Characterization of the complete mitochondrial genome sequence of wild yak (Bos mutus).

PubMed

Chunnian, Liang; Wu, Xiaoyun; Ding, Xuezhi; Wang, Hongbo; Guo, Xian; Chu, Min; Bao, Pengjia; Yan, Ping

2016-11-01

Wild yak is a special breed in China and it is regarded as an important genetic resource for sustainably developing the animal husbandry in Tibetan area and enriching region's biodiversity. The complete mitochondrial genome of wild yak (16,322 bp in length) displayed 37 typical animal mitochondrial genes and A + T-rich (61.01%), with an overall G + C content of only 38.99%. It contained a non-coding control region (D-loop), 13 protein-coding genes, two rRNA genes, and 22 tRNA genes. Most of the genes have ATG initiation codons, whereas ND2, ND3, and ND5 genes start with ATA and were encoded on H-strand. The gene order of wild yak mitogenome is identical to that observed in most other vertebrates. The complete mitochondrial genome sequence of wild yak reported here could provide valuable information for developing genetic markers and phylogenetic analysis in yak.

Fulminant neurological deterioration in a neonate with Leigh syndrome due to a maternally transmitted missense mutation in the mitochondrial ND3 gene.

PubMed

Leshinsky-Silver, E; Lev, D; Tzofi-Berman, Z; Cohen, S; Saada, A; Yanoov-Sharav, M; Gilad, E; Lerman-Sagie, T

2005-08-26

Leigh syndrome can result from both nuclear and mitochondrial DNA defects. Mutations in complex V genes of the respiratory chain were considered until recently as the most frequent cause for mitochondrial inherited Leigh syndrome, while gene defects in complex I were related to recessive Leigh syndrome. Recently few reports of mutations in the mitochondrial-encoded complex I subunit genes causing Leigh syndrome have been reported. We describe a 1-month-old baby who acutely deteriorated, with abrupt onset of brainstem dysfunction, due to basal ganglia lesions extending to the brainstem. A muscle biopsy demonstrated complex I deficiency. Subsequent analysis of the mitochondrial genome revealed a homoplastic T10191C mutation in the ND3 gene (in blood and muscle), resulting in a substitution of serine to proline. Hair root analysis revealed a 50% mutant load, reflecting heteroplasmy in early embryonic stages. The mutation was also detected in his mother (5%). Western blot analysis revealed a decrease of the 20 kDa subunit (likely ND6) and of the 30 kDa subunit (NDUFA9), which is probably due to instability attributed to the inability to form subcomplexes with ND3. This is the first description of infantile Leigh syndrome due to a maternally transmitted T10191C substitution in ND3 and not due to a de novo mutation. This mutation is age and tissue dependent and therefore may not be amenable to prenatal testing.

Genome features of Pseudomonas putida LS46, a novel polyhydroxyalkanoate producer and its comparison with other P. putida strains

PubMed Central

2014-01-01

A novel strain of Pseudomonas putida LS46 was isolated from wastewater on the basis of its ability to synthesize medium chain-length polyhydroxyalkanoates (mcl-PHAs). P.putida LS46 was differentiated from other P.putida strains on the basis of cpn60 (UT). The complete genome of P.putida LS46 was sequenced and annotated. Its chromosome is 5,86,2556 bp in size with GC ratio of 61.69. It is encoding 5316 genes, including 7 rRNA genes and 76 tRNA genes. Nucleotide sequence data of the complete P. putida LS46 genome was compared with nine other P. putida strains (KT2440, F1, BIRD-1, S16, ND6, DOT-T1E, UW4, W619 and GB-1) identified either as biocontrol agents or as bioremediation agents and isolated from different geographical region and different environment. BLASTn analysis of whole genome sequences of the ten P. putida strains revealed nucleotide sequence identities of 86.54 to 97.52%. P.putida genome arrangement was LS46 highly similar to P.putida BIRD1 and P.putida ND6 but was markedly different than P.putida DOT-T1E, P.putida UW4 and P.putida W619. Fatty acid biosynthesis (fab), fatty acid degradation (fad) and PHA synthesis genes were highly conserved among biocontrol and bioremediation P.putida strains. Six genes in pha operon of P. putida LS46 showed >98% homology at gene and proteins level. It appears that polyhydroxyalkanoate (PHA) synthesis is an intrinsic property of P. putida and was not affected by its geographic origin. However, all strains, including P. putida LS46, were different from one another on the basis of house keeping genes, and presence of plasmid, prophages, insertion sequence elements and genomic islands. While P. putida LS46 was not selected for plant growth promotion or bioremediation capacity, its genome also encoded genes for root colonization, pyoverdine synthesis, oxidative stress (present in other soil isolates), degradation of aromatic compounds, heavy metal resistance and nicotinic acid degradation, manganese (Mn II) oxidation. Genes for toluene or naphthalene degradation found in the genomes of P. putida F1, DOT-T1E, and ND6 were absent in the P. putida LS46 genome. Heavy metal resistant genes encoded by the P. putida W619 genome were also not present in the P. putida LS46 genome. Despite the overall similarity among genome of P.putida strains isolated for different applications and from different geographical location a number of differences were observed in genome arrangement, occurrence of transposon, genomic islands and prophage. It appears that P.putida strains had a common ancestor and by acquiring some specific genes by horizontal gene transfer it differed from other related strains. PMID:25401060

Metabolic Interfaces of Mercury Methylation Proteins in Desulfovibrio sp. ND132

NASA Astrophysics Data System (ADS)

Wall, J. D.; Bridou, R.; Smith, S. D.; Mok, K.; Widner, F.; Johs, A.; Parks, J.; Pierce, E. M.; Elias, D. A.; Gilmour, C. C.; Taga, M.

2015-12-01

Two genes necessary for microbial production of the neurotoxin methylmercury have been identified; hgcA encoding a corrinoid methyltransferase and hgcB, a ferredoxin-like protein. To date, all microbes possessing orthologs of these genes that have been tested are capable of methylating mercury; whereas, organisms lacking hgcA and hgcB are not. Also of interest is the observation that confirmed mercury-methylating microbes are all considered anaerobes although not members of a specific phylogenetic group. They are found scattered in the genomes of methanogens, Firmicutes, and Deltaproteobacteria. Methylation has not been demonstrated to provide protection of the microbes to mercury exposure. To determine the source of evolutionary pressure for acquisition and maintenance of these genes, we are seeking to understand whether there is a second function of the proteins. We are seeking evidence for the metabolic source(s) of the methyl group and for competing reactions. We have found that deletion of the metH gene encoding a tetrahydrofolate methyltransferase in Desulfovibrio sp. ND132 decreases the mercury methylation capacity by ca. 95%, consistent with an interpretation that this enzyme is involved in the pathway for the methyl group for HgcA. In addition, the corrinoid present in HgcA and the MetH of ND132 is strictly dependent on nicotinate nucleotide:5,6-dimethylbenzimidazole phosphoribosyltransferase encoded by the cobT gene, linking methionine biosynthesis with mercury methylation at a second level. Additional methyl transferases have not been found to be necessary for this function. While earlier evidence was provided for an involvement of the CO dehydrogenase/acetylCoA synthase, this enzyme is not universally present in methylating strains unlike the pathway for methionine synthesis.

Mitochondrial genome of Pteronotus personatus (Chiroptera: Mormoopidae): comparison with selected bats and phylogenetic considerations.

PubMed

López-Wilchis, Ricardo; Del Río-Portilla, Miguel Ángel; Guevara-Chumacero, Luis Manuel

2017-02-01

We described the complete mitochondrial genome (mitogenome) of the Wagner's mustached bat, Pteronotus personatus, a species belonging to the family Mormoopidae, and compared it with other published mitogenomes of bats (Chiroptera). The mitogenome of P. personatus was 16,570 bp long and contained a typically conserved structure including 13 protein-coding genes, 22 transfer RNA genes, two ribosomal RNA genes, and one control region (D-loop). Most of the genes were encoded on the H-strand, except for eight tRNA and the ND6 genes. The order of protein-coding and rRNA genes was highly conserved in all mitogenomes. All protein-coding genes started with an ATG codon, except for ND2, ND3, and ND5, which initiated with ATA, and terminated with the typical stop codon TAA/TAG or the codon AGA. Phylogenetic trees constructed using Maximum Parsimony, Maximum Likelihood, and Bayesian inference methods showed an identical topology and indicated the monophyly of different families of bats (Mormoopidae, Phyllostomidae, Vespertilionidae, Rhinolophidae, and Pteropopidae) and the existence of two major clades corresponding to the suborders Yangochiroptera and Yinpterochiroptera. The mitogenome sequence provided here will be useful for further phylogenetic analyses and population genetic studies in mormoopid bats.

The complete mitochondrial genome of Glaucidium brodiei (Strigiformes: Strigidae).

PubMed

Sun, Xiaonan; Zhou, Wenliang; Sun, Zhonglou; Qian, Lifu; Zhang, Yanan; Pan, Tao; Zhang, Baowei

2016-07-01

In this paper, the complete mitochondrial genome of Glaucidium brodiei is sequenced and reported for the first time. The mitochondrial genome is a circular molecule of 17,318 bp in length, consisting of 13 protein-coding genes (PCGs), 22 transfer RNA genes, 2 ribosomal RNA genes and a control region. Overall base composition of the complete mitochondrial DNA is A (29.9%), G (14.1%), C (32.1%) and T (23.9%), the percentage of A and T (53.8%) is slightly higher than G and C (46.2%). All the genes in G. brodiei are distributed on the H-strand, except for the ND6 subunit gene and nine tRNA genes, which are encoded on the L-strand.

Leigh disease presenting in utero due to a novel missense mutation in the mitochondrial DNA-ND3.

PubMed

Leshinsky-Silver, Esther; Lev, Dorit; Malinger, Gustavo; Shapira, Daniel; Cohen, Sarit; Lerman-Sagie, Tally; Saada, Ann

2010-05-01

Leigh syndrome can be caused by defects in both nuclear and mitochondrial genes involved in energy metabolism. Recently, an increasing number of mutations in mitochondrial DNA encoding regions, especially in NADH dehydrogenase (respiratory chain complex I) subunits, have been reported as causative of early onset Leigh syndrome. We describe a patient whose fetal brain ultrasound demonstrated periventricular pseudocyst suggestive of a possible mitochondrial disorder who presented postnatally with Leigh syndrome. A muscle biopsy demonstrated a partial decrease in complex I and pyruvate dehydrogenase (PDH-E1 alpha) activity. Sequencing of the PDH-E1 alpha gene did not reveal any mutation. Sequencing of the mtDNA revealed a novel heteroplasmic G10254A (D66N) mutation in the ND3 gene. This change results in a substitution of aspartic acid to asparagine in a highly conserved domain of the ND3 subunit. The mutation could not be detected in the mother's blood or urine sediment. Blue native gel electrophoresis of muscle mitochondria revealed a normal size, albeit a decreased level of complex I. The G10254A substitution in the mtDNA-ND3 gene is another cause of maternally inherited Leigh syndrome. This case demonstrates that periventricular pseudocysts may be the initial in utero presentation in patients with mitochondrial disorders. We emphasize the importance of screening the mtDNA in pediatric patients as the first step in molecular diagnosis of Leigh syndrome. (c) 2010 Elsevier Inc. All rights reserved.

The complete mitochondrial genome of black-footed ferret, Mustela nigripes (Mustela, Mustelinae).

PubMed

Zhao, Ren-Bin; Zhou, Chao-Yang; Lu, Zhi-Xiang; Hu, Peng; Liu, Jian-Qiong; Tan, Wei-Wei; Yang, Tong-Hua

2016-05-01

In this study, the complete mitochondrial genome sequence of black-footed ferret, Mustela nigripes, is determined for the first time. This mitogenome is 16,556 bp in length and contains 13 protein-coding genes, 22 transfer RNA genes, 2 ribosomal RNA genes, and 1 control region (D-loop). The overall base composition is A (32.9%), C (26.1%), G (13.8%), and T (27.2%), so the percentage of A and T (60.1%) is higher than that of G and C. Most of the genes are encoded on H-strand, except for the ND6 subunit gene and six tRNA genes. The complete mitochondrial genome sequence reported here would be useful for further phylogenetic analysis and conservation genetic studies in M. nigripes.

Mitochondrial DNA sequence data reveals association of haplogroup U with psychosis in bipolar disorder.

PubMed

Frye, Mark A; Ryu, Euijung; Nassan, Malik; Jenkins, Gregory D; Andreazza, Ana C; Evans, Jared M; McElroy, Susan L; Oglesbee, Devin; Highsmith, W Edward; Biernacka, Joanna M

2017-01-01

Converging genetic, postmortem gene-expression, cellular, and neuroimaging data implicate mitochondrial dysfunction in bipolar disorder. This study was conducted to investigate whether mitochondrial DNA (mtDNA) haplogroups and single nucleotide variants (SNVs) are associated with sub-phenotypes of bipolar disorder. MtDNA from 224 patients with Bipolar I disorder (BPI) was sequenced, and association of sequence variations with 3 sub-phenotypes (psychosis, rapid cycling, and adolescent illness onset) was evaluated. Gene-level tests were performed to evaluate overall burden of minor alleles for each phenotype. The haplogroup U was associated with a higher risk of psychosis. Secondary analyses of SNVs provided nominal evidence for association of psychosis with variants in the tRNA, ND4 and ND5 genes. The association of psychosis with ND4 (gene that encodes NADH dehydrogenase 4) was further supported by gene-level analysis. Preliminary analysis of mtDNA sequence data suggests a higher risk of psychosis with the U haplogroup and variation in the ND4 gene implicated in electron transport chain energy regulation. Further investigation of the functional consequences of this mtDNA variation is encouraged. Copyright © 2016. Published by Elsevier Ltd.

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

Leshinsky-Silver, E.; Mitochondrial Disease Center, Wolfson Medical Center, Holon; E-mail: leshinsky@wolfson.health.gov.il

Leigh syndrome can result from both nuclear and mitochondrial DNA defects. Mutations in complex V genes of the respiratory chain were considered until recently as the most frequent cause for mitochondrial inherited Leigh syndrome, while gene defects in complex I were related to recessive Leigh syndrome. Recently few reports of mutations in the mitochondrial-encoded complex I subunit genes causing Leigh syndrome have been reported. We describe a 1-month-old baby who acutely deteriorated, with abrupt onset of brainstem dysfunction, due to basal ganglia lesions extending to the brainstem. A muscle biopsy demonstrated complex I deficiency. Subsequent analysis of the mitochondrial genomemore » revealed a homoplastic T10191C mutation in the ND3 gene (in blood and muscle), resulting in a substitution of serine to proline. Hair root analysis revealed a 50% mutant load, reflecting heteroplasmy in early embryonic stages. The mutation was also detected in his mother (5%). Western blot analysis revealed a decrease of the 20 kDa subunit (likely ND6) and of the 30 kDa subunit (NDUFA9), which is probably due to instability attributed to the inability to form subcomplexes with ND3. This is the first description of infantile Leigh syndrome due to a maternally transmitted T10191C substitution in ND3 and not due to a de novo mutation. This mutation is age and tissue dependent and therefore may not be amenable to prenatal testing.« less

Differences in respiration between dormant and non-dormant buds suggest the involvement of ABA in the development of endodormancy in grapevines.

PubMed

Parada, Francisca; Noriega, Ximena; Dantas, Débora; Bressan-Smith, Ricardo; Pérez, Francisco J

2016-08-20

Grapevine buds (Vitis vinifera L) enter endodormancy (ED) after perceiving the short-day (SD) photoperiod signal and undergo metabolic changes that allow them to survive the winter temperatures. In the present study, we observed an inverse relationship between the depth of ED and the respiration rate of grapevine buds. Moreover, the respiration of dormant and non-dormant buds differed in response to temperature and glucose, two stimuli that normally increase respiration in plant tissues. While respiration in non-dormant buds rose sharply in response to both stimuli, respiration in dormant buds was only slightly affected. This suggests that a metabolic inhibitor is present. Here, we propose that the plant hormone abscisic acid (ABA) could be this inhibitor. ABA inhibits respiration in non-dormant buds and represses the expression of respiratory genes, such as ALTERNATIVE NADH DEHYDROGENASE (VaND1, VvaND2), CYTOCHROME OXIDASE (VvCOX6) and CYTOCHROME C (VvCYTC), and induces the expression of VvSnRK1, a gene encoding a member of a highly conserved family of protein kinases that act as energy sensors and regulate gene expression in response to energy depletion. In addition to inducing ED the SD-photoperiod up-regulated the expression of VvNCED, a gene that encodes a key enzyme in ABA synthesis. Taken together, these results suggest that ABA through the mediation of VvSnRK1, could play a key role in the regulation of the metabolic changes accompanying the entry into ED of grapevine buds. Copyright © 2016 Elsevier GmbH. All rights reserved.

Molecular dissection of Norrie disease.

PubMed

Berger, W

1998-01-01

Norrie disease (ND) is a severe form of congenital blindness accompanied by mental retardation and/or deafness in at least one third of the patients. This article summarizes advances in the molecular genetic analysis of this disease during the last 13 years, including mapping and cloning of the human gene and the generation and characterization of a mouse model. Genetic linkage studies and physical mapping strategies have assigned the ND locus to the proximal short arm of the human X chromosome. The identification of chromosomal rearrangements in several patients, such as microdeletions, enabled the isolation of the ND gene by a positional cloning approach. Numerous point mutations in this gene have been identified in three distinct clinical entities: (1) ND, (2) familial and sporadic exudative vitreoretinopathy, and (3) retinopathy of prematurity. The gene encodes a relatively small protein, consisting of 133 amino acids. The function of the gene product is yet unknown, although homologies with known proteins and molecular modelling data suggest a role in the regulation of cell interaction or differentiation processes. A mouse model has been generated to shed more light on early pathogenic events involved in ND and allelic disorders. The mouse homologous protein is highly identical (94%) to the human polypeptide. The gene is expressed in the neuronal layers of the mouse retina, the cerebellum and olfactory epithelium. Mutant mice show snowflake-like opacities within the vitreous, dysgenesis of the ganglion cell layer and occasionally degeneration of photoreceptor cells. The mouse phenotype does not include phthisis bulbi and, overall, resembles a mild form of ND. Electrophysiological studies revealed a severely altered electroretinogram b-wave. These results suggest a primary defect in the inner neuronal layers of the retina. Defects in the vitreous and photoreceptor cell layer are most likely secondary effects. Further histological, functional and molecular studies of the mouse model are needed to provide additional information on disease associated pathways.

Herpes Simplex Virus Is Equipped with RNA- and Protein-Based Mechanisms To Repress Expression of ATRX, an Effector of Intrinsic Immunity

PubMed Central

Jurak, Igor; Silverstein, Leah B.; Sharma, Mayuri

2012-01-01

Intrinsic immunity is a first-line intracellular defense against virus infection, and viruses have evolved mechanisms to counteract it. During herpes simplex virus (HSV) infection, nuclear domain 10 (ND10) components localize adjacent to incoming viral genomes and generate a repressive environment for viral gene expression. Here, we found that the ND10 component, alpha-thalassemia/mental retardation syndrome X-linked (ATRX) protein, is predicted to be a target of HSV-1 miR-H1 and HSV-2 miR-H6. These microRNAs (miRNAs) share a seed sequence and are abundant during lytic infection. Mimics of both miRNAs could deplete endogenous ATRX, and an miR-H1 mimic could repress the expression of a reporter linked to the 3′ untranslated region of ATRX mRNA, identifying a cellular mRNA targeted by an HSV miRNA. Interestingly, ATRX protein and its mRNA were depleted in cells lytically infected with HSV, and ATRX protein was also depleted in cells infected with human cytomegalovirus. However, infection with an HSV-1 mutant lacking miR-H1 still resulted in ATRX depletion. This depletion was sensitive to a proteasome inhibitor and was largely ablated by a deletion of the gene encoding the immediate-early ICP0 protein. Additionally, a deletion of the gene encoding the tegument protein Vhs ablated most of the depletion of ATRX mRNA. Thus, HSV is equipped with multiple mechanisms to limit the expression of ATRX. As ATRX is implicated in repression of lytic viral gene expression, our results suggest roles for these different mechanisms during various phases of HSV infection. PMID:22787211

Herpes simplex virus is equipped with RNA- and protein-based mechanisms to repress expression of ATRX, an effector of intrinsic immunity.

PubMed

Jurak, Igor; Silverstein, Leah B; Sharma, Mayuri; Coen, Donald M

2012-09-01

Intrinsic immunity is a first-line intracellular defense against virus infection, and viruses have evolved mechanisms to counteract it. During herpes simplex virus (HSV) infection, nuclear domain 10 (ND10) components localize adjacent to incoming viral genomes and generate a repressive environment for viral gene expression. Here, we found that the ND10 component, alpha-thalassemia/mental retardation syndrome X-linked (ATRX) protein, is predicted to be a target of HSV-1 miR-H1 and HSV-2 miR-H6. These microRNAs (miRNAs) share a seed sequence and are abundant during lytic infection. Mimics of both miRNAs could deplete endogenous ATRX, and an miR-H1 mimic could repress the expression of a reporter linked to the 3' untranslated region of ATRX mRNA, identifying a cellular mRNA targeted by an HSV miRNA. Interestingly, ATRX protein and its mRNA were depleted in cells lytically infected with HSV, and ATRX protein was also depleted in cells infected with human cytomegalovirus. However, infection with an HSV-1 mutant lacking miR-H1 still resulted in ATRX depletion. This depletion was sensitive to a proteasome inhibitor and was largely ablated by a deletion of the gene encoding the immediate-early ICP0 protein. Additionally, a deletion of the gene encoding the tegument protein Vhs ablated most of the depletion of ATRX mRNA. Thus, HSV is equipped with multiple mechanisms to limit the expression of ATRX. As ATRX is implicated in repression of lytic viral gene expression, our results suggest roles for these different mechanisms during various phases of HSV infection.

Complete sequence and gene organization of the mitochondrial genome of Asio flammeus (Strigiformes, strigidae).

PubMed

Zhang, Yanan; Song, Tao; Pan, Tao; Sun, Xiaonan; Sun, Zhonglou; Qian, Lifu; Zhang, Baowei

2016-07-01

The complete sequence of the mitochondrial genome was determined for Asio flammeus, which is distributed widely in geography. The length of the complete mitochondrial genome was 18,966 bp, containing 2 rRNA genes, 22 tRNA genes, 13 protein-coding genes (PCGs), and 1 non-coding region (D-loop). All the genes were distributed on the H-strand, except for the ND6 subunit gene and eight tRNA genes which were encoded on the L-strand. The D-loop of A. flammeus contained many tandem repeats of varying lengths and repeat numbers. The molecular-based phylogeny showed that our species acted as the sister group to A. capensis and the supported Asio was the monophyletic group.

Maternal Betaine Supplementation during Gestation Enhances Expression of mtDNA-Encoded Genes through D-Loop DNA Hypomethylation in the Skeletal Muscle of Newborn Piglets.

PubMed

Jia, Yimin; Song, Haogang; Gao, Guichao; Cai, Demin; Yang, Xiaojing; Zhao, Ruqian

2015-11-25

Betaine has been widely used in animal and human nutrition to promote muscle growth and performance, yet it remains unknown whether maternal betaine supplementation during gestation affects the metabolic characteristics of neonatal skeletal muscles. In the present study, feeding sows with betaine-supplemented diets throughout gestation significantly upregulated the expression of mtDNA-encoded OXPHOS genes (p < 0.05), including COX1, COX2, and ND5, in the muscle of newborn piglets, which was associated with enhanced mitochondrial COX enzyme activity (p < 0.05). Concurrently, maternal betaine supplementation increased the plasma betaine concentration and muscle expression of methyl transfer enzymes (p < 0.05), BHMT and GNMT, in offspring piglets. Nevertheless, Dnmt3a was downregulated at the level of both mRNA and protein, which was associated with a hypomethylated mtDNA D-loop region (p < 0.05). These results suggest that maternal betaine supplementation during gestation enhances expression of mtDNA-encoded genes through D-loop DNA hypomethylation in the skeletal muscle of newborn piglets.

The complete mitochondrial genome and phylogenetic analysis of the giant panda (Ailuropoda melanoleuca).

PubMed

Peng, Rui; Zeng, Bo; Meng, Xiuxiang; Yue, Bisong; Zhang, Zhihe; Zou, Fangdong

2007-08-01

The complete mitochondrial genome sequence of the giant panda, Ailuropoda melanoleuca, was determined by the long and accurate polymerase chain reaction (LA-PCR) with conserved primers and primer walking sequence methods. The complete mitochondrial DNA is 16,805 nucleotides in length and contains two ribosomal RNA genes, 13 protein-coding genes, 22 transfer RNA genes and one control region. The total length of the 13 protein-coding genes is longer than the American black bear, brown bear and polar bear by 3 amino acids at the end of ND5 gene. The codon usage also followed the typical vertebrate pattern except for an unusual ATT start codon, which initiates the NADH dehydrogenase subunit 5 (ND5) gene. The molecular phylogenetic analysis was performed on the sequences of 12 concatenated heavy-strand encoded protein-coding genes, and suggested that the giant panda is most closely related to bears.

Partial Gene Cloning and Enzyme Structure Modeling of Exolevanase Fragment from Bacillus subtilis

NASA Astrophysics Data System (ADS)

Azhar, M.; Natalia, D.; Syukur, S.; Andriani, N.; Jamsari, J.

2018-04-01

Inulin hydrolysis thermophilic and thermotolerant bacteria are potential sources of inulin hydrolysis enzymes. Partial gene that encodes inulin hydrolysis enzymes had been isolated from Bacillus subtilis using polymerase chain reaction (PCR) method with the DPE.slFandDPE.eR degenerative primers. The partial gene was cloned into pGEM-T Easy vector with E. coli as host cells and analyzed using BLASTx, CrustalW2, and Phyre2 programs. Size of thepartial gene had been found539 bp that encoded 179aminoacid residues of protein fragment. The sequences of protein fragment was more similar to exolevanase than exoinulinase. The protein fragment had conserved motif FSGS, and specific hits GH32 β-fructosidase. It had three residues of active site and five residues of substrate binding. The active site on the protein fragment were D (1-WLNDP-5), D (125-FRDPK-129) and E (177-WEC-179). Substrate binding on the protein fragment were ND (1-WLNDP-5), Q (18-FYQY-21), FS (60-FSGS-63) RD (125-FRDPK-129) and E (177-WEC-179).

The sequence and organization of complete mitochondrial genome of the yellowfin tuna, Thunnus albacares (Bonnaterre, 1788).

PubMed

Pang, Jiaohui; Cheng, Qiqun; Sun, Dandan; Zhang, Heng; Jin, Shaofei

2016-09-01

Yellowfin tuna (Thunnus albacares) is one of the most important economic fishes around the world. In the present study, we determined the complete mitochondrial DNA sequence and organization of T. albacares. The entire mitochondrial genome is a circular-molecule of 16,528 bp in length, which encodes 37 genes in all. These genes comprise 13 protein-coding genes (ATP6 and 8, COI-III, Cytb, ND1-6 and 4 L), 22 transfer RNA genes (tRNAs), and 2 ribosomal RNA genes (12S and 16S rRNAs). The complete mitochondrial genome sequence of T. albacares can provide basic information for the studies on molecular taxonomy and conservation genetics of teleost fishes.

Positive and purifying selection in mitochondrial genomes of a bird with mitonuclear discordance.

PubMed

Morales, Hernán E; Pavlova, Alexandra; Joseph, Leo; Sunnucks, Paul

2015-06-01

Diversifying selection on metabolic pathways can reduce intraspecific gene flow and promote population divergence. An opportunity to explore this arises from mitonuclear discordance observed in an Australian bird Eopsaltria australis. Across >1500 km, nuclear differentiation is low and latitudinally structured by isolation by distance, whereas two highly divergent, parapatric mitochondrial lineages (>6.6% in ND2) show a discordant longitudinal geographic pattern and experience different climates. Vicariance, incomplete lineage sorting and sex-biased dispersal were shown earlier to be unlikely drivers of the mitonuclear discordance; instead, natural selection on a female-linked trait was the preferred hypothesis. Accordingly, here we tested for signals of positive, divergent selection on mitochondrial genes in E. australis. We used codon models and physicochemical profiles of amino acid replacements to analyse complete mitochondrial genomes of the two mitochondrial lineages in E. australis, its sister species Eopsaltria griseogularis, and outgroups. We found evidence of positive selection on at least five amino acids, encoded by genes of two oxidative phosphorylation pathway complexes NADH dehydrogenase (ND4 and ND4L) and cytochrome bc1 (cyt-b) against a background of widespread purifying selection on all mitochondrial genes. Three of these amino acid replacements were fixed in ND4 of the geographically most widespread E. australis lineage. The other two replacements were fixed in ND4L and cyt-b of the geographically more restricted E. australis lineage. We discuss whether this selection may reflect local environmental adaptation, a by-product of other selective processes, or genetic incompatibilities, and propose how these hypotheses can be tested in future. © 2015 John Wiley & Sons Ltd.

Two new mutations in the MT-TW gene leading to the disruption of the secondary structure of the tRNA(Trp) in patients with Leigh syndrome.

PubMed

Mkaouar-Rebai, Emna; Chamkha, Imen; Kammoun, Fatma; Kammoun, Thouraya; Aloulou, Hajer; Hachicha, Mongia; Triki, Chahnez; Fakhfakh, Faiza

2009-07-01

Leigh syndrome is a progressive neurodegenerative disorder occurring in infancy and childhood characterized in most cases by a psychomotor retardation, optic atrophy, ataxia, dystonia, failure to thrive, seizures and respiratory failure. In this study, we performed a systematic sequence analysis of mitochondrial genes associated with LS in Tunisian patients. We sequenced the encoded complex I units: ND2, ND3, ND4, ND5 and ND6 genes and the mitochondrial ATPase 6, tRNA(Val), tRNA(Leu(UUR)), tRNA(Trp) and tRNA(Lys) genes in 10 unrelated patients with Leigh syndrome. We revealed the presence of 34 reported polymorphisms, nine novel nucleotide variants and two new mutations (T5523G and A5559G) in the tested patients. These two mutations were localized in two conserved regions of the tRNA(Trp) and affect, respectively, the D-stem and the T-stem of the mitochondrial tRNA leading to a disruption of the secondary structure of this tRNA. SSP-PCR analysis showed that the T5523G and A5559G mutations were present with respective heteroplasmic rates of 66% and 43 %. We report here the first mutational screening of mitochondrial mutations in Tunisian patients with Leigh syndrome which described two novel mutations associated with this disorder.

Complete mitochondrial genome of the a rare subspecies of genus Bos, Tianzhu white yak from Tibetan area in China.

PubMed

E, Guangxin; Na, Ri-Su; Zhao, Yong-Ju; Gao, Hui-Jiang; An, Tian-Wu; Huang, Yong-Fu

2016-01-01

The population of domestic yak, Tianzhu white yak, from Tibetan area in China is considered as a rare Bos grunniens species. We first determined and annotated its complete mitochondrial genome. The mitogenome is 16,319 bp in length, consisting of 13 protein-coding genes, 22 transfer RNA (tRNA) genes, 2 ribosomal RNA (rRNA) genes and a control region. As in other mammals, most mitochondrial genes are encoded on the heavy strand, except for ND6 and eight tRNA genes, which are encoded on the light strand. Its overall base composition is A: 33.7%, T: 27.2%, C: 25.8% and G: 13.2%. The complete mitogenome of the new subspecies of Bos grunniens could provide an important data to further explore the taxonomic status of the subspecies.

The importance of biochemical and genetic findings in the diagnosis of atypical Norrie disease.

PubMed

Rodríguez-Muñoz, Ana; García-García, Gema; Menor, Francisco; Millán, José M; Tomás-Vila, Miguel; Jaijo, Teresa

2018-01-26

Norrie disease (ND) is a rare X-linked disorder characterized by bilateral congenital blindness. ND is caused by a mutation in the Norrie disease pseudoglioma (NDP) gene, which encodes a 133-amino acid protein called norrin. Intragenic deletions including NDP and adjacent genes have been identified in ND patients with a more severe neurologic phenotype. We report the biochemical, molecular, clinical and radiological features of two unrelated affected males with a deletion including NDP and MAO genes. Biochemical and genetic analyses were performed to understand the atypical phenotype and radiological findings. Biogenic amines in cerebrospinal fluid (CSF) were measured by high-performance liquid chromatography. The coding exons of NDP gene were amplified by polymerase chain reaction. Multiplex ligation-dependent probe amplification and chromosomal microarray were carried out on both affected males. Computed tomography and magnetic resonance imaging were performed on the two patients. In one patient, the serotonin and catecholamine metabolite levels in CSF were virtually undetectable. In both patients, genetic studies revealed microdeletions in the Xp11.3 region, involving the NDP, MAOA and MAOB genes. Radiological examination demonstrated brain and cerebellar atrophy. We suggest that alterations caused by MAO deficit may remain during the first years of life. Clinical phenotype, biochemical findings and neuroimaging can guide the genetic study in patients with atypical ND and help us to a better understanding of this disease.

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.

The Mitochondrial Genomes of Aquila fasciata and Buteo lagopus (Aves, Accipitriformes): Sequence, Structure and Phylogenetic Analyses

PubMed Central

Jiang, Lan; Chen, Juan; Wang, Ping; Ren, Qiongqiong; Yuan, Jian; Qian, Chaoju; Hua, Xinghong; Guo, Zhichun; Zhang, Lei; Yang, Jianke; Wang, Ying; Zhang, Qin; Ding, Hengwu; Bi, De; Zhang, Zongmeng; Wang, Qingqing; Chen, Dongsheng; Kan, Xianzhao

2015-01-01

The family Accipitridae is one of the largest groups of non-passerine birds, including 68 genera and 243 species globally distributed. In the present study, we determined the complete mitochondrial sequences of two species of accipitrid, namely Aquila fasciata and Buteo lagopus, and conducted a comparative mitogenome analysis across the family. The mitogenome length of A. fasciata and B. lagopus are 18,513 and 18,559 bp with an A + T content of 54.2% and 55.0%, respectively. For both the two accipitrid birds mtDNAs, obvious positive AT-skew and negative GC-skew biases were detected for all 12 PCGs encoded by the H strand, whereas the reverse was found in MT-ND6 encoded by the L strand. One extra nucleotide‘C’is present at the position 174 of MT-ND3 gene of A. fasciata, which is not observed at that of B. lagopus. Six conserved sequence boxes in the Domain II, named boxes F, E, D, C, CSBa, and CSBb, respectively, were recognized in the CRs of A. fasciata and B. lagopus. Rates and patterns of mitochondrial gene evolution within Accipitridae were also estimated. The highest dN/dS was detected for the MT-ATP8 gene (0.32493) among Accipitridae, while the lowest for the MT-CO1 gene (0.01415). Mitophylogenetic analysis supported the robust monophyly of Accipitriformes, and Cathartidae was basal to the balance of the order. Moreover, we performed phylogenetic analyses using two other data sets (two mitochondrial loci, and combined nuclear and mitochondrial loci). Our results indicate that the subfamily Aquilinae and all currently polytypic genera of this subfamily are monophyletic. These two novel mtDNA data will be useful in refining the phylogenetic relationships and evolutionary processes of Accipitriformes. PMID:26295156

The complete mitochondrial genome of Chinese green hydra, Hydra sinensis (Hydroida: Hydridae).

PubMed

Pan, Hong-Chun; Qian, Xiao-Cheng; Li, Ping; Li, Xiao-Fei; Wang, An-Tai

2014-02-01

The complete mitochondrial genome of Chinese green hydra, Hydra sinensis (Hydroida: Hydridae) is a linear molecule of 16,189 bp in length, containing 13 protein-coding genes, small and large subunit ribosomal RNAs, methionine and tryptophan transfer RNAs, a pseudogene consisting of a partial copy of COI and terminal sequences at two ends of the linear mitochondrial DNA. The A + T content of the overall base composition of H-strand is 77.2% (T: 41.7%; C: 10.9%; A: 35.5%; and G: 11.9%). COI and ND1 genes begin with GTG as start codon, while other 11 protein-coding genes start with a typical ATG initiation codon. COII, ATP8, ATP6, COIII, ND5, ND6, ND3, ND1, ND4 and COI genes are terminated with TAA as stop codon, ND4L ends with TAG, ND2 ends with TA and Cyt b ends with T.

The complete mitochondrial genome of Pomacea canaliculata (Gastropoda: Ampullariidae).

PubMed

Zhou, Xuming; Chen, Yu; Zhu, Shanliang; Xu, Haigen; Liu, Yan; Chen, Lian

2016-01-01

The mitochondrial genome of Pomacea canaliculata (Gastropoda: Ampullariidae) is the first complete mtDNA sequence reported in the genus Pomacea. The total length of mtDNA is 15,707 bp, which containing 13 protein-coding genes, 2 ribosomal RNAs, 22 transfer RNAs, and a 359 bp non-coding region. The A + T content of the overall base composition of H-strand is 71.7% (T: 41%, C: 12.7%, A: 30.7%, G: 15.6%). ATP6, ATP8, CO1, CO2, ND1-3, ND5, ND6, ND4L and Cyt b genes begin with ATG as start codon, CO3 and ND4 begin with ATA. ATP8, CO2-3, ND4L, ND2-6 and Cyt b genes are terminated with TAA as stop codon, ATP6, ND1, and CO1 end with TAG. A long non-coding region is found and a 23 bp repeat unit repeat 11 times in this region.

Juvenile Leigh syndrome, optic atrophy, ataxia, dystonia, and epilepsy due to T14487C mutation in the mtDNA-ND6 gene: a mitochondrial syndrome presenting from birth to adolescence.

PubMed

Leshinsky-Silver, Esther; Shuvalov, Ruslan; Inbar, Shani; Cohen, Sarit; Lev, Dorit; Lerman-Sagie, Tally

2011-04-01

An increasing number of reports describe mutations in mitochondrial DNA coding regions, especially in mitochondrial DNA- encoded nicotinamide adenine dinucleotide dehydrogenase subunit genes of the respiratory chain complex I, as causing early-onset Leigh syndrome. The authors report the molecular findings in a 24-year-old patient with juvenile-onset Leigh syndrome presenting with optic atrophy, ataxia dystonia, and epilepsy. A brain magnetic resonance imaging revealed bilateral basal ganglia and thalamic hypointensities, and a magnetic resonance spectroscopy revealed an increased lactate peak. The authors identified a T14487C change causing M63V substitution in the mitochondrial ND6 gene. The mutation was heteroplasmic in muscle and blood samples, with different mutation loads, and was absent in the patient's mother's urine and blood samples. They suggest that the T14487C mtDNA mutation should be analyzed in Leigh syndrome, presenting with optic atrophy, ataxia, dystonia, and epilepsy, regardless of age.

Mouse Y-linked Zfy1 and Zfy2 are expressed during the male-specific interphase between meiosis I and meiosis II and promote the 2nd meiotic division.

PubMed

Vernet, Nadège; Mahadevaiah, Shantha K; Yamauchi, Yasuhiro; Decarpentrie, Fanny; Mitchell, Michael J; Ward, Monika A; Burgoyne, Paul S

2014-06-01

Mouse Zfy1 and Zfy2 encode zinc finger transcription factors that map to the short arm of the Y chromosome (Yp). They have previously been shown to promote meiotic quality control during pachytene (Zfy1 and Zfy2) and at the first meiotic metaphase (Zfy2). However, from these previous studies additional roles for genes encoded on Yp during meiotic progression were inferred. In order to identify these genes and investigate their function in later stages of meiosis, we created three models with diminishing Yp and Zfy gene complements (but lacking the Y-long-arm). Since the Y-long-arm mediates pairing and exchange with the X via their pseudoautosomal regions (PARs) we added a minute PAR-bearing X chromosome derivative to enable formation of a sex bivalent, thus avoiding Zfy2-mediated meiotic metaphase I (MI) checkpoint responses to the unpaired (univalent) X chromosome. Using these models we obtained definitive evidence that genetic information on Yp promotes meiosis II, and by transgene addition identified Zfy1 and Zfy2 as the genes responsible. Zfy2 was substantially more effective and proved to have a much more potent transactivation domain than Zfy1. We previously established that only Zfy2 is required for the robust apoptotic elimination of MI spermatocytes in response to a univalent X; the finding that both genes potentiate meiosis II led us to ask whether there was de novo Zfy1 and Zfy2 transcription in the interphase between meiosis I and meiosis II, and this proved to be the case. X-encoded Zfx was also expressed at this stage and Zfx over-expression also potentiated meiosis II. An interphase between the meiotic divisions is male-specific and we previously hypothesised that this allows meiosis II critical X and Y gene reactivation following sex chromosome silencing in meiotic prophase. The interphase transcription and meiosis II function of Zfx, Zfy1 and Zfy2 validate this hypothesis.

Mouse Y-Linked Zfy1 and Zfy2 Are Expressed during the Male-Specific Interphase between Meiosis I and Meiosis II and Promote the 2nd Meiotic Division

PubMed Central

Vernet, Nadège; Mahadevaiah, Shantha K.; Yamauchi, Yasuhiro; Decarpentrie, Fanny; Mitchell, Michael J.; Ward, Monika A.; Burgoyne, Paul S.

2014-01-01

Mouse Zfy1 and Zfy2 encode zinc finger transcription factors that map to the short arm of the Y chromosome (Yp). They have previously been shown to promote meiotic quality control during pachytene (Zfy1 and Zfy2) and at the first meiotic metaphase (Zfy2). However, from these previous studies additional roles for genes encoded on Yp during meiotic progression were inferred. In order to identify these genes and investigate their function in later stages of meiosis, we created three models with diminishing Yp and Zfy gene complements (but lacking the Y-long-arm). Since the Y-long-arm mediates pairing and exchange with the X via their pseudoautosomal regions (PARs) we added a minute PAR-bearing X chromosome derivative to enable formation of a sex bivalent, thus avoiding Zfy2-mediated meiotic metaphase I (MI) checkpoint responses to the unpaired (univalent) X chromosome. Using these models we obtained definitive evidence that genetic information on Yp promotes meiosis II, and by transgene addition identified Zfy1 and Zfy2 as the genes responsible. Zfy2 was substantially more effective and proved to have a much more potent transactivation domain than Zfy1. We previously established that only Zfy2 is required for the robust apoptotic elimination of MI spermatocytes in response to a univalent X; the finding that both genes potentiate meiosis II led us to ask whether there was de novo Zfy1 and Zfy2 transcription in the interphase between meiosis I and meiosis II, and this proved to be the case. X-encoded Zfx was also expressed at this stage and Zfx over-expression also potentiated meiosis II. An interphase between the meiotic divisions is male-specific and we previously hypothesised that this allows meiosis II critical X and Y gene reactivation following sex chromosome silencing in meiotic prophase. The interphase transcription and meiosis II function of Zfx, Zfy1 and Zfy2 validate this hypothesis. PMID:24967676

Rapid identification of a novel complex I MT-ND3 m.10134C>A mutation in a Leigh syndrome patient.

PubMed

Miller, David K; Menezes, Minal J; Simons, Cas; Riley, Lisa G; Cooper, Sandra T; Grimmond, Sean M; Thorburn, David R; Christodoulou, John; Taft, Ryan J

2014-01-01

Leigh syndrome (LS) is a rare progressive multi-system neurodegenerative disorder, the genetics of which is frequently difficult to resolve. Rapid determination of the genetic etiology of LS in a 5-year-old girl facilitated inclusion in Edison Pharmaceutical's phase 2B clinical trial of EPI-743. SNP-arrays and high-coverage whole exome sequencing were performed on the proband, both parents and three unaffected siblings. Subsequent multi-tissue targeted high-depth mitochondrial sequencing was performed using custom long-range PCR amplicons. Tissue-specific mutant load was also assessed by qPCR. Complex I was interrogated by spectrophotometric enzyme assays and Western Blot. No putatively causal mutations were identified in nuclear-encoded genes. Analysis of low-coverage off-target mitochondrial reads revealed a previously unreported mitochondrial mutation in the proband in MT-ND3 (m.10134C>A, p.Q26K), a Complex I mitochondrial gene previously associated with LS. Targeted investigations demonstrated that this mutation was 1% heteroplasmic in the mother's blood and homoplasmic in the proband's blood, fibroblasts, liver and muscle. Enzyme assays revealed decreased Complex I activity. The identification of this novel LS MT-ND3 variant, the genomics of which was accomplished in less than 3.5 weeks, indicates that rapid genomic approaches may prove useful in time-sensitive cases with an unresolved genetic diagnosis.

Why Words are Hard for Adults with Developmental Language Impairments

PubMed Central

McGregor, Karla K.; Licandro, Ulla; Arenas, Richard; Eden, Nichole; Stiles, Derek; Bean, Allison; Walker, Elizabeth

2013-01-01

PURPOSE: To determine whether word learning problems associated with developmental language impairment (LI) reflect deficits in encoding or subsequent remembering of forms and meanings. METHOD: Sixty-nine 18-25-year-olds with LI or without (ND) took tests to measure learning of 16 word forms and meanings immediately after training (encoding) and 12-hours, 24-hours, and 1-week later (remembering). Half of the participants trained in the morning and half in the evening. RESULTS: At immediate posttest, those with LI performed more poorly on form and meaning than those with ND. Poor performance was more likely among those with more severe LI. The LI and ND groups demonstrated no difference in remembering word meanings over one week. In both groups, participants who trained in the evening, and therefore slept shortly after training, demonstrated greater gains in meaning recall than those who trained in the morning. In contrast, the LI-ND gap for word form recall widened over the week. CONCLUSIONS: Some adults with LI have encoding deficits that limit the addition of word forms and meanings to the lexicon. Similarities and differences in patterns of remembering in the LI and ND groups motivate the hypothesis that consolidation of declarative memory is a strength for adults with LI. PMID:24023376

A variant of Leber hereditary optic neuropathy characterized by recovery of vision and by an unusual mitochondrial genetic etiology.

PubMed Central

Mackey, D; Howell, N

1992-01-01

The Tas2 and Vic2 Australian families are affected with a variant of Leber hereditary optic neuropathy (LHON). The risk of developing the optic neuropathy shows strict maternal inheritance, and the ophthalmological changes in affected family members are characteristic of LHON. However, in contrast to the common form of the disease, members of these two families show a high frequency of vision recovery. To ascertain the mitochondrial genetic etiology of the LHON in these families, both (a) the the nucleotide sequences of the seven mitochondrial genes encoding subunits of respiratory-chain complex I and (b) the mitochondrial cytochrome b gene were determined for representatives of both families. Neither family carries any of the previously identified primary mitochondrial LHON mutations: ND4/11778, ND1/3460, or ND1/4160. Instead, both LHON families carry multiple nucleotide changes in the mitochondrial complex I genes, which produce conservative amino acid changes. From the available sequence data, it is inferred that the Vic2 and Tas2 LHON families are phylogenetically related to each other and to a cluster of LHON families in which mutations in the mitochondrial cytochrome b gene have been hypothesized to play a primary etiological role. However, sequencing analysis establishes that the Vic2 and Tas2 LHON families do not carry these cytochrome b mutations. There are two hypotheses to account for the unusual mitochondrial genetic etiology of the LHON in the Tas2 and Vic2 LHON families. One possibility is that there is a primary LHON mutation within the mitochondrial genome but that it is at a site that was not included in the sequencing analyses. Alternatively, the disease in these families may result from the cumulative effects of multiple secondary LHON mutations that have less severe phenotypic consequences. PMID:1463007

A variant of Leber hereditary optic neuropathy characterized by recovery of vision and by an unusual mitochondrial genetic etiology

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

Mackey, D.; Howell, N.

1992-12-01

The Tas2 and Vic2 Australian families are affected with a variant of Leber hereditary optic neuropathy (LHON). The risk of developing the optic neuropathy shows strict maternal inheritance, and the opthalmological changes in affected family members are characteristic of LHON. However, in contrast to the common form of the disease, members of these two families show a high frequency of vision recovery. To ascertain the mitochondrial genetic etiology of the LHON in these families, both (a) the nucleotide sequences of the seven mitochondrial genes encoding subunits of respiratory-chain complex I and (b) the mitochondrial cytochrome b gene were determined formore » representatives of both families. Neither family carries any of the previously identified primary mitochondrial LHON mutations: ND4/11778, ND1/3460, or ND1/4160. Instead, both LHON families carry multiple nucleotide changes in the mitochondrial complex I genes, which produce conservative amino acid changes. From the available sequence data, it is inferred that the Vic2 and Tas2 LHON families are phylogenetically related to each other and to a cluster of LHON families in which mutations in the mitochondrial cytochrome b gene have been hypothesized to play a primary etiological role. However, sequencing analysis establishes that the Vic2 and Tas2 LHON families do not carry these cytochrome b mutations. There are two hypotheses to account for the unusual mitochondrial genetic etiology of the LHON in the Tas2 and Vic2 LHON families. One possibility is that there is a primary LHON mutation within the mitochondrial genome but that it is at a site that was not included in the sequencing analyses. Alternatively, the disease in these families may result from the cumulative effects of multiple secondary LHON mutations that have less severe phenotypic consequences. 29 refs., 3 figs., 3 tabs.« less

The complete mitochondrial genome of the redeye mullet Liza haematocheila (Teleostei, Mugilidae).

PubMed

Chen, Jianhua; Li, Yinglei; Chen, Haigang; Yan, Binlun; Meng, Xueping

2015-01-01

The complete mitochondrial sequence of the redeye mullet Liza haematocheila has been determined. The circle genome is 16,822 bp in size, and consists of 13 protein-coding genes, 2 rRNA genes, 22 tRNA genes and a control region. The gene order and composition of L. haematocheila was similar to that of most other teleosts. The base composition of H-strand is 26.42% (A), 26.38% (T), 16.72% (G) and 30.47% (C), with an AT content of 52.8%. All genes are encoded on the heavy strand with the exception of ND6 and eight tRNA genes. The mitochondrial genome of L. haematocheila presented will be in favor of resolving phylogenetic relationships within the family Scatophagidae and the Mugiliformes.

Complete mitochondrial genome of Palawan peacock-pheasant Polyplectron napoleonis (Galliformes, Phasianidae).

PubMed

Quach, Tommy; Brooks, Daniel M; Miranda, Hector C

2016-01-01

The complete mitochondrial genome of the Palawan peacock-pheasant Polyplectron napoleonis is 16,710 bp and contains 13 protein-coding genes, 2 rRNA genes, 22 tRNA genes and a control-region. All protein-coding genes use the standard ATG start codon, except for cox1 which has GTG start codon. Seven out of 13 PCGs have TAA stop codons, two have AGG (cox1 and nd6), and three PCGs (nd2, cox2 and nd4) have incomplete stop codon of just T- - nucleotide.

Homology-dependent Gene Silencing in Paramecium

PubMed Central

Ruiz, Françoise; Vayssié, Laurence; Klotz, Catherine; Sperling, Linda; Madeddu, Luisa

1998-01-01

Microinjection at high copy number of plasmids containing only the coding region of a gene into the Paramecium somatic macronucleus led to a marked reduction in the expression of the corresponding endogenous gene(s). The silencing effect, which is stably maintained throughout vegetative growth, has been observed for all Paramecium genes examined so far: a single-copy gene (ND7), as well as members of multigene families (centrin genes and trichocyst matrix protein genes) in which all closely related paralogous genes appeared to be affected. This phenomenon may be related to posttranscriptional gene silencing in transgenic plants and quelling in Neurospora and allows the efficient creation of specific mutant phenotypes thus providing a potentially powerful tool to study gene function in Paramecium. For the two multigene families that encode proteins that coassemble to build up complex subcellular structures the analysis presented herein provides the first experimental evidence that the members of these gene families are not functionally redundant. PMID:9529389

Codon usage and expression level of human mitochondrial 13 protein coding genes across six continents.

