EAE, rather than generally suppressing the immune system, but does not induce a direct neuro-protective effect against EAE. Copyright © 2017 Elsevier Inc. All rights reserved.

Consequences of missense mutations for dimerization and turnover of alanine:glyoxylate aminotransferase: study of a spectrum of mutations.

PubMed

Coulter-Mackie, M B; Lian, Q

2006-12-01

Alanine:glyoxylate aminotransferase (AGT) is a liver peroxisomal enzyme, deficiency of which results in primary hyperoxaluria type 1 (PH1). More than 65 PH1-related mutations are now documented in the AGT gene (AGXT), of which about 50% are missense. We have generated a spectrum of 15 missense changes including the most common PH1 mutation, G170R, and expressed them on the appropriate background of the major or minor allele, in an Escherichia coli overexpression system and in a rabbit reticulocyte transcription/translation system. We have investigated their effects on enzyme activity, dimerization, aggregation, and turnover. The effect of pyridoxal phosphate (PLP) on dimerization and stability was also investigated. Although all 15 mutant AGTs were expressed as intact proteins in E. coli, only three: G41R and G41V on the major allele, and the common mutation G170R, resulted in significant amounts of enzymatic activity. Dimerization failure was a frequent observation (13/15) except for G41V and D183N. Dimerization was poor with S187F but was substantially improved with PLP. Proteasome-mediated protein degradation was observed for all the mutations except G41R on the major allele, G41V, D183N, G170R, and S218L. Increases in the stability of the mutant enzymes in the presence of PLP were small; however, G41R on the minor allele showed a direct relationship between its half life and the concentration of PLP. The minor allele AGT product and many of the mutants were subject to a limited non-proteasomal proteolytic cleavage when ATP was depleted.

Novel variants in PAX6 gene caused congenital aniridia in two Chinese families.

PubMed

Zhang, R; Linpeng, S; Wei, X; Li, H; Huang, Y; Guo, J; Wu, Q; Liang, D; Wu, L

2017-06-01

PurposeTo reveal the underlying genetic defect in two four-generation Chinese families with aniridia and explore the pathologic mechanism.MethodsFull ophthalmic examinations were performed in two families with aniridia. The PAX6 gene was directly sequenced in patients of two families, and the detected variants were screened in unaffected family members and two hundred unrelated healthy controls. Real-time quantitative PCR was used to explore pathologic mechanisms of the two variants.ResultsAniridia, cataract, and oscillatory nystagmus were observed in patients of the two families. In addition, we observed corneal opacity and microphthalmus in family 1, and strabismus, left ectopia lentis, microphthalmus, and microcornea in family 2. Sanger sequencing detected a novel 1-bp duplication (c.50dupA) in family 1 and a novel 2-bp splice site deletion (c.765+1_765+2delGT) in family 2. Sequencing of cDNA indicated skipping of exon 9 caused by the splice site deletion, being predicted to cause a premature stop codon, as well as the duplication. The PAX6 mRNA significantly lower in patients with aniridia than in unaffected family members in both families, suggesting that the duplication and splice site deletion caused nonsense-mediated mRNA decay.ConclusionsOur study identified two novel PAX6 variants in two families with aniridia and revealed the pathogenicity of the variants; this would expand the variant spectrum of PAX6 and help us better understand the molecular basis of aniridia, thus facilitating genetic counseling.

Mobile group II intron based gene targeting in Lactobacillus plantarum WCFS1.

PubMed

Sasikumar, Ponnusamy; Paul, Eldho; Gomathi, Sivasamy; Abhishek, Albert; Sasikumar, Sundaresan; Selvam, Govindan Sadasivam

2016-10-01

The usage of recombinant lactic acid bacteria for delivery of therapeutic proteins to the mucosa has been emerging. In the present study, an attempt was made to engineer a thyA mutant of Lactobacillus plantarum (L. plantarum) using lactococcal group II intron Ll.LtrB for the development of biologically contained recombinant L. plantarum for prevention of calcium oxalate stone disease. The 3 kb Ll.LtrB intron donor cassettes from the source vector pACD4C was PCR amplified, ligated into pSIP series of lactobacillus vector pLp_3050sAmyA, yielding a novel vector pLpACD4C (8.6 kb). The quantitative real-time PCR experiment shows 94-fold increased expression of Ll.LtrB intron and 14-fold increased expression of ltrA gene in recombinant L. plantarum containing pLpACD4C. In order to target the thyA gene, the potential intron RNA binding sites in the thyA gene of L. plantarum was predicted with help of computer algorithm. The insertion location 188|189s of thyA gene (lowest E-0.134) was chosen and the wild type intron Ll.LtrB was PCR modified, yielding a retargeted intron of pLpACDthyA. The retargeted intron was expressed by using induction peptide (sppIP), subsequently the integration of intron in thyA gene was identified by PCR screening and finally ThyA - mutant of L. plantarum (ThyA18) was detected. In vitro growth curve result showed that in the absence of thymidine, colony forming units of mutant ThyA18 was decreased, whereas high thymidine concentration (10 μM) supported the growth of the culture until saturation. In conclusion, ThyA - mutant of L. plantarum (ThyA18) constructed in this study will be used as a biologically contained recombinant probiotic to deliver oxalate decarboxylase into the lumen for treatment of hyperoxaluria and calcium oxalate stone deposition. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Exposure to Nickel, Chromium, or Cadmium Causes Distinct Changes in the Gene Expression Patterns of a Rat Liver Derived Cell Line

DTIC Science & Technology

2011-11-16

protein A (Rpa2), the minichromosome maintenance complex component genes which encode helicases, DNA ligase (Lig1), DNA polymerase e ( Pole and Pole2...and DNA polymerase d ( Pold1 and Pold2 ) are all up-regulated as a result of exposure to chromium (Figure 6), suggesting that there is an increase in...Exposure to Nickel, Chromium, or Cadmium Causes Distinct Changes in the Gene Expression Patterns of a Rat Liver Derived Cell Line Matthew G

Prison Literacy Project Handbook.

ERIC Educational Resources Information Center

Kops, Joan, Ed.

This handbook records the creation, development and growth, and stumbling blocks and successes of the Prison Literacy Project (PLP). It is intended to serve as a model for other community groups that are developing their own literacy projects. The handbook provides a history and philosophy of PLP, states PLP's vision and purpose, discusses need,…

Homozygosity mapping reveals new nonsense mutation in the FAM161A gene causing autosomal recessive retinitis pigmentosa in a Palestinian family.

PubMed

Zobor, Ditta; Balousha, Ghassan; Baumann, Britta; Wissinger, Bernd

2014-01-01

Retinitis pigmentosa (RP) is a heterogenous group of inherited retinal degenerations caused by mutations in at least 45 genes. Recently, the FAM161A gene was identified as the causative gene for RP28, an autosomal recessive form of RP. We performed a clinical and molecular genetic study of a consanguineous Palestinian family with two three siblings affected with retinitis pigmentosa. DNA samples were collected from the index patient, his father, his affected sister, and two non-affected brothers. DNA sample from the index was subjected to high resolution genome-wide SNP array. Assuming identity-by-descent in this consanguineous family we applied homozygosity mapping to identify disease causing genes. The index patient reported night blindness since the age of 20 years, followed by moderate disease progression with decrease of peripheral vision, the development of photophobia and later on reduced central vision. At the age of 40 his visual acuity was counting fingers (CF) for both eyes, color discrimination was not possible and his visual fields were severely constricted. Funduscopic examination revealed a typical appearance of advanced RP with optic disc pallor, narrowed retinal vessels, bone-spicule like pigmentary changes in the mid-periphery and atrophic changes in the macula. His younger affected brother (37 years) was reported with overall milder symptoms, while the youngest sister (21 years) reported problems only with night vision. Applying high-density SNP arrays we identified several homozygous genomic regions one of which included the recently identified FAM161A gene mutated in RP28-linked autosomal recessive RP. Sequencing analysis revealed the presence of a novel homozygous nonsense mutation, c.1003C>T/p.R335X in the index patient and the affected sister. We identified an RP28-linked RP family in the Palestinian population caused by a novel nonsense mutation in FAM161A. RP in this family shows a typical disease onset with moderate to rapid progression

Homozygosity mapping reveals new nonsense mutation in the FAM161A gene causing autosomal recessive retinitis pigmentosa in a Palestinian family

PubMed Central

Zobor, Ditta; Balousha, Ghassan; Baumann, Britta

2014-01-01

Purpose: Retinitis pigmentosa (RP) is a heterogenous group of inherited retinal degenerations caused by mutations in at least 45 genes. Recently, the FAM161A gene was identified as the causative gene for RP28, an autosomal recessive form of RP. Methods: We performed a clinical and molecular genetic study of a consanguineous Palestinian family with two three siblings affected with retinitis pigmentosa. DNA samples were collected from the index patient, his father, his affected sister, and two non-affected brothers. DNA sample from the index was subjected to high resolution genome-wide SNP array. Assuming identity-by-descent in this consanguineous family we applied homozygosity mapping to identify disease causing genes. Results: The index patient reported night blindness since the age of 20 years, followed by moderate disease progression with decrease of peripheral vision, the development of photophobia and later on reduced central vision. At the age of 40 his visual acuity was counting fingers (CF) for both eyes, color discrimination was not possible and his visual fields were severely constricted. Funduscopic examination revealed a typical appearance of advanced RP with optic disc pallor, narrowed retinal vessels, bone-spicule like pigmentary changes in the mid-periphery and atrophic changes in the macula. His younger affected brother (37 years) was reported with overall milder symptoms, while the youngest sister (21 years) reported problems only with night vision. Applying high-density SNP arrays we identified several homozygous genomic regions one of which included the recently identified FAM161A gene mutated in RP28-linked autosomal recessive RP. Sequencing analysis revealed the presence of a novel homozygous nonsense mutation, c.1003C>T/p.R335X in the index patient and the affected sister. Conclusion: We identified an RP28-linked RP family in the Palestinian population caused by a novel nonsense mutation in FAM161A. RP in this family shows a typical disease

Gitelman or Bartter type 3 syndrome? A case of distal convoluted tubulopathy caused by CLCNKB gene mutation

PubMed Central

Cruz, António José; Castro, Alexandra

2013-01-01

A 32-year-old woman with no significant medical history was sent to our consultation due to hypokalaemia (<3.0 mmol/l). Her main complaints were longstanding polyuria and nocturia. Physical examination was normal. Basic investigations showed normal renal function, low serum potassium (2.7 mmol/l) and magnesium (0.79 mmol/l), metabolic alkalosis (pH 7.54; bicarbonate 32.5 mmol/l), elevated urinary potassium (185 mmol/24 h) and normal urinary calcium (246 mg/24 h). Thiazide test revealed blunted response. Chronic vomiting and the abuse of diuretics were excluded. Genetic tests for SLC12A3 gene mutation described in Gitelman syndrome (GS) came negative. CLCNKB gene mutation analysis present in both GS and Bartter (BS) type 3 syndromes was positive. The patient is now being treated with potassium and magnesium oral supplements, ramipril and spironolactone with stable near-normal potassium and magnesium levels. This article presents the case of a patient with hypokalaemia caused by CLCNKB gene mutation hard to categorise as GS or BS type 3. PMID:23345488

Gitelman or Bartter type 3 syndrome? A case of distal convoluted tubulopathy caused by CLCNKB gene mutation.

PubMed

Cruz, António José; Castro, Alexandra

2013-01-22

A 32-year-old woman with no significant medical history was sent to our consultation due to hypokalaemia (<3.0 mmol/l). Her main complaints were longstanding polyuria and nocturia. Physical examination was normal. Basic investigations showed normal renal function, low serum potassium (2.7 mmol/l) and magnesium (0.79 mmol/l), metabolic alkalosis (pH 7.54; bicarbonate 32.5 mmol/l), elevated urinary potassium (185 mmol/24 h) and normal urinary calcium (246 mg/24 h). Thiazide test revealed blunted response. Chronic vomiting and the abuse of diuretics were excluded. Genetic tests for SLC12A3 gene mutation described in Gitelman syndrome (GS) came negative. CLCNKB gene mutation analysis present in both GS and Bartter (BS) type 3 syndromes was positive. The patient is now being treated with potassium and magnesium oral supplements, ramipril and spironolactone with stable near-normal potassium and magnesium levels. This article presents the case of a patient with hypokalaemia caused by CLCNKB gene mutation hard to categorise as GS or BS type 3.

Silencing of the ACC synthase gene ACACS2 causes delayed flowering in pineapple [Ananas comosus (L.) Merr.].

PubMed

Trusov, Yuri; Botella, José Ramón

2006-01-01

Flowering is a crucial developmental stage in the plant life cycle. A number of different factors, from environmental to chemical, can trigger flowering. In pineapple, and other bromeliads, it has been proposed that flowering is triggered by a small burst of ethylene production in the meristem in response to environmental cues. A 1-amino-cyclopropane-1-carboxylate synthase (ACC synthase) gene has been cloned from pineapple (ACACS2), which is induced in the meristem under the same environmental conditions that induce flowering. Two transgenic pineapple lines have been produced containing co-suppression constructs designed to down-regulate the expression of the ACACS2 gene. Northern analysis revealed that the ACACS2 gene was silenced in a number of transgenic plants in both lines. Southern hybridization revealed clear differences in the methylation status of silenced versus non-silenced plants by the inability of a methylation-sensitive enzyme to digest within the ACACS2 DNA extracted from silenced plants, indicating that methylation is the cause of the observed co-suppression of the ACACS2 gene. Flowering characteristics of the transgenic plants were studied under field conditions in South East Queensland, Australia. Flowering dynamics studies revealed significant differences in flowering behaviour, with transgenic plants exhibiting silencing showing a marked delay in flowering when compared with non-silenced transgenic plants and control non-transformed plants. It is argued that the ACACS2 gene is one of the key contributors towards triggering 'natural flowering' in mature pineapples under commercial field conditions.

Hyperthyroidism caused by a germline activating mutation of the thyrotropin receptor gene: difficulties in diagnosis and therapy.

PubMed

Bertalan, Rita; Sallai, Agnes; Sólyom, János; Lotz, Gábor; Szabó, István; Kovács, Balázs; Szabó, Eva; Patócs, Attila; Rácz, Károly

2010-03-01

Germline activating mutations of the thyrotropin receptor (TSHR) gene have been considered as the only known cause of sporadic nonautoimmune hyperthyroidism in the pediatric population. Here we describe the long-term follow-up and evaluation of a patient with sporadic nonautoimmune primary hyperthyroidism who was found to have a de novo germline activating mutation of the TSHR gene. The patient was an infant who presented at the age of 10 months in an unconscious state with exsiccation, wet skin, fever, and tachycardia. Nonautoimmune primary hyperthyroidism was diagnosed, and brain magnetic resonance imaging and computed tomography showed also Arnold-Chiari malformation type I. Continuous propylthiouracil treatment resulted in a prolonged clinical cure lasting for 10 years. At the age of 11 years and 5 months the patient underwent subtotal thyroidectomy because of symptoms of trachea compression caused by a progressive multinodular goiter. However, 2 months after surgery, hormonal evaluation indicated recurrent hyperthyroidism and the patient was treated with propylthiouracil during the next 4 years. At the age of 15 years the patient again developed symptoms of trachea compression. Radioiodine treatment resulted in a regression of the recurrent goiter and a permanent cure of hyperthyroidism without relapse during the last 3 years of his follow-up. Sequencing of exon 10 of the TSHR gene showed a de novo heterozygous germline I630L mutation, which has been previously described as activating mutation at somatic level in toxic thyroid nodules. The I630L mutation of the TSHR gene occurs not only at somatic level in toxic thyroid nodules, but also its presence in germline is associated with nonautoimmune primary hyperthyroidism. Our case report demonstrates that in this disorder a continuous growth of the thyroid occurs without any evidence of elevated TSH due to antithyroid drug overdosing. This may justify previous recommendations for early treatment of affected

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

Salmonella typhimurium PtsJ is a novel MocR-like transcriptional repressor involved in regulating the vitamin B6 salvage pathway.

PubMed

Tramonti, Angela; Milano, Teresa; Nardella, Caterina; di Salvo, Martino L; Pascarella, Stefano; Contestabile, Roberto

2017-02-01

The vitamin B 6 salvage pathway, involving pyridoxine 5'-phosphate oxidase (PNPOx) and pyridoxal kinase (PLK), recycles B 6 vitamers from nutrients and protein turnover to produce pyridoxal 5'-phosphate (PLP), the catalytically active form of the vitamin. Regulation of this pathway, widespread in living organisms including humans and many bacteria, is very important to vitamin B 6 homeostasis but poorly understood. Although some information is available on the enzymatic regulation of PNPOx and PLK, little is known on their regulation at the transcriptional level. In the present work, we identified a new MocR-like regulator, PtsJ from Salmonella typhimurium, which controls the expression of the pdxK gene encoding one of the two PLKs expressed in this organism (PLK1). Analysis of pdxK expression in a ptsJ knockout strain demonstrated that PtsJ acts as a transcriptional repressor. This is the first case of a MocR-like regulator acting as repressor of its target gene. Expression and purification of PtsJ allowed a detailed characterisation of its effector and DNA-binding properties. PLP is the only B 6 vitamer acting as effector molecule for PtsJ. A DNA-binding region composed of four repeated nucleotide sequences is responsible for binding of PtsJ to its target promoter. Analysis of binding stoichiometry revealed that protein subunits/DNA molar ratio varies from 4 : 1 to 2 : 1, depending on the presence or absence of PLP. Structural characteristics of DNA transcriptional factor-binding sites suggest that PtsJ binds DNA according to a different model with respect to other characterised members of the MocR subgroup. © 2016 Federation of European Biochemical Societies.

[Gene doping: gene transfer and possible molecular detection].

PubMed

Argüelles, Carlos Francisco; Hernández-Zamora, Edgar

2007-01-01

The use of illegal substances in sports to enhance athletic performance during competition has caused international sports organizations such as the COI and WADA to take anti doping measures. A new doping method know as gene doping is defined as "the non-therapeutic use of genes, genetic elements and/or cells that have the capacity to enhance athletic performance". However, gene doping in sports is not easily identified and can cause serious consequences. Molecular biology techniques are needed in order to distinguish the difference between a "normal" and an "altered" genome. Further, we need to develop new analytic methods and biological molecular techniques in anti-doping laboratories, and design programs that avoid the non therapeutic use of genes.

Molecular insights into primary hyperoxaluria type 1 pathogenesis.

PubMed

Cellini, Barbara; Oppici, Elisa; Paiardini, Alessandro; Montioli, Riccardo

2012-01-01

Primary hyperoxaluria type 1 (PH1) is a rare autosomal recessive disorder of glyoxylate metabolism caused by the deficiency of liver peroxisomal alanine:glyoxylate aminotransferase (AGT), a pyridoxal 5'-phosphate (PLP)-dependent enzyme. The PH1 pathogenesis is mostly due to single point mutations (more than 150 so far identified) on the AGXT gene, and is characterized by a marked heterogeneity in terms of genotype, enzymatic and clinical phenotypes. This article presents an up to date review of selected aspects of the biochemical properties of the two allelic forms of AGT and of some PH1-causing variants. These recent discoveries highlight the effects at the protein level of the pathogenic mutations, and, together with previous cell biology and clinical data, (i) improve the understanding of the molecular basis of PH1 pathogenesis, and (ii) help to delineate perspectives for predicting the response to pyridoxine treatment or for suggesting new strategies for PH1 patients bearing the analyzed mutations.

Heterozygous Germline Mutations in the CBL Tumor-Suppressor Gene Cause a Noonan Syndrome-like Phenotype

PubMed Central

Martinelli, Simone; De Luca, Alessandro; Stellacci, Emilia; Rossi, Cesare; Checquolo, Saula; Lepri, Francesca; Caputo, Viviana; Silvano, Marianna; Buscherini, Francesco; Consoli, Federica; Ferrara, Grazia; Digilio, Maria C.; Cavaliere, Maria L.; van Hagen, Johanna M.; Zampino, Giuseppe; van der Burgt, Ineke; Ferrero, Giovanni B.; Mazzanti, Laura; Screpanti, Isabella; Yntema, Helger G.; Nillesen, Willy M.; Savarirayan, Ravi; Zenker, Martin; Dallapiccola, Bruno; Gelb, Bruce D.; Tartaglia, Marco

2010-01-01

RAS signaling plays a key role in controlling appropriate cell responses to extracellular stimuli and participates in early and late developmental processes. Although enhanced flow through this pathway has been established as a major contributor to oncogenesis, recent discoveries have revealed that aberrant RAS activation causes a group of clinically related developmental disorders characterized by facial dysmorphism, a wide spectrum of cardiac disease, reduced growth, variable cognitive deficits, ectodermal and musculoskeletal anomalies, and increased risk for certain malignancies. Here, we report that heterozygous germline mutations in CBL, a tumor-suppressor gene that is mutated in myeloid malignancies and encodes a multivalent adaptor protein with E3 ubiquitin ligase activity, can underlie a phenotype with clinical features fitting or partially overlapping Noonan syndrome (NS), the most common condition of this disease family. Independent CBL mutations were identified in two sporadic cases and two families from among 365 unrelated subjects who had NS or suggestive features and were negative for mutations in previously identified disease genes. Phenotypic heterogeneity and variable expressivity were documented. Mutations were missense changes altering evolutionarily conserved residues located in the RING finger domain or the linker connecting this domain to the N-terminal tyrosine kinase binding domain, a known mutational hot spot in myeloid malignancies. Mutations were shown to affect CBL-mediated receptor ubiquitylation and dysregulate signal flow through RAS. These findings document that germline mutations in CBL alter development to cause a clinically variable condition that resembles NS and that possibly predisposes to malignancies. PMID:20619386

Evidence that the lung Adenocarcinoma EML4-ALK fusion gene is not caused by exposure to secondhand tobacco smoke during childhood.

PubMed

Ryan, Bríd M; Wang, Yi; Jen, Jin; Yi, Eunhee S; Olivo-Marston, Susan; Yang, Ping; Harris, Curtis C

2014-07-01

The EML4-ALK fusion gene is more frequently found in younger, never smoking patients with lung cancer. Meanwhile, never smokers exposed to secondhand tobacco smoke (SHS) during childhood are diagnosed at a younger age compared with never smoking patients with lung cancer who are not exposed. We, therefore, hypothesized that SHS, which can induce DNA damage, is associated with the EML4-ALK fusion gene. We compared the frequency of the EML4-ALK fusion gene among 197 never smoker patients with lung cancer with and without a history of exposure to SHS during childhood at Mayo Clinic. The EML4-ALK fusion gene was detected in 33% of cases from never smokers with a history of SHS exposure during childhood, whereas 47% of never smoking lung cancer cases without a history of childhood SHS exposure tested positive for the fusion gene. The EML4-ALK fusion gene is not enriched in tumors from individuals exposed to SHS during childhood. These data suggest that childhood exposure to SHS is not a significant etiologic cause of the EML4-ALK fusion gene in lung cancer. ©2014 American Association for Cancer Research.

A novel A781V mutation in the CSF1R gene causes hereditary diffuse leucoencephalopathy with axonal spheroids☆

PubMed Central

Ahmed, Rebekah; Guerreiro, Rita; Rohrer, Jonathan D.; Guven, Gamze; Rossor, Martin N.; Hardy, John; Fox, Nick C.

2013-01-01

We report a family with a novel CSF1R mutation causing hereditary diffuse leucoencephalopathy with axonal spheroids. Family members presented with neuropsychiatric and behavioural symptoms, with subsequent development of motor symptoms and gait disturbance. MRI brain showed extensive white matter change with a frontal predominance and associated atrophy in two members of the family. Genetic testing revealed a novel mutation c.2342C > T (p.A781V) in the CSF1R gene in two brothers of the family. This report highlights the difficulties in diagnosing HDLS and discusses the indications for testing for mutations in the CSF1R gene. PMID:23816250

Ablation of XP-V gene causes adipose tissue senescence and metabolic abnormalities

PubMed Central

Chen, Yih-Wen; Harris, Robert A.; Hatahet, Zafer; Chou, Kai-ming

2015-01-01

Obesity and the metabolic syndrome have evolved to be major health issues throughout the world. Whether loss of genome integrity contributes to this epidemic is an open question. DNA polymerase η (pol η), encoded by the xeroderma pigmentosum (XP-V) gene, plays an essential role in preventing cutaneous cancer caused by UV radiation-induced DNA damage. Herein, we demonstrate that pol η deficiency in mice (pol η−/−) causes obesity with visceral fat accumulation, hepatic steatosis, hyperleptinemia, hyperinsulinemia, and glucose intolerance. In comparison to WT mice, adipose tissue from pol η−/− mice exhibits increased DNA damage and a greater DNA damage response, indicated by up-regulation and/or phosphorylation of ataxia telangiectasia mutated (ATM), phosphorylated H2AX (γH2AX), and poly[ADP-ribose] polymerase 1 (PARP-1). Concomitantly, increased cellular senescence in the adipose tissue from pol η−/− mice was observed and measured by up-regulation of senescence markers, including p53, p16Ink4a, p21, senescence-associated (SA) β-gal activity, and SA secretion of proinflammatory cytokines interleukin 6 (IL-6) and tumor necrosis factor α (TNF-α) as early as 4 wk of age. Treatment of pol η−/− mice with a p53 inhibitor, pifithrin-α, reduced adipocyte senescence and attenuated the metabolic abnormalities. Furthermore, elevation of adipocyte DNA damage with a high-fat diet or sodium arsenite exacerbated adipocyte senescence and metabolic abnormalities in pol η−/− mice. In contrast, reduction of adipose DNA damage with N-acetylcysteine or metformin ameliorated cellular senescence and metabolic abnormalities. These studies indicate that elevated DNA damage is a root cause of adipocyte senescence, which plays a determining role in the development of obesity and insulin resistance. PMID:26240351

Functional architecture of two exclusively late stage pollen-specific promoters in rice (Oryza sativa L.).

PubMed

Yan, Shuo; Wang, Zhongni; Liu, Yuan; Li, Wei; Wu, Feng; Lin, Xuelei; Meng, Zheng

2015-07-01

Late stage pollen-specific promoters are important tools in crop molecular breeding. Several such promoters, and their functional motifs, have been well characterized in dicotyledonous plants such as tomato and tobacco. However, knowledge about the functional architecture of such promoters is limited in the monocotyledonous plant rice. Here, pollen-late-stage-promoter 1 (PLP1) and pollen-late-stage-promoter 2 (PLP2) were characterized using a stable transformation system in rice. Histochemical staining showed that the two promoters exclusively drive GUS expression in late-stage pollen grains in rice. 5' deletion analysis revealed that four regions, including the -1159 to -720 and the -352 to -156 regions of PLP1 and the -740 to -557 and the -557 to -339 regions of PLP2, are important in maintaining the activity and specificity of these promoters. Motif mutation analysis indicated that 'AGAAA' and 'CAAT' motifs in the -740 to -557 region of PLP2 act as enhancers in the promoter. Gain of function experiments indicated that the novel TA-rich motif 'TACATAA' and 'TATTCAT' in the core region of the PLP1 and PLP2 promoters is necessary, but not sufficient, for pollen-specific expression in rice. Our results provide evidence that the enhancer motif 'AGAAA' is conserved in the pollen-specific promoters of both monocots and eudicots, but that some functional architecture characteristics are different.

Permanent neonatal diabetes caused by a homozygous nonsense mutation in the glucokinase gene.

PubMed

Rubio-Cabezas, O; Díaz González, F; Aragonés, A; Argente, J; Campos-Barros, A

2008-06-01

Glucokinase deficiency is an unfrequent cause of permanent neonatal diabetes (PND), as only seven patients have been reported, either homozygous for a missense or frameshift mutation or compound heterozygous for both of them. We report here the first known case caused by a homozygous nonsense mutation (Y61X) in the glucokinase gene (GCK) that introduces a premature stop codon, generating a truncated protein that is predicted to be completely inactive as it lacks both the glucose- and the adenosine triphosphate-binding sites. The proband, born to consanguineous parents, was a full-term, intra-uterine growth-retarded male newborn who presented with a glycaemia of 129 mg/dL (7.16 mmol/L) on his second day of life, increasing thereafter up to 288 mg/dL (15.98 mmol/L) and 530 mg/dL (29.41 mmol/L) over the next 24 h, in the face of low serum insulin (<3 muIU/mL; <20.83 pmol/L). He was put on insulin on the third day of life. Insulin has never been discontinued since then. The patient was tested negative for anti-insulin and islet cell antibodies at age 5 months. His father had non-progressive, impaired fasting glucose for several years. The mother was found to be mildly hyperglycaemic only when her glucose was checked after the child was diagnosed. In conclusion, biallelic GCK loss should be considered as a potential cause of PND in children born to consanguineous parents, even if they are not known to be diabetic at the time of PND presentation.

Congenital analbuminemia caused by a novel aberrant splicing in the albumin gene

PubMed Central

Caridi, Gianluca; Dagnino, Monica; Erdeve, Omer; Di Duca, Marco; Yildiz, Duran; Alan, Serdar; Atasay, Begum; Arsan, Saadet; Campagnoli, Monica; Galliano, Monica; Minchiotti, Lorenzo

2014-01-01

Introduction: Congenital analbuminemia is a rare autosomal recessive disorder manifested by the presence of a very low amount of circulating serum albumin. It is an allelic heterogeneous defect, caused by variety of mutations within the albumin gene in homozygous or compound heterozygous state. Herein we report the clinical and molecular characterization of a new case of congenital analbuminemia diagnosed in a female newborn of consanguineous (first degree cousins) parents from Ankara, Turkey, who presented with a low albumin concentration (< 8 g/L) and severe clinical symptoms. Materials and methods: The albumin gene of the index case was screened by single-strand conformation polymorphism, heteroduplex analysis, and direct DNA sequencing. The effect of the splicing mutation was evaluated by examining the cDNA obtained by reverse transcriptase - polymerase chain reaction (RT-PCR) from the albumin mRNA extracted from proband’s leukocytes. Results: DNA sequencing revealed that the proband is homozygous, and both parents are heterozygous, for a novel G>A transition at position c.1652+1, the first base of intron 12, which inactivates the strongly conserved GT dinucleotide at the 5′ splice site consensus sequence of this intron. The splicing defect results in the complete skipping of the preceding exon (exon 12) and in a frame-shift within exon 13 with a premature stop codon after the translation of three mutant amino acid residues. Conclusions: Our results confirm the clinical diagnosis of congenital analbuminemia in the proband and the inheritance of the trait and contribute to shed light on the molecular genetics of analbuminemia. PMID:24627724

Congenital analbuminemia caused by a novel aberrant splicing in the albumin gene.

PubMed

Caridi, Gianluca; Dagnino, Monica; Erdeve, Omer; Di Duca, Marco; Yildiz, Duran; Alan, Serdar; Atasay, Begum; Arsan, Saadet; Campagnoli, Monica; Galliano, Monica; Minchiotti, Lorenzo

2014-01-01

Congenital analbuminemia is a rare autosomal recessive disorder manifested by the presence of a very low amount of circulating serum albumin. It is an allelic heterogeneous defect, caused by variety of mutations within the albumin gene in homozygous or compound heterozygous state. Herein we report the clinical and molecular characterization of a new case of congenital analbuminemia diagnosed in a female newborn of consanguineous (first degree cousins) parents from Ankara, Turkey, who presented with a low albumin concentration (< 8 g/L) and severe clinical symptoms. The albumin gene of the index case was screened by single-strand conformation polymorphism, heteroduplex analysis, and direct DNA sequencing. The effect of the splicing mutation was evaluated by examining the cDNA obtained by reverse transcriptase - polymerase chain reaction (RT-PCR) from the albumin mRNA extracted from proband's leukocytes. DNA sequencing revealed that the proband is homozygous, and both parents are heterozygous, for a novel G>A transition at position c.1652+1, the first base of intron 12, which inactivates the strongly conserved GT dinucleotide at the 5' splice site consensus sequence of this intron. The splicing defect results in the complete skipping of the preceding exon (exon 12) and in a frame-shift within exon 13 with a premature stop codon after the translation of three mutant amino acid residues. Our results confirm the clinical diagnosis of congenital analbuminemia in the proband and the inheritance of the trait and contribute to shed light on the molecular genetics of analbuminemia.

Multiple Genes Cause Postmating Prezygotic Reproductive Isolation in the Drosophila virilis Group.

PubMed

Ahmed-Braimah, Yasir H

2016-12-07

Understanding the genetic basis of speciation is a central problem in evolutionary biology. Studies of reproductive isolation have provided several insights into the genetic causes of speciation, especially in taxa that lend themselves to detailed genetic scrutiny. Reproductive barriers have usually been divided into those that occur before zygote formation (prezygotic) and after (postzygotic), with the latter receiving a great deal of attention over several decades. Reproductive barriers that occur after mating but before zygote formation [postmating prezygotic (PMPZ)] are especially understudied at the genetic level. Here, I present a phenotypic and genetic analysis of a PMPZ reproductive barrier between two species of the Drosophila virilis group: D. americana and D. virilis This species pair shows strong PMPZ isolation, especially when D. americana males mate with D. virilis females: ∼99% of eggs laid after these heterospecific copulations are not fertilized. Previous work has shown that the paternal loci contributing to this incompatibility reside on two chromosomes, one of which (chromosome 5) likely carries multiple factors. The other (chromosome 2) is fixed for a paracentric inversion that encompasses nearly half the chromosome. Here, I present two results. First, I show that PMPZ in this species cross is largely due to defective sperm storage in heterospecific copulations. Second, using advanced intercross and backcross mapping approaches, I identify genomic regions that carry genes capable of rescuing heterospecific fertilization. I conclude that paternal incompatibility between D. americana males and D. virilis females is underlain by four or more genes on chromosomes 2 and 5. Copyright © 2016 Ahmed-Braimah.

Functional vitamin B-6 status and long-term mortality in renal transplant recipients.

PubMed

Minović, Isidor; van der Veen, Anna; van Faassen, Martijn; Riphagen, Ineke J; van den Berg, Else; van der Ley, Claude; Gomes-Neto, António W; Geleijnse, Johanna M; Eggersdorfer, Manfred; Navis, Gerjan J; Kema, Ido P; Bakker, Stephan Jl

2017-12-01

Background: Low plasma concentrations of pyridoxal 5'-phosphate (PLP) are common in renal transplant recipients (RTRs) and confer increased risk of long-term mortality. To our knowledge, it is not known whether low plasma PLP concentrations have functional (i.e., intracellular) consequences and, if so, whether such consequences are associated with increased risk of mortality. Objectives: We assessed the association of plasma PLP with functional vitamin B-6 status and explored the potential association of functional vitamin B-6 status with long-term mortality in RTRs. Design: In a longitudinal cohort of 678 stable RTRs with a median follow-up of 5.3 y (IQR: 4.8-6.1 y) and 297 healthy controls, PLP, plasma 3-hydroxykynurenine (3-HK), and xanthurenic acid (XA) were analyzed via validated assays. PLP was used as direct biomarker for vitamin B-6 status, and the 3-HK:XA ratio was used as functional biomarker of vitamin B-6 status with a higher ratio reflecting worse functional vitamin B-6 status. Results: Median PLP, 3-HK, and XA concentrations were 41 nmol/L (IQR: 29-60 nmol/L), 40.1 nmol/L (IQR: 33.0-48.0 nmol/L), and 19.1 nmol/L (IQR: 14.5-24.9 nmol/L), respectively, in healthy controls compared with 29 nmol/L (IQR: 17-50 nmol/L), 61.5 nmol/L (IQR: 45.6-86.5 nmol/L), and 25.5 nmol/L (IQR: 17.2-40.0 nmol/L), respectively, in RTRs (all P < 0.001). RTRs had a higher median 3-HK:XA ratio (2.38; IQR: 1.68-3.49) than did healthy controls (2.13; IQR: 1.63-2.71) ( P < 0.05). In RTRs, the 3-HK:XA ratio was inversely associated with plasma PLP (β = -0.21, P < 0.001). Moreover, a higher 3-HK:XA ratio was independently associated with increased risk of all-cause mortality (HR per SD increment: 1.30; 95% CI: 1.13, 1.49), cancer mortality (HR per SD increment: 1.47; 95% CI: 1.12, 1.95), and infectious disease mortality (HR per SD increment: 1.50; 95% CI: 1.21, 1.86) in RTRs. Conclusions: Vitamin B-6-deficient RTRs have a worse functional vitamin B-6 status than do healthy controls

UMD-USHbases: a comprehensive set of databases to record and analyse pathogenic mutations and unclassified variants in seven Usher syndrome causing genes.