PubMed

Chakraborty, Supriyo; Uddin, Arif; Mazumder, Tarikul Huda; Choudhury, Monisha Nath; Malakar, Arup Kumar; Paul, Prosenjit; Halder, Binata; Deka, Himangshu; Mazumder, Gulshana Akthar; Barbhuiya, Riazul Ahmed; Barbhuiya, Masuk Ahmed; Devi, Warepam Jesmi

2017-12-02

The study of codon usage coupled with phylogenetic analysis is an important tool to understand the genetic and evolutionary relationship of a gene. The 13 protein coding genes of human mitochondria are involved in electron transport chain for the generation of energy currency (ATP). However, no work has yet been reported on the codon usage of the mitochondrial protein coding genes across six continents. To understand the patterns of codon usage in mitochondrial genes across six different continents, we used bioinformatic analyses to analyze the protein coding genes. The codon usage bias was low as revealed from high ENC value. Correlation between codon usage and GC3 suggested that all the codons ending with G/C were positively correlated with GC3 but vice versa for A/T ending codons with the exception of ND4L and ND5 genes. Neutrality plot revealed that for the genes ATP6, COI, COIII, CYB, ND4 and ND4L, natural selection might have played a major role while mutation pressure might have played a dominant role in the codon usage bias of ATP8, COII, ND1, ND2, ND3, ND5 and ND6 genes. Phylogenetic analysis indicated that evolutionary relationships in each of 13 protein coding genes of human mitochondria were different across six continents and further suggested that geographical distance was an important factor for the origin and evolution of 13 protein coding genes of human mitochondria. Copyright © 2017 Elsevier B.V. and Mitochondria Research Society. All rights reserved.

CRISPR-LbCpf1 prevents choroidal neovascularization in a mouse model of age-related macular degeneration.

PubMed

Koo, Taeyoung; Park, Sung Wook; Jo, Dong Hyun; Kim, Daesik; Kim, Jin Hyoung; Cho, Hee-Yeon; Kim, Jeungeun; Kim, Jeong Hun; Kim, Jin-Soo

2018-05-10

LbCpf1, derived from Lachnospiraceae bacterium ND2006, is a CRISPR RNA-guided endonuclease and holds promise for therapeutic applications. Here we show that LbCpf1 can be used for therapeutic gene editing in a mouse model of age-related macular degeneration (AMD). The intravitreal delivery of LbCpf1, targeted to two angiogenesis-associated genes encoding vascular endothelial growth factor A (Vegfa) and hypoxia inducing factor 1a (Hif1a), using adeno-associated virus, led to efficient gene disruption with no apparent off-target effects in the retina and retinal pigment epithelium (RPE) cells. Importantly, LbCpf1 targeted to Vegfa or Hif1a in RPE cells reduced the area of laser-induced choroidal neovascularization as efficiently as aflibercept, an anti-VEGF drug currently used in the clinic, without inducing cone dysfunction. Unlike aflibercept, LbCpf1 targeted to Vegfa or Hif1a achieved a long-term therapeutic effect on CNV, potentially avoiding repetitive injections. Taken together, these results indicate that LbCpf1-mediated in vivo genome editing to ablate pathologic angiogenesis provides an effective strategy for the treatment of AMD and other neovascularization-associated diseases.

Human AZU-1 gene, variants thereof and expressed gene products

DOEpatents

Chen, Huei-Mei; Bissell, Mina

2004-06-22

A human AZU-1 gene, mutants, variants and fragments thereof. Protein products encoded by the AZU-1 gene and homologs encoded by the variants of AZU-1 gene acting as tumor suppressors or markers of malignancy progression and tumorigenicity reversion. Identification, isolation and characterization of AZU-1 and AZU-2 genes localized to a tumor suppressive locus at chromosome 10q26, highly expressed in nonmalignant and premalignant cells derived from a human breast tumor progression model. A recombinant full length protein sequences encoded by the AZU-1 gene and nucleotide sequences of AZU-1 and AZU-2 genes and variant and fragments thereof. Monoclonal or polyclonal antibodies specific to AZU-1, AZU-2 encoded protein and to AZU-1, or AZU-2 encoded protein homologs.

Polyglycerol-functionalized nanodiamond as a platform for gene delivery: Derivatization, characterization, and hybridization with DNA

PubMed Central

Zhao, Li; Nakae, Yuki; Qin, Hongmei; Ito, Tadamasa; Kimura, Takahide; Kojima, Hideto; Chan, Lawrence

2014-01-01

Summary A gene vector consisting of nanodiamond, polyglycerol, and basic polypeptide (ND-PG-BPP) has been designed, synthesized, and characterized. The ND-PG-BPP was synthesized by PG functionalization of ND through ring-opening polymerization of glycidol on the ND surface, multistep organic transformations (–OH → –OTs (tosylate) → –N3) in the PG layer, and click conjugation of the basic polypeptides (Arg8, Lys8 or His8) terminated with propargyl glycine. The ND-PG-BPP exhibited good dispersibility in water (>1.0 mg/mL) and positive zeta potential ranging from +14.2 mV to +44.1 mV at neutral pH in Milli-Q water. It was confirmed by gel retardation assay that ND-PG-Arg8 and ND-PG-Lys8 with higher zeta potential hybridized with plasmid DNA (pDNA) through electrostatic attraction, making them promising as nonviral vectors for gene delivery. PMID:24778723

Novel genomic rearrangements mediated by multiple genetic elements in Streptococcus pyogenes M23ND confer potential for evolutionary persistence

PubMed Central

Bao, Yun-Juan; Liang, Zhong; Mayfield, Jeffrey A.; McShan, William M.; Lee, Shaun W.; Ploplis, Victoria A.; Castellino, Francis J.

2016-01-01

Symmetric genomic rearrangements around replication axes in genomes are commonly observed in prokaryotic genomes, including Group A Streptococcus (GAS). However, asymmetric rearrangements are rare. Our previous studies showed that the hypervirulent invasive GAS strain, M23ND, containing an inactivated transcriptional regulator system, covRS, exhibits unique extensive asymmetric rearrangements, which reconstructed a genomic structure distinct from other GAS genomes. In the current investigation, we identified the rearrangement events and examined the genetic consequences and evolutionary implications underlying the rearrangements. By comparison with a close phylogenetic relative, M18-MGAS8232, we propose a molecular model wherein a series of asymmetric rearrangements have occurred in M23ND, involving translocations, inversions and integrations mediated by multiple factors, viz., rRNA-comX (factor for late competence), transposons and phage-encoded gene segments. Assessments of the cumulative gene orientations and GC skews reveal that the asymmetric genomic rearrangements did not affect the general genomic integrity of the organism. However, functional distributions reveal re-clustering of a broad set of CovRS-regulated actively transcribed genes, including virulence factors and metabolic genes, to the same leading strand, with high confidence (p-value ~10−10). The re-clustering of the genes suggests a potential selection advantage for the spatial proximity to the transcription complexes, which may contain the global transcriptional regulator, CovRS, and other RNA polymerases. Their proximities allow for efficient transcription of the genes required for growth, virulence and persistence. A new paradigm of survival strategies of GAS strains is provided through multiple genomic rearrangements, while, at the same time, maintaining genomic integrity. PMID:27329479

Technical adequacy of bisulfite sequencing and pyrosequencing for detection of mitochondrial DNA methylation: Sources and avoidance of false-positive detection.

PubMed

Owa, Chie; Poulin, Matthew; Yan, Liying; Shioda, Toshi

2018-01-01

The existence of cytosine methylation in mammalian mitochondrial DNA (mtDNA) is a controversial subject. Because detection of DNA methylation depends on resistance of 5'-modified cytosines to bisulfite-catalyzed conversion to uracil, examined parameters that affect technical adequacy of mtDNA methylation analysis. Negative control amplicons (NCAs) devoid of cytosine methylation were amplified to cover the entire human or mouse mtDNA by long-range PCR. When the pyrosequencing template amplicons were gel-purified after bisulfite conversion, bisulfite pyrosequencing of NCAs did not detect significant levels of bisulfite-resistant cytosines (brCs) at ND1 (7 CpG sites) or CYTB (8 CpG sites) genes (CI95 = 0%-0.94%); without gel-purification, significant false-positive brCs were detected from NCAs (CI95 = 4.2%-6.8%). Bisulfite pyrosequencing of highly purified, linearized mtDNA isolated from human iPS cells or mouse liver detected significant brCs (~30%) in human ND1 gene when the sequencing primer was not selective in bisulfite-converted and unconverted templates. However, repeated experiments using a sequencing primer selective in bisulfite-converted templates almost completely (< 0.8%) suppressed brC detection, supporting the false-positive nature of brCs detected using the non-selective primer. Bisulfite-seq deep sequencing of linearized, gel-purified human mtDNA detected 9.4%-14.8% brCs for 9 CpG sites in ND1 gene. However, because all these brCs were associated with adjacent non-CpG brCs showing the same degrees of bisulfite resistance, DNA methylation in this mtDNA-encoded gene was not confirmed. Without linearization, data generated by bisulfite pyrosequencing or deep sequencing of purified mtDNA templates did not pass the quality control criteria. Shotgun bisulfite sequencing of human mtDNA detected extremely low levels of CpG methylation (<0.65%) over non-CpG methylation (<0.55%). Taken together, our study demonstrates that adequacy of mtDNA methylation analysis using methods dependent on bisulfite conversion needs to be established for each experiment, taking effects of incomplete bisulfite conversion and template impurity or topology into consideration.

Nucleotide sequences of two genomic DNAs encoding peroxidase of Arabidopsis thaliana.

PubMed

Intapruk, C; Higashimura, N; Yamamoto, K; Okada, N; Shinmyo, A; Takano, M

1991-02-15

The peroxidase (EC 1.11.1.7)-encoding gene of Arabidopsis thaliana was screened from a genomic library using a cDNA encoding a neutral isozyme of horseradish, Armoracia rusticana, peroxidase (HRP) as a probe, and two positive clones were isolated. From the comparison with the sequences of the HRP-encoding genes, we concluded that two clones contained peroxidase-encoding genes, and they were named prxCa and prxEa. Both genes consisted of four exons and three introns; the introns had consensus nucleotides, GT and AG, at the 5' and 3' ends, respectively. The lengths of each putative exon of the prxEa gene were the same as those of the HRP-basic-isozyme-encoding gene, prxC3, and coded for 349 amino acids (aa) with a sequence homology of 89% to that encoded by prxC3. The prxCa gene was very close to the HRP-neutral-isozyme-encoding gene, prxC1b, and coded for 354 aa with 91% homology to that encoded by prxC1b. The aa sequence homology was 64% between the two peroxidases encoded by prxCa and prxEa.

Degradation of Benzene by Pseudomonas veronii 1YdBTEX2 and 1YB2 Is Catalyzed by Enzymes Encoded in Distinct Catabolism Gene Clusters.

PubMed

de Lima-Morales, Daiana; Chaves-Moreno, Diego; Wos-Oxley, Melissa L; Jáuregui, Ruy; Vilchez-Vargas, Ramiro; Pieper, Dietmar H

2016-01-01

Pseudomonas veronii 1YdBTEX2, a benzene and toluene degrader, and Pseudomonas veronii 1YB2, a benzene degrader, have previously been shown to be key players in a benzene-contaminated site. These strains harbor unique catabolic pathways for the degradation of benzene comprising a gene cluster encoding an isopropylbenzene dioxygenase where genes encoding downstream enzymes were interrupted by stop codons. Extradiol dioxygenases were recruited from gene clusters comprising genes encoding a 2-hydroxymuconic semialdehyde dehydrogenase necessary for benzene degradation but typically absent from isopropylbenzene dioxygenase-encoding gene clusters. The benzene dihydrodiol dehydrogenase-encoding gene was not clustered with any other aromatic degradation genes, and the encoded protein was only distantly related to dehydrogenases of aromatic degradation pathways. The involvement of the different gene clusters in the degradation pathways was suggested by real-time quantitative reverse transcription PCR. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

AMPD1 gene polymorphism and the vasodilatory response to ischemia.

PubMed

Hand, Brian D; Roth, Stephen M; Roltsch, Mark H; Park, Jung-Jun; Kostek, Matthew C; Ferrell, Robert E; Brown, Michael D

2006-09-05

Peripheral vasculature resistance can play an important role in affecting blood pressure and the development of cardiovascular disease. A better understanding of the genes that encode vasodilators, such as adenosine, will provide insight into the mechanisms underlying cardiovascular disease. We tested whether the adenosine monophosphate deaminase-1 (AMPD1) C34T gene polymorphism was associated with the vasodilatory response to ischemia in Caucasian females aged 18-35 years. Blood samples (n = 58) were analyzed for the C34T variant and resulted in the following genotype groups: CC (n = 45) and CT (n = 13). Mean blood pressure (MBP), heart rate, and forearm blood flow (FBF) measured by venous occlusion plethysmography were measured at baseline and at 1 (peak FBF), 2 and 3 min of vasodilation during reactive hyperemia following 5 min of arm ischemia. To control for interindividual variability in baseline FBF and forearm vascular resistance (FVR) the percent change in FBF and FVR were calculated for each min. The percent decrease in FVR was significantly greater in the CT compared to the CC genotype group (-40+/-4% vs. -24+/-3%, P = 0.01) during the 2nd min of reactive hyperemia. The percent increase in FBF tended to be greater in the CT compared to the CC genotype group (+69+/-9% vs. +42+/-9%, P = 0.07) during the 2nd min of reactive hyperemia after adjustment for percent body fat. Consistent with previous findings of increased production of adenosine during exercise in individuals carrying a T allele, our findings suggest that the AMPD1 C34T polymorphism is associated with vasodilatory response to ischemia in the peripheral vasculature because individuals with the T allele had a greater vasodilatory response to ischemia.

Evidence for Sexual Reproduction: Identification, Frequency, and Spatial Distribution of Venturia effusa (Pecan Scab) Mating Type Idiomorphs.

PubMed

Young, Carolyn A; Bock, Clive H; Charlton, Nikki D; Mattupalli, Chakradhar; Krom, Nick; Bowen, Joanna K; Templeton, Matthew; Plummer, Kim M; Wood, Bruce W

2018-05-10

Venturia effusa (syn. Fusicladium effusum), causal agent of pecan scab, is the most prevalent pathogen of pecan (Carya illinoinensis), causing severe yield losses in the southeastern United States. V. effusa is currently known only by its asexual (conidial) stage. However, the degree and distribution of genetic diversity observed within and among populations of V. effusa are typical of a sexually reproducing fungal pathogen, and comparable with other dothideomycetes with a known sexual stage, including the closely related apple scab pathogen, V. inaequalis. Using the mating type (MAT) idiomorphs from V. inaequalis, we identified a single MAT gene, MAT1-1-1, in a draft genome of V. effusa. The MAT1-1-1 locus is flanked by two conserved genes encoding a DNA lyase (APN2) and a hypothetical protein. The MAT locus spanning the flanking genes was amplified and sequenced from a subset of 14 isolates, of which 7 contained MAT1-1-1 and the remaining samples contained MAT1-2-1. A multiplex polymerase chain reaction screen was developed to amplify MAT1-1-1, MAT1-2-1, and a conserved reference gene encoding β-tubulin, and used to screen 784 monoconidial isolates of V. effusa collected from 11 populations of pecan across the southeastern United States. A hierarchical sampling protocol representing region, orchard, and tree allowed for analysis of MAT structure at different spatial scales. Analysis of this collection revealed the frequency of the MAT idiomorphs is in a 1:1 equilibrium of MAT1-1:MAT1-2. The apparent equilibrium of the MAT idiomorphs provides impetus for a renewed effort to search for the sexual stage of V. effusa. [Formula: see text] Copyright © 2018 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license .

The mitochondrial genome of Cethosia biblis (Drury) (Lepidoptera: Nymphalidae).

PubMed

Xin, Tianrong; Li, Lei; Yao, Chengyi; Wang, Yayu; Zou, Zhiwen; Wang, Jing; Xia, Bin

2016-07-01

We present the complete mitogenome of Cethosia biblis (Drury) (Lepidoptera: Nymphalidae) in this article. The mitogenome was a circle molecular consisting of 15,286 nucleotides, 37 genes, and an A + T-rich region. The order of 37 genes was typical of insect mitochondrial DNA sequences described to date. The overall base composition of the genome is A (37.41%), T (42.80%), C (11.87%), and G (7.91%) with an A + T-rich hallmark as that of other invertebrate mitochondrial genomes. The start codon was mainly ATA in most of the mitochondrial protein-coding genes such as ND2, COI, ATP8, ND3, ND5, ND4, ND6, and ND1, but COII, ATP6, COIII, ND4L, and Cob genes employing ATG. The stop codon was TAA in all the protein-coding genes. The A + T region is located between 12S rRNA and tRNA(M)(et). The phylogenetic relationships of Lepidoptera species were constructed based on the nucleotides sequences of 13 PCGs of mitogenomes using the neighbor-joining method. The molecular-based phylogeny supported the traditional morphological classification on relationships within Lepidoptera species.

Adaptation of the Mitochondrial Genome in Cephalopods: Enhancing Proton Translocation Channels and the Subunit Interactions

PubMed Central

Almeida, Daniela; Maldonado, Emanuel; Vasconcelos, Vitor; Antunes, Agostinho

2015-01-01

Mitochondrial protein-coding genes (mt genes) encode subunits forming complexes of crucial cellular pathways, including those involved in the vital process of oxidative phosphorylation (OXPHOS). Despite the vital role of the mitochondrial genome (mt genome) in the survival of organisms, little is known with respect to its adaptive implications within marine invertebrates. The molluscan Class Cephalopoda is represented by a marine group of species known to occupy contrasting environments ranging from the intertidal to the deep sea, having distinct metabolic requirements, varied body shapes and highly advanced visual and nervous systems that make them highly competitive and successful worldwide predators. Thus, cephalopods are valuable models for testing natural selection acting on their mitochondrial subunits (mt subunits). Here, we used concatenated mt genes from 17 fully sequenced mt genomes of diverse cephalopod species to generate a robust mitochondrial phylogeny for the Class Cephalopoda. We followed an integrative approach considering several branches of interest–covering cephalopods with distinct morphologies, metabolic rates and habitats–to identify sites under positive selection and localize them in the respective protein alignment and/or tridimensional structure of the mt subunits. Our results revealed significant adaptive variation in several mt subunits involved in the energy production pathway of cephalopods: ND5 and ND6 from Complex I, CYTB from Complex III, COX2 and COX3 from Complex IV, and in ATP8 from Complex V. Furthermore, we identified relevant sites involved in protein-interactions, lining proton translocation channels, as well as disease/deficiencies related sites in the aforementioned complexes. A particular case, revealed by this study, is the involvement of some positively selected sites, found in Octopoda lineage in lining proton translocation channels (site 74 from ND5) and in interactions between subunits (site 507 from ND5) of Complex I. PMID:26285039

Inheritance of the complete mitochondrial genomes Cyprinus capio furong(♀) × Cyprinus carpio var.singguonensis(♂).

PubMed

Peng, Huizhen; Liu, Qiaolin; Xiao, Tiaoyi

2016-09-01

In this study, 15 sets of primers were used to amplify contiguous, overlapping segments of the complete mitochondrial DNA (mtDNA) of C. capio furong(♀) × C. carpio var.singguonensis(♂) in order to characterize and compare their mitochondrial genomes. The total length of the mitochondrial genome was 16,581 bp and deposited in the GenBank with the accession number KP210473. The organization of the mitochondrial genomes contained 37 genes (13 protein-coding genes, 2 ribosomal RNA and 22 transfer RNAs) and a major non-coding control region which was similar to those reported mitochondrial genomes. Most genes were encoded on the H-strand, except for the ND6 and 8 tRNA genes, encoding on the L-strand. The nucleotide skewness for the coding strands of C. capio furong(♀) × C. carpio var.singguonensis(♂) (AT-skew = 0.12, GC-skew = -0.27) were biased toward T and G. The complete mitogenome may provide important date for the study of genetic mechanism of C. capio furong(♀) × C. carpio var.singguonensis(♂).

Encoding Deficits Impede Word Learning and Memory in Adults with Developmental Language Disorders

ERIC Educational Resources Information Center

McGregor, Karla K.; Gordon, Katherine; Eden, Nichole; Arbisi-Kelm, Tim; Oleson, Jacob

2017-01-01

Purpose: The aim of this study was to determine whether the word-learning challenges associated with developmental language disorder (DLD) result from encoding or retention deficits. Method In Study 1, 59 postsecondary students with DLD and 60 with normal development (ND) took the California Verbal Learning Test-Second Edition, Adult Version…

A complete mitochondrial genome sequence of the wild two-humped camel (Camelus bactrianus ferus): an evolutionary history of camelidae

PubMed Central

Cui, Peng; Ji, Rimutu; Ding, Feng; Qi, Dan; Gao, Hongwei; Meng, He; Yu, Jun; Hu, Songnian; Zhang, Heping

2007-01-01

Background The family Camelidae that evolved in North America during the Eocene survived with two distinct tribes, Camelini and Lamini. To investigate the evolutionary relationship between them and to further understand the evolutionary history of this family, we determined the complete mitochondrial genome sequence of the wild two-humped camel (Camelus bactrianus ferus), the only wild survivor of the Old World camel. Results The mitochondrial genome sequence (16,680 bp) from C. bactrianus ferus contains 13 protein-coding, two rRNA, and 22 tRNA genes as well as a typical control region; this basic structure is shared by all metazoan mitochondrial genomes. Its protein-coding region exhibits codon usage common to all mammals and possesses the three cryptic stop codons shared by all vertebrates. C. bactrianus ferus together with the rest of mammalian species do not share a triplet nucleotide insertion (GCC) that encodes a proline residue found only in the nd1 gene of the New World camelid Lama pacos. This lineage-specific insertion in the L. pacos mtDNA occurred after the split between the Old and New World camelids suggests that it may have functional implication since a proline insertion in a protein backbone usually alters protein conformation significantly, and nd1 gene has not been seen as polymorphic as the rest of ND family genes among camelids. Our phylogenetic study based on complete mitochondrial genomes excluding the control region suggested that the divergence of the two tribes may occur in the early Miocene; it is much earlier than what was deduced from the fossil record (11 million years). An evolutionary history reconstructed for the family Camelidae based on cytb sequences suggested that the split of bactrian camel and dromedary may have occurred in North America before the tribe Camelini migrated from North America to Asia. Conclusion Molecular clock analysis of complete mitochondrial genomes from C. bactrianus ferus and L. pacos suggested that the two tribes diverged from their common ancestor about 25 million years ago, much earlier than what was predicted based on fossil records. PMID:17640355

Genetic variants and early cigarette smoking and nicotine dependence phenotypes in adolescents.

PubMed

O'Loughlin, Jennifer; Sylvestre, Marie-Pierre; Labbe, Aurélie; Low, Nancy C; Roy-Gagnon, Marie-Hélène; Dugas, Erika N; Karp, Igor; Engert, James C

2014-01-01

While the heritability of cigarette smoking and nicotine dependence (ND) is well-documented, the contribution of specific genetic variants to specific phenotypes has not been closely examined. The objectives of this study were to test the associations between 321 tagging single-nucleotide polymorphisms (SNPs) that capture common genetic variation in 24 genes, and early smoking and ND phenotypes in novice adolescent smokers, and to assess if genetic predictors differ across these phenotypes. In a prospective study of 1294 adolescents aged 12-13 years recruited from ten Montreal-area secondary schools, 544 participants who had smoked at least once during the 7-8 year follow-up provided DNA. 321 single-nucleotide polymorphisms (SNPs) in 24 candidate genes were tested for an association with number of cigarettes smoked in the past 3 months, and with five ND phenotypes (a modified version of the Fagerstrom Tolerance Questionnaire, the ICD-10 and three clusters of ND symptoms representing withdrawal symptoms, use of nicotine for self-medication, and a general ND/craving symptom indicator). The pattern of SNP-gene associations differed across phenotypes. Sixteen SNPs in seven genes (ANKK1, CHRNA7, DDC, DRD2, COMT, OPRM1, SLC6A3 (also known as DAT1)) were associated with at least one phenotype with a p-value <0.01 using linear mixed models. After permutation and FDR adjustment, none of the associations remained statistically significant, although the p-values for the association between rs557748 in OPRM1 and the ND/craving and self-medication phenotypes were both 0.076. Because the genetic predictors differ, specific cigarette smoking and ND phenotypes should be distinguished in genetic studies in adolescents. Fifteen of the 16 top-ranked SNPs identified in this study were from loci involved in dopaminergic pathways (ANKK1/DRD2, DDC, COMT, OPRM1, and SLC6A3). Dopaminergic pathways may be salient during early smoking and the development of ND.

The complete mitochondrial genome of the Aluterus monoceros.

PubMed

Li, Wenshen; Zhang, Guoqing; Wen, Xin; Wang, Qian; Chen, Guohua

2016-07-01

The complete mitochondrial genome of Aluterus monoceros (A. monoceros) has been sequenced. The mitochondrial genome of A. monoceros is 16,429 bp in length, consisting of 22 tRNA genes, 2 rRNA genes, 13 protein-coding genes and a D-loop region (Gen Bank accession number KP637022). The base A + T of the mitochondrial genome is 63.25%, including 33.16% of A, 30.09% of T and 20.74% of C. Twelve protein-coding genes start with a standard ATG as the initiation codon, expect for the COXI, which begins with GTG. Some of the termination codons are incomplete T or TA, except for the ND1, COXI, ATP8, ND4L1, ND5 and ND6, which stop with TAA. Construction of phylogenetic trees based on the entire mitochondrial genome sequence of 14 Tetrodontiformes species constructed has suggested that A. monoceros has closer relationship with Acreichthys tomentosus and Monacanthus chinensis, and they constitute a sister group.

A novel Norrie disease pseudoglioma gene mutation, c.-1_2delAAT, responsible for Norrie disease in a Chinese family.

PubMed

Zhang, Xin-Yu; Jiang, Wei-Ying; Chen, Lu-Ming; Chen, Su-Qin

2013-01-01

To investigate the genetic findings and phenotypic characteristics of a Chinese family with Norrie disease (ND). Molecular genetic analysis and clinical examinations were performed on a Chinese family with ND. Mutations in the Norrie disease pseudoglioma (NDP) gene were detected by direct sequencing. Haplotypes were constructed and compared with the phenotypes in the family. Evolutionary comparisons and mutant open reading frame (ORF) prediction were also undertaken. Two family members with ocular manifestations were diagnosed with ND. No signs of sensorineural hearing loss were observed in either patient, while one of them showed signs of mild mental retardation. A novel heterozygous mutation in the NDP gene, c.-1_2delAAT, was detected in both patients. The mutation and the mutation bearing haplotype co-segregated with the ND phenotype in males and was transmitted from their mothers and/or grandmothers (II:2). The male without ND did not harbor the mutation. The mutation occurred at the highly conserved nucleotides. ORF finder predicted that the mutation would lead to the production of a truncated protein that lacks the first 11 N-terminal amino acids. A novel mutation, c.-1_2delAAT in the NDP gene, was identified in a Chinese family with ND. This mutation caused ND without obvious sensorineural hearing loss. Mental disorder was found in one but not the other patients. The clinical heterogeneity in the family indicated that other genetic variants and epigenetic factors may also play a role in the disease presentation.

A novel Norrie disease pseudoglioma gene mutation, c.-1_2delAAT, responsible for Norrie disease in a Chinese family

PubMed Central

Zhang, Xin-Yu; Jiang, Wei-Ying; Chen, Lu-Ming; Chen, Su-Qin

2013-01-01

AIM To investigate the genetic findings and phenotypic characteristics of a Chinese family with Norrie disease (ND). METHODS Molecular genetic analysis and clinical examinations were performed on a Chinese family with ND. Mutations in the Norrie disease pseudoglioma (NDP) gene were detected by direct sequencing. Haplotypes were constructed and compared with the phenotypes in the family. Evolutionary comparisons and mutant open reading frame (ORF) prediction were also undertaken. RESULTS Two family members with ocular manifestations were diagnosed with ND. No signs of sensorineural hearing loss were observed in either patient, while one of them showed signs of mild mental retardation. A novel heterozygous mutation in the NDP gene, c.-1_2delAAT, was detected in both patients. The mutation and the mutation bearing haplotype co-segregated with the ND phenotype in males and was transmitted from their mothers and/or grandmothers (II:2). The male without ND did not harbor the mutation. The mutation occurred at the highly conserved nucleotides. ORF finder predicted that the mutation would lead to the production of a truncated protein that lacks the first 11 N-terminal amino acids. CONCLUSION A novel mutation, c.-1_2delAAT in the NDP gene, was identified in a Chinese family with ND. This mutation caused ND without obvious sensorineural hearing loss. Mental disorder was found in one but not the other patients. The clinical heterogeneity in the family indicated that other genetic variants and epigenetic factors may also play a role in the disease presentation. PMID:24392318

The complete mitochondrial genome of Gobiobotia filifer (Teleostei, Cypriniformes: Cyprinidae).

PubMed

Li, Qiang; Liu, Ya; Zhou, Jian; Gong, Quan; Li, Hua; Lai, Jiansheng; Li, Lianman

2016-09-01

The Gobiobotia filifer is a small economic fish which distributes in the upstream of Yangtze River and its distributaries. For the environmental pollution and overfishing, its population declined drastically in recent decades, so it is essential to protect its resource. In this study, the complete mitochondrial genome sequence of G. filifer was determined with PCR technology, which contains 13 protein-coding genes, 22 tRNA genes, two rRNA genes, and a non-coding control region with the total length of 16,613 bp. The order and composition of genes were similar to most of the other teleost fish. Most of the genes were encoded on heavy strand, except for ND6 genes and eight tRNAs. Just like most other vertebrates, the bias of G and C has been found in different genes/regions. The complete mitochondrial genome sequence of G. filifer would contribute to better understand evolution of this lineage, population genetics, and will help administrative department to make rules and laws to protect this lineage.

The complete mitochondrial genome of Liobagrus marginatus (Teleostei, Siluriformes: Amblycipitidae).

PubMed

Li, Qiang; Du, Jun; Liu, Ya; Zhou, Jian; Ke, Hongyu; Liu, Chao; Liu, Guangxun

2014-04-01

The Liobagrus marginatus is an economic fish which distribute in the upstream of Yangtze river and its distributary. For its taste fresh, environmental pollution and overfishing, its population declined drastically and body miniaturization in recent decades, so it is essential to protect its resource. In this study, the complete mitochondrial genome sequence of Liobagrus marginatus was sequenced, which contains 22 tRNA genes, 13 protein-coding genes, 2 rRNA genes, and a non-coding control region with the total length of 16,497 bp. The gene arrangement and composition are similar to most of other fish. Most of the genes are encoded on heavy-strand, except for eight tRNA and ND6 genes. Just like most other vertebrates, the bias of G and C has been found in statistics results of different genes/regions. The complete mitochondrial genome sequence of Liobagrus marginatus would contribute to better understand population genetics, evolution of this lineage, and will help administrative departments to make rules and laws to protect it.

[Construction, expression and characterization of the fusion gene of super-antigen SEA and single chain Fv of the ND-1 monoclonal antibody against human colorectal cancer].

PubMed

Chen, Hang; Li, Li; Fang, Jin

2012-04-01

To construct and express the recombinant ND-1-scFv/SEA, a fusion protein of superantigen (staphylococcal enterotoxinA, SEA) and single-chain variable fragment of monoclonal antibody ND-1 against human clolorectal carcinoma, and to enhance the targeted killing effect of SEA. The expression of the fusion protein was induced in E.coli M15 by IPTG. Ni-NTA resin affinity chromatography was used to separate and purify the expressed product. The specific binding activity of the purified ND-1-scFv/SEA protein was examined by indirect immunofluorescence assay and the targeted-cytotoxicity was determined using MTT assay. The expressing vector of fusion gene ND-1scFv/SEA was constructed successfully. ND-1-scFv/SEA protein retained a high binding affinity to antigen-positive human colorectal cancer cell CCL-187 and had a stronger capability to activate PBMC and kill the target cells compared to SEA alone, with a killing rate of 91% at 4 μg/mL. ND-1-scFv/SEA fusion protein could specifically target colorectal cancer cell, enhance the activity of kill tumor cell and has potential applications in the targeted therapy of colorectal cancer.

The complete mitochondrial genome of Acanthosaura lepidogaster (Squamata: Agamidae).

PubMed

Yu, Xiu-Li; Du, Yu; Yao, Yun-Tao; Lin, Chi-Xian; Lin, Long-Hui

2017-03-01

In this paper, we report the complete mitochondrial genome of Acanthosaura lepidogaster (Squamata, Agamidae), which is a circular molecule of 16 899 bp in size and consists of 13 protein-coding genes, 2 ribosomal RNAs, 22 transfer RNAs, and a control region. The overall base composition is as follows: T (22.8%), C (30.5%), A (32.3%), and G (14.4%). We constructed a phylogeny that included for 10 species of Leiolepidinae lizards and one outgroup Leiocephalus personatus constructed in BEAST, based on 15 mitochondrial genes (12S, 16S, ND1, ND2, COI, COII, ATP8, ATP6, COIII, ND3, ND4L, ND4, ND5, ND6, and cytochrome b). The topology of the phylogenetic tree is broadly similar to that mentioned by Pyron et al.

Novel genes in brain tissues of EAE-induced normal and obese mice: Upregulation of metal ion-binding protein genes in obese-EAE mice.

PubMed

Hasan, Mahbub; Seo, Ji-Eun; Rahaman, Khandoker Asiqur; Min, Hophil; Kim, Ki Hun; Park, Ju-Hyung; Sung, Changmin; Son, Junghyun; Kang, Min-Jung; Jung, Byung Hwa; Park, Won Sang; Kwon, Oh-Seung

2017-02-20

Experimental autoimmune encephalomyelitis (EAE) is an inflammatory autoimmune disease of the central nervous system resulting from degeneration of the myelin sheath. This study is aimed to identify differentially expressed genes (DEGs) in the brain of EAE-induced normal diet (ND) mice and high-fat diet (HFD)-induced obese mice, and to identify novel genes responsible for elucidating the mechanism of the disease. Purified mRNA samples from the brain tissue were analyzed for gene microarray and validated by real-time RT-PCR. DEGs were identified if significant changes greater than 1.5-fold or less than 0.66-fold were observed (p<0.05). Pathway construction and functional categorization were performed using the Kyoto encyclopedia of genes and genomes pathways and gene ontology (GO) analysis. HFD-EAE mice showed more severe disease symptoms than ND-EAE mice. From GO study, fold changes of HFD-EAE to ND-EAE genes indicated that the genes were significantly associated to the pathways related with the immune response, antigen presentation, and complement activation. The genes related with metal ion-binding proteins were upregulated in HFD-EAE and ND-EAE mice. Upregulation of Cul9, Mast2, and C4b expression is significantly higher in HFD-EAE mice than ND-EAE mice. Cul9, Mast2, C4b, Psmb8, Ly86, and Ms4a6d were significantly upregulated in both ND- and HFD-EAE mice. Fcgr4, S3-12, Gca, and Zdhhc4 were upregulated only in ND-EAE, and Xlr4b was upregulated only in HFD-EAE mice. And significant upregulated genes of metal ion-binding proteins (Cul9 and Mast2) were observed in HFD-EAE mice. Copyright © 2016 IBRO. Published by Elsevier Ltd. All rights reserved.

Intraspecific Genetic Variation and Phylogenetic Analysis of Dirofilaria immitis Samples from Western China Using Complete ND1 and 16S rDNA Gene Sequences

PubMed Central

Liu, Tianyu; Liang, Yinan; Zhong, Xiuqin; Wang, Ning; Hu, Dandan; Zhou, Xuan; Gu, Xiaobin; Peng, Xuerong; Yang, Guangyou

2014-01-01

Dirofilaria immitis (heartworm) is the causative agent of an important zoonotic disease that is spread by mosquitoes. In this study, molecular and phylogenetic characterization of D. immitis were performed based on complete ND1 and 16S rDNA gene sequences, which provided the foundation for more advanced molecular diagnosis, prevention, and control of heartworm diseases. The mutation rate and evolutionary divergence in adult heartworm samples from seven dogs in western China were analyzed to obtain information on genetic diversity and variability. Phylogenetic relationships were inferred using both maximum parsimony (MP) and Bayes methods based on the complete gene sequences. The results suggest that D. immitis formed an independent monophyletic group in which the 16S rDNA gene has mutated more rapidly than has ND1. PMID:24639299

Insight into the mobilome of Aeromonas strains.

PubMed

Piotrowska, Marta; Popowska, Magdalena

2015-01-01

The mobilome is a pool of genes located within mobile genetic elements (MGE), such as plasmids, IS elements, transposons, genomic/pathogenicity islands, and integron-associated gene cassettes. These genes are often referred to as "flexible" and may encode virulence factors, toxic compounds as well as resistance to antibiotics. The phenomenon of MGE transfer between bacteria, known as horizontal gene transfer (HGT), is well documented. The genes present on MGE are subject to continuous processes of evolution and environmental changes, largely induced or significantly accelerated by man. For bacteria, the only chance of survival in an environment contaminated with toxic chemicals, heavy metals and antibiotics is the acquisition of genes providing the ability to survive in such conditions. The process of acquiring and spreading antibiotic resistance genes (ARG) is of particular significance, as it is important for the health of humans and animals. Therefore, it is important to thoroughly study the mobilome of Aeromonas spp. that is widely distributed in various environments, causing many diseases in fishes and humans. This review discusses the recently published information on MGE prevalent in Aeromonas spp. with special emphasis on plasmids belonging to different incompatibility groups, i.e., IncA/C, IncU, IncQ, IncF, IncI, and ColE-type. The vast majority of plasmids carry a number of different transposons (Tn3, Tn21, Tn1213, Tn1721, Tn4401), the 1st, 2nd, or 3rd class of integrons, IS elements (e.g., IS26, ISPa12, ISPa13, ISKpn8, ISKpn6) and encode determinants such as antibiotic and mercury resistance genes, as well as virulence factors. Although the actual role of Aeromonas spp. as a human pathogen remains controversial, species of this genus may pose a serious risk to human health. This is due to the considerable potential of their mobilome, particularly in terms of antibiotic resistance and the possibility of the horizontal transfer of resistance genes.

Insight into the mobilome of Aeromonas strains

PubMed Central

Piotrowska, Marta; Popowska, Magdalena

2015-01-01

The mobilome is a pool of genes located within mobile genetic elements (MGE), such as plasmids, IS elements, transposons, genomic/pathogenicity islands, and integron-associated gene cassettes. These genes are often referred to as “flexible” and may encode virulence factors, toxic compounds as well as resistance to antibiotics. The phenomenon of MGE transfer between bacteria, known as horizontal gene transfer (HGT), is well documented. The genes present on MGE are subject to continuous processes of evolution and environmental changes, largely induced or significantly accelerated by man. For bacteria, the only chance of survival in an environment contaminated with toxic chemicals, heavy metals and antibiotics is the acquisition of genes providing the ability to survive in such conditions. The process of acquiring and spreading antibiotic resistance genes (ARG) is of particular significance, as it is important for the health of humans and animals. Therefore, it is important to thoroughly study the mobilome of Aeromonas spp. that is widely distributed in various environments, causing many diseases in fishes and humans. This review discusses the recently published information on MGE prevalent in Aeromonas spp. with special emphasis on plasmids belonging to different incompatibility groups, i.e., IncA/C, IncU, IncQ, IncF, IncI, and ColE-type. The vast majority of plasmids carry a number of different transposons (Tn3, Tn21, Tn1213, Tn1721, Tn4401), the 1st, 2nd, or 3rd class of integrons, IS elements (e.g., IS26, ISPa12, ISPa13, ISKpn8, ISKpn6) and encode determinants such as antibiotic and mercury resistance genes, as well as virulence factors. Although the actual role of Aeromonas spp. as a human pathogen remains controversial, species of this genus may pose a serious risk to human health. This is due to the considerable potential of their mobilome, particularly in terms of antibiotic resistance and the possibility of the horizontal transfer of resistance genes. PMID:26074893

Tobacco habit: historical, cultural, neurobiological, and genetic features of people's relationship with an addictive drug.

PubMed

Russo, Patrizia; Nastrucci, Candida; Alzetta, Giulio; Szalai, Clara

2011-01-01

This article reviews the cultural history of man's relationship with tobacco and the steps in the discovery of tobacco addiction. Nicotine dependence (ND) or nicotine addiction (NA), among other forms of drug addiction, continues to be a significant public health problem in the world, as it is associated with major severe diseases such as cardiovascular disease and cancer. Evidence for a genetic influence on smoking behavior and ND has prompted a search for susceptibility genes. Proof has recently accumulated that single nucleotide polymorphisms (SNPs) in the genetic region encoding the nicotinic acetylcholine receptor (nAChR) subunits α5, α3, and β4 are associated with smoking and ND. In this review, we consider tobacco as the archetype of substance addiction and describe the evolution of the tobacco habit from elite users to lower socioeconomic abusers (by mass marketing and specific targeting of vulnerable groups by the tobacco industry) to exemplify detrimental behavior with major threats to public health. Finally, we discuss the reasons for the difficulty of quitting addictions/habits and highlight possible solutions.

Selenium Pretreatment Alleviated LPS-Induced Immunological Stress Via Upregulation of Several Selenoprotein Encoding Genes in Murine RAW264.7 Cells.

PubMed

Wang, Longqiong; Jing, Jinzhong; Yan, Hui; Tang, Jiayong; Jia, Gang; Liu, Guangmang; Chen, Xiaoling; Tian, Gang; Cai, Jingyi; Shang, Haiying; Zhao, Hua

2018-04-18

This study was conducted to profile selenoprotein encoding genes in mouse RAW264.7 cells upon lipopolysaccharide (LPS) challenge and integrate their roles into immunological regulation in response to selenium (Se) pretreatment. LPS was used to develop immunological stress in macrophages. Cells were pretreated with different levels of Se (0, 0.5, 1.0, 1.5, 2.0 μmol Se/L) for 2 h, followed by LPS (100 ng/mL) stimulation for another 3 h. The mRNA expression of 24 selenoprotein encoding genes and 9 inflammation-related genes were investigated. The results showed that LPS (100 ng/mL) effectively induced immunological stress in RAW264.7 cells with induced inflammation cytokines, IL-6 and TNF-α, mRNA expression, and cellular secretion. LPS increased (P < 0.05) mRNA profiles of 9 inflammation-related genes in cells, while short-time Se pretreatment modestly reversed (P < 0.05) the LPS-induced upregulation of 7 genes (COX-2, ICAM-1, IL-1β, IL-6, IL-10, iNOS, and MCP-1) and further increased (P < 0.05) expression of IFN-β and TNF-α in stressed cells. Meanwhile, LPS decreased (P < 0.05) mRNA levels of 18 selenoprotein encoding genes and upregulated mRNA levels of TXNRD1 and TXNRD3 in cells. Se pretreatment recovered (P < 0.05) expression of 3 selenoprotein encoding genes (GPX1, SELENOH, and SELENOW) in a dose-dependent manner and increased (P < 0.05) expression of another 5 selenoprotein encoding genes (SELENOK, SELENOM, SELENOS, SELENOT, and TXNRD2) only at a high level (2.0 μmol Se/L). Taken together, LPS-induced immunological stress in RAW264.7 cells accompanied with the global downregulation of selenoprotein encoding genes and Se pretreatment alleviated immunological stress via upregulation of a subset of selenoprotein encoding genes.

An evolutionary switch in ND2 enables Src kinase regulation of NMDA receptors

NASA Astrophysics Data System (ADS)

Scanlon, David P.; Bah, Alaji; Krzeminski, Mickaël; Zhang, Wenbo; Leduc-Pessah, Heather L.; Dong, Yi Na; Forman-Kay, Julie D.; Salter, Michael W.

2017-05-01

The non-receptor tyrosine kinase Src is a key signalling hub for upregulating the function of N-methyl D-aspartate receptors (NMDARs). Src is anchored within the NMDAR complex via NADH dehydrogenase subunit 2 (ND2), a mitochondrially encoded adaptor protein. The interacting regions between Src and ND2 have been broadly identified, but the interaction between ND2 and the NMDAR has remained elusive. Here we generate a homology model of ND2 and dock it onto the NMDAR via the transmembrane domain of GluN1. This interaction is enabled by the evolutionary loss of three helices in bilaterian ND2 proteins compared to their ancestral homologues. We experimentally validate our model and demonstrate that blocking this interaction with an ND2 fragment identified in our experimental studies prevents Src-mediated upregulation of NMDAR currents in neurons. Our findings establish the mode of interaction between an NMDAR accessory protein with one of the core subunits of the receptor.

Cyclic stretch-induced the cytoskeleton rearrangement and gene expression of cytoskeletal regulators in human periodontal ligament cells.

PubMed

Wu, Yaqin; Zhuang, Jiabao; Zhao, Dan; Zhang, Fuqiang; Ma, Jiayin; Xu, Chun

2017-10-01

This study aimed to explore the mechanism of the stretch-induced cell realignment and cytoskeletal rearrangement by identifying several mechanoresponsive genes related to cytoskeletal regulators in human PDL cells. After the cells were stretched by 1, 10 and 20% strains for 0.5, 1, 2, 4, 6, 12 or 24 h, the changes of the morphology and content of microfilaments were recorded and calculated. Meanwhile, the expression of 84 key genes encoding cytoskeletal regulators after 6 and 24 h stretches with 20% strain was detected by using real-time PCR array. Western blot was applied to identify the protein expression level of several cytoskeletal regulators encoded by these differentially expressed genes. The confocal fluorescent staining results confirmed that stretch-induced realignment of cells and rearrangement of microfilaments. Among the 84 genes screened, one gene was up-regulated while two genes were down-regulated after 6 h stretch. Meanwhile, three genes were up-regulated while two genes were down-regulated after 24 h stretch. These genes displaying differential expression included genes regulating polymerization/depolymerization of microfilaments (CDC42EP2, FNBP1L, NCK2, PIKFYVE, WASL), polymerization/depolymerization of microtubules (STMN1), interacting between microfilaments and microtubules (MACF1), as well as a phosphatase (PPP1R12B). Among the proteins encoded by these genes, the protein expression level of Cdc42 effector protein-2 (encoded by CDC42EP2) and Stathmin-1 (encoded by STMN1) was down-regulated, while the protein expression level of N-WASP (encoded by WASL) was up-regulated. The present study confirmed the cyclic stretch-induced cellular realignment and rearrangement of microfilaments in the human PDL cells and indicated several force-sensitive genes with regard to cytoskeletal regulators.

Healthy Nordic diet downregulates the expression of genes involved in inflammation in subcutaneous adipose tissue in individuals with features of the metabolic syndrome.

PubMed

Kolehmainen, Marjukka; Ulven, Stine M; Paananen, Jussi; de Mello, Vanessa; Schwab, Ursula; Carlberg, Carsten; Myhrstad, Mari; Pihlajamäki, Jussi; Dungner, Elisabeth; Sjölin, Eva; Gunnarsdottir, Ingibjörg; Cloetens, Lieselotte; Landin-Olsson, Mona; Akesson, Björn; Rosqvist, Fredrik; Hukkanen, Janne; Herzig, Karl-Heinz; Dragsted, Lars O; Savolainen, Markku J; Brader, Lea; Hermansen, Kjeld; Risérus, Ulf; Thorsdottir, Inga; Poutanen, Kaisa S; Uusitupa, Matti; Arner, Peter; Dahlman, Ingrid

2015-01-01

Previously, a healthy Nordic diet (ND) has been shown to have beneficial health effects close to those of Mediterranean diets. The objective was to explore whether the ND has an impact on gene expression in abdominal subcutaneous adipose tissue (SAT) and whether changes in gene expression are associated with clinical and biochemical effects. Obese adults with features of the metabolic syndrome underwent an 18- to 24-wk randomized intervention study comparing the ND with the control diet (CD) (the SYSDIET study, carried out within Nordic Centre of Excellence of the Systems Biology in Controlled Dietary Interventions and Cohort Studies). The present study included participants from 3 Nordic SYSDIET centers [Kuopio (n = 20), Lund (n = 18), and Oulu (n = 18)] with a maximum weight change of ±4 kg, highly sensitive C-reactive protein concentration <10 mg/L at the beginning and the end of the intervention, and baseline body mass index (in kg/m²) <38. SAT biopsy specimens were obtained before and after the intervention and subjected to global transcriptome analysis with Gene 1.1 ST Arrays (Affymetrix). Altogether, 128 genes were differentially expressed in SAT between the ND and CD (nominal P < 0.01; false discovery rate, 25%). These genes were overrepresented in pathways related to immune response (adjusted P = 0.0076), resulting mainly from slightly decreased expression in the ND and increased expression in the CD. Immune-related pathways included leukocyte trafficking and macrophage recruitment (e.g., interferon regulatory factor 1, CD97), adaptive immune response (interleukin32, interleukin 6 receptor), and reactive oxygen species (neutrophil cytosolic factor 1). Interestingly, the regulatory region of the 128 genes was overrepresented for binding sites for the nuclear transcription factor κB. A healthy Nordic diet reduces inflammatory gene expression in SAT compared with a control diet independently of body weight change in individuals with features of the metabolic syndrome. © 2015 American Society for Nutrition.

NHE10, a novel osteoclast-specific member of the Na{sup +}/H{sup +} exchanger family, regulates osteoclast differentiation and survival

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

Lee, Seoung Hoon; Kim, Taesoo; Park, Eui-Soon

2008-05-02

Bone homeostasis is tightly regulated by the balanced actions of osteoblasts (OBs) and osteoclasts (OCs). We previously analyzed the gene expression profile of OC differentiation using a cDNA microarray, and identified a novel osteoclastogenic gene candidate, clone OCL-1-E7 [J. Rho, C.R. Altmann, N.D. Socci, L. Merkov, N. Kim, H. So, O. Lee, M. Takami, A.H. Brivanlou, Y. Choi, Gene expression profiling of osteoclast differentiation by combined suppression subtractive hybridization (SSH) and cDNA microarray analysis, DNA Cell Biol. 21 (2002) 541-549]. In this study, we have isolated full-length cDNAs corresponding to this clone from mice and humans to determine the functionalmore » roles of this gene in osteoclastogenesis. The full-length cDNA of OCL-1-E7 encodes 12 membrane-spanning domains that are typical of isoforms of the Na{sup +}/H{sup +} exchangers (NHEs), indicating that this clone is a novel member of the NHE family (hereafter referred to as NHE10). Here, we show that NHE10 is highly expressed in OCs in response to receptor activator of nuclear factor-{kappa}B ligand signaling and is required for OC differentiation and survival.« less

Partial kinetoplast-mitochondrial gene organization and expression in the respiratory deficient plant trypanosomatid Phytomonas serpens.