PubMed

Baux, David; Faugère, Valérie; Larrieu, Lise; Le Guédard-Méreuze, Sandie; Hamroun, Dalil; Béroud, Christophe; Malcolm, Sue; Claustres, Mireille; Roux, Anne-Françoise

2008-08-01

Using the Universal Mutation Database (UMD) software, we have constructed "UMD-USHbases", a set of relational databases of nucleotide variations for seven genes involved in Usher syndrome (MYO7A, CDH23, PCDH15, USH1C, USH1G, USH3A and USH2A). Mutations in the Usher syndrome type I causing genes are also recorded in non-syndromic hearing loss cases and mutations in USH2A in non-syndromic retinitis pigmentosa. Usher syndrome provides a particular challenge for molecular diagnostics because of the clinical and molecular heterogeneity. As many mutations are missense changes, and all the genes also contain apparently non-pathogenic polymorphisms, well-curated databases are crucial for accurate interpretation of pathogenicity. Tools are provided to assess the pathogenicity of mutations, including conservation of amino acids and analysis of splice-sites. Reference amino acid alignments are provided. Apparently non-pathogenic variants in patients with Usher syndrome, at both the nucleotide and amino acid level, are included. The UMD-USHbases currently contain more than 2,830 entries including disease causing mutations, unclassified variants or non-pathogenic polymorphisms identified in over 938 patients. In addition to data collected from 89 publications, 15 novel mutations identified in our laboratory are recorded in MYO7A (6), CDH23 (8), or PCDH15 (1) genes. Information is given on the relative involvement of the seven genes, the number and distribution of variants in each gene. UMD-USHbases give access to a software package that provides specific routines and optimized multicriteria research and sorting tools. These databases should assist clinicians and geneticists seeking information about mutations responsible for Usher syndrome.

NK cells are intrinsically functional in pigs with Severe Combined Immunodeficiency (SCID) caused by spontaneous mutations in the Artemis gene

USDA-ARS?s Scientific Manuscript database

We have identified Severe Combined Immunodeficiency (SCID) in a line of Yorkshire pigs at Iowa State University. These SCID pigs lack B-cells and T-cells, but possess Natural Killer (NK) cells. This SCID phenotype is caused by recessive mutations in the Artemis gene. Interestingly, two human tumor c...

Mirror therapy for phantom limb pain: brain changes and the role of body representation.

PubMed

Foell, J; Bekrater-Bodmann, R; Diers, M; Flor, H

2014-05-01

Phantom limb pain (PLP) is a common consequence of amputation and is difficult to treat. Mirror therapy (MT), a procedure utilizing the visual recreation of movement of a lost limb by moving the intact limb in front of a mirror, has been shown to be effective in reducing PLP. However, the neural correlates of this effect are not known. We investigated the effects of daily mirror training over 4 weeks in 13 chronic PLP patients after unilateral arm amputation. Eleven participants performed hand and lip movements during a functional magnetic resonance imaging (fMRI) measurement before and after MT. The location of neural activity in primary somatosensory cortex during these tasks was used to assess brain changes related to treatment. The treatment caused a significant reduction of PLP (average decrease of 27%). Treatment effects were predicted by a telescopic distortion of the phantom, with those patients who experienced a telescope profiting less from treatment. fMRI data analyses revealed a relationship between change in pain after MT and a reversal of dysfunctional cortical reorganization in primary somatosensory cortex. Pain reduction after mirror training was also related to a decrease of activity in the inferior parietal cortex (IPC). Experienced body appearance seems to be an important predictor of mirror treatment effectiveness. Maladaptive changes in cortical organization are reversed during mirror treatment, which also alters activity in the IPC, a region involved in painful perceptions and in the perceived relatedness to an observed limb. © 2013 The Authors. European Journal of Pain published by John Wiley & Sons Ltd on behalf of European Pain Federation - EFIC®.

Insufficiency of copper ion homeostasis causes freeze-thaw injury of yeast cells as revealed by indirect gene expression analysis.

PubMed

Takahashi, Shunsuke; Ando, Akira; Takagi, Hiroshi; Shima, Jun

2009-11-01

Saccharomyces cerevisiae is exposed to freeze-thaw stress in commercial processes, including frozen dough baking. Cell viability and fermentation activity after a freeze-thaw cycle were dramatically decreased due to freeze-thaw injury. Because this type of injury involves complex phenomena, the injury mechanisms are not fully understood. We examined freeze-thaw injury by indirect gene expression analysis during postthaw incubation after freeze-thaw treatment using DNA microarray profiling. The results showed that genes involved in the homeostasis of metal ions were frequently contained in genes that were upregulated, depending on the freezing period. We assessed the phenotype of deletion mutants of the metal ion homeostasis genes that exhibited freezing period-dependent upregulation and found that the strains with deletion of the MAC1 and CTR1 genes involved in copper ion homeostasis exhibited freeze-thaw sensitivity, suggesting that copper ion homeostasis is required for freeze-thaw tolerance. We found that supplementation with copper ions during postthaw incubation increased intracellular superoxide dismutase activity and intracellular levels of reactive oxygen species were decreased. Moreover, cell viability was increased by supplementation with copper ions. These results suggest that insufficiency of copper ion homeostasis may be one of the causes of freeze-thaw injury.

JPRS Report, Science & Technology, USSR: Life Sciences.

DTIC Science & Technology

1988-02-12

polypeptide chain frag- ments inside protein membranes or on their surfaces using bacteriorhodopsin as the test object. Purple membranes, partially...outside the membrane or close to its surface . A model was developed from these data which involved folding of certain regions of bacteriorhodopsin...into hepatic endothelial and Kupffer cells. These findings point to the putative usefulness of the PLP approach in gene therapy. Figures h

Overexpression of mouse TTF-2 gene causes cleft palate

PubMed Central

Meng, Tian; Shi, Jia-Yu; Wu, Min; Wang, Yan; Li, Ling; Liu, Yan; Zheng, Qian; Huang, Lei; Shi, Bing

2012-01-01

In humans, mutations of the gene encoding for thyroid transcription factor-2 (TTF-2 or FOXE1) result in Bamforth syndrome. Bamforth syndrome is characterized by agenesis, cleft palate, spiky hair and choanal atresia. TTF-2 null mice (TTF-2−/−) also exhibit cleft palate, suggesting its involvement in the palatogenesis. However, the molecular pathology and genetic regulation by TTF2 remain largely unknown. In the present study, the recombinant expression vector pBROAD3-TTF-2 containing the promoter of the mouse ROSA26 gene was created to form the structural gene of mouse TTF-2 and was microinjected into the male pronuclei of fertilized ova. Sequence analysis confirmed that the TTF-2 transgenic mouse model was established successfully. The transgenic mice displayed a phenotype of cleft palate. In addition, we found that TTF-2 was highly expressed in the medial edge epithelium (MEE) from the embryonic day 12.5 (E12.5) to E14.5 in TTF-2 transgenic mice. These observations suggest that overexpression of TTF-2 during palatogenesis may contribute to formation of cleft palate. PMID:22304410

Two novel mutations in the sixth transmembrane segment of the thyrotropin receptor gene causing hyperfunctioning thyroid nodules.

PubMed

Gozu, Hulya; Avsar, Melike; Bircan, Rifat; Claus, Maren; Sahin, Serap; Sezgin, Ozlem; Deyneli, Oguzhan; Paschke, Ralf; Cirakoglu, Beyazit; Akalin, Sema

2005-04-01

Autonomously functioning thyroid nodules (AFTNs) can present as hyperfunctioning adenomas or toxic multinodular goiters. In the last decade, a large number of activating mutations have been identified in the thyrotropin receptor (TSHR) gene in autonomously functioning thyroid nodules. Most have been situated close to, or within the sixth transmembrane segment and third intracellular loop of the TSHR where the receptor interacts with the Gs protein. In this study we describe two novel mutations in the sixth transmembrane segment of the TSHR causing hyperfunctioning thyroid nodules. Genomic DNAs were isolated from four hyperfunctioning thyroid nodules, normal tissues and peripheral leukocytes of two patients with toxic multinodular goiter. After amplifying the related regions, TSHR and G(s)alpha genes were analyzed by single-strand conformation polymorphism (SSCP) analysis. The precise localization of the mutations was identified by automatic DNA sequence analysis. Functional studies were done by site-directed mutagenesis and transfection of a mutant construct into COS-7 cells. We identified two novel TSHR mutations in two hyperfunctioning thyroid nodules: Phe631Val in the first patient and Iso630Met in the second patient. Both mutant receptors display an increase in constitutive stimulation of basal cyclic adenosine monophosphate (cAMP) levels compared to the wild-type receptor. This confirms that these mutant receptors cause hyperfunctioning thyroid nodules.

Inflammatory peeling skin syndrome caused by homozygous genomic deletion in the PSORS1 region encompassing the CDSN gene.

PubMed

Ishida-Yamamoto, Akemi; Furio, Laetitia; Igawa, Satomi; Honma, Masaru; Tron, Elodie; Malan, Valerie; Murakami, Masamoto; Hovnanian, Alain

2014-01-01

Peeling skin syndrome (PSS) type B is a rare recessive genodermatosis characterized by lifelong widespread, reddish peeling of the skin with pruritus. The disease is caused by small-scale mutations in the Corneodesmosin gene (CDSN) leading to premature termination codons. We report for the first time a Japanese case resulting from complete deletion of CDSN. Corneodesmosin was undetectable in the epidermis, and CDSN was unamplifiable by PCR. QMPSF analysis demonstrated deletion of CDSN exons inherited from each parent. Deletion mapping using microsatellite haplotyping, CGH array and PCR analysis established that the genomic deletion spanned 49-72 kb between HCG22 and TCF19, removing CDSN as well as five other genes within the psoriasis susceptibility region 1 (PSORS1) on 6p21.33. This observation widens the spectrum of molecular defects underlying PSS type B and shows that loss of these five genes from the PSORS1 region does not result in an additional cutaneous phenotype. © 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Digenic mutational inheritance of the integrin alpha 7 and the myosin heavy chain 7B genes causes congenital myopathy with left ventricular non-compact cardiomyopathy.

PubMed

Esposito, Teresa; Sampaolo, Simone; Limongelli, Giuseppe; Varone, Antonio; Formicola, Daniela; Diodato, Daria; Farina, Olimpia; Napolitano, Filomena; Pacileo, Giuseppe; Gianfrancesco, Fernando; Di Iorio, Giuseppe

2013-06-21

We report an Italian family in which the proband showed a severe phenotype characterized by the association of congenital fiber type disproportion (CFTD) with a left ventricular non-compaction cardiomyopathy (LVNC). This study was focused on the identification of the responsible gene/s. Using the whole-exome sequencing approach, we identified the proband homozygous missense mutations in two genes, the myosin heavy chain 7B (MYH7B) and the integrin alpha 7 (ITGA7). Both genes are expressed in heart and muscle tissues, and both mutations were predicted to be deleterious and were not found in the healthy population.The R890C mutation in the MYH7B gene segregated with the LVNC phenotype in the examined family. It was also found in one unrelated patient affected by LVNC, confirming a causative role in cardiomyopathy.The E882K mutation in the ITGA7 gene, a key component of the basal lamina of muscle fibers, was found only in the proband, suggesting a role in CFTD. This study identifies two novel disease genes. Mutation in MYH7B causes a classical LVNC phenotype, whereas mutation in ITGA7 causes CFTD. Both phenotypes represent alterations of skeletal and cardiac muscle maturation and are usually not severe. The severe phenotype of the proband is most likely due to a synergic effect of these two mutations.This study provides new insights into the genetics underlying Mendelian traits and demonstrates a role for digenic inheritance in complex phenotypes.

A novel mutation in the EDAR gene causes severe autosomal recessive hypohidrotic ectodermal dysplasia.

PubMed

Henningsen, Emil; Svendsen, Mathias Tiedemann; Lildballe, Dorte Launholt; Jensen, Peter Kjestrup Axel

2014-08-01

We report on a 2-year-old girl presenting with a severe form of hypohidrotic ectodermal dysplasia (HED). The patient presented with hypotrichosis, anodontia, hypohidrosis, frontal bossing, prominent lips and ears, dry, pale skin, and dermatitis. The patient had chronic rhinitis with malodorous nasal discharge. The girl was the second born child of first-cousin immigrants from Northern Iraq. A novel homozygous mutation (c.84delC) in the EDAR gene was identified. This mutation most likely causes a frameshift in the protein product (p.S29fs*74). This results in abolition of all ectodysplasin-mediated NF-kB signalling. This complete loss-of-function mutation likely accounts for the severe clinical abnormalities in ectodermal structures in the described patient. © 2014 Wiley Periodicals, Inc.

Two new mutations in the glucose-6-phosphatase gene cause glycogen storage disease in Hungarian patients.

PubMed

Parvari, R; Lei, K J; Szonyi, L; Narkis, G; Moses, S; Chou, J Y

1997-01-01

Glycogen storage disease type 1a (von Gierke disease, GSD-1A) is caused by the deficiency of microsomal glucose-6-phosphatase (G6Pase) activity which catalyzes the final common step of glycogenolysis and gluconeogenesis. The cloning of the G6Pase cDNA and characterization of the human G6Pase gene enabled the identification of the mutations causing GSD-1a. This, in turn, allows the development of non-invasive DNA-based diagnosis that provides reliable carrier testing and prenatal diagnosis. Here we report on two new mutations E110Q and D38V causing GSD-1a in two Hungarian patients. The analyses of these mutations by site-directed mutagenesis followed by transient expression assays demonstrated that E110Q retains 17% of G6Pase enzymatic activity while the D38V abolishes the enzymatic activity. The patient with the E110Q has G222R as his other mutation. G222R was also shown to preserve about 4% of the G6Pase enzymatic activity. Nevertheless, the patient presented with the classical severe symptomatology of the GSD-1a.

Short-term hyperglycaemia causes non-reversible changes in arterial gene expression in a fully 'switchable' in vivo mouse model of diabetes.

PubMed

Zervou, S; Wang, Y-F; Laiho, A; Gyenesei, A; Kytömäki, L; Hermann, R; Abouna, S; Epstein, D; Pelengaris, S; Khan, M

2010-12-01

Irreversible arterial damage due to early effects of hypo- or hyperglycaemia could account for the limited success of glucose-lowering treatments in preventing cardiovascular disease (CVD) events. We hypothesised that even brief hypo- or hyperglycaemia could adversely affect arterial gene expression and that these changes, moreover, might not be fully reversible. By controlled activation of a 'switchable' c-Myc transgene in beta cells, adult pIns-c-MycER(TAM) mice were rendered transiently hypo- and then hyperglycaemic, after which they were allowed to recover for up to 3 months. Immediate and sequential changes in aortic global gene expression from normal glycaemia through hypo- and hyperglycaemia to recovery were assessed. Gene expression was compared with that of normoglycaemic transgenic and tamoxifen-treated wild-type controls. Overall, expression of 95 genes was significantly affected by moderate hypoglycaemia (glucose down to 2.5 mmol/l), whereas over 769 genes were affected by hyperglycaemia. Genes and pathways activated included several involved in atherogenic processes, such as inflammation and arterial calcification. Although expression of many genes recovered to initial pre-exposure levels when hyperglycaemia was corrected (74.9%), in one in four genes this did not occur. Quantitative reverse transcriptase PCR and immunohistochemistry verified the gene expression patterns of key molecules, as shown by global gene arrays. Short-term exposure to hyperglycaemia can cause deleterious and persistent changes in arterial gene expression in vivo. Brief hypoglycaemia also adversely affects gene expression, although less substantially. Together, these results suggest that early correction of hyperglycaemia and avoidance of hypoglycaemia may both be necessary to avoid excess CVD risk in diabetes.

Two novel mutations in the PPIB gene cause a rare pedigree of osteogenesis imperfecta type IX.

PubMed

Jiang, Yu; Pan, Jingxin; Guo, Dongwei; Zhang, Wei; Xie, Jie; Fang, Zishui; Guo, Chunmiao; Fang, Qun; Jiang, Weiying; Guo, Yibin

2017-06-01

Osteogenesis imperfecta (OI) is a rare genetic skeletal disorder characterized by increased bone fragility and vulnerability to fractures. PPIB is identified as a candidate gene for OI-IX, here we detect two pathogenic mutations in PPIB and analyze the genotype-phenotype correlation in a Chinese family with OI. Next-generation sequencing (NGS) was used to screen the whole exome of the parents of proband. Screening of variation frequency, evolutionary conservation comparisons, pathogenicity evaluation, and protein structure prediction were conducted to assess the pathogenicity of the novel mutations. Sanger sequencing was used to confirm the candidate variants. RTQ-PCR was used to analyze the PPIB gene expression. All mutant genes screened out by NGS were excluded except PPIB. Two novel heterozygous PPIB mutations (father, c.25A>G; mother, c.509G>A) were identified in relation to osteogenesis imperfecta type IX. Both mutations were predicted to be pathogenic by bioinformatics analysis and RTQ-PCR analysis revealed downregulated PPIB expression in the two carriers. We report a rare pedigree with an autosomal recessive osteogenesis imperfecta type IX (OI-IX) caused by two novel PPIB mutations identified for the first time in China. The current study expands our knowledge of PPIB mutations and their associated phenotypes, and provides new information on the genetic defects associated with this disease for clinical diagnosis. Copyright © 2017 Elsevier B.V. All rights reserved.

Functional Interaction between Phosducin-like Protein 2 and Cytosolic Chaperonin Is Essential for Cytoskeletal Protein Function and Cell Cycle Progression

PubMed Central

Stirling, Peter C.; Srayko, Martin; Takhar, Karam S.; Pozniakovsky, Andrei; Hyman, Anthony A.

2007-01-01

The C haperonin Containing Tcp1 (CCT) maintains cellular protein folding homeostasis in the eukaryotic cytosol by assisting the biogenesis of many proteins, including actins, tubulins, and regulators of the cell cycle. Here, we demonstrate that the essential and conserved eukaryotic phosducin-like protein 2 (PhLP2/PLP2) physically interacts with CCT and modulates its folding activity. Consistent with this functional interaction, temperature-sensitive alleles of Saccharomyces cerevisiae PLP2 exhibit cytoskeletal and cell cycle defects. We uncovered several high-copy suppressors of the plp2 alleles, all of which are associated with G1/S cell cycle progression but which do not appreciably affect cytoskeletal protein function or fully rescue the growth defects. Our data support a model in which Plp2p modulates the biogenesis of several CCT substrates relating to cell cycle and cytoskeletal function, which together contribute to the essential function of PLP2. PMID:17429077

Screening and analysis of genes expressed upon infection of broad bean with Clover yellow vein virus causing lethal necrosis.

PubMed

Nakahara, Kenji S; Kitazawa, Hiroaki; Atsumi, Go; Choi, Sun Hee; Suzuki, Yuji; Uyeda, Ichiro

2011-07-18

Clover yellow vein virus (ClYVV) causes lethal systemic necrosis in legumes, including broad bean (Vicia faba) and pea (Pisum sativum). To identify host genes involved in necrotic symptom expression after ClYVV infection, we screened cDNA fragments in which expression was changed in advance of necrotic symptom expression in broad bean (V. faba cv. Wase) using the differential display technique and secondarily with Northern blot analysis. Expression changes were confirmed in 20 genes, and the six that exhibited the most change were analyzed further. These six genes included a gene that encodes a putative nitrate-induced NOI protein (VfNOI), and another was homologous to an Arabidopsis gene that encodes a glycine- and proline-rich protein GPRP (VfGPRP). We recently reported that necrotic symptom development in ClYVV-infected pea is associated with expression of salicylic acid (SA)-dependent pathogenesis-related (PR) proteins and requires SA-dependent host responses. Interestingly, VfNOI and VfGPRP expression was correlated with that of the putative SA-dependent PR proteins in ClYVV-infected broad bean. However, broad bean infected with a recombinant ClYVV expressing the VfGPRP protein showed weaker symptoms and less viral multiplication than that infected with ClYVV expressing the GFP protein. These results imply that VfGPRP plays a role in defense against ClYVV rather than in necrotic symptom expression.

Knockdown of Zebrafish Lumican Gene (zlum) Causes Scleral Thinning and Increased Size of Scleral Coats*

PubMed Central

Yeh, Lung-Kun; Liu, Chia-Yang; Kao, Winston W.-Y.; Huang, Chang-Jen; Hu, Fung-Rong; Chien, Chung-Liang; Wang, I-Jong

2010-01-01

The lumican gene (lum), which encodes one of the major keratan sulfate proteoglycans (KSPGs) in the vertebrate cornea and sclera, has been linked to axial myopia in humans. In this study, we chose zebrafish (Danio rerio) as an animal model to elucidate the role of lumican in the development of axial myopia. The zebrafish lumican gene (zlum) spans ∼4.6 kb of the zebrafish genome. Like human (hLUM) and mouse (mlum), zlum consists of three exons, two introns, and a TATA box-less promoter at the 5′-flanking region of the transcription initiation site. Sequence analysis of the cDNA predicts that zLum encodes 344 amino acids. zLum shares 51% amino acid sequence identity with human lumican. Similar to hLUM and mlum, zlum mRNA is expressed in the eye and many other tissues, such as brain, muscle, and liver as well. Transgenic zebrafish harboring an enhanced GFP reporter gene construct downstream of a 1.7-kb zlum 5′-flanking region displayed enhanced GFP expression in the cornea and sclera, as well as throughout the body. Down-regulation of zlum expression by antisense zlum morpholinos manifested ocular enlargement resembling axial myopia due to disruption of the collagen fibril arrangement in the sclera and resulted in scleral thinning. Administration of muscarinic receptor antagonists, e.g. atropine and pirenzepine, effectively subdued the ocular enlargement caused by morpholinos in in vivo zebrafish larvae assays. The observation suggests that zebrafish can be used as an in vivo model for screening compounds in treating myopia. PMID:20551313

A Mutation in the Rett Syndrome Gene, MECP2, Causes X-Linked Mental Retardation and Progressive Spasticity in Males

PubMed Central

Meloni, Ilaria; Bruttini, Mirella; Longo, Ilaria; Mari, Francesca; Rizzolio, Flavio; D’Adamo, Patrizia; Denvriendt, Koenraad; Fryns, Jean-Pierre; Toniolo, Daniela; Renieri, Alessandra

2000-01-01

Heterozygous mutations in the X-linked MECP2 gene cause Rett syndrome, a severe neurodevelopmental disorder of young females. Only one male presenting an MECP2 mutation has been reported; he survived only to age 1 year, suggesting that mutations in MECP2 are male lethal. Here we report a three-generation family in which two affected males showed severe mental retardation and progressive spasticity, previously mapped in Xq27.2-qter. Two obligate carrier females showed either normal or borderline intelligence, simulating an X-linked recessive trait. The two males and the two obligate carrier females presented a mutation in the MECP2 gene, demonstrating that, in males, MECP2 can be responsible for severe mental retardation associated with neurological disorders. PMID:10986043

Mutations in the LHX2 gene are not a frequent cause of micro/anophthalmia

PubMed Central

Desmaison, Annaïck; Vigouroux, Adeline; Rieubland, Claudine; Peres, Christine; Calvas, Patrick

2010-01-01

Purpose Microphthalmia and anophthalmia are at the severe end of the spectrum of abnormalities in ocular development. A few genes (orthodenticle homeobox 2 [OTX2], retina and anterior neural fold homeobox [RAX], SRY-box 2 [SOX2], CEH10 homeodomain-containing homolog [CHX10], and growth differentiation factor 6 [GDF6]) have been implicated mainly in isolated micro/anophthalmia but causative mutations of these genes explain less than a quarter of these developmental defects. The essential role of the LIM homeobox 2 (LHX2) transcription factor in early eye development has recently been documented. We postulated that mutations in this gene could lead to micro/anophthalmia, and thus performed molecular screening of its sequence in patients having micro/anophthalmia. Methods Seventy patients having non-syndromic forms of colobomatous microphthalmia (n=25), isolated microphthalmia (n=18), or anophthalmia (n=17), and syndromic forms of micro/anophthalmia (n=10) were included in this study after negative molecular screening for OTX2, RAX, SOX2, and CHX10 mutations. Mutation screening of LHX2 was performed by direct sequencing of the coding sequences and intron/exon boundaries. Results Two heterozygous variants of unknown significance (c.128C>G [p.Pro43Arg]; c.776C>A [p.Pro259Gln]) were identified in LHX2 among the 70 patients. These variations were not identified in a panel of 100 control patients of mixed origins. The variation c.776C>A (p.Pro259Gln) was considered as non pathogenic by in silico analysis, while the variation c.128C>G (p.Pro43Arg) considered as deleterious by in silico analysis and was inherited from the asymptomatic father. Conclusions Mutations in LHX2 do not represent a frequent cause of micro/anophthalmia. PMID:21203406

Mutations in the LHX2 gene are not a frequent cause of micro/anophthalmia.

PubMed

Desmaison, Annaïck; Vigouroux, Adeline; Rieubland, Claudine; Peres, Christine; Calvas, Patrick; Chassaing, Nicolas

2010-12-18

Microphthalmia and anophthalmia are at the severe end of the spectrum of abnormalities in ocular development. A few genes (orthodenticle homeobox 2 [OTX2], retina and anterior neural fold homeobox [RAX], SRY-box 2 [SOX2], CEH10 homeodomain-containing homolog [CHX10], and growth differentiation factor 6 [GDF6]) have been implicated mainly in isolated micro/anophthalmia but causative mutations of these genes explain less than a quarter of these developmental defects. The essential role of the LIM homeobox 2 (LHX2) transcription factor in early eye development has recently been documented. We postulated that mutations in this gene could lead to micro/anophthalmia, and thus performed molecular screening of its sequence in patients having micro/anophthalmia. Seventy patients having non-syndromic forms of colobomatous microphthalmia (n=25), isolated microphthalmia (n=18), or anophthalmia (n=17), and syndromic forms of micro/anophthalmia (n=10) were included in this study after negative molecular screening for OTX2, RAX, SOX2, and CHX10 mutations. Mutation screening of LHX2 was performed by direct sequencing of the coding sequences and intron/exon boundaries. Two heterozygous variants of unknown significance (c.128C>G [p.Pro43Arg]; c.776C>A [p.Pro259Gln]) were identified in LHX2 among the 70 patients. These variations were not identified in a panel of 100 control patients of mixed origins. The variation c.776C>A (p.Pro259Gln) was considered as non pathogenic by in silico analysis, while the variation c.128C>G (p.Pro43Arg) considered as deleterious by in silico analysis and was inherited from the asymptomatic father. Mutations in LHX2 do not represent a frequent cause of micro/anophthalmia.

Latest improvements in microbolometer thin film packaging: paving the way for low-cost consumer applications

NASA Astrophysics Data System (ADS)

Yon, J. J.; Dumont, G.; Goudon, V.; Becker, S.; Arnaud, A.; Cortial, S.; Tisse, C. L.

2014-06-01

Silicon-based vacuum packaging is a key enabling technology for achieving affordable uncooled Infrared Focal Plane Arrays (IRFPA) required by a promising mass market that shows momentum for some extensive consumer applications, such as automotive driving assistance, smart presence localization and building management. Among the various approaches studied worldwide, CEA, LETI in partnership with ULIS is committed to the development of a unique technology referred to as PLP (Pixel Level Packaging). In this PLP technology, each bolometer pixel is sealed under vacuum using a transparent thin film deposition on wafer. PLP operates as an array of hermetic micro caps above the focal plane, each enclosing a single microbolometer. In continuation of our on-going studies on PLP for regular QVGA IRFPAs, this paper emphasizes on the innate scalability of the technology which was successfully demonstrated through the development of an 80 × 80 pixel IRFPA. The relevance of the technology with regard to the two formats is discussed, considering both performance and cost issues. We show that the suboptimal fill factor inherent to the PLP arrangement is not so critical when considering smaller arrays preferably fitted for consumer applications. The discussion is supported with the electro-optical performance measurements of the PLP-based 80×80 demonstrator.

Memantine for the Treatment of Phantom Limb Pain: A Systematic Review.

PubMed

Loy, Brittany M; Britt, Rachel B; Brown, Jamie N

2016-12-01

Phantom limb pain (PLP) occurs in up to 85% of patients who have undergone an amputation and remains difficult to treat. Memantine is a N-Methyl-d-aspartate receptor antagonist that has shown benefit in pain syndromes. The objective of this systematic review is to evaluate the evidence for the use of memantine in the treatment of acute and chronic PLP. MEDLINE (1956 to May 2016) and Embase (1957 to May 2016) were queried for articles that characterized the clinical outcomes of patient(s) treated with memantine for PLP. The initial search identified 185 studies and case reports. After screening, eight articles were included. One prospective study, a case report, and two case series demonstrated benefit with memantine in the treatment of acute PLP. However, in chronic PLP that persisted for over 1 year, four prospective studies failed to demonstrate significant analgesic effects with memantine. Memantine was well tolerated in all studies. Memantine appears to be a reasonable option to trial in a patient with a recent amputation or who has failed or cannot tolerate other analgesics. Additional research is needed to further determine the role of memantine in the treatment and prevention of PLP and to identify the population most likely to gain benefit.

Interaction between vitamin B6 metabolism, nitrogen metabolism and autoimmunity.

PubMed

Colinas, Maite; Fitzpatrick, Teresa B

2016-01-01

The essential micronutrient vitamin B6 is best known in its enzymatic cofactor form, pyridoxal 5'-phosphate (PLP). However, vitamin B6 comprises the amine pyridoxamine 5'-phosphate (PMP) and the alcohol pyridoxine 5'-phosphate (PNP) in addition to PLP, as well as their corresponding non-phosphorylated forms. The different B6 forms (called vitamers) are enzymatically interconverted in a ubiquitous salvage pathway. Recently, we have shown that balancing the ratio of the different B6 vitamers in particular PMP by the PMP/PNP oxidase PDX3 is essential for growth and development in Arabidopsis thaliana. Intriguingly, nitrate to ammonium conversion is impaired in pdx3 mutants, such that the mutants become ammonium-dependent, suggesting an interaction between vitamin B6 and nitrogen metabolism. In addition, we found a strong up-regulation of genes related to plant defense. Here, we further show that pdx3 mutants display a temperature-sensitive phenotype that is typical of autoimmune mutants and is possibly connected to the impaired nitrogen metabolism.

Severe manifestation of Bartter syndrome Type IV caused by a novel insertion mutation in the BSND gene.

PubMed

de Pablos, Augusto Luque; García-Nieto, Victor; López-Menchero, Jesús C; Ramos-Trujillo, Elena; González-Acosta, Hilaria; Claverie-Martín, Félix

2014-05-01

Bartter syndrome Type IV is a rare subtype of the Bartter syndromes that leads to both severe renal salt wasting and sensorineural deafness. This autosomal recessive disease is caused by mutations in the gene encoding barttin, BSND, an essential subunit of the ClC-K chloride channels expressed in renal and inner ear epithelia. Patients differ in the severity of renal symptoms, which appears to depend on the modification of channel function by the mutant barttin. To date, only a few BSND mutations have been reported, most of which are missense or nonsense mutations. In this study, we report the identification of the first insertion mutation, p.W102Vfs*7, in the BSND gene of a newborn girl with acute clinical symptoms including early-onset chronic renal failure. The results support previous data indicating that mutations that are predicted to abolish barttin expression are associated with a severe phenotype and early onset renal failure.

Autosomal recessive posterior column ataxia with retinitis pigmentosa caused by novel mutations in the FLVCR1 gene.

PubMed

Shaibani, Aziz; Wong, Lee-Jun; Wei Zhang, Victor; Lewis, Richard Alan; Shinawi, Marwan

2015-01-01

Posterior column ataxia with retinitis pigmentosa (PCARP) is an autosomal recessive disorder characterized by severe sensory ataxia, muscle weakness and atrophy, and progressive pigmentary retinopathy. Recently, mutations in the FLVCR1 gene were described in four families with this condition. We investigated the molecular basis and studied the phenotype of PCARP in a new family. The proband is a 33-year-old woman presented with sensory polyneuropathy and retinitis pigmentosa (RP). The constellation of clinical findings with normal metabolic and genetic evaluation, including mitochondrial DNA (mtDNA) analysis and normal levels of phytanic acid and vitamin E, prompted us to seek other causes of our patient's condition. Sequencing of FLVCR1 in the proband and targeted mutation testing in her two affected siblings revealed two novel variants, c.1547G > A (p.R516Q) and c.1593+5_+8delGTAA predicted, respectively, to be highly conserved throughout evolution and affecting the normal splicing, therefore, deleterious. This study supports the pathogenic role of FLVCR1 in PCARP and expands the molecular and clinical spectra of PCARP. We show for the first time that nontransmembrane domain (TMD) mutations in the FLVCR1 can cause PCARP, suggesting different mechanisms for pathogenicity. Our clinical data reveal that impaired sensation can be part of the phenotypic spectrum of PCARP. This study along with previously reported cases suggests that targeted sequencing of the FLVCR1 gene should be considered in patients with severe sensory ataxia, RP, and peripheral sensory neuropathy.

Cloning of soybean genes induced during hypersensitive cell death caused by syringolide elicitor.

PubMed

Hagihara, Takuya; Hashi, Masaru; Takeuchi, Yoji; Yamaoka, Naoto

2004-02-01

Syringolide elicitors produced by bacteria expressing Pseudomonas syringae pv. glycinea avirulence gene D (avrD) induce hypersensitive cell death (HCD) only in soybean (Glycine max [L.] Merr.) plants carrying the Rpg4 disease resistance gene. Employing a differential display method, we isolated 13 gene fragments induced in cultured cells of a soybean cultivar Harosoy (Rpg4) treated with syringolides. Several genes for isolated fragments were induced by syringolides in an rpg4 cultivar Acme as well as in Harosoy; however, the genes for seven fragments designated as SIH (for syringolide-induced/ HCD associated) were induced exclusively or strongly in Harosoy. cDNA clones for SIH genes were obtained from a cDNA library of Harosoy treated with syringolide. Several sequences are homologous to proteins associated with plant defense responses. The SIH genes did not respond to a non-specific beta-glucan elicitor, which induces phytoalexin accumulation but not HCD, suggesting that the induction of the SIH genes is specific for the syringolide-Harosoy interaction. HCD and the induction of SIH genes by syringolides were independent of H(2)O(2). On the other hand, Ca(2+) was required for HCD and the induction of some SIH genes. These results suggest that the induction of SIH genes by syringolides could be activated through the syringolide-specific signaling pathway and the SIH gene products may play an important role(s) in the processes of HCD induced by syringolides.

A case report of novel mutation in PRF1 gene, which causes familial autosomal recessive hemophagocytic lymphohistiocytosis.

PubMed

Bordbar, Mohammad Reza; Modarresi, Farzaneh; Farazi Fard, Mohammad Ali; Dastsooz, Hassan; Shakib Azad, Nader; Faghihi, Mohammad Ali

2017-05-03

Hemophagocytic Lymphohistiocytosis (HLH) is a life-threatening immunodeficiency and multi-organ disease that affects people of all ages and ethnic groups. Common symptoms and signs of this disease are high fever, hepatosplenomegaly, and cytopenias. Familial form of HLH disease, which is an autosomal recessive hematological disorder is due to disease-causing mutations in several genes essential for NK and T-cell granule-mediated cytotoxic function. For an effective cytotoxic response from cytotoxic T lymphocyte or NK cell encountering an infected cell or tumor cell, different processes are required, including trafficking, docking, priming, membrane fusion, and entry of cytotoxic granules into the target cell leading to apoptosis. Therefore, genes involved in these steps play important roles in the pathogenesis of HLH disease which include PRF1, UNC13D (MUNC13-4), STX11, and STXBP2 (MUNC18-2). Here, we report a novel missense mutation in an 8-year-old boy suffered from hepatosplenomegaly, hepatitis, epilepsy and pancytopenia. The patient was born to a first-cousin parents with no previous documented disease in his parents. To identify mutated gene in the proband, Whole Exome Sequencing (WES) utilizing next generation sequencing was used on an Illumina HiSeq 2000 platform on DNA sample from the patient. Results showed a novel deleterious homozygous missense mutation in PRF1 gene (NM_001083116: exon3: c. 1120 T > G, p.W374G) in the patient and then using Sanger sequencing it was confirmed in the proband and his parents. Since his parents were heterozygous for the identified mutation, autosomal recessive pattern of inheritance was confirmed in the family. Our study identified a rare new pathogenic missense mutation in PRF1 gene in patient with HLH disease and it is the first report of mutation in PRF1 in Iranian patients with this disease.

Acute abdomen due to group A streptococcus bacteremia caused by an isolate with a mutation in the csrS gene.

PubMed

Kaneko, Masahiko; Maruta, Masaki; Shikata, Hisaharu; Hanayama, Masakazu; Ikebe, Tadayoshi

2015-11-01

Streptococcus pyogenes (group A streptococcus) is an aerobic gram-positive coccus that causes infections ranging from non-invasive pharyngitis to severely invasive necrotizing fasciitis. Mutations in csrS/csrR and rgg, negative regulator genes of group A streptococcus, are crucial factors in the pathogenesis of streptococcal toxic shock syndrome, which is a severe, invasive infection characterized by sudden onset of shock and multiorgan failure, resulting in a high mortality rate. Here we present a case of group A streptococcal bacteremia in a 28-year-old Japanese woman with no relevant previous medical history. The patient developed progressive abdominal symptoms that may have been due to spontaneous bacterial peritonitis, followed by a state of shock, which did not fulfill the proposed criteria for streptococcal toxic shock. The isolate was found to harbor a mutation in the negative regulator csrS gene, whereas the csrR and rgg genes were intact. It was noteworthy that this strain carrying a csrS mutation had caused group A streptococcal bacteremia characterized by acute abdomen as the presenting symptom in a young individual who had been previously healthy. This case indicates that group A streptococcus with csrS mutations has potential virulence factors that are associated with the onset of group A streptococcal bacteremia that does not meet the diagnostic criteria for streptococcal toxic shock syndrome. Copyright © 2015 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

Mutation in the epsilon subunit of the cytosolic chaperonin-containing t-complex peptide-1 (Cct5) gene causes autosomal recessive mutilating sensory neuropathy with spastic paraplegia.

PubMed

Bouhouche, A; Benomar, A; Bouslam, N; Chkili, T; Yahyaoui, M

2006-05-01

Mutilating sensory neuropathy with spastic paraplegia is a very rare disease with both autosomal dominant and recessive modes of inheritance. We previously mapped the locus of the autosomal recessive form to a 25 cM interval between markers D5S2048 and D5S648 on chromosome 5p. In this candidate interval, the Cct5 gene encoding the epsilon subunit of the cytosolic chaperonin-containing t-complex peptide-1 (CCT) was the most obvious candidate gene since mutation in the Cct4 gene encoding the CCT delta subunit has been reported to be associated with autosomal recessive mutilating sensory neuropathy in mutilated foot (mf) rat mutant. A consanguineous Moroccan family with four patients displaying mutilating sensory neuropathy associated with spastic paraplegia was investigated. To identify the disease causing gene, the 11 coding exons of the Cct5 gene were screened for mutations by direct sequencing in all family members including the four patients, parents, and six at risk relatives. Sequence analysis of the Cct5 gene revealed a missense A492G mutation in exon 4 that results in the substitution of a highly conserved histidine for arginine amino acid 147. Interestingly, R147 was absent in 384 control matched chromosomes tested. This is the first disease causing mutation that has been identified in the human CCT subunit genes; the mf rat mutant could serve as an animal model for studying these chaperonopathies.

Characterization of Resistance Genes and Plasmids from Outbreaks and Illness Clusters Caused by Salmonella Resistant to Ceftriaxone in the United States, 2011-2012.

PubMed

Folster, Jason P; Grass, Julian E; Bicknese, Amelia; Taylor, Julia; Friedman, Cindy R; Whichard, Jean M

2017-03-01

Salmonella is an important cause of foodborne illness; however, quickly identifying the source of these infections can be difficult, and source identification is a crucial step in preventing additional illnesses. Although most infections are self-limited, invasive salmonellosis may require antimicrobial treatment. Ceftriaxone, an extended-spectrum cephalosporin, is commonly used for treatment of salmonellosis. Previous studies have identified a correlation between the food animal/retail meat source of ceftriaxone-resistant Salmonella and the type of resistance gene and plasmid it carries. In this study, we examined seven outbreaks of ceftriaxone-resistant Salmonella infections, caused by serotypes Typhimurium, Newport, Heidelberg, and Infantis. All isolates were positive for a plasmid-encoded bla CMY gene. Plasmid incompatibility typing identified five IncI1 and two IncA/C plasmids. Both outbreaks containing bla CMY -IncA/C plasmids were linked to consumption of cattle products. Three of five outbreaks with bla CMY -IncI1 (ST12) plasmids were linked to a poultry source. The remaining IncI1 outbreaks were associated with ground beef (ST20) and tomatoes (ST12). In addition, we examined isolates from five unsolved clusters of ceftriaxone-resistant Salmonella infections and used our plasmid-encoded gene findings to predict the source. Overall, we identified a likely association between the source of ceftriaxone-resistant Salmonella outbreaks and the type of resistance gene/plasmid it carries.

The effects of mirror therapy on pain and motor control of phantom limb in amputees: A systematic review.

PubMed

Barbin, J; Seetha, V; Casillas, J M; Paysant, J; Pérennou, D

2016-09-01

Phantom limb pain (PLP) is a major problem after limb amputation. Mirror therapy (MT) is a non-pharmacological treatment using representations of movement, the efficacy of which in reducing PLP remains to be clarified. Here, we present the first systematic review on MT efficacy in PLP and phantom limb movement (PLM) in amputees (lower or upper limb). A search on Medline, Cochrane Database and Embase, crossing the keywords "Phantom Limb" and "Mirror Therapy" found studies which were read and analyzed according the PRISMA statement. Twenty studies were selected, 12 on the subject of MT and PLP, 3 on MT and PLM, 5 on MT and both (PLP and PLM). Among these 20 studies, 5 were randomized controlled trials (163 patients), 6 prospective studies (55 patients), 9 case studies (40 patients) and methodologies were heterogeneous. Seventeen of the 18 studies reported the efficacy of MT on PLP, but with low levels of evidence. One randomized controlled trial did not show any significant effect of MT. As to the effect of MT on PLM, the 8 studies concerned reported effectiveness of MT: 4 with a low level of evidence and 4 with a high level of evidence. An alternative to visual illusion seems to be tactile or auditory stimulation. We cannot recommend MT as a first intention treatment in PLP. The level of evidence is insufficient. Further research is needed to assess the effect of MT on pain, prosthesis use, and body representation, and to standardize protocols. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

A Homozygous TPO Gene Duplication (c.1184_1187dup4) Causes Congenital Hypothyroidism in Three Siblings Born to a Consanguineous Family

PubMed Central

Cangul, Hakan; Aydin, Banu K.; Bas, Firdevs

2015-01-01

Congenital hypothyroidism (CH) is the most common neonatal endocrine disease, and germ-line mutations in the TPO gene cause the inherited form of the disease. Our aim in this study was to determine the genetic basis of congenital hypothyroidism in three affected children coming from a consanguineous Turkish family. Because CH is usually inherited in autosomal recessive manner in consanguineous/multicase families, we adopted a two-stage strategy of genetic linkage studies and targeted sequencing of the candidate genes. First, we investigated the potential genetic linkage of the family to any known CH locus, using microsatellite markers, and then screened for mutations in linked-gene by conventional sequencing. The family showed potential linkage to the TPO gene and we detected a homozygous duplication (c.1184_1187dup4) in all cases. The mutation segregated with disease status in the family. This study confirms the pathogenicity of the c.1184_1187dup4 mutation in the TPO gene and helps establish a genotype/phenotype correlation associated with this mutation. It also highlights the importance of molecular genetic studies in the definitive diagnosis and accurate classification of CH. PMID:27617131

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

CRISPR Knockout of the HuR Gene Causes a Xenograft Lethal Phenotype

PubMed Central

Lal, Shruti; Cheung, Edwin C.; Zarei, Mahsa; Preet, Ranjan; Chand, Saswati N.; Mambelli-Lisboa, Nicole C.; Romeo, Carmella; Stout, Matthew C.; Londin, Eric; Goetz, Austin; Lowder, Cinthya Y.; Nevler, Avinoam; Yeo, Charles J.; Campbell, Paul M.; Winter, Jordan M.; Dixon, Dan A.; Brody, Jonathan R.

2017-01-01

Pancreatic ductal adenocarcinoma (PDA) is the third leading cause of cancer related deaths in the U.S., while colorectal cancer (CRC) is the third most common cancer. The RNA binding protein HuR (ELAVL1), supports a pro-oncogenic network in gastrointestinal (GI) cancer cells through enhanced HuR expression. Using a publically available database, HuR expression levels were determined to be increased in primary PDA and CRC tumor cohorts as compared to normal pancreas and colon tissues, respectively. CRISPR/Cas9 technology was successfully used to delete the HuR gene in both PDA (MIA PaCa-2 and Hs 766T) and CRC (HCT116) cell lines. HuR deficiency has a mild phenotype, in vitro, as HuR-deficient MIA PaCa-2 (MIA.HuR-KO(−/−)) cells had increased apoptosis when compared to isogenic wild-type (MIA.HuR-WT(+/+)) cells. Using this isogenic system, mRNAs were identified that specifically bound to HuR and were required for transforming a 2D culture into 3D (i.e., organoids). Importantly, HuR-deficient MIA PaCa-2 and Hs 766T cells were unable to engraft tumors in vivo compared to control HuR-proficient cells, demonstrating a unique xenograft lethal phenotype. While not as a dramatic phenotype, CRISPR knockout HuR HCT116 colon cancer cells (HCT.HuR-KO(−/−)) showed significantly reduced in vivo tumor growth compared to controls (HCT.HuR-WT(+/+)). Finally, HuR deletion affects KRAS activity and controls a subset of pro-oncogenic genes. Implications The work reported here supports the notion that targeting HuR is a promising therapeutic strategy to treat GI malignancies. PMID:28242812

CRISPR Knockout of the HuR Gene Causes a Xenograft Lethal Phenotype.

PubMed

Lal, Shruti; Cheung, Edwin C; Zarei, Mahsa; Preet, Ranjan; Chand, Saswati N; Mambelli-Lisboa, Nicole C; Romeo, Carmella; Stout, Matthew C; Londin, Eric; Goetz, Austin; Lowder, Cinthya Y; Nevler, Avinoam; Yeo, Charles J; Campbell, Paul M; Winter, Jordan M; Dixon, Dan A; Brody, Jonathan R

2017-06-01

Pancreatic ductal adenocarcinoma (PDA) is the third leading cause of cancer-related deaths in the United States, whereas colorectal cancer is the third most common cancer. The RNA-binding protein HuR (ELAVL1) supports a pro-oncogenic network in gastrointestinal (GI) cancer cells through enhanced HuR expression. Using a publically available database, HuR expression levels were determined to be increased in primary PDA and colorectal cancer tumor cohorts as compared with normal pancreas and colon tissues, respectively. CRISPR/Cas9 technology was successfully used to delete the HuR gene in both PDA (MIA PaCa-2 and Hs 766T) and colorectal cancer (HCT116) cell lines. HuR deficiency has a mild phenotype, in vitro , as HuR-deficient MIA PaCa-2 (MIA.HuR-KO (-/-) ) cells had increased apoptosis when compared with isogenic wild-type (MIA.HuR-WT (+/+) ) cells. Using this isogenic system, mRNAs were identified that specifically bound to HuR and were required for transforming a two-dimensional culture into three dimensional (i.e., organoids). Importantly, HuR-deficient MIA PaCa-2 and Hs 766T cells were unable to engraft tumors in vivo compared with control HuR-proficient cells, demonstrating a unique xenograft lethal phenotype. Although not as a dramatic phenotype, CRISPR knockout HuR HCT116 colon cancer cells (HCT.HuR-KO (-/-) ) showed significantly reduced in vivo tumor growth compared with controls (HCT.HuR-WT (+/+) ). Finally, HuR deletion affects KRAS activity and controls a subset of pro-oncogenic genes. Implications: The work reported here supports the notion that targeting HuR is a promising therapeutic strategy to treat GI malignancies. Mol Cancer Res; 15(6); 696-707. ©2017 AACR . ©2017 American Association for Cancer Research.

Autism and Genes

ERIC Educational Resources Information Center

National Institutes of Health, 2005

2005-01-01

This document defines and discusses autism and how genes play a role in the condition. Answers to the following questions are covered: (1) What are genes? (2) What is autism? (3) What causes autism? (4) Why study genes to learn about autism? (5) How do researchers look for the genes involved in autism? (screen the whole genome; conduct cytogenetic…

Polymorphisms in metabolism and repair genes affects DNA damage caused by open-cast coal mining exposure.

PubMed

Espitia-Pérez, Lyda; Sosa, Milton Quintana; Salcedo-Arteaga, Shirley; León-Mejía, Grethel; Hoyos-Giraldo, Luz Stella; Brango, Hugo; Kvitko, Katia; da Silva, Juliana; Henriques, João A P

2016-09-15

Increasing evidence suggest that occupational exposure to open-cast coal mining residues like dust particles, heavy metals and Polycyclic Aromatic Hydrocarbons (PAHs) may cause a wide range of DNA damage and genomic instability that could be associated to initial steps in cancer development and other work-related diseases. The aim of our study was to evaluate if key polymorphisms in metabolism genes CYP1A1Msp1, GSTM1Null, GSTT1Null and DNA repair genes XRCC1Arg194Trp and hOGG1Ser326Cys could modify individual susceptibility to adverse coal exposure effects, considering the DNA damage (Comet assay) and micronucleus formation in lymphocytes (CBMN) and buccal mucosa cells (BMNCyt) as endpoints for genotoxicity. The study population is comprised of 200 healthy male subjects, 100 open-cast coal-mining workers from "El Cerrejón" (world's largest open-cast coal mine located in Guajira - Colombia) and 100 non-exposed referents from general population. The data revealed a significant increase of CBMN frequency in peripheral lymphocytes of occupationally exposed workers carrying the wild-type variant of GSTT1 (+) gene. Exposed subjects carrying GSTT1null polymorphism showed a lower micronucleus frequency compared with their positive counterparts (FR: 0.83; P=0.04), while BMNCyt, frequency and Comet assay parameters in lymphocytes: Damage Index (DI) and percentage of DNA in the tail (Tail % DNA) were significantly higher in exposed workers with the GSTM1Null polymorphism. Other exfoliated buccal mucosa abnormalities related to cell death (Karyorrhexis and Karyolysis) were increased in GSTT/M1Null carriers. Nuclear buds were significantly higher in workers carrying the CYP1A1Msp1 (m1/m2, m2/m2) allele. Moreover, BMNCyt frequency and Comet assay parameters were significantly lower in exposed carriers of XRCC1Arg194Trp (Arg/Trp, Trp/Trp) and hOGG1Ser326Cys (Ser/Cys, Cys/Cys), thereby providing new data to the increasing evidence about the protective role of these polymorphisms

ROAM mutations causing increased expression of yeast genes: their activation by signals directed toward conjugation functions and their formation by insertion of tyl repetitive elements

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

Errede, B.; Cardillo, T.S.; Wever, G.

1980-01-01

Mechanisms available to eukaryotic organisms for the coordinate regulation of gene expression are being examined by genetic and biochemical characterization of an unusual mutation, CYC7-H2, which causes overproduction of iso-2-cytochrome c in the yeast Saccharomyces cerevisiae. The CYC7-H2 mutation causes approximately a twenty fold overproduction of iso-2-cytochrome c in haploid strains but only a one to four fold overproduction in MATa/MAT..cap alpha.. diploid strains. This regulation of overproduction has been characterized as a response to signals controlling conjugation in yeast. The CYC7-H2 mutation is closely related to other regulatory mutations occurring at the cargA, cargB and DUR1,2 loci which aremore » the structural genes for arginase, ornithine transaminase and urea amidolyase, respectively. Similar to the CYC7-H2 mutation, the mutations designated cargA/sup +/O/sup h/, cargB/sup +/O/sup h/ and durO/sup h/ cause constitutive production of their respective gene products at much lower levels in MATa/MAT..cap alpha.. diploid strains than in the corresponding haploid strains. Observations characterizing the regulation of overproduction in the CYC7-H2 mutant are presented with the additional and parallel observations for the O/sup h/ mutants.« less

Transient protein-protein interactions perturb E. coli metabolome and cause gene dosage toxicity

PubMed Central

Bhattacharyya, Sanchari; Bershtein, Shimon; Yan, Jin; Argun, Tijda; Gilson, Amy I; Trauger, Sunia A; Shakhnovich, Eugene I

2016-01-01

Gene dosage toxicity (GDT) is an important factor that determines optimal levels of protein abundances, yet its molecular underpinnings remain unknown. Here, we demonstrate that overexpression of DHFR in E. coli causes a toxic metabolic imbalance triggered by interactions with several functionally related enzymes. Though deleterious in the overexpression regime, surprisingly, these interactions are beneficial at physiological concentrations, implying their functional significance in vivo. Moreover, we found that overexpression of orthologous DHFR proteins had minimal effect on all levels of cellular organization – molecular, systems, and phenotypic, in sharp contrast to E. coli DHFR. Dramatic difference of GDT between ‘E. coli’s self’ and ‘foreign’ proteins suggests the crucial role of evolutionary selection in shaping protein-protein interaction (PPI) networks at the whole proteome level. This study shows how protein overexpression perturbs a dynamic metabolon of weak yet potentially functional PPI, with consequences for the metabolic state of cells and their fitness. DOI: http://dx.doi.org/10.7554/eLife.20309.001 PMID:27938662

Structural dynamics of a methionine γ-lyase for calicheamicin biosynthesis: Rotation of the conserved tyrosine stacking with pyridoxal phosphate

DOE PAGES

Cao, Hongnan; Tan, Kemin; Wang, Fengbin; ...

2016-04-29

CalE6 from Micromonospora echinospora is a (pyridoxal 50 phosphate) PLP-dependent methionine γ-lyase involved in the biosynthesis of calicheamicins. Here, we report the crystal structure of a CalE6 2-(N-morpholino)ethanesulfonic acid complex showing ligand-induced rotation of Tyr100, which stacks with PLP, resembling the corresponding tyrosine rotation of true catalytic intermediates of CalE6 homologs. Elastic network modeling and crystallographic ensemble refinement reveal mobility of the N-terminal loop, which involves both tetrameric assembly and PLP binding. Modeling and comparative structural analysis of PLP-dependent enzymes involved in Cys/Met metabolism shine light on the functional implications of the intrinsic dynamic properties of CalE6 in catalysis andmore » holoenzyme maturation.« less

Further insight into the phenotype associated with a mutation in the ORC6 gene, causing Meier-Gorlin syndrome 3.

PubMed

Shalev, Stavit Allon; Khayat, Morad; Etty, Daniel-Spiegl; Elpeleg, Orly

2015-03-01

Mutations in genes encoding the origin recognition complex subunits cause Meier-Gorlin syndrome. The disease manifests a triad of short stature, small ears, and small and/or absent patellae with variable expressivity. We report on the identification of a homozygous deleterious mutation in the ORC6 gene in previously described fetuses at the severe end of the Meier-Gorlin spectrum. The phenotype included severe intrauterine growth retardation, dislocation of knees, gracile bones, clubfeet, and small mandible and chest. To date, the clinical presentation of ORC6-associated Meier-Gorlin syndrome has been mild compared to other the phenotype associated with other loci. The present report expands the clinical phenotype associated with ORC6 mutations to include severely abnormal embryological development suggesting a possible genotype-phenotype correlation. © 2015 Wiley Periodicals, Inc.

Exonization of an Intronic LINE-1 Element Causing Becker Muscular Dystrophy as a Novel Mutational Mechanism in Dystrophin Gene

PubMed Central

Gonçalves, Ana; Coelho, Teresa; Melo-Pires, Manuel; Sousa, Mário

2017-01-01

A broad mutational spectrum in the dystrophin (DMD) gene, from large deletions/duplications to point mutations, causes Duchenne/Becker muscular dystrophy (D/BMD). Comprehensive genotyping is particularly relevant considering the mutation-centered therapies for dystrophinopathies. We report the genetic characterization of a patient with disease onset at age 13 years, elevated creatine kinase levels and reduced dystrophin labeling, where multiplex-ligation probe amplification (MLPA) and genomic sequencing failed to detect pathogenic variants. Bioinformatic, transcriptomic (real time PCR, RT-PCR), and genomic approaches (Southern blot, long-range PCR, and single molecule real-time sequencing) were used to characterize the mutation. An aberrant transcript was identified, containing a 103-nucleotide insertion between exons 51 and 52, with no similarity with the DMD gene. This corresponded to the partial exonization of a long interspersed nuclear element (LINE-1), disrupting the open reading frame. Further characterization identified a complete LINE-1 (~6 kb with typical hallmarks) deeply inserted in intron 51. Haplotyping and segregation analysis demonstrated that the mutation had a de novo origin. Besides underscoring the importance of mRNA studies in genetically unsolved cases, this is the first report of a disease-causing fully intronic LINE-1 element in DMD, adding to the diversity of mutational events that give rise to D/BMD. PMID:28972564

Exonization of an Intronic LINE-1 Element Causing Becker Muscular Dystrophy as a Novel Mutational Mechanism in Dystrophin Gene.

PubMed

Gonçalves, Ana; Oliveira, Jorge; Coelho, Teresa; Taipa, Ricardo; Melo-Pires, Manuel; Sousa, Mário; Santos, Rosário

2017-10-03

A broad mutational spectrum in the dystrophin ( DMD ) gene, from large deletions/duplications to point mutations, causes Duchenne/Becker muscular dystrophy (D/BMD). Comprehensive genotyping is particularly relevant considering the mutation-centered therapies for dystrophinopathies. We report the genetic characterization of a patient with disease onset at age 13 years, elevated creatine kinase levels and reduced dystrophin labeling, where multiplex-ligation probe amplification (MLPA) and genomic sequencing failed to detect pathogenic variants. Bioinformatic, transcriptomic (real time PCR, RT-PCR), and genomic approaches (Southern blot, long-range PCR, and single molecule real-time sequencing) were used to characterize the mutation. An aberrant transcript was identified, containing a 103-nucleotide insertion between exons 51 and 52, with no similarity with the DMD gene. This corresponded to the partial exonization of a long interspersed nuclear element (LINE-1), disrupting the open reading frame. Further characterization identified a complete LINE-1 (~6 kb with typical hallmarks) deeply inserted in intron 51. Haplotyping and segregation analysis demonstrated that the mutation had a de novo origin. Besides underscoring the importance of mRNA studies in genetically unsolved cases, this is the first report of a disease-causing fully intronic LINE-1 element in DMD , adding to the diversity of mutational events that give rise to D/BMD.

Autosomal Recessive Hypotrichosis with Woolly Hair Caused by a Mutation in the Keratin 25 Gene Expressed in Hair Follicles.

PubMed

Zernov, Nikolay V; Skoblov, Mikhail Y; Marakhonov, Andrey V; Shimomura, Yutaka; Vasilyeva, Tatyana A; Konovalov, Fedor A; Abrukova, Anna V; Zinchenko, Rena A

2016-06-01

Hypotrichosis is an abnormal condition characterized by decreased hair density and various defects in hair structure and growth patterns. In particular, in woolly hair, hypotrichosis is characterized by a tightly curled structure and abnormal growth. In this study, we present a detailed comparative examination of individuals affected by autosomal-recessive hypotrichosis (ARH), which distinguishes two types of ARH. Earlier, we demonstrated that exon 4 deletion in the lipase H gene caused an ARH (hypotrichosis 7; MIM: 604379) in populations of the Volga-Ural region of Russia. Screening for this mutation in all affected individuals revealed its presence only in the group with the hypotrichosis 7 phenotype. Other patients formed a separate group of woolly hair-associated ARH, with a homozygous missense mutation c.712G>T (p.Val238Leu) in a highly conserved position of type I keratin KRT25 (K25). Haplotype analysis indicated a founder effect. An expression study in the HaCaT cell line demonstrated a deleterious effect of the p.Val238Leu mutation on the formation of keratin intermediate filaments. Hence, we have identified a previously unreported missense mutation in the KRT25 gene causing ARH with woolly hair. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

A mutation in the insulin receptor gene that impairs transport of the receptor to the plasma membrane and causes insulin-resistant diabetes.

PubMed Central

Accili, D; Frapier, C; Mosthaf, L; McKeon, C; Elbein, S C; Permutt, M A; Ramos, E; Lander, E; Ullrich, A; Taylor, S I

1989-01-01

Insulin binds to a receptor on the cell surface, thereby triggering a biological response within the target cell. Mutations in the insulin receptor gene can render the cell resistant to the biological action of insulin. We have studied a family in which two sisters have a genetic form of insulin-resistant diabetes mellitus. The technique of homozygosity mapping has been used to demonstrate that the mutation causing diabetes in this consanguineous family is genetically linked to the insulin receptor gene. The two insulin-resistant sisters are homozygous for a mutation encoding substitution of valine for phenylalanine at position 382 in the alpha-subunit of the insulin receptor. Transfection of mutant insulin receptor cDNA into NIH3T3 cells demonstrated that the Val382 mutation impaired post-translational processing and retarded transport of the insulin receptor to the plasma membrane. Thus, the mutation causes insulin resistance by decreasing the number of insulin receptors on the surface of the patients' cells. Images PMID:2573522

A NOVEL MUTATION IN THE HCN4 GENE CAUSES SYMPTOMATIC SINUS BRADYCARDIA IN MOROCCAN JEWS

PubMed Central

Laish-Farkash, Avishag; Brass, Dovrat; Marek-Yagel, Dina; Pras, Elon; Dascal, Nathan; Antzelevitch, Charles; Nof, Eyal; Reznik, Haya; Eldar, Michael; Glikson, Michael; Luria, David

2010-01-01

Objectives To conduct a clinical, genetic and functional analysis of three unrelated families with familial sinus bradycardia (FSB). Background Mutations in the hyperpolarization-activated nucleotide-gated channel (HCN4) are known to be associated with FSB. Methods and Results Three males of Moroccan Jewish descent were hospitalized: one survived an out-of-hospital cardiac arrest and 2 presented with weakness and presyncopal events. All 3 had significant sinus bradycardia, also found in other first-degree relatives, with a segregation suggesting autosomal-dominant inheritance. All had normal response to exercise and normal heart structure. Sequencing of the HCN4 gene in all patients revealed a C to T transition at nucleotide position 1454, which resulted in an alanine to valine change (A485V) in the ion channel pore found in most of their bradycardiac relatives, but not in 150 controls. Functional expression of the mutated ion channel in Xenopus oocytes and in human embryonic kidney 293 cells revealed profoundly reduced function and synthesis of the mutant channel compared to wild-type. Conclusions We describe a new mutation in the HCN4 gene causing symptomatic FSB in 3 unrelated individuals of similar ethnic background that may indicate unexplained FSB in this ethnic group. This profound functional defect is consistent with the symptomatic phenotype. PMID:20662977

A novel cysteine desulfurase influencing organosulfur compounds in Lentinula edodes

PubMed Central

Liu, Ying; Lei, Xiao-Yu; Chen, Lian-Fu; Bian, Yin-Bing; Yang, Hong; Ibrahim, Salam A.; Huang, Wen

2015-01-01

Organosulfur compounds are the basis for the unique aroma of Lentinula edodes, and cysteine sulfoxide lyase (C-S lyase) is the key enzyme in this trait. The enzyme from Alliium sativum has been crystallized and well-characterized; however, there have been no reports of the characterization of fungi C-S lyase at the molecular level. We identified a L. edodes C-S lyase (Lecsl), cloned a gene of Csl encoded Lecsl and then combined modeling, simulations, and experiments to understand the molecular basis of the function of Lecsl. Our analysis revealed Lecsl to be a novel cysteine desulfurase and not a type of cysteine sulfoxide lyase. The pyridoxal-5-phosphate (PLP) molecule bonded tightly to Lecsl to form a Lecsl-PLP complex. Moreover, the Lecsl had one active center that served to bind two kinds of substrates, S-methyl-L-cysteine sulfoxide and L-cysteine, and had both cysteine sulfoxide lyase and cysteine desulfurase activity. We found that the amino acid residue Asn393 was essential for the catalytic activity of Lecsl and that the gene Csl encoded a novel cysteine desulfurase to influence organosulfur compounds in L. edodes. Our results provide a new insight into understanding the formation of the unique aroma of L. edodes. PMID:26054293

A novel cysteine desulfurase influencing organosulfur compounds in Lentinula edodes.

PubMed

Liu, Ying; Lei, Xiao-Yu; Chen, Lian-Fu; Bian, Yin-Bing; Yang, Hong; Ibrahim, Salam A; Huang, Wen

2015-06-09

Organosulfur compounds are the basis for the unique aroma of Lentinula edodes, and cysteine sulfoxide lyase (C-S lyase) is the key enzyme in this trait. The enzyme from Alliium sativum has been crystallized and well-characterized; however, there have been no reports of the characterization of fungi C-S lyase at the molecular level. We identified a L. edodes C-S lyase (Lecsl), cloned a gene of Csl encoded Lecsl and then combined modeling, simulations, and experiments to understand the molecular basis of the function of Lecsl. Our analysis revealed Lecsl to be a novel cysteine desulfurase and not a type of cysteine sulfoxide lyase. The pyridoxal-5-phosphate (PLP) molecule bonded tightly to Lecsl to form a Lecsl-PLP complex. Moreover, the Lecsl had one active center that served to bind two kinds of substrates, S-methyl-L-cysteine sulfoxide and L-cysteine, and had both cysteine sulfoxide lyase and cysteine desulfurase activity. We found that the amino acid residue Asn393 was essential for the catalytic activity of Lecsl and that the gene Csl encoded a novel cysteine desulfurase to influence organosulfur compounds in L. edodes. Our results provide a new insight into understanding the formation of the unique aroma of L. edodes.

A mechanism for exon skipping caused by nonsense or missense mutations in BRCA1 and other genes.

PubMed

Liu, H X; Cartegni, L; Zhang, M Q; Krainer, A R

2001-01-01

Point mutations can generate defective and sometimes harmful proteins. The nonsense-mediated mRNA decay (NMD) pathway minimizes the potential damage caused by nonsense mutations. In-frame nonsense codons located at a minimum distance upstream of the last exon-exon junction are recognized as premature termination codons (PTCs), targeting the mRNA for degradation. Some nonsense mutations cause skipping of one or more exons, presumably during pre-mRNA splicing in the nucleus; this phenomenon is termed nonsense-mediated altered splicing (NAS), and its underlying mechanism is unclear. By analyzing NAS in BRCA1, we show here that inappropriate exon skipping can be reproduced in vitro, and results from disruption of a splicing enhancer in the coding sequence. Enhancers can be disrupted by single nonsense, missense and translationally silent point mutations, without recognition of an open reading frame as such. These results argue against a nuclear reading-frame scanning mechanism for NAS. Coding-region single-nucleotide polymorphisms (cSNPs) within exonic splicing enhancers or silencers may affect the patterns or efficiency of mRNA splicing, which may in turn cause phenotypic variability and variable penetrance of mutations elsewhere in a gene.

Inflammation, vitamin B6 and related pathways.

PubMed

Ueland, Per Magne; McCann, Adrian; Midttun, Øivind; Ulvik, Arve

2017-02-01

The active form of vitamin B6, pyridoxal 5'-phosphate (PLP), serves as a co-factor in more than 150 enzymatic reactions. Plasma PLP has consistently been shown to be low in inflammatory conditions; there is a parallel reduction in liver PLP, but minor changes in erythrocyte and muscle PLP and in functional vitamin B6 biomarkers. Plasma PLP also predicts the risk of chronic diseases like cardiovascular disease and some cancers, and is inversely associated with numerous inflammatory markers in clinical and population-based studies. Vitamin B6 intake and supplementation improve some immune functions in vitamin B6-deficient humans and experimental animals. A possible mechanism involved is mobilization of vitamin B6 to the sites of inflammation where it may serve as a co-factor in pathways producing metabolites with immunomodulating effects. Relevant vitamin B6-dependent inflammatory pathways include vitamin B6 catabolism, the kynurenine pathway, sphingosine 1-phosphate metabolism, the transsulfuration pathway, and serine and glycine metabolism. Copyright © 2016 Elsevier Ltd. All rights reserved.

Upregulation of proinflammatory genes in skin lesions may be the cause of keloid formation (Review)

PubMed Central

DONG, XIANGLIN; MAO, SHAOLIN; WEN, HAO

2013-01-01

It was previously demonstrated that the main cause behind keloid formation may be keloid fibroblast abnormalities, which are closely associated with the microenvironment of the keloid lesion. The post-traumatic and chronic inflammation of the keloid lesion area suggest that inflammatory mediators play an important role in the keloid microenvironment and are crucial for keloid fibroblast abnormalities. In this study, we hypothesized that the mechanism underlying keloid formation may involve the continuous upregulation of proinflammatory gene expression in keloid lesions. This hypothesis may explain the inflammatory response, invasive growth and recurrence following resection of keloids, as well as the selective localization of keloids in specific parts of a patient’s body and the differences in localization among different patients. PMID:24649037

A case report: a heterozygous deletion (2791_2805 del) in exon 18 of the filamin C gene causing filamin C-related myofibrillar myopathies in a Chinese family.

PubMed

Miao, Jing; Su, Fei-Fei; Liu, Xue-Mei; Wei, Xiao-Jing; Yuan, Yun; Yu, Xue-Fan

2018-06-04

Filamin C-related myofibrillar myopathies (MFM) are progressive skeletal myopathies with an autosomal dominant inheritance pattern. The conditions are caused by mutations of the filamin C gene (FLNC) located in the chromosome 7q32-q35 region. Genetic variations in the FLNC gene result in various clinical phenotypes. We describe a 43-year-old woman who suffered filamin C-related MFM, with symptoms first presenting in the proximal muscles of the lower limbs and eventually spreading to the upper limbs and distal muscles. The patient's serum level of creatine kinase was mildly increased. Mildy myopathic changes in the electromyographic exam and moderate lipomatous alterations in lower limb MRI were found. Histopathological examination revealed increased muscle fiber size variability, disturbances in oxidative enzyme activity, and the presence of abnormal protein aggregates and vacuoles in some muscle fibers. Ultrastructural analysis showed inclusions composed of thin filaments and interspersed granular densities. DNA sequencing analysis detected a novel 15-nucleotide deletion (c.2791_2805del, p.931_935del) in the FLNC gene. The patient's father, sister, brother, three paternal aunts, one paternal uncle, and the uncle's son also had slowly progressive muscle weakness, and thus, we detected an autosomal dominant inheritance pattern of the disorder. A novel heterogeneous 15-nucleotide deletion (c.2791_2805del, p.931_935del) in the Ig-like domain 7 of the FLNC gene was found to cause filamin C-related MFM. This deletion in the FLNC gene causes protein aggregation, abnormalities in muscle structure, and impairment in muscle fiber function, which leads to muscle weakness.