PubMed

Maslov, D A; Nawathean, P; Scheel, J

1999-04-30

In plant-dwelling trypanosomatids from the genus Phytomonas, mitochondrial functions, such as cytochrome mediated respiration, ATP production and Krebs cycle, are missing, and cell energetics is based on the glycolysis. Using Blue Native/Tricine-SDS two-dimensional gel electrophoretic analysis, we observed that mitochondrial respiratory Complexes III (cytochrome bc1) and IV (cytochrome c oxidase) were absent in Phytomonas serpens; however, Complex V (ATPase) was present. A deletion of the genes for cytochrome c oxidase subunit III (COIII) and apocytochrome b (Cyb) was identified within the 6234 bp sequenced region of the 31 kb maxicircle kinetoplast DNA. Genes, found in this region, include 12S and 9S ribosomal RNAs, subunits 7, 8 and 9 of NADH dehydrogenase (ND7, ND8 and ND9) and subunit 6 of ATPase (A6 or MURF4), as well as the genes (MURF1, MURF5 and G3) with unknown function. Most genes are actively transcribed and some mRNAs are edited. Fully edited mRNAs for A6 and G3 were abundant, while edited ND7 transcripts were rare, and only partially edited and pre-edited transcripts for ND8 were detected. The data show that the mitochondrial genome of P. serpens is functional, although its functions may be limited to expressing the ATPase and, possibly, NADH dehydrogenase complexes.

The organization of the fuc regulon specifying L-fucose dissimilation in Escherichia coli K12 as determined by gene cloning.

PubMed

Chen, Y M; Zhu, Y; Lin, E C

1987-12-01

In Escherichia coli the six known genes specifying the utilization of L-fucose as carbon and energy source cluster at 60.2 min and constitute a regulon. These genes include fucP (encoding L-fucose permease), fucI (encoding L-fucose isomerase), fucK (encoding L-fuculose kinase), fucA (encoding L-fuculose 1-phosphate aldolase), fucO (encoding L-1,2-propanediol oxidoreductase), and fucR (encoding the regulatory protein). In this study the fuc genes were cloned and their positions on the chromosome were established by restriction endonuclease and complementation analyses. Clockwise, the gene order is: fucO-fucA-fucP-fucI-fucK-fucR. The operons comprising the structural genes and the direction of transcription were determined by complementation analysis and Southern blot hybridization. The fucPIK and fucA operons are transcribed clockwise. The fucO operon is transcribed counterclockwise. The fucR gene product activates the three structural operons in trans.

Contiguous deletion of the NDP, MAOA, MAOB, and EFHC2 genes in a patient with Norrie disease, severe psychomotor retardation and myoclonic epilepsy.

PubMed

Rodriguez-Revenga, L; Madrigal, I; Alkhalidi, L S; Armengol, L; González, E; Badenas, C; Estivill, X; Milà, M

2007-05-01

Norrie disease (ND) is an X-linked disorder, inherited as a recessive trait that, therefore, mostly affects males. The gene responsible for ND, called NDP, maps to the short arm of chromosome X (Xp11.4-p11.3). We report here an atypical case of ND, consisting of a patient harboring a large submicroscopic deletion affecting not only the NDP gene but also the MAOA, MAOB, and EFHC2 genes. Microarray comparative genomic hybridization (CGH) analysis showed that 11 consecutive bacterial artificial chromosome (BAC) clones, mapping around the NDP gene, were deleted. These clones span a region of about 1 Mb on Xp11.3. The deletion was ascertained by fluorescent in situ hybridization (FISH) analysis with different BAC clones located within the region. Clinical features of the proband include bilateral retinal detachment, microcephaly, severe psychomotor retardation without verbal language skills acquired, and epilepsy. The identification and molecular characterization of this case reinforces the idea of a new contiguous gene syndrome that would explain the complex phenotype shared by atypical ND patients.

Epigenetic Testing for Breast Cancer Risk Stratification

DTIC Science & Technology

2012-10-01

The genes selected for this validation were: ER-POS: GSTP1 , HBA2, BNC1, and WDR66 ER-NEG: IRF7, PECI, ARTN, VCAN, ADM, LIPG, and PLAU Figure 1...6 maintained in only a small fraction of the tumor cells. Only BNC1, CCNA1, and GSTP1 show noticeable expansion of the methylated population in...ADM 0.820 0.609 ARTN 0.192 0.138 GSTP1 0.218 0.070 LIPG 0.090 0.011 CCNA1 0.655 0.428 VCAN 0.301 0.128 IRF7 0.483 0.496 HBA2 cND ND PLAU ND ND

An initial investigation of associations between dopamine-linked genetic variation and smoking motives in African Americans.

PubMed

Bidwell, L C; McGeary, J E; Gray, J C; Palmer, R H C; Knopik, V S; MacKillop, J

2015-11-01

Nicotine dependence (ND) is a heterogeneous phenotype with complex genetic influences that may vary across ethnicities. The use of intermediate phenotypes may clarify genetic influences and reveal specific etiological pathways. Prior work in European Americans has found that the four Primary Dependence Motives (PDM) subscales (Automaticity, Craving, Loss of Control, and Tolerance) of the Wisconsin Inventory of Smoking Motives represent core features of nicotine dependence and are promising intermediate phenotypes for understanding genetic pathways to ND. However, no studies have examined PDM as an intermediate phenotype in African American smokers, an ethnic population that displays unique patterns of smoking and genetic variation. In the current study, 268 African American daily smokers completed a phenotypic assessment and provided a sample of DNA. Associations among haplotypes in the NCAM1-TTC12-ANKK1-DRD2 gene cluster, a dopamine-related gene region associated with ND, PDM intermediate phenotypes, and ND were examined. Dopamine-related genetic variation in the DBH and COMT genes was also considered on an exploratory basis. Mediational analysis was used to test the indirect pathway from genetic variation to smoking motives to nicotine dependence. NCAM1-TTC12-ANKK1-DRD2 region variation was significantly associated with the Automaticity subscale and, further, Automaticity significantly mediated associations among NCAM1-TTC12-ANKK1-DRD2 cluster variants and ND. DBH was also significantly associated with Automaticity, Craving, and Tolerance; Automaticity and Tolerance also served as mediators of the DBH-ND relationship. These results suggest that PDM, Automaticity in particular, may be a viable intermediate phenotype for understanding dopamine-related genetic influences on ND in African American smokers. Findings support a model in which putatively dopaminergic variants exert influence on ND through an effect on patterns of automatic routinized smoking. Copyright © 2015 Elsevier Inc. All rights reserved.

Excess amino acid polymorphism in mitochondrial DNA: contrasts among genes from Drosophila, mice, and humans.

PubMed

Rand, D M; Kann, L M

1996-07-01

Recent studies of mitochondrial DNA (mtDNA) variation in mammals and Drosophila have shown an excess of amino acid variation within species (replacement polymorphism) relative to the number of silent and replacement differences fixed between species. To examine further this pattern of nonneutral mtDNA evolution, we present sequence data for the ND3 and ND5 genes from 59 lines of Drosophila melanogaster and 29 lines of D. simulans. Of interest are the frequency spectra of silent and replacement polymorphisms, and potential variation among genes and taxa in the departures from neutral expectations. The Drosophila ND3 and ND5 data show no significant excess of replacement polymorphism using the McDonald-Kreitman test. These data are in contrast to significant departures from neutrality for the ND3 gene in mammals and other genes in Drosophila mtDNA (cytochrome b and ATPase 6). Pooled across genes, however, both Drosophila and human mtDNA show very significant excesses of amino acid polymorphism. Silent polymorphisms at ND5 show a significantly higher variance in frequency than replacement polymorphisms, and the latter show a significant skew toward low frequencies (Tajima's D = -1.954). These patterns are interpreted in light of the nearly neutral theory where mildly deleterious amino acid haplotypes are observed as ephemeral variants within species but do not contribute to divergence. The patterns of polymorphism and divergence at charge-altering amino acid sites are presented for the Drosophila ND5 gene to examine the evolution of functionally distinct mutations. Excess charge-altering polymorphism is observed at the carboxyl terminal and excess charge-altering divergence is detected at the amino terminal. While the mildly deleterious model fits as a net effect in the evolution of nonrecombining mitochondrial genomes, these data suggest that opposing evolutionary pressures may act on different regions of mitochondrial genes and genomes.

Leaderless mRNAs are circularized in Chlamydomonas reinhardtii mitochondria.

PubMed

Cahoon, A Bruce; Qureshi, Ali A

2018-06-01

The mitochondrial genome of Chlamydomonas reinhardtii encodes eight protein coding genes transcribed on two polycistronic primary transcripts. The mRNAs are endonucleolytically cleaved from these transcripts directly upstream of their AUG start codons, creating leaderless mRNAs with 3' untranslated regions (UTR) comprised of most or all of their downstream intergenic regions. In this report, we provide evidence that these processed linear mRNAs are circularized, which places the 3' UTR upstream of the 5' start codon, creating a leader sequence ex post facto. The circular mRNAs were found to be ribosome associate by polysome profiling experiments suggesting they are translated. Sequencing of the 3'-5' junctions of the circularized mRNAs found the intra-molecular ligations occurred between fully processed 5' ends (the start AUG) and a variable 3' terminus. For five genes (cob, cox, nd2, nd4, and nd6), some of the 3' ends maintained an oligonucleotide addition during ligation, and for two of them, cob and nd6, these 3' termini were the most commonly recovered sequence. Previous reports have shown that after cleavage, three untemplated oligonucleotide additions may occur on the 3' termini of these mRNAs-adenylation, uridylylation, or cytidylation. These results suggest oligo(U) and oligo(C) additions may be part of the maturation process since they are maintained in the circular mRNAs. Circular RNAs occur in organisms across the biological spectrum, but their purpose in some systems, such as organelles (mitochondria and chloroplasts) is unclear. We hypothesize, that in C. reinhardtii mitochondria it may create a leader sequence to facilitate translation initiation, which may negate the need for an alternative translation initiation mechanism in this system, as previously speculated. In addition, circularization may play a protective role against exonucleases, and/or increase translational productivity.

Chromobacterium spp. harbour Ambler class A β-lactamases showing high identity with KPC.

PubMed

Gudeta, Dereje Dadi; Bortolaia, Valeria; Jayol, Aurélie; Poirel, Laurent; Nordmann, Patrice; Guardabassi, Luca

2016-06-01

The origin of KPC is unknown. The aim of this study was to detect progenitors of KPC in silico and to functionally verify their β-lactam hydrolysis activity. The sequence of KPC-2 was used to mine the NCBI protein sequence database. The best non-KPC hits were analysed by amino acid (aa) alignment and phylogenetic tree construction. Genes encoding KPC-2 homologues were expressed in Escherichia coli. The carbapenemase activities of the recombinant strains were characterized by the CarbaNP test and UV spectrophotometry and MICs of selected β-lactams were determined. Genes encoding the closest KPC-2 homologues were identified on the chromosome of Chromobacterium piscinae strain ND17 (CRP-1, 76% aa identity), Chromobacterium sp. C-61 (CRS-1, 70% aa identity) and Chromobacterium haemolyticum DSM19808 (CRH-1, 69% aa identity). All three Chromobacterium β-lactamases were phylogenetically more related to KPC than to other Ambler class A β-lactamases. The 27 bp region preceding the start codon of blaCRP-1 displayed high nucleotide identity to the corresponding region upstream from blaKPC (74%). Heterologous expression of blaCRP-1 and to a lesser extent of blaCRH-1 in E. coli significantly increased the MICs of meropenem and most cephalosporins. The CarbaNP test was positive for both recombinant strains, but spectrophotometric analysis confirmed higher carbapenemase activity for CRP-1-producing clones. The recovery of three class A β-lactamases with up to 76% aa identity to KPC from distinct Chromobacterium species is highly indicative of the role played by this genus in the evolution of KPC. © The Author 2016. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Paralogous ALT1 and ALT2 Retention and Diversification Have Generated Catalytically Active and Inactive Aminotransferases in Saccharomyces cerevisiae

PubMed Central

Peñalosa-Ruiz, Georgina; Aranda, Cristina; Ongay-Larios, Laura; Colon, Maritrini; Quezada, Hector; Gonzalez, Alicia

2012-01-01

Background Gene duplication and the subsequent divergence of paralogous pairs play a central role in the evolution of novel gene functions. S. cerevisiae possesses two paralogous genes (ALT1/ALT2) which presumably encode alanine aminotransferases. It has been previously shown that Alt1 encodes an alanine aminotransferase, involved in alanine metabolism; however the physiological role of Alt2 is not known. Here we investigate whether ALT2 encodes an active alanine aminotransferase. Principal Findings Our results show that although ALT1 and ALT2 encode 65% identical proteins, only Alt1 displays alanine aminotransferase activity; in contrast ALT2 encodes a catalytically inert protein. ALT1 and ALT2 expression is modulated by Nrg1 and by the intracellular alanine pool. ALT1 is alanine-induced showing a regulatory profile of a gene encoding an enzyme involved in amino acid catabolism, in agreement with the fact that Alt1 is the sole pathway for alanine catabolism present in S. cerevisiae. Conversely, ALT2 expression is alanine-repressed, indicating a role in alanine biosynthesis, although the encoded-protein has no alanine aminotransferase enzymatic activity. In the ancestral-like yeast L. kluyveri, the alanine aminotransferase activity was higher in the presence of alanine than in the presence of ammonium, suggesting that as for ALT1, LkALT1 expression could be alanine-induced. ALT2 retention poses the questions of whether the encoded protein plays a particular function, and if this function was present in the ancestral gene. It could be hypotesized that ALT2 diverged after duplication, through neo-functionalization or that ALT2 function was present in the ancestral gene, with a yet undiscovered function. Conclusions ALT1 and ALT2 divergence has resulted in delegation of alanine aminotransferase activity to Alt1. These genes display opposed regulatory profiles: ALT1 is alanine-induced, while ALT2 is alanine repressed. Both genes are negatively regulated by the Nrg1 repressor. Presented results indicate that alanine could act as ALT2 Nrg1-co-repressor. PMID:23049841

Novel mutation at the initiation codon in the Norrie disease gene in two Japanese families.

PubMed

Isashiki, Y; Ohba, N; Yanagita, T; Hokita, N; Doi, N; Nakagawa, M; Ozawa, M; Kuroda, N

1995-01-01

We have identified a new mutation of Norrie disease (ND) gene in two Japanese males from unrelated families; they showed typical ocular features of ND but no mental retardation or hearing impairment. A mutation was found in both patients at the initiation codon of exon 2 of the ND gene (ATG to GTG), with otherwise normal nucleotide sequences. Their mothers had the normal and mutant types of the gene, which was expected for heterozygotes of the disease. The mutation of the initiation codon would cause the failure of ND gene expression or a defect in translation thereby truncating the amino terminus of ND protein. In view of the rarity and marked heterogeneity of mutations in the ND gene, the present apparently unrelated Japanese families who have lived in the same area for over two centuries presumably share the origin of the mutation.

Aluminium induced oxidative stress results in decreased mitochondrial biogenesis via modulation of PGC-1α expression

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

Sharma, Deep Raj; Sunkaria, Aditya; Wani, Willayat Yousuf

The present investigation was carried out to elucidate a possible molecular mechanism related to the effects of aluminium-induced oxidative stress on various mitochondrial respiratory complex subunits with special emphasis on the role of Peroxisome proliferator activated receptor gamma co-activator 1α (PGC-1α) and its downstream targets i.e. Nuclear respiratory factor-1(NRF-1), Nuclear respiratory factor-2(NRF-2) and Mitochondrial transcription factor A (Tfam) in mitochondrial biogenesis. Aluminium lactate (10 mg/kg b.wt./day) was administered intragastrically to rats for 12 weeks. After 12 weeks of exposure, we found an increase in ROS levels, mitochondrial DNA oxidation and decrease in citrate synthase activity in the Hippocampus (HC) andmore » Corpus striatum (CS) regions of rat brain. On the other hand, there was a decrease in the mRNA levels of the mitochondrial encoded subunits–NADH dehydrogenase (ND) subunits i.e. ND1, ND2, ND3, Cytochrome b (Cytb), Cytochrome oxidase (COX) subunits i.e. COX1, COX3, ATP synthase (ATPase) subunit 6 along with reduced expression of nuclear encoded subunits COX4, COX5A, COX5B of Electron transport chain (ETC). Besides, a decrease in mitochondrial DNA copy number and mitochondrial content in both regions of rat brain was observed. The PGC-1α was down-regulated in aluminium treated rats along with NRF-1, NRF-2 and Tfam, which act downstream from PGC-1α in aluminium treated rats. Electron microscopy results revealed a significant increase in the mitochondrial swelling, loss of cristae, chromatin condensation and decreases in mitochondrial number in case of aluminium treated rats as compared to control. So, PGC-1α seems to be a potent target for aluminium neurotoxicity, which makes it an almost ideal target to control or limit the damage that has been associated with the defective mitochondrial function seen in neurodegenerative diseases. - Highlights: • Aluminium decreases the mRNA levels of mitochondrial and nuclear encoded subunits. • It decreases the mtDNA copy number and mitochondrial content in rat brain. • It down-regulates the mRNA and protein levels of PGC-1α, NRF-1, NRF-2 and Tfam. • It also disturbs the mitochondrial or nuclear architecture of neurons. • Finally it also decreases mitochondrial number in HC and CS regions of rat brain.« less

Identification of missense mutations in the Norrie disease gene associated with advanced retinopathy of prematurity.

PubMed

Shastry, B S; Pendergast, S D; Hartzer, M K; Liu, X; Trese, M T

1997-05-01

Retinopathy of prematurity (ROP) is a retinal vascular disease occurring in infants with short gestational age and low birth weight and can lead to retinal detachment (ROP stages 4 and 5). X-linked familial exudative vitreoretinopathy is phenotypically similar to ROP and has been associated with mutations in the Norrie disease (ND) gene in some cases. To determine if similar mutations in the ND gene may play a role in the development of advanced ROP. Clinical examination and molecular genetic analysis were performed on 16 children, including 2 dizygotic and 1 monozygotic twin pairs, and their parents from 13 families. Sequencing of the amplified products revealed missense mutations (R121W and L108P) in the third exon of the ND gene in 4 patients. These mutations were not present in an unaffected premature twin, 2 children with regressed stage 3 ROP, the parents, or in 50 unrelated healthy control subjects. These findings suggest that mutations in the ND gene may play a role in the development of severe ROP in premature infants.

Mitochondrial Gene Expression Profiles Are Associated with Maternal Psychosocial Stress in Pregnancy and Infant Temperament

PubMed Central

Lambertini, Luca; Chen, Jia; Nomura, Yoko

2015-01-01

Background Gene-environment interactions mediate through the placenta and shape the fetal brain development. Between the environmental determinants of the fetal brain, maternal psychosocial stress in pregnancy has been shown to negatively influence the infant temperament development. This in turn may have adverse consequences on the infant neurodevelopment extending throughout the entire life-span. However little is known about the underlying biological mechanisms of the effects of maternal psychosocial stress in pregnancy on infant temperament. Environmental stressors such as maternal psychosocial stress in pregnancy activate the stress response cascade that in turn drives the increase in the cellular energy demand of vital organs with high metabolic rates such as, in pregnancy, the placenta. Key players of the stress response cascade are the mitochondria. Results Here, we tested the expression of all 13 protein-coding genes encoded by the mitochondria in 108 placenta samples from the Stress in Pregnancy birth cohort, a study that aims at determining the influence of in utero exposure to maternal psychosocial stress in pregnancy on infant temperament. We showed that the expression of the protein-coding mitochondrial-encoded gene MT-ND2 was positively associated with indices of maternal psychosocial stress in pregnancy including Prenatal Perceived Stress (β = 0.259; p-regression = 0.004; r2-regression = 0.120), State Anxiety (β = 0.218; p-regression = 0.003; r2-regression = 0.153), Trait Anxiety (β = 0.262; p-regression = 0.003; r2-regression = 0.129) and Pregnancy Anxiety Total (β = 0.208; p-regression = 0.010; r2-regression = 0.103). In the meantime MT-ND2 was negatively associated with the infant temperament indices of Activity Level (β = -0.257; p-regression = 0.008; r2-regression = 0.165) and Smile and Laughter (β = -0.286; p-regression = 0.036; r2-regression = 0.082). Additionally, MT-ND6 was associated with the maternal psychosocial stress in pregnancy index of Prenatal Perceived Stress (β = -0.231; p-regression = 0.004; r2-regression = 0.120), while MT-CO2 was associated with the maternal psychosocial stress in pregnancy indices of State Anxiety (β = 0.206; p-regression = 0.003; r2-regression = 0.153) and Trait Anxiety (β = 0.205; p-regression = 0.003; r2-regression = 0.129). Conclusions Our data support the role of mitochondria in responding to maternal psychosocial stress in pregnancy, as assessed in placenta, while also suggesting an important role for the mitochondria in the infant temperament development. PMID:26418562

Mitochondrial Gene Expression Profiles Are Associated with Maternal Psychosocial Stress in Pregnancy and Infant Temperament.

PubMed

Lambertini, Luca; Chen, Jia; Nomura, Yoko

2015-01-01

Gene-environment interactions mediate through the placenta and shape the fetal brain development. Between the environmental determinants of the fetal brain, maternal psychosocial stress in pregnancy has been shown to negatively influence the infant temperament development. This in turn may have adverse consequences on the infant neurodevelopment extending throughout the entire life-span. However little is known about the underlying biological mechanisms of the effects of maternal psychosocial stress in pregnancy on infant temperament. Environmental stressors such as maternal psychosocial stress in pregnancy activate the stress response cascade that in turn drives the increase in the cellular energy demand of vital organs with high metabolic rates such as, in pregnancy, the placenta. Key players of the stress response cascade are the mitochondria. Here, we tested the expression of all 13 protein-coding genes encoded by the mitochondria in 108 placenta samples from the Stress in Pregnancy birth cohort, a study that aims at determining the influence of in utero exposure to maternal psychosocial stress in pregnancy on infant temperament. We showed that the expression of the protein-coding mitochondrial-encoded gene MT-ND2 was positively associated with indices of maternal psychosocial stress in pregnancy including Prenatal Perceived Stress (β = 0.259; p-regression = 0.004; r2-regression = 0.120), State Anxiety (β = 0.218; p-regression = 0.003; r2-regression = 0.153), Trait Anxiety (β = 0.262; p-regression = 0.003; r2-regression = 0.129) and Pregnancy Anxiety Total (β = 0.208; p-regression = 0.010; r2-regression = 0.103). In the meantime MT-ND2 was negatively associated with the infant temperament indices of Activity Level (β = -0.257; p-regression = 0.008; r2-regression = 0.165) and Smile and Laughter (β = -0.286; p-regression = 0.036; r2-regression = 0.082). Additionally, MT-ND6 was associated with the maternal psychosocial stress in pregnancy index of Prenatal Perceived Stress (β = -0.231; p-regression = 0.004; r2-regression = 0.120), while MT-CO2 was associated with the maternal psychosocial stress in pregnancy indices of State Anxiety (β = 0.206; p-regression = 0.003; r2-regression = 0.153) and Trait Anxiety (β = 0.205; p-regression = 0.003; r2-regression = 0.129). Our data support the role of mitochondria in responding to maternal psychosocial stress in pregnancy, as assessed in placenta, while also suggesting an important role for the mitochondria in the infant temperament development.

Complete mitochondrial genome of Yangtze River wild common carp (Cyprinus carpio haematopterus) and Russian scattered scale mirror carp (Cyprinus carpio carpio).

PubMed

Hu, Guang Fu; Liu, Xiang Jiang; Zou, Gui Wei; Li, Zhong; Liang, Hong-Wei; Hu, Shao-Na

2016-01-01

We sequenced the complete mitogenomes of (Cyprinus carpio haematopterus) and Russian scattered scale mirror carp (Cyprinus carpio carpio). Comparison of these two mitogenomes revealed that the mitogenomes of these two common carp strains were remarkably similar in genome length, gene order and content, and AT content. There were only 55 bp variations in 16,581 nucleotides. About 1 bp variation was located in rRNAs, 2 bp in tRNAs, 9 bp in the control region and 43 bp in protein-coding genes. Furthermore, forty-three variable nucleotides in the protein-coding genes of the two strains led to four variable amino acids, which were located in the ND2, ATPase 6, ND5 and ND6 genes, respectively.

The Bacillus subtilis ywjI (glpX) gene encodes a class II fructose-1,6-bisphosphatase, functionally equivalent to the class III Fbp enzyme.

PubMed

Jules, Matthieu; Le Chat, Ludovic; Aymerich, Stéphane; Le Coq, Dominique

2009-05-01

We present here experimental evidence that the Bacillus subtilis ywjI gene encodes a class II fructose-1,6-bisphosphatase, functionally equivalent to the fbp-encoded class III enzyme, and constitutes with the upstream gene, murAB, an operon transcribed at the same level under glycolytic or gluconeogenic conditions.

The Bacillus subtilis ywjI (glpX) Gene Encodes a Class II Fructose-1,6-Bisphosphatase, Functionally Equivalent to the Class III Fbp Enzyme▿

PubMed Central

Jules, Matthieu; Le Chat, Ludovic; Aymerich, Stéphane; Le Coq, Dominique

2009-01-01

We present here experimental evidence that the Bacillus subtilis ywjI gene encodes a class II fructose-1,6-bisphosphatase, functionally equivalent to the fbp-encoded class III enzyme, and constitutes with the upstream gene, murAB, an operon transcribed at the same level under glycolytic or gluconeogenic conditions. PMID:19270101

Two Thai families with Norrie disease (ND): association of two novel missense mutations with severe ND phenotype, seizures, and a manifesting carrier.

PubMed

Yamada, K; Limprasert, P; Ratanasukon, M; Tengtrisorn, S; Yingchareonpukdee, J; Vasiknanonte, P; Kitaoka, T; Ghadami, M; Niikawa, N; Kishino, T

2001-04-15

We describe two Thai families with Norrie disease (ND) in three generations, including 10 affected males and one manifesting female. All affected males in each family had severely defective eye development with complete loss of vision. In addition, three male patients (one from family 1 and two from family 2) suffered from epilepsy, and one female carrier from one family manifested blindness with phthisis bulbi in her right eye. Mutation analysis of the ND gene (NDP) revealed two different novel missense mutations (L16P and S75P) that co-segregated with ND in each family, suggesting that the newly appearing proline at codon 16 or codon 75 alters the conformation of the ND protein and contributes to the severe phenotype of ND in each family. Other studies suggest that epileptic seizures or growth retardation that is associated with ND is the consequence of loss of contiguous genes, because most such patients had deletions extending beyond the Norrie locus. Our finding that the three affected males in the two families with the missense mutations had epilepsy does not support a contiguous gene effect, but favors the pleiotropism of NDP, at least as far as the epileptic manifestation is concerned. The unilateral blindness in the female carrier may have been due to non-random X-inactivation. Copyright 2001 Wiley-Liss, Inc.

Accumulation of Mitochondrial DNA Common Deletion Since The Preataxic Stage of Machado-Joseph Disease.

PubMed

Raposo, Mafalda; Ramos, Amanda; Santos, Cristina; Kazachkova, Nadiya; Teixeira, Balbina; Bettencourt, Conceição; Lima, Manuela

2018-04-21

Molecular alterations reflecting pathophysiologic changes thought to occur many years before the clinical onset of Machado-Joseph disease (MJD)/spinocerebellar ataxia type 3 (SCA3), a late-onset polyglutamine disorder, remain unidentified. The absence of molecular biomarkers hampers clinical trials, which lack sensitive measures of disease progression, preventing the identification of events occurring prior to clinical onset. Our aim was to analyse the mtDNA content and the amount of the common deletion (m.8482_13460del4977) in a cohort of 16 preataxic MJD mutation carriers, 85 MJD patients and 101 apparently healthy age-matched controls. Relative expression levels of RPPH1, MT-ND1 and MT-ND4 genes were assessed by quantitative real-time PCR. The mtDNA content was calculated as the difference between the expression levels of a mitochondrial gene (MT-ND1) and a nuclear gene (RPPH1); the amount of mtDNA common deletion was calculated as the difference between expression levels of a deleted (MT-ND4) and an undeleted (MT-ND1) mitochondrial genes. mtDNA content in MJD carriers was similar to that of healthy age-matched controls, whereas the percentage of the common deletion was significantly increased in MJD subjects, and more pronounced in the preclinical stage (p < 0.05). The BCL2/BAX ratio was decreased in preataxic carriers compared to controls, suggesting that the mitochondrial-mediated apoptotic pathway is altered in MJD. Our findings demonstrate for the first time that accumulation of common deletion starts in the preclinical stage. Such early alterations provide support to the current understanding that any therapeutic intervention in MJD should start before the overt clinical phenotype.

Next-generation sequencing of the yellowfin tuna mitochondrial genome reveals novel phylogenetic relationships within the genus Thunnus.

PubMed

Guo, Liang; Li, Mingming; Zhang, Heng; Yang, Sen; Chen, Xinghan; Meng, Zining; Lin, Haoran

2016-05-01

Recently, the next-generation sequencing (NGS) technology has become a powerful tool for sequencing the teleost mitochondrial genome (mitogenome). Here, we used this technology to determine the mitogenome of the yellowfin tuna (Thunnus albacares). A total of 41,378 reads were generated by Illumina platform with an average depth of 250×. The mitogenome (16,528 bp in length) contained 37 mitochondrial genes with the similar gene order to other typical teleosts. These mitochondrial genes were encoded on the heavy strand except for ND6 and eight tRNA genes. The result of phylogenetic analysis supported two distinct clades dividing the genus Thunnus, but the tuna species of these two genetic clades were different from that of two recognized subgenus based on anatomical characters and geographical distribution. Our results might help to understand the structure, function, and evolutionary history of the yellowfin tuna mitogenome and also provide valuable new insights for phylogenetic affinity of tuna species.

Bisphenol A-Associated Alterations in the Expression and Epigenetic Regulation of Genes Encoding Xenobiotic Metabolizing Enzymes in Human Fetal Liver

PubMed Central

Nahar, Muna S.; Kim, Jung H.; Sartor, Maureen A.; Dolinoy, Dana C.

2014-01-01

Alterations in xenobiotic metabolizing enzyme (XME) expression across the life course, along with genetic, nutritional, and environmental regulation, can influence how organisms respond to toxic insults. In this study, we investigated the hypothesis that in utero exposure to the endocrine active compound, bisphenol A (BPA), influences expression and epigenetic regulation of phase I and II XME genes during development. Using healthy 1st to 2nd trimester human fetal liver specimens quantified for internal BPA levels, we examined XME gene expression using PCR Array (n =8) and RNA-sequencing (n =12) platforms. Of the greater than 160 XME genes assayed, 2 phase I and 12 phase II genes exhibited significantly reduced expression with higher BPA levels, including isoforms from the carboxylesterase, catechol O-methyltransferase, glutathione S-transferase, sulfotransferase, and UDP-glucuronosyltransferase families. When the promoters of these candidate genes were evaluated in silico, putative binding sites for the E-twenty-six (ETS) and activator protein1 (AP1) related transcription factor families were identified and unique to 97% of all candidate transcripts. Interestingly, many ETS binding sites contain cytosine-guanine dinucleotides (CpGs) within their consensus sequences. Thus, quantitative analysis of CpG methylation of three candidate genes was conducted across n =50 samples. Higher BPA levels were associated with increased site-specific methylation at COMT (P <0.005) and increased average methylation at SULT2A1 (P <0.020) promoters. While toxicological studies have traditionally focused on high-dose effects and hormonal receptor mediated regulation, our findings suggest the importance of low-dose effects and nonclassical mechanisms of endocrine disruption during development. PMID:24214726

The complete mitochondrial genome of Percocypris pingi (Teleostei, Cypriniformes).

PubMed

Li, Yanping; Wang, Jinjin; Peng, Zuogang

2013-02-01

Percocypris pingi is an endemic and economic fish species only found in the upper Yangtze River basin in China. It has become endangered in recent years due to overfishing and/or dam construction. However, the available genetic data are still scarce for this species. Here, we sequenced the complete mitochondrial genome sequence of P. pingi using long polymerase chain reactions. The complete mitogenome sequence has 16,586 bp and contains the usual 13 protein-coding genes, 2 ribosomal RNA genes, 22 transfer RNA (tRNA) genes, and 1 control region, the gene composition and order of which are similar to most of other vertebrates. Most mitochondrial genes except ND6 and eight tRNAs are encoded on the heavy strand. The overall base composition of the heavy strand is 30.9% A, 25.7% T, 26.6% C, and 16.8% G with a slight AT bias of 56.6%. There are seven regions of gene overlaps totaling 23 bp and 11 intergenic spacer regions totaling 35 bp. Combined with the COI barcoding region sequences of other 25 cyprinids, the phylogenetic position of P. pingi was estimated using neighbor-joining method. The results showed that P. pingi had a close phylogenetic relationship with the species from genus Schizothorax. This mitogenome sequence data of P. pingi would provide the fundamental genetic data for further conservation genetic studies for this endangered fish species.

Trichoderma genes

DOEpatents

Foreman, Pamela [Los Altos, CA; Goedegebuur, Frits [Vlaardingen, NL; Van Solingen, Pieter [Naaldwijk, NL; Ward, Michael [San Francisco, CA

2012-06-19

Described herein are novel gene sequences isolated from Trichoderma reesei. Two genes encoding proteins comprising a cellulose binding domain, one encoding an arabionfuranosidase and one encoding an acetylxylanesterase are described. The sequences, CIP1 and CIP2, contain a cellulose binding domain. These proteins are especially useful in the textile and detergent industry and in pulp and paper industry.

Characterization of the complete mitochondrial genome of Acanthoscelides obtectus (Coleoptera: Chrysomelidae: Bruchinae) with phylogenetic analysis.

PubMed

Yao, Jie; Yang, Hong; Dai, Renhuai

2017-10-01

Acanthoscelides obtectus is a common species of the subfamily Bruchinae and a worldwide-distributed seed-feeding beetle. The complete mitochondrial genome of A. obtectus is 16,130 bp in length with an A + T content of 76.4%. It contains a positive AT skew and a negative GC skew. The mitogenome of A. obtectus contains 13 protein-coding genes (PCGs), 22 tRNA genes, two rRNA genes and a non-coding region (D-loop). All PCGs start with an ATN codon, and seven (ND3, ATP6, COIII, ND3, ND4L, ND6, and Cytb) of them terminate with TAA, while the remaining five (COI, COII, ND1, ND4, and ND5) terminate with a single T, ATP8 terminates with TGA. Except tRNA Ser , the secondary structures of 21 tRNAs that can be folded into a typical clover-leaf structure were identified. The secondary structures of lrRNA and srRNA were also predicted in this study. There are six domains with 48 helices in lrRNA and three domains with 32 helices in srRNA. The control region of A. obtectus is 1354 bp in size with the highest A + T content (83.5%) in a mitochondrial gene. Thirteen PCGs in 19 species have been used to infer their phylogenetic relationships. Our results show that A. obtectus belongs to the family Chrysomelidae (subfamily-Bruchinae). This is the first study on phylogenetic analyses involving the mitochondrial genes of A. obtectus and could provide basic data for future studies of mitochondrial genome diversities and the evolution of related insect lineages.

Complete mitochondrial genome of Cynopterus sphinx (Pteropodidae: Cynopterus).

PubMed

Li, Linmiao; Li, Min; Wu, Zhengjun; Chen, Jinping

2015-01-01

We have characterized the complete mitochondrial genome of Cynopterus sphinx (Pteropodidae: Cynopterus) and described its organization in this study. The total length of C. sphinx complete mitochondrial genome was 16,895 bp with the base composition of 32.54% A, 14.05% G, 25.82% T and 27.59% C. The complete mitochondrial genome included 13 protein-coding genes, 22 tRNA genes, 2 rRNA genes (12S rRNA and 16S rRNA) and 1 control region (D-loop). The control region was 1435 bp long with the sequence CATACG repeat 64 times. Three protein-coding genes (ND1, COI and ND4) were ended with incomplete stop codon TA or T.

Aluminium induced oxidative stress results in decreased mitochondrial biogenesis via modulation of PGC-1α expression.

PubMed

Sharma, Deep Raj; Sunkaria, Aditya; Wani, Willayat Yousuf; Sharma, Reeta Kumari; Kandimalla, Ramesh J L; Bal, Amanjit; Gill, Kiran Dip

2013-12-01

The present investigation was carried out to elucidate a possible molecular mechanism related to the effects of aluminium-induced oxidative stress on various mitochondrial respiratory complex subunits with special emphasis on the role of Peroxisome proliferator activated receptor gamma co-activator 1α (PGC-1α) and its downstream targets i.e. Nuclear respiratory factor-1(NRF-1), Nuclear respiratory factor-2(NRF-2) and Mitochondrial transcription factor A (Tfam) in mitochondrial biogenesis. Aluminium lactate (10mg/kgb.wt./day) was administered intragastrically to rats for 12 weeks. After 12 weeks of exposure, we found an increase in ROS levels, mitochondrial DNA oxidation and decrease in citrate synthase activity in the Hippocampus (HC) and Corpus striatum (CS) regions of rat brain. On the other hand, there was a decrease in the mRNA levels of the mitochondrial encoded subunits-NADH dehydrogenase (ND) subunits i.e. ND1, ND2, ND3, Cytochrome b (Cytb), Cytochrome oxidase (COX) subunits i.e. COX1, COX3, ATP synthase (ATPase) subunit 6 along with reduced expression of nuclear encoded subunits COX4, COX5A, COX5B of Electron transport chain (ETC). Besides, a decrease in mitochondrial DNA copy number and mitochondrial content in both regions of rat brain was observed. The PGC-1α was down-regulated in aluminium treated rats along with NRF-1, NRF-2 and Tfam, which act downstream from PGC-1α in aluminium treated rats. Electron microscopy results revealed a significant increase in the mitochondrial swelling, loss of cristae, chromatin condensation and decreases in mitochondrial number in case of aluminium treated rats as compared to control. So, PGC-1α seems to be a potent target for aluminium neurotoxicity, which makes it an almost ideal target to control or limit the damage that has been associated with the defective mitochondrial function seen in neurodegenerative diseases. © 2013.

Identification of chitinolytic bacteria isolated from shrimp pond sediment and characterization of their chitinase encoding gene

NASA Astrophysics Data System (ADS)

Triwijayani, A. U.; Puspita, I. D.; Murwantoko; Ustadi

2018-03-01

Chitinolytic bacteria are a group of bacteria owning enzymes that able to hydrolyze chitin. Previously, we isolated chitinolytic bacteria from shrimp pond sediment in Bantul, Yogyakarta, and obtained five isolates showing high chitinolytic index named as isolate PT1, PT2, PT5, PT6 and PB2. The aims of this study were to identify chitinolytic bacteria isolated from shrimp pond sediment and to characterize the chitinase encoding gene from each isolate. The molecular technique was performed by amplification of 16S rDNA, amplification of chitinase encoding gene and sequence analysis. Two chitinolytic bacteria of PT1 and PT2 were similar to Aeromonas bivalvium strain D15, PT5 to Pseudomonas stutzeri strain BD-2.2.1, PT6 to Serratia marcescens strain FZSF02 and PB2 to Streptomyces misionensis strain OsiRt-1. The comparison of chitinase encoding gene between three isolates with those in Gen Bank shows that PT1 had similar sequences with the chi1 gene in Aeromonas sp. 17m, PT2 with chi1 gene in A. caviae (CB101) and PT6 with chiB gene in S. Marcescens (BJL200).

Nano-nutrition of chicken embryos. The effect of in ovo administration of diamond nanoparticles and L-glutamine on molecular responses in chicken embryo pectoral muscles.

PubMed

Grodzik, Marta; Sawosz, Filip; Sawosz, Ewa; Hotowy, Anna; Wierzbicki, Mateusz; Kutwin, Marta; Jaworski, Sławomir; Chwalibog, André

2013-11-20

It has been demonstrated that the content of certain amino acids in eggs is not sufficient to fully support embryonic development. One possibility to supply the embryo with extra nutrients and energy is in ovo administration of nutrients. Nanoparticles of diamond are highly biocompatible non-toxic carbonic structures, and we hypothesized that bio-complexes of diamond nanoparticles with L-glutamine may affect molecular responses in breast muscle. The objective of the investigation was to evaluate the effect of diamond nanoparticle (ND) and L-glutamine (Gln) on expression of growth and differentiation factors of chicken embryo pectoral muscles. ND, Gln, and Gln/ND solutions (50 mg/L) were injected into fertilized broiler chicken eggs at the beginning of embryogenesis. Muscle tissue was dissected at day 20 of incubation and analysed for gene expression of FGF2, VEGF-A, and MyoD1. ND and especially Gln/ND up-regulated expression of genes related to muscle cell proliferation (FGF2) and differentiation (MyoD1). Furthermore, the ratio between FGF2 and MyoD1 was highest in the Gln/ND group. At the end of embryogenesis, Gln/ND enhanced both proliferation and differentiation of pectoral muscle cells and differentiation dominated over proliferation. These preliminary results suggest that the bio-complex of glutamine and diamond nanoparticles may accelerate growth and maturation of muscle cells.

Correlation of ophthalmic examination with carrier status in females potentially harboring a severe Norrie disease gene mutation.

PubMed

Khan, Arif O; Aldahmesh, Mohammed A; Meyer, Brian

2008-04-01

To correlate ophthalmic findings with carrier status for a severe Norrie disease (ND) gene mutation (C95F). Prospective interventional case series. Six potential carriers and 1 obligate carrier from a family harboring the mutation. An ophthalmologist blind to the pedigree performed a full ophthalmic examination for the 7 asymptomatic family members. A peripheral blood sample was collected from each for ND gene sequencing. Ophthalmic examination findings (with attention to the presence or absence of retinal findings) and results of ND gene sequencing. Three carriers were identified by molecular genetics, and all 3 of them had peripheral retinal abnormality. However, 3 of the 4 genetically identified noncarriers also exhibited peripheral retinal abnormality. Two of these noncarriers with retinal findings were the offspring of a confirmed noncarrier. The genetically identified noncarrier with a normal peripheral retinal examination was the daughter of an obligate carrier. The presence of peripheral retinal changes was not useful for carrier prediction in a family harboring ND. There are likely additional loci responsible for phenotypic expression.

A highly divergent gene cluster in honey bees encodes a novel silk family.

PubMed

Sutherland, Tara D; Campbell, Peter M; Weisman, Sarah; Trueman, Holly E; Sriskantha, Alagacone; Wanjura, Wolfgang J; Haritos, Victoria S

2006-11-01

The pupal cocoon of the domesticated silk moth Bombyx mori is the best known and most extensively studied insect silk. It is not widely known that Apis mellifera larvae also produce silk. We have used a combination of genomic and proteomic techniques to identify four honey bee fiber genes (AmelFibroin1-4) and two silk-associated genes (AmelSA1 and 2). The four fiber genes are small, comprise a single exon each, and are clustered on a short genomic region where the open reading frames are GC-rich amid low GC intergenic regions. The genes encode similar proteins that are highly helical and predicted to form unusually tight coiled coils. Despite the similarity in size, structure, and composition of the encoded proteins, the genes have low primary sequence identity. We propose that the four fiber genes have arisen from gene duplication events but have subsequently diverged significantly. The silk-associated genes encode proteins likely to act as a glue (AmelSA1) and involved in silk processing (AmelSA2). Although the silks of honey bees and silkmoths both originate in larval labial glands, the silk proteins are completely different in their primary, secondary, and tertiary structures as well as the genomic arrangement of the genes encoding them. This implies independent evolutionary origins for these functionally related proteins.

Mutational Analysis of the Rift Valley Fever Virus Glycoprotein Precursor Proteins for Gn Protein Expression

PubMed Central

Phoenix, Inaia; Lokugamage, Nandadeva; Nishiyama, Shoko; Ikegami, Tetsuro

2016-01-01

The Rift Valley fever virus (RVFV) M-segment encodes the 78 kD, NSm, Gn, and Gc proteins. The 1st AUG generates the 78 kD-Gc precursor, the 2nd AUG generates the NSm-Gn-Gc precursor, and the 3rd AUG makes the NSm’-Gn-Gc precursor. To understand biological changes due to abolishment of the precursors, we quantitatively measured Gn secretion using a reporter assay, in which a Gaussia luciferase (gLuc) protein is fused to the RVFV M-segment pre-Gn region. Using the reporter assay, the relative expression of Gn/gLuc fusion proteins was analyzed among various AUG mutants. The reporter assay showed efficient secretion of Gn/gLuc protein from the precursor made from the 2nd AUG, while the removal of the untranslated region upstream of the 2nd AUG (AUG2-M) increased the secretion of the Gn/gLuc protein. Subsequently, recombinant MP-12 strains encoding mutations in the pre-Gn region were rescued, and virological phenotypes were characterized. Recombinant MP-12 encoding the AUG2-M mutation replicated slightly less efficiently than the control, indicating that viral replication is further influenced by the biological processes occurring after Gn expression, rather than the Gn abundance. This study showed that, not only the abolishment of AUG, but also the truncation of viral UTR, affects the expression of Gn protein by the RVFV M-segment. PMID:27231931

Mutational Analysis of the Rift Valley Fever Virus Glycoprotein Precursor Proteins for Gn Protein Expression.

PubMed

Phoenix, Inaia; Lokugamage, Nandadeva; Nishiyama, Shoko; Ikegami, Tetsuro

2016-05-24

The Rift Valley fever virus (RVFV) M-segment encodes the 78 kD, NSm, Gn, and Gc proteins. The 1st AUG generates the 78 kD-Gc precursor, the 2nd AUG generates the NSm-Gn-Gc precursor, and the 3rd AUG makes the NSm'-Gn-Gc precursor. To understand biological changes due to abolishment of the precursors, we quantitatively measured Gn secretion using a reporter assay, in which a Gaussia luciferase (gLuc) protein is fused to the RVFV M-segment pre-Gn region. Using the reporter assay, the relative expression of Gn/gLuc fusion proteins was analyzed among various AUG mutants. The reporter assay showed efficient secretion of Gn/gLuc protein from the precursor made from the 2nd AUG, while the removal of the untranslated region upstream of the 2nd AUG (AUG2-M) increased the secretion of the Gn/gLuc protein. Subsequently, recombinant MP-12 strains encoding mutations in the pre-Gn region were rescued, and virological phenotypes were characterized. Recombinant MP-12 encoding the AUG2-M mutation replicated slightly less efficiently than the control, indicating that viral replication is further influenced by the biological processes occurring after Gn expression, rather than the Gn abundance. This study showed that, not only the abolishment of AUG, but also the truncation of viral UTR, affects the expression of Gn protein by the RVFV M-segment.

CIP1 polypeptides and their uses

DOEpatents

Foreman, Pamela [Los Altos, CA; Van Solingen, Pieter [Naaldwijk, NL; Goedegebuur, Frits [Vlaardingen, NL; Ward, Michael [San Francisco, CA

2011-04-12

Described herein are novel gene sequences isolated from Trichoderma reesei. Two genes encoding proteins comprising a cellulose binding domain, one encoding an arabionfuranosidase and one encoding an acetylxylanesterase are described. The sequences, CIP1 and CIP2, contain a cellulose binding domain. These proteins are especially useful in the textile and detergent industry and in pulp and paper industry.

Mollusk genes encoding lysine tRNA (UUU) contain introns.

PubMed

Matsuo, M; Abe, Y; Saruta, Y; Okada, N

1995-11-20

New intron-containing genes encoding tRNAs were discovered when genomic DNA isolated from various animal species was amplified by the polymerase chain reaction (PCR) with primers based on sequences of rabbit tRNA(Lys). From sequencing analysis of the products of PCR, we found that introns are present in several genes encoding tRNA(Lys) in mollusks, such as Loligo bleekeri (squid) and Octopus vulgaris (octopus). These introns were specific to genes encoding tRNA(Lys)(CUU) and were not present in genes encoding tRNA(Lys)(CUU). In addition, the sequences of the introns were different from one another. To confirm the results of our initial experiments, we isolated and sequenced genes encoding tRNA(Lys)(CUU) and tRNA(Lys)(UUU). The gene for tRNA(Lys)(UUU) from squid contained an intron, whose sequence was the same as that identified by PCR, and the gene formed a cluster with a corresponding pseudogene. Several DNA regions of 2.1 kb containing this cluster appeared to be tandemly arrayed in the squid genome. By contrast, the gene encoding tRNA(Lys)(CUU) did not contain an intron, as shown also by PCR. The tRNA(Lys)(UUU) that corresponded to the analyzed gene was isolated and characterized. The present study provides the first example of an intron-containing gene encoding a tRNA in mollusks and suggests the universality of introns in such genes in higher eukaryotes.

Characterization of two genes encoding the Mycobacterium tuberculosis ribonucleotide reductase small subunit.

PubMed Central

Yang, F; Curran, S C; Li, L S; Avarbock, D; Graf, J D; Chua, M M; Lu, G; Salem, J; Rubin, H

1997-01-01

Two nrdF genes, nrdF1 and nrdF2, encoding the small subunit (R2) of ribonucleotide reductase (RR) from Mycobacterium tuberculosis have 71% identity at the amino acid level and are both highly homologous with Salmonella typhimurium R2F. The calculated molecular masses of R2-1 and R2-2 are 36,588 (322 amino acids [aa]) and 36,957 (324 aa) Da, respectively. Western blot analysis of crude M. tuberculosis extracts indicates that both R2s are expressed in vivo. Recombinant R2-2 is enzymatically active when assayed with pure recombinant M. tuberculosis R1 subunit. Both ATP and dATP are activators for CDP reduction up to 2 and 1 mM, respectively. The gene encoding M. tuberculosis R2-1, nrdF1, is not linked to nrdF2, nor is either gene linked to the gene encoding the large subunit, M. tuberculosis nrdE. The gene encoding MTP64 was found downstream from nrdF1, and the gene encoding alcohol dehydrogenase was found downstream from nrdF2. A nrdA(Ts) strain of E. coli (E101) could be complemented by simultaneous transformation with M. tuberculosis nrdE and nrdF2. An M. tuberculosis nrdF2 variant in which the codon for the catalytically necessary tyrosine was replaced by the phenylalanine codon did not complement E101 when cotransformed with M. tuberculosis nrdE. Similarly, M. tuberculosis nrdF1 and nrdE did not complement E101. Activity of recombinant M. tuberculosis RR was inhibited by incubating the enzyme with a peptide corresponding to the 7 C-terminal amino acid residues of the R2-2 subunit. M. tuberculosis is a species in which a nrdEF system appears to encode the biologically active species of RR and also the only bacterial species identified so far in which class I RR subunits are not arranged on an operon. PMID:9335290

Draft Genome Sequence of Ezakiella peruensis Strain M6.X2, a Human Gut Gram-Positive Anaerobic Coccus.

PubMed

Diop, Awa; Diop, Khoudia; Tomei, Enora; Raoult, Didier; Fenollar, Florence; Fournier, Pierre-Edouard

2018-03-01

We report here the draft genome sequence of Ezakiella peruensis strain M6.X2 T The draft genome is 1,672,788 bp long and harbors 1,589 predicted protein-encoding genes, including 26 antibiotic resistance genes with 1 gene encoding vancomycin resistance. The genome also exhibits 1 clustered regularly interspaced short palindromic repeat region and 333 genes acquired by horizontal gene transfer. Copyright © 2018 Diop et al.

A single amino acid insertion in the WRKY domain of the Arabidopsis TIR-NBS-LRR-WRKY-type disease resistance protein SLH1 (sensitive to low humidity 1) causes activation of defense responses and hypersensitive cell death.