Osteopoikilosis and multiple exostoses caused by novel mutations in LEMD3 and EXT1 genes respectively - coincidence within one family

PubMed Central

2010-01-01

Background Osteopoikilosis is a rare autosomal dominant genetic disorder, characterised by the occurrence of the hyperostotic spots preferentially localized in the epiphyses and metaphyses of the long bones, and in the carpal and tarsal bones [1]. Heterozygous LEMD3 gene mutations were shown to be the primary cause of the disease [2]. Association of the primarily asymptomatic osteopokilosis with connective tissue nevi of the skin is categorized as Buschke-Ollendorff syndrome (BOS) [3]. Additionally, osteopoikilosis can coincide with melorheostosis (MRO), a more severe bone disease characterised by the ectopic bone formation on the periosteal and endosteal surface of the long bones [4-6]. However, not all MRO affected individuals carry germ-line LEMD3 mutations [7]. Thus, the genetic cause of MRO remains unknown. Here we describe a familial case of osteopoikilosis in which a novel heterozygous LEMD3 mutation coincides with a novel mutation in EXT1, a gene involved in aetiology of multiple exostosis syndrome. The patients affected with both LEMD3 and EXT1 gene mutations displayed typical features of the osteopoikilosis. There were no additional skeletal manifestations detected however, various non-skeletal pathologies coincided in this group. Methods We investigated LEMD3 and EXT1 in the three-generation family from Poland, with 5 patients affected with osteopoikilosis and one child affected with multiple exostoses. Results We found a novel c.2203C > T (p.R735X) mutation in exon 9 of LEMD3, resulting in a premature stop codon at amino acid position 735. The mutation co-segregates with the osteopoikilosis phenotype and was not found in 200 ethnically matched controls. Another new substitution G > A was found in EXT1 gene at position 1732 (cDNA) in Exon 9 (p.A578T) in three out of five osteopoikilosis affected family members. Evolutionary conservation of the affected amino acid suggested possible functional relevance, however no additional skeletal manifestations were

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.

The D153del Mutation in GNB3 Gene Causes Tissue Specific Signalling Patterns and an Abnormal Renal Morphology in Rge Chickens

PubMed Central

Tummala, Hemanth; Fleming, Stewart; Hocking, Paul M.; Wehner, Daniel; Naseem, Zahid; Ali, Manir; Inglehearn, Christopher F.; Zhelev, Nikolai; Lester, Douglas H.

2011-01-01

Background The GNB3 gene is expressed in cone but not rod photoreceptors of vertebrates, where it acts as the β transducin subunit in the colour visual transduction process. A naturally occurring mutation ‘D153del’ in the GNB3 gene causes the recessively inherited blinding phenotype retinopathy globe enlarged (rge) disease in chickens. GNB3 is however also expressed in most other vertebrate tissues suggesting that the D153del mutation may exert pathological effects that outlie from eye. Principal Findings Recombinant studies in COS-7 cells that were transfected with normal and mutant recombinant GNB3 constructs and subjected to cycloheximide chase showed that the mutant GNB3d protein had a much shorter half life compared to normal GNB3. GNB3 codes for the Gβ3 protein subunit that, together with different Gγ and Gα subunits, activates and regulates phosphorylation cascades in different tissues. As expected, the relative levels of cGMP and cAMP secondary messengers and their activated kinases such as MAPK, AKT and GRK2 were also found to be altered significantly in a tissue specific manner in rge chickens. Histochemical analysis on kidney tissue sections, from rge homozygous affected chickens, showed the chickens had enlargement of the glomerular capsule, causing glomerulomegaly and tubulointerstitial inflammation whereas other tissues (brain, heart, liver, pancreas) were unaffected. Significance These findings confirm that the D153del mutation in GNB3 gene targets GNB3 protein to early degradation. Lack of GNB3 signalling causes reduced phosphorylation activity of ERK2 and AKT leading to severe pathological phenotypes such as blindness and renal abnormalities in rge chickens. PMID:21887213

Coupling of Peripheral Tolerance to Endogenous Interleukin 10 Promotes Effective Modulation of Myelin-Activated T Cells and Ameliorates Experimental Allergic Encephalomyelitis

PubMed Central

Legge, Kevin L.; Min, Booki; Bell, J. Jeremiah; Caprio, Jacque C.; Li, Lequn; Gregg, Randal K.; Zaghouani, Habib

2000-01-01

Several immune-based approaches are being considered for modulation of inflammatory T cells and amelioration of autoimmune diseases. The most recent strategies include simulation of peripheral self-tolerance by injection of adjuvant free antigen, local delivery of cytokines by genetically altered T cells, and interference with the function of costimulatory molecules. Although promising results have been obtained from these studies that define mechanisms of T cell modulation, efficacy, practicality, and toxicity, concerns remain unsolved, thereby justifying further investigations to define alternatives for effective downregulation of aggressive T cells. In prior studies, we demonstrated that an immunoglobulin (Ig) chimera carrying the encephalitogenic proteolipid protein (PLP)1 peptide corresponding to amino acid sequence 139–151 of PLP, Ig-PLP1, is presented to T cells ∼100-fold better than free PLP1. Here, we demonstrate that aggregation endows Ig-PLP1 with an additional feature, namely, induction of interleukin (IL)-10 production by macrophages and dendritic cells, both of which are antigen-presenting cells (APCs). These functions synergize in vivo and drive effective modulation of autoimmunity. Indeed, it is shown that animals with ongoing active experimental allergic encephalomyelitis dramatically reduce the severity of their paralysis when treated with adjuvant free aggregated Ig-PLP1. Moreover, IL-10 displays bystander antagonism on unrelated autoreactive T cells, allowing for reversal of disease involving multiple epitopes. Therefore, aggregated Ig-PLP1 likely brings together a peripheral T cell tolerance mechanism emanating from peptide presentation by APCs expressing suboptimal costimulatory molecules and IL-10 bystander suppression to drive a dual-modal T cell modulation system effective for reversal of autoimmunity involving several epitopes and diverse T cell specificities. PMID:10859329

A Mild Version of Danon Disease Caused by a Newly Recognized Mutation in the Lysosome-associated Membrane Protein-2 Gene.

PubMed

Kyaw, Htoo; Shaik, Fatima; Lin, Aung Naing; Shinnar, Meir

2018-02-04

We present the case of a patient with dilated cardiomyopathy caused by a novel mutation in the lysosome-associated membrane protein-2 (LAMP-2) gene. Patients with pathogenic mutations of this gene typically suffer from Danon disease - a condition that leads to cognitive decline, severe skeletal myopathy, and severe hypertrophic cardiomyopathy. Our patient's presentation and clinical course, however, is different and much less severe than other patients with this disease. He did not suffer from neurologic and musculoskeletal complications. He is also possibly the longest-known survivor of this disease without a heart transplant. This disease is unfamiliar to many physicians, and our case highlights the importance of an awareness of this disorder, particularly because of its implications for both the patient and his family.

A hybrid QM/MM simulation study of intramolecular proton transfer in the pyridoxal 5'-phosphate in the active site of transaminase: influence of active site interaction on proton transfer.

PubMed

Dutta Banik, Sindrila; Chandra, Amalendu

2014-09-25

Pyridoxal 5'-phosphate (PLP) Schiff base, a versatile cofactor, exhibits a tautomeric equilibrium that involves an intramolecular proton transfer between the N-protonated zwitterionic ketoenamine tautomer and the O-protonated covalent enolimine tautomer. It has been postulated that for the catalytic activity, the PLP has to be in the zwitterionic ketoenamine tautomeric form. However, the exact position of the tautomeric equilibrium of Schiff base in the active site of PLP-dependent enzyme is not known yet. In the present work, we investigated the tautomeric equilibrium for the external aldimine state of PLP aspartate (PLP-Asp) Schiff base in the active site of aspartate aminotransferase (AspAT) using combined quantum mechanical and molecular mechanical simulations. The main focus of the present study is to analyze the factors that control the tautomeric equilibrium in the active sites of various PLP-dependent enzymes. The results show that the ketoenamine tautomer is more preferred than the enolimine tautomer both in the gas and aqueous phases as well as in the active site of AspAT. Current simulations show that the active site of AspAT is more suitable for the ketoenamine tautomer compared to the enolimine tautomer. Interestingly, the Tyr225 acts as a proton donor to the phenolic oxygen in the ketoenamine tautomer, while in the covalent enolimine tautomer, it acts as a proton acceptor to the phenolic oxygen. Finally, the metadynamics study confirms this result. The calculated free energy barrier is about 7.5 kcal/mol. A comparative analysis of the microenvironment created by the active site residues of three different PLP-dependent enzymes (aspartate aminotransferase, Dopa decarboxylase, and Ala-racemase) has been carried out to understand the controlling factor(s) of the tautomeric equilibrium. The analysis shows that the intermolecular hydrogen bonding between active site residues and the phenolic oxygen of PLP shifts the tautomeric equilibrium toward the N

Characterization of Resistance Genes and Plasmids from Outbreaks and Illness Clusters Caused by Salmonella Resistant to Ceftriaxone in the United States, 2011–2012

PubMed Central

Folster, Jason P.; Grass, Julian E.; Bicknese, Amelia; Taylor, Julia; Friedman, Cindy R.; Whichard, Jean M.

2017-01-01

Salmonella is an important cause of foodborne illness; however, quickly identifying the source of these infections can be difficult, and source identification is a crucial step in preventing additional illnesses. Although most infections are self-limited, invasive salmonellosis may require antimicrobial treatment. Ceftriaxone, an extended-spectrum cephalosporin, is commonly used for treatment of salmonellosis. Previous studies have identified a correlation between the food animal/retail meat source of ceftriaxone-resistant Salmonella and the type of resistance gene and plasmid it carries. In this study, we examined seven outbreaks of ceftriaxone-resistant Salmonella infections, caused by serotypes Typhimurium, Newport, Heidelberg, and Infantis. All isolates were positive for a plasmid-encoded blaCMY gene. Plasmid incompatibility typing identified five IncI1 and two IncA/C plasmids. Both outbreaks containing blaCMY-IncA/C plasmids were linked to consumption of cattle products. Three of five outbreaks with blaCMY-IncI1 (ST12) plasmids were linked to a poultry source. The remaining IncI1 outbreaks were associated with ground beef (ST20) and tomatoes (ST12). Additionally, we examined isolates from five unsolved clusters of ceftriaxone-resistant Salmonella infections and used our plasmid encoded gene findings to predict the source. Overall, we identified a likely association between the source of ceftriaxone-resistant Salmonella outbreaks and the type of resistance gene/plasmid it carries. PMID:27828730

Regulation of gene expression in plasmid ColE1: delayed expression of the kil gene.

PubMed Central

Zhang, S P; Yan, L F; Zubay, G

1988-01-01

cea, imm, and kil are a cluster of three functionally related genes of the plasmid ColE1. The cea and kil genes are in the same inducible operon, with transcription being initiated from a promoter adjacent to the cea gene. The imm gene is located between the cea and kil genes, but it is transcribed in the opposite direction. Complementary interaction between the imm mRNA and the anti-imm sequences in the middle of the cea-kil transcript causes a pronounced delay in expression of the kil gene when the cea-kil operon is induced. A segment in the overlapping region between the cea and imm genes causes delayed expression of the kil gene in the absence of imm gene transcription. This delay effect increases the yields of colicin synthesized in induced cells. Images PMID:3142845

The Mycobacterium tuberculosis Complex has a Pathway for the Biosynthesis of 4-Formamido-4,6-Dideoxy-d-Glucose.

PubMed

Brown, Haley A; Vinogradov, Evgeny; Gilbert, Michel; Holden, Hazel M

2018-05-15

Recent studies have demonstrated that the O-antigens of some pathogenic bacteria such as Brucella abortus, Francisella tularensis, and Campylobacter jejuni contain quite unusual N-formylated sugars (3-formamido-3,6-dideoxy-d-glucose or 4-formamido-4,6-dideoxy-d-glucose). Typically, four enzymes are required for the formation of such sugars: a thymidylyltransferase, a 4,6-dehydratase, a pyridoxal 5'-phosphate or PLP-dependent aminotransferase, and an N-formyltransferase. To date, there have been no published reports of N-formylated sugars associated with Mycobacterium tuberculosis. A recent investigation from our laboratories, however, has demonstrated that one gene product from M. tuberculosis, Rv3404c, functions as a sugar N-formyltransferase. Given that M. tuberculosis produces l-rhamnose, both a thymidylyltransferase (Rv0334) and a 4,6-dehydratase (Rv3464) required for its formation have been identified. Thus, there is one remaining enzyme needed for the production of an N-formylated sugar in M. tuberculosis, namely a PLP-dependent aminotransferase. Here we demonstrate that the M. tuberculosis rv3402c gene encodes such an enzyme. Our data prove that M. tuberculosis contains all of the enzymatic activities required for the formation of dTDP-4-formamido-4,6-dideoxy-d-glucose. Indeed, the rv3402c gene product likely contributes to virulence or persistence during infection, though its temporal expression and location remain to be determined. This article is protected by copyright. All rights reserved. © 2018 The Protein Society.

Identification of the first intragenic deletion of the PITX2 gene causing an Axenfeld-Rieger Syndrome: case report.

PubMed

de la Houssaye, Guillaume; Bieche, Ivan; Roche, Olivier; Vieira, Véronique; Laurendeau, Ingrid; Arbogast, Laurence; Zeghidi, Hatem; Rapp, Philippe; Halimi, Philippe; Vidaud, Michel; Dufier, Jean-Louis; Menasche, Maurice; Abitbol, Marc

2006-11-29

Axenfeld-Rieger syndrome (ARS) is characterized by bilateral congenital abnormalities of the anterior segment of the eye associated with abnormalities of the teeth, midface, and umbilicus. Most cases of ARS are caused by mutations in the genes encoding PITX2 or FOXC1. Here we describe a family affected by a severe form of ARS. Two members of this family (father and daughter) presented with typical ARS and developed severe glaucoma. The ocular phenotype was much more severe in the daughter than in the father. Magnetic resonance imaging (MRI) detected an aggressive form of meningioma in the father. There was no mutation in the PITX2 gene, determined by exon screening. We identified an intragenic deletion by quantitative genomic PCR analysis and characterized this deletion in detail. Our findings implicate the first intragenic deletion of the PITX2 gene in the pathogenesis of a severe form of ARS in an affected family. This study stresses the importance of a systematic search for intragenic deletions in families affected by ARS and in sporadic cases for which no mutations in the exons or introns of PITX2 have been found. The molecular genetics of some ARS pedigrees should be re-examined with enzymes that can amplify medium and large genomic fragments.

Identification of the first intragenic deletion of the PITX2 gene causing an Axenfeld-Rieger Syndrome: case report

PubMed Central

de la Houssaye, Guillaume; Bieche, Ivan; Roche, Olivier; Vieira, Véronique; Laurendeau, Ingrid; Arbogast, Laurence; Zeghidi, Hatem; Rapp, Philippe; Halimi, Philippe; Vidaud, Michel; Dufier, Jean-Louis; Menasche, Maurice; Abitbol, Marc

2006-01-01

Background Axenfeld-Rieger syndrome (ARS) is characterized by bilateral congenital abnormalities of the anterior segment of the eye associated with abnormalities of the teeth, midface, and umbilicus. Most cases of ARS are caused by mutations in the genes encoding PITX2 or FOXC1. Here we describe a family affected by a severe form of ARS. Case presentation Two members of this family (father and daughter) presented with typical ARS and developed severe glaucoma. The ocular phenotype was much more severe in the daughter than in the father. Magnetic resonance imaging (MRI) detected an aggressive form of meningioma in the father. There was no mutation in the PITX2 gene, determined by exon screening. We identified an intragenic deletion by quantitative genomic PCR analysis and characterized this deletion in detail. Conclusion Our findings implicate the first intragenic deletion of the PITX2 gene in the pathogenesis of a severe form of ARS in an affected family. This study stresses the importance of a systematic search for intragenic deletions in families affected by ARS and in sporadic cases for which no mutations in the exons or introns of PITX2 have been found. The molecular genetics of some ARS pedigrees should be re-examined with enzymes that can amplify medium and large genomic fragments. PMID:17134502

Haplodeficiency of Klotho Gene Causes Arterial Stiffening via Upregulation of Scleraxis Expression and Induction of Autophagy.

PubMed

Chen, Kai; Zhou, Xiaoli; Sun, Zhongjie

2015-11-01

The prevalence of arterial stiffness increases with age, whereas the level of the aging-suppressor protein klotho decreases with age. The objective of this study is to assess whether haplodeficiency of klotho gene causes arterial stiffness and to investigate the underlying mechanism. Pulse wave velocity, a direct measure of arterial stiffness, was increased significantly in klotho heterozygous (klotho(+/-)) mice versus their age-matched wild-type (WT) littermates, suggesting that haplodeficiency of klotho causes arterial stiffening. Notably, plasma aldosterone levels were elevated significantly in klotho(+/-) mice. Treatment with eplerenone (6 mg/kg per day IP), an aldosterone receptor blocker, abolished klotho deficiency-induced arterial stiffening in klotho(+/-) mice. Klotho deficiency was associated with increased collagen and decreased elastin contents in the media of aortas. In addition, arterial matrix metalloproteinase-2, matrix metalloproteinase-9, and transforming growth factor-β1 expression and myofibroblast differentiation were increased in klotho(+/-) mice. These klotho deficiency-related changes can be blocked by eplerenone. Protein expression of scleraxis, a transcription factor for collagen synthesis, and LC3-II/LC3-I, an index of autophagy, were upregulated in aortas of klotho(+/-) mice, which can be abolished by eplerenone. In cultured mouse aortic smooth muscle cells, aldosterone increased collagen-1 expression that can be completely eliminated by small interfering RNA knockdown of scleraxis. Interestingly, aldosterone decreased elastin levels in smooth muscle cells, which can be abolished by small interfering RNA knockdown of Beclin-1, an autophagy-related gene. In conclusion, this study demonstrated for the first time that klotho deficiency-induced arterial stiffening may involve aldosterone-mediated upregulation of scleraxis and induction of autophagy, which led to increased collagen-1 expression and decreased elastin levels, respectively. �

Human amyloid β peptide and tau co-expression impairs behavior and causes specific gene expression changes in Caenorhabditis elegans.

PubMed

Wang, Chenyin; Saar, Valeria; Leung, Ka Lai; Chen, Liang; Wong, Garry

2018-01-01

Alzheimer's disease (AD) is a progressive neurodegenerative disorder characterized by the presence of extracellular amyloid plaques consisting of Amyloid-β peptide (Aβ) aggregates and neurofibrillary tangles formed by aggregation of hyperphosphorylated microtubule-associated protein tau. We generated a novel invertebrate model of AD by crossing Aβ1-42 (strain CL2355) with either pro-aggregating tau (strain BR5270) or anti-aggregating tau (strain BR5271) pan-neuronal expressing transgenic Caenorhabditis elegans. The lifespan and progeny viability of the double transgenic strains were significantly decreased compared with wild type N2 (P<0.0001). In addition, co-expression of these transgenes interfered with neurotransmitter signaling pathways, caused deficits in chemotaxis associative learning, increased protein aggregation visualized by Congo red staining, and increased neuronal loss. Global transcriptomic RNA-seq analysis revealed 248 up- and 805 down-regulated genes in N2 wild type versus Aβ1-42+pro-aggregating tau animals, compared to 293 up- and 295 down-regulated genes in N2 wild type versus Aβ1-42+anti-aggregating tau animals. Gene set enrichment analysis of Aβ1-42+pro-aggregating tau animals uncovered up-regulated annotation clusters UDP-glucuronosyltransferase (5 genes, P<4.2E-4), protein phosphorylation (5 genes, P<2.60E-02), and aging (5 genes, P<8.1E-2) while the down-regulated clusters included nematode cuticle collagen (36 genes, P<1.5E-21). RNA interference of 13 available top up-regulated genes in Aβ1-42+pro-aggregating tau animals revealed that F-box family genes and nep-4 could enhance life span deficits and chemotaxis deficits while Y39G8C.2 (TTBK2) could suppress these behaviors. Comparing the list of regulated genes from C. elegans to the top 60 genes related to human AD confirmed an overlap of 8 genes: patched homolog 1, PTCH1 (ptc-3), the Rab GTPase activating protein, TBC1D16 (tbc-16), the WD repeat and FYVE domain-containing protein 3

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.

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

Matchmaking facilitates the diagnosis of an autosomal-recessive mitochondrial disease caused by biallelic mutation of the tRNA isopentenyltransferase (TRIT1) gene.

PubMed

Kernohan, Kristin D; Dyment, David A; Pupavac, Mihaela; Cramer, Zvi; McBride, Arran; Bernard, Genevieve; Straub, Isabella; Tetreault, Martine; Hartley, Taila; Huang, Lijia; Sell, Erick; Majewski, Jacek; Rosenblatt, David S; Shoubridge, Eric; Mhanni, Aziz; Myers, Tara; Proud, Virginia; Vergano, Samanta; Spangler, Brooke; Farrow, Emily; Kussman, Jennifer; Safina, Nicole; Saunders, Carol; Boycott, Kym M; Thiffault, Isabelle

2017-05-01

Deleterious variants in the same gene present in two or more families with overlapping clinical features provide convincing evidence of a disease-gene association; this can be a challenge in the study of ultrarare diseases. To facilitate the identification of additional families, several groups have created "matching" platforms. We describe four individuals from three unrelated families "matched" by GeneMatcher and MatchMakerExchange. Individuals had microcephaly, developmental delay, epilepsy, and recessive mutations in TRIT1. A single homozygous mutation in TRIT1 associated with similar features had previously been reported in one family. The identification of these individuals provides additional evidence to support TRIT1 as the disease-causing gene and interprets the variants as "pathogenic." TRIT1 functions to modify mitochondrial tRNAs and is necessary for protein translation. We show that dysfunctional TRIT1 results in decreased levels of select mitochondrial proteins. Our findings confirm the TRIT1 disease association and advance the phenotypic and molecular understanding of this disorder. © 2017 Wiley Periodicals, Inc.

Toxoplasma Gondii Infection of Chicken Embryos Causes Retinal Changes and Modulates HSP90B1 Gene Expression: A Promising Ocular Toxoplasmosis Model.

PubMed

Nasaré, Alex M; Tedesco, Roberto C; Cristovam, Priscila C; Cenedese, Marcos A; Galisteo, Andrés J; Andrade, Heitor F; Gomes, José Álvaro P; Guimarães, Érik V; Barbosa, Helene S; Alonso, Luis G

2015-12-01

HSP90B1 is a gene that codifies heat shock protein 108 (HSP108) that belongs to a group of proteins induced under stress situation, and it has close relation with the nervous system, especially in the retina. Toxoplasma gondii causes ocular toxoplasmosis that has been associated with a late manifestation of the congenital toxoplasmosis although experimental models show that morphological alterations are already present during embryological development. Here, we used 18 eyes of Gallus domesticus embryos in 7th and 20th embryonic days to establish a model of congenital ocular toxoplasmosis, experimentally infected in its fifth day correlating with HSP90B1 gene expression. Embryos' eyes were histologically evaluated, and gene expression was performed by real-time polymerase chain reaction (PCR). Our data showed parasite present in the choroid, unusual migration of retinal pigment epithelium, and chorioretinal scars, and a tendency to a lower expression of the HSP90B1 gene upon experimental infection. This is a promising model to better understand T. gondii etiopathogeny.

Genetic recombination as a major cause of mutagenesis in the human globin gene clusters.

PubMed

Borg, Joseph; Georgitsi, Marianthi; Aleporou-Marinou, Vassiliki; Kollia, Panagoula; Patrinos, George P

2009-12-01

Homologous recombination is a frequent phenomenon in multigene families and as such it occurs several times in both the alpha- and beta-like globin gene families. In numerous occasions, genetic recombination has been previously implicated as a major mechanism that drives mutagenesis in the human globin gene clusters, either in the form of unequal crossover or gene conversion. Unequal crossover results in the increase or decrease of the human globin gene copies, accompanied in the majority of cases with minor phenotypic consequences, while gene conversion contributes either to maintaining sequence homogeneity or generating sequence diversity. The role of genetic recombination, particularly gene conversion in the evolution of the human globin gene families has been discussed elsewhere. Here, we summarize our current knowledge and review existing experimental evidence outlining the role of genetic recombination in the mutagenic process in the human globin gene families.

Mutations of Cystic Fibrosis Transmembrane Conductance Regulator Gene Cause a Monocyte-Selective Adhesion Deficiency.

PubMed

Sorio, Claudio; Montresor, Alessio; Bolomini-Vittori, Matteo; Caldrer, Sara; Rossi, Barbara; Dusi, Silvia; Angiari, Stefano; Johansson, Jan E; Vezzalini, Marzia; Leal, Teresinha; Calcaterra, Elisa; Assael, Baroukh M; Melotti, Paola; Laudanna, Carlo

2016-05-15

Cystic fibrosis (CF) is a common genetic disease caused by mutations of the cystic fibrosis transmembrane conductance regulator (CFTR) gene. Persistent lung inflammation, characterized by increasing polymorphonuclear leukocyte recruitment, is a major cause of the decline in respiratory function in patients with CF and is a leading cause of morbidity and mortality. CFTR is expressed in various cell types, including leukocytes, but its involvement in the regulation of leukocyte recruitment is unknown. We evaluated whether CF leukocytes might present with alterations in cell adhesion and migration, a key process governing innate and acquired immune responses. We used ex vivo adhesion and chemotaxis assays, flow cytometry, immunofluorescence, and GTPase activity assays in this study. We found that chemoattractant-induced activation of β1 and β2 integrins and of chemotaxis is defective in mononuclear cells isolated from patients with CF. In contrast, polymorphonuclear leukocyte adhesion and chemotaxis were normal. The functionality of β1 and β2 integrins was restored by treatment of CF monocytes with the CFTR-correcting drugs VRT325 and VX809. Moreover, treatment of healthy monocytes with the CFTR inhibitor CFTR(inh)-172 blocked integrin activation by chemoattractants. In a murine model of lung inflammation, we found that integrin-independent migration of CF monocytes into the lung parenchyma was normal, whereas, in contrast, integrin-dependent transmigration into the alveolar space was impaired. Finally, signal transduction analysis showed that, in CF monocytes, chemoattractant-triggered activation of RhoA and CDC42 Rho small GTPases (controlling integrin activation and chemotaxis, respectively) was strongly deficient. Altogether, these data highlight the critical regulatory role of CFTR in integrin activation by chemoattractants in monocytes and identify CF as a new, cell type-selective leukocyte adhesion deficiency disease, providing new insights into CF

Decreased necrotizing fasciitis capacity caused by a single nucleotide mutation that alters a multiple gene virulence axis

PubMed Central

Olsen, Randall J.; Sitkiewicz, Izabela; Ayeras, Ara A.; Gonulal, Vedia E.; Cantu, Concepcion; Beres, Stephen B.; Green, Nicole M.; Lei, Benfang; Humbird, Tammy; Greaver, Jamieson; Chang, Ellen; Ragasa, Willie P.; Montgomery, Charles A.; Cartwright, Joiner; McGeer, Allison; Low, Donald E.; Whitney, Adeline R.; Cagle, Philip T.; Blasdel, Terry L.; DeLeo, Frank R.; Musser, James M.

2010-01-01

Single-nucleotide changes are the most common cause of natural genetic variation among members of the same species, but there is remarkably little information bearing on how they alter bacterial virulence. We recently discovered a single-nucleotide mutation in the group A Streptococcus genome that is epidemiologically associated with decreased human necrotizing fasciitis (“flesh-eating disease”). Working from this clinical observation, we find that wild-type mtsR function is required for group A Streptococcus to cause necrotizing fasciitis in mice and nonhuman primates. Expression microarray analysis revealed that mtsR inactivation results in overexpression of PrsA, a chaperonin involved in posttranslational maturation of SpeB, an extracellular cysteine protease. Isogenic mutant strains that overexpress prsA or lack speB had decreased secreted protease activity in vivo and recapitulated the necrotizing fasciitis-negative phenotype of the ΔmtsR mutant strain in mice and monkeys. mtsR inactivation results in increased PrsA expression, which in turn causes decreased SpeB secreted protease activity and reduced necrotizing fasciitis capacity. Thus, a naturally occurring single-nucleotide mutation dramatically alters virulence by dysregulating a multiple gene virulence axis. Our discovery has broad implications for the confluence of population genomics and molecular pathogenesis research. PMID:20080771

Decreased necrotizing fasciitis capacity caused by a single nucleotide mutation that alters a multiple gene virulence axis.

PubMed

Olsen, Randall J; Sitkiewicz, Izabela; Ayeras, Ara A; Gonulal, Vedia E; Cantu, Concepcion; Beres, Stephen B; Green, Nicole M; Lei, Benfang; Humbird, Tammy; Greaver, Jamieson; Chang, Ellen; Ragasa, Willie P; Montgomery, Charles A; Cartwright, Joiner; McGeer, Allison; Low, Donald E; Whitney, Adeline R; Cagle, Philip T; Blasdel, Terry L; DeLeo, Frank R; Musser, James M

2010-01-12

Single-nucleotide changes are the most common cause of natural genetic variation among members of the same species, but there is remarkably little information bearing on how they alter bacterial virulence. We recently discovered a single-nucleotide mutation in the group A Streptococcus genome that is epidemiologically associated with decreased human necrotizing fasciitis ("flesh-eating disease"). Working from this clinical observation, we find that wild-type mtsR function is required for group A Streptococcus to cause necrotizing fasciitis in mice and nonhuman primates. Expression microarray analysis revealed that mtsR inactivation results in overexpression of PrsA, a chaperonin involved in posttranslational maturation of SpeB, an extracellular cysteine protease. Isogenic mutant strains that overexpress prsA or lack speB had decreased secreted protease activity in vivo and recapitulated the necrotizing fasciitis-negative phenotype of the DeltamtsR mutant strain in mice and monkeys. mtsR inactivation results in increased PrsA expression, which in turn causes decreased SpeB secreted protease activity and reduced necrotizing fasciitis capacity. Thus, a naturally occurring single-nucleotide mutation dramatically alters virulence by dysregulating a multiple gene virulence axis. Our discovery has broad implications for the confluence of population genomics and molecular pathogenesis research.

Domain Organization and Evolution of the Highly Divergent 5′ Coding Region of Genomes of Arteriviruses, Including the Novel Possum Nidovirus

PubMed Central

Gulyaeva, Anastasia; Hoogendoorn, Erik; Giles, Julia; Samborskiy, Dmitry

2017-01-01

ABSTRACT In five experimentally characterized arterivirus species, the 5′-end genome coding region encodes the most divergent nonstructural proteins (nsp's), nsp1 and nsp2, which include papain-like proteases (PLPs) and other poorly characterized domains. These are involved in regulation of transcription, polyprotein processing, and virus-host interaction. Here we present results of a bioinformatics analysis of this region of 14 arterivirus species, including that of the most distantly related virus, wobbly possum disease virus (WPDV), determined by a modified 5′ rapid amplification of cDNA ends (RACE) protocol. By combining profile-profile comparisons and phylogeny reconstruction, we identified an association of the four distinct domain layouts of nsp1-nsp2 with major phylogenetic lineages, implicating domain gain, including duplication, and loss in the early nsp1 evolution. Specifically, WPDV encodes highly divergent homologs of PLP1a, PLP1b, PLP1c, and PLP2, with PLP1a lacking the catalytic Cys residue, but does not encode nsp1 Zn finger (ZnF) and “nuclease” domains, which are conserved in other arteriviruses. Unexpectedly, our analysis revealed that the only catalytically active nsp1 PLP of equine arteritis virus (EAV), known as PLP1b, is most similar to PLP1c and thus is likely to be a PLP1b paralog. In all non-WPDV arteriviruses, PLP1b/c and PLP1a show contrasting patterns of conservation, with the N- and C-terminal subdomains, respectively, being enriched with conserved residues, which is indicative of different functional specializations. The least conserved domain of nsp2, the hypervariable region (HVR), has its size varied 5-fold and includes up to four copies of a novel PxPxPR motif that is potentially recognized by SH3 domain-containing proteins. Apparently, only EAV lacks the signal that directs −2 ribosomal frameshifting in the nsp2 coding region. IMPORTANCE Arteriviruses comprise a family of mammalian enveloped positive-strand RNA viruses

Mutated-leptin gene transfer induces increases in body weight by electroporation and hydrodynamics-based gene delivery in mice.

PubMed

Xiang, Lan; Murai, Atsushi; Muramatsu, Tatsuo

2005-12-01

To investigate whether in vivo gene transfer causes leptin-antagonistic effects on food intake, animal body weight and fat tissue weight, the R128Q mutated-leptin gene, an R to Q substitution at position 128 of mouse leptin, was transferred into mouse liver and leg muscle by electroporation and hydrodynamics-based gene delivery. Mutated-leptin gene transfer by electroporation caused significant increases in body weight at 5 days and after (5.4% increase relative to control; p<0.05). Hydrodynamics-based gene delivery of the mutated-leptin gene also caused an increase in body weight (3.0% increase relative to control; p<0.05). Mutated-leptin gene transfer by electroporation significantly increased the tissue weight of epididymal white fat and neuropeptide Y mRNA expression in the hypothalamus compared with those of the control group 3 weeks after gene transfer (p<0.05). These results suggest that mutated-leptin gene transfer successfully produced leptin-antagonistic effects by modulating the central regulator of energy homeostasis. Also, the extent of leptin-antagonistic effects by electroporation was much higher than hydrodynamics-based gene delivery, with at least single gene transfer.

ALK1 expression in oral lichen planus: a possible relation to microvessel density.

PubMed

Hazzaa, Hala H A; El-Wakeel, Naglaa M; Attia, Enas A S; Abo Hager, Eman A

2016-05-01

To assess the expression of activin receptor-like kinase 1 (ALK1) and investigate its possible relationship with microvessel density (MVD) in different forms of oral lichen planus (OLP) compared to controls' biopsies. Biopsies from 20 reticular/papular OLP (R/PLP), 20 atrophic/erosive OLP (A/ELP) patients, and 20 healthy subjects were immunohistochemically analyzed and statistically compared and correlated for ALK1 expression and MVD as assessed by CD34 expression. All OLP specimens revealed the presence of positive cytoplasmic CD34 immunostaining in endothelial cells, with statistically high significant MVD in each of R/PLP (Median; M = 4.40) and A/ELP (M = 7.69) compared to controls (M = 1.16) (P < 0.001). Statistically significant MVD was found in A/ELP compared to R/PLP (P < 0.001). All control specimens revealed negative ALK1 immunostaining of the few inflammatory cells found, while 85% of A/ELP cases and 70% of R/PLP cases showed positively immunostained sections for ALK-1, with statistically significant higher ALK1 expression In A/ELP (M = 1.95) compared to R/PLP (M = 0.86) (P = 0.005). No significant correlation between CD34 and ALK1 was detected in R/PLP (r = 0.081), while a barely moderate positive correlation was found in A/ELP (r = 0.396). ALK1 expression and MVD are increased in OLP, particularly in A/ELP type. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Contrasting invertebrate immune defense behaviors caused by a single gene, the Caenorhabditis elegans neuropeptide receptor gene npr-1.

PubMed

Nakad, Rania; Snoek, L Basten; Yang, Wentao; Ellendt, Sunna; Schneider, Franziska; Mohr, Timm G; Rösingh, Lone; Masche, Anna C; Rosenstiel, Philip C; Dierking, Katja; Kammenga, Jan E; Schulenburg, Hinrich

2016-04-11

The invertebrate immune system comprises physiological mechanisms, physical barriers and also behavioral responses. It is generally related to the vertebrate innate immune system and widely believed to provide nonspecific defense against pathogens, whereby the response to different pathogen types is usually mediated by distinct signalling cascades. Recent work suggests that invertebrate immune defense can be more specific at least at the phenotypic level. The underlying genetic mechanisms are as yet poorly understood. We demonstrate in the model invertebrate Caenorhabditis elegans that a single gene, a homolog of the mammalian neuropeptide Y receptor gene, npr-1, mediates contrasting defense phenotypes towards two distinct pathogens, the Gram-positive Bacillus thuringiensis and the Gram-negative Pseudomonas aeruginosa. Our findings are based on combining quantitative trait loci (QTLs) analysis with functional genetic analysis and RNAseq-based transcriptomics. The QTL analysis focused on behavioral immune defense against B. thuringiensis, using recombinant inbred lines (RILs) and introgression lines (ILs). It revealed several defense QTLs, including one on chromosome X comprising the npr-1 gene. The wildtype N2 allele for the latter QTL was associated with reduced defense against B. thuringiensis and thus produced an opposite phenotype to that previously reported for the N2 npr-1 allele against P. aeruginosa. Analysis of npr-1 mutants confirmed these contrasting immune phenotypes for both avoidance behavior and nematode survival. Subsequent transcriptional profiling of C. elegans wildtype and npr-1 mutant suggested that npr-1 mediates defense against both pathogens through p38 MAPK signaling, insulin-like signaling, and C-type lectins. Importantly, increased defense towards P. aeruginosa seems to be additionally influenced through the induction of oxidative stress genes and activation of GATA transcription factors, while the repression of oxidative stress genes

Horizontal gene transfer and gene dosage drives adaptation to wood colonization in a tree pathogen

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

Dhillon, Braham; Feau, Nicolas; Aerts, Andrea L.

Some of the most damaging tree diseases are caused by pathogens that induce cankers, a stem deformation often lethal. To investigate the cause of this adaptation, we sequenced the genomes of poplar pathogens that do and do not cause cankers. We found a unique cluster of genes that produce secondary metabolites and are co-activated when the canker pathogen is grown on poplar wood and leaves. The gene genealogy is discordant with the species phylogeny, showing a signature of horizontal transfer from fungi associated with wood decay. Furthermore, genes encoding hemicellulose-degrading enzymes are up-regulated on poplar wood chips, with some havingmore » been acquired horizontally. In conclusion, we propose that adaptation to colonize poplar woody stems is the result of acquisition of these genes.« less

Horizontal gene transfer and gene dosage drives adaptation to wood colonization in a tree pathogen

DOE PAGES

Dhillon, Braham; Feau, Nicolas; Aerts, Andrea L.; ...

2015-03-02

Some of the most damaging tree diseases are caused by pathogens that induce cankers, a stem deformation often lethal. To investigate the cause of this adaptation, we sequenced the genomes of poplar pathogens that do and do not cause cankers. We found a unique cluster of genes that produce secondary metabolites and are co-activated when the canker pathogen is grown on poplar wood and leaves. The gene genealogy is discordant with the species phylogeny, showing a signature of horizontal transfer from fungi associated with wood decay. Furthermore, genes encoding hemicellulose-degrading enzymes are up-regulated on poplar wood chips, with some havingmore » been acquired horizontally. In conclusion, we propose that adaptation to colonize poplar woody stems is the result of acquisition of these genes.« less

Exposure to Nickel, Chromium, or Cadmium Causes Distinct Changes in the Gene Expression Patterns of a Rat Liver Derived Cell Line

PubMed Central

Permenter, Matthew G.; Lewis, John A.; Jackson, David A.

2011-01-01

Many heavy metals, including nickel (Ni), cadmium (Cd), and chromium (Cr) are toxic industrial chemicals with an exposure risk in both occupational and environmental settings that may cause harmful outcomes. While these substances are known to produce adverse health effects leading to disease or health problems, the detailed mechanisms remain unclear. To elucidate the processes involved in the toxicity of nickel, cadmium, and chromium at the molecular level and to perform a comparative analysis, H4-II-E-C3 rat liver-derived cell lines were treated with soluble salts of each metal using concentrations derived from viability assays, and gene expression patterns were determined with DNA microarrays. We identified both common and unique biological responses to exposure to the three metals. Nickel, cadmium, chromium all induced oxidative stress with both similar and unique genes and pathways responding to this stress. Although all three metals are known to be genotoxic, evidence for DNA damage in our study only exists in response to chromium. Nickel induced a hypoxic response as well as inducing genes involved in chromatin structure, perhaps by replacing iron in key proteins. Cadmium distinctly perturbed genes related to endoplasmic reticulum stress and invoked the unfolded protein response leading to apoptosis. With these studies, we have completed the first gene expression comparative analysis of nickel, cadmium, and chromium in H4-II-E-C3 cells. PMID:22110744

COL4A4 gene study of a European population: description of new mutations causing autosomal dominant Alport syndrome.

PubMed

Rosado, Consolación; Bueno, Elena; Felipe, Carmen; González-Sarmiento, Rogelio

2014-01-01

Autosomal forms of Alport syndrome represent 20% of all patients (15% recessive and 5% dominant). They are caused by mutations in the COL4A3 and COL4A4 genes, which encode a-3 and a-4 collagen IV chains of the glomerular basement membrane, cochlea and eye. Thin basement membrane nephropathy may affect up to 1% of the population. The pattern of inheritance in the 40% of cases is the same as autosomal dominant Alport syndrome: heterozygous mutations in these genes. The aim of this study is to detect new pathogenic mutations in the COL4A4 gene in the patients previously diagnosed with autosomal Alport syndrome and thin basement membrane nephropathy in our hospital. We conducted a clinical and genetic study in eleven patients belonging to six unrelated families with aforementioned clinical symptoms and a negative study of COL4A3 gene. The molecular study was made by conformation of sensitive gel electrophoresis (CSGE) and direct sequencing of the fragments that show an altered electrophoretic migration pattern. We found two pathogenic mutations, not yet described: IVS3 + 1G > C is a replacement of Guanine to Cytosine in position +1 of intron 3, in the splicing region, which leads to a pathogenic mutation. c.4267C > T; p.P1423S is a missense mutation, also considered pathogenic. We also found seven new polymorphisms. We describe two new pathogenic mutations, responsible for autosomal dominant Alport syndrome. The other families of the study were undiagnosed owing to problems in the method employed and the possibility of mutations in other genes, giving rise to other diseases with similar symptoms.

Molecular characterization of novel pyridoxal-5'-phosphate-dependent enzymes from the human microbiome.

PubMed

Fleischman, Nicholas M; Das, Debanu; Kumar, Abhinav; Xu, Qingping; Chiu, Hsiu-Ju; Jaroszewski, Lukasz; Knuth, Mark W; Klock, Heath E; Miller, Mitchell D; Elsliger, Marc-André; Godzik, Adam; Lesley, Scott A; Deacon, Ashley M; Wilson, Ian A; Toney, Michael D

2014-08-01

Pyridoxal-5'-phosphate or PLP, the active form of vitamin B6, is a highly versatile cofactor that participates in a large number of mechanistically diverse enzymatic reactions in basic metabolism. PLP-dependent enzymes account for ∼1.5% of most prokaryotic genomes and are estimated to be involved in ∼4% of all catalytic reactions, making this an important class of enzymes. Here, we structurally and functionally characterize three novel PLP-dependent enzymes from bacteria in the human microbiome: two are from Eubacterium rectale, a dominant, nonpathogenic, fecal, Gram-positive bacteria, and the third is from Porphyromonas gingivalis, which plays a major role in human periodontal disease. All adopt the Type I PLP-dependent enzyme fold and structure-guided biochemical analysis enabled functional assignments as tryptophan, aromatic, and probable phosphoserine aminotransferases. © 2014 The Protein Society.

Hydrocephalus due to multiple ependymal malformations is caused by mutations in the MPDZ gene.

PubMed

Saugier-Veber, Pascale; Marguet, Florent; Lecoquierre, François; Adle-Biassette, Homa; Guimiot, Fabien; Cipriani, Sara; Patrier, Sophie; Brasseur-Daudruy, Marie; Goldenberg, Alice; Layet, Valérie; Capri, Yline; Gérard, Marion; Frébourg, Thierry; Laquerrière, Annie

2017-05-01

Congenital hydrocephalus is considered as either acquired due to haemorrhage, infection or neoplasia or as of developmental nature and is divided into two subgroups, communicating and obstructive. Congenital hydrocephalus is either syndromic or non-syndromic, and in the latter no cause is found in more than half of the patients. In patients with isolated hydrocephalus, L1CAM mutations represent the most common aetiology. More recently, a founder mutation has also been reported in the MPDZ gene in foetuses presenting massive hydrocephalus, but the neuropathology remains unknown. We describe here three novel homozygous null mutations in the MPDZ gene in foetuses whose post-mortem examination has revealed a homogeneous phenotype characterized by multiple ependymal malformations along the aqueduct of Sylvius, the third and fourth ventricles as well as the central canal of the medulla, consisting in multifocal rosettes with immature cell accumulation in the vicinity of ependymal lining early detached from the ventricular zone. MPDZ also named MUPP1 is an essential component of tight junctions which are expressed from early brain development in the choroid plexuses and ependyma. Alterations in the formation of tight junctions within the ependyma very likely account for the lesions observed and highlight for the first time that primary multifocal ependymal malformations of the ventricular system is genetically determined in humans. Therefore, MPDZ sequencing should be performed when neuropathological examination reveals multifocal ependymal rosette formation within the aqueduct of Sylvius, of the third and fourth ventricles and of the central canal of the medulla.

Terahertz spectroscopy for the isothermal detection of bacterial DNA by magnetic bead-based rolling circle amplification.

PubMed

Yang, Xiang; Yang, Ke; Zhao, Xiang; Lin, Zhongquan; Liu, Zhiyong; Luo, Sha; Zhang, Yang; Wang, Yunxia; Fu, Weiling

2017-12-04

The demand for rapid and sensitive bacterial detection is continuously increasing due to the significant requirements of various applications. In this study, a terahertz (THz) biosensor based on rolling circle amplification (RCA) was developed for the isothermal detection of bacterial DNA. The synthetic bacterium-specific sequence of 16S rDNA hybridized with a padlock probe (PLP) that contains a sequence fully complementary to the target sequence at the 5' and 3' ends. The linear PLP was circularized by ligation to form a circular PLP upon recognition of the target sequence; then the capture probe (CP) immobilized on magnetic beads (MBs) acted as a primer to initialize RCA. As DNA molecules are much less absorptive than water molecules in the THz range, the RCA products on the surface of the MBs cause a significant decrease in THz absorption, which can be sensitively probed by THz spectroscopy. Our results showed that 0.12 fmol of synthetic bacterial DNA and 0.05 ng μL -1 of genomic DNA could be effectively detected using this assay. In addition, the specificity of this strategy was demonstrated by its low signal response to interfering bacteria. The proposed strategy not only represents a new method for the isothermal detection of the target bacterial DNA but also provides a general methodology for sensitive and specific DNA biosensing using THz spectroscopy.

Organization and evolution of mating-type genes in three Stagonosporopsis species causing gummy stem blight of cucurbits and leaf spot and dry rot of papaya.

PubMed

Li, Hao-Xi; Gottilla, Thomas M; Brewer, Marin Talbot

2017-10-01

Population divergence and speciation of closely related lineages can result from reproductive differences leading to genetic isolation. An increasing number of fungal diseases of plants and animals have been determined to be caused by morphologically indistinguishable species that are genetically distinct, thereby representing cryptic species. We were interested in identifying if mating systems among three Stagonosporopsis species (S. citrulli, S. cucurbitacearum, and S. caricae) causing gummy stem blight (GSB) of cucurbits or leaf spot and dry rot of papaya differed, possibly underlying species divergence. Additionally, we were interested in identifying evolutionary pressures acting on the genes controlling mating in these fungi. The mating-type loci (MAT1) of three isolates from each of the three species were identified in draft genome sequences. For the three species, MAT1 was structurally identical and contained both mating-type genes necessary for sexual reproduction, which suggests that all three species are homothallic. However, both MAT1-1-1 and MAT1-2-1 were divergent among species showing rapid evolution with a much greater number of amino acid-changing substitutions detected for the reproductive genes compared with genes flanking MAT1. Positive selection was detected in MAT1-2-1, especially in the highly conserved high mobility group (MATA_HMG-box) domain. Thus, the mating-type genes are rapidly evolving in GSB fungi, but a difference in mating systems among the three species does not underlie their divergence. Copyright © 2017 British Mycological Society. Published by Elsevier Ltd. All rights reserved.

A mutation in the LAMC2 gene causes the Herlitz junctional epidermolysis bullosa (H-JEB) in two French draft horse breeds.

PubMed

Milenkovic, Dragan; Chaffaux, Stéphane; Taourit, Sead; Guérin, Gérard

2003-01-01

Epidermolysis bullosa (EB) is a heterogeneous group of inherited diseases characterised by skin blistering and fragility. In humans, one of the most severe forms of EB known as Herlitz-junctional EB (H-JEB), is caused by mutations in the laminin 5 genes. EB has been described in several species, like cattle, sheep, dogs, cats and horses where the mutation, a cytosine insertion in exon 10 of the LAMC2 gene, was very recently identified in Belgian horses as the mutation responsible for JEB. In this study, the same mutation was found to be totally associated with the JEB phenotype in two French draft horse breeds, Trait Breton and Trait Comtois. This result provides breeders a molecular test to better manage their breeding strategies by genetic counselling.

Male infertility and its genetic causes.

PubMed

Miyamoto, Toshinobu; Minase, Gaku; Okabe, Kimika; Ueda, Hiroto; Sengoku, Kazuo

2015-10-01

Infertility is a serious social problem in advanced nations, with male factor infertility accounting for approximately half of all cases of infertility. Here, we aim to discuss our laboratory results in the context of recent literature on critical genes residing on the Y chromosome or autosomes that play important roles in human spermatogenesis. The PubMed database was systematically searched using the following keywords: 'genetics of male factor infertility'; 'male infertility genes', 'genetics of spermatogenesis' to retrieve information for this review. Striking progress has recently been made in the elucidation of mechanisms of spermatogenesis using knockout mouse models. This information has, in many cases, not been directly translatable to humans. Nevertheless, mutations in several critical genes have been shown to cause male infertility. We discuss here the contribution to male factor infertility of a number of genes identified in the azoospermia factor (AZF) region on the Y chromosome, as well as the autosomally located genes: SYKP3, KLHL10, AURKC and SPATA16. Non-obstructive azoospermia is the most severe form of azoospermia. However, the presence of spermatozoa can only be confirmed through procedures, which may prove to be unnecessary. Elucidation of the genes underlying male factor infertility, and thereby a better understanding of the mechanisms that cause it, will result in more tailored, evidence-based decisions in treatment of patients. © 2015 Japan Society of Obstetrics and Gynecology.

Altered Gene Expression in Three Plant Species in Response to Treatment with Nep1, a Fungal Protein That Causes Necrosis

PubMed Central

Keates, Sarah E.; Kostman, Todd A.; Anderson, James D.; Bailey, Bryan A.

2003-01-01

Nep1 is an extracellular fungal protein that causes necrosis when applied to many dicotyledonous plants, including invasive weed species. Using transmission electron microscopy, it was determined that application of Nep1 (1.0 μg mL–1, 0.1% [v/v] Silwet-L77) to Arabidopsis and two invasive weed species, spotted knapweed (Centaurea maculosa) and dandelion (Taraxacum officinale), caused a reduction in the thickness of the cuticle and a breakdown of chloroplasts 1 to 4 h after treatment. Membrane breakdown was most severe in cells closest to the surface of application. Differential display was used to isolate cDNA clones from the three species showing differential expression in response to Nep1 treatment. Differential gene expression was observed for a putative serpin (CmSER-1) and a calmodulin-like (CmCAL-1) protein from spotted knapweed, and a putative protein phosphatase 2C (ToPP2C-1) and cytochrome P-450 (ToCYP-1) protein from dandelion. In addition, differential expression was observed for genes coding for a putative protein kinase (AtPK-1), a homolog (AtWI-12) of wound-induced WI12, a homolog (AtLEA-1) of late embryogenesis abundant LEA-5, a WRKY-18 DNA-binding protein (AtWRKY-18), and a phospholipase D (AtPLD-1) from Arabidopsis. Genes showing elevated mRNA levels in Nep1-treated (5 μg mL–1, 0.1% [v/v] Silwet-L77) leaves 15 min after Nep1 treatment included CmSER-1 and CmCAL-1 for spotted knapweed, ToCYP-1 and CmCAL-1 for dandelion, and AtPK-1, AtWRKY-18, AtWI-12, and AtLEA-1 for Arabidopsis. Levels of mRNA for AtPLD-1 (Arabidopsis) and ToPP2C-1 (dandelion) decreased rapidly in Silwet-l77-treated plants between 15 min and 4 h of treatment, but were maintained or decreased more slowly over time in Nep1-treated (5 μg mL–1, 0.1% [v/v] Silwet-L77) leaves. In general, increases in mRNA band intensities were in the range of two to five times, with only ToCYP-1 in dandelion exceeding an increase of 10 times. The identified genes have been shown to be involved

Role of zebrafish cytochrome P450 CYP1C genes in the reduced mesencephalic vein blood flow caused by activation of AHR2

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

Kubota, Akira, E-mail: akubota@whoi.edu; Biology Department, Woods Hole Oceanographic Institution, Woods Hole, MA 02543; Stegeman, John J.

2011-06-15

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) causes various signs of toxicity in early life stages of vertebrates through activation of the aryl hydrocarbon receptor (AHR). We previously reported a sensitive and useful endpoint of TCDD developmental toxicity in zebrafish, namely a decrease in blood flow in the dorsal midbrain, but downstream genes involved in the effect are not known. The present study addressed the role of zebrafish cytochrome P450 1C (CYP1C) genes in association with a decrease in mesencephalic vein (MsV) blood flow. The CYP1C subfamily was recently discovered in fish and includes the paralogues CYP1C1 and CYP1C2, both of which are induced viamore » AHR2 in zebrafish embryos. We used morpholino antisense oligonucleotides (MO or morpholino) to block initiation of translation of the target genes. TCDD-induced mRNA expression of CYP1Cs and a decrease in MsV blood flow were both blocked by gene knockdown of AHR2. Gene knockdown of CYP1C1 by two different morpholinos and CYP1C2 by two different morpholinos, but not by their 5 nucleotide-mismatch controls, was effective in blocking reduced MsV blood flow caused by TCDD. The same CYP1C-MOs prevented reduction of blood flow in the MsV caused by {beta}-naphthoflavone (BNF), representing another class of AHR agonists. Whole-mount in situ hybridization revealed that mRNA expression of CYP1C1 and CYP1C2 was induced by TCDD most strongly in branchiogenic primordia and pectoral fin buds. In situ hybridization using head transverse sections showed that TCDD increased the expression of both CYP1Cs in endothelial cells of blood vessels, including the MsV. These results indicate a potential role of CYP1C1 and CYP1C2 in the local circulation failure induced by AHR2 activation in the dorsal midbrain of the zebrafish embryo. - Research Highlights: > We examine the roles of zebrafish CYP1C1 and CYP1C2 in TCDD developmental toxicity. > TCDD induces mRNA expression of both CYP1Cs in the mesencephalic vein. > Knockdown

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.

Fanconi anemia: causes and consequences of genetic instability.

PubMed

Kalb, R; Neveling, K; Nanda, I; Schindler, D; Hoehn, H

2006-01-01

Fanconi anemia (FA) is a rare recessive disease that reflects the cellular and phenotypic consequences of genetic instability: growth retardation, congenital malformations, bone marrow failure, high risk of neoplasia, and premature aging. At the cellular level, manifestations of genetic instability include chromosomal breakage, cell cycle disturbance, and increased somatic mutation rates. FA cells are exquisitely sensitive towards oxygen and alkylating drugs such as mitomycin C or diepoxybutane, pointing to a function of FA genes in the defense against reactive oxygen species and other DNA damaging agents. FA is caused by biallelic mutations in at least 12 different genes which appear to function in the maintenance of genomic stability. Eight of the FA proteins form a nuclear core complex with a catalytic function involving ubiquitination of the central FANCD2 protein. The posttranslational modification of FANCD2 promotes its accumulation in nuclear foci, together with known DNA maintenance proteins such as BRCA1, BRCA2, and the RAD51 recombinase. Biallelic mutations in BRCA2 cause a severe FA-like phenotype, as do biallelic mutations in FANCD2. In fact, only leaky or hypomorphic mutations in this central group of FA genes appear to be compatible with life birth and survival. The newly discovered FANCJ (= BRIP1) and FANCM (= Hef ) genes correspond to known DNA-maintenance genes (helicase resp. helicase-associated endonuclease for fork-structured DNA). These genes provide the most convincing evidence to date of a direct involvement of FA genes in DNA repair functions associated with the resolution of DNA crosslinks and stalled replication forks. Even though genetic instability caused by mutational inactivation of the FANC genes has detrimental effects for the majority of FA patients, around 20% of patients appear to benefit from genetic instability since genetic instability also increases the chance of somatic reversion of their constitutional mutations. Intragenic

A mutation in the gamma actin 1 (ACTG1) gene causes autosomal dominant hearing loss (DFNA20/26)

PubMed Central

van Wijk, E; Krieger, E; Kemperman, M; De Leenheer, E M R; Huygen, P; Cremers, C; Cremers, F; Kremer, H

2003-01-01

Linkage analysis in a multigenerational family with autosomal dominant hearing loss yielded a chromosomal localisation of the underlying genetic defect in the DFNA20/26 locus at 17q25-qter. The 6-cM critical region harboured the γ-1-actin (ACTG1) gene, which was considered an attractive candidate gene because actins are important structural elements of the inner ear hair cells. In this study, a Thr278Ile mutation was identified in helix 9 of the modelled protein structure. The alteration of residue Thr278 is predicted to have a small but significant effect on the γ 1 actin structure owing to its close proximity to a methionine residue at position 313 in helix 11. Met313 has no space in the structure to move away. Moreover, the Thr278 residue is highly conserved throughout eukaryotic evolution. Using a known actin structure the mutation could be predicted to impair actin polymerisation. These findings strongly suggest that the Thr278Ile mutation in ACTG1 represents the first disease causing germline mutation in a cytoplasmic actin isoform. PMID:14684684

The effects of passive leg press training on jumping performance, speed, and muscle power.

PubMed

Liu, Chiang; Chen, Chuan-Shou; Ho, Wei-Hua; Füle, Róbert János; Chung, Pao-Hung; Shiang, Tzyy-Yuang

2013-06-01

Passive leg press (PLP) training was developed based on the concepts of the stretch-shortening cycle (SSC) and the benefits of high muscle contraction velocity. Passive leg press training enables lower limb muscle groups to apply a maximum downward force against a platform moved up and down at high frequency by an electric motor. Thus, these muscle groups accomplished both concentric and eccentric isokinetic contractions in a passive, rapid, and repetitive manner. This study investigates the effects of 10 weeks of PLP training at high and low movement frequencies have on jumping performance, speed, and muscle power. The authors selected 30 college students who had not performed systematic resistance training in the previous 6 months, including traditional resistance training at a squat frequency of 0.5 Hz, PLP training at a low frequency of 0.5 Hz, and PLP training at a high frequency of 2.5 Hz, and randomly divided them into 3 groups (n = 10). The participants' vertical jump, drop jump, 30-m sprint performance, explosive force, and SSC efficiency were tested under the same experimental procedures at pre- and post-training. Results reveal that high-frequency PLP training significantly increased participants' vertical jump, drop jump, 30-m sprint performance, instantaneous force, peak power, and SSC efficiency (p < 0.05). Additionally, their change rate abilities were substantially superior to those of the traditional resistance training (p < 0.05). The low-frequency PLP training significantly increased participants' vertical jump, 30-m sprint performance, instantaneous force, and peak power (p < 0.05). However, traditional resistance training only increased participants' 30-m sprint performance and peak power (p < 0.05). The findings suggest that jump performance, speed, and muscle power significantly improved after 10 weeks of PLP training at high movement frequency. A PLP training machine powered by an electrical motor enables muscles of the lower extremities to

Novel Compound Heterozygous CLCNKB Gene Mutations (c.1755A>G/ c.848_850delTCT) Cause Classic Bartter Syndrome.

PubMed

Wang, Chunli; Chen, Ying; Zheng, Bixia; Zhu, Mengshu; Fan, Jia; Wang, Juejin; Jia, Zhanjun; Huang, Songming; Zhang, Aihua

2018-02-14

Inactivated variants in CLCNKB gene encoding the basolateral chloride channel ClC-Kb cause classic Bartter syndrome characterized by hypokalemic metabolic alkalosis and hyperreninemic hyperaldosteronism. Here we identified two cBS siblings presenting hypokalemia in a Chinese family due to novel compound heterozygous CLCNKB mutations (c.848_850delTCT/c.1755A>G). Compound heterozygosity was confirmed by amplifying and sequencing the patient's genomic DNA. The synonymous mutation c.1755A>G (Thr585Thr) was located at +2bp from the 5' splice donor site in exon 15, further transcript analysis demonstrated that this single nucleotide mutation causes exclusion of exon 15 in the cDNA from the proband and his mother. Furthermore, we investigated the expression and protein trafficking change of c.848_850delTCT (TCT) and exon 15 deletion（E15）mutation in vitro. The E15 mutation markedly decreased the expression of ClC-Kb and resulted in a low-molecular-weight band (~55kD) trapping in the endoplasmic reticulum, while the TCT mutant only decreased the total and plasma membrane ClC-Kb protein expression but did not affect the subcellular localization. Finally, we studied the physiological functions of mutations by using whole-cell patch clamp and found that E15 or TCT mutation decreased the current of ClC-Kb/barttin channel. These results suggested that the compound defective mutations of CLCNKB gene are the molecular mechanism of the two cBS siblings.

Mutations in the Caenorhabditis elegans orthologs of human genes required for mitochondrial tRNA modification cause similar electron transport chain defects but different nuclear responses.

PubMed

Navarro-González, Carmen; Moukadiri, Ismaïl; Villarroya, Magda; López-Pascual, Ernesto; Tuck, Simon; Armengod, M-Eugenia

2017-07-01

Several oxidative phosphorylation (OXPHOS) diseases are caused by defects in the post-transcriptional modification of mitochondrial tRNAs (mt-tRNAs). Mutations in MTO1 or GTPBP3 impair the modification of the wobble uridine at position 5 of the pyrimidine ring and cause heart failure. Mutations in TRMU affect modification at position 2 and cause liver disease. Presently, the molecular basis of the diseases and why mutations in the different genes lead to such different clinical symptoms is poorly understood. Here we use Caenorhabditis elegans as a model organism to investigate how defects in the TRMU, GTPBP3 and MTO1 orthologues (designated as mttu-1, mtcu-1, and mtcu-2, respectively) exert their effects. We found that whereas the inactivation of each C. elegans gene is associated with a mild OXPHOS dysfunction, mutations in mtcu-1 or mtcu-2 cause changes in the expression of metabolic and mitochondrial stress response genes that are quite different from those caused by mttu-1 mutations. Our data suggest that retrograde signaling promotes defect-specific metabolic reprogramming, which is able to rescue the OXPHOS dysfunction in the single mutants by stimulating the oxidative tricarboxylic acid cycle flux through complex II. This adaptive response, however, appears to be associated with a biological cost since the single mutant worms exhibit thermosensitivity and decreased fertility and, in the case of mttu-1, longer reproductive cycle. Notably, mttu-1 worms also exhibit increased lifespan. We further show that mtcu-1; mttu-1 and mtcu-2; mttu-1 double mutants display severe growth defects and sterility. The animal models presented here support the idea that the pathological states in humans may initially develop not as a direct consequence of a bioenergetic defect, but from the cell's maladaptive response to the hypomodification status of mt-tRNAs. Our work highlights the important association of the defect-specific metabolic rewiring with the pathological phenotype

Novel large deletion in AVPR2 gene causing copy number variation in a patient with X-linked nephrogenic diabetes insipidus.

PubMed

Cho, Sun Young; Law, Chun Yiu; Ng, Kwok Leung; Lam, Ching Wan

2016-04-01

The diagnosis of cranial and nephrogenic diabetes insipidus (DI) can be clinically challenging. The application of molecular genetic analysis can help in resolving diagnostic difficulties. A 3 month-old boy presented with recurrent polyuria was admitted to Intensive Care Unit and was treated as DI. The patient also had a strong family history of polyuria affecting his maternal uncles. Molecular genetic analysis using Single Nucleotide Polymorphism (SNP) array detected a large deletion located at Xq28 region and the breakpoint was identified using PCR and Sanger sequencing. An 11,535 bp novel deletion affecting the entire APVR2 gene and the last intron and exon of the ARHGAP4 gene was confirmed. This large deletion is likely due to the 7-bp microhomology sequence at the junctions of both 5' and 3' breakpoints. No disease-causing mutation was identified for AQP2. We report a novel deletion in a Chinese patient with congenital nephrogenic DI. We suggested that patients with suspected congenital DI should undergo genetic analysis of AVPR2 and AQP2 genes. A definitive diagnosis can benefit patient by treatment of hydrochlorothiazide and amiloride and avoiding unnecessary investigations. Copyright © 2016 Elsevier B.V. All rights reserved.

Follow-up Findings in a Turkish Girl with Pseudohypoparathyroidism Type Ia Caused by a Novel Heterozygous Mutation in the GNAS Gene.

PubMed

Şahin, Sezgin; Hiort, Olaf; Thiele, Susanne; Evliyaoğlu, Olcay; Tüysüz, Beyhan

2017-03-01

Pseudohypoparathyroidism type Ia (PHP-Ia) is characterized by multihormone resistance and an Albright hereditary osteodystrophy (AHO) phenotype. It is caused by heterozygous mutations in GNAS gene. Clinical and biochemical findings of a female PHP-Ia patient were evaluated from age of diagnosis (6.5 years) to 14.5 years of age. The patient had short stature, brachydactyly, and subcutaneous heterotopic ossifications. Serum calcium and phosphorus levels were normal, but parathyroid hormone levels were high. Based on the typical clinical findings of AHO phenotype and biochemical findings, she was diagnosed as having PHP-Ia. A novel heterozygous mutation (c.128T>C) was found in the GNAS gene. Follow-up examinations revealed resistance to thyroid-stimulating hormone and a bioinactive growth hormone. Clinicians should take into consideration PHP-Ia in patients referred with short stature, and patients with an AHO phenotype must be further evaluated for hormone resistance, GNAS gene mutation, Gsα activity. To our knowledge, this is the first case report describing bioinactive growth hormone in PHP-Ia.

Mutation update of transcription factor genes FOXE3, HSF4, MAF, and PITX3 causing cataracts and other developmental ocular defects.

PubMed

Anand, Deepti; Agrawal, Smriti A; Slavotinek, Anne; Lachke, Salil A

2018-04-01

Mutations in the transcription factor genes FOXE3, HSF4, MAF, and PITX3 cause congenital lens defects including cataracts that may be accompanied by defects in other components of the eye or in nonocular tissues. We comprehensively describe here all the variants in FOXE3, HSF4, MAF, and PITX3 genes linked to human developmental defects. A total of 52 variants for FOXE3, 18 variants for HSF4, 20 variants for MAF, and 19 variants for PITX3 identified so far in isolated cases or within families are documented. This effort reveals FOXE3, HSF4, MAF, and PITX3 to have 33, 16, 18, and 7 unique causal mutations, respectively. Loss-of-function mutant animals for these genes have served to model the pathobiology of the associated human defects, and we discuss the currently known molecular function of these genes, particularly with emphasis on their role in ocular development. Finally, we make the detailed FOXE3, HSF4, MAF, and PITX3 variant information available in the Leiden Online Variation Database (LOVD) platform at https://www.LOVD.nl/FOXE3, https://www.LOVD.nl/HSF4, https://www.LOVD.nl/MAF, and https://www.LOVD.nl/PITX3. Thus, this article informs on key variants in transcription factor genes linked to cataract, aphakia, corneal opacity, glaucoma, microcornea, microphthalmia, anterior segment mesenchymal dysgenesis, and Ayme-Gripp syndrome, and facilitates their access through Web-based databases. © 2018 Wiley Periodicals, Inc.

A large homozygous deletion in the SAMHD1 gene causes atypical Aicardi–Goutiéres syndrome associated with mtDNA deletions

PubMed Central

Leshinsky-Silver, Esther; Malinger, Gustavo; Ben-Sira, Liat; Kidron, Dvora; Cohen, Sarit; Inbar, Shani; Bezaleli, Tali; Levine, Arie; Vinkler, Chana; Lev, Dorit; Lerman-Sagie, Tally

2011-01-01

Aicardi–Goutiéres syndrome (AGS) is a genetic neurodegenerative disorder with clinical symptoms mimicking a congenital viral infection. Five causative genes have been described: three prime repair exonuclease1 (TREX1), ribonucleases H2A, B and C, and most recently SAM domain and HD domain 1 (SAMHD1). We performed a detailed clinical and molecular characterization of a family with autosomal recessive neurodegenerative disorder showing white matter destruction and calcifications, presenting in utero and associated with multiple mtDNA deletions. A muscle biopsy was normal and did not show any evidence of respiratory chain dysfunction. Southern blot analysis of tissue from a living child and affected fetuses demonstrated multiple mtDNA deletions. Molecular analysis of genes involved in mtDNA synthesis and maintenance (POLGα, POLGβ, Twinkle, ANT1, TK2, SUCLA1 and DGOUK) revealed normal sequences. Sequencing of TREX1 and ribonucleases H2A, B and C failed to reveal any mutations. Whole-genome homozygosity mapping revealed a candidate region containing the SAMHD1 gene. Sequencing of the gene in the affected child and two affected fetuses revealed a large deletion (9 kb), spanning the promoter, exon1 and intron 1. The parents were found to be heterozygous for this deletion. The identification of a homozygous large deletion in the SAMHD1 gene causing atypical AGS with multiple mtDNA deletions may add information regarding the involvement of mitochondria in self-activation of innate immunity by cell intrinsic components. PMID:21102625

ABCD syndrome is caused by a homozygous mutation in the EDNRB gene.

PubMed

Verheij, Joke B G M; Kunze, Jürgen; Osinga, Jan; van Essen, Anthonie J; Hofstra, Robert M W

2002-03-15

ABCD syndrome is an autosomal recessive syndrome characterized by albinism, black lock, cell migration disorder of the neurocytes of the gut (Hirschsprung disease [HSCR]), and deafness. This phenotype clearly overlaps with the features of the Shah-Waardenburg syndrome, comprising sensorineural deafness; hypopigmentation of skin, hair, and irides; and HSCR. Therefore, we screened DNA of the index patient of the ABCD syndrome family for mutations in the endothelin B receptor (EDNRB) gene, a gene known to be involved in Shah-Waardenburg syndrome. A homozygous nonsense mutation in exon 3 (R201X) of the EDNRB gene was found. We therefore suggest that ABCD syndrome is not a separate entity, but an expression of Shah-Waardenburg syndrome.

Novel mutations in the TULP1 gene causing autosomal recessive retinitis pigmentosa.

PubMed

Paloma, E; Hjelmqvist, L; Bayés, M; García-Sandoval, B; Ayuso, C; Balcells, S; Gonzàlez-Duarte, R

2000-03-01

To assess the contribution of TULP1 to autosomal recessive retinitis pigmentosa (arRP). Fifteen exons of the gene were screened by single-strand conformation polymorphism analysis of 7 (of 49) arRP pedigrees showing cosegregation with TULP1 locus markers. In one of the seven families two allelic mutations, IVS4-2delAGA and c.937delC, were found in exons 5 and 10, respectively. Two novel mutations in TULP1 were found to be associated with arRP. That they both compromise the gene product supports their pathogenicity. This gene was present in no more than 2% of a panel of 49 Spanish families affected by arRP.