PubMed

Noutoshi, Yoshiteru; Ito, Takuya; Seki, Motoaki; Nakashita, Hideo; Yoshida, Shigeo; Marco, Yves; Shirasu, Ken; Shinozaki, Kazuo

2005-09-01

In this study we characterized the sensitive to low humidity 1 (slh1) mutant of Arabidopsis ecotype No-0 which exhibits normal growth on agar plate medium but which on transfer to soil shows growth arrest and development of necrotic lesions. cDNA microarray hybridization and RNA gel blot analysis revealed that genes associated with activation of disease resistance were upregulated in the slh1 mutants in response to conditions of low humidity. Furthermore, the slh1 mutants accumulate callose, autofluorescent compounds and salicylic acid (SA). We demonstrate that SA is required for the slh1 phenotype but not PAD4 or NPR1. SLH1 was isolated by map-based cloning and it encodes a resistance (R)-like protein consisting of a domain with Toll and interleukin-1 receptor homology (TIR), a nucleotide-binding domain (NB), leucine-rich repeats (LRR) and a carboxy-terminal WRKY domain. SLH1 is identical to the R gene RRS1-R of the Arabidopsis ecotype Nd-1, a gene which confers resistance to the bacterial pathogen Ralstonia solanacearum GMI1000 and also functions as an R gene to this pathogen in No-0. We identified a 3 bp insertion mutation in slh1 that results in the addition of a single amino acid in the WRKY domain; thereby impairing its DNA-binding activity. Our data suggest that SLH1 disease resistance signaling may be negatively regulated by its WRKY domain in the R protein and that the constitutive defense activation conferred by the slh1 mutation is inhibited by conditions of high humidity.

Alterations in mitochondrial DNA copy number and the activities of electron transport chain complexes and pyruvate dehydrogenase in the frontal cortex from subjects with autism

PubMed Central

Gu, F; Chauhan, V; Kaur, K; Brown, W T; LaFauci, G; Wegiel, J; Chauhan, A

2013-01-01

Autism is a neurodevelopmental disorder associated with social deficits and behavioral abnormalities. Recent evidence suggests that mitochondrial dysfunction and oxidative stress may contribute to the etiology of autism. This is the first study to compare the activities of mitochondrial electron transport chain (ETC) complexes (I–V) and pyruvate dehydrogenase (PDH), as well as mitochondrial DNA (mtDNA) copy number in the frontal cortex tissues from autistic and age-matched control subjects. The activities of complexes I, V and PDH were most affected in autism (n=14) being significantly reduced by 31%, 36% and 35%, respectively. When 99% confidence interval (CI) of control group was taken as a reference range, impaired activities of complexes I, III and V were observed in 43%, 29% and 43% of autistic subjects, respectively. Reduced activities of all five ETC complexes were observed in 14% of autistic cases, and the activities of multiple complexes were decreased in 29% of autistic subjects. These results suggest that defects in complexes I and III (sites of mitochondrial free radical generation) and complex V (adenosine triphosphate synthase) are more prevalent in autism. PDH activity was also reduced in 57% of autistic subjects. The ratios of mtDNA of three mitochondrial genes ND1, ND4 and Cyt B (that encode for subunits of complexes I and III) to nuclear DNA were significantly increased in autism, suggesting a higher mtDNA copy number in autism. Compared with the 95% CI of the control group, 44% of autistic children showed higher copy numbers of all three mitochondrial genes examined. Furthermore, ND4 and Cyt B deletions were observed in 44% and 33% of autistic children, respectively. This study indicates that autism is associated with mitochondrial dysfunction in the brain. PMID:24002085

Complete mitochondrial genome of the Kwangtung skate: Dipturus kwangtungensis (Rajiformes, Rajidae).

PubMed

Jeong, Dageum; Kim, Sung; Kim, Choong-Gon; Lee, Youn-Ho

2015-01-01

The complete sequence of mitochondrial DNA of a Kwangtung skate, Dipturus kwangtungensis, was determined as being circular molecules of 16,912 bp including 2 rRNA, 22 tRNA, 13 protein coding genes (PCGs) and a control region. The arrangement of the PCGs is the same as that found in other Rajidae species. The nucleotide of L-strand which encodes most of the proteins is composed of 30.2% A, 27.4% C, 28.2% T and 14.2% G with a bias toward A+T slightly. Twelve of 13 PCGs are initiated by the ATG codon while COX1 starts with GTG. Only ND4 harbors the incomplete termination codon, TA. All tRNA genes have a typical clover-leaf structure of mitochondrial tRNA with the exception of tRNA(Ser)AGY, which has a reduced DHU arm. This mitogenome is the first report for a species of the genus Dipturus, which will become an important source of information on the phylogenetic relationship and the evolution of the genus Dipturus within the family Rajidae.

Two pheromone precursor genes are transcriptionally expressed in the homothallic ascomycete Sordaria macrospora.

PubMed

Pöggeler, S

2000-06-01

In order to analyze the involvement of pheromones in cell recognition and mating in a homothallic fungus, two putative pheromone precursor genes, named ppg1 and ppg2, were isolated from a genomic library of Sordaria macrospora. The ppg1 gene is predicted to encode a precursor pheromone that is processed by a Kex2-like protease to yield a pheromone that is structurally similar to the alpha-factor of the yeast Saccharomyces cerevisiae. The ppg2 gene encodes a 24-amino-acid polypeptide that contains a putative farnesylated and carboxy methylated C-terminal cysteine residue. The sequences of the predicted pheromones display strong structural similarity to those encoded by putative pheromones of heterothallic filamentous ascomycetes. Both genes are expressed during the life cycle of S. macrospora. This is the first description of pheromone precursor genes encoded by a homothallic fungus. Southern-hybridization experiments indicated that ppg1 and ppg2 homologues are also present in other homothallic ascomycetes.

Innate responses to gene knockouts impact overlapping gene networks and vary with respect to resistance to viral infection.

PubMed

Liu, Yonghong; Liu, Yuanyuan; Wu, Jiaming; Roizman, Bernard; Zhou, Grace Guoying

2018-04-03

Analyses of the levels of mRNAs encoding IFIT1, IFI16, RIG-1, MDA5, CXCL10, LGP2, PUM1, LSD1, STING, and IFNβ in cell lines from which the gene encoding LGP2, LSD1, PML, HDAC4, IFI16, PUM1, STING, MDA5, IRF3, or HDAC 1 had been knocked out, as well as the ability of these cell lines to support the replication of HSV-1, revealed the following: ( i ) Cell lines lacking the gene encoding LGP2, PML, or HDAC4 (cluster 1) exhibited increased levels of expression of partially overlapping gene networks. Concurrently, these cell lines produced from 5 fold to 12 fold lower yields of HSV-1 than the parental cells. ( ii ) Cell lines lacking the genes encoding STING, LSD1, MDA5, IRF3, or HDAC 1 (cluster 2) exhibited decreased levels of mRNAs of partially overlapping gene networks. Concurrently, these cell lines produced virus yields that did not differ from those produced by the parental cell line. The genes up-regulated in cell lines forming cluster 1, overlapped in part with genes down-regulated in cluster 2. The key conclusions are that gene knockouts and subsequent selection for growth causes changes in expression of multiple genes, and hence the phenotype of the cell lines cannot be ascribed to a single gene; the patterns of gene expression may be shared by multiple knockouts; and the enhanced immunity to viral replication by cluster 1 knockout cell lines but not by cluster 2 cell lines suggests that in parental cells, the expression of innate resistance to infection is specifically repressed.

Photocontrol of the expression of genes encoding chlorophyll a/b binding proteins and small subunit of ribulose-1,5-bisphosphate carboxylase in etiolated seedlings of Lycopersicon esculentum (L. ) and Nicotiana tabacum (L. )

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

Wehmeyer, B.; Cashmore, A.R.; Schaefer, E.

Phytochrome and the blue ultraviolet-A photoreceptor control light-induced expression of genes encoding the chlorophyll a/b binding protein of photosystem II and photosystem I and the genes for the small subunit of the ribulose-1,5-bisphosphate carboxylase in etiolated seedlings of Lycopersicon esculentum (tomato) and Nicotiana tabacum (tobacco). A high irradiance response also controls the induction of these genes. Genes encoding photosystem II- and I-associated chlorophyll a/b binding proteins both exhibit a transient rapid increase in expression in response to light pulse or to continuous irradiation. In contrast, genes encoding the small subunit exhibit a continuous increase in expression in response to light.more » These distinct expression characteristics are shown to reflect differences at the level of transcription.« less

Molecular cloning and expression of heteromeric ACCase subunit genes from Jatropha curcas.

PubMed

Gu, Keyu; Chiam, Huihui; Tian, Dongsheng; Yin, Zhongchao

2011-04-01

Acetyl-CoA carboxylase (ACCase) catalyzes the biotin-dependent carboxylation of acetyl-CoA to produce malonyl-CoA, which is the essential first step in the biosynthesis of long-chain fatty acids. ACCase exists as a multi-subunit enzyme in most prokaryotes and the chloroplasts of most plants and algae, while it is present as a multi-domain enzyme in the endoplasmic reticulum of most eukaryotes. The heteromeric ACCase of higher plants consists of four subunits: an α-subunit of carboxyltransferase (α-CT, encoded by accA gene), a biotin carboxyl carrier protein (BCCP, encoded by accB gene), a biotin carboxylase (BC, encoded by accC gene) and a β-subunit of carboxyltransferase (β-CT, encoded by accD gene). In this study, we cloned and characterized the genes accA, accB1, accC and accD that encode the subunits of heteromeric ACCase in Jatropha (Jatropha curcas), a potential biofuel plant. The full-length cDNAs of the four subunit genes were isolated from a Jatropha cDNA library and by using 5' RACE, whereas the genomic clones were obtained from a Jatropha BAC library. They encode a 771 amino acid (aa) α-CT, a 286-aa BCCP1, a 537-aa BC and a 494-aa β-CT, respectively. The single-copy accA, accB1 and accC genes are nuclear genes, while the accD gene is located in chloroplast genome. Jatropha α-CT, BCCP1, BC and β-CT show high identity to their homologues in other higher plants at amino acid level and contain all conserved domains for ACCase activity. The accA, accB1, accC and accD genes are temporally and spatially expressed in the leaves and endosperm of Jatropha plants, which are regulated by plant development and environmental factors. Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

Complete Mitochondrial Genome Sequences of Chinese Indigenous Sheep with Different Tail Types and an Analysis of Phylogenetic Evolution in Domestic Sheep.

PubMed

Fan, Hongying; Zhao, Fuping; Zhu, Caiye; Li, Fadi; Liu, Jidong; Zhang, Li; Wei, Caihong; Du, Lixin

2016-05-01

China has a long history of sheep (Ovis aries [O. aries]) breeding and an abundance of sheep genetic resources. Knowledge of the complete O. aries mitogenome should facilitate the study of the evolutionary history of the species. Therefore, the complete mitogenome of O. aries was sequenced and annotated. In order to characterize the mitogenomes of 3 Chinese sheep breeds (Altay sheep [AL], Shandong large-tailed sheep [SD], and small-tailed Hulun Buir sheep [sHL]), 19 sets of primers were employed to amplify contiguous, overlapping segments of the complete mitochondrial DNA (mtDNA) sequence of each breed. The sizes of the complete mitochondrial genomes of the sHL, AL, and SD breeds were 16,617 bp, 16,613 bp, and 16,613 bp, respectively. The mitochondrial genomes were deposited in the GenBank database with accession numbers KP702285 (AL sheep), KP981378 (SD sheep), and KP981380 (sHL sheep) respectively. The organization of the 3 analyzed sheep mitochondrial genomes was similar, with each consisting of 22 tRNA genes, 2 rRNA genes (12S rRNA and 16S rRNA), 13 protein-coding genes, and 1 control region (D-loop). The NADH dehydrogenase subunit 6 (ND6) and 8 tRNA genes were encoded on the light strand, whereas the rest of the mitochondrial genes were encoded on the heavy strand. The nucleotide skewness of the coding strands of the 3 analyzed mitogenomes was biased toward A and T. We constructed a phylogenetic tree using the complete mitogenomes of each type of sheep to allow us to understand the genetic relationships between Chinese breeds of O. aries and those developed and utilized in other countries. Our findings provide important information regarding the O. aries mitogenome and the evolutionary history of O. aries inside and outside China. In addition, our results provide a foundation for further exploration of the taxonomic status of O. aries.

Complete Mitochondrial Genome Sequences of Chinese Indigenous Sheep with Different Tail Types and an Analysis of Phylogenetic Evolution in Domestic Sheep

PubMed Central

Fan, Hongying; Zhao, Fuping; Zhu, Caiye; Li, Fadi; Liu, Jidong; Zhang, Li; Wei, Caihong; Du, Lixin

2016-01-01

China has a long history of sheep (Ovis aries [O. aries]) breeding and an abundance of sheep genetic resources. Knowledge of the complete O. aries mitogenome should facilitate the study of the evolutionary history of the species. Therefore, the complete mitogenome of O. aries was sequenced and annotated. In order to characterize the mitogenomes of 3 Chinese sheep breeds (Altay sheep [AL], Shandong large-tailed sheep [SD], and small-tailed Hulun Buir sheep [sHL]), 19 sets of primers were employed to amplify contiguous, overlapping segments of the complete mitochondrial DNA (mtDNA) sequence of each breed. The sizes of the complete mitochondrial genomes of the sHL, AL, and SD breeds were 16,617 bp, 16,613 bp, and 16,613 bp, respectively. The mitochondrial genomes were deposited in the GenBank database with accession numbers KP702285 (AL sheep), KP981378 (SD sheep), and KP981380 (sHL sheep) respectively. The organization of the 3 analyzed sheep mitochondrial genomes was similar, with each consisting of 22 tRNA genes, 2 rRNA genes (12S rRNA and 16S rRNA), 13 protein-coding genes, and 1 control region (D-loop). The NADH dehydrogenase subunit 6 (ND6) and 8 tRNA genes were encoded on the light strand, whereas the rest of the mitochondrial genes were encoded on the heavy strand. The nucleotide skewness of the coding strands of the 3 analyzed mitogenomes was biased toward A and T. We constructed a phylogenetic tree using the complete mitogenomes of each type of sheep to allow us to understand the genetic relationships between Chinese breeds of O. aries and those developed and utilized in other countries. Our findings provide important information regarding the O. aries mitogenome and the evolutionary history of O. aries inside and outside China. In addition, our results provide a foundation for further exploration of the taxonomic status of O. aries. PMID:26954183

Intrafamilial variability of the ocular phenotype in a Polish family with a missense mutation (A63D) in the Norrie disease gene.

PubMed

Zaremba, J; Feil, S; Juszko, J; Myga, W; van Duijnhoven, G; Berger, W

1998-09-01

To describe the phenotypic variability in a Polish Norrie disease (ND) family associated with the missense mutation A63D. A patient with spared vision from a Polish ND family underwent detailed ophthalmological examinations including slit-lamp biomicroscopy, ultrasound (USG), angiography, Goldmann kinetic visual field, and electroretinography (ERG). Mutation screening was carried out using the single-strand conformation polymorphism (SSCP) technique and subsequent DNA sequencing of the coding part of the ND gene. A mutation was detected (exon 3, A63D) in a large Polish family with 12 affected males, all but one presenting with classical ND symptoms. In one male, partially preserved vision was observed up to 40 years of age (distance acuity of the right eye 1/50 and left eye 2/50). Slit-lamp examination revealed remnants of a persistent primary vitreous and hyaloid artery. Upon angiography, the retina was vascularized within the posterior pole but not in the periphery. The ERG revealed pathological changes characteristic for chorioretinal degenerations. Within one family, individuals with identical sequence alterations in the ND gene can show remarkable phenotypic variability of the ocular symptoms. These findings indicate the involvement of additional factors (epigenetic or genetic) in ocular pathogenesis of ND.

[Expression changes of major outer membrane protein antigens in Leptospira interrogans during infection and its mechanism].

PubMed

Zheng, Linli; Ge, Yumei; Hu, Weilin; Yan, Jie

2013-03-01

To determine expression changes of major outer membrane protein(OMP) antigens of Leptospira interrogans serogroup Icterohaemorrhagiae serovar Lai strain Lai during infection of human macrophages and its mechanism. OmpR encoding genes and OmpR-related histidine kinase (HK) encoding gene of L.interrogans strain Lai and their functional domains were predicted using bioinformatics technique. mRNA level changes of the leptospiral major OMP-encoding genes before and after infection of human THP-1 macrophages were detected by real-time fluorescence quantitative RT-PCR. Effects of the OmpR-encoding genes and HK-encoding gene on the expression of leptospiral OMPs during infection were determined by HK-peptide antiserum block assay and closantel inhibitive assays. The bioinformatics analysis indicated that LB015 and LB333 were referred to OmpR-encoding genes of the spirochete, while LB014 might act as a OmpR-related HK-encoding gene. After the spirochete infecting THP-1 cells, mRNA levels of leptospiral lipL21, lipL32 and lipL41 genes were rapidly and persistently down-regulated (P <0.01), whereas mRNA levels of leptospiral groEL, mce, loa22 and ligB genes were rapidly but transiently up-regulated (P<0.01). The treatment with closantel and HK-peptide antiserum partly reversed the infection-based down-regulated mRNA levels of lipL21 and lipL48 genes (P <0.01). Moreover, closantel caused a decrease of the infection-based up-regulated mRNA levels of groEL, mce, loa22 and ligB genes (P <0.01). Expression levels of L.interrogans strain Lai major OMP antigens present notable changes during infection of human macrophages. There is a group of OmpR-and HK-encoding genes which may play a major role in down-regulation of expression levels of partial OMP antigens during infection.

Norrie gene product is necessary for regression of hyaloid vessels.

PubMed

Ohlmann, Anne V; Adamek, Edith; Ohlmann, Andreas; Lütjen-Drecoll, Elke

2004-07-01

To investigate the nature and origin of the vitreous membranes in mice with knock-out of the Norrie gene product (ND mice). Eighty-two eyes of ND mice of different age groups (postnatal day [P]0-13 months) and 95 age-matched wild-type control mice were investigated. In vitreoretinal wholemounts and in sagittal sections, vessels and free cells were visualized by labeling for lectin. In addition, staining with a marker for macrophages (F4/80) and collagen XVIII/endostatin known to be involved in regression of hyaloid vessels was performed for light and electron microscopic investigations. Endostatin expression was confirmed by Western blot analysis. Wild-type controls showed the typical pattern of hyaloid vessels, their regression and concomitantly retinal vasculogenesis and angiogenesis. Hyaloid vessels all stained for endostatin, whereas retinal vessels remained unstained. In ND mice, 1 to 5 days after birth, the hyaloid and retinal vasculatures were comparable to that in control mice. The hyaloid vessels also stained for endostatin. Numerous F4/80-positive cells were present adjacent to the vessels. With increasing age, only a few connecting branches of the hyaloid vessels regressed. Even in old mice most of the hyaloid vessels persisted. The vessels still stained for endostatin. Retinal angiogenesis was impaired. Retrolental membranes in ND mice consist of persistent hyaloid vessels, indicating that the ND gene product is important for the process of regression of these vessels. The ND gene product neither influences endostatin expression nor the presence of macrophages.

Novel mutations in Norrie disease gene in Japanese patients with Norrie disease and familial exudative vitreoretinopathy.

PubMed

Kondo, Hiroyuki; Qin, Minghui; Kusaka, Shunji; Tahira, Tomoko; Hasebe, Haruyuki; Hayashi, Hideyuki; Uchio, Eiichi; Hayashi, Kenshi

2007-03-01

To search for mutations in the Norrie disease gene (NDP) in Japanese patients with familial exudative vitreoretinopathy (FEVR) and Norrie disease (ND) and to delineate the mutation-associated clinical features. Direct sequencing after polymerase chain reaction of all exons of the NDP gene was performed on blood collected from 62 probands (31 familial and 31 simplex) with FEVR, from 3 probands with ND, and from some of their family members. The clinical symptoms and signs in the patients with mutations were assessed. X-inactivation in the female carriers was examined in three FEVR families by using leukocyte DNA. Four novel mutations-I18K, K54N, R115L, and IVS2-1G-->A-and one reported mutation, R97P, in the NDP gene were identified in six families. The severity of vitreoretinopathy varied among these patients. Three probands with either K54N or R115L had typical features of FEVR, whereas the proband with R97P had those of ND. Families with IVS2-1G-->A exhibited either ND or FEVR characteristics. A proband with I18K presented with significant phenotypic heterogeneity between the two eyes. In addition, affected female carriers in a family harboring the K54N mutation presented with different degrees of vascular abnormalities in the periphery of the retina. X-inactivation profiles indicated that the skewing was not significantly different between affected and unaffected women. These observations indicate that mutations of the NDP gene can cause ND and 6% of FEVR cases in the Japanese population. The X-inactivation assay with leukocytes may not be predictive of the presence of a mutation in affected female carriers.

L-Theanine Content and Related Gene Expression: Novel Insights into Theanine Biosynthesis and Hydrolysis among Different Tea Plant (Camellia sinensis L.) Tissues and Cultivars

PubMed Central

Liu, Zhi-Wei; Wu, Zhi-Jun; Li, Hui; Wang, Yong-Xin; Zhuang, Jing

2017-01-01

L-Theanine content has tissues and cultivars specificity in tea plant (Camellia sinensis L.), the correlations of theanine metabolic related genes expression profiles with theanine contents were explored in this study. L-theanine contents in the bud and 1st leaf, 2nd leaf, 3rd leaf, old leaf, stem, and lateral root were determined by HPLC from three C. sinensis cultivars, namely ‘Huangjinya’, ‘Anjibaicha’, and ‘Yingshuang’, respectively. The theanine contents in leaves and root of ‘Huangjinya’ were the highest, followed by ‘Anjibaicha’, and ‘Yingshuang’. The theanine contents in the leaves reduced as the leaf mature gradually, and in stem were the least. Seventeen genes encoding enzymes involved in theanine metabolism were identified from GenBank and our tea transcriptome database, including CsTS1, CsTS2, CsGS1, CsGS2, CsGOGAT-Fe, CsGOGAT-NAD(P)H, CsGDH1, CsGDH2, CsALT, CsSAMDC, CsADC, CsCuAO, CsPAO, CsNiR, CsNR, CsGGT1, and CsGGT3. The transcript profiles of those seventeen genes in the different tissues of three tea plant cultivars were analyzed comparatively. Among the different cultivars, the transcript levels of most selected genes in ‘Huangjinya’ were significantly higher than that in the ‘Anjibaicha’ and ‘Yingshuang’. Among the different tissues, the transcript levels of CsTS2, CsGS1, and CsGDH2 almost showed positive correlation with the theanine contents, while the other genes showed negative correlation with the theanine contents in most cases. The theanine contents showed correlations with related genes expression levels among cultivars and tissues of tea plant, and were determined by the integrated effect of the metabolic related genes. PMID:28439281

Distribution and Evolution of Yersinia Leucine-Rich Repeat Proteins

PubMed Central

Hu, Yueming; Huang, He; Hui, Xinjie; Cheng, Xi; White, Aaron P.

2016-01-01

Leucine-rich repeat (LRR) proteins are widely distributed in bacteria, playing important roles in various protein-protein interaction processes. In Yersinia, the well-characterized type III secreted effector YopM also belongs to the LRR protein family and is encoded by virulence plasmids. However, little has been known about other LRR members encoded by Yersinia genomes or their evolution. In this study, the Yersinia LRR proteins were comprehensively screened, categorized, and compared. The LRR proteins encoded by chromosomes (LRR1 proteins) appeared to be more similar to each other and different from those encoded by plasmids (LRR2 proteins) with regard to repeat-unit length, amino acid composition profile, and gene expression regulation circuits. LRR1 proteins were also different from LRR2 proteins in that the LRR1 proteins contained an E3 ligase domain (NEL domain) in the C-terminal region or an NEL domain-encoding nucleotide relic in flanking genomic sequences. The LRR1 protein-encoding genes (LRR1 genes) varied dramatically and were categorized into 4 subgroups (a to d), with the LRR1a to -c genes evolving from the same ancestor and LRR1d genes evolving from another ancestor. The consensus and ancestor repeat-unit sequences were inferred for different LRR1 protein subgroups by use of a maximum parsimony modeling strategy. Structural modeling disclosed very similar repeat-unit structures between LRR1 and LRR2 proteins despite the different unit lengths and amino acid compositions. Structural constraints may serve as the driving force to explain the observed mutations in the LRR regions. This study suggests that there may be functional variation and lays the foundation for future experiments investigating the functions of the chromosomally encoded LRR proteins of Yersinia. PMID:27217422

A family of oculofaciocardiodental syndrome (OFCD) with a novel BCOR mutation and genomic rearrangements involving NHS.

PubMed

Kondo, Yukiko; Saitsu, Hirotomo; Miyamoto, Toshinobu; Nishiyama, Kiyomi; Tsurusaki, Yoshinori; Doi, Hiroshi; Miyake, Noriko; Ryoo, Na-Kyung; Kim, Jeong Hun; Yu, Young Suk; Matsumoto, Naomichi

2012-03-01

Oculofaciocardiodental syndrome (OFCD) is an X-linked dominant disorder associated with male lethality, presenting with congenital cataract, dysmorphic face, dental abnormalities and septal heart defects. Mutations in BCOR (encoding BCL-6-interacting corepressor) cause OFCD. Here, we report on a Korean family with common features of OFCD including bilateral 2nd-3rd toe syndactyly and septal heart defects in three affected females (mother and two daughters). Through the mutation screening and copy number analysis using genomic microarray, we identified a novel heterozygous mutation, c.888delG, in the BCOR gene and two interstitial microduplications at Xp22.2-22.13 and Xp21.3 in all the three affected females. The BCOR mutation may lead to a premature stop codon (p.N297IfsX80). The duplication at Xp22.2-22.13 involved the NHS gene causative for Nance-Horan syndrome, which is an X-linked disorder showing similar clinical features with OFCD in affected males, and in carrier females with milder presentation. Considering the presence of bilateral 2nd-3rd toe syndactyly and septal heart defects, which is unique to OFCD, the mutation in BCOR is likely to be the major determinant for the phenotypes in this family.

The prrF-Encoded Small Regulatory RNAs Are Required for Iron Homeostasis and Virulence of Pseudomonas aeruginosa

PubMed Central

Reinhart, Alexandria A.; Powell, Daniel A.; Nguyen, Angela T.; O'Neill, Maura; Djapgne, Louise; Wilks, Angela; Ernst, Robert K.

2014-01-01

Pseudomonas aeruginosa is an opportunistic pathogen that requires iron to cause infection, but it also must regulate the uptake of iron to avoid iron toxicity. The iron-responsive PrrF1 and PrrF2 small regulatory RNAs (sRNAs) are part of P. aeruginosa's iron regulatory network and affect the expression of at least 50 genes encoding iron-containing proteins. The genes encoding the PrrF1 and PrrF2 sRNAs are encoded in tandem in P. aeruginosa, allowing for the expression of a distinct, heme-responsive sRNA named PrrH that appears to regulate genes involved in heme metabolism. Using a combination of growth, mass spectrometry, and gene expression analysis, we showed that the ΔprrF1,2 mutant, which lacks expression of the PrrF and PrrH sRNAs, is defective for both iron and heme homeostasis. We also identified phuS, encoding a heme binding protein involved in heme acquisition, and vreR, encoding a previously identified regulator of P. aeruginosa virulence genes, as novel targets of prrF-mediated heme regulation. Finally, we showed that the prrF locus encoding the PrrF and PrrH sRNAs is required for P. aeruginosa virulence in a murine model of acute lung infection. Moreover, we showed that inoculation with a ΔprrF1,2 deletion mutant protects against future challenge with wild-type P. aeruginosa. Combined, these data demonstrate that the prrF-encoded sRNAs are critical regulators of P. aeruginosa virulence. PMID:25510881

Molecular cloning and characterization of two novel genes from hexaploid wheat that encode double PR-1 domains coupled with a receptor-like protein kinase

USDA-ARS?s Scientific Manuscript database

Hexaploid wheat (Triticum aestivum L.) contains at least 23 TaPr-1 genes encoding the group 1 pathogenesis-related (PR-1) proteins as identified in our previous work. Here we report the cloning and characterization of TaPr-1-rk1 and TaPr-1-rk2, two novel genes closely related to the wheat PR-1 famil...

Chlorella viruses contain genes encoding a complete polyamine biosynthetic pathway

PubMed Central

Baumann, Sascha; Sander, Adrianne; Gurnon, James R.; Yanai-Balser, Giane; VanEtten, James L.; Piotrowski, Markus

2007-01-01

Two genes encoding the putative polyamine biosynthetic enzymes agmatine iminohydrolase (AIH) and N-carbamoylputrescine amidohydrolase (CPA) were cloned from the chloroviruses PBCV-1, NY-2A and MT325. They were expressed in Escherichia coli to form C-terminal (His)6-tagged proteins and the recombinant proteins were purified by Ni2+- binding affinity chromatography. The biochemical properties of the two enzymes are similar to AIH and CPA enzymes from Arabidopsis thaliana and Pseudomonas aeruginosa. Together with the previously known virus genes encoding ornithine/arginine decarboxlyase (ODC/ADC) and homospermidine synthase, the chloroviruses have genes that encode a complete set of functional enzymes that synthesize the rare polyamine homospermidine from arginine via agmatine, N-carbamoylputrescine and putrescine. The PBCV-1 aih and cpa genes are expressed early during virus infection together with the odc/adc gene, suggesting that biosynthesis of putrescine is important in early stages of viral replication. The aih and cpa genes are widespread in the chlorella viruses. PMID:17101165

Molecular analysis of the NDP gene in two families with Norrie disease.

PubMed

Rivera-Vega, M Refugio; Chiñas-Lopez, Silvet; Vaca, Ana Luisa Jimenez; Arenas-Sordo, M Luz; Kofman-Alfaro, Susana; Messina-Baas, Olga; Cuevas-Covarrubias, Sergio Alberto

2005-04-01

To describe the molecular defects in the Norrie disease protein (NDP) gene in two families with Norrie disease (ND). We analysed two families with ND at molecular level through polymerase chain reaction, DNA sequence analysis and GeneScan. Two molecular defects found in the NDP gene were: a missense mutation (265C > G) within codon 97 that resulted in the interchange of arginine by proline, and a partial deletion in the untranslated 3' region of exon 3 of the NDP gene. Clinical findings were more severe in the family that presented the partial deletion. We also diagnosed the carrier status of one daughter through GeneScan; this method proved to be a useful tool for establishing female carriers of ND. Here we report two novel mutations in the NDP gene in Mexican patients and propose that GeneScan is a viable mean of establishing ND carrier status.

The molecular genetics of Usher syndrome.

PubMed

Ahmed, Z M; Riazuddin, S; Riazuddin, S; Wilcox, E R

2003-06-01

Association of sensorineural deafness and progressive retinitis pigmentosa with and without a vestibular abnormality is the hallmark of Usher syndrome and involves at least 12 loci among three different clinical subtypes. Genes identified for the more commonly inherited loci are USH2A (encoding usherin), MYO7A (encoding myosin VIIa), CDH23 (encoding cadherin 23), PCDH15 (encoding protocadherin 15), USH1C (encoding harmonin), USH3A (encoding clarin 1), and USH1G (encoding SANS). Transcripts from all these genes are found in many tissues/cell types other than the inner ear and retina, but all are uniquely critical for retinal and cochlear cell function. Many of these protein products have been demonstrated to have direct interactions with each other and perform an essential role in stereocilia homeostasis.

Evolutionary analysis of hydrophobin gene family in two wood-degrading basidiomycetes, Phlebia brevispora and Heterobasidion annosum s.l.

PubMed Central

2013-01-01

Background Hydrophobins are small secreted cysteine-rich proteins that play diverse roles during different phases of fungal life cycle. In basidiomycetes, hydrophobin-encoding genes often form large multigene families with up to 40 members. The evolutionary forces driving hydrophobin gene expansion and diversification in basidiomycetes are poorly understood. The functional roles of individual genes within such gene families also remain unclear. The relationship between the hydrophobin gene number, the genome size and the lifestyle of respective fungal species has not yet been thoroughly investigated. Here, we present results of our survey of hydrophobin gene families in two species of wood-degrading basidiomycetes, Phlebia brevispora and Heterobasidion annosum s.l. We have also investigated the regulatory pattern of hydrophobin-encoding genes from H. annosum s.s. during saprotrophic growth on pine wood as well as on culture filtrate from Phlebiopsis gigantea using micro-arrays. These data are supplemented by results of the protein structure modeling for a representative set of hydrophobins. Results We have identified hydrophobin genes from the genomes of two wood-degrading species of basidiomycetes, Heterobasidion irregulare, representing one of the microspecies within the aggregate H. annosum s.l., and Phlebia brevispora. Although a high number of hydrophobin-encoding genes were observed in H. irregulare (16 copies), a remarkable expansion of these genes was recorded in P. brevispora (26 copies). A significant expansion of hydrophobin-encoding genes in other analyzed basidiomycetes was also documented (1–40 copies), whereas contraction through gene loss was observed among the analyzed ascomycetes (1–11 copies). Our phylogenetic analysis confirmed the important role of gene duplication events in the evolution of hydrophobins in basidiomycetes. Increased number of hydrophobin-encoding genes appears to have been linked to the species’ ecological strategy, with the non-pathogenic fungi having increased numbers of hydrophobins compared with their pathogenic counterparts. However, there was no significant relationship between the number of hydrophobin-encoding genes and genome size. Furthermore, our results revealed significant differences in the expression levels of the 16 H. annosum s.s. hydrophobin-encoding genes which suggest possible differences in their regulatory patterns. Conclusions A considerable expansion of the hydrophobin-encoding genes in basidiomycetes has been observed. The distribution and number of hydrophobin-encoding genes in the analyzed species may be connected to their ecological preferences. Results of our analysis also have shown that H. annosum s.l. hydrophobin-encoding genes may be under positive selection. Our gene expression analysis revealed differential expression of H. annosum s.s. hydrophobin genes under different growth conditions, indicating their possible functional diversification. PMID:24188142

Nandrolone decanoate interferes with testosterone biosynthesis altering blood-testis barrier components.

PubMed

Barone, Rosario; Pitruzzella, Alessandro; Marino Gammazza, Antonella; Rappa, Francesca; Salerno, Monica; Barone, Fulvio; Sangiorgi, Claudia; D'Amico, Daniela; Locorotondo, Nicola; Di Gaudio, Francesca; Cipolloni, Luigi; Di Felice, Valentina; Schiavone, Stefania; Rapisarda, Venerando; Sani, Gabriele; Tambo, Amos; Cappello, Francesco; Turillazzi, Emanuela; Pomara, Cristoforo

2017-08-01

The aim of this study was to investigate whether nandrolone decanoate (ND) use affects testosterone production and testicular morphology in a model of trained and sedentary mice. A group of mice underwent endurance training while another set led a sedentary lifestyle and were freely mobile within cages. All experimental groups were treated with either ND or peanut oil at different doses for 6 weeks. Testosterone serum levels were measured via liquid chromatography-mass spectrometry. Western blot analysis and quantitative real-time PCR were utilized to determine gene and protein expression levels of the primary enzymes implicated in testosterone biosynthesis and gene expression levels of the blood-testis barrier (BTB) components. Immunohistochemistry and immunofluorescence were conducted for testicular morphological evaluation. The study demonstrated that moderate to high doses of ND induced a diminished serum testosterone level and altered the expression level of the key steroidogenic enzymes involved in testosterone biosynthesis. At the morphological level, ND induced degradation of the BTB by targeting the tight junction protein-1 (TJP1). ND stimulation deregulated metalloproteinase-9, metalloproteinase-2 (MMP-2) and the tissue inhibitor of MMP-2. Moreover, ND administration resulted in a mislocalization of mucin-1. In conclusion, ND abuse induces a decline in testosterone production that is unable to regulate the internalization and redistribution of TJP1 and may induce the deregulation of other BTB constituents via the inhibition of MMP-2. ND may well be considered as both a potential inducer of male infertility and a potential risk factor to a low endogenous bioavailable testosterone. © 2017 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

Nano-Nutrition of Chicken Embryos. The Effect of in Ovo Administration of Diamond Nanoparticles and l-Glutamine on Molecular Responses in Chicken Embryo Pectoral Muscles

PubMed Central

Grodzik, Marta; Sawosz, Filip; Sawosz, Ewa; Hotowy, Anna; Wierzbicki, Mateusz; Kutwin, Marta; Jaworski, Sławomir; Chwalibog, André

2013-01-01

It has been demonstrated that the content of certain amino acids in eggs is not sufficient to fully support embryonic development. One possibility to supply the embryo with extra nutrients and energy is in ovo administration of nutrients. Nanoparticles of diamond are highly biocompatible non-toxic carbonic structures, and we hypothesized that bio-complexes of diamond nanoparticles with l-glutamine may affect molecular responses in breast muscle. The objective of the investigation was to evaluate the effect of diamond nanoparticle (ND) and l-glutamine (Gln) on expression of growth and differentiation factors of chicken embryo pectoral muscles. ND, Gln, and Gln/ND solutions (50 mg/L) were injected into fertilized broiler chicken eggs at the beginning of embryogenesis. Muscle tissue was dissected at day 20 of incubation and analysed for gene expression of FGF2, VEGF-A, and MyoD1. ND and especially Gln/ND up-regulated expression of genes related to muscle cell proliferation (FGF2) and differentiation (MyoD1). Furthermore, the ratio between FGF2 and MyoD1 was highest in the Gln/ND group. At the end of embryogenesis, Gln/ND enhanced both proliferation and differentiation of pectoral muscle cells and differentiation dominated over proliferation. These preliminary results suggest that the bio-complex of glutamine and diamond nanoparticles may accelerate growth and maturation of muscle cells. PMID:24264045

A rapid screening with direct sequencing from blood samples for the diagnosis of Leigh syndrome.

PubMed

Shimbo, Hiroko; Takagi, Mariko; Okuda, Mitsuko; Tsuyusaki, Yu; Takano, Kyoko; Iai, Mizue; Yamashita, Sumimasa; Murayama, Kei; Ohtake, Akira; Goto, Yu-Ichi; Aida, Noriko; Osaka, Hitoshi

2014-01-01

Large numbers of genes are responsible for Leigh syndrome (LS), making genetic confirmation of LS difficult. We screened our patients with LS using a limited set of 21 primers encompassing the frequently reported gene for the respiratory chain complexes I (ND1-ND6, and ND4L), IV(SURF1), and V(ATP6) and the pyruvate dehydrogenase E1α-subunit. Of 18 LS patients, we identified mutations in 11 patients, including 7 in mDNA (two with ATP6), 4 in nuclear (three with SURF1). Overall, we identified mutations in 61% of LS patients (11/18 individuals) in this cohort. Sanger sequencing with our limited set of primers allowed us a rapid genetic confirmation of more than half of the LS patients and it appears to be efficient as a primary genetic screening in this cohort.

Genome-Wide Architecture of Disease Resistance Genes in Lettuce

PubMed Central

Christopoulou, Marilena; Wo, Sebastian Reyes-Chin; Kozik, Alex; McHale, Leah K.; Truco, Maria-Jose; Wroblewski, Tadeusz; Michelmore, Richard W.

2015-01-01

Genome-wide motif searches identified 1134 genes in the lettuce reference genome of cv. Salinas that are potentially involved in pathogen recognition, of which 385 were predicted to encode nucleotide binding-leucine rich repeat receptor (NLR) proteins. Using a maximum-likelihood approach, we grouped the NLRs into 25 multigene families and 17 singletons. Forty-one percent of these NLR-encoding genes belong to three families, the largest being RGC16 with 62 genes in cv. Salinas. The majority of NLR-encoding genes are located in five major resistance clusters (MRCs) on chromosomes 1, 2, 3, 4, and 8 and cosegregate with multiple disease resistance phenotypes. Most MRCs contain primarily members of a single NLR gene family but a few are more complex. MRC2 spans 73 Mb and contains 61 NLRs of six different gene families that cosegregate with nine disease resistance phenotypes. MRC3, which is 25 Mb, contains 22 RGC21 genes and colocates with Dm13. A library of 33 transgenic RNA interference tester stocks was generated for functional analysis of NLR-encoding genes that cosegregated with disease resistance phenotypes in each of the MRCs. Members of four NLR-encoding families, RGC1, RGC2, RGC21, and RGC12 were shown to be required for 16 disease resistance phenotypes in lettuce. The general composition of MRCs is conserved across different genotypes; however, the specific repertoire of NLR-encoding genes varied particularly of the rapidly evolving Type I genes. These tester stocks are valuable resources for future analyses of additional resistance phenotypes. PMID:26449254

A mutation affecting carbon catabolite repression suppresses growth defects in pyruvate carboxylase mutants from Saccharomyces cerevisiae.

PubMed

Blázquez, M A; Gamo, F J; Gancedo, C

1995-12-18

Yeasts with disruptions in the genes PYC1 and PYC2 encoding the isoenzymes of pyruvate carboxylase cannot grow in a glucose-ammonium medium (Stucka et al. (1991) Mol. Gen. Genet. 229, 307-315). We have isolated a dominant mutation, BPC1-1, that allows growth in this medium of yeasts with interrupted PYC1 and PYC2 genes. The BPC1-1 mutation abolishes catabolite repression of a series of genes and allows expression of the enzymes of the glyoxylate cycle during growth in glucose. A functional glyoxylate cycle is necessary for suppression as a disruption of gene ICL1 encoding isocitrate lyase abolished the phenotypic effect of BPC1-1 on growth in glucose-ammonium. Concurrent expression from constitutive promoters of genes ICL1 and MLS1 (encoding malate synthase) also suppressed the growth phenotype of pyc1 pyc2 mutants. The mutation BPC1-1 is either allelic or closely linked to the mutation DGT1-1.

Electrochemical detection of the MT-ND6 gene and its enzymatic digestion: application in human genomic sample.

PubMed

Mazloum-Ardakani, Mohammad; Ahmadi, Roya; Heidari, Mohammad Mehdi; Sheikh-Mohseni, Mohammad Ali

2014-06-15

A simple electrochemical biosensor was developed for the detection of the mitochondrial NADH dehydrogenase 6 gene (MT-ND6) and its enzymatic digestion by BamHI enzyme. This biosensor was fabricated by modification of a glassy carbon electrode with gold nanoparticles (AuNPs/GCE) and a probe oligonucleotide (ssDNA/AuNPs/GCE). The probe, which is a thiolated segment of the MT-ND6 gene, was deposited by self-assembling immobilization on AuNPs/GCE. Two indicators including methylene blue (MB) and neutral red (NR) were used as the electroactive indicators and the electrochemical response of the modified electrode was measured by differential pulse voltammetry. The proposed biosensor can detect the complementary sequences of the MT-ND6 gene. Also the modified electrode was used for the detection of an enzymatic digestion process by BamHI enzyme. The electrochemical biosensor can detect the MT-ND6 gene and its enzymatic digestion in polymerase chain reaction (PCR)-amplified DNA extracted from human blood. Also the biosensor was used directly for detection of the MT-ND6 gene in all of the human genome. Copyright © 2014 Elsevier Inc. All rights reserved.

Association between N142D genetic polymorphism of GSTO2 and susceptibility to colorectal cancer.

PubMed

Masoudi, Mohammad; Saadat, Iraj; Omidvari, Shahpour; Saadat, Mostafa

2011-10-01

Expression pattern analysis has been revealed that glutathione S-transferase omega 2 (GSTO2, a member of class omega) is ubiquitously expressed. Over expression of GSTO2 induced apoptosis. The gene encoding GSTO2 was localized to human chromosome 10q24.3, a region that may harbor gene(s) involved in the developing of colorectal cancer. To investigate the association between GSTO2 N142D genetic polymorphism and susceptibility to colorectal cancer the present study was done. We studied 63 (26 females, 37 males) colorectal cancer patients and 126 (52 females, 74 males) healthy individuals. The control subjects were frequency matched for age and gender with the colorectal cancer group. The genotypes were performed using RFLP-PCR method. The ND and DD genotypes were not associated with risk of colorectal cancer, in comparison with the NN genotype. Family history for cancer in the first degree of relatives significantly differed between cases and controls (P = 0.012). The profiles of GSTO2 genotypes and family history in control and cancerous groups were compared to each other. Subjects with NN genotype and positive family history significantly were at high risk to develop colorectal cancer in comparison with subjects with DD or ND genotypes and negative family history (P = 0.003). Present findings indicating that GSTO2 NN genotype increase the risk of colorectal cancer in persons with positive family history for cancer in the first degree relatives.

Amplification of Chromosome 1q Genes Encoding the Phosphoinositide Signalling Enzymes PI4KB, AKT3, PIP5K1A and PI3KC2B in Breast Cancer

PubMed Central

Waugh, Mark G.

2014-01-01

Little is known about the possible oncogenic roles of genes encoding for the phosphatidylinositol 4-kinases, a family of enzymes that regulate an early step in phosphoinositide signalling. To address this issue, the mutational status of all four human phosphatidylinositol 4-kinases genes was analyzed across 852 breast cancer samples using the COSMIC data resource. Point mutations in the phosphatidylinositol 4-kinase genes were uncommon and appeared in less than 1% of the patient samples however, 62% of the tumours had increases in gene copy number for PI4KB which encodes the phosphatidylinositol 4-kinase IIIbeta isozyme. Extending this analysis to subsequent enzymes in the phosphoinositide signalling cascades revealed that the only PIP5K1A, PI3KC2B and AKT3 genes exhibited similar patterns of gene copy number variation. By comparison, gene copy number increases for established oncogenes such as EGFR and HER2/Neu were only evident in 20% of the samples. The PI4KB, PIP5K1A, PI3KC2B and AKT3 genes are related in that they all localize to chromosome 1q which is often structurally and numerically abnormal in breast cancer. These results demonstrate that a gene quartet encoding a potential phosphoinositide signalling pathway is amplified in a subset of breast cancers. PMID:25368680

Salinivibrio siamensis sp. nov., from fermented fish (pla-ra) in Thailand.

PubMed

Chamroensaksri, Nitcha; Tanasupawat, Somboon; Akaracharanya, Ancharida; Visessanguan, Wonnop; Kudo, Takuji; Itoh, Takashi

2009-04-01

A Gram-negative, facultatively anaerobic, moderately halophilic bacterium, strain ND1-1(T), was isolated from fermented fish (pla-ra) in Thailand. The cells were curved rods, motile and non-endospore-forming. The novel strain grew optimally at 37 degrees C, at pH 8 and in the presence of 9-10 % (w/v) NaCl. The predominant respiratory lipoquinone was Q-8. The major cellular fatty acids were C(16 : 0) and C(12 : 0). Polar lipid analysis revealed the presence of phosphatidylethanolamine, phosphatidylglycerol and diphosphatidylglycerol. The DNA G+C content was 49.0 mol%. Comparative 16S rRNA gene sequence analyses indicated that strain ND1-1(T) was closely related to Salinivibrio costicola, which comprises three subspecies, and Salinivibrio proteolyticus with gene sequence similarities of 98.3-98.6 %. Strain ND1-1(T) showed low levels of DNA-DNA relatedness with S. costicola subsp. costicola JCM 15095(T) (33.2 %), S. costicola subsp. alcaliphilus DSM 16359(T) (38.4 %), S. costicola subsp. vallismortis JCM 15096(T) (59.7 %), and S. proteolyticus AF-2004(T) (42.1 %). On the basis of the physiological and biochemical characteristics and the molecular data presented, strain ND1-1(T) should be classified as a novel species of the genus Salinivibrio for which the name Salinivibrio siamensis sp. nov. is proposed. The type strain is ND1-1(T) (=JCM 14472(T)=PCU 301(T)=TISTR 1810(T)).

Gene Cluster Encoding Cholate Catabolism in Rhodococcus spp.

PubMed Central

Wilbrink, Maarten H.; Casabon, Israël; Stewart, Gordon R.; Liu, Jie; van der Geize, Robert; Eltis, Lindsay D.

2012-01-01

Bile acids are highly abundant steroids with important functions in vertebrate digestion. Their catabolism by bacteria is an important component of the carbon cycle, contributes to gut ecology, and has potential commercial applications. We found that Rhodococcus jostii RHA1 grows well on cholate, as well as on its conjugates, taurocholate and glycocholate. The transcriptome of RHA1 growing on cholate revealed 39 genes upregulated on cholate, occurring in a single gene cluster. Reverse transcriptase quantitative PCR confirmed that selected genes in the cluster were upregulated 10-fold on cholate versus on cholesterol. One of these genes, kshA3, encoding a putative 3-ketosteroid-9α-hydroxylase, was deleted and found essential for growth on cholate. Two coenzyme A (CoA) synthetases encoded in the cluster, CasG and CasI, were heterologously expressed. CasG was shown to transform cholate to cholyl-CoA, thus initiating side chain degradation. CasI was shown to form CoA derivatives of steroids with isopropanoyl side chains, likely occurring as degradation intermediates. Orthologous gene clusters were identified in all available Rhodococcus genomes, as well as that of Thermomonospora curvata. Moreover, Rhodococcus equi 103S, Rhodococcus ruber Chol-4 and Rhodococcus erythropolis SQ1 each grew on cholate. In contrast, several mycolic acid bacteria lacking the gene cluster were unable to grow on cholate. Our results demonstrate that the above-mentioned gene cluster encodes cholate catabolism and is distinct from a more widely occurring gene cluster encoding cholesterol catabolism. PMID:23024343

The feasibility of using magnetic nanoparticles modified as gene vector.

PubMed

Chen, D; Tang, Q; Xue, W; Wang, X

2010-06-01

To evaluate the feasibility of using magnetic nanoparticles (MNPs) as gene vector and the effect of magnetic field on efficiency of transfection. Magnetic nanoparticles were prepared by controlling some chemical reaction parameters through a partially reduction precipitation method with ferric chloride aqueous solution as precursor material. The surface of particles was modified by polyethyleneimine (PEI) agents. The appearance, the size distribution, structure and phase constitute of MNPs were characterized by Transmission electron microscope (TEM), X-ray diffraction (XRD); the potential of absorbing DNA of MNPs was analysed by electrophoresis. Transfection was determined by delivering reporter gene, PGL2-control encoding luciferase, to different cell lines using MNPs-PLL as vector. The effect of magnetic field on the efficiency of transfection was determined using Nd-Fe-B permanent magnet. Foreign gene could be delivered to various cell lines by MNPs-PLL and expressed with high efficiency but the transfection efficiency and time course varied in the different cell lines studied. Magnetic field could enhance the efficiency of transfection by 5-10 fold. MNPs- PLL can be used as a novel non-viral gene vector in vitro, which offers a basis for gene delivery in vivo.

Functional characterization of MAT1-1-specific mating-type genes in the homothallic ascomycete Sordaria macrospora provides new insights into essential and nonessential sexual regulators.

PubMed

Klix, V; Nowrousian, M; Ringelberg, C; Loros, J J; Dunlap, J C; Pöggeler, S

2010-06-01

Mating-type genes in fungi encode regulators of mating and sexual development. Heterothallic ascomycete species require different sets of mating-type genes to control nonself-recognition and mating of compatible partners of different mating types. Homothallic (self-fertile) species also carry mating-type genes in their genome that are essential for sexual development. To analyze the molecular basis of homothallism and the role of mating-type genes during fruiting-body development, we deleted each of the three genes, SmtA-1 (MAT1-1-1), SmtA-2 (MAT1-1-2), and SmtA-3 (MAT1-1-3), contained in the MAT1-1 part of the mating-type locus of the homothallic ascomycete species Sordaria macrospora. Phenotypic analysis of deletion mutants revealed that the PPF domain protein-encoding gene SmtA-2 is essential for sexual reproduction, whereas the alpha domain protein-encoding genes SmtA-1 and SmtA-3 play no role in fruiting-body development. By means of cross-species microarray analysis using Neurospora crassa oligonucleotide microarrays hybridized with S. macrospora targets and quantitative real-time PCR, we identified genes expressed under the control of SmtA-1 and SmtA-2. Both genes are involved in the regulation of gene expression, including that of pheromone genes.

Functional Characterization of MAT1-1-Specific Mating-Type Genes in the Homothallic Ascomycete Sordaria macrospora Provides New Insights into Essential and Nonessential Sexual Regulators▿†

PubMed Central

Klix, V.; Nowrousian, M.; Ringelberg, C.; Loros, J. J.; Dunlap, J. C.; Pöggeler, S.

2010-01-01

Mating-type genes in fungi encode regulators of mating and sexual development. Heterothallic ascomycete species require different sets of mating-type genes to control nonself-recognition and mating of compatible partners of different mating types. Homothallic (self-fertile) species also carry mating-type genes in their genome that are essential for sexual development. To analyze the molecular basis of homothallism and the role of mating-type genes during fruiting-body development, we deleted each of the three genes, SmtA-1 (MAT1-1-1), SmtA-2 (MAT1-1-2), and SmtA-3 (MAT1-1-3), contained in the MAT1-1 part of the mating-type locus of the homothallic ascomycete species Sordaria macrospora. Phenotypic analysis of deletion mutants revealed that the PPF domain protein-encoding gene SmtA-2 is essential for sexual reproduction, whereas the α domain protein-encoding genes SmtA-1 and SmtA-3 play no role in fruiting-body development. By means of cross-species microarray analysis using Neurospora crassa oligonucleotide microarrays hybridized with S. macrospora targets and quantitative real-time PCR, we identified genes expressed under the control of SmtA-1 and SmtA-2. Both genes are involved in the regulation of gene expression, including that of pheromone genes. PMID:20435701

Structure of the Elastin-Contractile Units in the Thoracic Aorta and How Genes That Cause Thoracic Aortic Aneurysms and Dissections Disrupt This Structure.

PubMed

Karimi, Ashkan; Milewicz, Dianna M

2016-01-01

The medial layer of the aorta confers elasticity and strength to the aortic wall and is composed of alternating layers of smooth muscle cells (SMCs) and elastic fibres. The SMC elastin-contractile unit is a structural unit that links the elastin fibres to the SMCs and is characterized by the following: (1) layers of elastin fibres that are surrounded by microfibrils; (2) microfibrils that bind to the integrin receptors in focal adhesions on the cell surface of the SMCs; and (3) SMC contractile filaments that are linked to the focal adhesions on the inner side of the membrane. The genes that are altered to cause thoracic aortic aneurysms and aortic dissections encode proteins involved in the structure or function of the SMC elastin-contractile unit. Included in this gene list are the genes encoding protein that are structural components of elastin fibres and microfibrils, FBN1, MFAP5, ELN, and FBLN4. Also included are genes that encode structural proteins in the SMC contractile unit, including ACTA2, which encodes SMC-specific α-actin and MYH11, which encodes SMC-specific myosin heavy chain, along with MYLK and PRKG1, which encode kinases that control SMC contraction. Finally, mutations in the gene encoding the protein linking integrin receptors to the contractile filaments, FLNA, also predispose to thoracic aortic disease. Thus, these data suggest that functional SMC elastin-contractile units are important for maintaining the structural integrity of the aorta. Copyright © 2016 Canadian Cardiovascular Society. Published by Elsevier Inc. All rights reserved.

Detection of β-lactamase encoding genes in feces, soil and water from a Brazilian pig farm.

PubMed

Furlan, João Pedro Rueda; Stehling, Eliana Guedes

2018-01-10

β-lactam antibiotics are widely used for the treatment of different types of infections worldwide and the resistance to these antibiotics has grown sharply, which is of great concern. Resistance to β-lactams in gram-negative bacteria is mainly due to the production of β-lactamases, which are classified according to their functional activities. The aim of this study was to verify the presence of β-lactamases encoding genes in feces, soil, and water from a Brazilian pig farm. Different β-lactamases encoding genes were found, including bla CTX-M-Gp1 , bla CTX-M-Gp9 , bla SHV , bla OXA-1-like , bla GES , and bla VEB . The bla SHV and bla CTX-M-Gp1 genes have been detected in all types of samples, indicating the spread of β-lactam resistant bacteria among farm pigs and the environment around them. These results indicate that β-lactamase encoding genes belonging to the cloxacillinase, ESBL, and carbapenemase and they have high potential to spread in different sources, due to the fact that genes are closely related to mobile genetic elements, especially plasmids.

Genome-Wide Identification and Mapping of NBS-Encoding Resistance Genes in Solanum tuberosum Group Phureja

PubMed Central

Lozano, Roberto; Ponce, Olga; Ramirez, Manuel; Mostajo, Nelly; Orjeda, Gisella

2012-01-01

The majority of disease resistance (R) genes identified to date in plants encode a nucleotide-binding site (NBS) and leucine-rich repeat (LRR) domain containing protein. Additional domains such as coiled-coil (CC) and TOLL/interleukin-1 receptor (TIR) domains can also be present. In the recently sequenced Solanum tuberosum group phureja genome we used HMM models and manual curation to annotate 435 NBS-encoding R gene homologs and 142 NBS-derived genes that lack the NBS domain. Highly similar homologs for most previously documented Solanaceae R genes were identified. A surprising ∼41% (179) of the 435 NBS-encoding genes are pseudogenes primarily caused by premature stop codons or frameshift mutations. Alignment of 81.80% of the 577 homologs to S. tuberosum group phureja pseudomolecules revealed non-random distribution of the R-genes; 362 of 470 genes were found in high density clusters on 11 chromosomes. PMID:22493716

Mutations of the Norrie gene in Korean ROP infants.

PubMed

Kim, Jeong Hun; Yu, Young Suk; Kim, Jiyeon; Park, Seong Sup

2002-12-01

The present study was conducted to evaluate if there is a Norrie disease gene (ND gene) mutation involved in the retinopathy of prematurity (ROP), and to identify the possibility of a genetic abnormality that may be linked to the presence of ROP. Nineteen premature Korean infants, with a low birth weight (1500 g or less) or low gestational age (32 weeks or less), were included in the study. Eighteen infants had ROP, and the other did not. Genomic DNA was isolated from the peripheral blood leukocytes of these patients, and all three exons and their flanking areas, all known ND gene mutations regions, were evaluated following amplification by a polymerase chain reaction, but no ND gene mutations were detected. Any disagreement between the relationship of ROP to the ND gene mutation will need to be clarified by further investigation.

Bioinformatics analysis and detection of gelatinase encoded gene in Lysinibacillussphaericus

NASA Astrophysics Data System (ADS)

Repin, Rul Aisyah Mat; Mutalib, Sahilah Abdul; Shahimi, Safiyyah; Khalid, Rozida Mohd.; Ayob, Mohd. Khan; Bakar, Mohd. Faizal Abu; Isa, Mohd Noor Mat

2016-11-01

In this study, we performed bioinformatics analysis toward genome sequence of Lysinibacillussphaericus (L. sphaericus) to determine gene encoded for gelatinase. L. sphaericus was isolated from soil and gelatinase species-specific bacterium to porcine and bovine gelatin. This bacterium offers the possibility of enzymes production which is specific to both species of meat, respectively. The main focus of this research is to identify the gelatinase encoded gene within the bacteria of L. Sphaericus using bioinformatics analysis of partially sequence genome. From the research study, three candidate gene were identified which was, gelatinase candidate gene 1 (P1), NODE_71_length_93919_cov_158.931839_21 which containing 1563 base pair (bp) in size with 520 amino acids sequence; Secondly, gelatinase candidate gene 2 (P2), NODE_23_length_52851_cov_190.061386_17 which containing 1776 bp in size with 591 amino acids sequence; and Thirdly, gelatinase candidate gene 3 (P3), NODE_106_length_32943_cov_169.147919_8 containing 1701 bp in size with 566 amino acids sequence. Three pairs of oligonucleotide primers were designed and namely as, F1, R1, F2, R2, F3 and R3 were targeted short sequences of cDNA by PCR. The amplicons were reliably results in 1563 bp in size for candidate gene P1 and 1701 bp in size for candidate gene P3. Therefore, the results of bioinformatics analysis of L. Sphaericus resulting in gene encoded gelatinase were identified.

PUM1 is a biphasic negative regulator of innate immunity genes by suppressing LGP2.

PubMed

Liu, Yonghong; Qu, Linlin; Liu, Yuanyuan; Roizman, Bernard; Zhou, Grace Guoying

2017-08-15

PUM1 is an RNA binding protein shown to regulate the stability and function of mRNAs bearing a specific sequence. We report the following: ( i ) A key function of PUM1 is that of a repressor of key innate immunity genes by repressing the expression of LGP2. Thus, between 12 and 48 hours after transfection of human cells with siPUM1 RNA there was an initial (phase 1) upsurge of transcripts encoding LGP2, CXCL10, IL6, and PKR. This was followed 24 hours later (phase 2) by a significant accumulation of mRNAs encoding RIG-I, SP100, MDA5, IFIT1, PML, STING, and IFNβ. The genes that were not activated encoded HDAC4 and NF-κB1. ( ii ) Simultaneous depletion of PUM1 and LGP2, CXCL10, or IL6 revealed that up-regulation of phase 1 and phase 2 genes was the consequence of up-regulation of LGP2. ( iii ) IFNβ produced 48-72 hours after transfection of siPUM1 was effective in up-regulating LGP2 and phase 2 genes and reducing the replication of HSV-1 in untreated cells. ( iv ) Because only half of genes up-regulated in phase 1 and 2 encode mRNAs containing PUM1 binding sites, the upsurge in gene expression could not be attributed solely to stabilization of mRNAs in the absence of PUM1. ( v ) Lastly, depletion of PUM2 does not result in up-regulation of phase 1 or phase 2 genes. The results of the studies presented here indicate that PUM1 is a negative regulator of LGP2, a master regulator of innate immunity genes expressed in a cascade fashion.

The genetic signature of recent speciation in manta rays (Manta alfredi and M. birostris).

PubMed

Kashiwagi, Tom; Marshall, Andrea D; Bennett, Michael B; Ovenden, Jennifer R

2012-07-01

Manta rays have been taxonomically revised as two species, Manta alfredi and M. birostris, on the basis of morphological and meristic data, yet the two species occur in extensive mosaic sympatry. We analysed the genetic signatures of the species boundary using a portion of the nuclear RAG1 (681 base pairs), mitochondrial CO1 (574 bp) and ND5 genes (1188 bp). The assay with CO1 sequences, widely used in DNA barcoding, failed to distinguish the two species. The two species were clearly distinguishable, however, with no shared RAG1 or ND5 haplotypes. The species were reciprocally monophyletic for RAG1, but paraphyletic for ND5 sequences. Qualitative evidence and statistical inferences using the 'Isolation-with-Migration models' indicated that these results were better explained with post-divergence gene flow in the recent past rather than incomplete lineage sorting with zero gene flow since speciation. An estimate of divergence time was less than 0.5 Ma with an upper confidence limit of within 1 Ma. Recent speciation of highly mobile species in the marine environment is of great interest, as it suggests that speciation may have occurred in the absence of long-term physical barriers to gene flow. We propose that the ecologically driven forces such as habitat choice played a significant role in speciation in manta rays. Copyright © 2012 Elsevier Inc. All rights reserved.

Mitochondrial Gene Therapy: Advances in Mitochondrial Gene Cloning, Plasmid Production, and Nanosystems Targeted to Mitochondria.

PubMed

Coutinho, Eduarda; Batista, Cátia; Sousa, Fani; Queiroz, João; Costa, Diana

2017-03-06

Mitochondrial gene therapy seems to be a valuable and promising strategy to treat mitochondrial disorders. The use of a therapeutic vector based on mitochondrial DNA, along with its affinity to the site of mitochondria, can be considered a powerful tool in the reestablishment of normal mitochondrial function. In line with this and for the first time, we successfully cloned the mitochondrial gene ND1 that was stably maintained in multicopy pCAG-GFP plasmid, which is used to transform E. coli. This mitochondrial-gene-based plasmid was encapsulated into nanoparticles. Furthermore, the functionalization of nanoparticles with polymers, such as cellulose or gelatin, enhances their overall properties and performance for gene therapy. The fluorescence arising from rhodamine nanoparticles in mitochondria and a fluorescence microscopy study show pCAG-GFP-ND1-based nanoparticles' cell internalization and mitochondria targeting. The quantification of GFP expression strongly supports this finding. This work highlights the viability of gene therapy based on mitochondrial DNA instigating further in vitro research and clinical translation.

Impact assessment of bisphenol A on lignin-modifying enzymes by basidiomycete Trametes versicolor.

PubMed

Takamiya, Minako; Magan, Naresh; Warner, Philip J

2008-06-15

The impact of different concentrations of bisphenol A (BPA) was evaluated on growth of the white-rot basidiomycete, Trametes versicolor, and on the expression of genes encoding lignin-modifying enzyme (LME) activities. Effective doses (EDs) were obtained from fungal growth rate to monitor LME activities and the expression levels of their encoding genes. The fungus showed mycelial growth at concentrations of up to 300 microg ml(-1) of BPA with an ED50 value of 185 microg ml(-1). The LME activities were stimulated by BPA concentrations up to 300 microg ml(-1). The lignin peroxidase (LIP) encoding gene may be sensitive to BPA stress.

Two Bombyx mori acetylcholinesterase genes influence motor control and development in different ways

USDA-ARS?s Scientific Manuscript database

Among its other biological roles, acetylcholinesterase (AChE, EC 3.1.1.7), encoded by two ace genes in most insects, catalyses the breakdown of acetylcholine, thereby terminating synaptic transmission. ace1 encodes the synaptic enzyme and ace2 has other essential actions in many insect species, such...

Insertion and deletion mutations in the dinucleotide repeat region of the Norrie disease gene in patients with advanced retinopathy of prematurity.

PubMed

Hiraoka, M; Berinstein, D M; Trese, M T; Shastry, B S

2001-01-01

Retinopathy of prematurity (ROP) is a leading cause of blindness in premature children. It is a multifactorial disorder which causes fibrovascular tissue changes that affect the retina in low birth-weight and short gestational age infants. To determine the prevalence of Norrie disease (ND) gene mutations, clinical examination and molecular genetic analyses were performed in 100 pre-term babies of different ethnic backgrounds who developed advanced ROP. The leukocyte DNA was extracted, amplified by the polymerase chain reaction (PCR), and analyzed by single-strand conformation polymorphism (SSCP), G/T and C/A scanning, and by DNA sequencing. All three exons, including splice sites and the 3'-untranslated region, were screened. Of the 100 patients analyzed, 2 patients with advanced ROP showed a mobility shift in the DNA. In 1 patient, this mobility shift was caused by the insertion of an additional 12-bp CT repeat in exon 1, and in the second patient, there was a 14-bp deletion in the same exon of the ND gene, as evidenced by direct sequencing of the amplified products. Similar analyses of exons 2 and 3 and the 3'-untranslated region failed to detect additional mutations in the gene. None of the 130 normal, unrelated controls revealed similar changes. Taking into account the above results, as well as those of other studies, it appears that the ND gene mutations can account for 3% of cases of advanced ROP. Although the ND gene is not frequently involved in advanced ROP, the present large-scale study further supports the hypothesis that genetic influences may play an important role in the development of severe ROP in some premature infants.

Interactive Effects of Dietary Lipid and Phenotypic Feed Efficiency on the Expression of Nuclear and Mitochondrial Genes Involved in the Mitochondrial Electron Transport Chain in Rainbow Trout

PubMed Central

Eya, Jonathan C.; Ukwuaba, Vitalis O.; Yossa, Rodrigue; Gannam, Ann L.

2015-01-01

A 2 × 3 factorial study was conducted to evaluate the effects of dietary lipid level on the expression of mitochondrial and nuclear genes involved in electron transport chain in all-female rainbow trout Oncorhynchus mykiss. Three practical diets with a fixed crude protein content of 40%, formulated to contain 10% (40/10), 20% (40/20) and 30% (40/30) dietary lipid, were fed to apparent satiety to triplicate groups of either low-feed efficient (F120; 217.66 ± 2.24 g initial average mass) or high-feed efficient (F136; 205.47 ± 1.27 g) full-sib families of fish, twice per day, for 90 days. At the end of the experiment, the results showed that there is an interactive effect of the dietary lipid levels and the phenotypic feed efficiency (growth rate and feed efficiency) on the expression of the mitochondrial genes nd1 (NADH dehydrogenase subunit 1), cytb (Cytochrome b), cox1 (Cytochrome c oxidase subunits 1), cox2 (Cytochrome c oxidase subunits 2) and atp6 (ATP synthase subunit 6) and nuclear genes ucp2α (uncoupling proteins 2 alpha), ucp2β (uncoupling proteins 2 beta), pparα (peroxisome proliferator-activated receptor alpha), pparβ (peroxisome proliferatoractivated receptor beta) and ppargc1α (proliferator-activated receptor gamma coactivator 1 alpha) in fish liver, intestine and muscle, except on ppargc1α in the muscle which was affected by the diet and the family separately. Also, the results revealed that the expression of mitochondrial genes is associated with that of nuclear genes involved in electron transport chain in fish liver, intestine and muscle. Furthermore, this work showed that the expression of mitochondrial genes parallels with the expression of genes encoding uncoupling proteins (UCP) in the liver and the intestine of rainbow trout. This study for the first time presents the molecular basis of the effects of dietary lipid level on mitochondrial and nuclear genes involved in mitochondrial electron transport chain in fish. PMID:25853266

The phosphoenolpyruvate/phosphate translocator is required for phenolic metabolism, palisade cell development, and plastid-dependent nuclear gene expression.

PubMed

Streatfield, S J; Weber, A; Kinsman, E A; Häusler, R E; Li, J; Post-Beittenmiller, D; Kaiser, W M; Pyke, K A; Flügge, U I; Chory, J

1999-09-01

The Arabidopsis chlorophyll a/b binding protein (CAB) gene underexpressed 1 (cue1) mutant underexpresses light-regulated nuclear genes encoding chloroplast-localized proteins. cue1 also exhibits mesophyll-specific chloroplast and cellular defects, resulting in reticulate leaves. Both the gene underexpression and the leaf cell morphology phenotypes are dependent on light intensity. In this study, we determine that CUE1 encodes the plastid inner envelope phosphoenolpyruvate/phosphate translocator (PPT) and define amino acid residues that are critical for translocator function. The biosynthesis of aromatics is compromised in cue1, and the reticulate phenotype can be rescued by feeding aromatic amino acids. Determining that CUE1 encodes PPT indicates the in vivo role of the translocator in metabolic partitioning and reveals a mesophyll cell-specific requirement for the translocator in Arabidopsis leaves. The nuclear gene expression defects in cue1 suggest that a light intensity-dependent interorganellar signal is modulated through metabolites dependent on a plastid supply of phosphoenolpyruvate.

Identification of a Polymorphic Gene, BCL2A1, Encoding Two Novel Hematopoietic Lineage-specific Minor Histocompatibility Antigens

PubMed Central

Akatsuka, Yoshiki; Nishida, Tetsuya; Kondo, Eisei; Miyazaki, Mikinori; Taji, Hirohumi; Iida, Hiroatsu; Tsujimura, Kunio; Yazaki, Makoto; Naoe, Tomoki; Morishima, Yasuo; Kodera, Yoshihisa; Kuzushima, Kiyotaka; Takahashi, Toshitada

2003-01-01

We report the identification of two novel minor histocompatibility antigens (mHAgs), encoded by two separate single nucleotide polymorphisms on a single gene, BCL2A1, and restricted by human histocompatibility leukocyte antigen (HLA)-A*2402 (the most common HLA-A allele in Japanese) and B*4403, respectively. Two cytotoxic T lymphocyte (CTL) clones specific for these mHAgs were first isolated from two distinct recipients after hematopoietic cell transplantation. Both clones lyse only normal and malignant cells within the hematopoietic lineage. To localize the gene encoding the mHAgs, two-point linkage analysis was performed on the CTL lytic patterns of restricting HLA-transfected B lymphoblastoid cell lines obtained from Centre d'Etude du Polymorphisme Humain. Both CTL clones showed a completely identical lytic pattern for 4 pedigrees and the gene was localized within a 3.6-cM interval of 15q24.3–25.1 region that encodes at least 46 genes. Of those, only BCL2A1 has been reported to be expressed in hematopoietic cells and possess three nonsynonymous nucleotide changes. Minigene transfection and epitope reconstitution assays with synthetic peptides identified both HLA-A*2402– and B*4403-restricted mHAg epitopes to be encoded by distinct polymorphisms within BCL2A1. PMID:12771180

IQCJ-SCHIP1, a novel fusion transcript encoding a calmodulin-binding IQ motif protein

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

Kwasnicka-Crawford, Dorota A.; Carson, Andrew R.; Scherer, Stephen W.

The existence of transcripts that span two adjacent, independent genes is considered rare in the human genome. This study characterizes a novel human fusion gene named IQCJ-SCHIP1. IQCJ-SCHIP1 is the longest isoform of a complex transcriptional unit that bridges two separate genes that encode distinct proteins, IQCJ, a novel IQ motif containing protein and SCHIP1, a schwannomin interacting protein that has been previously shown to interact with the Neurofibromatosis type 2 (NF2) protein. IQCJ-SCHIP1 is located on the chromosome 3q25 and comprises a 1692-bp transcript encompassing 11 exons spanning 828 kb of the genomic DNA. We show that IQCJ-SCHIP1 mRNAmore » is highly expressed in the brain. Protein encoded by the IQCJ-SCHIP1 gene was localized to cytoplasm and actin-rich regions and in differentiated PC12 cells was also seen in neurite extensions.« less

Cloning and sequence analysis of the LEU2 homologue gene from Pichia anomala.

PubMed

De la Rosa, J M; Pérez, J A; Gutiérrez, F; González, J M; Ruiz, T; Rodríguez, L

2001-11-01

The Pichia anomala LEU2 gene (PaLEU2) was isolated by complementation of a leu2 Saccharomyces cerevisiae mutant. The cloned gene also allowed growth of a Escherichia coli leuB mutant in leucine-lacking medium, indicating that it encodes a product able to complement the beta-isopropylmalate dehydrogenase deficiency of the mutants. The sequenced DNA fragment contains a complete ORF of 1092 bp, and the deduced polypeptide shares significant homologies with the products of the LEU2 genes from S. cerevisiae (84% identity) and other yeast species. A sequence resembling the GC-rich palindrome motif identified in the 5' region of S. cerevisiae LEU2 gene as the binding site for the transcription activating factor encoded by the LEU3 gene was found at the promoter region. In addition, upstream of the PaLEU2 the 3'-terminal half of a gene of the same orientation, encoding a homologue of the S. cerevisiae NFS1/SPL1 gene that encodes a mitochondrial cysteine desulphurase involved in both tRNA processing and mitochondrial metabolism, was found. The genomic organization of the PaNFS1-PaLEU2 gene pair is similar to that found in several other yeast species, including S. cerevisiae and Candida albicans, except that in some of them the LEU2 gene appears in the reverse orientation. Copyright 2001 John Wiley & Sons, Ltd.

Phorbol 12-myristate 13-acetate enhances nm23 gene expression in murine melanocytes but not in syngeneic B16-BL6 melanoma variants.

PubMed

Huijzer, J C; McFarland, M; Niles, R M; Meadows, G G

1996-03-01

The nm23 gene has been described as a potential metastasis suppressor gene in certain rodent and human tumors. We previously demonstrated that tyrosine and phenylalanine restriction suppresses metastatic heterogeneity of B16-BL6 murine melanoma and selects for tumor variants with decreased metastatic potential. In this study, we investigated nm23 expression in the highly metastatic B16-BL6 (ND) melanoma, its nutritionally derived poorly metastatic (LT) variant, and the syngeneic non-tumorigenic Mel-ab melanocytes. No differences in nm23 expression were observed between ND and LT cells, and nm23 expression varied between different isolates. Previously, we showed that metastatic potential of 1-ND cells decreases and is not altered in 1-LT cells after prolonged in vitro cell passage; however, nm23 expression is equivalently increased by 2-fold. In 2-ND and 2-LT cells, expression of nm23 is not different at higher in vitro cell passage. Expression of nm23 decreased about 2-fold when phorbol 12-myristate 13-acetate (PMA) was removed from Mel-ab cells, which induces these cells to become quiescent. Although membrane-associated protein kinase C (PKC) activity decreased after prolonged PMA treatment in all cells, neither nm23 expression nor proliferation of ND and LT cells was affected by PMA. These data indicate that nm23 expression is related to proliferative activity rather than to the suppression of metastatic potential.

Molecular comparison of the structural proteins encoding gene clusters of two related Lactobacillus delbrueckii bacteriophages.

PubMed Central

Vasala, A; Dupont, L; Baumann, M; Ritzenthaler, P; Alatossava, T

1993-01-01

Virulent phage LL-H and temperate phage mv4 are two related bacteriophages of Lactobacillus delbrueckii. The gene clusters encoding structural proteins of these two phages have been sequenced and further analyzed. Six open reading frames (ORF-1 to ORF-6) were detected. Protein sequencing and Western immunoblotting experiments confirmed that ORF-3 (g34) encoded the main capsid protein Gp34. The presence of a putative late promoter in front of the phage LL-H g34 gene was suggested by primer extension experiments. Comparative sequence analysis between phage LL-H and phage mv4 revealed striking similarities in the structure and organization of this gene cluster, suggesting that the genes encoding phage structural proteins belong to a highly conservative module. Images PMID:8497043

CovRS-Regulated Transcriptome Analysis of a Hypervirulent M23 Strain of Group A Streptococcus pyogenes Provides New Insights into Virulence Determinants.

PubMed

Bao, Yun-Juan; Liang, Zhong; Mayfield, Jeffrey A; Lee, Shaun W; Ploplis, Victoria A; Castellino, Francis J

2015-10-01

The two-component control of virulence (Cov) regulator (R)-sensor (S) (CovRS) regulates the virulence of Streptococcus pyogenes (group A Streptococcus [GAS]). Inactivation of CovS during infection switches the pathogenicity of GAS to a more invasive form by regulating transcription of diverse virulence genes via CovR. However, the manner in which CovRS controls virulence through expression of extended gene families has not been fully determined. In the current study, the CovS-regulated gene expression profiles of a hypervirulent emm23 GAS strain (M23ND/CovS negative [M23ND/CovS(-)]) and a noninvasive isogenic strain (M23ND/CovS(+)), under different growth conditions, were investigated. RNA sequencing identified altered expression of ∼ 349 genes (18% of the chromosome). The data demonstrated that M23ND/CovS(-) achieved hypervirulence by allowing enhanced expression of genes responsible for antiphagocytosis (e.g., hasABC), by abrogating expression of toxin genes (e.g., speB), and by compromising gene products with dispensable functions (e.g., sfb1). Among these genes, several (e.g., parE and parC) were not previously reported to be regulated by CovRS. Furthermore, the study revealed that CovS also modulated the expression of a broad spectrum of metabolic genes that maximized nutrient utilization and energy metabolism during growth and dissemination, where the bacteria encounter large variations in available nutrients, thus restructuring metabolism of GAS for adaption to diverse growth environments. From constructing a genome-scale metabolic model, we identified 16 nonredundant metabolic gene modules that constitute unique nutrient sources. These genes were proposed to be essential for pathogen growth and are likely associated with GAS virulence. The genome-wide prediction of genes associated with virulence identifies new candidate genes that potentially contribute to GAS virulence. The CovRS system modulates transcription of ∼ 18% of the genes in the Streptococcus pyogenes genome. Mutations that inactivate CovR or CovS enhance the virulence of this bacterium. We determined complete transcriptomes of a naturally CovS-inactivated invasive deep tissue isolate of an emm23 strain of S. pyogenes (M23ND) and its complemented avirulent variant (CovS(+)). We identified diverse virulence genes whose altered expression revealed a genetic switching of a nonvirulent form of M23ND to a highly virulent strain. Furthermore, we also systematically uncovered for the first time the comparative levels of expression of a broad spectrum of metabolic genes, which reflected different metabolic needs of the bacterium as it invaded deeper tissue of the human host. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Amino acid substitutions in the VanS sensor of the VanA-type vancomycin-resistant Enterococcus strains result in high-level vancomycin resistance and low-level teicoplanin resistance.

PubMed

Hashimoto, Y; Tanimoto, K; Ozawa, Y; Murata, T; Ike, Y

2000-04-15

The vancomycin-resistant enterococci GV1, GV2 and GV3, which were isolated from droppings from broiler farms in Japan have been characterized as VanA-type VRE, which express high-level vancomycin resistance (256 or 512 microg ml(-1), MIC) and low-level teicoplanin resistance (1 or 2 microg ml(-1), MIC). The vancomycin resistances were encoded on plasmids. The vancomycin resistance conjugative plasmid pMG2 was isolated from the GV2 strain. The VanA determinant of pMG2 showed the same genetic organization as that of the VanA genes encoded on the representative transposon Tn1546, which comprises vanRSHAXYZ. The nucleotide sequences of all the genes, except the gene related to the vanS gene on Tn1546, were completely identical to the genes encoded on Tn1546. Three amino acid substitutions in the N-terminal region of the deduced VanS were detected in the nucleotide sequence of vanS encoded on pMG2. There were also three amino acid substitutions in the vanS gene of the GV1 and GV3 strains in the same positions as in the vanS gene of pMG2. Vancomycin induced the increased teicoplanin resistance in these strains.

Elucidating the genotype-phenotype relationships and network perturbations of human shared and specific disease genes from an evolutionary perspective.

PubMed

Begum, Tina; Ghosh, Tapash Chandra

2014-10-05

To date, numerous studies have been attempted to determine the extent of variation in evolutionary rates between human disease and nondisease (ND) genes. In our present study, we have considered human autosomal monogenic (Mendelian) disease genes, which were classified into two groups according to the number of phenotypic defects, that is, specific disease (SPD) gene (one gene: one defect) and shared disease (SHD) gene (one gene: multiple defects). Here, we have compared the evolutionary rates of these two groups of genes, that is, SPD genes and SHD genes with respect to ND genes. We observed that the average evolutionary rates are slow in SHD group, intermediate in SPD group, and fast in ND group. Group-to-group evolutionary rate differences remain statistically significant regardless of their gene expression levels and number of defects. We demonstrated that disease genes are under strong selective constraint if they emerge through edgetic perturbation or drug-induced perturbation of the interactome network, show tissue-restricted expression, and are involved in transmembrane transport. Among all the factors, our regression analyses interestingly suggest the independent effects of 1) drug-induced perturbation and 2) the interaction term of expression breadth and transmembrane transport on protein evolutionary rates. We reasoned that the drug-induced network disruption is a combination of several edgetic perturbations and, thus, has more severe effect on gene phenotypes. © The Author(s) 2014. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

Identification of unique expression signatures and therapeutic targets in esophageal squamous cell carcinoma

PubMed Central

2012-01-01

Background Esophageal squamous cell carcinoma (ESCC), the predominant histological subtype of esophageal cancer, is characterized by high mortality. Previous work identified important mRNA expression differences between normal and tumor cells; however, to date there are limited ex vivo studies examining expression changes occurring during normal esophageal squamous cell differentiation versus those associated with tumorigenesis. In this study, we used a unique tissue microdissection strategy and microarrays to measure gene expression profiles associated with cell differentiation versus tumorigenesis in twelve cases of patient-matched normal basal squamous epithelial cells (NB), normal differentiated squamous epithelium (ND), and squamous cell cancer. Class comparison and pathway analysis were used to compare NB versus tumor in a search for unique therapeutic targets. Results As a first step towards this goal, gene expression profiles and pathways were evaluated. Overall, ND expression patterns were markedly different from NB and tumor; whereas, tumor and NB were more closely related. Tumor showed a general decrease in differentially expressed genes relative to NB as opposed to ND that exhibited the opposite trend. FSH and IgG networks were most highly dysregulated in normal differentiation and tumorigenesis, respectively. DNA repair pathways were generally elevated in NB and tumor relative to ND indicating involvement in both normal and pathological growth. PDGF signaling pathway and 12 individual genes unique to the tumor/NB comparison were identified as therapeutic targets, and 10 associated ESCC gene-drug pairs were identified. We further examined the protein expression level and the distribution patterns of four genes: ODC1, POSTN, ASPA and IGF2BP3. Ultimately, three genes (ODC1, POSTN, ASPA) were verified to be dysregulated in the same pattern at both the mRNA and protein levels. Conclusions These data reveal insight into genes and molecular pathways mediating ESCC development and provide information potentially useful in designing novel therapeutic interventions for this tumor type. PMID:22280838

Detection with synthetic oligonucleotide probes of nucleotide sequence variations in the genes encoding enterotoxins of Escherichia coli.

PubMed Central

Nishibuchi, M; Murakami, A; Arita, M; Jikuya, H; Takano, J; Honda, T; Miwatani, T

1989-01-01

We examined variations in the genes encoding heat-stable enterotoxin (ST) and heat-labile enterotoxin (LT) in 88 strains of Escherichia coli isolated from individuals with traveler's diarrhea to find suitable sequences for use as oligonucleotide probes. Four oligonucleotide probes of the gene encoding ST of human origin (STIb or STh), one oligonucleotide probe of the gene encoding ST of porcine origin (STIa or STp), and three oligonucleotide probes of the gene encoding LT of human origin (LTIh) were used in DNA colony hybridization tests. In 15 of 22 strains possessing the STh gene and 28 of 42 strains producing LT, the sequences of all regions tested were identical to the published sequences. One region in the STh gene examined with a 18-mer probe was relatively well conserved and was shown to be closely associated with the enterotoxicity of the E. coli strains in suckling mice. This oligonucleotide, however, hybridized with strains of Vibrio cholerae O1, V. parahaemolyticus, and Yersinia enterocolitica that gave negative results in the suckling mouse assay. PMID:2685027

Deregulation of Rab and Rab Effector Genes in Bladder Cancer

PubMed Central

Ho, Joel R.; Chapeaublanc, Elodie; Kirkwood, Lisa; Nicolle, Remy; Benhamou, Simone; Lebret, Thierry; Allory, Yves; Southgate, Jennifer; Radvanyi, François; Goud, Bruno

2012-01-01

Growing evidence indicates that Rab GTPases, key regulators of intracellular transport in eukaryotic cells, play an important role in cancer. We analysed the deregulation at the transcriptional level of the genes encoding Rab proteins and Rab-interacting proteins in bladder cancer pathogenesis, distinguishing between the two main progression pathways so far identified in bladder cancer: the Ta pathway characterized by a high frequency of FGFR3 mutation and the carcinoma in situ pathway where no or infrequent FGFR3 mutations have been identified. A systematic literature search identified 61 genes encoding Rab proteins and 223 genes encoding Rab-interacting proteins. Transcriptomic data were obtained for normal urothelium samples and for two independent bladder cancer data sets corresponding to 152 and 75 tumors. Gene deregulation was analysed with the SAM (significant analysis of microarray) test or the binomial test. Overall, 30 genes were down-regulated, and 13 were up-regulated in the tumor samples. Five of these deregulated genes (LEPRE1, MICAL2, RAB23, STXBP1, SYTL1) were specifically deregulated in FGFR3-non-mutated muscle-invasive tumors. No gene encoding a Rab or Rab-interacting protein was found to be specifically deregulated in FGFR3-mutated tumors. Cluster analysis showed that the RAB27 gene cluster (comprising the genes encoding RAB27 and its interacting partners) was deregulated and that this deregulation was associated with both pathways of bladder cancer pathogenesis. Finally, we found that the expression of KIF20A and ZWINT was associated with that of proliferation markers and that the expression of MLPH, MYO5B, RAB11A, RAB11FIP1, RAB20 and SYTL2 was associated with that of urothelial cell differentiation markers. This systematic analysis of Rab and Rab effector gene deregulation in bladder cancer, taking relevant tumor subgroups into account, provides insight into the possible roles of Rab proteins and their effectors in bladder cancer pathogenesis. This approach is applicable to other group of genes and types of cancer. PMID:22724020

Increased mitochondrial-encoded gene transcription in immortal DF-1 cells.

PubMed

Kim, H; You, S; Kim, I J; Farris, J; Foster, L K; Foster, D N

2001-05-01

We have established, in continuous cell culture, a spontaneously immortalized chicken embryo fibroblast (CEF) cell line (DF-1) as well as several other immortal CEF cell lines. The immortal DF-1 cells divided more rapidly than primary and other immortal CEF cells. To identify the genes involved in rapidly dividing DF-1 cells, we have used differential display RT-PCR. Of the numerous genes analyzed, three mitochondrial-encoded genes (ATPase 8/6, 16S rRNA, and cytochrome b) were shown to express at higher levels in DF-1 cells compared to primary and other immortal CEF cells. The inhibition of mitochondrial translation by treatment with chloramphenicol markedly decreased ATP production and cell proliferation in DF-1 cells, while not affecting growth in either primary or other immortal CEF cells. This result suggests a correlation between rapid cell proliferation and the increased mitochondrial respiratory functions. We also determined that the increased transcription of mitochondrial-encoded genes in DF-1 cells is due to increased de novo transcript synthesis as shown by mitochondrial run-on assays, and not the result of either increased mitochondrial biogenesis or mitochondrial transcript half-lives. Together, the present studies suggest that the transcriptional activation of mitochondrial-encoded genes and the elevated respiratory function should be one of the characteristics of rapidly dividing immortal cells. Copyright 2001 Academic Press.

Polyamidoamine-Decorated Nanodiamonds as a Hybrid Gene Delivery Vector and siRNA Structural Characterization at the Charged Interfaces.

PubMed

Lim, Dae Gon; Rajasekaran, Nirmal; Lee, Dukhee; Kim, Nam Ah; Jung, Hun Soon; Hong, Sungyoul; Shin, Young Kee; Kang, Eunah; Jeong, Seong Hoon

2017-09-20

Nanodiamonds have been discovered as a new exogenous material source in biomedical applications. As a new potent form of nanodiamond (ND), polyamidoamine-decorated nanodiamonds (PAMAM-NDs) were prepared for E7 or E6 oncoprotein-suppressing siRNA gene delivery for high risk human papillomavirus-induced cervical cancer, such as types 16 and 18. It is critical to understand the physicochemical properties of siRNA complexes immobilized on cationic solid ND surfaces in the aspect of biomolecular structural and conformational changes, as the new inert carbon material can be extended into the application of a gene delivery vector. A spectral study of siRNA/PAMAM-ND complexes using differential scanning calorimetry and circular dichroism spectroscopy proved that the hydrogen bonding and electrostatic interactions between siRNA and PAMAM-NDs decreased endothermic heat capacity. Moreover, siRNA/PAMAM-ND complexes showed low cell cytotoxicity and significant suppressing effects for forward target E6 and E7 oncogenic genes, proving functional and therapeutic efficacy. The cellular uptake of siRNA/PAMAM-ND complexes at 8 h was visualized by macropinocytes and direct endosomal escape of the siRNA/PAMAM-ND complexes. It is presumed that PAMAM-NDs provided a buffering cushion to adjust the pH and hard mechanical stress to escape endosomes. siRNA/PAMAM-ND complexes provide a potential organic/inorganic hybrid material source for gene delivery carriers.

A Novel Mitochondrial DNA Deletion in Patient with Pearson Syndrome.

PubMed

Khasawneh, Rame; Alsokhni, Hala; Alzghoul, Bayan; Momani, Asim; Abualsheikh, Nazih; Kamal, Nazmi; Qatawneh, Mousa

2018-04-01

Arteriovenous Pearson syndrome is a very rare multisystemic mitochondrial disease characterized by sideroblastic anemia and exocrine pancreatic insufficiency. It is usually fatal in infancy. We reported a four-month-old infant presented with fever and pancytopenia. Bone marrow examination showed hypoplastic changes and sideroblastic features. Molecular Study showed a novel hetroplasmic mitochondrial deletions (m. 10760 -m. 15889+) in multiple genes (ND4,ND5,ND6, CYTB). In our patient the pathogenic mutation was 5.1 kb heteroplasmic deletions in multiple genes that are important and crucial for intact oxidative phosphorylation pathway and ATP production in the mitochondrial DNA. This mutation was not reported in literature including the mitomap.org website (which was last edited on Nov 30, 2017 and accessed on Jan 13, 2018).

The bglA Gene of Aspergillus kawachii Encodes Both Extracellular and Cell Wall-Bound β-Glucosidases

PubMed Central

Iwashita, Kazuhiro; Nagahara, Tatsuya; Kimura, Hitoshi; Takano, Makoto; Shimoi, Hitoshi; Ito, Kiyoshi

1999-01-01

We cloned the genomic DNA and cDNA of bglA, which encodes β-glucosidase in Aspergillus kawachii, based on a partial amino acid sequence of purified cell wall-bound β-glucosidase CB-1. The nucleotide sequence of the cloned bglA gene revealed a 2,933-bp open reading frame with six introns that encodes an 860-amino-acid protein. Based on the deduced amino acid sequence, we concluded that the bglA gene encodes cell wall-bound β-glucosidase CB-1. The amino acid sequence exhibited high levels of homology with the amino acid sequences of fungal β-glucosidases classified in subfamily B. We expressed the bglA cDNA in Saccharomyces cerevisiae and detected the recombinant β-glucosidase in the periplasm fraction of the recombinant yeast. A. kawachii can produce two extracellular β-glucosidases (EX-1 and EX-2) in addition to the cell wall-bound β-glucosidase. A. kawachii in which the bglA gene was disrupted produced none of the three β-glucosidases, as determined by enzyme assays and a Western blot analysis. Thus, we concluded that the bglA gene encodes both extracellular and cell wall-bound β-glucosidases in A. kawachii. PMID:10584016

A novel nonsense mutation in the NDP gene in a Chinese family with Norrie disease.

PubMed

Liu, Deyuan; Hu, Zhengmao; Peng, Yu; Yu, Changhong; Liu, Yalan; Mo, Xiaoyun; Li, Xiaoping; Lu, Lina; Xu, Xiaojuan; Su, Wei; Pan, Qian; Xia, Kun

2010-12-08

Norrie disease (ND), a rare X-linked recessive disorder, is characterized by congenital blindness and, occasionally, mental retardation and hearing loss. ND is caused by the Norrie Disease Protein gene (NDP), which codes for norrin, a cysteine-rich protein involved in ocular vascular development. Here, we report a novel mutation of NDP that was identified in a Chinese family in which three members displayed typical ND symptoms and other complex phenotypes, such as cerebellar atrophy, motor disorders, and mental disorders. We conducted an extensive clinical examination of the proband and performed a computed tomography (CT) scan of his brain. Additionally, we performed ophthalmic examinations, haplotype analyses, and NDP DNA sequencing for 26 individuals from the proband's extended family. The proband's computed tomography scan, in which the fifth ventricle could be observed, indicated cerebellar atrophy. Genome scans and haplotype analyses traced the disease to chromosome Xp21.1-p11.22. Mutation screening of the NDP gene identified a novel nonsense mutation, c.343C>T, in this region. Although recent research has shown that multiple different mutations can be responsible for the ND phenotype, additional research is needed to understand the mechanism responsible for the diverse phenotypes caused by mutations in the NDP gene.

A novel nonsense mutation in the NDP gene in a Chinese family with Norrie disease

PubMed Central

Liu, Deyuan; Hu, Zhengmao; Peng, Yu; Yu, Changhong; Liu, Yalan; Mo, Xiaoyun; Li, Xiaoping; Lu, Lina; Xu, Xiaojuan; Su, Wei; Pan, Qian

2010-01-01

Purpose Norrie disease (ND), a rare X-linked recessive disorder, is characterized by congenital blindness and, occasionally, mental retardation and hearing loss. ND is caused by the Norrie Disease Protein gene (NDP), which codes for norrin, a cysteine-rich protein involved in ocular vascular development. Here, we report a novel mutation of NDP that was identified in a Chinese family in which three members displayed typical ND symptoms and other complex phenotypes, such as cerebellar atrophy, motor disorders, and mental disorders. Methods We conducted an extensive clinical examination of the proband and performed a computed tomography (CT) scan of his brain. Additionally, we performed ophthalmic examinations, haplotype analyses, and NDP DNA sequencing for 26 individuals from the proband’s extended family. Results The proband’s computed tomography scan, in which the fifth ventricle could be observed, indicated cerebellar atrophy. Genome scans and haplotype analyses traced the disease to chromosome Xp21.1-p11.22. Mutation screening of the NDP gene identified a novel nonsense mutation, c.343C>T, in this region. Conclusions Although recent research has shown that multiple different mutations can be responsible for the ND phenotype, additional research is needed to understand the mechanism responsible for the diverse phenotypes caused by mutations in the NDP gene. PMID:21179243

Genomic characterization of an extensively-drug resistance Salmonella enterica serotype Indiana strain harboring blaNDM-1 gene isolated from a chicken carcass in China.

PubMed

Wang, Wei; Peng, Zixin; Baloch, Zulqarnain; Hu, Yujie; Xu, Jin; Zhang, Wenhui; Fanning, Séamus; Li, Fengqin

2017-11-01

The objective of this study was to genetically characterize the antimicrobial resistance mechanisms of Salmonella enterica serotype Indiana C629 isolated from a chicken carcass in China in 2014. Antimicrobial susceptibility against a panel of 23 antimicrobial agents was carried out on Salmonella enterica serotype Indiana C629 and assessed according to CLSI standards. Whole-genome sequencing of this isolate was conducted to obtain the complete genome of S. Indiana. Salmonella Indiana C629 expressed an XDR phenotype being resistant to more than 20 antimicrobial agents, including imipenem and meropenem. From the analysis of the resistance mechanisms, two mutations were identified in subunit A of DNA gyrase within the quinolone resistance determining region, in addition to the acquisition of mobile efflux pumps encoding oqxA/B/R. Additionally, four beta-lactamases resistance genes (bla CTX-M-65 , bla TEM-1 , bla OXA-1 , and bla NDM-1 ), five aminoglycosides resistance genes (aac(3)-IV, aac(6')-Ib-cr, aadA2, aadA5, and aph(4)-Ia), two phenicol resistance genes (catB3 and floR), and five trimethoprim/sulfamethoxazole resistance genes (sul1/2/3 and dfrA12/17) were also identified. A total of 191 virulence genes were identified. Among them, 57 belonged to type-three secretion system (T3SS) encoding genes, 55 belonged to fimbrial adherence encoding genes, and 39 belonged to flagella-encoding genes CONCLUSIONS: This study demonstrated that multi-resistance mechanisms consistent with an XDR-phenotype, along with various virulence encoding genes of a S. Indiana strain in China These findings highlight the importance of cooperation among different sectors in order to monitor the spread of resistant pathogens among food animal, foods of animal origin and human beings that might further take measures to protect consumers' health. Copyright © 2017 Elsevier GmbH. All rights reserved.

Evidence for a Dual Antiviral Role of the Major Nuclear Domain 10 Component Sp100 during the Immediate-Early and Late Phases of the Human Cytomegalovirus Replication Cycle ▿

PubMed Central

Tavalai, Nina; Adler, Martina; Scherer, Myriam; Riedl, Yvonne; Stamminger, Thomas

2011-01-01

In recent studies, the nuclear domain 10 (ND10) components PML and hDaxx were identified as cellular restriction factors that inhibit the initiation of human cytomegalovirus (HCMV) replication. The antiviral function of ND10, however, is antagonized by the IE1 protein, which induces ND10 disruption. Here we show that IE1 not only de-SUMOylates PML immediately upon infection but also directly targets Sp100. IE1 expression alone was sufficient to downregulate endogenous Sp100 independently of the presence of PML. Moreover, cotransfection experiments revealed that IE1 negatively interferes with the SUMOylation of all Sp100 isoforms. The modulation of Sp100 at immediate-early (IE) times of infection, indeed, seemed to have an in vivo relevance for HCMV replication, since knockdown of Sp100 resulted in more cells initiating the viral gene expression program. In addition, we observed that Sp100 was degraded in a proteasome-dependent manner at late times postinfection, suggesting that Sp100 may play an additional antiviral role during the late phase. Infection experiments conducted with Sp100 knockdown human foreskin fibroblasts (HFFs) confirmed this hypothesis: depletion of Sp100 resulted in augmented release of progeny virus particles compared to that from control cells. Consistent with this observation, we noted increased amounts of viral late gene products in the absence of Sp100. Importantly, this elevated late gene expression was not dependent on enhanced viral IE gene expression. Taken together, our data provide evidence that Sp100 is the first ND10-related factor identified that not only possesses the potential to restrict the initial stage of infection but also inhibits HCMV replication during the late phase. PMID:21734036

Restricted utilization of germ-line VH genes in rabbits: implications for inheritance of VH allotypes and generation of antibody diversity.

PubMed

Knight, K L; Becker, R S; DiPietro, L A

1991-01-01

The presence of inherited VH region allotypic specificities, a1, a2 or a3, on nearly all rabbit immunoglobulins has presented a paradox. We know the germline contains hundreds of VH genes, and if we assume that most of these are used in the generation of antibody diversity, then we must ask how have the a allotype-encoding regions been maintained over time? On the other hand, if we assume that only one (or a small number) of these VH gene(s) is (are) used in VDJ gene rearrangements, then, how is antibody diversity generated? To address these questions, we have cloned and determined the nucleotide sequence of the 3'-most germline VH genes from the a1, a2 and a3 chromosomes and shown in each case that the 3'-most H gene, VH1-a1, VH1-a2, or VH1-a3, encodes an a1, a2 or a3 VH region, respectively. Analysis of rearranged VDJ genes from leukemic B cells showed that VH1 was utilized in these rearrangements. Based on these data, we propose that the allelic inheritance of the VH allotypes is explained by the preferential usage of the VH1 gene in VDJ rearrangements. Support for this hypothesis was obtained from analysis of the mutant rabbit Alicia in which most serum Ig molecules do not have VHa allotypic specificities, but instead have so-called VHa-negative Ig molecules. In this rabbit, VH1 is not expressed as it has been deleted. Analysis of cDNA clones from spleen of Alicia rabbits suggests that the expressed VHa-negative molecules also are encoded by a single germline VH gene. Thus, we suggest that nearly all rabbit VH regions are encoded by one to two germline VH genes and that antibody diversity is generated primarily by somatic hypermutation and gene conversion.

Direct cloning of the trxB gene that encodes thioredoxin reductase.

PubMed Central

Russel, M; Model, P

1985-01-01

A strain was constructed which contains mutations in the genes encoding thioredoxin (trxA) and thioredoxin reductase (trxB) such that filamentous phage f1 cannot grow. The complementation of either mutation with its wild-type allele permits phage growth. We used this strain to select f1 phage which contain a cloned trxB gene. The location of the gene on the cloned fragment was determined, and its protein product was identified. Plasmid subclones that contain this gene overproduce thioredoxin reductase. Images PMID:2989245

Gene disruption in Trichoderma atroviride via Agrobacterium-mediated transformation.

PubMed

Zeilinger, Susanne

2004-02-01

A modified Agrobacterium-mediated transformation method for the efficient disruption of two genes encoding signaling compounds of the mycoparasite Trichoderma atroviride is described, using the hph gene of Escherichia coli as selection marker. The transformation vectors contained about 1 kb of 5' and 3' non-coding regions from the tmk1 (encoding a MAP kinase) or tga3 (encoding an alpha-subunit of a heterotrimeric G protein) target loci flanking a selection marker. Transformation of fungal conidia and selection on hygromycin-containing media applying an overlay-based procedure, which overcomes the lack of formation of distinct single colonies by the fungus, led to stable clones for both disruption constructs. Southern and PCR analyses proved gene disruption by single-copy homologous integration with a frequency of approximately 60% for both genes; and the loss of tmk1 and tga3 transcript formation in the disruptants was demonstrated by RT-PCR.