Sanfilippo syndrome: causes, consequences, and treatments

PubMed Central

Fedele, Anthony O

2015-01-01

Sanfilippo syndrome, or mucopolysaccharidosis (MPS) type III, refers to one of five autosomal recessive, neurodegenerative lysosomal storage disorders (MPS IIIA to MPS IIIE) whose symptoms are caused by the deficiency of enzymes involved exclusively in heparan sulfate degradation. The primary characteristic of MPS III is the degeneration of the central nervous system, resulting in mental retardation and hyperactivity, typically commencing during childhood. The significance of the order of events leading from heparan sulfate accumulation through to downstream changes in the levels of biomolecules within the cell and ultimately the (predominantly neuropathological) clinical symptoms is not well understood. The genes whose deficiencies cause the MPS III subtypes have been identified, and their gene products, as well as a selection of disease-causing mutations, have been characterized to varying degrees with respect to both frequency and direct biochemical consequences. A number of genetic and biochemical diagnostic methods have been developed and adopted by diagnostic laboratories. However, there is no effective therapy available for any form of MPS III, with treatment currently limited to clinical management of neurological symptoms. The availability of animal models for all forms of MPS III, whether spontaneous or generated via gene targeting, has contributed to improved understanding of the MPS III subtypes, and has provided and will deliver invaluable tools to appraise emerging therapies. Indeed, clinical trials to evaluate intrathecally-delivered enzyme replacement therapy in MPS IIIA patients, and gene therapy for MPS IIIA and MPS IIIB patients are planned or underway. PMID:26648750

A mutated dph3 gene causes sensitivity of Schizosaccharomyces pombe cells to cytotoxic agents.

PubMed

Villahermosa, Desirée; Knapp, Karen; Fleck, Oliver

2017-12-01

Dph3 is involved in diphthamide modification of the eukaryotic translation elongation factor eEF2 and in Elongator-mediated modifications of tRNAs, where a 5-methoxycarbonyl-methyl moiety is added to wobble uridines. Lack of such modifications affects protein synthesis due to inaccurate translation of mRNAs at ribosomes. We have discovered that integration of markers at the msh3 locus of Schizosaccharomyces pombe impaired the function of the nearby located dph3 gene. Such integrations rendered cells sensitive to the cytotoxic drugs hydroxyurea and methyl methanesulfonate. We constructed dph3 and msh3 strains with mutated ATG start codons (ATGmut), which allowed investigating drug sensitivity without potential interference by marker insertions. The dph3-ATGmut and a dph3::loxP-ura4-loxM gene disruption strain, but not msh3-ATGmut, turned out to be sensitive to hydroxyurea and methyl methanesulfonate, likewise the strains with cassettes integrated at the msh3 locus. The fungicide sordarin, which inhibits diphthamide modified eEF2 of Saccharomyces cerevisiae, barely affected survival of wild type and msh3Δ S. pombe cells, while the dph3Δ mutant was sensitive. The msh3-ATG mutation, but not dph3Δ or the dph3-ATG mutation caused a defect in mating-type switching, indicating that the ura4 marker at the dph3 locus did not interfere with Msh3 function. We conclude that Dph3 is required for cellular resistance to the fungicide sordarin and to the cytotoxic drugs hydroxyurea and methyl methanesulfonate. This is likely mediated by efficient translation of proteins in response to DNA damage and replication stress.

Diabetes Causing Gene, Kruppel-Like Factor 11, Modulates the Antinociceptive Response of Chronic Ethanol Intake

PubMed Central

Ou, Xiao-Ming; Udemgba, Chinelo; Wang, Niping; Dai, Xiaoli; Lomberk, Gwen; Seo, Seungmae; Urrutia, Raul; Wang, Junming; Duncan, Jeremy; Harris, Sharonda; Fairbanks, Carolyn A.; Zhang, Xiao

2017-01-01

Background Alcohol (ethanol) is an antinociceptive agent, working in part, by reducing sensitivity to painful stimuli. The transcription factor, Kruppel-like factor 11 (KLF11), a human diabetes-causing gene that also regulates the neurotransmitter-metabolic enzymes monoamine oxidase (MAOs), has recently been identified as an ethanol-inducible gene. However, its role in antinociception remains unknown. Consequently, we investigated the function of KLF11 in chronic ethanol-induced antinociception using a genetically engineered knockout mouse model. Methods Wild-type (Klf11+/+) and KLF11 knockout (Klf11−/−) mice were fed a liquid diet containing ethanol for 28 days with increasing amounts of ethanol from 0% up to a final concentration of 6.4%, representing a final diet containing 36% of calories primarily from ethanol. Control mice from both genotypes were fed liquid diet without ethanol for 28 days. The ethanol-induced antinociceptive effect was determined using the tail-flick test before and after ethanol exposure (on day 29). In addition, the enzyme activity and mRNA levels of MAO A and MAO B were measured by Real-time RT-PCR and enzyme assays, respectively. Results Ethanol produced an antinociceptive response to thermal pain in Klf11+/+ mice, as expected. In contrast, deletion of KLF11 in the Klf11−/− mice abolished the ethanol-induced antinociceptive effect. The mRNA and protein levels of KLF11were significantly increased in the brain prefrontal cortex of Klf11+/+ mice exposed to ethanol compared to control Klf11+/+ mice. Furthermore, MAO enzyme activities were affected differently in Klf11 wild-type versus Klf11 knockout mice exposed to chronic ethanol. Chronic ethanol intake significantly increased MAO-B activity in Klf1+/+ mice. Conclusions The data show KLF11 modulation of ethanol-induced antinociception. The KLF11-targeted MAO B enzyme, may contribute more significantly to ethanol-induced antinociception. Thus, this study revealed a new role for the

The identification of a gene (Cwp1), silenced during Solanum evolution, which causes cuticle microfissuring and dehydration when expressed in tomato fruit.

PubMed

Hovav, Ran; Chehanovsky, Noam; Moy, Michal; Jetter, Reinhard; Schaffer, Arthur A

2007-11-01

One of the most intriguing phenomena of fleshy fruit is the ability to maintain high water content at maturity, even following harvest. This is accomplished by a fruit cuticle that is highly impermeable to water diffusion. In this paper, we report on a novel genotype of tomato, developed via introgression from the wild species Solanum habrochaites, which is characterized by microfissuring of the fruit cuticle and dehydration of the mature fruit. The microfissure/dehydration phenotype is inherited as a single gene, termed Cwp1 (cuticular water permeability). The gene was fine mapped, and its identity was determined by map-based cloning and differential expression analysis in near-isogenic lines. Causality of the Cwp1 gene was shown by the heterologous transgenic expression of the gene in the cultivated tomato, which caused a microfissured fruit cuticle leading to dehydrated fruit. Cwp1 encodes for a protein of unidentified function in the DUF833 domain family. The gene is expressed in the fruit epidermis of the dehydrating genotype harbouring the wild-species introgression, but not in the cultivated tomato. It is expressed only in the primitive green-fruited wild tomato species, but is not expressed in the cultivated Solanum lycopersicum and the closely related Solanum cheesmaniae and Solanum pimpinellifolium, indicating a pre-adaptive role for Cwp1 silencing in the evolution and domestication of the cultivated tomato.

Direct visualization of critical hydrogen atoms in a pyridoxal 5'-phosphate enzyme.

PubMed

Dajnowicz, Steven; Johnston, Ryne C; Parks, Jerry M; Blakeley, Matthew P; Keen, David A; Weiss, Kevin L; Gerlits, Oksana; Kovalevsky, Andrey; Mueser, Timothy C

2017-10-16

Enzymes dependent on pyridoxal 5'-phosphate (PLP, the active form of vitamin B 6 ) perform a myriad of diverse chemical transformations. They promote various reactions by modulating the electronic states of PLP through weak interactions in the active site. Neutron crystallography has the unique ability of visualizing the nuclear positions of hydrogen atoms in macromolecules. Here we present a room-temperature neutron structure of a homodimeric PLP-dependent enzyme, aspartate aminotransferase, which was reacted in situ with α-methylaspartate. In one monomer, the PLP remained as an internal aldimine with a deprotonated Schiff base. In the second monomer, the external aldimine formed with the substrate analog. We observe a deuterium equidistant between the Schiff base and the C-terminal carboxylate of the substrate, a position indicative of a low-barrier hydrogen bond. Quantum chemical calculations and a low-pH room-temperature X-ray structure provide insight into the physical phenomena that control the electronic modulation in aspartate aminotransferase.Pyridoxal 5'-phosphate (PLP) is a ubiquitous co factor for diverse enzymes, among them aspartate aminotransferase. Here the authors use neutron crystallography, which allows the visualization of the positions of hydrogen atoms, and computation to characterize the catalytic mechanism of the enzyme.

Isolation, purification, and structural features of a polysaccharide from Phellinus linteus and its hypoglycemic effect in alloxan-induced diabetic mice.

PubMed

Zhao, Chao; Liao, Zunsheng; Wu, Xiaoqi; Liu, Yanling; Liu, Xiaoyan; Lin, Zhanxi; Huang, Yifan; Liu, Bin

2014-05-01

Phellinus linteus is a medicinal mushroom that has been used in Oriental countries for centuries for its antitumor, antioxidant, immunomodulatory, and biological activity on hyperglycemia. A water-soluble crude polysaccharide was extracted using hot water from P. linteus mycelia grown under submerged culture. An orthogonal experiment was used to optimize the extraction conditions of P. linteus mycelia polysaccharides (PLP). The crude polysaccharide was purified using DEAE Sephadex A-50 and Sephadex G-200 chromatography. Fourier transform infrared (FT-IR) spectroscopy and nuclear magnetic resonance ((1) H NMR) spectroscopy were used to investigate the structure of the purified P. linteus polysaccharide (PLP-I), revealing that it was mainly a branched-type glycan with both α- and β-linkages and a pyranoid sugar ring conformation. PLP orally administered at 100 mg/kg body weight/d could significantly reduce the blood glucose level by 35.60% in alloxan-induced diabetic mice. The results of an oral glucose tolerance test (OGTT) revealed that PLP had an effect on glucose disposal after 28 d of treatment. The result revealed that PLP from a submerged culture of P. linteus mycelia possessed potent hypoglycemic properties. The polysaccharide may be useful as a functional food additive and a hypoglycemic agent. © 2014 Institute of Food Technologists®

A prospective evaluation of serum kynurenine metabolites and risk of pancreatic cancer.

PubMed

Huang, Joyce Y; Butler, Lesley M; Midttun, Øivind; Ulvik, Arve; Wang, Renwei; Jin, Aizhen; Gao, Yu-Tang; Ueland, Per M; Koh, Woon-Puay; Yuan, Jian-Min

2018-01-01

Serum pyridoxal 5'-phosphate (PLP), the active form of vitamin B6, is associated with reduced risk of pancreatic cancer. Data on functional measures of vitamin B6 status and risk of pancreatic cancer is lacking. A nested case-control study involving 187 incident cases of pancreatic cancer and 362 individually matched controls were conducted within two prospective cohorts to evaluate the associations between kynurenine metabolites in pre-diagnostic serum samples and risk of pancreatic cancer. Higher serum concentrations of 3-hydroxyanthranilic acid (HAA) and the HAA:3-hydroxykynurenine (HK) ratio (a measure for in vivo functional status of PLP) were significantly associated with reduced risk of pancreatic cancer. Compared with the lowest tertile, odds ratios (95% confidence intervals) of pancreatic cancer for the highest tertile was 0.62 (0.39, 1.01) for HAA, and 0.59 (0.35-0.98) for the HAA:HK ratio, after adjustment for potential confounders and serum PLP (both Ps for trend<0.05). The kynurenine:tryptophan ratio or neopterin was not significantly associated with pancreatic cancer risk. The inverse association between HAA or the HAA:HK ratio and risk of pancreatic cancer supports the notion that functional status of PLP may be a more important measure than circulating PLP alone for the development of pancreatic cancer.

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

Lentiginous phenotypes caused by diverse pathogenic genes (SASH1 and PTPN11): clinical and molecular discrimination.

PubMed

Zhang, J; Cheng, R; Liang, J; Ni, C; Li, M; Yao, Z

2016-10-01

Pathogenic mutations in genes (SASH1 and PTPN11) can cause a rare genetic disorder associated with pigmentation defects and the well-known LEOPARD syndrome, respectively. Both conditions presented with lentiginous phenotypes. The aim of this study was to arrive at definite diagnoses of three Chinese boys with clinically suspected lentigines-related syndromes. ADAR1, ABCB6, SASH1 and PTPN11 were candidate genes for mutational screening. Sanger sequencing was performed to identify the mutations, whereas bioinformatic analysis was used to predict the pathogenicity of novel missense mutations. Two novel mutations c.1537A>C (p.Ser513Arg) and 1527_1530dupAAGT (p.Leu511Lysfs*21) in SASH1 and a common p.Thr468Met mutation in PTPN11 were detected in three pediatric patients with lentiginous phenotypes, respectively. Comparisons between clinical presentations showed that SASH1-related phenotypes can exhibit hyper- and hypopigmentation on the trunk and extremities, similar to dyschromatosis, while scattered café au-lait spots usually appeared in PTPN11-related LEOPARD syndrome. Furthermore, the similarity in the clinical presentations of Peutz-Jeghers syndrome, Laugier-Hunziker syndrome, xeroderma pigmentosum, neurofibromatosis type I, suggesting that these conditions should be added into the differential diagnoses of lentiginous phenotypes. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Selective ablation of Ppp1cc gene in testicular germ cells causes oligo-teratozoospermia and infertility in mice.

PubMed

Sinha, Nilam; Puri, Pawan; Nairn, Angus C; Vijayaraghavan, Srinivasan

2013-11-01

The four isoforms of serine/threonine phosphoprotein phosphatase 1 (PP1), derived from three genes, are among the most conserved proteins known. The Ppp1cc gene encodes two alternatively spliced variants, PP1 gamma1 (PPP1CC1) and PP1 gamma2 (PPP1CC2). Global deletion of the Ppp1cc gene, which causes loss of both isoforms, results in male infertility due to impaired spermatogenesis. This phenotype was assumed to be due to the loss of PPP1CC2, which is abundant in testis. While PPP1CC2 is predominant, other PP1 isoforms are also expressed in testis. Given the significant homology between the four PP1 isoforms, the lack of compensation by the other PP1 isoforms for loss of one, only in testis, is surprising. Here we document, for the first time, expression patterns of the PP1 isoforms in postnatal developing and adult mouse testis. The timing and sites of testis expression of PPP1CC1 and PPP1CC2 in testis are nonoverlapping. PPP1CC2 is the only one of the four PP1 isoforms not detected in sertoli cells and spermatogonia. Conversely, PPP1CC2 may be the only PP1 isoform expressed in postmeiotic germ cells. Deletion of the Ppp1cc gene in germ cells at the differentiated spermatogonia stage of development and beyond in Stra8 promoter-driven Cre transgenic mice results in oligo-terato-asthenozoospermia and male infertility, thus phenocopying global Ppp1cc null (-/-) mice. Taken together, these results confirm that spermatogenic defects observed in the global Ppp1cc knockout mice and in mice expressing low levels of PPP1CC2 in testis are due to compromised functions of PPP1CC2 in meiotic and postmeiotic germ cells.

Spermine oxidase maintains basal skeletal muscle gene expression and fiber size and is strongly repressed by conditions that cause skeletal muscle atrophy

PubMed Central

Bongers, Kale S.; Fox, Daniel K.; Kunkel, Steven D.; Stebounova, Larissa V.; Murry, Daryl J.; Pufall, Miles A.; Ebert, Scott M.; Dyle, Michael C.; Bullard, Steven A.; Dierdorff, Jason M.

2014-01-01

Skeletal muscle atrophy is a common and debilitating condition that remains poorly understood at the molecular level. To better understand the mechanisms of muscle atrophy, we used mouse models to search for a skeletal muscle protein that helps to maintain muscle mass and is specifically lost during muscle atrophy. We discovered that diverse causes of muscle atrophy (limb immobilization, fasting, muscle denervation, and aging) strongly reduced expression of the enzyme spermine oxidase. Importantly, a reduction in spermine oxidase was sufficient to induce muscle fiber atrophy. Conversely, forced expression of spermine oxidase increased muscle fiber size in multiple models of muscle atrophy (immobilization, fasting, and denervation). Interestingly, the reduction of spermine oxidase during muscle atrophy was mediated by p21, a protein that is highly induced during muscle atrophy and actively promotes muscle atrophy. In addition, we found that spermine oxidase decreased skeletal muscle mRNAs that promote muscle atrophy (e.g., myogenin) and increased mRNAs that help to maintain muscle mass (e.g., mitofusin-2). Thus, in healthy skeletal muscle, a relatively low level of p21 permits expression of spermine oxidase, which helps to maintain basal muscle gene expression and fiber size; conversely, during conditions that cause muscle atrophy, p21 expression rises, leading to reduced spermine oxidase expression, disruption of basal muscle gene expression, and muscle fiber atrophy. Collectively, these results identify spermine oxidase as an important positive regulator of muscle gene expression and fiber size, and elucidate p21-mediated repression of spermine oxidase as a key step in the pathogenesis of skeletal muscle atrophy. PMID:25406264

Tyramine and phenylethylamine biosynthesis by food bacteria.

PubMed

Marcobal, Angela; De las Rivas, Blanca; Landete, José María; Tabera, Laura; Muñoz, Rosario

2012-01-01

Tyramine poisoning is caused by the ingestion of food containing high levels of tyramine, a biogenic amine. Any foods containing free tyrosine are subject to tyramine formation if poor sanitation and low quality foods are used or if the food is subject to temperature abuse or extended storage time. Tyramine is generated by decarboxylation of the tyrosine through tyrosine decarboxylase (TDC) enzymes derived from the bacteria present in the food. Bacterial TDC have been only unequivocally identified and characterized in Gram-positive bacteria, especially in lactic acid bacteria. Pyridoxal phosphate (PLP)-dependent TDC encoding genes (tyrDC) appeared flanked by a similar genetic organization in several species of lactic acid bacteria, suggesting a common origin by a single mobile genetic element. Bacterial TDC are also able to decarboxylate phenylalanine to produce phenylethylamine (PEA), another biogenic amine. The molecular knowledge of the genes involved in tyramine production has led to the development of molecular methods for the detection of bacteria able to produce tyramine and PEA. These rapid and simple methods could be used for the analysis of the ability to form tyramine by bacteria in order to evaluate the potential risk of tyramine biosynthesis in food products.

A matter of phylogenetic scale: Distinguishing incomplete lineage sorting from lateral gene transfer as the cause of gene tree discord in recent versus deep diversification histories.

PubMed

Knowles, L Lacey; Huang, Huateng; Sukumaran, Jeet; Smith, Stephen A

2018-03-01

Discordant gene trees are commonly encountered when sequences from thousands of loci are applied to estimate phylogenetic relationships. Several processes contribute to this discord. Yet, we have no methods that jointly model different sources of conflict when estimating phylogenies. An alternative to analyzing entire genomes or all the sequenced loci is to identify a subset of loci for phylogenetic analysis. If we can identify data partitions that are most likely to reflect descent from a common ancestor (i.e., discordant loci that indeed reflect incomplete lineage sorting [ILS], as opposed to some other process, such as lateral gene transfer [LGT]), we can analyze this subset using powerful coalescent-based species-tree approaches. Test data sets were simulated where discord among loci could arise from ILS and LGT. Data sets where analyzed using the newly developed program CLASSIPHY (Huang et al., ) to assess whether our ability to distinguish the cause of discord among loci varied when ILS and LGT occurred in the recent versus deep past and whether the accuracy of these inferences were affected by the mutational process. We show that accuracy of probabilistic classification of individual loci by the cause of discord differed when ILS and LGT events occurred more recently compared with the distant past and that the signal-to-noise ratio arising from the mutational process contributes to difficulties in inferring LGT data partitions. We discuss our findings in terms of the promise and limitations of identifying subsets of loci for species-tree inference that will not violate the underlying coalescent model (i.e., data partitions in which ILS, and not LGT, contributes to discord). We also discuss the empirical implications of our work given the many recalcitrant nodes in the tree of life (e.g., origins of angiosperms, amniotes, or Neoaves), and recent arguments for concatenating loci. © 2018 Botanical Society of America.

Evaluation of androgen receptor gene as a candidate gene in female androgenetic alopecia.

PubMed

el-Samahy, May H; Shaheen, Maha A; Saddik, Dina E B; Abdel-Fattah, Nermeen S A; el-Sawi, Mohammad A; Mahran, Manal Z; Shehab, Abeer A A

2009-06-01

Genetic polymorphisms of the androgen receptor (AR) gene have been studied in male androgenetic alopecia (AGA); however, little is known about gene polymorphism and female AGA. To evaluate the AR gene as a candidate gene for female AGA. Thirty premenopausal Egyptian female patients with AGA (mean age, 32.3 +/- 7 years) and 11 age- and sex-matched controls were included. All subjects underwent laboratory and pelvic ultrasound evaluation to exclude other precipitating cause(s) of hair loss. Scalp biopsy was taken and the AR gene was evaluated using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). According to Ludwig's classification, all patients had type II AGA. Statistical analysis showed no statistically significant difference in genotype (chi(2) = 5.513, P > or = 0.05) or allele frequency (chi(2) = 1.312, P > or = 0.05) between patients and controls. There was also no statistically significant difference between the genotype and allele frequency with disease duration. In contrast with male AGA, no association was found between type II AGA in Egyptian women and the AR gene. Therefore, the genetic study of this gene does not serve as a biomarker for the identification of women with a predisposition to AGA.

Mild myelin disruption elicits early alteration in behavior and proliferation in the subventricular zone.

PubMed

Gould, Elizabeth A; Busquet, Nicolas; Shepherd, Douglas; Dietz, Robert M; Herson, Paco S; Simoes de Souza, Fabio M; Li, Anan; George, Nicholas M; Restrepo, Diego; Macklin, Wendy B

2018-02-13

Myelin, the insulating sheath around axons, supports axon function. An important question is the impact of mild myelin disruption. In the absence of the myelin protein proteolipid protein (PLP1), myelin is generated but with age, axonal function/maintenance is disrupted. Axon disruption occurs in Plp1 -null mice as early as 2 months in cortical projection neurons. High-volume cellular quantification techniques revealed a region-specific increase in oligodendrocyte density in the olfactory bulb and rostral corpus callosum that increased during adulthood. A distinct proliferative response of progenitor cells was observed in the subventricular zone (SVZ), while the number and proliferation of parenchymal oligodendrocyte progenitor cells was unchanged. This SVZ proliferative response occurred prior to evidence of axonal disruption. Thus, a novel SVZ response contributes to the region-specific increase in oligodendrocytes in Plp1 -null mice. Young adult Plp1- null mice exhibited subtle but substantial behavioral alterations, indicative of an early impact of mild myelin disruption. © 2018, Gould et al.

Mild myelin disruption elicits early alteration in behavior and proliferation in the subventricular zone

PubMed Central

Gould, Elizabeth A; Busquet, Nicolas; Shepherd, Douglas; Dietz, Robert M; Herson, Paco S; Simoes de Souza, Fabio M; Li, Anan; George, Nicholas M

2018-01-01

Myelin, the insulating sheath around axons, supports axon function. An important question is the impact of mild myelin disruption. In the absence of the myelin protein proteolipid protein (PLP1), myelin is generated but with age, axonal function/maintenance is disrupted. Axon disruption occurs in Plp1-null mice as early as 2 months in cortical projection neurons. High-volume cellular quantification techniques revealed a region-specific increase in oligodendrocyte density in the olfactory bulb and rostral corpus callosum that increased during adulthood. A distinct proliferative response of progenitor cells was observed in the subventricular zone (SVZ), while the number and proliferation of parenchymal oligodendrocyte progenitor cells was unchanged. This SVZ proliferative response occurred prior to evidence of axonal disruption. Thus, a novel SVZ response contributes to the region-specific increase in oligodendrocytes in Plp1-null mice. Young adult Plp1-null mice exhibited subtle but substantial behavioral alterations, indicative of an early impact of mild myelin disruption. PMID:29436368

Structure of the Mitochondrial Aminolevulinic Acid Synthase, a Key Heme Biosynthetic Enzyme.

PubMed

Brown, Breann L; Kardon, Julia R; Sauer, Robert T; Baker, Tania A

2018-04-03

5-Aminolevulinic acid synthase (ALAS) catalyzes the first step in heme biosynthesis. We present the crystal structure of a eukaryotic ALAS from Saccharomyces cerevisiae. In this homodimeric structure, one ALAS subunit contains covalently bound cofactor, pyridoxal 5'-phosphate (PLP), whereas the second is PLP free. Comparison between the subunits reveals PLP-coupled reordering of the active site and of additional regions to achieve the active conformation of the enzyme. The eukaryotic C-terminal extension, a region altered in multiple human disease alleles, wraps around the dimer and contacts active-site-proximal residues. Mutational analysis demonstrates that this C-terminal region that engages the active site is important for ALAS activity. Our discovery of structural elements that change conformation upon PLP binding and of direct contact between the C-terminal extension and the active site thus provides a structural basis for investigation of disruptions in the first step of heme biosynthesis and resulting human disorders. Copyright © 2018 Elsevier Ltd. All rights reserved.

Direct visualization of critical hydrogen atoms in a pyridoxal 5'-phosphate enzyme

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

Dajnowicz, Steven; Johnston, Ryne C.; Parks, Jerry M.

Enzymes dependent on pyridoxal 5'-phosphate (PLP, the active form of vitamin B6) perform a myriad of diverse chemical transformations. They promote various reactions by modulating the electronic states of PLP through weak interactions in the active site. Neutron crystallography has the unique ability of visualizing the nuclear positions of hydrogen atoms in macromolecules. Here we present a room-temperature neutron structure of a homodimeric PLP-dependent enzyme, aspartate aminotransferase, which was reacted in situ with α-methylaspartate. In one monomer, the PLP remained as an internal aldimine with a deprotonated Schiff base. In the second monomer, the external aldimine formed with the substratemore » analog. We observe a deuterium equidistant between the Schiff base and the C-terminal carboxylate of the substrate, a position indicative of a low-barrier hydrogen bond. As a result, quantum chemical calculations and a low-pH room-temperature X-ray structure provide insight into the physical phenomena that control the electronic modulation in aspartate aminotransferase.« less

Direct visualization of critical hydrogen atoms in a pyridoxal 5'-phosphate enzyme

DOE PAGES

Dajnowicz, Steven; Johnston, Ryne C.; Parks, Jerry M.; ...

2017-10-16

Enzymes dependent on pyridoxal 5'-phosphate (PLP, the active form of vitamin B6) perform a myriad of diverse chemical transformations. They promote various reactions by modulating the electronic states of PLP through weak interactions in the active site. Neutron crystallography has the unique ability of visualizing the nuclear positions of hydrogen atoms in macromolecules. Here we present a room-temperature neutron structure of a homodimeric PLP-dependent enzyme, aspartate aminotransferase, which was reacted in situ with α-methylaspartate. In one monomer, the PLP remained as an internal aldimine with a deprotonated Schiff base. In the second monomer, the external aldimine formed with the substratemore » analog. We observe a deuterium equidistant between the Schiff base and the C-terminal carboxylate of the substrate, a position indicative of a low-barrier hydrogen bond. As a result, quantum chemical calculations and a low-pH room-temperature X-ray structure provide insight into the physical phenomena that control the electronic modulation in aspartate aminotransferase.« less

A novel deletion mutation in RS1 gene caused X-linked juvenile retinoschisis in a Chinese family.

PubMed

Huang, Y; Mei, L; Gui, B; Su, W; Liang, D; Wu, L; Pan, Q

2014-11-01

X-linked juvenile retinoschisis (XLRS), a leading cause of juvenile macular degeneration, is characterized by a spoke-wheel pattern in the macular region of the retina and splitting of the neurosensory retina. This study aimed to identify the underlying genetic defect in a Chinese family with XLRS. The proband underwent complete ophthalmic examinations, including fundus examination, fundus autofluorescence, and optical coherence tomography. DNA extracted from proband and his younger brother was screened for mutations in RS1 gene. The detected RS1 mutation was tested in all available family members and 200 healthy controls. Reduced visual acuity, spoke-wheel pattern at the fovea, and split retina were observed in the proband. A novel frameshift mutation c.206-207delTG in the RS1 gene, leading to a truncated protein (p.L69fs16X), was identified in the proband and his younger brother. This mutation was not found in any unaffected member or in the healthy controls. The mother of the proband was hemizygous for this mutant allele. We identified a novel causative mutation of RS1 in a Chinese family with XLRS. This finding expands the mutation spectrum of RS1 and provides evidence for a phenotype-genotype study in XLRS.

A novel deletion mutation in RS1 gene caused X-linked juvenile retinoschisis in a Chinese family

PubMed Central

Huang, Y; Mei, L; Gui, B; Su, W; Liang, D; Wu, L; Pan, Q

2014-01-01

Purpose X-linked juvenile retinoschisis (XLRS), a leading cause of juvenile macular degeneration, is characterized by a spoke-wheel pattern in the macular region of the retina and splitting of the neurosensory retina. This study aimed to identify the underlying genetic defect in a Chinese family with XLRS. Methods The proband underwent complete ophthalmic examinations, including fundus examination, fundus autofluorescence, and optical coherence tomography. DNA extracted from proband and his younger brother was screened for mutations in RS1 gene. The detected RS1 mutation was tested in all available family members and 200 healthy controls. Results Reduced visual acuity, spoke-wheel pattern at the fovea, and split retina were observed in the proband. A novel frameshift mutation c.206-207delTG in the RS1 gene, leading to a truncated protein (p.L69fs16X), was identified in the proband and his younger brother. This mutation was not found in any unaffected member or in the healthy controls. The mother of the proband was hemizygous for this mutant allele. Conclusions We identified a novel causative mutation of RS1 in a Chinese family with XLRS. This finding expands the mutation spectrum of RS1 and provides evidence for a phenotype–genotype study in XLRS. PMID:25168411

Vitamin B-6 and colorectal cancer risk: a prospective population-based study using 3 distinct plasma markers of vitamin B-6 status.

PubMed

Gylling, Björn; Myte, Robin; Schneede, Jörn; Hallmans, Göran; Häggström, Jenny; Johansson, Ingegerd; Ulvik, Arve; Ueland, Per M; Van Guelpen, Bethany; Palmqvist, Richard

2017-04-01

Background: Higher plasma concentrations of the vitamin B-6 marker pyridoxal 5'-phosphate (PLP) have been associated with reduced colorectal cancer (CRC) risk. Inflammatory processes, including vitamin B-6 catabolism, could explain such findings. Objective: We investigated 3 biomarkers of vitamin B-6 status in relation to CRC risk. Design: This was a prospective case-control study of 613 CRC cases and 1190 matched controls nested within the Northern Sweden Health and Disease Study ( n = 114,679). Participants were followed from 1985 to 2009, and the median follow-up from baseline to CRC diagnosis was 8.2 y. PLP, pyridoxal, pyridoxic acid (PA), 3-hydroxykynurenine, and xanthurenic acids (XAs) were measured in plasma with the use of liquid chromatography-tandem mass spectrometry. We calculated relative and absolute risks of CRC for PLP and the ratios 3-hydroxykynurenine:XA (HK:XA), an inverse marker of functional vitamin B-6 status, and PA:(PLP + pyridoxal) (PAr), a marker of inflammation and oxidative stress and an inverse marker of vitamin B-6 status. Results: Plasma PLP concentrations were associated with a reduced CRC risk for the third compared with the first quartile and for PLP sufficiency compared with deficiency [OR: 0.60 (95% CI: 0.44, 0.81) and OR: 0.55 (95% CI: 0.37, 0.81), respectively]. HK:XA and PAr were both associated with increased CRC risk [OR: 1.48 (95% CI: 1.08, 2.02) and OR: 1.50 (95% CI: 1.10, 2.04), respectively] for the fourth compared with the first quartile. For HK:XA and PAr, the findings were mainly observed in study participants with <10.5 y of follow-up between sampling and diagnosis. Conclusions: Vitamin B-6 deficiency as measured by plasma PLP is associated with a clear increase in CRC risk. Furthermore, our analyses of novel markers of functional vitamin B-6 status and vitamin B-6-associated oxidative stress and inflammation suggest a role in tumor progression rather than initiation. © 2017 American Society for Nutrition.

Activation of the hexosamine pathway causes oxidative stress and abnormal embryo gene expression: involvement in diabetic teratogenesis.

PubMed

Horal, Melissa; Zhang, Zhiquan; Stanton, Robert; Virkamäki, Antti; Loeken, Mary R

2004-08-01

Oxidative stress is critical to the teratogenic effects of diabetic pregnancy, yet the specific biochemical pathways responsible for oxidative stress have not been fully elucidated. The hexosamine pathway is activated in many tissues during diabetes and could contribute to oxidative stress by inhibiting the pentose shunt pathway, thereby diminishing production of the cellular antioxidant, reduced glutathione (GSH). To test the hypothesis that activation of the hexosamine pathway might contribute to the teratogenic effects of diabetic pregnancy, pregnant mice were injected with glucose, to induce hyperglycemia, or glucosamine, to directly activate the hexosamine pathway. Embryo tissue fragments were also cultured in physiological glucose, high glucose, or physiological glucose plus glucosamine, to test effects on oxidative stress and embryo gene expression. Glucosamine increased hexosamine synthesis and inhibited pentose shunt activity. There was a trend for transient hyperglycemia to have the same effects, but they did not reach statistical significance. However, both glucose and glucosamine significantly decreased GSH, and increased oxidative stress, as indicated by 2',7'-dichloro-dihydrofluorescein fluorescence. Glucose and glucosamine inhibited expression of Pax-3, a gene required for neural tube closure both in vivo and in vitro, and increased neural tube defects (NTDs) in vivo; these effects were prevented by GSH ethyl ester. High glucose and glucosamine inhibited Pax-3 expression by embryo culture, but culture in glutamine-free media to block the hexosamine pathway prevented the inhibition of Pax-3 expression by high glucose. Activation of the hexosamine pathway causes oxidative stress through depletion of GSH and consequent disruption of embryo gene expression. Activation of this pathway may contribute to diabetic teratogenesis.

Tsc2 gene inactivation causes a more severe epilepsy phenotype than Tsc1 inactivation in a mouse model of tuberous sclerosis complex.

PubMed

Zeng, Ling-Hui; Rensing, Nicholas R; Zhang, Bo; Gutmann, David H; Gambello, Michael J; Wong, Michael

2011-02-01

Tuberous Sclerosis Complex (TSC) is an autosomal dominant, multi-system disorder, typically involving severe neurological symptoms, such as epilepsy, cognitive deficits and autism. Two genes, TSC1 and TSC2, encoding the proteins hamartin and tuberin, respectively, have been identified as causing TSC. Although there is a substantial overlap in the clinical phenotype produced by TSC1 and TSC2 mutations, accumulating evidence indicates that TSC2 mutations cause more severe neurological manifestations than TSC1 mutations. In this study, the neurological phenotype of a novel mouse model involving conditional inactivation of the Tsc2 gene in glial-fibrillary acidic protein (GFAP)-positive cells (Tsc2(GFAP1)CKO mice) was characterized and compared with previously generated Tsc1(GFAP1)CKO mice. Similar to Tsc1(GFAP1)CKO mice, Tsc2(GFAP1)CKO mice exhibited epilepsy, premature death, progressive megencephaly, diffuse glial proliferation, dispersion of hippocampal pyramidal cells and decreased astrocyte glutamate transporter expression. However, Tsc2(GFAP1)CKO mice had an earlier onset and higher frequency of seizures, as well as significantly more severe histological abnormalities, compared with Tsc1(GFAP1)CKO mice. The differences between Tsc1(GFAP1)CKO and Tsc2(GFAP1)CKO mice were correlated with higher levels of mammalian target of rapamycin (mTOR) activation in Tsc2(GFAP1)CKO mice and were reversed by the mTOR inhibitor, rapamycin. These findings provide novel evidence in mouse models that Tsc2 mutations intrinsically cause a more severe neurological phenotype than Tsc1 mutations and suggest that the difference in phenotype may be related to the degree to which Tsc1 and Tsc2 inactivation causes abnormal mTOR activation.