Identification of a Novel Dioxygenase Involved in Metabolism of o-Xylene, Toluene, and Ethylbenzene by Rhodococcus sp. Strain DK17

PubMed Central

Kim, Dockyu; Chae, Jong-Chan; Zylstra, Gerben J.; Kim, Young-Soo; Kim, Seong-Ki; Nam, Myung Hee; Kim, Young Min; Kim, Eungbin

2004-01-01

Rhodococcus sp. strain DK17 is able to grow on o-xylene, benzene, toluene, and ethylbenzene. DK17 harbors at least two megaplasmids, and the genes encoding the initial steps in alkylbenzene metabolism are present on the 330-kb pDK2. The genes encoding alkylbenzene degradation were cloned in a cosmid clone and sequenced completely to reveal 35 open reading frames (ORFs). Among the ORFs, we identified two nearly exact copies (one base difference) of genes encoding large and small subunits of an iron sulfur protein terminal oxygenase that are 6 kb apart from each other. Immediately downstream of one copy of the dioxygenase genes (akbA1a and akbA2a) is a gene encoding a dioxygenase ferredoxin component (akbA3), and downstream of the other copy (akbA1b and akbA2b) are genes putatively encoding a meta-cleavage pathway. RT-PCR experiments show that the two copies of the dioxygenase genes are operonic with the downstream putative catabolic genes and that both operons are induced by o-xylene. When expressed in Escherichia coli, AkbA1a-AkbA2a-AkbA3 transformed o-xylene into 2,3- and 3,4-dimethylphenol. These were apparently derived from an unstable o-xylene cis-3,4-dihydrodiol, which readily dehydrates. This indicates a single point of attack of the dioxygenase on the aromatic ring. In contrast, attack of AkbA1a-AkbA2a-AkbA3 on ethylbenzene resulted in the formation of two different cis-dihydrodiols resulting from an oxidation at the 2,3 and the 3,4 positions on the aromatic ring, respectively. PMID:15574904

Isolation of Nicotiana plumbaginifolia cDNAs encoding isoforms of serine acetyltransferase and O-acetylserine (thiol) lyase in a yeast two-hybrid system with Escherichia coli cysE and cysK genes as baits.

PubMed

Liszewska, Frantz; Gaganidze, Dali; Sirko, Agnieszka

2005-01-01

We applied the yeast two-hybrid system for screening of a cDNA library of Nicotiana plumbaginifolia for clones encoding plant proteins interacting with two proteins of Escherichia coli: serine acetyltransferase (SAT, the product of cysE gene) and O-acetylserine (thiol)lyase A, also termed cysteine synthase (OASTL-A, the product of cysK gene). Two plant cDNA clones were identified when using the cysE gene as a bait. These clones encode a probable cytosolic isoform of OASTL and an organellar isoform of SAT, respectively, as indicated by evolutionary trees. The second clone, encoding SAT, was identified independently also as a "prey" when using cysK as a bait. Our results reveal the possibility of applying the two-hybrid system for cloning of plant cDNAs encoding enzymes of the cysteine synthase complex in the two-hybrid system. Additionally, using genome walking sequences located upstream of the sat1 cDNA were identified. Subsequently, in silico analyses were performed aiming towards identification of the potential signal peptide and possible location of the deduced mature protein encoded by sat1.

Isolation and characterization of polygalacturonase genes (pecA and pecB) from Aspergillus flavus.

PubMed Central

Whitehead, M P; Shieh, M T; Cleveland, T E; Cary, J W; Dean, R A

1995-01-01

Two genes, pecA and pecB, encoding endopolyglacturonases were cloned from a highly aggressive strain of Aspergillus flavus. The pecA gene consisted of 1,228 bp encoding a protein of 363 amino acids with a predicted molecular mass of 37.6 kDa, interrupted by two introns of 58 and 81 bp in length. Accumulation of pecA mRNA in both pectin- or glucose-grown mycelia in the highly aggressive strain matched the activity profile of a pectinase previously identified as P2c. Transformants of a weakly aggressive strain containing a functional copy of the pecA gene produced P2c in vitro, confirming that pecA encodes P2c. The coding region of pecB was determined to be 1,217 bp in length interrupted by two introns of 65 and 54 bp in length. The predicted protein of 366 amino acids had an estimated molecular mass of 38 kDa. Transcripts of this gene accumulated in mycelia grown in medium containing pectin alone, never in mycelia grown in glucose-containing medium, for both highly and weakly aggressive strains. Thus, pecB encodes the activity previously identified as P1 or P3. pecA and pecB share a high degree of sequence identity with polygalacturonase genes from Aspergillus parasiticus and Aspergillus oryzae, further establishing the close relationships between members of the A. flavus group. Conservation of intron positions in these genes also indicates that they share a common ancestor with genes encoding endopolyglacturonases of Aspergillus niger. PMID:7574642

Genetic and Environmental Overlap between Chinese and English Reading-Related Skills in Chinese Children

ERIC Educational Resources Information Center

Wong, Simpson W. L.; Chow, Bonnie Wing-Yin; Ho, Connie Suk-Han; Waye, Mary M. Y.; Bishop, Dorothy V. M.

2014-01-01

This twin study examined the relative contributions of genes and environment on 2nd language reading acquisition of Chinese-speaking children learning English. We examined whether specific skills-visual word recognition, receptive vocabulary, phonological awareness, phonological memory, and speech discrimination-in the 1st and 2nd languages have…

Mitochondrial functions modulate neuroendocrine, metabolic, inflammatory, and transcriptional responses to acute psychological stress

PubMed Central

Picard, Martin; McManus, Meagan J.; Gray, Jason D.; Nasca, Carla; Moffat, Cynthia; Kopinski, Piotr K.; Seifert, Erin L.; McEwen, Bruce S.; Wallace, Douglas C.

2015-01-01

The experience of psychological stress triggers neuroendocrine, inflammatory, metabolic, and transcriptional perturbations that ultimately predispose to disease. However, the subcellular determinants of this integrated, multisystemic stress response have not been defined. Central to stress adaptation is cellular energetics, involving mitochondrial energy production and oxidative stress. We therefore hypothesized that abnormal mitochondrial functions would differentially modulate the organism’s multisystemic response to psychological stress. By mutating or deleting mitochondrial genes encoded in the mtDNA [NADH dehydrogenase 6 (ND6) and cytochrome c oxidase subunit I (COI)] or nuclear DNA [adenine nucleotide translocator 1 (ANT1) and nicotinamide nucleotide transhydrogenase (NNT)], we selectively impaired mitochondrial respiratory chain function, energy exchange, and mitochondrial redox balance in mice. The resulting impact on physiological reactivity and recovery from restraint stress were then characterized. We show that mitochondrial dysfunctions altered the hypothalamic–pituitary–adrenal axis, sympathetic adrenal–medullary activation and catecholamine levels, the inflammatory cytokine IL-6, circulating metabolites, and hippocampal gene expression responses to stress. Each mitochondrial defect generated a distinct whole-body stress-response signature. These results demonstrate the role of mitochondrial energetics and redox balance as modulators of key pathophysiological perturbations previously linked to disease. This work establishes mitochondria as stress-response modulators, with implications for understanding the mechanisms of stress pathophysiology and mitochondrial diseases. PMID:26627253

Material and methods to increase plant growth and yield

DOEpatents

Kirst, Matias

2015-09-15

The present invention relates to materials and methods for modulating growth rates, yield, and/or resistance to drought conditions in plants. In one embodiment, a method of the invention comprises increasing expression of an hc1 gene (or a homolog thereof that provides for substantially the same activity), or increasing expression or activity of the protein encoded by an hc1 gene thereof, in a plant, wherein expression of the hc1 gene or expression or activity of the protein encoded by an hc1 gene results in increased growth rate, yield, and/or drought resistance in the plant.

Materials and methods to increase plant growth and yield

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

Kirst, Matias

The present invention relates to materials and methods for modulating growth rates, yield, and/or resistance to drought conditions in plants. In one embodiment, a method of the invention comprises increasing expression of an hc1 gene (or a homolog thereof that provides for substantially the same activity), or increasing expression or activity of the protein encoded by an hc1 gene thereof, in a plant, wherein expression of the hc1 gene or expression or activity of the protein encoded by an hc1 gene results in increased growth rate, yield, and/or drought resistance in the plant.

[Divergence of paralogous growth-hormone-encoding genes and their promoters in Salmonidae].

PubMed

Kamenskaya, D N; Pankova, M V; Atopkin, D M; Brykov, V A

2017-01-01

In many fish species, including salmonids, the growth-hormone is encoded by two duplicated paralogous genes, gh1 and gh2. Both genes were already in place at the time of divergence of species in this group. A comparison of the entire sequence of these genes of salmonids has shown that their conserved regions are associated with exons, while their most variable regions correspond to introns. Introns C and D include putative regulatory elements (sites Pit-1, CRE, and ERE), that are also conserved. In chars, the degree of polymorphism of gh2 gene is 2-3 times as large as that in gh1 gene. However, a comparison across all Salmonidae species would not extent this observation to other species. In both these chars' genes, the promoters are conserved mainly because they correspond to putative regulatory sequences (TATA box, binding sites for the pituitary transcription factor Pit-1 (F1-F4), CRE, GRE and RAR/RXR elements). The promoter of gh2 gene has a greater degree of polymorphism compared with gh1 gene promoter in all investigated species of salmonids. The observed differences in the rates of accumulation of changes in growth hormone encoding paralogs could be explained by differences in the intensity of selection.

Structural evolution of the 4/1 genes and proteins in non-vascular and lower vascular plants.

PubMed

Morozov, Sergey Y; Milyutina, Irina A; Bobrova, Vera K; Ryazantsev, Dmitry Y; Erokhina, Tatiana N; Zavriev, Sergey K; Agranovsky, Alexey A; Solovyev, Andrey G; Troitsky, Alexey V

2015-12-01

The 4/1 protein of unknown function is encoded by a single-copy gene in most higher plants. The 4/1 protein of Nicotiana tabacum (Nt-4/1 protein) has been shown to be alpha-helical and predominantly expressed in conductive tissues. Here, we report the analysis of 4/1 genes and the encoded proteins of lower land plants. Sequences of a number of 4/1 genes from liverworts, lycophytes, ferns and gymnosperms were determined and analyzed together with sequences available in databases. Most of the vascular plants were found to encode Magnoliophyta-like 4/1 proteins exhibiting previously described gene structure and protein properties. Identification of the 4/1-like proteins in hornworts, liverworts and charophyte algae (sister lineage to all land plants) but not in mosses suggests that 4/1 proteins are likely important for plant development but not required for a primary metabolic function of plant cell. Copyright © 2015 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.

A second cistron in the CACNA1A gene encodes a transcription factor that mediates cerebellar development and SCA6

PubMed Central

Du, Xiaofei; Wang, Jun; Zhu, Haipeng; Rinaldo, Lorenzo; Lamar, Kay-Marie; Palmenberg, Ann C.; Hansel, Christian; Gomez, Christopher M.

2014-01-01

SUMMARY The CACNA1A gene, encoding the voltage-gated calcium channel subunit α1A, is involved in pre- and postsynaptic Ca2+ signaling, gene expression, and several genetic neurological disorders. We found that CACNA1A employs a novel strategy to directly coordinate a gene expression program, using a bicistronic mRNA bearing a cryptic internal ribosomal entry site (IRES). The first cistron encodes the well-characterized α1A subunit. The second expresses a newly-recognized transcription factor, α1ACT, that coordinates expression of a program of genes involved in neural and Purkinje cell development. α1ACT also contains the polyglutamine (polyQ) tract that, when expanded, causes spinocerebellar ataxia type 6 (SCA6). When expressed as an independent polypeptide, α1ACT, bearing an expanded polyQ tract, lacks transcription factor function and neurite outgrowth properties, causes cell death in culture, and leads to ataxia and cerebellar atrophy in transgenic mice. Suppression of CACNA1A IRES function in SCA6 may be a potential therapeutic strategy. PMID:23827678

Arxula adeninivorans (Blastobotrys adeninivorans) — A Dimorphic Yeast of Great Biotechnological Potential

NASA Astrophysics Data System (ADS)

Böer, Erik; Steinborn, Gerhard; Florschütz, Kristina; Körner, Martina; Gellissen, Gerd; Kunze, Gotthard

The dimorphic ascomycetous yeast Arxula adeninivorans exhibits some unusual properties. Being a thermo- and halotolerant species it is able to assimilate and ferment many compounds as sole carbon and/or nitrogen source. It utilises n-alkanes and is capable of degrading starch. Due to these unusual biochemical properties A. adeninivorans can be exploited as a gene donor for the production of enzymes with attractive biotechnological characteristics. Examples of A. adeninivorans-derived genes that are overexpressed include the ALIP1 gene encoding a secretory lipase, the AINV encoding invertase, the AXDH encoding xylitol dehydrogenase and the APHY encoding a secretory phosphatase with phytase activity.

The complete mitochondrial genome of the black star fat minnow (Rhynchocypris semotilus), an endemic and endangered fish of Korea.

PubMed

Yu, Jeong-Nam; Kim, Byung-Jik; Kim, Changmu; Yeo, Joo-Hong; Kim, Soonok

2017-01-01

The Black star fat minnow (Rhynchocypris semotilus) is an endemic and critically endangered freshwater fish in Korea. Its genome was 16 605 bp long and consisted of 13 protein-coding genes (PCG), two rRNA genes, 22 tRNA genes, and a control region. The gene order and the composition of R. semotilus were similar to that of most other vertebrates. Four overlapping regions in ATP8/ATP6, ATP6/COX3, ND4L/ND4, and ND5/ND6, among the 13 PCGs were found. The control region was located between the tRNA-Pro and tRNA-Phe genes and was determined to be 935 bp in length with the 3' end containing a 12 TA-repeat sequence. Phylogenetic analysis suggested that R. semotilus is most closely related to R. oxycephalus.

Activation of Pathogenesis-related Genes by the Rhizobacterium, Bacillus sp. JS, Which Induces Systemic Resistance in Tobacco Plants.

PubMed

Kim, Ji-Seong; Lee, Jeongeun; Lee, Chan-Hui; Woo, Su Young; Kang, Hoduck; Seo, Sang-Gyu; Kim, Sun-Hyung

2015-06-01

Plant growth promoting rhizobacteria (PGPR) are known to confer disease resistance to plants. Bacillus sp. JS demonstrated antifungal activities against five fungal pathogens in in vitro assays. To verify whether the volatiles of Bacillus sp. JS confer disease resistance, tobacco leaves pre-treated with the volatiles were damaged by the fungal pathogen, Rhizoctonia solani and oomycete Phytophthora nicotianae. Pre-treated tobacco leaves had smaller lesion than the control plant leaves. In pathogenesis-related (PR) gene expression analysis, volatiles of Bacillus sp. JS caused the up-regulation of PR-2 encoding β-1,3-glucanase and acidic PR-3 encoding chitinase. Expression of acidic PR-4 encoding chitinase and acidic PR-9 encoding peroxidase increased gradually after exposure of the volatiles to Bacillus sp. JS. Basic PR-14 encoding lipid transfer protein was also increased. However, PR-1 genes, as markers of salicylic acid (SA) induced resistance, were not expressed. These results suggested that the volatiles of Bacillus sp. JS confer disease resistance against fungal and oomycete pathogens through PR genes expression.

Fusagene vectors: a novel strategy for the expression of multiple genes from a single cistron.

PubMed

Gäken, J; Jiang, J; Daniel, K; van Berkel, E; Hughes, C; Kuiper, M; Darling, D; Tavassoli, M; Galea-Lauri, J; Ford, K; Kemeny, M; Russell, S; Farzaneh, F

2000-12-01

Transduction of cells with multiple genes, allowing their stable and co-ordinated expression, is difficult with the available methodologies. A method has been developed for expression of multiple gene products, as fusion proteins, from a single cistron. The encoded proteins are post-synthetically cleaved and processed into each of their constituent proteins as individual, biologically active factors. Specifically, linkers encoding cleavage sites for the Golgi expressed endoprotease, furin, have been incorporated between in-frame cDNA sequences encoding different secreted or membrane bound proteins. With this strategy we have developed expression vectors encoding multiple proteins (IL-2 and B7.1, IL-4 and B7.1, IL-4 and IL-2, IL-12 p40 and p35, and IL-12 p40, p35 and IL-2 ). Transduction and analysis of over 100 individual clones, derived from murine and human tumour cell lines, demonstrate the efficient expression and biological activity of each of the encoded proteins. Fusagene vectors enable the co-ordinated expression of multiple gene products from a single, monocistronic, expression cassette.

Cloning and characterization of the nagA gene that encodes beta-n-acetylglucosaminidase from Aspergillus nidulans and its expression in Aspergillus oryzae.

PubMed

Kim, Sunhwa; Matsuo, Ichiro; Ajisaka, Katsumi; Nakajima, Harushi; Kitamoto, Katsuhiko

2002-10-01

We isolated a beta-N-acetylglucosaminidase encoding gene and its cDNA from the filamentous fungus Aspergillus nidulans, and designated it nagA. The nagA gene contained no intron and encoded a polypeptide of 603 amino acids with a putative 19-amino acid signal sequence. The deduced amino acid sequence was very similar to the sequence of Candida albicans Hex1 and Trichoderma harzianum Nag1. Yeast cells containing the nagA cDNA under the control of the GAL1 promoter expressed beta-N-acetylglucosaminidase activity. The chromosomal nagA gene of A. nidulans was disrupted by replacement with the argB marker gene. The disruptant strains expressed low levels of beta-N-acetylglucosaminidase activity and showed poor growth on a medium containing chitobiose as a carbon source. Aspergillus oryzae strain carrying the nagA gene under the control of the improved glaA promoter produced large amounts of beta-N-acetylglucosaminidase in a wheat bran solid culture.

Structure, Expression, Chromosomal Location and Product of the Gene Encoding Adh2 in Petunia

PubMed Central

Gregerson, R. G.; Cameron, L.; McLean, M.; Dennis, P.; Strommer, J.

1993-01-01

In most higher plants the genes encoding alcohol dehydrogenase comprise a small gene family, usually with two members. The Adh1 gene of Petunia has been cloned and analyzed, but a second identifiable gene was not recovered from any of three genomic libraries. We have therefore employed the polymerase chain reaction to obtain the major portion of a second Adh gene. From sequence, mapping and northern data we conclude this gene encodes ADH2, the major anaerobically inducible Adh gene of Petunia. The availability of both Adh1 and Adh2 from Petunia has permitted us to compare their structures and patterns of expression to those of the well-studied Adh genes of maize, of which one is highly expressed developmentally, while both are induced in response to hypoxia. Despite their evolutionary distance, evidenced by deduced amino acid sequence as well as taxonomic classification, the pairs of genes are regulated in strikingly similar ways in maize and Petunia. Our findings suggest a significant biological basis for the regulatory strategy employed by these distant species for differential expression of multiple Adh genes. PMID:8096485

[Polymorphism of genes encoding proteins of DNA repair vs. occupational and environmental exposure to lead, arsenic and pesticides].

PubMed

Bukowski, Karol; Woźniak, Katarzyna

2018-03-09

Genetic polymorphism is associated with the occurrence of at least 2 different alleles in the locus with a frequency higher than 1% in the population. Among polymorphisms we can find single nucleotide polymorphism (SNP) and polymorphism of variable number of tandem repeats. The presence of certain polymorphisms in genes encoding DNA repair enzymes is associated with the speed and efficiency of DNA repair and can protect or expose humans to the effects provoked by xenobiotics. Chemicals, such as lead, arsenic pesticides are considered to exhibit strong toxicity. There are many different polymorphisms in genes encoding DNA repair enzymes, which determine the speed and efficiency of DNA damage repair induced by these xenobiotics. In the case of lead, the influence of various polymorphisms, such as APE1 (apurinic/apyrimidinic endonuclease 1) (rs1130409), hOGG1 (human 8-oxoguanine glycosylase) (rs1052133), XRCC1 (X-ray repair cross-complementing protein group 1) (rs25487), XRCC1 (rs1799782) and XRCC3 (X-ray repair cross-complementing protein group 3) (rs861539) were described. For arsenic polymorphisms, such as ERCC2 (excision repair cross-complementing) (rs13181), XRCC3 (rs861539), APE1 (rs1130409) and hOGG1 (rs1052133) were examined. As to pesticides, separate and combined effects of polymorphisms in genes encoding DNA repair enzymes, such as XRCC1 (rs1799782), hOGG1 (rs1052133), XRCC4 (X-ray repair cross-complementing protein group 4) (rs28360135) and the gene encoding the detoxification enzyme PON1 paraoxonase (rs662) were reported. Med Pr 2018;69(2):225-235. This work is available in Open Access model and licensed under a CC BY-NC 3.0 PL license.

[Participation of GDNF, LIMK1 signal pathways and heat shock proteins in processes of Drosophila learning and memory formation].

PubMed

Nikitina, E A; Medvedeva, A V; Dolgaia, Iu F; Korochkin, L I; Pavlova, G V; Savvateeva-Popova, E V

2012-01-01

Molecular mechanisms of the synapse and dendrite maintenance and their disturbance in psychiatric and neurodegenerative diseases (ND) are intensively studied in searching for target genes of therapeutic actions. It is suggested that glia, alongside with well-studied pre- and postsynaptic neurons, is the third, poorly studied partner in synaptic transmission (the tripartite synapse) that is involved in the positive feedback between the first two partners. This bidirectional coupling between presynaptic neurons and their postsynaptic targets involve neurotrophins (NTF), such as glial cell-derived neurotrophic factor (GDNF) that is produced LIM kinase 1 (LIMK1, the key enzyme of actin remodeling). The cytoplasmic domain of neuregulins interacts with LIMK1. Since neurons and axons that do not receive a sufficient NTF amount are at risk of degeneration and synapse elimination, GDNF seems to be the best studied factor of the ND therapy. The delivery of GDNF stem cells to the neurodegeneration locus is very efficient. There has been proposed a new approach based on use of Drosophila heat shock (hs) promoter. This promoter responds to the mammalian body temperature as to the shock factor resulting in the constant expression of the GDNF gene. The Drosophila models allow studying any given component of the bidirectional communication between pre- and postsynaptic neurons in development of the main diagnostic ND symptom, such as defective memory resulted from synaptic atrophy. In the present study we used the Drosophila stocks imitating different disturbances of the nervous system: Canton-S (wild type), GDNF (transgenic flies that carry human glial-cell-line derived nerve factor (GDNF) gene under hs promoter), l(1)ts403 with dusturbance of HSPs mRNA extranuclear transport, a defect of intracellular stress report, and agn(ts3) mutation in LIMK1 gene. We have revealed functional connections at the behavioral level (learning/memory) depending on the GDNF and LIMK1 brain expression and HSPs transduction that might provide targets for complex approaches for the ND treatment.

AtMRP1 gene of Arabidopsis encodes a glutathione S-conjugate pump: isolation and functional definition of a plant ATP-binding cassette transporter gene.

PubMed

Lu, Y P; Li, Z S; Rea, P A

1997-07-22

Because plants produce cytotoxic compounds to which they, themselves, are susceptible and are exposed to exogenous toxins (microbial products, allelochemicals, and agrochemicals), cell survival is contingent on mechanisms for detoxifying these agents. One detoxification mechanism is the glutathione S-transferase-catalyzed glutathionation of the toxin, or an activated derivative, and transport of the conjugate out of the cytosol. We show here that a transporter responsible for the removal of glutathione S-conjugates from the cytosol, a specific Mg2+-ATPase, is encoded by the AtMRP1 gene of Arabidopsis thaliana. The sequence of AtMRP1 and the transport capabilities of membranes prepared from yeast cells transformed with plasmid-borne AtMRP1 demonstrate that this gene encodes an ATP-binding cassette transporter competent in the transport of glutathione S-conjugates of xenobiotics and endogenous substances, including herbicides and anthocyanins.

Linkage of the gene that encodes the alpha 1 chain of type V collagen (COL5A1) to type II Ehlers-Danlos syndrome (EDS II).

PubMed

Loughlin, J; Irven, C; Hardwick, L J; Butcher, S; Walsh, S; Wordsworth, P; Sykes, B

1995-09-01

Ehlers-Danlos syndrome (EDS) is a group of heritable disorders of connective tissue with skin, ligaments and blood vessels being the main sites affected. The commonest variant (EDS II) exhibits an autosomal dominant mode of inheritance and is characterized by joint hypermobility, cigarette paper scars, lax skin and excessive bruising. As yet no gene has been linked to EDS II, nor has linkage been established to a specific region of the genome. However, several candidate genes encoding proteins of the extracellular matrix have been excluded. Using an intragenic simple sequence repeat polymorphism, we report linkage of the COL5A1 gene, which encodes the alpha 1(V) chain of type V collagen, to EDS II. A maximum LOD score (Zmax) for linkage of 8.3 at theta = 0.00 was generated for a single large pedigree.

Adjustment of Trehalose Metabolism in Wine Saccharomyces cerevisiae Strains To Modify Ethanol Yields

PubMed Central

Rossouw, D.; Heyns, E. H.; Setati, M. E.; Bosch, S.

2013-01-01

The ability of Saccharomyces cerevisiae to efficiently produce high levels of ethanol through glycolysis has been the focus of much scientific and industrial activity. Despite the accumulated knowledge regarding glycolysis, the modification of flux through this pathway to modify ethanol yields has proved difficult. Here, we report on the systematic screening of 66 strains with deletion mutations of genes encoding enzymes involved in central carbohydrate metabolism for altered ethanol yields. Five of these strains showing the most prominent changes in carbon flux were selected for further investigation. The genes were representative of trehalose biosynthesis (TPS1, encoding trehalose-6-phosphate synthase), central glycolysis (TDH3, encoding glyceraldehyde-3-phosphate dehydrogenase), the oxidative pentose phosphate pathway (ZWF1, encoding glucose-6-phosphate dehydrogenase), and the tricarboxylic acid (TCA) cycle (ACO1 and ACO2, encoding aconitase isoforms 1 and 2). Two strains exhibited lower ethanol yields than the wild type (tps1Δ and tdh3Δ), while the remaining three showed higher ethanol yields. To validate these findings in an industrial yeast strain, the TPS1 gene was selected as a good candidate for genetic modification to alter flux to ethanol during alcoholic fermentation in wine. Using low-strength promoters active at different stages of fermentation, the expression of the TPS1 gene was slightly upregulated, resulting in a decrease in ethanol production and an increase in trehalose biosynthesis during fermentation. Thus, the mutant screening approach was successful in terms of identifying target genes for genetic modification in commercial yeast strains with the aim of producing lower-ethanol wines. PMID:23793638

Cloning and characterization of the metE gene encoding S-adenosylmethionine synthetase from Bacillus subtilis.

PubMed Central

Yocum, R R; Perkins, J B; Howitt, C L; Pero, J

1996-01-01

The metE gene, encoding S-adenosylmethionine synthetase (EC 2.5.1.6) from Bacillus subtilis, was cloned in two steps by normal and inverse PCR. The DNA sequence of the metE gene contains an open reading frame which encodes a 400-amino-acid sequence that is homologous to other known S-adenosylmethionine synthetases. The cloned gene complements the metE1 mutation and integrates at or near the chromosomal site of metE1. Expression of S-adenosylmethionine synthetase is reduced by only a factor of about 2 by exogenous methioinine. Overproduction of S-adenosylmethionine synthetase from a strong constitutive promoter leads to methionine auxotrophy in B. subtilis, suggesting that S-adenosylmethionine is a corepressor of methionine biosynthesis in B. subtilis, as others have already shown for Escherichia coli. PMID:8755891

Cloning and characterization of the metE gene encoding S-adenosylmethionine synthetase from Bacillus subtilis.

PubMed

Yocum, R R; Perkins, J B; Howitt, C L; Pero, J

1996-08-01

The metE gene, encoding S-adenosylmethionine synthetase (EC 2.5.1.6) from Bacillus subtilis, was cloned in two steps by normal and inverse PCR. The DNA sequence of the metE gene contains an open reading frame which encodes a 400-amino-acid sequence that is homologous to other known S-adenosylmethionine synthetases. The cloned gene complements the metE1 mutation and integrates at or near the chromosomal site of metE1. Expression of S-adenosylmethionine synthetase is reduced by only a factor of about 2 by exogenous methioinine. Overproduction of S-adenosylmethionine synthetase from a strong constitutive promoter leads to methionine auxotrophy in B. subtilis, suggesting that S-adenosylmethionine is a corepressor of methionine biosynthesis in B. subtilis, as others have already shown for Escherichia coli.

Genome-wide analysis identifies colonic genes differentially associated with serum leptin and insulin concentrations in C57BL/6J mice fed a high-fat diet.

PubMed

Kim, Sung-Eun; Choo, Jinsil; Yoon, Joon; Chu, Jae Ryang; Bae, Yun Jung; Lee, Seungyeoun; Park, Taesung; Sung, Mi-Kyung

2017-01-01

Obesity-induced chronic inflammation is known to increase the risk of ulcerative colitis, Crohn's disease, and colorectal cancer. Accumulating evidence suggests that leptin and insulin are key molecules linking obesity with diseases of the lower intestine. Here, we identified serum phenotype-associated genes in the colon of diet-induced obese mice as early biomarkers of obesity-associated colonic diseases. C57BL/6J mice were fed with either normal diet (ND, 15% of fat calories) or high-fat diet (HFD, 45% of fat calories) for 8 weeks. Serum concentrations of insulin, insulin-like growth factor-1 (IGF-1), leptin, and adiponectin were measured as obesity-related phenotypic markers. Genome-wide gene expression profiles of colon tissue were determined, followed by statistical analyses to detect differentially expressed and serum phenotype-associated genes. HFD-fed mice showed higher serum concentrations of leptin (P < 0.001) and insulin (P < 0.01) than those in the ND group, whereas serum IGF-1 and adiponectin concentrations did not differ between the two dietary groups. Among differentially expressed genes affected by HFD, 135, 128, 110, and 341 genes were associated with serum levels of leptin, insulin, IGF-1, and adiponectin, respectively. We identified 17 leptin-associated genes and 4 insulin-associated genes that inversely responded to HFD and ND. Among these, leptin-associated Peli3 (Pellino E3 ubiquitin protein ligase family member 3), Creb1 (cAMP responsive element binding protein 1), and Enpp2 (ectonucleotide pyrophosphatase/phosphodiesterase 2, autotaxin) and insulin-associated Centg1 (AGAP2, ArfGAP with GTPase domain) are reported to play a role either in obesity or colonic diseases. mRNA expression of these genes was validated by RT-qPCR. Our data suggest Peli3, Creb1, Enpp2, and Centg1 as potential early biomarker candidates for obesity-induced pathophysiological changes in the colon. Future studies verifying the function of these candidates are needed for the prevention, early detection, and treatment of colon diseases.

Genome-wide analysis identifies colonic genes differentially associated with serum leptin and insulin concentrations in C57BL/6J mice fed a high-fat diet

PubMed Central

Yoon, Joon; Chu, Jae Ryang; Bae, Yun Jung; Lee, Seungyeoun; Park, Taesung; Sung, Mi-Kyung

2017-01-01

Obesity-induced chronic inflammation is known to increase the risk of ulcerative colitis, Crohn’s disease, and colorectal cancer. Accumulating evidence suggests that leptin and insulin are key molecules linking obesity with diseases of the lower intestine. Here, we identified serum phenotype-associated genes in the colon of diet-induced obese mice as early biomarkers of obesity-associated colonic diseases. C57BL/6J mice were fed with either normal diet (ND, 15% of fat calories) or high-fat diet (HFD, 45% of fat calories) for 8 weeks. Serum concentrations of insulin, insulin-like growth factor-1 (IGF-1), leptin, and adiponectin were measured as obesity-related phenotypic markers. Genome-wide gene expression profiles of colon tissue were determined, followed by statistical analyses to detect differentially expressed and serum phenotype-associated genes. HFD-fed mice showed higher serum concentrations of leptin (P < 0.001) and insulin (P < 0.01) than those in the ND group, whereas serum IGF-1 and adiponectin concentrations did not differ between the two dietary groups. Among differentially expressed genes affected by HFD, 135, 128, 110, and 341 genes were associated with serum levels of leptin, insulin, IGF-1, and adiponectin, respectively. We identified 17 leptin-associated genes and 4 insulin-associated genes that inversely responded to HFD and ND. Among these, leptin-associated Peli3 (Pellino E3 ubiquitin protein ligase family member 3), Creb1 (cAMP responsive element binding protein 1), and Enpp2 (ectonucleotide pyrophosphatase/phosphodiesterase 2, autotaxin) and insulin-associated Centg1 (AGAP2, ArfGAP with GTPase domain) are reported to play a role either in obesity or colonic diseases. mRNA expression of these genes was validated by RT-qPCR. Our data suggest Peli3, Creb1, Enpp2, and Centg1 as potential early biomarker candidates for obesity-induced pathophysiological changes in the colon. Future studies verifying the function of these candidates are needed for the prevention, early detection, and treatment of colon diseases. PMID:28170448

Differential conservation of transcriptional domains of mammalian Prophet of Pit-1 proteins revealed by structural studies of the bovine gene and comparative functional analysis of the protein.

PubMed

Showalter, Aaron D; Smith, Timothy P L; Bennett, Gary L; Sloop, Kyle W; Whitsett, Julie A; Rhodes, Simon J

2002-05-29

The Prophet of Pit-1 (PROP1) gene encodes a paired class homeodomain transcription factor that is exclusively expressed in the developing mammalian pituitary gland. PROP1 function is essential for anterior pituitary organogenesis, and heritable mutations in the gene are associated with combined pituitary hormone deficiency in human patients and animals. By cloning the bovine PROP1 gene and by comparative analysis, we demonstrate that the homeodomains and carboxyl termini of mammalian PROP1 proteins are highly conserved while the amino termini are diverged. Whereas the carboxyl termini of the human and bovine PROP1 proteins contain potent transcriptional activation domains, the amino termini and homeodomains have repressive activities. The bovine PROP1 gene has four exons and three introns and maps to a region of chromosome seven carrying a quantitative trait locus affecting ovulation rate. Two alleles of the bovine gene were found that encode distinct protein products with different DNA binding and transcriptional activities. These experiments demonstrate that mammalian PROP1 genes encode proteins with complex regulatory capacities and that modest changes in protein sequence can significantly alter the activity of this pituitary developmental transcription factor.

Induction of apoptosis in cells expressing exogenous Hippi, a molecular partner of huntingtin-interacting protein Hip1.

PubMed

Majumder, Pritha; Chattopadhyay, Biswanath; Mazumder, Arindam; Das, Pradeep; Bhattacharyya, Nitai P

2006-05-01

To decipher the pathway of apoptosis induction downstream to caspase-8 activation by exogenous expression of Hippi, an interactor of huntingtin-interacting protein Hip1, we studied apoptosis in HeLa and Neuro2A cells expressing GFP-tagged Hippi. Nuclear fragmentation, caspase-1, caspase-8, caspase-9/caspase-6 and caspase-3 activation were increased significantly in Hippi expressing cells. Cleavage of Bid, release of cytochrome c and apoptosis inducing factor (AIF) from mitochondria were also increased in GFP-Hippi expressing cells. It was observed that caspase-1 and caspase-8 activation was earlier than caspase-3 activation and nuclear fragmentation. Expression of caspase-1, caspase-3 and caspase-7 was increased while anti-apoptotic gene Bcl-2 and mitochondrial genes ND1 and ND4 were reduced in Hippi expressing cells. Besides, the expression SDHA and SDHB, nuclear genes, subunits of mitochondrial complex II were decreased in GFP-Hippi expressing cells. Taken together, we concluded that Hippi expression induced apoptosis by releasing AIF and cytochrome c from mitochondria, activation of caspase-1 and caspase-3, and altering the expression of apoptotic genes and genes involved in mitochondrial complex I and II.

Gene analysis of steroid 5 alpha-reductase 1 in hyperandrogenic women.

PubMed

Eminović, Izet; Komel, Radovan; Prezelj, Janez; Karamehić, Jasenko; Gavrankapetanović, Faris; Heljić, Becir

2005-08-01

To examine the gene encoding for 5alpha-reductase type 1 in hyperandrogenic women, and assess the association of its eventual mutations or polymorphisms with the development of the hyperandrogenic female pattern. Sixteen hyperandrogenic women were included in the study. Single-stranded conformation polymorphism analysis (SSCP) and DNA sequencing were performed after polymerase chain reaction amplification of each of the 5 exons of the SRD5A1 gene in both hyperandrogenic and control group (16 participants). Sequence analysis identified the existence of many polymorphisms; in codon 24 of exon 1, GGC (Gly) into GAC (Asp); in codon 30 of exon 1, CGG (Arg) into CGC (Arg); in exon 3 codon 169, ACA to ACG (both encoding for threonine); in exon 5, AGA to AGG (both encoding for arginine, codon 260); and T/C polymorphism in intron 2. Polymorphisms were found in both groups. Polymorphisms of SRD5A1 gene were the same in both hyperandrogenic and healthy women, indicating no significant associations of genetic polymorphisms/variations of SRD5A1 gene with clinical manifestations of hyperandrogenic disorders in women.

Reactive oxygen species-generating mitochondrial DNA mutation up-regulates hypoxia-inducible factor-1alpha gene transcription via phosphatidylinositol 3-kinase-Akt/protein kinase C/histone deacetylase pathway.

PubMed

Koshikawa, Nobuko; Hayashi, Jun-Ichi; Nakagawara, Akira; Takenaga, Keizo

2009-11-27

Lewis lung carcinoma-derived high metastatic A11 cells constitutively overexpress hypoxia-inducible factor (HIF)-1alpha mRNA compared with low metastatic P29 cells. Because A11 cells exclusively possess a G13997A mutation in the mitochondrial NADH dehydrogenase subunit 6 (ND6) gene, we addressed here a causal relationship between the ND6 mutation and the activation of HIF-1alpha transcription, and we investigated the potential mechanism. Using trans-mitochondrial cybrids between A11 and P29 cells, we found that the ND6 mutation was directly involved in HIF-1alpha mRNA overexpression. Stimulation of HIF-1alpha transcription by the ND6 mutation was mediated by overproduction of reactive oxygen species (ROS) and subsequent activation of phosphatidylinositol 3-kinase (PI3K)-Akt and protein kinase C (PKC) signaling pathways. The up-regulation of HIF-1alpha transcription was abolished by mithramycin A, an Sp1 inhibitor, but luciferase reporter and chromatin immunoprecipitation assays indicated that Sp1 was necessary but not sufficient for HIF-1alpha mRNA overexpression in A11 cells. On the other hand, trichostatin A, a histone deacetylase (HDAC) inhibitor, markedly suppressed HIF-1alpha transcription in A11 cells. In accordance with this, HDAC activity was high in A11 cells but low in P29 cells and in A11 cells treated with the ROS scavenger ebselene, the PI3K inhibitor LY294002, and the PKC inhibitor Ro31-8220. These results suggest that the ROS-generating ND6 mutation increases HIF-1alpha transcription via the PI3K-Akt/PKC/HDAC pathway, leading to HIF-1alpha protein accumulation in hypoxic tumor cells.

Characterization of the Genes Encoding the Cytosolic and Plastidial Forms of ADP-Glucose Pyrophosphorylase in Wheat Endosperm1

PubMed Central

Burton, Rachel A.; Johnson, Philip E.; Beckles, Diane M.; Fincher, Geoffrey B.; Jenner, Helen L.; Naldrett, Mike J.; Denyer, Kay

2002-01-01

In most species, the synthesis of ADP-glucose (Glc) by the enzyme ADP-Glc pyrophosphorylase (AGPase) occurs entirely within the plastids in all tissues so far examined. However, in the endosperm of many, if not all grasses, a second form of AGPase synthesizes ADP-Glc outside the plastid, presumably in the cytosol. In this paper, we show that in the endosperm of wheat (Triticum aestivum), the cytosolic form accounts for most of the AGPase activity. Using a combination of molecular and biochemical approaches to identify the cytosolic and plastidial protein components of wheat endosperm AGPase we show that the large and small subunits of the cytosolic enzyme are encoded by genes previously thought to encode plastidial subunits, and that a gene, Ta.AGP.S.1, which encodes the small subunit of the cytosolic form of AGPase, also gives rise to a second transcript by the use of an alternate first exon. This second transcript encodes an AGPase small subunit with a transit peptide. However, we could not find a plastidial small subunit protein corresponding to this transcript. The protein sequence of the purified plastidial small subunit does not match precisely to that encoded by Ta.AGP.S.1 or to the predicted sequences of any other known gene from wheat or barley (Hordeum vulgare). Instead, the protein sequence is most similar to those of the plastidial small subunits from chickpea (Cicer arietinum) and maize (Zea mays) and rice (Oryza sativa) seeds. These data suggest that the gene encoding the major plastidial small subunit of AGPase in wheat endosperm has yet to be identified. PMID:12428011

Cloning and characterization of largemouth bass ( Micropterus salmoides) myostatin encoding gene and its promoter

NASA Astrophysics Data System (ADS)

Li, Shengjie; Bai, Junjie; Wang, Lin

2008-08-01

Myostatin or GDF-8, a member of the transforming growth factor-β (TGF-β) superfamily, has been demonstrated to be a negative regulator of skeletal muscle mass in mammals. In the present study, we obtained a 5.64 kb sequence of myostatin encoding gene and its promoter from largemouth bass ( Micropterus salmoides). The myostatin encoding gene consisted of three exons (488 bp, 371 bp and 1779 bp, respectively) and two introns (390 bp and 855 bp, respectively). The intron-exon boundaries were conservative in comparison with those of mammalian myostatin encoding genes, whereas the size of introns was smaller than that of mammals. Sequence analysis of 1.569 kb of the largemouth bass myostatin gene promoter region revealed that it contained two TATA boxes, one CAAT box and nine putative E-boxes. Putative muscle growth response elements for myocyte enhancer factor 2 (MEF2), serum response factor (SRF), activator protein 1 (AP1), etc., and muscle-specific Mt binding site (MTBF) were also detected. Some of the transcription factor binding sites were conserved among five teleost species. This information will be useful for studying the transcriptional regulation of myostatin in fish.

A member of a new plant gene family encoding a meprin and TRAF homology (MATH) domain-containing protein is involved in restriction of long distance movement of plant viruses

PubMed Central

Cosson, Patrick; Sofer, Luc; Schurdi-Levraud, Valérie

2010-01-01

Restriction of long distance movement of several potyviruses in Arabidopsis thaliana is controlled by at least three dominant restricted TEV movement (RTM) genes, named RTM1, RTM2 and RTM3 and acts as a non-conventional resistance. RTM1 encodes a protein belonging to the jacalin family and RTM2 encodes a protein which has similarities to small heat shock proteins. The recent cloning of RTM3 which encodes a protein belonging to an unknown protein family of 29 members that has a meprin and TRAF homology (MATH) domain in its N-terminal region and a coiled-coil (CC) domain at its C-terminal end is an important breakthrough for a better understanding of this resistance process. Not only the third gene involved in this resistance has been identified and has allowed revealing a new gene family in plant but the discovery that the RTM3 protein interacts directly with RTM1 strongly suggests that the RTM proteins form a multimeric complex. However, these data also highlight striking similarities of the RTM resistance with the well known R-gene mediated resistance. PMID:20930558

Structural and transcriptional characterization of a novel member of the soybean urease gene family.

PubMed

Wiebke-Strohm, Beatriz; Ligabue-Braun, Rodrigo; Rechenmacher, Ciliana; De Oliveira-Busatto, Luisa Abruzzi; Carlini, Célia Regina; Bodanese-Zanettini, Maria Helena

2016-04-01

In plants, ureases have been related to urea degradation, to defense against pathogenic fungi and phytophagous insects, and to the soybean-Bradyrhizobium japonicum symbiosis. Two urease isoforms have been described for soybean: the embryo-specific, encoded by Eu1 gene, and the ubiquitous urease, encoded by Eu4. A third urease-encoding locus exists in the completed soybean genome. The gene was designated Eu5 and the putative product of its ORF as SBU-III. Phylogenetic analysis shows that 41 plant, moss and algal ureases have diverged from a common ancestor protein, but ureases from monocots, eudicots and ancient species have evolved independently. Genomes of ancient organisms present a single urease-encoding gene and urease-encoding gene duplication has occurred independently along the evolution of some eudicot species. SBU-III has a shorter amino acid sequence, since many gaps are found when compared to other sequences. A mutation in a highly conserved amino acid residue suggests absence of ureolytic activity, but the overall protein architecture remains very similar to the other ureases. The expression profile of urease-encoding genes in different organs and developmental stages was determined by RT-qPCR. Eu5 transcripts were detected in seeds one day after dormancy break, roots of young plants and embryos of developing seeds. Eu1 and Eu4 transcripts were found in all analyzed organs, but Eu4 expression was more prominent in seeds one day after dormancy break whereas Eu1 predominated in developing seeds. The evidence suggests that SBU-III may not be involved in nitrogen availability to plants, but it could be involved in other biological role(s). Copyright © 2016 Elsevier Masson SAS. All rights reserved.

Co-overexpressing a plasma membrane and a vacuolar membrane sodium/proton antiporter significantly improves salt tolerance in transgenic Arabidopsis plants.

USDA-ARS?s Scientific Manuscript database

The Arabidopsis gene AtNHX1 encodes a vacuolar membrane bound sodium/proton (Sodium/Hydrogen) antiporter that transports sodium into the vacuole and exports hydrogen into the cytoplasm. The Arabidopsis gene SOS1 encodes a plasma membrane bound sodium/hydrogen antiporter that exports sodium to the ex...

Escherichia coli yjjPB genes encode a succinate transporter important for succinate production.

PubMed

Fukui, Keita; Nanatani, Kei; Hara, Yoshihiko; Yamakami, Suguru; Yahagi, Daiki; Chinen, Akito; Tokura, Mitsunori; Abe, Keietsu

2017-09-01

Under anaerobic conditions, Escherichia coli produces succinate from glucose via the reductive tricarboxylic acid cycle. To date, however, no genes encoding succinate exporters have been established in E. coli. Therefore, we attempted to identify genes encoding succinate exporters by screening an E. coli MG1655 genome library. We identified the yjjPB genes as candidates encoding a succinate transporter, which enhanced succinate production in Pantoea ananatis under aerobic conditions. A complementation assay conducted in Corynebacterium glutamicum strain AJ110655ΔsucE1 demonstrated that both YjjP and YjjB are required for the restoration of succinate production. Furthermore, deletion of yjjPB decreased succinate production in E. coli by 70% under anaerobic conditions. Taken together, these results suggest that YjjPB constitutes a succinate transporter in E. coli and that the products of both genes are required for succinate export.

RNA-Seq Analysis of the Expression of Genes Encoding Cell Wall Degrading Enzymes during Infection of Lupin (Lupinus angustifolius) by Phytophthora parasitica

PubMed Central

Blackman, Leila M.; Cullerne, Darren P.; Torreña, Pernelyn; Taylor, Jen; Hardham, Adrienne R.

2015-01-01

RNA-Seq analysis has shown that over 60% (12,962) of the predicted transcripts in the Phytophthora parasitica genome are expressed during the first 60 h of lupin root infection. The infection transcriptomes included 278 of the 431 genes encoding P. parasitica cell wall degrading enzymes. The transcriptome data provide strong evidence of global transcriptional cascades of genes whose encoded proteins target the main categories of plant cell wall components. A major cohort of pectinases is predominantly expressed early but as infection progresses, the transcriptome becomes increasingly dominated by transcripts encoding cellulases, hemicellulases, β-1,3-glucanases and glycoproteins. The most highly expressed P. parasitica carbohydrate active enzyme gene contains two CBM1 cellulose binding modules and no catalytic domains. The top 200 differentially expressed genes include β-1,4-glucosidases, β-1,4-glucanases, β-1,4-galactanases, a β-1,3-glucanase, an α-1,4-polygalacturonase, a pectin deacetylase and a pectin methylesterase. Detailed analysis of gene expression profiles provides clues as to the order in which linkages within the complex carbohydrates may come under attack. The gene expression profiles suggest that (i) demethylation of pectic homogalacturonan occurs before its deacetylation; (ii) cleavage of the backbone of pectic rhamnogalacturonan I precedes digestion of its side chains; (iii) early attack on cellulose microfibrils by non-catalytic cellulose-binding proteins and enzymes with auxiliary activities may facilitate subsequent attack by glycosyl hydrolases and enzymes containing CBM1 cellulose-binding modules; (iv) terminal hemicellulose backbone residues are targeted after extensive internal backbone cleavage has occurred; and (v) the carbohydrate chains on glycoproteins are degraded late in infection. A notable feature of the P. parasitica infection transcriptome is the high level of transcription of genes encoding enzymes that degrade β-1,3-glucanases during middle and late stages of infection. The results suggest that high levels of β-1,3-glucanases may effectively degrade callose as it is produced by the plant during the defence response. PMID:26332397

RNA-Seq Analysis of the Expression of Genes Encoding Cell Wall Degrading Enzymes during Infection of Lupin (Lupinus angustifolius) by Phytophthora parasitica.

PubMed

Blackman, Leila M; Cullerne, Darren P; Torreña, Pernelyn; Taylor, Jen; Hardham, Adrienne R

2015-01-01

RNA-Seq analysis has shown that over 60% (12,962) of the predicted transcripts in the Phytophthora parasitica genome are expressed during the first 60 h of lupin root infection. The infection transcriptomes included 278 of the 431 genes encoding P. parasitica cell wall degrading enzymes. The transcriptome data provide strong evidence of global transcriptional cascades of genes whose encoded proteins target the main categories of plant cell wall components. A major cohort of pectinases is predominantly expressed early but as infection progresses, the transcriptome becomes increasingly dominated by transcripts encoding cellulases, hemicellulases, β-1,3-glucanases and glycoproteins. The most highly expressed P. parasitica carbohydrate active enzyme gene contains two CBM1 cellulose binding modules and no catalytic domains. The top 200 differentially expressed genes include β-1,4-glucosidases, β-1,4-glucanases, β-1,4-galactanases, a β-1,3-glucanase, an α-1,4-polygalacturonase, a pectin deacetylase and a pectin methylesterase. Detailed analysis of gene expression profiles provides clues as to the order in which linkages within the complex carbohydrates may come under attack. The gene expression profiles suggest that (i) demethylation of pectic homogalacturonan occurs before its deacetylation; (ii) cleavage of the backbone of pectic rhamnogalacturonan I precedes digestion of its side chains; (iii) early attack on cellulose microfibrils by non-catalytic cellulose-binding proteins and enzymes with auxiliary activities may facilitate subsequent attack by glycosyl hydrolases and enzymes containing CBM1 cellulose-binding modules; (iv) terminal hemicellulose backbone residues are targeted after extensive internal backbone cleavage has occurred; and (v) the carbohydrate chains on glycoproteins are degraded late in infection. A notable feature of the P. parasitica infection transcriptome is the high level of transcription of genes encoding enzymes that degrade β-1,3-glucanases during middle and late stages of infection. The results suggest that high levels of β-1,3-glucanases may effectively degrade callose as it is produced by the plant during the defence response.

Molecular cloning of a Candida albicans gene (SSB1) coding for a protein related to the Hsp70 family.

PubMed

Maneu, V; Cervera, A M; Martinez, J P; Gozalbo, D

1997-06-15

We have cloned and sequenced a Candida albicans gene (SSB1) encoding a potential member of the heat-shock protein seventy (hsp70) family. The protein encoded by this gene contains 613 amino acids and shows a high degree (85%) of sequence identity to the ssb subfamily (ssb1 and ssb2) of the Saccharomyces cerevisiae hsp70 family. The transcribed mRNA (2.1 kb) is present in similar amounts both in yeast and germ tube cells of C. albicans.

Loss of functional K+ channels encoded by ether-à-go-go-related genes in mouse myometrium prior to labour onset

PubMed Central

Greenwood, I A; Yeung, S Y; Tribe, R M; Ohya, S

2009-01-01

There is a growing appreciation that ion channels encoded by the ether-à-go-go-related gene family have a functional impact in smooth muscle in addition to their accepted role in cardiac myocytes and neurones. This study aimed to assess the expression of ERG1–3 (KCNH1–3) genes in the murine myometrium (smooth muscle layer of the uterus) and determine the functional impact of the ion channels encoded by these genes in pregnant and non-pregnant animals. Quantitative RT-PCR did not detect message for ERG2 and 3 in whole myometrial tissue extracts. In contrast, message for two isoforms of mERG1 were readily detected with mERG1a more abundant than mERG1b. In isometric tension studies of non-pregnant myometrium, the ERG channel blockers dofetilide (1 μm), E4031 (1 μm) and Be-KM1 (100 nm) increased spontaneous contractility and ERG activators (PD118057 and NS1643) inhibited spontaneous contractility. In contrast, neither ERG blockade nor activation had any effect on the inherent contractility in myometrium from late pregnant (19 days gestation) animals. Moreover, dofetilide-sensitive K+ currents with distinctive ‘hooked’ kinetics were considerably smaller in uterine myocytes from late pregnant compared to non-pregnant animals. Expression of mERG1 isoforms did not alter throughout gestation or upon delivery, but the expression of genes encoding auxillary subunits (KCNE) were up-regulated considerably. This study provides the first evidence for a regulation of ERG-encoded K+ channels as a precursor to late pregnancy physiological activity. PMID:19332483

The ANGULATA7 gene encodes a DnaJ-like zinc finger-domain protein involved in chloroplast function and leaf development in Arabidopsis.

PubMed

Muñoz-Nortes, Tamara; Pérez-Pérez, José Manuel; Ponce, María Rosa; Candela, Héctor; Micol, José Luis

2017-03-01

The characterization of mutants with altered leaf shape and pigmentation has previously allowed the identification of nuclear genes that encode plastid-localized proteins that perform essential functions in leaf growth and development. A large-scale screen previously allowed us to isolate ethyl methanesulfonate-induced mutants with small rosettes and pale green leaves with prominent marginal teeth, which were assigned to a phenotypic class that we dubbed Angulata. The molecular characterization of the 12 genes assigned to this phenotypic class should help us to advance our understanding of the still poorly understood relationship between chloroplast biogenesis and leaf morphogenesis. In this article, we report the phenotypic and molecular characterization of the angulata7-1 (anu7-1) mutant of Arabidopsis thaliana, which we found to be a hypomorphic allele of the EMB2737 gene, which was previously known only for its embryonic-lethal mutations. ANU7 encodes a plant-specific protein that contains a domain similar to the central cysteine-rich domain of DnaJ proteins. The observed genetic interaction of anu7-1 with a loss-of-function allele of GENOMES UNCOUPLED1 suggests that the anu7-1 mutation triggers a retrograde signal that leads to changes in the expression of many genes that normally function in the chloroplasts. Many such genes are expressed at higher levels in anu7-1 rosettes, with a significant overrepresentation of those required for the expression of plastid genome genes. Like in other mutants with altered expression of plastid-encoded genes, we found that anu7-1 exhibits defects in the arrangement of thylakoidal membranes, which appear locally unappressed. © 2016 The Authors The Plant Journal © 2016 John Wiley & Sons Ltd.

Identification and Characterization of Three Differentially Expressed Genes, Encoding S-Adenosylhomocysteine Hydrolase, Methionine Aminopeptidase, and a Histone-Like Protein, in the Toxic Dinoflagellate Alexandrium fundyense†

PubMed Central

Taroncher-Oldenburg, Gaspar; Anderson, Donald M.

2000-01-01

Genes showing differential expression related to the early G1 phase of the cell cycle during synchronized circadian growth of the toxic dinoflagellate Alexandrium fundyense were identified and characterized by differential display (DD). The determination in our previous work that toxin production in Alexandrium is relegated to a narrow time frame in early G1 led to the hypothesis that transcriptionally up- or downregulated genes during this subphase of the cell cycle might be related to toxin biosynthesis. Three genes, encoding S-adenosylhomocysteine hydrolase (Sahh), methionine aminopeptidase (Map), and a histone-like protein (HAf), were isolated. Sahh was downregulated, while Map and HAf were upregulated, during the early G1 phase of the cell cycle. Sahh and Map encoded amino acid sequences with about 90 and 70% similarity to those encoded by several eukaryotic and prokaryotic Sahh and Map genes, respectively. The partial Map sequence also contained three cobalt binding motifs characteristic of all Map genes. HAf encoded an amino acid sequence with 60% similarity to those of two histone-like proteins from the dinoflagellate Crypthecodinium cohnii Biecheler. This study documents the potential of applying DD to the identification of genes that are related to physiological processes or cell cycle events in phytoplankton under conditions where small sample volumes represent an experimental constraint. The identification of an additional 21 genes with various cell cycle-related DD patterns also provides evidence for the importance of pretranslational or transcriptional regulation in dinoflagellates, contrary to previous reports suggesting the possibility that translational mechanisms are the primary means of circadian regulation in this group of organisms. PMID:10788388

Molecular characterization and expression of the M6 gene of grass carp hemorrhage virus (GCHV), an aquareovirus.

PubMed

Qiu, T; Lu, R H; Zhang, J; Zhu, Z Y

2001-07-01

The complete nucleotide sequence of M6 gene of grass carp hemorrhage virus (GCHV) was determined. It is 2039 nucleotides in length and contains a single large open reading frame that could encode a protein of 648 amino acids with predicted molecular mass of 68.7 kDa. Amino acid sequence comparison revealed that the protein encoded by GCHV M6 is closely related to the protein mu1 of mammalian reovirus. The M6 gene, encoding the major outer-capsid protein, was expressed using the pET fusion protein vector in Escherichia coli and detected by Western blotting using chicken anti-GCHV immunoglobulin (IgY). The result indicates that the protein encoded by M6 may share a putative Asn-42-Pro-43 proteolytic cleavage site with mu1.

Gene encoding γ-carbonic anhydrase is cotranscribed with argC and induced in response to stationary phase and high CO2 in Azospirillum brasilense Sp7

PubMed Central

2010-01-01

Background Carbonic anhydrase (CA) is a ubiquitous enzyme catalyzing the reversible hydration of CO2 to bicarbonate, a reaction underlying diverse biochemical and physiological processes. Gamma class carbonic anhydrases (γ-CAs) are widespread in prokaryotes but their physiological roles remain elusive. At present, only γ-CA of Methanosarcina thermophila (Cam) has been shown to have CA activity. Genome analysis of a rhizobacterium Azospirillum brasilense, revealed occurrence of ORFs encoding one β-CA and two γ-CAs. Results One of the putative γ-CA encoding genes of A. brasilense was cloned and overexpressed in E. coli. Electrometric assays for CA activity of the whole cell extracts overexpressing recombinant GCA1 did not show CO2 hydration activity. Reverse transcription-PCR analysis indicated that gca1 in A. brasilense is co-transcribed with its upstream gene annotated as argC, which encodes a putative N-acetyl-γ-glutamate-phosphate reductase. 5'-RACE also demonstrated that there was no transcription start site between argC and gca1, and the transcription start site located upstream of argC transcribed both the genes (argC-gca1). Using transcriptional fusions of argC-gca1 upstream region with promoterless lacZ, we further demonstrated that gca1 upstream region did not have any promoter and its transcription occurred from a promoter located in the argC upstream region. The transcription of argC-gca1 operon was upregulated in stationary phase and at elevated CO2 atmosphere. Conclusions This study shows lack of CO2 hydration activity in a recombinant protein expressed from a gene predicted to encode a γ-carbonic anhydrase in A. brasilense although it cross reacts with anti-Cam antibody raised against a well characterized γ-CA. The organization and regulation of this gene along with the putative argC gene suggests its involvement in arginine biosynthetic pathway instead of the predicted CO2 hydration. PMID:20598158

Gene encoding gamma-carbonic anhydrase is cotranscribed with argC and induced in response to stationary phase and high CO2 in Azospirillum brasilense Sp7.

PubMed

Kaur, Simarjot; Mishra, Mukti N; Tripathi, Anil K

2010-07-04

Carbonic anhydrase (CA) is a ubiquitous enzyme catalyzing the reversible hydration of CO2 to bicarbonate, a reaction underlying diverse biochemical and physiological processes. Gamma class carbonic anhydrases (gamma-CAs) are widespread in prokaryotes but their physiological roles remain elusive. At present, only gamma-CA of Methanosarcina thermophila (Cam) has been shown to have CA activity. Genome analysis of a rhizobacterium Azospirillum brasilense, revealed occurrence of ORFs encoding one beta-CA and two gamma-CAs. One of the putative gamma-CA encoding genes of A. brasilense was cloned and overexpressed in E. coli. Electrometric assays for CA activity of the whole cell extracts overexpressing recombinant GCA1 did not show CO2 hydration activity. Reverse transcription-PCR analysis indicated that gca1 in A. brasilense is co-transcribed with its upstream gene annotated as argC, which encodes a putative N-acetyl-gamma-glutamate-phosphate reductase. 5'-RACE also demonstrated that there was no transcription start site between argC and gca1, and the transcription start site located upstream of argC transcribed both the genes (argC-gca1). Using transcriptional fusions of argC-gca1 upstream region with promoterless lacZ, we further demonstrated that gca1 upstream region did not have any promoter and its transcription occurred from a promoter located in the argC upstream region. The transcription of argC-gca1 operon was upregulated in stationary phase and at elevated CO2 atmosphere. This study shows lack of CO2 hydration activity in a recombinant protein expressed from a gene predicted to encode a gamma-carbonic anhydrase in A. brasilense although it cross reacts with anti-Cam antibody raised against a well characterized gamma-CA. The organization and regulation of this gene along with the putative argC gene suggests its involvement in arginine biosynthetic pathway instead of the predicted CO2 hydration.

Mitochondrial DNA sequence analysis of four Alzheimer`s and Parkinson`s disease patients

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

Brown, M.D.; Shoffner, J.M.; Wallace, D.C.

1996-01-22

The mitochondrial DNA (mtDNA) sequence was determined on 3 patients with Alzheimer`s disease (AD) exhibiting AD plus Parkinson`s disease (PD) neuropathologic changes and one patient with PD. Patient mtDNA sequences were compared to the standard Cambridge sequence to identify base changes. In the first AD + PD patient, 2 of the 15 nucleotide substitutions may contribute to the neuropathology, a nucleotide pair (np) 4336 transition in the tRNA{sup Gln} gene found 7.4 times more frequently in patients than in controls, and a unique np 721 transition in the 12S rRNA gene which was not found in 70 other patients ormore » 905 controls. In the second AD + PD patient, 27 nucleotide substitutions were detected, including an np 3397 transition in the ND1 gene which converts a conserved methionine to a valine. In the third AD + PD patient, 2 polymorphic base substitutions frequently found at increased frequency in Leber`s hereditary optic neuropathy patients were observed, an np 4216 transition in ND1 and an np 13708 transition in the ND5 gene. For the PD patient, 2 novel variants were observed among 25 base substitutions, an np 1709 substitution in the 16S rRNA gene and an np 15851 missense mutation in the cytb gene. Further studies will be required to demonstrate a casual role for these base substitutions in neurodegenerative disease. 68 refs., 2 tabs.« less

Molecular cloning and characterization of alpha - galactosidase gene from Glaciozyma antarctica

NASA Astrophysics Data System (ADS)

Moheer, Reyad Qaed Al; Bakar, Farah Diba Abu; Murad, Abdul Munir Abdul

2015-09-01

Psychrophilic enzymes are proteins produced by psychrophilic organisms which recently are the limelight for industrial applications. A gene encoding α-galactosidase from a psychrophilic yeast, Glaciozyma antarctica PI12 which belongs to glycoside hydrolase family 27, was isolated and analyzed using several bioinformatic tools. The cDNA of the gene with the size of 1,404-bp encodes a protein with 467 amino acid residues. Predicted molecular weight of protein was 48.59 kDa and hence we name the gene encoding α-galactosidase as GAL48. We found that the predicted protein sequences possessed signal peptide sequence and are highly conserved among other fungal α-galactosidase.

Spreading of genes encoding enterotoxins, haemolysins, adhesin and biofilm among methicillin resistant Staphylococcus aureus strains with staphylococcal cassette chromosome mec type IIIA isolated from burn patients.

PubMed

Motallebi, Mitra; Jabalameli, Fereshteh; Asadollahi, Kheirollah; Taherikalani, Morovat; Emaneini, Mohammad

2016-08-01

The emergence of antibiotic-resistant Staphylococcus aureus in particular methicillin-resistant S. aureus (MRSA) is an important concern in burn medical centers either in Iran or worldwide. A total of 128 S. aureus isolates were collected from wound infection of burn patients during June 2013 to June 2014. Multiplex-polymerase chain reaction (MPCR) assay was performed for the characterization of the staphylococcal cassette chromosome mec (SCCmec). Genes encoding virulence factors and biofilm were targeted by PCR. Of 128 S. aureus isolates, 77 (60.1%) isolates were MRSA. Fifty four (70.1%) isolates were identified as SCCmec type IIIA. The most frequently detected toxin genes among MRSA isolates with SCCmec type IIIA were sea (64.1%) and hla (51.8%). The rate of coexistence of sea with hla and sea with hla and hlb was 37% and12.9%, respectively. The sec, eta, tst, pvl, hla and hlb genes were not detected in any of the MRSA isolates. The most prevalent genes encoding biofilm was eno, found in 61.1% of isolates, followed by fib and icaA found in 48.1% and 38.8% of the isolates, respectively. The rate of coexistence of fib + eno + icaA + icaD and fib + eno was 20.3% and 9.2%, respectively. The ebps gene was not detected in any of the isolates. In conclusion, our study indicated that the sea, hla, fib and icaA were most frequent genes encoding virulence factors among MRSA with SCCmec type IIIA isolated from burn wound infection. Moreover, the results of this study shows that the rate of coexistence of genes encoding different virulence factor were high. Copyright © 2016 Elsevier Ltd. All rights reserved.

Long-term persistence of a single Legionella pneumophila strain possessing the mip gene in a municipal shower despite repeated cycles of chlorination.

PubMed

Cooper, I R; White, J; Mahenthiralingam, E; Hanlon, G W

2008-10-01

The ability of Legionella pneumophila to colonise domestic water systems is a primary cause of outbreaks of Legionnaire's disease in humans. World Health Organization guidelines recommend that drinking water is chlorinated to between 0.2 and 1mg/L [Chlorine in drinking-water. Guidelines for drinking-water quality, 2nd edn. Geneva: World Health Organization; 1996], but L. pneumophila is repeatedly isolated from chlorinated water systems, indicating that this treatment is not effective at preventing colonisation. Current UK guidelines recommend a one-off treatment of 20-50mg/L of free chlorine to remove the bacteria. In this study we report on the persistence of L. pneumophila serogroup 1 in a domestic shower system despite repeated cycles of chlorination at 50mg/L for 1h exposure time, over the course of two and a half years. Persisting isolates were subjected to in-vitro phenotypic analyses and polymerase chain reaction analysis for the toxin-encoding mip gene. Random amplified polymorphic DNA typing was also performed to determine whether the isolates recovered on different occasions were the same strain. We found that seven isolates of L. pneumophila recovered over a two-and-a-half year period are the same genetically defined strain, indicating that the bacteria can persist despite repeated cycles of chlorination after each successive isolation.

Structure of CARB-4 and AER-1 CarbenicillinHydrolyzing β-Lactamases

PubMed Central

Sanschagrin, François; Bejaoui, Noureddine; Levesque, Roger C.

1998-01-01

We determined the nucleotide sequences of blaCARB-4 encoding CARB-4 and deduced a polypeptide of 288 amino acids. The gene was characterized as a variant of group 2c carbenicillin-hydrolyzing β-lactamases such as PSE-4, PSE-1, and CARB-3. The level of DNA homology between the bla genes for these β-lactamases varied from 98.7 to 99.9%, while that between these genes and blaCARB-4 encoding CARB-4 was 86.3%. The blaCARB-4 gene was acquired from some other source because it has a G+C content of 39.1%, compared to a G+C content of 67% for typical Pseudomonas aeruginosa genes. DNA sequencing revealed that blaAER-1 shared 60.8% DNA identity with blaPSE-3 encoding PSE-3. The deduced AER-1 β-lactamase peptide was compared to class A, B, C, and D enzymes and had 57.6% identity with PSE-3, including an STHK tetrad at the active site. For CARB-4 and AER-1, conserved canonical amino acid boxes typical of class A β-lactamases were identified in a multiple alignment. Analysis of the DNA sequences flanking blaCARB-4 and blaAER-1 confirmed the importance of gene cassettes acquired via integrons in bla gene distribution. PMID:9687391

A study of Staphylococcus aureusnasal carriage, antibacterial resistance and virulence factor encoding genes in a tertiary care hospital, Kayseri, Turkey.

PubMed

Oguzkaya-Artan, M; Artan, C; Baykan, Z; Sakalar, C; Turan, A; Aksu, H

2015-01-01

This study was to determine the virulence encoding genes, and the antibiotic resistance patterns of the Staphylococcus aureus isolates, which were isolated from the nasal samples of chest clinic patients. The nasal samples of the in-patients (431) and out-patients (1857) in Kayseri Training and Research Hospital's Chest Clinic, Kayseri, Turkey, were cultured on CHROMagar (Biolife, Italiana) S. aureus, and subcultured on sheep blood agar for the isolation of S. aureus. Disc diffusion method was used for antimicrobial susceptibility testing. The occurrence of the staphylococcal virulence encoding genes (enterotoksins [sea, seb, sec, see, seg, seh, sei, sej], fibronectin-binding proteins A, B [fnbA, fnbB], toxic shock syndrome toxin-1 [tst]) were detected by polymerase chain reaction. Forty-five of the 55 (81.8%) S. aureus isolates from inpatients, and 319 (90.6%) isolates from tested 352 out-patient's isolates were suspected to all the antibiotics tested. methicillin-resistant S. aureus (MRSA) was detected in 1.2% of S. aureus isolates. Rifampin, trimethoprim-sulfamethoxazole, clindamycin, erythromycin, gentamicin resistance rates were 1.2%, 1.7%, 2.0%, 8.8%, and 1.2%, respectively. The isolates were susceptible to teicoplanin and vancomycin. The genes most frequently found were tst (92.7%), seg (85.8%), sea (83.6%), fnbA (70.9%). There was no statistical significance detected between MRSA and mecA-negative S. aureus isolates in encoding genes distribution (P > 0.05). Our results show that virulence factor encoding genes were prevalent in patients with S. aureus carriage, whereas antibiotic resistance was low. These virulence determinants may increase the risk for subsequent invasive infections in carriers.

The Drosophila pigmentation gene pink (p) encodes a homologue of human Hermansky-Pudlak syndrome 5 (HPS5).

PubMed

Falcón-Pérez, Juan M; Romero-Calderón, Rafael; Brooks, Elizabeth S; Krantz, David E; Dell'Angelica, Esteban C

2007-02-01

Lysosome-related organelles comprise a group of specialized intracellular compartments that include melanosomes and platelet dense granules (in mammals) and eye pigment granules (in insects). In humans, the biogenesis of these organelles is defective in genetic disorders collectively known as Hermansky-Pudlak syndrome (HPS). Patients with HPS-2, and two murine HPS models, carry mutations in genes encoding subunits of adaptor protein (AP)-3. Other genes mutated in rodent models include those encoding VPS33A and Rab38. Orthologs of all of these genes in Drosophila melanogaster belong to the 'granule group' of eye pigmentation genes. Other genes associated with HPS encode subunits of three complexes of unknown function, named biogenesis of lysosome-related organelles complex (BLOC)-1, -2 and -3, for which the Drosophila counterparts had not been characterized. Here, we report that the gene encoding the Drosophila ortholog of the HPS5 subunit of BLOC-2 is identical to the granule group gene pink (p), which was first studied in 1910 but had not been identified at the molecular level. The phenotype of pink mutants was exacerbated by mutations in AP-3 subunits or in the orthologs of VPS33A and Rab38. These results validate D. melanogaster as a genetic model to study the function of the BLOCs.

Cloning and Sequence Analysis of Vibrio halioticoli Genes Encoding Three Types of Polyguluronate Lyase.

PubMed

Sugimura; Sawabe; Ezura

2000-01-01

The alginate lyase-coding genes of Vibrio halioticoli IAM 14596(T), which was isolated from the gut of the abalone Haliotis discus hannai, were cloned using plasmid vector pUC 18, and expressed in Escherichia coli. Three alginate lyase-positive clones, pVHB, pVHC, and pVHE, were obtained, and all clones expressed the enzyme activity specific for polyguluronate. Three genes, alyVG1, alyVG2, and alyVG3, encoding polyguluronate lyase were sequenced: alyVG1 from pVHB was composed of a 1056-bp open reading frame (ORF) encoding 352 amino acid residues; alyVG2 gene from pVHC was composed of a 993-bp ORF encoding 331 amino acid residues; and alyVG3 gene from pVHE was composed of a 705-bp ORF encoding 235 amino acid residues. Comparison of nucleotide and deduced amino acid sequences among AlyVG1, AlyVG2, and AlyVG3 revealed low homologies. The identity value between AlyVG1 and AlyVG2 was 18.7%, and that between AlyVG2 and AlyVG3 was 17.0%. A higher identity value (26.0%) was observed between AlyVG1 and AlyVG3. Sequence comparison among known polyguluronate lyases including AlyVG1, AlyVG2, and AlyVG3 also did not reveal an identical region in these sequences. However, AlyVG1 showed the highest identity value (36.2%) and the highest similarity (73.3%) to AlyA from Klebsiella pneumoniae. A consensus region comprising nine amino acid (YFKAGXYXQ) in the carboxy-terminal region previously reported by Mallisard and colleagues was observed only in AlyVG1 and AlyVG2.

Systematic Analysis and Comparison of Nucleotide-Binding Site Disease Resistance Genes in a Diploid Cotton Gossypium raimondii

PubMed Central

Wei, Hengling; Li, Wei; Sun, Xiwei; Zhu, Shuijin; Zhu, Jun

2013-01-01

Plant disease resistance genes are a key component of defending plants from a range of pathogens. The majority of these resistance genes belong to the super-family that harbors a Nucleotide-binding site (NBS). A number of studies have focused on NBS-encoding genes in disease resistant breeding programs for diverse plants. However, little information has been reported with an emphasis on systematic analysis and comparison of NBS-encoding genes in cotton. To fill this gap of knowledge, in this study, we identified and investigated the NBS-encoding resistance genes in cotton using the whole genome sequence information of Gossypium raimondii. Totally, 355 NBS-encoding resistance genes were identified. Analyses of the conserved motifs and structural diversity showed that the most two distinct features for these genes are the high proportion of non-regular NBS genes and the high diversity of N-termini domains. Analyses of the physical locations and duplications of NBS-encoding genes showed that gene duplication of disease resistance genes could play an important role in cotton by leading to an increase in the functional diversity of the cotton NBS-encoding genes. Analyses of phylogenetic comparisons indicated that, in cotton, the NBS-encoding genes with TIR domain not only have their own evolution pattern different from those of genes without TIR domain, but also have their own species-specific pattern that differs from those of TIR genes in other plants. Analyses of the correlation between disease resistance QTL and NBS-encoding resistance genes showed that there could be more than half of the disease resistance QTL associated to the NBS-encoding genes in cotton, which agrees with previous studies establishing that more than half of plant resistance genes are NBS-encoding genes. PMID:23936305

Characterization of a Gene Encoding Clathrin Heavy Chain in Maize Up-Regulated by Salicylic Acid, Abscisic Acid and High Boron Supply

PubMed Central

Zeng, Mu-Heng; Liu, Sheng-Hong; Yang, Miao-Xian; Zhang, Ya-Jun; Liang, Jia-Yong; Wan, Xiao-Rong; Liang, Hong

2013-01-01

Clathrin, a three-legged triskelion composed of three clathrin heavy chains (CHCs) and three light chains (CLCs), plays a critical role in clathrin-mediated endocytosis (CME) in eukaryotic cells. In this study, the genes ZmCHC1 and ZmCHC2 encoding clathrin heavy chain in maize were cloned and characterized for the first time in monocots. ZmCHC1 encodes a 1693-amino acid-protein including 29 exons and 28 introns, and ZmCHC2 encodes a 1746-amino acid-protein including 28 exons and 27 introns. The high similarities of gene structure, protein sequences and 3D models among ZmCHC1, and Arabidopsis AtCHC1 and AtCHC2 suggest their similar functions in CME. ZmCHC1 gene is predominantly expressed in maize roots instead of ubiquitous expression of ZmCHC2. Consistent with a typical predicted salicylic acid (SA)-responsive element and four predicted ABA-responsive elements (ABREs) in the promoter sequence of ZmCHC1, the expression of ZmCHC1 instead of ZmCHC2 in maize roots is significantly up-regulated by SA or ABA, suggesting that ZmCHC1 gene may be involved in the SA signaling pathway in maize defense responses. The expressions of ZmCHC1 and ZmCHC2 genes in maize are down-regulated by azide or cold treatment, further revealing the energy requirement of CME and suggesting that CME in plants is sensitive to low temperatures. PMID:23880865

Effect of Roux-en-Y gastric bypass on the distribution and hormone expression of small-intestinal enteroendocrine cells in obese patients with type 2 diabetes.

PubMed

Rhee, Nicolai A; Wahlgren, Camilla D; Pedersen, Jens; Mortensen, Brynjulf; Langholz, Ebbe; Wandall, Erik P; Friis, Steffen U; Vilmann, Peter; Paulsen, Sarah J; Kristiansen, Viggo B; Jelsing, Jacob; Dalbøge, Louise S; Poulsen, Steen S; Holst, Jens J; Vilsbøll, Tina; Knop, Filip K

2015-10-01

We studied the impact of Roux-en-Y gastric bypass (RYGB) on the density and hormonal gene expression of small-intestinal enteroendocrine cells in obese patients with type 2 diabetes. Twelve patients with diabetes and 11 age- and BMI-matched controls underwent RYGB followed by enteroscopy ~10 months later. Mucosal biopsies taken during surgery and enteroscopy were immunohistochemically stained for glucagon-like peptide-1 (GLP-1), peptide YY (PYY), cholecystokinin (CCK), glucose-dependent insulinotropic polypeptide (GIP) and prohormone convertase 2 (PC2) and the expression of GCG (encoding preproglucagon), PYY, CCK, GIP, GHRL (encoding ghrelin), SCT (encoding secretin), NTS (encoding neurotensin) and NR1H4 (encoding farnesoid X receptor) was evaluated. The density of cells immunoreactive for GLP-1, CCK and GIP increased in patients after RYGB and the density of those immunoreactive for GLP-1, PYY, CCK and PC2 increased in controls. In both groups, GHRL, SCT and GIP mRNA was reduced after RYGB while PYY, CCK, NTS and NR1H4 gene expression was unaltered. GCG mRNA was upregulated in both groups. Numerous alterations in the distribution of enteroendocrine cells and their expression of hormonal genes are seen after RYGB and include increased density of GLP-1-, PYY-, CCK-, GIP- and PC2-positive cells, reduced gene expression of GHRL, SCT and GIP and increased expression of GCG.

The euryhaline yeast Debaryomyces hansenii has two catalase genes encoding enzymes with differential activity profile.

PubMed

Segal-Kischinevzky, Claudia; Rodarte-Murguía, Beatriz; Valdés-López, Victor; Mendoza-Hernández, Guillermo; González, Alicia; Alba-Lois, Luisa

2011-03-01

Debaryomyces hansenii is a spoilage yeast able to grow in a variety of ecological niches, from seawater to dairy products. Results presented in this article show that (i) D. hansenii has an inherent resistance to H2O2 which could be attributed to the fact that this yeast has a basal catalase activity which is several-fold higher than that observed in Saccharomyces cerevisiae under the same culture conditions, (ii) D. hansenii has two genes (DhCTA1 and DhCTT1) encoding two catalase isozymes with a differential enzymatic activity profile which is not strictly correlated with a differential expression profile of the encoding genes.

Regulation of Neurospora crassa cell wall remodeling via the cot-1 pathway is mediated by gul-1.

PubMed

Herold, Inbal; Yarden, Oded

2017-02-01

Impairment of the Neurospora crassa Nuclear DBF2-related kinase-encoding gene cot-1 results in pleiotropic effects, including abnormally thick hyphal cell walls and septa. An increase in the transcript abundance of genes encoding chitin and glucan synthases and the chitinase gh18-5, but not the cell wall integrity pathway transcription factor rlm-1, accompany the phenotypic changes observed. Deletion of chs-5 or chs-7 in a cot-1 background results in a reduction of hyperbranching frequency characteristic of the cot-1 parent. gul-1 (a homologue of the yeast SSD1 gene) encodes a translational regulator and has been shown to partially suppress cot-1. We demonstrate that the high expression levels of the cell wall remodeling genes analyzed is curbed, and reaches near wild type levels, when gul-1 is inactivated. This is accompanied by morphological changes that include reduced cell wall thickness and restoration of normal chitin levels. We conclude that gul-1 is a mediator of cell wall remodeling within the cot-1 pathway.

Genome-wide comparative analysis of NBS-encoding genes between Brassica species and Arabidopsis thaliana.

PubMed

Yu, Jingyin; Tehrim, Sadia; Zhang, Fengqi; Tong, Chaobo; Huang, Junyan; Cheng, Xiaohui; Dong, Caihua; Zhou, Yanqiu; Qin, Rui; Hua, Wei; Liu, Shengyi

2014-01-03

Plant disease resistance (R) genes with the nucleotide binding site (NBS) play an important role in offering resistance to pathogens. The availability of complete genome sequences of Brassica oleracea and Brassica rapa provides an important opportunity for researchers to identify and characterize NBS-encoding R genes in Brassica species and to compare with analogues in Arabidopsis thaliana based on a comparative genomics approach. However, little is known about the evolutionary fate of NBS-encoding genes in the Brassica lineage after split from A. thaliana. Here we present genome-wide analysis of NBS-encoding genes in B. oleracea, B. rapa and A. thaliana. Through the employment of HMM search and manual curation, we identified 157, 206 and 167 NBS-encoding genes in B. oleracea, B. rapa and A. thaliana genomes, respectively. Phylogenetic analysis among 3 species classified NBS-encoding genes into 6 subgroups. Tandem duplication and whole genome triplication (WGT) analyses revealed that after WGT of the Brassica ancestor, NBS-encoding homologous gene pairs on triplicated regions in Brassica ancestor were deleted or lost quickly, but NBS-encoding genes in Brassica species experienced species-specific gene amplification by tandem duplication after divergence of B. rapa and B. oleracea. Expression profiling of NBS-encoding orthologous gene pairs indicated the differential expression pattern of retained orthologous gene copies in B. oleracea and B. rapa. Furthermore, evolutionary analysis of CNL type NBS-encoding orthologous gene pairs among 3 species suggested that orthologous genes in B. rapa species have undergone stronger negative selection than those in B .oleracea species. But for TNL type, there are no significant differences in the orthologous gene pairs between the two species. This study is first identification and characterization of NBS-encoding genes in B. rapa and B. oleracea based on whole genome sequences. Through tandem duplication and whole genome triplication analysis in B. oleracea, B. rapa and A. thaliana genomes, our study provides insight into the evolutionary history of NBS-encoding genes after divergence of A. thaliana and the Brassica lineage. These results together with expression pattern analysis of NBS-encoding orthologous genes provide useful resource for functional characterization of these genes and genetic improvement of relevant crops.

Ornithorhynchus anatinus (platypus) links the evolution of immunoglobulin genes in eutherian mammals and nonmammalian tetrapods.

PubMed

Zhao, Yaofeng; Cui, Huiting; Whittington, Camilla M; Wei, Zhiguo; Zhang, Xiaofeng; Zhang, Ziding; Yu, Li; Ren, Liming; Hu, Xiaoxiang; Zhang, Yaping; Hellman, Lars; Belov, Katherine; Li, Ning; Hammarström, Lennart

2009-09-01

The evolutionary origins of mammalian immunoglobulin H chain isotypes (IgM, IgD, IgG, IgE, and IgA) are still incompletely understood as these isotypes differ considerably in structure and number from their counterparts in nonmammalian tetrapods. We report in this study that the platypus (Ornithorhynchus anatinus) Ig H chain constant region gene locus contains eight Ig encoding genes, which are arranged in an mu-delta-omicron-gamma2-gamma1-alpha1-epsilon-alpha2 order, spanning a total of approximately 200 kb DNA, encoding six distinct isotypes. The omicron (omicron for Ornithorhynchus) gene encodes a novel Ig H chain isotype that consists of four constant region domains and a hinge, and is structurally different from any of the five known mammalian Ig classes. This gene is phylogenetically related to upsilon (epsilon) and gamma, and thus appears to be a structural intermediate between these two genes. The platypus delta gene encodes ten heavy chain constant region domains, lacks a hinge region and is similar to IgD in amphibians and fish, but strikingly different from that in eutherian mammals. The platypus Ig H chain isotype repertoire thus shows a unique combination of genes that share similarity both to those of nonmammalian tetrapods and eutherian animals and demonstrates how phylogenetically informative species can be used to reconstruct the evolutionary history of functionally important genes.

Cloning, sequence, and disruption of the Saccharomyces diastaticus DAR1 gene encoding a glycerol-3-phosphate dehydrogenase.

PubMed Central

Wang, H T; Rahaim, P; Robbins, P; Yocum, R R

1994-01-01

The Saccharomyces diastaticus DAR1 gene was cloned by complementation in an Escherichia coli strain auxogrophic for glycerol-3-phosphate. DAR1 encodes an NADH-dependent dihydroxyacetone phosphate reductase (sn-glycerol-3-phosphate dehydrogenase [G3PDase; EC 1.1.1.8]) homologous to several other eukaryotic G3PDases. DAR1 is distinct from GUT2, which encodes a glucose-repressed mitochondrial G3PDase, but is identical to GPD1 from S. cerevisiae, a close relative of S. diastaticus. The level of DAR1-encoded G3PDase was increased about threefold in a medium of high osmolarity. Disruption of DAR1 in a haploid S. cerevisiae was not lethal but led to a decrease in cytoplasmic NADH-dependent G3PDase activity, an increase in osmotic sensitivity, and a 25% reduction in glycerol secretion from cells grown anaerobically on glucose. PMID:7961476

Cloning, sequence, and disruption of the Saccharomyces diastaticus DAR1 gene encoding a glycerol-3-phosphate dehydrogenase.

PubMed

Wang, H T; Rahaim, P; Robbins, P; Yocum, R R

1994-11-01

The Saccharomyces diastaticus DAR1 gene was cloned by complementation in an Escherichia coli strain auxogrophic for glycerol-3-phosphate. DAR1 encodes an NADH-dependent dihydroxyacetone phosphate reductase (sn-glycerol-3-phosphate dehydrogenase [G3PDase; EC 1.1.1.8]) homologous to several other eukaryotic G3PDases. DAR1 is distinct from GUT2, which encodes a glucose-repressed mitochondrial G3PDase, but is identical to GPD1 from S. cerevisiae, a close relative of S. diastaticus. The level of DAR1-encoded G3PDase was increased about threefold in a medium of high osmolarity. Disruption of DAR1 in a haploid S. cerevisiae was not lethal but led to a decrease in cytoplasmic NADH-dependent G3PDase activity, an increase in osmotic sensitivity, and a 25% reduction in glycerol secretion from cells grown anaerobically on glucose.

A Cluster of Five Genes Essential for the Utilization of Dihydroxamate Xenosiderophores in Synechocystis sp. PCC 6803.

PubMed

Obando S, Tobias A; Babykin, Michael M; Zinchenko, Vladislav V

2018-05-21

The unicellular freshwater cyanobacterium Synechocystis sp. PCC 6803 is capable of using dihydroxamate xenosiderophores, either ferric schizokinen (FeSK) or a siderophore of the filamentous cyanobacterium Anabaena variabilis ATCC 29413 (SAV), as the sole source of iron in the TonB-dependent manner. The fecCDEB1-schT gene cluster encoding a siderophore transport system that is involved in the utilization of FeSK and SAV in Synechocystis sp. PCC 6803 was identified. The gene schT encodes TonB-dependent outer membrane transporter, whereas the remaining four genes encode the ABC-type transporter FecB1CDE formed by the periplasmic binding protein FecB1, the transmembrane permease proteins FecC and FecD, and the ATPase FecE. Inactivation of any of these genes resulted in the inability of cells to utilize FeSK and SAV. Our data strongly suggest that Synechocystis sp. PCC 6803 can readily internalize Fe-siderophores via the classic TonB-dependent transport system.

Regulation of sugar transport and metabolism by the Candida albicans Rgt1 transcriptional repressor.

PubMed

Sexton, Jessica A; Brown, Victoria; Johnston, Mark

2007-10-01

The ability of the fungal pathogen Candida albicans to cause systemic infections depends in part on the function of Hgt4, a cell surface sugar sensor. The orthologues of Hgt4 in Saccharomyces cerevisiae, Snf3 and Rgt2, initiate a signalling cascade that inactivates Rgt1, a transcriptional repressor of genes encoding hexose transporters. To determine whether Hgt4 functions similarly through the C. albicans orthologue of Rgt1, we analysed Cargt1 deletion mutants. We found that Cargt1 mutants are sensitive to the glucose analogue 2-deoxyglucose, a phenotype probably due to uncontrolled expression of genes encoding glucose transporters. Indeed, transcriptional profiling revealed that expression of about two dozen genes, including multiple HGT genes encoding hexose transporters, is increased in the Cargt1 mutant in the absence of sugars, suggesting that CaRgt1 represses expression of several HGT genes under this condition. Some of the HGT genes (probably encoding high-affinity transporters) are also repressed by high levels of glucose, and we show that this repression is mediated by CaMig1, the orthologue of the major glucose-activated repressor in S. cerevisiae, but not by its paralogue CaMig2. Therefore, CaRgt1 and CaMig1 collaborate to control expression of C. albicans hexose transporters in response to different levels of sugars. We were surprised to find that CaRgt1 also regulates expression of GAL1, suggesting that regulation of galactose metabolism in C. albicans is unconventional. Finally, Cargt1 mutations cause cells to hyperfilament, and suppress the hypofilamented phenotype of an hgt4 mutant, indicating that the Hgt4 glucose sensor may affect filamentation by modulating sugar import and metabolism via CaRgt1. Copyright 2007 John Wiley & Sons, Ltd.

In silico analysis of β-mannanases and β-mannosidase from Aspergillus flavus and Trichoderma virens UKM1

NASA Astrophysics Data System (ADS)

Yee, Chai Sin; Murad, Abdul Munir Abdul; Bakar, Farah Diba Abu

2013-11-01

A gene encoding an endo-β-1,4-mannanase from Trichoderma virens UKM1 (manTV) and Aspergillus flavus UKM1 (manAF) was analysed with bioinformatic tools. In addition, A. flavus NRRL 3357 genome database was screened for a β-mannosidase gene and analysed (mndA-AF). These three genes were analysed to understand their gene properties. manTV and manAF both consists of 1,332-bp and 1,386-bp nucleotides encoding 443 and 461 amino acid residues, respectively. Both the endo-β-1,4-mannanases belong to the glycosyl hydrolase family 5 and contain a carbohydrate-binding module family 1 (CBM1). On the other hand, mndA-AF which is a 2,745-bp gene encodes a protein sequence of 914 amino acid residues. This β-mannosidase belongs to the glycosyl hydrolase family 2. Predicted molecular weight of manTV, manAF and mndA-AF are 47.74 kDa, 49.71 kDa and 103 kDa, respectively. All three predicted protein sequences possessed signal peptide sequence and are highly conserved among other fungal β-mannanases and β-mannosidases.

Novel and recurrent mutations in the C1NH gene of Arab patients affected with hereditary angioedema.

PubMed

Faiyaz-Ul-Haque, Muhammad; Al-Gazlan, Sulaiman; Abalkhail, Halah A; Al-Abdulatif, Ahmad; Toulimat, Mohamed; Peltekova, Iskra; Khaliq, Agha M R; Al-Dayel, Fouad; Zaidi, Syed H E

2010-01-01

Autosomal dominant hereditary angioedema (HAE) results in episodes of subcutaneous edema in any body part and/or submucosal edema of the upper respiratory or gastrointestinal tracts. This disorder is caused by mutations in the C1NH gene, many of which have been described primarily in European patients. However, the genetic cause of HAE in Middle Eastern Arab patients has not yet been determined. Four unrelated Arab families, in which 15 patients were diagnosed with HAE, were studied. DNA from 13 patients was analyzed for mutations in the C1NH gene by DNA sequencing. Three novel and 2 recurrent mutations were identified in the C1NH gene of HAE patients. In family 1, the patient was heterozygous for a novel c.856C>T and a recurrent c.1361T>A missense mutation encoding for p.Arg264Cys and p.Val432Glu, respectively. In patients from family 2, a novel c.509C>T missense mutation encoding for a p.Ser148Phe was identified. In patients from family 3, a novel c.1142delC nonsense mutation encoding for a p.Ala359AlafsX15 was discovered. In family 4, a recurrent c.1397G>A missense mutation encoding for a p.Arg444His was present. This is the first ever report of C1NH gene mutations in Middle Eastern Arab patients. Our study suggests that, despite the numerous existing mutations in the C1NH gene, there are novel and recurrent mutations in HAE patients of non-European origin. We conclude that the spectrum of C1NH gene mutations in HAE patients is wider due to the likely presence of novel and recurrent mutations in patients of other ethnicities. 2009 S. Karger AG, Basel.

Whole-Exome Sequencing Identifies Rare and Low-Frequency Coding Variants Associated with LDL Cholesterol

PubMed Central

Lange, Leslie A.; Hu, Youna; Zhang, He; Xue, Chenyi; Schmidt, Ellen M.; Tang, Zheng-Zheng; Bizon, Chris; Lange, Ethan M.; Smith, Joshua D.; Turner, Emily H.; Jun, Goo; Kang, Hyun Min; Peloso, Gina; Auer, Paul; Li, Kuo-ping; Flannick, Jason; Zhang, Ji; Fuchsberger, Christian; Gaulton, Kyle; Lindgren, Cecilia; Locke, Adam; Manning, Alisa; Sim, Xueling; Rivas, Manuel A.; Holmen, Oddgeir L.; Gottesman, Omri; Lu, Yingchang; Ruderfer, Douglas; Stahl, Eli A.; Duan, Qing; Li, Yun; Durda, Peter; Jiao, Shuo; Isaacs, Aaron; Hofman, Albert; Bis, Joshua C.; Correa, Adolfo; Griswold, Michael E.; Jakobsdottir, Johanna; Smith, Albert V.; Schreiner, Pamela J.; Feitosa, Mary F.; Zhang, Qunyuan; Huffman, Jennifer E.; Crosby, Jacy; Wassel, Christina L.; Do, Ron; Franceschini, Nora; Martin, Lisa W.; Robinson, Jennifer G.; Assimes, Themistocles L.; Crosslin, David R.; Rosenthal, Elisabeth A.; Tsai, Michael; Rieder, Mark J.; Farlow, Deborah N.; Folsom, Aaron R.; Lumley, Thomas; Fox, Ervin R.; Carlson, Christopher S.; Peters, Ulrike; Jackson, Rebecca D.; van Duijn, Cornelia M.; Uitterlinden, André G.; Levy, Daniel; Rotter, Jerome I.; Taylor, Herman A.; Gudnason, Vilmundur; Siscovick, David S.; Fornage, Myriam; Borecki, Ingrid B.; Hayward, Caroline; Rudan, Igor; Chen, Y. Eugene; Bottinger, Erwin P.; Loos, Ruth J.F.; Sætrom, Pål; Hveem, Kristian; Boehnke, Michael; Groop, Leif; McCarthy, Mark; Meitinger, Thomas; Ballantyne, Christie M.; Gabriel, Stacey B.; O’Donnell, Christopher J.; Post, Wendy S.; North, Kari E.; Reiner, Alexander P.; Boerwinkle, Eric; Psaty, Bruce M.; Altshuler, David; Kathiresan, Sekar; Lin, Dan-Yu; Jarvik, Gail P.; Cupples, L. Adrienne; Kooperberg, Charles; Wilson, James G.; Nickerson, Deborah A.; Abecasis, Goncalo R.; Rich, Stephen S.; Tracy, Russell P.; Willer, Cristen J.; Gabriel, Stacey B.; Altshuler, David M.; Abecasis, Gonçalo R.; Allayee, Hooman; Cresci, Sharon; Daly, Mark J.; de Bakker, Paul I.W.; DePristo, Mark A.; Do, Ron; Donnelly, Peter; Farlow, Deborah N.; Fennell, Tim; Garimella, Kiran; Hazen, Stanley L.; Hu, Youna; Jordan, Daniel M.; Jun, Goo; Kathiresan, Sekar; Kang, Hyun Min; Kiezun, Adam; Lettre, Guillaume; Li, Bingshan; Li, Mingyao; Newton-Cheh, Christopher H.; Padmanabhan, Sandosh; Peloso, Gina; Pulit, Sara; Rader, Daniel J.; Reich, David; Reilly, Muredach P.; Rivas, Manuel A.; Schwartz, Steve; Scott, Laura; Siscovick, David S.; Spertus, John A.; Stitziel, Nathaniel O.; Stoletzki, Nina; Sunyaev, Shamil R.; Voight, Benjamin F.; Willer, Cristen J.; Rich, Stephen S.; Akylbekova, Ermeg; Atwood, Larry D.; Ballantyne, Christie M.; Barbalic, Maja; Barr, R. Graham; Benjamin, Emelia J.; Bis, Joshua; Boerwinkle, Eric; Bowden, Donald W.; Brody, Jennifer; Budoff, Matthew; Burke, Greg; Buxbaum, Sarah; Carr, Jeff; Chen, Donna T.; Chen, Ida Y.; Chen, Wei-Min; Concannon, Pat; Crosby, Jacy; Cupples, L. Adrienne; D’Agostino, Ralph; DeStefano, Anita L.; Dreisbach, Albert; Dupuis, Josée; Durda, J. Peter; Ellis, Jaclyn; Folsom, Aaron R.; Fornage, Myriam; Fox, Caroline S.; Fox, Ervin; Funari, Vincent; Ganesh, Santhi K.; Gardin, Julius; Goff, David; Gordon, Ora; Grody, Wayne; Gross, Myron; Guo, Xiuqing; Hall, Ira M.; Heard-Costa, Nancy L.; Heckbert, Susan R.; Heintz, Nicholas; Herrington, David M.; Hickson, DeMarc; Huang, Jie; Hwang, Shih-Jen; Jacobs, David R.; Jenny, Nancy S.; Johnson, Andrew D.; Johnson, Craig W.; Kawut, Steven; Kronmal, Richard; Kurz, Raluca; Lange, Ethan M.; Lange, Leslie A.; Larson, Martin G.; Lawson, Mark; Lewis, Cora E.; Levy, Daniel; Li, Dalin; Lin, Honghuang; Liu, Chunyu; Liu, Jiankang; Liu, Kiang; Liu, Xiaoming; Liu, Yongmei; Longstreth, William T.; Loria, Cay; Lumley, Thomas; Lunetta, Kathryn; Mackey, Aaron J.; Mackey, Rachel; Manichaikul, Ani; Maxwell, Taylor; McKnight, Barbara; Meigs, James B.; Morrison, Alanna C.; Musani, Solomon K.; Mychaleckyj, Josyf C.; Nettleton, Jennifer A.; North, Kari; O’Donnell, Christopher J.; O’Leary, Daniel; Ong, Frank; Palmas, Walter; Pankow, James S.; Pankratz, Nathan D.; Paul, Shom; Perez, Marco; Person, Sharina D.; Polak, Joseph; Post, Wendy S.; Psaty, Bruce M.; Quinlan, Aaron R.; Raffel, Leslie J.; Ramachandran, Vasan S.; Reiner, Alexander P.; Rice, Kenneth; Rotter, Jerome I.; Sanders, Jill P.; Schreiner, Pamela; Seshadri, Sudha; Shea, Steve; Sidney, Stephen; Silverstein, Kevin; Smith, Nicholas L.; Sotoodehnia, Nona; Srinivasan, Asoke; Taylor, Herman A.; Taylor, Kent; Thomas, Fridtjof; Tracy, Russell P.; Tsai, Michael Y.; Volcik, Kelly A.; Wassel, Chrstina L.; Watson, Karol; Wei, Gina; White, Wendy; Wiggins, Kerri L.; Wilk, Jemma B.; Williams, O. Dale; Wilson, Gregory; Wilson, James G.; Wolf, Phillip; Zakai, Neil A.; Hardy, John; Meschia, James F.; Nalls, Michael; Singleton, Andrew; Worrall, Brad; Bamshad, Michael J.; Barnes, Kathleen C.; Abdulhamid, Ibrahim; Accurso, Frank; Anbar, Ran; Beaty, Terri; Bigham, Abigail; Black, Phillip; Bleecker, Eugene; Buckingham, Kati; Cairns, Anne Marie; Caplan, Daniel; Chatfield, Barbara; Chidekel, Aaron; Cho, Michael; Christiani, David C.; Crapo, James D.; Crouch, Julia; Daley, Denise; Dang, Anthony; Dang, Hong; De Paula, Alicia; DeCelie-Germana, Joan; Drumm, Allen DozorMitch; Dyson, Maynard; Emerson, Julia; Emond, Mary J.; Ferkol, Thomas; Fink, Robert; Foster, Cassandra; Froh, Deborah; Gao, Li; Gershan, William; Gibson, Ronald L.; Godwin, Elizabeth; Gondor, Magdalen; Gutierrez, Hector; Hansel, Nadia N.; Hassoun, Paul M.; Hiatt, Peter; Hokanson, John E.; Howenstine, Michelle; Hummer, Laura K.; Kanga, Jamshed; Kim, Yoonhee; Knowles, Michael R.; Konstan, Michael; Lahiri, Thomas; Laird, Nan; Lange, Christoph; Lin, Lin; Lin, Xihong; Louie, Tin L.; Lynch, David; Make, Barry; Martin, Thomas R.; Mathai, Steve C.; Mathias, Rasika A.; McNamara, John; McNamara, Sharon; Meyers, Deborah; Millard, Susan; Mogayzel, Peter; Moss, Richard; Murray, Tanda; Nielson, Dennis; Noyes, Blakeslee; O’Neal, Wanda; Orenstein, David; O’Sullivan, Brian; Pace, Rhonda; Pare, Peter; Parker, H. Worth; Passero, Mary Ann; Perkett, Elizabeth; Prestridge, Adrienne; Rafaels, Nicholas M.; Ramsey, Bonnie; Regan, Elizabeth; Ren, Clement; Retsch-Bogart, George; Rock, Michael; Rosen, Antony; Rosenfeld, Margaret; Ruczinski, Ingo; Sanford, Andrew; Schaeffer, David; Sell, Cindy; Sheehan, Daniel; Silverman, Edwin K.; Sin, Don; Spencer, Terry; Stonebraker, Jackie; Tabor, Holly K.; Varlotta, Laurie; Vergara, Candelaria I.; Weiss, Robert; Wigley, Fred; Wise, Robert A.; Wright, Fred A.; Wurfel, Mark M.; Zanni, Robert; Zou, Fei; Nickerson, Deborah A.; Rieder, Mark J.; Green, Phil; Shendure, Jay; Akey, Joshua M.; Bustamante, Carlos D.; Crosslin, David R.; Eichler, Evan E.; Fox, P. Keolu; Fu, Wenqing; Gordon, Adam; Gravel, Simon; Jarvik, Gail P.; Johnsen, Jill M.; Kan, Mengyuan; Kenny, Eimear E.; Kidd, Jeffrey M.; Lara-Garduno, Fremiet; Leal, Suzanne M.; Liu, Dajiang J.; McGee, Sean; O’Connor, Timothy D.; Paeper, Bryan; Robertson, Peggy D.; Smith, Joshua D.; Staples, Jeffrey C.; Tennessen, Jacob A.; Turner, Emily H.; Wang, Gao; Yi, Qian; Jackson, Rebecca; Peters, Ulrike; Carlson, Christopher S.; Anderson, Garnet; Anton-Culver, Hoda; Assimes, Themistocles L.; Auer, Paul L.; Beresford, Shirley; Bizon, Chris; Black, Henry; Brunner, Robert; Brzyski, Robert; Burwen, Dale; Caan, Bette; Carty, Cara L.; Chlebowski, Rowan; Cummings, Steven; Curb, J. David; Eaton, Charles B.; Ford, Leslie; Franceschini, Nora; Fullerton, Stephanie M.; Gass, Margery; Geller, Nancy; Heiss, Gerardo; Howard, Barbara V.; Hsu, Li; Hutter, Carolyn M.; Ioannidis, John; Jiao, Shuo; Johnson, Karen C.; Kooperberg, Charles; Kuller, Lewis; LaCroix, Andrea; Lakshminarayan, Kamakshi; Lane, Dorothy; Lasser, Norman; LeBlanc, Erin; Li, Kuo-Ping; Limacher, Marian; Lin, Dan-Yu; Logsdon, Benjamin A.; Ludlam, Shari; Manson, JoAnn E.; Margolis, Karen; Martin, Lisa; McGowan, Joan; Monda, Keri L.; Kotchen, Jane Morley; Nathan, Lauren; Ockene, Judith; O’Sullivan, Mary Jo; Phillips, Lawrence S.; Prentice, Ross L.; Robbins, John; Robinson, Jennifer G.; Rossouw, Jacques E.; Sangi-Haghpeykar, Haleh; Sarto, Gloria E.; Shumaker, Sally; Simon, Michael S.; Stefanick, Marcia L.; Stein, Evan; Tang, Hua; Taylor, Kira C.; Thomson, Cynthia A.; Thornton, Timothy A.; Van Horn, Linda; Vitolins, Mara; Wactawski-Wende, Jean; Wallace, Robert; Wassertheil-Smoller, Sylvia; Zeng, Donglin; Applebaum-Bowden, Deborah; Feolo, Michael; Gan, Weiniu; Paltoo, Dina N.; Sholinsky, Phyliss; Sturcke, Anne

2014-01-01

Elevated low-density lipoprotein cholesterol (LDL-C) is a treatable, heritable risk factor for cardiovascular disease. Genome-wide association studies (GWASs) have identified 157 variants associated with lipid levels but are not well suited to assess the impact of rare and low-frequency variants. To determine whether rare or low-frequency coding variants are associated with LDL-C, we exome sequenced 2,005 individuals, including 554 individuals selected for extreme LDL-C (>98th or <2nd percentile). Follow-up analyses included sequencing of 1,302 additional individuals and genotype-based analysis of 52,221 individuals. We observed significant evidence of association between LDL-C and the burden of rare or low-frequency variants in PNPLA5, encoding a phospholipase-domain-containing protein, and both known and previously unidentified variants in PCSK9, LDLR and APOB, three known lipid-related genes. The effect sizes for the burden of rare variants for each associated gene were substantially higher than those observed for individual SNPs identified from GWASs. We replicated the PNPLA5 signal in an independent large-scale sequencing study of 2,084 individuals. In conclusion, this large whole-exome-sequencing study for LDL-C identified a gene not known to be implicated in LDL-C and provides unique insight into the design and analysis of similar experiments. PMID:24507775

Whole-exome sequencing identifies rare and low-frequency coding variants associated with LDL cholesterol.