Mutations in the PFN1 gene are not a common cause in patients with amyotrophic lateral sclerosis and frontotemporal lobar degeneration in France.

PubMed

Lattante, Serena; Le Ber, Isabelle; Camuzat, Agnès; Brice, Alexis; Kabashi, Edor

2013-06-01

Amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD) are 2 adult onset neurological disorders with overlapping symptoms and clinical characteristics. It is well established that they share a common pathologic and genetic background. Recently, mutations in profilin 1 gene (PFN1) have been identified in patients with familial ALS, suggesting a role for this gene in the pathogenesis of the disease. Based on this, we hypothesized that mutations in PFN1 might also contribute to FTLD disease. We studied a French cohort of 165 ALS/FTLD patients, without finding any variant. We conclude that mutations in PFN1 are not a common cause for ALS/FTLD in France. Copyright © 2013 Elsevier Inc. All rights reserved.

Identification of essential genes and synthetic lethal gene combinations in Escherichia coli K-12.

PubMed

Mori, Hirotada; Baba, Tomoya; Yokoyama, Katsushi; Takeuchi, Rikiya; Nomura, Wataru; Makishi, Kazuichi; Otsuka, Yuta; Dose, Hitomi; Wanner, Barry L

2015-01-01

Here we describe the systematic identification of single genes and gene pairs, whose knockout causes lethality in Escherichia coli K-12. During construction of precise single-gene knockout library of E. coli K-12, we identified 328 essential gene candidates for growth in complex (LB) medium. Upon establishment of the Keio single-gene deletion library, we undertook the development of the ASKA single-gene deletion library carrying a different antibiotic resistance. In addition, we developed tools for identification of synthetic lethal gene combinations by systematic construction of double-gene knockout mutants. We introduce these methods herein.

Two novel mutations in NOTCH3 gene causes cerebral autosomal dominant arteriopathy with subcritical infarct and leucoencephalopathy in two Chinese families.

PubMed

Zhu, Yuyou; Wang, Juan; Wu, Yuanbo; Wang, Guoping; Hu, Bai

2015-01-01

To investigate the genetic pathogenic causes of cerebral autosomal dominant arteriopathy with subcritical infarct and leucoencephalopathy (CADASIL) in two Chinese families, to provide the molecular basis for genetic counseling and antenatal diagnosis. The genetic mutation of gene NOTCH3 of propositus and family members was analyzed in these two CADASIL families by polymerase chain reaction and DNA sequencing technology directly. At the same time, the NOTCH3 gene mutation point of 100 healthy collators was detected, to explicit the pathogenic mutation by function prediction with Polyphen-2 and SIFT. Both propositus of the two families and patients with symptom were all accorded with the clinical features of CADASIL. It was shown by DNA sequencing that the 19(th) exon [c. 3043 T > A (p.Cys1015Ser)] in gene NOTCH3 of propositus, 2 patients (II3, III7), and a presymptomatic patient (IV1) in Family I all had heterozygosity missense mutation; and the 3(rd) exon [c.316T > G, p. (Cys106Gly)] in gene NOTCH3 of the propositus, a patient (IV3) and two presymptomatic patients (IV5, 6) in Family II all had heterozygosity missense mutation; and no mutations were detected in the 100 healthy collators. It was indicated by analyzing the function prediction that the mutation of [c. 3043 T > A (p.Cys1015Ser)] and [c.316T > G, p. (Cys106Gly)] may both influence encoding protein in NOTCH3. By analysis of the conservatism of mutation point in each species, these two basic groups were highly conserved. The heterozygosity missense mutation of 19(th) exon [c. 3043 T > A (p.Cys1015Ser)] and the 3(rd) exon [c.316T > G, p. (Cys106Gly)] in NOTCH3 gene are the new pathogenic mutations of CADASIL, and enriches the mutation spectrum of NOTCH3 gene.

Deubiquitinase function of arterivirus papain-like protease 2 suppresses the innate immune response in infected host cells.

PubMed

van Kasteren, Puck B; Bailey-Elkin, Ben A; James, Terrence W; Ninaber, Dennis K; Beugeling, Corrine; Khajehpour, Mazdak; Snijder, Eric J; Mark, Brian L; Kikkert, Marjolein

2013-02-26

Protein ubiquitination regulates important innate immune responses. The discovery of viruses encoding deubiquitinating enzymes (DUBs) suggests they remove ubiquitin to evade ubiquitin-dependent antiviral responses; however, this has never been conclusively demonstrated in virus-infected cells. Arteriviruses are economically important positive-stranded RNA viruses that encode an ovarian tumor (OTU) domain DUB known as papain-like protease 2 (PLP2). This enzyme is essential for arterivirus replication by cleaving a site within the viral replicase polyproteins and also removes ubiquitin from cellular proteins. To dissect this dual specificity, which relies on a single catalytic site, we determined the crystal structure of equine arteritis virus PLP2 in complex with ubiquitin (1.45 Å). PLP2 binds ubiquitin using a zinc finger that is uniquely integrated into an exceptionally compact OTU-domain fold that represents a new subclass of zinc-dependent OTU DUBs. Notably, the ubiquitin-binding surface is distant from the catalytic site, which allowed us to mutate this surface to significantly reduce DUB activity without affecting polyprotein cleavage. Viruses harboring such mutations exhibited WT replication kinetics, confirming that PLP2-mediated polyprotein cleavage was intact, but the loss of DUB activity strikingly enhanced innate immune signaling. Compared with WT virus infection, IFN-β mRNA levels in equine cells infected with PLP2 mutants were increased by nearly an order of magnitude. Our findings not only establish PLP2 DUB activity as a critical factor in arteriviral innate immune evasion, but the selective inactivation of DUB activity also opens unique possibilities for developing improved live attenuated vaccines against arteriviruses and other viruses encoding similar dual-specificity proteases.

A missense mutation in the arginine-vasopressin neurophysin-II gene causes autosomal dominant neurohypophyseal diabetes insipidus in a Chinese family.

PubMed

Ye, Dan; Dong, FengQin; Lu, WeiQin; Zhang, Zhe; Lu, XunLiang; Li, ChengJiang; Liu, YanNing

2013-06-01

Familial neurohypophyseal diabetes insipidus, an autosomal dominant disorder, is mostly caused by mutations in the genes that encode AVP or its intracellular binding protein, neurophysin-II. The mutations lead to aberrant preprohormone processing and progressive destruction of AVP-secreting cells, which gradually manifests a progressive polyuria and polydipsia during early childhood, and a disorder of water homeostasis. We characterized the clinical and biochemical features, and sequenced the AVP neurophysin-II(AVP-NPII) gene of the affected individuals with autosomal dominant neurohypophyseal diabetes insipidus(ADNDI)to determine whether this disease was genetically determined. We obtained the histories of eight affected and four unaffected family individuals. The diagnosis of ADNDI was established using a water deprivation test and exogenous AVP administration. For molecular analysis, genomic DNA was extracted and the AVP-NPII gene was amplified using polymerase chain reaction and sequenced. The eight affected individuals showed different spectra of age of onsets (7-15 years) and urine volumes (132-253 ml/kg/24 h). All affected individuals responded to vasopressin administration, with a resolution of symptoms and an increase in urine osmolality by more than 50%. The characteristic hyperintense signal in the posterior pituitary on T1-weighted magnetic resonance imaging was absent in six family members and present in one. Sequencing analysis revealed a missense heterozygous mutation 1516G > T (Gly17Val) in exon 2 of the AVP-NPII gene among the ADNDI individuals. We identified a missense mutation in the AVP-NPII gene and the same mutation showed different spectra of age of onsets and urine volumes in a new Chinese family with ADNDI. The mutation may provide a molecular basis for understanding the characteristics of NPII and add to our knowledge of the pathogenesis of ADNDI, which would allow the presymptomatic diagnosis of asymptomatic subjects. © 2012 John Wiley

A mitochondrial tRNA(His) gene mutation causing pigmentary retinopathy and neurosensorial deafness.

PubMed

Crimi, M; Galbiati, S; Perini, M P; Bordoni, A; Malferrari, G; Sciacco, M; Biunno, I; Strazzer, S; Moggio, M; Bresolin, N; Comi, G P

2003-04-08

We have identified a heteroplasmic G to A mutation at position 12,183 of the mitochondrial transfer RNA Histidine (tRNA(His)) gene in three related patients. These phenotypes varied according to mutation heteroplasmy: one had severe pigmentary retinopathy, neurosensorial deafness, testicular dysfunction, muscle hypotrophy, and ataxia; the other two had only retinal and inner ear involvement. The mutation is in a highly conserved region of the T(psi)C stem of the tRNA(His) gene and may alter secondary structure formation. This is the first described pathogenic, maternally inherited mutation of the mitochondrial tRNA(His) gene.

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.

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.

Nanoparticles for Retinal Gene Therapy

PubMed Central

Conley, Shannon M.; Naash, Muna I.

2010-01-01

Ocular gene therapy is becoming a well-established field. Viral gene therapies for the treatment of Leber’s congentinal amaurosis (LCA) are in clinical trials, and many other gene therapy approaches are being rapidly developed for application to diverse ophthalmic pathologies. Of late, development of non-viral gene therapies has been an area of intense focus and one technology, polymer-compacted DNA nanoparticles, is especially promising. However, development of pharmaceutically and clinically viable therapeutics depends not only on having an effective and safe vector but also on a practical treatment strategy. Inherited retinal pathologies are caused by mutations in over 220 genes, some of which contain over 200 individual disease-causing mutations, which are individually very rare. This review will focus on both the progress and future of nanoparticles and also on what will be required to make them relevant ocular pharmaceutics. PMID:20452457

Gene expression profiling and candidate gene resequencing identifies pathways and mutations important for malignant transformation caused by leukemogenic fusion genes.

PubMed

Novak, Rachel L; Harper, David P; Caudell, David; Slape, Christopher; Beachy, Sarah H; Aplan, Peter D

2012-12-01

NUP98-HOXD13 (NHD13) and CALM-AF10 (CA10) are oncogenic fusion proteins produced by recurrent chromosomal translocations in patients with acute myeloid leukemia (AML). Transgenic mice that express these fusions develop AML with a long latency and incomplete penetrance, suggesting that collaborating genetic events are required for leukemic transformation. We employed genetic techniques to identify both preleukemic abnormalities in healthy transgenic mice as well as collaborating events leading to leukemic transformation. Candidate gene resequencing revealed that 6 of 27 (22%) CA10 AMLs spontaneously acquired a Ras pathway mutation and 8 of 27 (30%) acquired an Flt3 mutation. Two CA10 AMLs acquired an Flt3 internal-tandem duplication, demonstrating that these mutations can be acquired in murine as well as human AML. Gene expression profiles revealed a marked upregulation of Hox genes, particularly Hoxa5, Hoxa9, and Hoxa10 in both NHD13 and CA10 mice. Furthermore, mir196b, which is embedded within the Hoxa locus, was overexpressed in both CA10 and NHD13 samples. In contrast, the Hox cofactors Meis1 and Pbx3 were differentially expressed; Meis1 was increased in CA10 AMLs but not NHD13 AMLs, whereas Pbx3 was consistently increased in NHD13 but not CA10 AMLs. Silencing of Pbx3 in NHD13 cells led to decreased proliferation, increased apoptosis, and decreased colony formation in vitro, suggesting a previously unexpected role for Pbx3 in leukemic transformation. Published by Elsevier Inc.

Building predictive gene signatures through simultaneous assessment of transcription factor activation and gene expression.

EPA Science Inventory

Building predictive gene signatures through simultaneous assessment of transcription factor activation and gene expression Exposure to many drugs and environmentally-relevant chemicals can cause adverse outcomes. These adverse outcomes, such as cancer, have been linked to mol...

NMR Crystallography of a Carbanionic Intermediate in Tryptophan Synthase: Chemical Structure, Tautomerization, and Reaction Specificity.

PubMed

Caulkins, Bethany G; Young, Robert P; Kudla, Ryan A; Yang, Chen; Bittbauer, Thomas J; Bastin, Baback; Hilario, Eduardo; Fan, Li; Marsella, Michael J; Dunn, Michael F; Mueller, Leonard J

2016-11-23

Carbanionic intermediates play a central role in the catalytic transformations of amino acids performed by pyridoxal-5'-phosphate (PLP)-dependent enzymes. Here, we make use of NMR crystallography-the synergistic combination of solid-state nuclear magnetic resonance, X-ray crystallography, and computational chemistry-to interrogate a carbanionic/quinonoid intermediate analogue in the β-subunit active site of the PLP-requiring enzyme tryptophan synthase. The solid-state NMR chemical shifts of the PLP pyridine ring nitrogen and additional sites, coupled with first-principles computational models, allow a detailed model of protonation states for ionizable groups on the cofactor, substrates, and nearby catalytic residues to be established. Most significantly, we find that a deprotonated pyridine nitrogen on PLP precludes formation of a true quinonoid species and that there is an equilibrium between the phenolic and protonated Schiff base tautomeric forms of this intermediate. Natural bond orbital analysis indicates that the latter builds up negative charge at the substrate C α and positive charge at C4' of the cofactor, consistent with its role as the catalytic tautomer. These findings support the hypothesis that the specificity for β-elimination/replacement versus transamination is dictated in part by the protonation states of ionizable groups on PLP and the reacting substrates and underscore the essential role that NMR crystallography can play in characterizing both chemical structure and dynamics within functioning enzyme active sites.

NMR Crystallography of a Carbanionic Intermediate in Tryptophan Synthase: Chemical Structure, Tautomerization, and Reaction Specificity

PubMed Central

2016-01-01

Carbanionic intermediates play a central role in the catalytic transformations of amino acids performed by pyridoxal-5′-phosphate (PLP)-dependent enzymes. Here, we make use of NMR crystallography—the synergistic combination of solid-state nuclear magnetic resonance, X-ray crystallography, and computational chemistry—to interrogate a carbanionic/quinonoid intermediate analogue in the β-subunit active site of the PLP-requiring enzyme tryptophan synthase. The solid-state NMR chemical shifts of the PLP pyridine ring nitrogen and additional sites, coupled with first-principles computational models, allow a detailed model of protonation states for ionizable groups on the cofactor, substrates, and nearby catalytic residues to be established. Most significantly, we find that a deprotonated pyridine nitrogen on PLP precludes formation of a true quinonoid species and that there is an equilibrium between the phenolic and protonated Schiff base tautomeric forms of this intermediate. Natural bond orbital analysis indicates that the latter builds up negative charge at the substrate Cα and positive charge at C4′ of the cofactor, consistent with its role as the catalytic tautomer. These findings support the hypothesis that the specificity for β-elimination/replacement versus transamination is dictated in part by the protonation states of ionizable groups on PLP and the reacting substrates and underscore the essential role that NMR crystallography can play in characterizing both chemical structure and dynamics within functioning enzyme active sites. PMID:27779384

Structures of Bacterial Biosynthetic Arginine Decarboxylases

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

F Forouhar; S Lew; J Seetharaman

2011-12-31

Biosynthetic arginine decarboxylase (ADC; also known as SpeA) plays an important role in the biosynthesis of polyamines from arginine in bacteria and plants. SpeA is a pyridoxal-5'-phosphate (PLP)-dependent enzyme and shares weak sequence homology with several other PLP-dependent decarboxylases. Here, the crystal structure of PLP-bound SpeA from Campylobacter jejuni is reported at 3.0 {angstrom} resolution and that of Escherichia coli SpeA in complex with a sulfate ion is reported at 3.1 {angstrom} resolution. The structure of the SpeA monomer contains two large domains, an N-terminal TIM-barrel domain followed by a {beta}-sandwich domain, as well as two smaller helical domains. Themore » TIM-barrel and {beta}-sandwich domains share structural homology with several other PLP-dependent decarboxylases, even though the sequence conservation among these enzymes is less than 25%. A similar tetramer is observed for both C. jejuni and E. coli SpeA, composed of two dimers of tightly associated monomers. The active site of SpeA is located at the interface of this dimer and is formed by residues from the TIM-barrel domain of one monomer and a highly conserved loop in the {beta}-sandwich domain of the other monomer. The PLP cofactor is recognized by hydrogen-bonding, {pi}-stacking and van der Waals interactions.« less

Prevalence and Predictors of Low Vitamin B6 Status in Healthy Young Adult Women in Metro Vancouver.

PubMed

Ho, Chia-Ling; Quay, Teo A W; Devlin, Angela M; Lamers, Yvonne

2016-09-01

Low periconceptional vitamin B6 (B6) status has been associated with an increased risk of preterm birth and early pregnancy loss. Given many pregnancies are unplanned; it is important for women to maintain an adequate B6 status throughout reproductive years. There is limited data on B6 status in Canadian women. This study aimed to assess the prevalence of B6 deficiency and predictors of B6 status in young adult women in Metro Vancouver. We included a convenience sample of young adult non-pregnant women (19-35 years; n = 202). Vitamin B6 status was determined using fasting plasma concentrations of pyridoxal 5'-phosphate (PLP). Mean (95% confidence interval) plasma PLP concentration was 61.0 (55.2, 67.3) nmol/L. The prevalence of B6 deficiency (plasma PLP < 20 nmol/L) was 1.5% and that of suboptimal B6 status (plasma PLP = 20-30 nmol/L) was 10.9%. Body mass index, South Asian ethnicity, relative dietary B6 intake, and the use of supplemental B6 were significant predictors of plasma PLP. The combined 12.4% prevalence of B6 deficiency and suboptimal status was lower than data reported in US populations and might be due to the high socioeconomic status of our sample. More research is warranted to determine B6 status in the general Canadian population.

AccuCopy quantification combined with pre-amplification of long-distance PCR for fast analysis of intron 22 inversion in haemophilia A.

PubMed

Ding, Qianlan; Wu, Xi; Lu, Yeling; Chen, Changming; Shen, Rui; Zhang, Xi; Jiang, Zhengwen; Wang, Xuefeng

2016-07-01

To develop a digitalized intron 22 inversion (Inv22) detection in patients with severe haemophilia A. The design included two tests: A genotyping test included two multiplex pre-amplification of LD-PCR (PLP) with two combinations of five primers to amplify wild-type and chimeric int22h alleles; a carrier mosaicism test was similar to the genotyping test except only amplification of chimeric int22h alleles by removing one primer from each of two combinations. AccuCopy detection was used to quantify PLP products. PLP product patterns in the genotyping test allowed identifying all known Inv22. Quantitative patterns accurately represented the product patterns. The results of 164 samples detected by the genotyping test were consistent with those obtained by LD-PCR detection. Limit of detection (LOD) of the carrier mosaicism test was at least 2% of heterozygous cells with Inv22. Performing the test in two obligate mothers with negative Inv22 from two sporadic pedigrees mosaic rates of blood and hair root of the mother from pedigree 1 were 8.3% and >20%, respectively and negative results were obtained in pedigree 2. AccuCopy quantification combined with PLP (AQ-PLP) method was confirmed to be rapid and reliable for genotyping Inv22 and highly sensitive to carrier mosaicism detection. Copyright © 2016 Elsevier B.V. All rights reserved.

Hospitalizations for Accidents and Injuries in the U.S. Navy. III. Time in Assignment and Seniority.

DTIC Science & Technology

1985-09-01

MICROCOPY RESOLUTION TEST CHART NATIONAL BJREAU OF STANARDS 1963 A 2i ... ........................ . . . . . . . . . . . . AD-A161 677 HOSPITALIZATIONS FOR...formula ( Lilienfeld and Lilien- feld, 19801: p ± 1.96 (p)(l-P) N Relative risk levels were then computed as the ratio of these rates; 95% confidence...financial change, etc.) which bring about a significant change * in an individual’s ongoing life pattern may cause psychological turrmoil that can

Diabetes-causing gene, kruppel-like factor 11, modulates the antinociceptive response of chronic ethanol intake.

PubMed

Ou, Xiao-Ming; Udemgba, Chinelo; Wang, Niping; Dai, Xiaoli; Lomberk, Gwen; Seo, Seungmae; Urrutia, Raul; Wang, Junming; Duncan, Jeremy; Harris, Sharonda; Fairbanks, Carolyn A; Zhang, Xiao

2014-02-01

Alcohol (EtOH [ethanol]) is an antinociceptive agent, working in part, by reducing sensitivity to painful stimuli. The transcription factor Kruppel-like factor 11 (KLF11), a human diabetes-causing gene that also regulates the neurotransmitter metabolic enzymes monoamine oxidase (MAO), has recently been identified as an EtOH-inducible gene. However, its role in antinociception remains unknown. Consequently, we investigated the function of KLF11 in chronic EtOH-induced antinociception using a genetically engineered knockout mouse model. Wild-type (Klf11(+/+) ) and KLF11 knockout (Klf11(-/-) ) mice were fed a liquid diet containing EtOH for 28 days with increasing amounts of EtOH from 0% up to a final concentration of 6.4%, representing a final diet containing 36% of calories primarily from EtOH. Control mice from both genotypes were fed liquid diet without EtOH for 28 days. The EtOH-induced antinociceptive effect was determined using the tail-flick test before and after EtOH exposure (on day 29). In addition, the enzyme activity and mRNA levels of MAO A and MAO B were measured by real-time RT-PCR and enzyme assays, respectively. EtOH produced an antinociceptive response to thermal pain in Klf11(+/+) mice, as expected. In contrast, deletion of KLF11 in the Klf11(-/-) mice abolished the EtOH-induced antinociceptive effect. The mRNA and protein levels of KLF11 were significantly increased in the brain prefrontal cortex of Klf11(+/+) mice exposed to EtOH compared with control Klf11(+/+) mice. Furthermore, MAO enzyme activities were affected differently in Klf11 wild-type versus Klf11 knockout mice exposed to chronic EtOH. Chronic EtOH intake significantly increased MAO B activity in Klf11(+/+) mice. The data show KLF11 modulation of EtOH-induced antinociception. The KLF11-targeted MAO B enzyme may contribute more significantly to EtOH-induced antinociception. Thus, this study revealed a new role for the KLF11 gene in the mechanisms underlying the antinociceptive

Amino-modified polystyrene nanoparticles affect signalling pathways of the sea urchin (Paracentrotus lividus) embryos.

PubMed

Pinsino, Annalisa; Bergami, Elisa; Della Torre, Camilla; Vannuccini, Maria Luisa; Addis, Piero; Secci, Marco; Dawson, Kenneth A; Matranga, Valeria; Corsi, Ilaria

2017-03-01

Polystyrene nanoparticles have been shown to pose serious risk to marine organisms including sea urchin embryos based on their surface properties and consequently behaviour in natural sea water. The aim of this study is to investigate the toxicity pathways of amino polystyrene nanoparticles (PS-NH 2 , 50 nm) in Paracentrotus lividus embryos in terms of development and signalling at both protein and gene levels. Two sub-lethal concentrations of 3 and 4 μg/mL of PS-NH 2 were used to expose sea urchin embryos in natural sea water (PS-NH 2 as aggregates of 143 ± 5 nm). At 24 and 48 h post-fertilisation (hpf) embryonic development was monitored and variations in the levels of key proteins involved in stress response and development (Hsp70, Hsp60, MnSOD, Phospho-p38 Mapk) as well as the modulation of target genes (Pl-Hsp70, Pl-Hsp60, Pl-Cytochrome b, Pl-p38 Mapk, Pl-Caspase 8, Pl-Univin) were measured. At 48 hpf various striking teratogenic effects were observed such as the occurrence of cells/masses randomly distributed, severe skeletal defects and delayed development. At 24 hpf a significant up-regulation of Pl-Hsp70, Pl-p38 Mapk, Pl-Univin and Pl-Cas8 genes was found, while at 48 hpf only for Pl-Univin was observed. Protein profile showed different patterns as a significant increase of Hsp70 and Hsp60 only after 48 hpf compared to controls. Conversely, P-p38 Mapk protein significantly increased at 24 hpf and decreased at 48 hpf. Our findings highlight that PS-NH 2 are able to disrupt sea urchin embryos development by modulating protein and gene profile providing new understandings into the signalling pathways involved.

The seed dormancy defect of Arabidopsis mutants lacking the transcript elongation factor TFIIS is caused by reduced expression of the DOG1 gene.

PubMed

Mortensen, Simon A; Grasser, Klaus D

2014-01-03

TFIIS is a transcript elongation factor that facilitates transcription by RNA polymerase II, as it assists the enzyme to bypass blocks to mRNA synthesis. Previously, we have reported that Arabidopsis plants lacking TFIIS exhibit reduced seed dormancy. Among the genes differentially expressed in tfIIs seeds, the DOG1 gene was identified that is a known QTL for seed dormancy. Here we have analysed plants that overexpress TFIIS in wild type background, or that harbour an additional copy of DOG1 in tfIIs mutant background. These experiments demonstrate that the down-regulation of DOG1 expression causes the seed dormancy phenotype of tfIIs mutants. Copyright © 2013 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

Oligonucleotide treatment causes flax β-glucanase up-regulation via changes in gene-body methylation.

PubMed

Wojtasik, Wioleta; Kulma, Anna; Boba, Aleksandra; Szopa, Jan

2014-10-05

Nowadays, the challenge for biotechnology is to develop tools for agriculture and industry to provide plants characterized by productivity and quality that will satisfy the growing demand for different kinds of natural products. To meet the challenge, the generation and application of genetically modified plants is justified. However, the strong social resistance to genetically modified organisms and restrictive regulations in European Union countries necessitated the development of a new technology for new plant types generation which uses the knowledge resulting from analysis of genetically modified plants to generate favourably altered plants while omitting the introduction of heterologous genes to their genome. Four-year experiments led to the development of a technology inducing heritable epigenetic gene activation without transgenesis. The method comprises the induction of changes in methylation/demethylation of the endogenous gene by the plant's treatment with short oligodeoxynucleotides antisense to the coding region. In vitro cultured plants and F3 generation flax plants overproducing the β-1,3-glucanase gene (EMO-βGlu flax) were characterized by up-regulation of β-glucanase and chitinase genes, decreases in the methylation of CCGG sequences in the β-glucanase gene and in total DNA methylation and, more importantly, reasonable resistance against Fusarium infection. In addition, EMO-βGlu flax obtained by this technology showed similar features as those obtained by genetic engineering. To our best knowledge, this is the first report on plant gene activation by treatment with oligodeoxynucleotides homologous to the coding region of the gene. Apart from the evident effectiveness, the most important issue is that the EMO method allows generation of favourably altered plants, whose cultivation makes the plant producer independent from the complicated procedure of obtaining an agreement on GMO release into the environment and whose products might be more

The T1048I mutation in ATP7A gene causes an unusual Menkes disease presentation

PubMed Central

2012-01-01

Background The ATP7A gene encodes the ATP7A protein, which is a trans-Golgi network copper transporter expressed in the brain and other organs. Mutations in this gene cause disorders of copper metabolism, such as Menkes disease. Here we describe the novel and unusual mutation (p.T1048I) in the ATP7A gene of a child with Menkes disease. The mutation affects a conserved DKTGT1048 phosphorylation motif that is involved in the catalytic activity of ATP7A. We also describe the clinical course and the response to copper treatment in this patient. Case presentation An 11-month-old male Caucasian infant was studied because of hypotonia, ataxia and global developmental delay. The patient presented low levels of serum copper and ceruloplasmin, and was shown to be hemizygous for the p.T1048I mutation in ATP7A. The diagnosis was confirmed when the patient was 18 months old, and treatment with copper-histidinate (Cu-His) was started immediately. The patient showed some neurological improvement and he is currently 8 years old. Because the p.T1048I mutation affects its catalytic site, we expected a complete loss of functional ATP7A and a classical Menkes disease presentation. However, the clinical course of the patient was mild, and he responded to Cu-His treatment, which suggests that this mutation leads to partial conservation of the activity of ATP7A. Conclusion This case emphasizes the important correlation between genotype and phenotype in patients with Menkes disease. The prognosis in Menkes disease is associated with early detection, early initiation of treatment and with the preservation of some ATP7A activity, which is necessary for Cu-His treatment response. The description of this new mutation and the response of the patient to Cu-His treatment will contribute to the growing body of knowledge about treatment response in Menkes disease. PMID:22992316

Researching the Effectiveness of a Science Professional Learning Programme Using a Proposed Curriculum Framework for Schools: A Case Study

ERIC Educational Resources Information Center

Paige, Kathryn; Zeegers, Yvonne; Lloyd, David; Roetman, Philip

2016-01-01

This paper reports on an action research-based professional learning programme (PLP) in which early career teachers volunteered to identify and then research an aspect of their science teaching practice. The PLP was facilitated by academics from the School of Education and the Barbara Hardy Institute at the University of South Australia. The…

The Personal Learning Planner: Collaboration through Online Learning and Publication

ERIC Educational Resources Information Center

Gibson, David; Sherry, Lorraine; Havelock, Bruce

2007-01-01

This paper discusses the online Personal Learning Planner (PLP) project underway at the National Institute of Community Innovations (NICI), one of the partners in the Teacher Education Network (TEN), a 2000 PT3 Catalyst grantee. The Web-based PLP provides a standards-linked "portfolio space" for both works in progress and demonstration collections…

Unusual cause of aborted sudden cardiac death in a teen athlete: homozygosity for the 4G allele of the plasminogen activase inhibitor type 1 gene.

PubMed

Phillips, Susie B; Batlivala, Sarosh; Knudson, Jarrod D

2015-10-01

Common aetiologies of sudden cardiac death in children include coronary anomalies, channelopathies, and cardiomyopathies. Less frequently, hypercoagulable states cause sudden arrest. We report an unusual case of aborted sudden cardiac death in a teenager, ultimately found to have homozygosity for the 4G allele of the plasminogen activase inhibitor type 1 gene.

The effect of in silico targeting Pseudomonas aeruginosa patatin-like protein D, for immunogenic administration.

PubMed

Chirani, Alireza Salimi; Majidzadeh, Robabeh; Pouriran, Ramin; Heidary, Mohsen; Nasiri, Mohammad Javad; Gholami, Mehrdad; Goudarzi, Mehdi; Omrani, Vahid Fallah

2018-02-05

The vaccine candidates that have been introduced for immunization against Pseudomonas aeruginosa (P. aeruginosa) strains are quite diverse. In fact, there has been no proper antigen to act as an effective immunogenic substance against this ubiquitous pathogen in the market as yet. The complications caused by this bacterium due to the rapid development of multiple drug resistant strains have led to clinical problems worldwide. P. aeruginosa encodes many specific virulence elements that could be used as appropriate vaccine candidates. Type Vd secretion system, also known as patatin-like protein D, is a novel P. aeruginosa auto-transporter system. It is known that cellular or humoral immune responses could be elevated by chimeric proteins carrying epitopes. It has been recognized that in silico tools are essential for the evaluation of new chimeric antigens. In this study, we have considered the patatin-like protein D (PlpD) molecule from P. aeruginosa and predicted some immunogenic properties of this strong cytotoxic phospholipase A2 with the use of in-depth computational and immunoinformatics assessment methods The novelty of our in silico study is the modeling and assessment of both humoral and cellular immune potential against the PlpD molecule. The molecule was considered by multiple sequence alignment and homology valuation. The extremely conserved regions in the PlpD were predicted. The allergenic and physicochemical property predictions on the PlpD state that the molecule is a non-allergic and stable molecule. High-resolution secondary and tertiary conformations were created. Indeed, the B-cell and T-cell epitope mapping on the chimeric target protein confirmed that the engineered protein contained a tremendous number of both B-cell and T-cell corresponding epitopes. This investigation magnificently attained the chimeric molecule as being a potent lipolytic enzyme composed of numerous B-cell and T-cell restricted epitopes and could induce both humoral and

In silico analysis of miRNA-mediated gene regulation in OCA and OA genes.

PubMed

Kamaraj, Balu; Gopalakrishnan, Chandrasekhar; Purohit, Rituraj

2014-12-01

Albinism is an autosomal recessive genetic disorder due to low secretion of melanin. The oculocutaneous albinism (OCA) and ocular albinism (OA) genes are responsible for melanin production and also act as a potential targets for miRNAs. The role of miRNA is to inhibit the protein synthesis partially or completely by binding with the 3'UTR of the mRNA thus regulating gene expression. In this analysis, we predicted the genetic variation that occurred in 3'UTR of the transcript which can be a reason for low melanin production thus causing albinism. The single nucleotide polymorphisms (SNPs) in 3'UTR cause more new binding sites for miRNA which binds with mRNA which leads to inhibit the translation process either partially or completely. The SNPs in the mRNA of OCA and OA genes can create new binding sites for miRNA which may control the gene expression and lead to hypopigmentation. We have developed a computational procedure to determine the SNPs in the 3'UTR region of mRNA of OCA (TYR, OCA2, TYRP1 and SLC45A2) and OA (GPR143) genes which will be a potential cause for albinism. We identified 37 SNPs in five genes that are predicted to create 87 new binding sites on mRNA, which may lead to abrogation of the translation process. Expression analysis confirms that these genes are highly expressed in skin and eye regions. It is well supported by enrichment analysis that these genes are mainly involved in eye pigmentation and melanin biosynthesis process. The network analysis also shows how the genes are interacting and expressing in a complex network. This insight provides clue to wet-lab researches to understand the expression pattern of OCA and OA genes and binding phenomenon of mRNA and miRNA upon mutation, which is responsible for inhibition of translation process at genomic levels.

Involvement of Vitamin B6 Biosynthesis Pathways in the Insecticidal Activity of Photorhabdus luminescens.

PubMed

Sato, Kazuki; Yoshiga, Toyoshi; Hasegawa, Koichi

2016-06-15

Photorhabdus luminescens is a Gram-negative entomopathogenic bacterium which symbiotically associates with the entomopathogenic nematode Heterorhabditis bacteriophora P. luminescens is highly virulent to many insects and nonsymbiotic nematodes, including Caenorhabditis elegans To understand the virulence mechanisms of P. luminescens, we obtained virulence-deficient and -attenuated mutants against C. elegans through a transposon-mutagenized library. From the genetic screening, we identified the pdxB gene, encoding erythronate-4-phosphate dehydrogenase, as required for de novo vitamin B6 biosynthesis. Mutation in pdxB caused growth deficiency of P. luminescens in nutrient-poor medium, which was restored under nutrient-rich conditions or by supplementation with pyridoxal 5'-phosphate (PLP), an active form of vitamin B6 Supplementation with three other B6 vitamers (pyridoxal, pyridoxine, and pyridoxamine) also restored the growth of the pdxB mutant, suggesting the existence of a salvage pathway for vitamin B6 biosynthesis in P. luminescens Moreover, supplementation with PLP restored the virulence-deficient phenotype against C. elegans Combining these results with the fact that pdxB mutation also caused attenuation of insecticidal activity, we concluded that the production of appropriate amounts of vitamin B6 is critical for P. luminescens pathogenicity. The Gram-negative entomopathogenic bacterium Photorhabdus luminescens symbiotically associates with the entomopathogenic nematode Heterorhabditis bacteriophora P. luminescens is highly virulent to many insects and nonsymbiotic nematodes, including Caenorhabditis elegans We have obtained several virulence-deficient and -attenuated P. luminescens mutants against C. elegans through genetic screening. From the genetic analysis, we present the vitamin B6 biosynthetic pathways in P. luminescens that are important for its insecticidal activity. Mutation in pdxB, encoding erythronate-4-phosphate dehydrogenase and required for

Inversion of exons 1-7 of the MSH2 gene is a frequent cause of unexplained Lynch syndrome in one local population.

PubMed

Rhees, Jennifer; Arnold, Mildred; Boland, C Richard

2014-06-01

Germline mutations in DNA mismatch repair (MMR) genes, such as MSH2, cause Lynch syndrome, an autosomal dominant predisposition to colorectal as well as other cancers. Our research clinic focuses on hereditary colorectal cancer, and over the past 9 years we have identified germline mutations in DNA MMR genes in 101 patients using commercial genetic reference laboratories. We also collected samples from twelve patients with absent MSH2 protein expression and microsatellite instability in tumor tissue, with a family history suggestive of Lynch syndrome, but negative germline test results. The most likely explanation for this set of results is that the germline testing did not detect true germline mutations in these patients. Two of our patients with failed commercial testing were later found to have deletions in the 3' region of EPCAM, the gene just upstream of MSH2, but no explanation could be found for inactivation of MSH2 in the other ten patients. We used allelic dropout in long PCR to look for potential regions of rearrangement in the MSH2 gene. This method detected a potential rearrangement breakpoint in the same region of MSH2 where one breakpoint of a 10 Mb inversion was reported previously. We tested these ten patients for this inversion. Six of 10 patients had the inversion, indicating the importance of including testing for this inversion in patients suspected of having MSH2-type Lynch syndrome in our population. Additionally, this method could be further developed to look for inversions in other genes where current methods of testing fail to find a causative mutation.

Common Variable Immunodeficiency Caused by FANC Mutations.

PubMed

Sekinaka, Yujin; Mitsuiki, Noriko; Imai, Kohsuke; Yabe, Miharu; Yabe, Hiromasa; Mitsui-Sekinaka, Kanako; Honma, Kenichi; Takagi, Masatoshi; Arai, Ayako; Yoshida, Kenichi; Okuno, Yusuke; Shiraishi, Yuichi; Chiba, Kenichi; Tanaka, Hiroko; Miyano, Satoru; Muramatsu, Hideki; Kojima, Seiji; Hira, Asuka; Takata, Minoru; Ohara, Osamu; Ogawa, Seishi; Morio, Tomohiro; Nonoyama, Shigeaki

2017-07-01

Common variable immunodeficiency (CVID) is the most common adult-onset primary antibody deficiency disease due to various causative genes. Several genes, which are known to be the cause of different diseases, have recently been reported as the cause of CVID in patients by performing whole exome sequencing (WES) analysis. Here, we found FANC gene mutations as a cause of adult-onset CVID in two patients. B cells were absent and CD4 + T cells were skewed toward CD45RO + memory T cells. T-cell receptor excision circles (TRECs) and signal joint kappa-deleting recombination excision circles (sjKRECs) were undetectable in both patients. Both patients had no anemia, neutropenia, or thrombocytopenia. Using WES, we identified compound heterozygous mutations of FANCE in one patient and homozygous mutation of FANCA in another patient. The impaired function of FANC protein complex was confirmed by a monoubiquitination assay and by chromosome fragility test. We then performed several immunological evaluations including quantitative lymphocyte analysis and TRECs/sjKRECs analysis for 32 individuals with Fanconi anemia (FA). In total, 22 FA patients (68.8%) were found to have immunological abnormalities, suggesting that such immunological findings may be common in FA patients. These data indicate that FANC mutations are involved in impaired lymphogenesis probably by the accumulation of DNA replication stress, leading to CVID. It is important to diagnose FA because it drastically changes clinical management. We propose that FANC mutations can cause isolated immunodeficiency in addition to bone marrow failure and malignancy.

Differential phosphorylation patterns of P-glycoprotein reconstituted into a proteoliposome system: insight into additional unconventional phosphorylation sites.

PubMed

Lelong-Rebel, Isabelle H; Cardarelli, Carol O

2005-01-01

Membrane vesicles from the multidrug-resistant KB-V1 and KB-C1 cell lines overexpressing P-glycoprotein (Pgp), responsible for pleiotropic chemotherapeutic agents resistance, were solubilized with octyl-glucoside (OG-EX) and further fractionated on DEAE-sepharose column with increased concentrations of NaCl. The fraction containing Pgp (F3) was reconstituted into proteoliposomes (F3-PLP). Comparisons of the phosphorylation levels of Pgp achieved throughout the purification and reconstitution steps were addressed in this study. The [delta32 P] ATP-driven phosphorylation of Pgp was strongly increased in OG-EX, decreased in F3 and not detected in F3-PLP, when compared to Pgp phosphorylation in native plasma membrane vesicles. [delta32 P]ATP-phosphorylation of Pgp in F3-PLP could be restored by exogenously added PKC or by the catalytic sub-unit of PKA. The vanadate-induced hyperphosphorylation effect on Pgp by [delta32 P]ATP observed with plasma membrane vesicles was maintained in OG-EX, but was lost in F3 and did not enable labelling in F3-PLP. Enhancement of [delta32 P]-labelling of native Pgp via [delta32 P]ATP combined with GTP was maintained and also triggered phosphorylation of purified/reconstituted Pgp in F3-PLP as well. Altogether, our data suggest differential phosphorylation patterns of the transporter linked to environmental molecular composition (lipids, presence of detergent) and structure (unfolded versus embedded). In addition, restoration by GTP of Pgp phosphorylation by [delta32 P]ATP in the frame of F3-PLP suggests intra-molecular modulations and hints that other phosphorylation sites and processes, different from the classic ones involving PKC and/or PKA, may participate in the transporter's mechanism.

The effects of graded motor imagery and its components on phantom limb pain and disability in upper and lower limb amputees: a systematic review protocol.

PubMed

Limakatso, Katleho; Corten, Lieselotte; Parker, Romy

2016-09-01

Phantom limb pain (PLP) is characterized by the anatomical shifting of neighbouring somatosensory and motor areas into a deafferented cortical area of the brain contralateral to the amputated limb. It has been shown that maladaptive neuroplasticity is positively correlated to the perception of PLP in amputees. Recent studies support the use of graded motor imagery (GMI) and its component to alleviate the severity of PLP and disability. However, there is insufficient collective empirical evidence exploring the effectiveness of these treatment modalities in amputees with PLP. This systematic review will therefore explore the effects of GMI and its individual components on PLP and disability in upper and lower limb amputees. We will utilize a customized search strategy to search PubMed, Cochrane Central register of Controlled Trials, MEDLINE, Embase, PsycINFO, PEDro, Scopus, CINAHL, LILACS, DARE, Africa-Wide Information and Web of Science. We will also look at clinicaltrials.gov ( http://www.clinicaltrials.gov/ ), Pactr.gov ( http://www.pactr.org/ ) and EU Clinical trials register ( https://www.clinicaltrialsregister.eu/ ) for ongoing research. Two independent reviewers will screen articles for methodological validity. Thereafter, data from included studies will be extracted by two independent reviewers through a customized pre-set data extraction sheet. Studies with a comparable intervention and outcome measure will be pooled for meta-analysis. Studies with high heterogeneity will be analysed through random effects model. A narrative data analysis will be considered where there is insufficient data to perform a meta-analysis. Several studies investigating the effectiveness of GMI and its different components on PLP have drawn contrasting conclusions regarding the efficacy and applicability of GMI in clinical practice. This systematic review will therefore gather and critically appraise all relevant data, to generate a substantial conclusion and recommendations for

Evaluation of vitamin B6 intake and status of 20- to 64-year-old Koreans.

PubMed

Kim, Young-Nam; Cho, Youn-Ok

2014-12-01

Recent research regarding vitamin B6 status including biochemical index is limited. Thus, this study estimated intakes and major food sources of vitamin B6; determined plasma pyridoxal 5'-phosphate (PLP); and assessed vitamin B6 status of Korean adults. Three consecutive 24-h diet recalls and fasting blood samples were collected from healthy 20- to 64-year-old adults (n = 254) living in the Seoul metropolitan area, cities of Kwangju and Gumi, Korea. Vitamin B6 intake and plasma PLP were analyzed by gender and by vitamin B6 supplementation. Pearson's correlation coefficient was used to determine associations of vitamin B6 intake and plasma PLP. The mean dietary and total (dietary plus supplemental) vitamin B6 intake was 1.94 ± 0.64 and 2.41 ± 1.45 mg/day, respectively. Median (50th percentile) dietary intake of men and women was 2.062 and 1.706 mg/day. Foods from plant sources provided 70.61% of dietary vitamin B6 intake. Only 6.3% of subjects consumed total vitamin B6 less than Estimated Average Requirements. Plasma PLP concentration of all subjects was 40.03 ± 23.71 nmol/L. The concentration of users of vitamin B6 supplements was significantly higher than that of nonusers (P < 0.001). Approximately 16% of Korean adults had PLP levels < 20 nmol/L, indicating a biochemical deficiency of vitamin B6, while 19.7% had marginal vitamin B6 status. Plasma PLP concentration showed positive correlation with total vitamin B6 intake (r = 0.40984, P < 0.0001). In this study, vitamin B6 intake of Korean adults was generally adequate. However, one-third of subjects had vitamin B6 deficiency or marginal status. Therefore, in some adults in Korea, consumption of vitamin B6-rich food sources should be encouraged.

75 FR 75870 - Airworthiness Directives; Airbus Model A300 Series Airplanes

Federal Register 2010, 2011, 2012, 2013, 2014

2010-12-07

... to the aeroplane has two load paths, a Primary Load Path (PLP) and a Secondary Load Path (SLP), which is only engaged in case of PLP failure. Following the design intent, engagement of the SLP leads to... representative flights have demonstrated that, when the SLP is engaged, it does not systematically jam the THSA...

75 FR 52652 - Airworthiness Directives; Airbus Model A300 Series Airplanes

Federal Register 2010, 2011, 2012, 2013, 2014

2010-08-27

... to the aeroplane has two load paths, a Primary Load Path (PLP) and a Secondary Load Path (SLP), which is only engaged in case of PLP failure. Following the design intent, engagement of the SLP leads to... representative flights have demonstrated that, when the SLP is engaged, it does not systematically jam the THSA...

Findings from an Evaluation of a Comprehensive Parent Education Program for School-Age Parents.

ERIC Educational Resources Information Center

Braun, Maureen H.; Fuscaldo, Diane P.

This study evaluates the impact of the Parent Linking Project III (PLP III) on its participants. The PLP III is a comprehensive program for teenage mothers which offers workshops in parenting, mother-child interaction, and vocational awareness, and provides day care services that allow teen mothers to take the opportunity to complete their high…

A new compound heterozygous frameshift mutation in the type II 3{beta}-hydroxysteroid dehydrogenase 3{beta}-HSD gene causes salt-wasting 3{beta}-HSD deficiency congenital adrenal hyperplasia

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

Zhang, L.; Sakkal-Alkaddour, S.; Chang, Ying T.

1996-01-01

We report a new compound heterozygous frameshift mutation in the type II 3{Beta}-hydroxysteroid dehydrogenase (3{beta}-HSD) gene in a Pakistanian female child with the salt-wasting form of 3{Beta}-HSD deficiency congenital adrenal hyperplasia. The etiology for her congenital adrenal hyperplasia was not defined. Although the family history suggested possible 3{beta}-HSd deficiency disorder, suppressed adrenal function caused by excess glucocorticoid therapy in this child at 7 yr of age did not allow hormonal diagnosis. To confirm 3{beta}-HSD deficiency, we sequenced the type II 3{beta}-HSD gene in the patient, her family, and the parents of her deceased paternal cousins. The type II 3{beta}-HSD genemore » region of a putative promotor, exons I, II, III, and IV, and exon-intron boundaries were amplified by PCR and sequenced in all subjects. The DNA sequence of the child revealed a single nucleotide deletion at codon 318 [ACA(Thr){r_arrow}AA] in exon IV in one allele, and two nucleotide deletions at codon 273 [AAA(Lys){r_arrow}A] in exon IV in the other allele. The remaining gene sequences were normal. The codon 318 mutation was found in one allele from the father, brother, and parents of the deceased paternal cousins. The codon 273 mutation was found in one allele of the mother and a sister. These findings confirmed inherited 3{beta}-HSD deficiency in the child caused by the compound heterozygous type II 3{beta}-HSD gene mutation. Both codons at codons 279 and 367, respectively, are predicted to result in an altered and truncated type II 3{beta}-HSD protein, thereby causing salt-wasting 3{beta}-HSD deficiency in the patient. 21 refs., 2 figs., 1 tab.« less

Graphite Based Electrode for ECG Monitoring: Evaluation under Freshwater and Saltwater Conditions.

PubMed

Thap, Tharoeun; Yoon, Kwon-Ha; Lee, Jinseok

2016-04-15

We proposed new electrodes that are applicable for electrocardiogram (ECG) monitoring under freshwater- and saltwater-immersion conditions. Our proposed electrodes are made of graphite pencil lead (GPL), a general-purpose writing pencil. We have fabricated two types of electrode: a pencil lead solid type (PLS) electrode and a pencil lead powder type (PLP) electrode. In order to assess the qualities of the PLS and PLP electrodes, we compared their performance with that of a commercial Ag/AgCl electrode, under a total of seven different conditions: dry, freshwater immersion with/without movement, post-freshwater wet condition, saltwater immersion with/without movement, and post-saltwater wet condition. In both dry and post-freshwater wet conditions, all ECG-recorded PQRST waves were clearly discernible, with all types of electrodes, Ag/AgCl, PLS, and PLP. On the other hand, under the freshwater- and saltwater-immersion conditions with/without movement, as well as post-saltwater wet conditions, we found that the proposed PLS and PLP electrodes provided better ECG waveform quality, with significant statistical differences compared with the quality provided by Ag/AgCl electrodes.

The Parkinson's disease death rate: carbidopa and vitamin B6.

PubMed

Hinz, Marty; Stein, Alvin; Cole, Ted

2014-01-01

The only indication for carbidopa and benserazide is the management of L-3,4-dihydroxyphenylalanine (L-dopa)-induced nausea. Both drugs irreversibly bind to and permanently deactivate pyridoxal 5'-phosphate (PLP), the active form of vitamin B6, and PLP-dependent enzymes. PLP is required for the function of over 300 enzymes and proteins. Virtually every major system in the body is impacted directly or indirectly by PLP. The administration of carbidopa and benserazide potentially induces a nutritional catastrophe. During the first 15 years of prescribing L-dopa, a decreasing Parkinson's disease death rate was observed. Then, in 1976, 1 year after US Food and Drug Administration approved the original L-dopa/carbidopa combination drug, the Parkinson's disease death rate started increasing. This trend has continued to the present, for 38 years and counting. The previous literature documents this increasing death rate, but no hypothesis has been offered concerning this trend. Carbidopa is postulated to contribute to the increasing Parkinson's disease death rate and to the classification of Parkinson's as a progressive neurodegenerative disease. It may contribute to L-dopa tachyphylaxis.

Charcot-Marie-Tooth disease type 2 caused by homozygous MME gene mutation superimposed by chronic inflammatory demyelinating polyneuropathy.

PubMed

Fujisawa, Miwako; Sano, Yasuteru; Omoto, Masatoshi; Ogasawara, Jyun-Ichi; Koga, Michiaki; Takashima, Hiroshi; Kanda, Takashi

2017-09-30

We report a 59-year-old Japanese male who developed gradually worsening weakness and numbness of distal four extremities since age 50. His parents were first cousins, and blood and cerebral spinal examinations were unremarkable. Homozygous mutation of MME gene was detected and thus he was diagnosed as autosomal-recessive Charcot-Marie-Tooth disease 2T (AR-CMT2T); however, electrophysiological examinations revealed scattered demyelinative changes including elongated terminal latency in several peripheral nerve trunks. Sural nerve biopsy showed endoneurial edema and a lot of thinly myelinated nerve fibers with uneven distribution of remnant myelinated fibers within and between fascicles. Immunoglobulin treatment was initiated considering the possibility of superimposed inflammation and demyelination, and immediate clinical as well as electrophysiological improvements were noted. Our findings indicate that AR-CMT2T caused by MME mutation predisposes to a superimposed inflammatory demyelinating neuropathy. This is the first report which documented the co-existence of CMT2 and chronic inflammatory demyelinating polyneuropathy (CIDP); however, in the peripheral nervous system, neprilysin, a product of MME gene, is more abundant in myelin sheath than in axonal component. The fragility of myelin sheath due to mutated neprilysin may trigger the detrimental immune response against peripheral myelin in this patient.

BMPR1B mutation causes Pierre Robin sequence

PubMed Central

Yao, Xu; Zhang, Rong; Yang, Hui; Zhao, Rui; Guo, Jihong; Jin, Ke; Mei, Haibo; Luo, Yongqi; Zhao, Liu; Tu, Ming; Zhu, Yimin

2017-01-01

Background We investigated a large family with Pierre Robin sequence (PRS). Aim of the study This study aims to determine the genetic cause of PRS. Results The reciprocal translocation t(4;6)(q22;p21) was identified to be segregated with PRS in a three-generation family. Whole-genome sequencing and Sanger sequencing successfully detected breakpoints in the intragenic regions of BMRP1B and GRM4. We hypothesized that PRS in this family was caused by (i) haploinsufficiency for BMPR1B or (ii) a gain of function mechanism mediated by the BMPR1B-GRM4 fusion gene. In an unrelated family, we identified another BMPR1B-splicing mutation that co-segregated with PRS. Conclusion We detected two BMPR1B mutations in two unrelated PRS families, suggesting that BMPR1B disruption is probably a cause of human PRS. Methods GTG banding, comparative genomic hybridization, whole-genome sequencing, and Sanger sequencing were performed to identify the gene causing PRS. PMID:28418932

Emergence of a Staphylococcus aureus Clone Resistant to Mupirocin and Fusidic Acid Carrying Exotoxin Genes and Causing Mainly Skin Infections

PubMed Central

Spiliopoulou, Iris; Spyridis, Nikolaos; Giormezis, Nikolaos; Kopsidas, John; Militsopoulou, Maria; Lebessi, Evangelia; Tsolia, Maria

2017-01-01

ABSTRACT Skin and soft tissue infections (SSTIs) caused by mupirocin-resistant Staphylococcus aureus strains have recently increased in number in our settings. We sought to evaluate the characteristics of these cases over a 43-month period. Data for all community-acquired staphylococcal infections caused by mupirocin-resistant strains were retrospectively reviewed. Genes encoding products producing high-level resistance (HLR) to mupirocin (mupA), fusidic acid resistance (fusB), resistance to macrolides and lincosamides (ermC and ermA), Panton-Valentine leukocidin (PVL) (lukS/lukF-PV), exfoliative toxins (eta and etb), and fibronectin binding protein A (fnbA) were investigated by PCRs in 102 selected preserved strains. Genotyping was performed by SCCmec and agr typing, whereas clonality was determined by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). A total of 437 cases among 2,137 staphylococcal infections were recorded in 2013 to 2016; they were all SSTIs with the exception of 1 case of primary bacteremia. Impetigo was the predominant clinical entity (371 cases [84.9%]), followed by staphylococcal scalded skin syndrome (21 cases [4.8%]), and there were no abscesses. The number of infections detected annually increased during the study years. All except 3 isolates were methicillin susceptible. The rates of HLR to mupirocin and constitutive resistance to clindamycin were 99% and 20.1%, respectively. Among the 102 tested strains, 100 (98%) were mupA positive and 97 (95%) were fusB positive, 26/27 clindamycin-resistant strains (96.3%) were ermA positive, 83 strains (81.4%) were lukS/lukF positive, 95 (93%) carried both eta and etb genes, and 99 (97%) were fnbA positive. Genotyping of methicillin-sensitive S. aureus (MSSA) strains revealed that 96/99 (96.7%) belonged to one main pulsotype, pulsotype 1, classified as sequence type 121 (ST121). The emergence of a single MSSA clone (ST121) causing impetigo was documented. Resistance to

Identification and characterization of a class III chitin synthase gene of Moniliophthora perniciosa, the fungus that causes witches' broom disease of cacao.

PubMed

Souza, Catiane S; Oliveira, Bruno M; Costa, Gustavo G L; Schriefer, Albert; Selbach-Schnadelbach, Alessandra; Uetanabaro, Ana Paula T; Pirovani, Carlos P; Pereira, Gonçalo A G; Taranto, Alex G; Cascardo, Júlio Cézar de M; Góes-Neto, Aristóteles

2009-08-01

Chitin synthase (CHS) is a glucosyltransferase that converts UDP-N-acetylglucosamine into chitin, one of the main components of fungal cell wall. Class III chitin synthases act directly in the formation of the cell wall. They catalyze the conversion of the immediate precursor of chitin and are responsible for the majority of chitin synthesis in fungi. As such, they are highly specific molecular targets for drugs that can inhibit the growth and development of fungal pathogens. In this work, we have identified and characterized a chitin synthase gene of Moniliophthora perniciosa (Mopchs) by primer walking. The complete gene sequence is 3,443 bp, interrupted by 13 small introns, and comprises a cDNA with an ORF with 2,739 bp, whose terminal region was experimentally determined, encoding a protein with 913 aa that harbors all the motifs and domains typically found in class III chitin synthases. This is the first report on the characterization of a chitin synthase gene, its mature transcription product, and its putative protein in basidioma and secondary mycelium stages of M. perniciosa, a basidiomycotan fungus that causes witches' broom disease of cacao.

A mutation in the MATP gene causes the cream coat colour in the horse

PubMed Central

Mariat, Denis; Taourit, Sead; Guérin, Gérard

2003-01-01

In horses, basic colours such as bay or chestnut may be partially diluted to buckskin and palomino, or extremely diluted to cream, a nearly white colour with pink skin and blue eyes. This dilution is expected to be controlled by one gene and we used both candidate gene and positional cloning strategies to identify the "cream mutation". A horse panel including reference colours was established and typed for different markers within or in the neighbourhood of two candidate genes. Our data suggest that the causal mutation, a G to A transition, is localised in exon 2 of the MATP gene leading to an aspartic acid to asparagine substitution in the encoded protein. This conserved mutation was also described in mice and humans, but not in medaka. PMID:12605854

A Putative Type III Secretion System Effector Encoded by the MA20_12780 Gene in Bradyrhizobium japonicum Is-34 Causes Incompatibility with Rj4 Genotype Soybeans

PubMed Central

Hashimoto, Syougo; Okizaki, Kouhei; Kanesaki, Yu; Yoshikawa, Hirofumi; Yamakawa, Takeo

2015-01-01

The nodulation of Bradyrhizobium japonicum Is-34 is restricted by Rj4 genotype soybeans (Glycine max). To identify the genes responsible for this incompatibility, Tn5 mutants of B. japonicum Is-34 that were able to overcome this nodulation restriction were obtained. Analysis of the Tn5 mutants revealed that Tn5 was inserted into a region containing the MA20_12780 gene. In addition, direct disruption of this gene using marker exchange overcame the nodulation restriction by Rj4 genotype soybeans. The MA20_12780 gene has a tts box motif in its upstream region, indicating a possibility that this gene encodes a type III secretion system (T3SS) effector protein. Bioinformatic characterization revealed that the MA20_12780 protein contains the small ubiquitin-like modifier (SUMO) protease domain of the C48 peptidase (ubiquitin-like protease 1 [Ulp1]) family. The results of the present study indicate that a putative T3SS effector encoded by the MA20_12780 gene causes the incompatibility with Rj4 genotype soybeans, and they suggest the possibility that the nodulation restriction of B. japonicum Is-34 may be due to Rj4 genotype soybeans recognizing the putative T3SS effector (MA20_12780 protein) as a virulence factor. PMID:26092458

Temporal parameters and time course of perceptual latency priming.

PubMed

Scharlau, Ingrid; Neumann, Odmar

2003-06-01

Visual stimuli (primes) reduce the perceptual latency of a target appearing at the same location (perceptual latency priming, PLP). Three experiments assessed the time course of PLP by masked and, in Experiment 3, unmasked primes. Experiments 1 and 2 investigated the temporal parameters that determine the size of priming. Stimulus onset asynchrony was found to exert the main influence accompanied by a small effect of prime duration. Experiment 3 used a large range of priming onset asynchronies. We suggest to explain PLP by the Asynchronous Updating Model which relates it to the asynchrony of 2 central coding processes, preattentive coding of basic visual features and attentional orienting as a prerequisite for perceptual judgments and conscious perception.

Gene expression meta-analysis identifies chromosomal regions and candidate genes involved in breast cancer metastasis.

PubMed

Thomassen, Mads; Tan, Qihua; Kruse, Torben A

2009-01-01

Breast cancer cells exhibit complex karyotypic alterations causing deregulation of numerous genes. Some of these genes are probably causal for cancer formation and local growth whereas others are causal for the various steps of metastasis. In a fraction of tumors deregulation of the same genes might be caused by epigenetic modulations, point mutations or the influence of other genes. We have investigated the relation of gene expression and chromosomal position, using eight datasets including more than 1200 breast tumors, to identify chromosomal regions and candidate genes possibly causal for breast cancer metastasis. By use of "Gene Set Enrichment Analysis" we have ranked chromosomal regions according to their relation to metastasis. Overrepresentation analysis identified regions with increased expression for chromosome 1q41-42, 8q24, 12q14, 16q22, 16q24, 17q12-21.2, 17q21-23, 17q25, 20q11, and 20q13 among metastasizing tumors and reduced gene expression at 1p31-21, 8p22-21, and 14q24. By analysis of genes with extremely imbalanced expression in these regions we identified DIRAS3 at 1p31, PSD3, LPL, EPHX2 at 8p21-22, and FOS at 14q24 as candidate metastasis suppressor genes. Potential metastasis promoting genes includes RECQL4 at 8q24, PRMT7 at 16q22, GINS2 at 16q24, and AURKA at 20q13.

A recessive contiguous gene deletion causing infantile hyperinsulinism, enteropathy and deafness identifies the Usher type 1C gene.

PubMed

Bitner-Glindzicz, M; Lindley, K J; Rutland, P; Blaydon, D; Smith, V V; Milla, P J; Hussain, K; Furth-Lavi, J; Cosgrove, K E; Shepherd, R M; Barnes, P D; O'Brien, R E; Farndon, P A; Sowden, J; Liu, X Z; Scanlan, M J; Malcolm, S; Dunne, M J; Aynsley-Green, A; Glaser, B

2000-09-01

Usher syndrome type 1 describes the association of profound, congenital sensorineural deafness, vestibular hypofunction and childhood onset retinitis pigmentosa. It is an autosomal recessive condition and is subdivided on the basis of linkage analysis into types 1A through 1E. Usher type 1C maps to the region containing the genes ABCC8 and KCNJ11 (encoding components of ATP-sensitive K + (KATP) channels), which may be mutated in patients with hyperinsulinism. We identified three individuals from two consanguineous families with severe hyperinsulinism, profound congenital sensorineural deafness, enteropathy and renal tubular dysfunction. The molecular basis of the disorder is a homozygous 122-kb deletion of 11p14-15, which includes part of ABCC8 and overlaps with the locus for Usher syndrome type 1C and DFNB18. The centromeric boundary of this deletion includes part of a gene shown to be mutated in families with type 1C Usher syndrome, and is hence assigned the name USH1C. The pattern of expression of the USH1C protein is consistent with the clinical features exhibited by individuals with the contiguous gene deletion and with isolated Usher type 1C.

Scarlet fever is caused by a limited number of Streptococcus pyogenes lineages and is associated with the exotoxin genes ssa, speA and speC.

PubMed

Silva-Costa, Catarina; Carriço, Joao A; Ramirez, Mario; Melo-Cristino, Jose

2014-03-01

Several outbreaks of scarlet fever caused by Streptococcus pyogenes were recently reported. Scarlet fever is historically considered a toxin-mediated disease, dependent on the production of the exotoxins SpeA and SpeC, but a strict association between scarlet fever and these exotoxins is not always detected. The aims of this study were to characterize the scarlet fever bacterial isolates recovered from patients in a Lisbon hospital and to identify any distinctive characteristics of such isolates. We characterized a collection of 303 pharyngeal S. pyogenes collected between 2002 and 2008. One-hundred and one were isolated from scarlet fever patients and 202 were associated to a diagnosis of tonsillo-pharyngitis. Isolates were characterized by T and emm typing, pulsed field gel electrophoresis profiling and superantigen gene profiling. The diversity of the scarlet fever isolates was lower than that of the pharyngitis isolates. Specific lineages of emm87, emm4 and emm3 were overrepresented in scarlet fever isolates but only 1 pulsed field gel electrophoresis major lineage was significantly associated with scarlet fever. Multivariate analysis indicated associations of ssa, speA and speC with scarlet fever. In nonoutbreak conditions, scarlet fever is caused by a number of distinct genetic lineages. The lower diversity of these isolates and the association with specific exotoxin genes indicates that some lineages are more prone to cause this presentation than others even in nonoutbreak conditions.

Effect of feeding glucose, fructose, and inulin on blood glucose and insulin concentrations in normal ponies and those predisposed to laminitis.

PubMed

Borer, K E; Bailey, S R; Menzies-Gow, N J; Harris, P A; Elliott, J

2012-09-01

Identification of ponies (Equus caballus) at increased risk of pasture-associated laminitis would aid in the prevention of the disease. Insulin resistance has been associated with laminitis and could be used to identify susceptible individuals. Insulin resistance may be diagnosed by feeding supplementary water-soluble carbohydrate (WSC) and measuring blood glucose and insulin concentrations. The aim of this study was to assess the glycemic and insulinemic responses of 7 normal (NP) and 5 previously laminitic (PLP), mixed breed, native UK ponies fed glucose, fructose, and inulin [1 g/(kg·d) for 3 d] or no supplementary WSC (control) in spring and fall after a 7-d adaptation to a pasture or hay diet. Blood samples were taken for 12 h after feeding on each day, and baseline and peak concentrations and area under the curve (AUC) for glucose and insulin were recorded. Linear mixed models were used for statistical analysis. Differences between PLP and NP groups were most marked after glucose feeding with differences in peak glucose (P = 0.02) and peak insulin (P = 0.016) concentrations. Season and diet adaptation also affected results. Peak concentrations of glucose and insulin occurred 2 to 4 h after WSC feeding. Peak insulin concentration was greater and more variable in fall, particularly in PLP adapted to fall pasture. Baseline glucose and insulin concentrations varied between individuals and with season and diet adaptation but were not greater in PLP than NP. Insulin AUC was greater in PLP than NP after feeding both glucose and fructose (P = 0.017), but there were no differences between PLP and NP in glucose AUC. Glycemic and insulinemic changes were less (P ≤ 0.05) after feeding fructose than glucose, although differences between PLP and NP were still evident. Minimal changes in glucose and insulin concentrations occurred after inulin feeding. Measurement of peak insulin 2 h after feeding of a single dose of glucose (1 g/kg) may be a simple and practical way to

Variation across mitochondrial gene trees provides evidence for systematic error: How much gene tree variation is biological?

PubMed

Richards, Emilie J; Brown, Jeremy M; Barley, Anthony J; Chong, Rebecca A; Thomson, Robert C

2018-02-19

The use of large genomic datasets in phylogenetics has highlighted extensive topological variation across genes. Much of this discordance is assumed to result from biological processes. However, variation among gene trees can also be a consequence of systematic error driven by poor model fit, and the relative importance of biological versus methodological factors in explaining gene tree variation is a major unresolved question. Using mitochondrial genomes to control for biological causes of gene tree variation, we estimate the extent of gene tree discordance driven by systematic error and employ posterior prediction to highlight the role of model fit in producing this discordance. We find that the amount of discordance among mitochondrial gene trees is similar to the amount of discordance found in other studies that assume only biological causes of variation. This similarity suggests that the role of systematic error in generating gene tree variation is underappreciated and critical evaluation of fit between assumed models and the data used for inference is important for the resolution of unresolved phylogenetic questions.

Mutation in the Monocarboxylate Transporter 12 Gene Affects Guanidinoacetate Excretion but Does Not Cause Glucosuria.

PubMed

Dhayat, Nasser; Simonin, Alexandre; Anderegg, Manuel; Pathare, Ganesh; Lüscher, Benjamin P; Deisl, Christine; Albano, Giuseppe; Mordasini, David; Hediger, Matthias A; Surbek, Daniel V; Vogt, Bruno; Sass, Jörn Oliver; Kloeckener-Gruissem, Barbara; Fuster, Daniel G

2016-05-01

A heterozygous mutation (c.643C>A; p.Q215X) in the monocarboxylate transporter 12-encoding gene MCT12 (also known as SLC16A12) that mediates creatine transport was recently identified as the cause of a syndrome with juvenile cataracts, microcornea, and glucosuria in a single family. Whereas the MCT12 mutation cosegregated with the eye phenotype, poor correlation with the glucosuria phenotype did not support a pathogenic role of the mutation in the kidney. Here, we examined MCT12 in the kidney and found that it resides on basolateral membranes of proximal tubules. Patients with MCT12 mutation exhibited reduced plasma levels and increased fractional excretion of guanidinoacetate, but normal creatine levels, suggesting that MCT12 may function as a guanidinoacetate transporter in vivo However, functional studies in Xenopus oocytes revealed that MCT12 transports creatine but not its precursor, guanidinoacetate. Genetic analysis revealed a separate, undescribed heterozygous mutation (c.265G>A; p.A89T) in the sodium/glucose cotransporter 2-encoding gene SGLT2 (also known as SLC5A2) in the family that segregated with the renal glucosuria phenotype. When overexpressed in HEK293 cells, the mutant SGLT2 transporter did not efficiently translocate to the plasma membrane, and displayed greatly reduced transport activity. In summary, our data indicate that MCT12 functions as a basolateral exit pathway for creatine in the proximal tubule. Heterozygous mutation of MCT12 affects systemic levels and renal handling of guanidinoacetate, possibly through an indirect mechanism. Furthermore, our data reveal a digenic syndrome in the index family, with simultaneous MCT12 and SGLT2 mutation. Thus, glucosuria is not part of the MCT12 mutation syndrome. Copyright © 2016 by the American Society of Nephrology.

Lithium-Responsive Seizure-Like Hyperexcitability Is Caused by a Mutation in the Drosophila Voltage-Gated Sodium Channel Gene paralytic

PubMed Central

Kasuya, Junko; Ueda, Atsushi; Iyengar, Atulya; Wu, Chun-Fang

2016-01-01

Abstract Shudderer (Shu) is an X-linked dominant mutation in Drosophila melanogaster identified more than 40 years ago. A previous study showed that Shu caused spontaneous tremors and defects in reactive climbing behavior, and that these phenotypes were significantly suppressed when mutants were fed food containing lithium, a mood stabilizer used in the treatment of bipolar disorder (Williamson, 1982). This unique observation suggested that the Shu mutation affects genes involved in lithium-responsive neurobiological processes. In the present study, we identified Shu as a novel mutant allele of the voltage-gated sodium (Nav) channel gene paralytic (para). Given that hypomorphic para alleles and RNA interference–mediated para knockdown reduced the severity of Shu phenotypes, Shu was classified as a para hypermorphic allele. We also demonstrated that lithium could improve the behavioral abnormalities displayed by other Nav mutants, including a fly model of the human generalized epilepsy with febrile seizures plus. Our electrophysiological analysis of Shu showed that lithium treatment did not acutely suppress Nav channel activity, indicating that the rescue effect of lithium resulted from chronic physiological adjustments to this drug. Microarray analysis revealed that lithium significantly alters the expression of various genes in Shu, including those involved in innate immune responses, amino acid metabolism, and oxidation-reduction processes, raising the interesting possibility that lithium-induced modulation of these biological pathways may contribute to such adjustments. Overall, our findings demonstrate that Nav channel mutants in Drosophila are valuable genetic tools for elucidating the effects of lithium on the nervous system in the context of neurophysiology and behavior. PMID:27844061