PubMed

Lange, Leslie A; Hu, Youna; Zhang, He; Xue, Chenyi; Schmidt, Ellen M; Tang, Zheng-Zheng; Bizon, Chris; Lange, Ethan M; Smith, Joshua D; Turner, Emily H; Jun, Goo; Kang, Hyun Min; Peloso, Gina; Auer, Paul; Li, Kuo-Ping; Flannick, Jason; Zhang, Ji; Fuchsberger, Christian; Gaulton, Kyle; Lindgren, Cecilia; Locke, Adam; Manning, Alisa; Sim, Xueling; Rivas, Manuel A; Holmen, Oddgeir L; Gottesman, Omri; Lu, Yingchang; Ruderfer, Douglas; Stahl, Eli A; Duan, Qing; Li, Yun; Durda, Peter; Jiao, Shuo; Isaacs, Aaron; Hofman, Albert; Bis, Joshua C; Correa, Adolfo; Griswold, Michael E; Jakobsdottir, Johanna; Smith, Albert V; Schreiner, Pamela J; Feitosa, Mary F; Zhang, Qunyuan; Huffman, Jennifer E; Crosby, Jacy; Wassel, Christina L; Do, Ron; Franceschini, Nora; Martin, Lisa W; Robinson, Jennifer G; Assimes, Themistocles L; Crosslin, David R; Rosenthal, Elisabeth A; Tsai, Michael; Rieder, Mark J; Farlow, Deborah N; Folsom, Aaron R; Lumley, Thomas; Fox, Ervin R; Carlson, Christopher S; Peters, Ulrike; Jackson, Rebecca D; van Duijn, Cornelia M; Uitterlinden, André G; Levy, Daniel; Rotter, Jerome I; Taylor, Herman A; Gudnason, Vilmundur; Siscovick, David S; Fornage, Myriam; Borecki, Ingrid B; Hayward, Caroline; Rudan, Igor; Chen, Y Eugene; Bottinger, Erwin P; Loos, Ruth J F; Sætrom, Pål; Hveem, Kristian; Boehnke, Michael; Groop, Leif; McCarthy, Mark; Meitinger, Thomas; Ballantyne, Christie M; Gabriel, Stacey B; O'Donnell, Christopher J; Post, Wendy S; North, Kari E; Reiner, Alexander P; Boerwinkle, Eric; Psaty, Bruce M; Altshuler, David; Kathiresan, Sekar; Lin, Dan-Yu; Jarvik, Gail P; Cupples, L Adrienne; Kooperberg, Charles; Wilson, James G; Nickerson, Deborah A; Abecasis, Goncalo R; Rich, Stephen S; Tracy, Russell P; Willer, Cristen J

2014-02-06

Elevated low-density lipoprotein cholesterol (LDL-C) is a treatable, heritable risk factor for cardiovascular disease. Genome-wide association studies (GWASs) have identified 157 variants associated with lipid levels but are not well suited to assess the impact of rare and low-frequency variants. To determine whether rare or low-frequency coding variants are associated with LDL-C, we exome sequenced 2,005 individuals, including 554 individuals selected for extreme LDL-C (>98(th) or <2(nd) percentile). Follow-up analyses included sequencing of 1,302 additional individuals and genotype-based analysis of 52,221 individuals. We observed significant evidence of association between LDL-C and the burden of rare or low-frequency variants in PNPLA5, encoding a phospholipase-domain-containing protein, and both known and previously unidentified variants in PCSK9, LDLR and APOB, three known lipid-related genes. The effect sizes for the burden of rare variants for each associated gene were substantially higher than those observed for individual SNPs identified from GWASs. We replicated the PNPLA5 signal in an independent large-scale sequencing study of 2,084 individuals. In conclusion, this large whole-exome-sequencing study for LDL-C identified a gene not known to be implicated in LDL-C and provides unique insight into the design and analysis of similar experiments. Copyright © 2014 The American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.

The Gene YALI0E20207g from Yarrowia lipolytica Encodes an N-Acetylglucosamine Kinase Implicated in the Regulated Expression of the Genes from the N-Acetylglucosamine Assimilatory Pathway

PubMed Central

Flores, Carmen-Lisset; Gancedo, Carlos

2015-01-01

The non-conventional yeast Yarrowia lipolytica possesses an ORF, YALI0E20207g, which encodes a protein with an amino acid sequence similar to hexokinases from different organisms. We have cloned that gene and determined several enzymatic properties of its encoded protein showing that it is an N-acetylglucosamine (NAGA) kinase. This conclusion was supported by the lack of growth in NAGA of a strain carrying a YALI0E20207g deletion. We named this gene YlNAG5. Expression of YlNAG5 as well as that of the genes encoding the enzymes of the NAGA catabolic pathway—identified by a BLAST search—was induced by this sugar. Deletion of YlNAG5 rendered that expression independent of the presence of NAGA in the medium and reintroduction of the gene restored the inducibility, indicating that YlNag5 participates in the transcriptional regulation of the NAGA assimilatory pathway genes. Expression of YlNAG5 was increased during sporulation and homozygous Ylnag5/Ylnag5 diploid strains sporulated very poorly as compared with a wild type isogenic control strain pointing to a participation of the protein in the process. Overexpression of YlNAG5 allowed growth in glucose of an Ylhxk1glk1 double mutant and produced, in a wild type background, aberrant morphologies in different media. Expression of the gene in a Saccharomyces cerevisiae hxk1 hxk2 glk1 triple mutant restored ability to grow in glucose. PMID:25816199

Arabidopsis ESK1 encodes a novel regulator of freezing tolerance.

PubMed

Xin, Zhanguo; Mandaokar, Ajin; Chen, Junping; Last, Robert L; Browse, John

2007-03-01

The eskimo1 (esk1) mutation of Arabidopsis resulted in a 5.5 degrees C improvement in freezing tolerance in the absence of cold acclimation. Here we show that the increase in freezing tolerance is not associated with any increase in the ability to survive drought or salt stresses, which are similar to freezing in their induction of cellular dehydration. Genome-wide comparisons of gene expression between esk1-1 and wild type indicate that mutations at esk1 result in altered expression of transcription factors and signaling components and of a set of stress-responsive genes. Interestingly, the list of 312 genes regulated by ESK1 shows greater overlap with sets of genes regulated by salt, osmotic and abscisic acid treatments than with genes regulated by cold acclimation or by the transcription factors CBF3 and ICE1, which have been shown to control genetic pathways for freezing tolerance. Map-based cloning identified the esk1 locus as At3g55990. The wild-type ESK1 gene encodes a 57-kDa protein and is a member of a large gene family of DUF231 domain proteins whose members encode a total of 45 proteins of unknown function. Our results indicate that ESK1 is a novel negative regulator of cold acclimation. Mutations in the ESK1 gene provide strong freezing tolerance through genetic regulation that is apparently very different from previously described genetic mechanisms of cold acclimation.

Topological and organizational properties of the products of house-keeping and tissue-specific genes in protein-protein interaction networks.

PubMed

Lin, Wen-Hsien; Liu, Wei-Chung; Hwang, Ming-Jing

2009-03-11

Human cells of various tissue types differ greatly in morphology despite having the same set of genetic information. Some genes are expressed in all cell types to perform house-keeping functions, while some are selectively expressed to perform tissue-specific functions. In this study, we wished to elucidate how proteins encoded by human house-keeping genes and tissue-specific genes are organized in human protein-protein interaction networks. We constructed protein-protein interaction networks for different tissue types using two gene expression datasets and one protein-protein interaction database. We then calculated three network indices of topological importance, the degree, closeness, and betweenness centralities, to measure the network position of proteins encoded by house-keeping and tissue-specific genes, and quantified their local connectivity structure. Compared to a random selection of proteins, house-keeping gene-encoded proteins tended to have a greater number of directly interacting neighbors and occupy network positions in several shortest paths of interaction between protein pairs, whereas tissue-specific gene-encoded proteins did not. In addition, house-keeping gene-encoded proteins tended to connect with other house-keeping gene-encoded proteins in all tissue types, whereas tissue-specific gene-encoded proteins also tended to connect with other tissue-specific gene-encoded proteins, but only in approximately half of the tissue types examined. Our analysis showed that house-keeping gene-encoded proteins tend to occupy important network positions, while those encoded by tissue-specific genes do not. The biological implications of our findings were discussed and we proposed a hypothesis regarding how cells organize their protein tools in protein-protein interaction networks. Our results led us to speculate that house-keeping gene-encoded proteins might form a core in human protein-protein interaction networks, while clusters of tissue-specific gene-encoded proteins are attached to the core at more peripheral positions of the networks.

Draft genome sequence of Actinotignum schaalii DSM 15541T: Genetic insights into the lifestyle, cell fitness and virulence.

PubMed

Yassin, Atteyet F; Langenberg, Stefan; Huntemann, Marcel; Clum, Alicia; Pillay, Manoj; Palaniappan, Krishnaveni; Varghese, Neha; Mikhailova, Natalia; Mukherjee, Supratim; Reddy, T B K; Daum, Chris; Shapiro, Nicole; Ivanova, Natalia; Woyke, Tanja; Kyrpides, Nikos C

2017-01-01

The permanent draft genome sequence of Actinotignum schaalii DSM 15541T is presented. The annotated genome includes 2,130,987 bp, with 1777 protein-coding and 58 rRNA-coding genes. Genome sequence analysis revealed absence of genes encoding for: components of the PTS systems, enzymes of the TCA cycle, glyoxylate shunt and gluconeogensis. Genomic data revealed that A. schaalii is able to oxidize carbohydrates via glycolysis, the nonoxidative pentose phosphate and the Entner-Doudoroff pathways. Besides, the genome harbors genes encoding for enzymes involved in the conversion of pyruvate to lactate, acetate and ethanol, which are found to be the end products of carbohydrate fermentation. The genome contained the gene encoding Type I fatty acid synthase required for de novo FAS biosynthesis. The plsY and plsX genes encoding the acyltransferases necessary for phosphatidic acid biosynthesis were absent from the genome. The genome harbors genes encoding enzymes responsible for isoprene biosynthesis via the mevalonate (MVA) pathway. Genes encoding enzymes that confer resistance to reactive oxygen species (ROS) were identified. In addition, A. schaalii harbors genes that protect the genome against viral infections. These include restriction-modification (RM) systems, type II toxin-antitoxin (TA), CRISPR-Cas and abortive infection system. A. schaalii genome also encodes several virulence factors that contribute to adhesion and internalization of this pathogen such as the tad genes encoding proteins required for pili assembly, the nanI gene encoding exo-alpha-sialidase, genes encoding heat shock proteins and genes encoding type VII secretion system. These features are consistent with anaerobic and pathogenic lifestyles. Finally, resistance to ciprofloxacin occurs by mutation in chromosomal genes that encode the subunits of DNA-gyrase (GyrA) and topisomerase IV (ParC) enzymes, while resistant to metronidazole was due to the frxA gene, which encodes NADPH-flavin oxidoreductase.

DNA methylation of miRNA-encoding genes in non-small cell lung cancer patients.

PubMed

Heller, Gerwin; Altenberger, Corinna; Steiner, Irene; Topakian, Thais; Ziegler, Barbara; Tomasich, Erwin; Lang, György; End-Pfützenreuter, Adelheid; Zehetmayer, Sonja; Döme, Balazs; Arns, Britt-Madeleine; Klepetko, Walter; Zielinski, Christoph C; Zöchbauer-Müller, Sabine

2018-03-23

De-regulated DNA methylation leading to transcriptional inactivation of certain genes occurs frequently in non-small cell lung cancers (NSCLC). Besides protein-encoding genes also microRNA (miRNA)-encoding genes may be targets for methylation in NSCLCs, however, the number of known methylated miRNA genes is still small. Thus, we investigated methylation of miRNA genes in primary tumours (TU) and corresponding non-malignant lung tissue samples (NL) of 50 NSCLC patients using methylated DNA immunoprecipitation followed by custom designed tiling microarray analyses (MeDIP-chip) and 252 differentially methylated probes between TU and NL samples were identified. These probes were annotated which resulted in the identification of 34 miRNA-encoding genes with increased methylation in TU specimens. While some of these miRNA-encoding genes were already known to be methylated in NSCLCs (e.g. miR-9-3, miR-124), methylation of the vast majority of them was unknown so far. We selected six miRNA genes (miR-10b, miR-1179, miR-137, miR-572, miR-3150b and miR-129-2) for gene-specific methylation analyses in TU and corresponding NL samples of 104 NSCLC patients and observed a statistically significant increase of methylation of these miRNA genes in TU samples (p<0.0001, respectively). In silico target prediction of the six miRNAs identified several oncogenic/cell proliferation promoting factors (e.g. CCNE1 as miR-1179 target). To investigate if miR-1179 indeed targets CCNE1, we transfected miR-1179 mimics into CCNE1 expressing NSCLC cells and observed down-regulated CCNE1 mRNA expression in these cells compared to control cells. Similar effects on Cyclin E1 expression were seen in Western blot analyses. In addition, we found a statistically significant growth reduction of NSCLC cells transfected with miR-1179 mimics compared to control cells. In conclusion, we identified many methylated miRNA genes in NSCLC patients and found that miR-1179 is a potential tumour cell growth suppressor in NSCLCs. Overall, our findings emphasize the impact of miRNA gene methylation on the pathogenesis of NSCLCs. This article is protected by copyright. All rights reserved.

Genomic polymorphism, recombination, and linkage disequilibrium in human major histocompatibility complex-encoded antigen-processing genes.

PubMed Central

van Endert, P M; Lopez, M T; Patel, S D; Monaco, J J; McDevitt, H O

1992-01-01

Recently, two subunits of a large cytosolic protease and two putative peptide transporter proteins were found to be encoded by genes within the class II region of the major histocompatibility complex (MHC). These genes have been suggested to be involved in the processing of antigenic proteins for presentation by MHC class I molecules. Because of the high degree of polymorphism in MHC genes, and previous evidence for both functional and polypeptide sequence polymorphism in the proteins encoded by the antigen-processing genes, we tested DNA from 27 consanguineous human cell lines for genomic polymorphism by restriction fragment length polymorphism (RFLP) analysis. These studies demonstrate a strong linkage disequilibrium between TAP1 and LMP2 RFLPs. Moreover, RFLPs, as well as a polymorphic stop codon in the telomeric TAP2 gene, appear to be in linkage disequilibrium with HLA-DR alleles and RFLPs in the HLA-DO gene. A high rate of recombination, however, seems to occur in the center of the complex, between the TAP1 and TAP2 genes. Images PMID:1360671

Marek's disease virus protein kinase gene identified within the short unique region of the viral genome is not essential for viral replication in cell culture and vaccine-induced immunity in chickens.

PubMed

Sakaguchi, M; Urakawa, T; Hirayama, Y; Miki, N; Yamamoto, M; Zhu, G S; Hirai, K

1993-07-01

The open reading frame (ORF) of 1206 bp within the short unique region (Us) of Marek's disease virus type 1 (MDV1) shows significant homology with the herpes simplex virus type 1 US3 gene encoding protein kinase (PK). The lacZ gene of Escherichia coli was inserted within the ORF, designated MDV1-US3, of MDV1 K544 strain DNA by homologous recombination. The plaque-purified recombinant MDV1 stably expressed the beta-galactosidase encoded by the inserted lacZ gene in infected cells and replicated well as the parental K544 strain. Antibodies against both MDV1 antigen and beta-galactosidase were detected in the sera of chickens immunized with recombinant MDV1. Chickens vaccinated with the recombinant MDV1 were protected from challenge with virulent MDV1. The MDV1 US3 gene expressed by a baculovirus vector encoded a 44-kDa protein. Mouse antisera against the 44-kDa protein reacted with two proteins of 44 and 45 kDa in extracts of cells infected with MDV1 but not with MDV types 2 or 3. The PK activity was detected in immune complexes of the anti-44-kDa sera with extracts of cells infected with MDV1 but not with the recombinant MDV1. Thus, PK encoded from the MDV1-US3 is not essential for virus replication in cell culture and vaccine-induced immunity.

A new yeast gene with a myosin-like heptad repeat structure.

PubMed

Kölling, R; Nguyen, T; Chen, E Y; Botstein, D

1993-03-01

We isolated a gene encoding a 218 kDa myosin-like protein from Saccharomyces cerevisiae using a monoclonal antibody directed against human platelet myosin as a probe. The protein sequence encoded by the MLP1 gene (for myosin-like protein) contains extensive stretches of a heptad-repeat pattern suggesting that the protein can form coiled coils typical of myosins. Immunolocalization experiments using affinity-purified antibodies raised against a TrpE-MLP1 fusion protein showed a dot-like structure adjacent to the nucleus in yeast cells bearing the MLP1 gene on a multicopy plasmid. In mouse epithelial cells the yeast anti-MLP1 antibodies stained the nucleus. Mutants bearing disruptions of the MLP1 gene were viable, but more sensitive to ultraviolet light than wild-type strains, suggesting an involvement of MLP1 in DNA repair. The MLP1 gene was mapped to chromosome 11, 25 cM from met1.

A cDNA from a mouse pancreatic beta cell encoding a putative transcription factor of the insulin gene.

PubMed Central

Walker, M D; Park, C W; Rosen, A; Aronheim, A

1990-01-01

Cell specific expression of the insulin gene is achieved through transcriptional mechanisms operating on multiple DNA sequence elements located in the 5' flanking region of the gene. Of particular importance in the rat insulin I gene are two closely similar 9 bp sequences (IEB1 and IEB2): mutation of either of these leads to 5-10 fold reduction in transcriptional activity. We have screened an expression cDNA library derived from mouse pancreatic endocrine beta cells with a radioactive DNA probe containing multiple copies of the IEB1 sequence. A cDNA clone (A1) isolated by this procedure encodes a protein which shows efficient binding to the IEB1 probe, but much weaker binding to either an unrelated DNA probe or to a probe bearing a single base pair insertion within the recognition sequence. DNA sequence analysis indicates a protein belonging to the helix-loop-helix family of DNA-binding proteins. The ability of the protein encoded by clone A1 to recognize a number of wild type and mutant DNA sequences correlates closely with the ability of each sequence element to support transcription in vivo in the context of the insulin 5' flanking DNA. We conclude that the isolated cDNA may encode a transcription factor that participates in control of insulin gene expression. Images PMID:2181401

Cloning and heterologous expression of the antibiotic peptide (ABP) genes from Rhizopus oligosporus NBRC 8631.

PubMed

Yamada, Osamu; Sakamoto, Kazutoshi; Tominaga, Mihoko; Nakayama, Tasuku; Koseki, Takuya; Fujita, Akiko; Akita, Osamu

2005-03-01

We carried out protein sequencing of purified Antibiotic Peptide (ABP), and cloned two genes encoding this peptide as abp1 and abp2, from Rhizopus oligosporus NBRC 8631. Both genes contain an almost identical 231-bp segment, with only 3 nucleotide substitutions, encoding a 77 amino acid peptide. The abp gene product comprises a 28 amino acid signal sequence and a 49 amino acid mature peptide. Northern blot analysis showed that at least one of the abp genes is transcribed in R. oligosporus NBRC 8631. A truncated form of abp1 encoding only the mature peptide was fused with the alpha-factor signal peptide and engineered for expression in Pichia pastoris SMD1168H. Culture broth of the recombinant Pichia displayed ABP activity against Bacillus subtilis NBRC 3335 after induction of heterologous gene expression. This result indicates that mature ABP formed the active structure without the aid of other factors from R. oligosporus, and was secreted.

Staphylococcus aureus nasal carriage in Ukraine: antibacterial resistance and virulence factor encoding genes.

PubMed

Netsvyetayeva, Irina; Fraczek, Mariusz; Piskorska, Katarzyna; Golas, Marlena; Sikora, Magdalena; Mlynarczyk, Andrzej; Swoboda-Kopec, Ewa; Marusza, Wojciech; Palmieri, Beniamino; Iannitti, Tommaso

2014-03-05

The number of studies regarding the incidence of multidrug resistant strains and distribution of genes encoding virulence factors, which have colonized the post-Soviet states, is considerably limited. The aim of the study was (1) to assess the Staphylococcus (S.) aureus nasal carriage rate, including Methicillin Resistant S. aureus (MRSA) strains in adult Ukrainian population, (2) to determine antibiotic resistant pattern and (3) the occurrence of Panton Valentine Leukocidine (PVL)-, Fibronectin-Binding Protein A (FnBPA)- and Exfoliative Toxin (ET)-encoding genes. Nasal samples for S. aureus culture were obtained from 245 adults. The susceptibility pattern for several classes of antibiotics was determined by disk diffusion method according to the European Committee on Antimicrobial Susceptibility Testing (EUCAST) guidelines. The virulence factor encoding genes, mecA, lukS-lukF, eta, etb, etd, fnbA, were detected by Polymerase Chain Reaction (PCR). The S. aureus nasal carriage rate was 40%. The prevalence of nasal MRSA carriage in adults was 3.7%. LukS-lukF genes were detected in over 58% of the strains. ET-encoding genes were detected in over 39% of the strains and the most prevalent was etd. The fnbA gene was detected in over 59% of the strains. All MRSA isolates tested were positive for the mecA gene. LukS-lukF genes and the etd gene were commonly co-present in MRSA, while lukS-lukF genes and the fnbA gene were commonly co-present in Methicillin Sensitive S. aureus (MSSA) isolates. No significant difference was detected between the occurrence of lukS-lukF genes (P > 0.05) and the etd gene (P > 0.05) when comparing MRSA and MSSA. The occurrence of the fnbA gene was significantly more frequent in MSSA strains (P < 0.05). In Ukraine, S. aureus is a common cause of infection. The prevalence of S. aureus nasal carriage in our cohort of patients from Ukraine was 40.4%. We found that 9.1% of the strains were classified as MRSA and all MRSA isolates tested positive for the mecA gene. We also observed a high prevalence of PVL- and ET- encoding genes among S. aureus nasal carriage strains. A systematic surveillance system can help prevent transmission and spread of drug resistant toxin producing S. aureus strains.

Wound induced Beta vulgaris polygalacturonase-inhibiting protein genes encode a longer leucine-rich repeat domain and inhibit fungal polygalacturonases

USDA-ARS?s Scientific Manuscript database

Polygalacturonase-inhibiting proteins (PGIPs) are leucine-rich repeat (LRR) proteins involved in plant defense. Sugar beet (Beta vulgaris L.) PGIP genes, BvPGIP1, BvPGIP2 and BvPGIP3, were isolated from two breeding lines, F1016 and F1010. Full-length cDNA sequences of the three BvPGIP genes encod...

Quercetin protects against aluminium induced oxidative stress and promotes mitochondrial biogenesis via activation of the PGC-1α signaling pathway.

PubMed

Sharma, Deep Raj; Sunkaria, Aditya; Wani, Willayat Yousuf; Sharma, Reeta Kumari; Verma, Deepika; Priyanka, Kumari; Bal, Amanjit; Gill, Kiran Dip

2015-12-01

The present investigation was carried out to elucidate a possible molecular mechanism related to the protective effect of quercetin administration against aluminium-induced oxidative stress on various mitochondrial respiratory complex subunits with special emphasis on the role of PGC-1α and its downstream targets, i.e. NRF-1, NRF-2 and Tfam in mitochondrial biogenesis. Aluminium lactate (10mg/kg b.wt./day) was administered intragastrically to rats, which were pre-treated with quercetin 6h before aluminium (10mg/kg b.wt./day, intragastrically) for 12 weeks. We found a decrease in ROS levels, mitochondrial DNA oxidation and citrate synthase activity in the hippocampus (HC) and corpus striatum (CS) regions of rat brain treated with quercetin. Besides this an increase in the mRNA levels of the mitochondrial encoded subunits - ND1, ND2, ND3, Cyt b, COX1, COX3 and ATPase6 along with increased expression of nuclear encoded subunits COX4, COX5A and COX5B of electron transport chain (ETC). In quercetin treated group an increase in the mitochondrial DNA copy number and mitochondrial content in both the regions of rat brain was observed. The PGC-1α was up regulated in quercetin treated rats along with NRF-1, NRF-2 and Tfam, which act downstream from PGC-1α. Electron microscopy results revealed a significant decrease in the mitochondrial cross-section area, mitochondrial perimeter length and increase in mitochondrial number in case of quercetin treated rats as compared to aluminium treated ones. Therefore it seems quercetin increases mitochondrial biogenesis and makes it an almost ideal flavanoid to control or limit the damage that has been associated with the defective mitochondrial function seen in many neurodegenerative diseases. Copyright © 2015 Elsevier Inc. All rights reserved.

Genome-wide comparative analysis of NBS-encoding genes in four Gossypium species

USDA-ARS?s Scientific Manuscript database

Nucleotide binding site (NBS) genes encode a large family of disease resistance (R) proteins in plants. The availability of genomic data of the two diploid cotton species, Gossypium arboreum and Gossypium raimondii, and the two allotetraploid cotton species, Gossypium hirsutum (TM-1) and Gossypium ...

Identification of three redundant segments responsible for herpes simplex virus 1 ICP0 to fuse with ND10 nuclear bodies.

PubMed

Zheng, Yi; Gu, Haidong

2015-04-01

Infected cell protein 0 (ICP0) of herpes simplex virus 1 (HSV-1) is a key regulator in both lytic and latent infections. In lytic infection, an important early event is the colocalization of ICP0 to nuclear domain 10 (ND10), the discrete nuclear bodies that impose restrictions on viral expression. ICP0 contains an E3 ubiquitin ligase that degrades promyelocytic leukemia protein (PML) and Sp100, two major components of ND10, and disperses ND10 to alleviate repression. We previously reported that the association between ICP0 and ND10 is a dynamic process that includes three steps: adhesion, fusion, and retention. ICP0 residues 245 to 474, defined as ND10 entry signal (ND10-ES), is a region required for the fusion step. Without ND10-ES, ICP0 adheres at the ND10 surface but fails to enter. In the present study, we focus on characterizing ND10-ES. Here we report the following. (i) Fusion of ICP0 with ND10 relies on specific sequences located within ND10-ES. Replacement of ND10-ES by the corresponding region from ORF61 of varicella-zoster virus did not rescue ND10 fusion. (ii) Three tandem ND10 fusion segments (ND10-FS1, ND10-FS2, and ND10-FS3), encompassing 200 amino acids within ND10-ES, redundantly facilitate fusion. Each of the three segments is sufficient to independently drive the fusion process, but none of the segments by themselves are necessary for ND10 fusion. Only when all three segments are deleted is fusion blocked. (iii) The SUMO interaction motif located within ND10-FS2 is not required for ND10 fusion but is required for the complete degradation of PML, suggesting that PML degradation and ND10 fusion are regulated by different molecular mechanisms. ND10 nuclear bodies are part of the cell-intrinsic antiviral defenses that restrict viral gene expression upon virus infection. As a countermeasure, infected cell protein 0 (ICP0) of herpes simplex virus 1 (HSV-1) localizes to ND10s, degrades the ND10 organizer, and disperses ND10 components in order to alleviate repression. We studied the ICP0-ND10 association to delineate elements important for this dynamic interaction and to understand its role in viral replication and host defense. In this work, we show that ICP0 contains three redundant segments to ensure an effective mergence of ICP0 with ND10 nuclear bodies. This is the first study to systematically investigate ICP0 elements that are important for ICP0-ND10 fusion. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Mutational screening in patients with profound sensorineural hearing loss and neurodevelopmental delay: Description of a novel m.3861A > C mitochondrial mutation in the MT-ND1 gene.

PubMed

Ammar, Marwa; Tabebi, Mouna; Sfaihi, Lamia; Alila-Fersi, Olfa; Maalej, Marwa; Felhi, Rahma; Chabchoub, Imen; Keskes, Leila; Hachicha, Mongia; Fakhfakh, Faiza; Mkaouar-Rebai, Emna

2016-06-10

Mitochondrial diseases caused by mitochondrial dysfunction are a clinically and genetically, heterogeneous group of disorders involving multiple organs, particularly tissues with high-energy demand. Hearing loss is a recognized symptom of a number of mitochondrial diseases and can result from neuronal or cochlear dysfunction. The tissue affected in this pathology is most probably the cochlear hair cells, which are essential for hearing function since they are responsible for maintaining the ionic gradients necessary for sound signal transduction. Several mitochondrial DNA mutations have been associated with hearing loss and since mitochondria are crucial for the cellular energy supply in many tissues, most of these mtDNA mutations affect several tissues and will cause syndromic hearing loss. In the present study, we described 2 patients with sensorineural hearing loss and neurodevelopmental delay in whom we tested mitochondrial genes described to be associated with syndromic hearing loss. One of these patients showed a novel heteroplasmic mitochondrial mutation m.3861A > C (W185C) which lead to a loss of stability of the ND1 protein since it created a new hydrogen bund between the unique created cystein C185 and the A182 residue. In the second patient, we detected two novel heteroplasmic variations m.12350C > A (T5N) and m.14351T > C (E108G) respectively in the MT-ND5 and the MT-ND6 genes. The TopPred II prediction for the E108G variation revealed a decrease of the hydrophobicity in the mutated MT-ND6. Copyright © 2016 Elsevier Inc. All rights reserved.

Large-scale, multi-genome analysis of alternate open reading frames in bacteria and archaea.

PubMed

Veloso, Felipe; Riadi, Gonzalo; Aliaga, Daniela; Lieph, Ryan; Holmes, David S

2005-01-01

Analysis of over 300,000 annotated genes in 105 bacterial and archaeal genomes reveals an unexpectedly high frequency of large (>300 nucleotides) alternate open reading frames (ORFs). Especially notable is the very high frequency of alternate ORFs in frames +3 and -1 (where the annotated gene is defined as frame +1). The occurrence of alternate ORFs is correlated with genomic G+C content and is strongly influenced by synonymous codon usage bias. The frequency of alternate ORFs in frame -1 is also influenced by the occurrence of codons encoding leucine and serine in frame +1. Although some alternate ORFs have been shown to encode proteins, many others are probably not expressed because they lack appropriate signals for transcription and translation. These latter can be mis-annotated by automatic gene finding programs leading to errors in public databases. Especially prone to mis-annotation is frame -1, because it exhibits a potential codon usage and theoretical capacity to encode proteins with an amino acid composition most similar to real genes. Some alternate ORFs are conserved across bacterial or archaeal species, and can give rise to misannotated "conserved hypothetical" genes, while others are unique to a genome and are misidentified as "hypothetical orphan" genes, contributing significantly to the orphan gene paradox.

β-Lactamase Genes of the Penicillin-Susceptible Bacillus anthracis Sterne Strain

PubMed Central

Chen, Yahua; Succi, Janice; Tenover, Fred C.; Koehler, Theresa M.

2003-01-01

Susceptibility to penicillin and other β-lactam-containing compounds is a common trait of Bacillus anthracis. β-lactam agents, particularly penicillin, have been used worldwide to treat anthrax in humans. Nonetheless, surveys of clinical and soil-derived strains reveal penicillin G resistance in 2 to 16% of isolates tested. Bacterial resistance to β-lactam agents is often mediated by production of one or more types of β-lactamases that hydrolyze the β-lactam ring, inactivating the antimicrobial agent. Here, we report the presence of two β-lactamase (bla) genes in the penicillin-susceptible Sterne strain of B. anthracis. We identified bla1 by functional cloning with Escherichia coli. bla1 is a 927-nucleotide (nt) gene predicted to encode a protein with 93.8% identity to the type I β-lactamase gene of Bacillus cereus. A second gene, bla2, was identified by searching the unfinished B. anthracis chromosome sequence database of The Institute for Genome Research for open reading frames (ORFs) predicted to encode β-lactamases. We found a partial ORF predicted to encode a protein with significant similarity to the carboxy-terminal end of the type II β-lactamase of B. cereus. DNA adjacent to the 5′ end of the partial ORF was cloned using inverse PCR. bla2 is a 768-nt gene predicted to encode a protein with 92% identity to the B. cereus type II enzyme. The bla1 and bla2 genes confer ampicillin resistance to E. coli and Bacillus subtilis when cloned individually in these species. The MICs of various antimicrobial agents for the E. coli clones indicate that the two β-lactamase genes confer different susceptibility profiles to E. coli; bla1 is a penicillinase, while bla2 appears to be a cephalosporinase. The β-galactosidase activities of B. cereus group species harboring bla promoter-lacZ transcriptional fusions indicate that bla1 is poorly transcribed in B. anthracis, B. cereus, and B. thuringiensis. The bla2 gene is strongly expressed in B. cereus and B. thuringiensis and weakly expressed in B. anthracis. Taken together, these data indicate that the bla1 and bla2 genes of the B. anthracis Sterne strain encode functional β-lactamases of different types, but gene expression is usually not sufficient to confer resistance to β-lactam agents. PMID:12533457

Cloning and sequencing the genes encoding goldfish and carp ependymin.

PubMed

Adams, D S; Shashoua, V E

1994-04-20

Ependymins (EPNs) are brain glycoproteins thought to function in optic nerve regeneration and long-term memory consolidation. To date, epn genes have been characterized in two orders of teleost fish. In this study, polymerase chain reactions (PCR) were used to amplify the complete 1.6-kb epn genes, gf-I and cc-I, from genomic DNA of Cypriniformes, goldfish and carp, respectively. Amplified bands were cloned and sequenced. Each gene consists of six exons and five introns. The exon portion of gf-I encodes a predicted 215-amino-acid (aa) protein previously characterized as GF-I, while cc-I encodes a predicted 215-aa protein 95% homologous to GF-I.

The complete mitochondrial genome sequence of Eimeria innocua (Eimeriidae, Coccidia, Apicomplexa).

PubMed

Hafeez, Mian Abdul; Vrba, Vladimir; Barta, John Robert

2016-07-01

The complete mitochondrial genome of Eimeria innocua KR strain (Eimeriidae, Coccidia, Apicomplexa) was sequenced. This coccidium infects turkeys (Meleagris gallopavo), Bobwhite quails (Colinus virginianus), and Grey partridges (Perdix perdix). Genome organization and gene contents were comparable with other Eimeria spp. infecting galliform birds. The circular-mapping mt genome of E. innocua is 6247 bp in length with three protein-coding genes (cox1, cox3, and cytb), 19 gene fragments encoding large subunit (LSU) rRNA and 14 gene fragments encoding small subunit (SSU) rRNA. Like other Apicomplexa, no tRNA was encoded. The mitochondrial genome of E. innocua confirms its close phylogenetic affinities to Eimeria dispersa.

Identification of the Pr1 Gene Product Completes the Anthocyanin Biosynthesis Pathway of Maize

PubMed Central

Sharma, Mandeep; Cortes-Cruz, Moises; Ahern, Kevin R.; McMullen, Michael; Brutnell, Thomas P.; Chopra, Surinder

2011-01-01

In maize, mutations in the pr1 locus lead to the accumulation of pelargonidin (red) rather than cyanidin (purple) pigments in aleurone cells where the anthocyanin biosynthetic pathway is active. We characterized pr1 mutation and isolated a putative F3′H encoding gene (Zmf3′h1) and showed by segregation analysis that the red kernel phenotype is linked to this gene. Genetic mapping using SNP markers confirms its position on chromosome 5L. Furthermore, genetic complementation experiments using a CaMV 35S::ZmF3′H1 promoter–gene construct established that the encoded protein product was sufficient to perform a 3′-hydroxylation reaction. The Zmf3′h1-specific transcripts were detected in floral and vegetative tissues of Pr1 plants and were absent in pr1. Four pr1 alleles were characterized: two carry a 24 TA dinucleotide repeat insertion in the 5′-upstream promoter region, a third has a 17-bp deletion near the TATA box, and a fourth contains a Ds insertion in exon1. Genetic and transcription assays demonstrated that the pr1 gene is under the regulatory control of anthocyanin transcription factors red1 and colorless1. The cloning and characterization of pr1 completes the molecular identification of all genes encoding structural enzymes of the anthocyanin pathway of maize. PMID:21385724

The Escherichia coli Serogroup O1 and O2 Lipopolysaccharides Are Encoded by Multiple O-antigen Gene Clusters.

PubMed

Delannoy, Sabine; Beutin, Lothar; Mariani-Kurkdjian, Patricia; Fleiss, Aubin; Bonacorsi, Stéphane; Fach, Patrick

2017-01-01

Escherichia coli strains belonging to serogroups O1 and O2 are frequently associated with human infections, especially extra-intestinal infections such as bloodstream infections or urinary tract infections. These strains can be associated with a large array of flagellar antigens. Because of their frequency and clinical importance, a reliable detection of E. coli O1 and O2 strains and also the frequently associated K1 capsule is important for diagnosis and source attribution of E. coli infections in humans and animals. By sequencing the O-antigen clusters of various O1 and O2 strains we showed that the serogroups O1 and O2 are encoded by different sets of O-antigen encoding genes and identified potentially new O-groups. We developed qPCR-assays to detect the various O1 and O2 variants and the K1-encoding gene. These qPCR assays proved to be 100% sensitive and 100% specific and could be valuable tools for the investigations of zoonotic and food-borne infection of humans with O1 and O2 extra-intestinal (ExPEC) or Shiga toxin-producing E. coli (STEC) strains.

The Escherichia coli Serogroup O1 and O2 Lipopolysaccharides Are Encoded by Multiple O-antigen Gene Clusters

PubMed Central

Delannoy, Sabine; Beutin, Lothar; Mariani-Kurkdjian, Patricia; Fleiss, Aubin; Bonacorsi, Stéphane; Fach, Patrick

2017-01-01

Escherichia coli strains belonging to serogroups O1 and O2 are frequently associated with human infections, especially extra-intestinal infections such as bloodstream infections or urinary tract infections. These strains can be associated with a large array of flagellar antigens. Because of their frequency and clinical importance, a reliable detection of E. coli O1 and O2 strains and also the frequently associated K1 capsule is important for diagnosis and source attribution of E. coli infections in humans and animals. By sequencing the O-antigen clusters of various O1 and O2 strains we showed that the serogroups O1 and O2 are encoded by different sets of O-antigen encoding genes and identified potentially new O-groups. We developed qPCR-assays to detect the various O1 and O2 variants and the K1-encoding gene. These qPCR assays proved to be 100% sensitive and 100% specific and could be valuable tools for the investigations of zoonotic and food-borne infection of humans with O1 and O2 extra-intestinal (ExPEC) or Shiga toxin-producing E. coli (STEC) strains. PMID:28224115

Virulence and extended-spectrum β-lactamase encoding genes in Escherichia coli recovered from chicken meat intended for hospitalized human consumption.

PubMed

Younis, Gamal A; Elkenany, Rasha M; Fouda, Mohamed A; Mostafa, Noura F

2017-10-01

This study describes the prevalence of Escherichia coli in frozen chicken meat intended for human consumption with emphasis on their virulence determinants through detection of the virulence genes and recognition of the extended-spectrum β-lactamase (ESBL) encoding genes ( bla OXA and bla TEM genes). A total of 120 frozen chicken meat samples were investigated for isolation of E. coli . All isolates were subjected to biochemical and serological tests. Eight serotypes isolated from samples were analyzed for the presence of various virulence genes ( stx1, stx2 , and eae A genes) using multiplex polymerase chain reaction (PCR) technique. Moreover, the strains were evaluated for the ESBL encoding genes ( bla TEM and bla OXA ). Overall, 11.66% (14/120) chicken meat samples carried E. coli according to cultural and biochemical properties. The most predominant serotypes were O78 and O128: H2 (21.5%, each), followed by O121: H7 and O44: H18. Molecular method detected that 2 strains (25%) harbored stx1 , 3 strains (37.5%) stx2 , and 3 strains (37.5%) both stx1 and stx2 , while 1 (12.5%) strain carried eae A gene. Particularly, only O26 serotype had all tested virulence genes ( stx1, stx2, and eae A ). The results revealed that all examined 8 serotypes were Shiga toxin-producing E. coli (STEC). The ESBL encoding genes ( bla TEM and bla OXA ) of STEC were detected in 4 (50%) isolates by multiplex PCR. The overall incidence of bla TEM and bla OXA genes was 3 (37.5%) and 2 (25%) isolates. The present study indicates the prevalence of virulent and ESBL-producing E. coli in frozen chicken meat intended for hospitalized human consumption due to poor hygienic measures and irregular use of antibiotics. Therefore, the basic instructions regarding good hygienic measures should be adapted to limit public health hazard.

ND3, ND1 and 39 kDa subunits are more exposed in the de-active form of bovine mitochondrial complex I

PubMed Central

Babot, Marion; Labarbuta, Paola; Birch, Amanda; Kee, Sara; Fuszard, Matthew; Botting, Catherine H.; Wittig, Ilka; Heide, Heinrich; Galkin, Alexander

2014-01-01

An intriguing feature of mitochondrial complex I from several species is the so-called A/D transition, whereby the idle enzyme spontaneously converts from the active (A) form to the de-active (D) form. The A/D transition plays an important role in tissue response to the lack of oxygen and hypoxic deactivation of the enzyme is one of the key regulatory events that occur in mitochondria during ischaemia. We demonstrate for the first time that the A/D conformational change of complex I does not affect the macromolecular organisation of supercomplexes in vitro as revealed by two types of native electrophoresis. Cysteine 39 of the mitochondrially-encoded ND3 subunit is known to become exposed upon de-activation. Here we show that even if complex I is a constituent of the I + III2 + IV (S1) supercomplex, cysteine 39 is accessible for chemical modification in only the D-form. Using lysine-specific fluorescent labelling and a DIGE-like approach we further identified two new subunits involved in structural rearrangements during the A/D transition: ND1 (MT-ND1) and 39 kDa (NDUFA9). These results clearly show that structural rearrangements during de-activation of complex I include several subunits located at the junction between hydrophilic and hydrophobic domains, in the region of the quinone binding site. De-activation of mitochondrial complex I results in concerted structural rearrangement of membrane subunits which leads to the disruption of the sealed quinone chamber required for catalytic turnover. PMID:24560811

Mitogenome Phylogenetics: The Impact of Using Single Regions and Partitioning Schemes on Topology, Substitution Rate and Divergence Time Estimation

PubMed Central

Duchêne, Sebastián; Archer, Frederick I.; Vilstrup, Julia; Caballero, Susana; Morin, Phillip A.

2011-01-01

The availability of mitochondrial genome sequences is growing as a result of recent technological advances in molecular biology. In phylogenetic analyses, the complete mitogenome is increasingly becoming the marker of choice, usually providing better phylogenetic resolution and precision relative to traditional markers such as cytochrome b (CYTB) and the control region (CR). In some cases, the differences in phylogenetic estimates between mitogenomic and single-gene markers have yielded incongruent conclusions. By comparing phylogenetic estimates made from different genes, we identified the most informative mitochondrial regions and evaluated the minimum amount of data necessary to reproduce the same results as the mitogenome. We compared results among individual genes and the mitogenome for recently published complete mitogenome datasets of selected delphinids (Delphinidae) and killer whales (genus Orcinus). Using Bayesian phylogenetic methods, we investigated differences in estimation of topologies, divergence dates, and clock-like behavior among genes for both datasets. Although the most informative regions were not the same for each taxonomic group (COX1, CYTB, ND3 and ATP6 for Orcinus, and ND1, COX1 and ND4 for Delphinidae), in both cases they were equivalent to less than a quarter of the complete mitogenome. This suggests that gene information content can vary among groups, but can be adequately represented by a portion of the complete sequence. Although our results indicate that complete mitogenomes provide the highest phylogenetic resolution and most precise date estimates, a minimum amount of data can be selected using our approach when the complete sequence is unavailable. Studies based on single genes can benefit from the addition of a few more mitochondrial markers, producing topologies and date estimates similar to those obtained using the entire mitogenome. PMID:22073275

Parametrial adipose tissue and metabolic dysfunctions induced by fructose-rich diet in normal and neonatal-androgenized adult female rats.

PubMed

Alzamendi, Ana; Castrogiovanni, Daniel; Ortega, Hugo H; Gaillard, Rolf C; Giovambattista, Andres; Spinedi, Eduardo

2010-03-01

Hyperandrogenemia predisposes an organism toward developing impaired insulin sensitivity. The aim of our study was to evaluate endocrine and metabolic effects during early allostasis induced by a fructose-rich diet (FRD) in normal (control; CT) and neonatal-androgenized (testosterone propionate; TP) female adult rats. CT and TP rats were fed either a normal diet (ND) or an FRD for 3 weeks immediately before the day of study, which was at age 100 days. Energy intake, body weight (BW), parametrial (PM) fat characteristics, and endocrine/metabolic biomarkers were then evaluated. Daily energy intake was similar in CT and TP rats regardless of the differences in diet. When compared with CT-ND rats, the TP-ND rats were heavier, had larger PM fat, and were characterized by basal hypoadiponectinemia and enhanced plasma levels of non-esterified fatty acid (NEFA), plasminogen activator inhibitor-1 (PAI-1), and leptin. FRD-fed CT rats, when compared with CT-ND rats, had high plasma levels of NEFA, triglyceride (TG), PAI-1, leptin, and adiponectin. The TP-FRD rats, when compared with TP-ND rats, displayed enhanced leptinemia and triglyceridemia, and were hyperinsulinemic, with glucose intolerance. The PM fat taken from TP rats displayed increase in the size of adipocytes, decrease in adiponectin (protein/gene), and a greater abundance of the leptin gene. PM adipocyte response to insulin was impaired in CT-FRD, TP-ND, and TP-FRD rats. A very short duration of isocaloric FRD intake in TP rats induced severe metabolic dysfunction at the reproductive age. Our study supports the hypothesis that the early-androgenized female rat phenotype is highly susceptible to developing endocrine/metabolic dysfunction. In turn, these abnormalities enhance the risk of metabolic syndrome, obesity, type 2 diabetes, and cardiovascular disease.

Expression of the ribulose-1,5-bisphosphate carboxylase large subunit gene and three small subunit genes in two cell types of maize leaves

PubMed Central

Sheen, Jenq-Yunn; Bogorad, Lawrence

1986-01-01

Transcripts of three distinct ribulose-1,5-bisphosphate carboxylase (RuBPC) small subunit (SS) genes account for ∼90% of the mRNA for this protein in maize leaves. Transcripts of two of them constitute >80% of the SS mRNA in 24-h greening maize leaves. The third gene contribute ∼10%. Transcripts of all three nuclear-encoded SS genes are detectable in bundle sheath (BSC) and mesophyll cells (MC) of etiolated maize leaves. The level of mRNA for each gene is different in etioplasts of MC but all drop during photoregulated development of chloroplasts in MC and follow a pattern of transitory rise and fall in BSC. The amounts of LS and SS proteins continue to increase steadily well after the mRNA levels reach their peaks in BSC. The molar ratio of mRNA for chloroplast-encoded RuBPC large subunit (LS) to the nuclear genome encoded SS is about 10:1 although LS and SS proteins are present in about equimolar amounts. ImagesFig. 1.Fig. 2.Fig. 3.Fig. 4.Fig. 5.Fig. 6. PMID:16453739

Survey of Genes Encoding Staphylococcal Enterotoxins, Toxic Shock Syndrome Toxin 1, and Exfoliative Toxins in Members of the Staphylococcus sciuri Group

PubMed Central

Dakić, Ivana; Vuković, Dragana; Stepanović, Srdjan; Hauschild, Tomasz; Ježek, Petr; Petráš, Petr; Morrison, Donald

2005-01-01

Genes encoding staphylococcal enterotoxins (sea to see, seg, and seh), toxic shock syndrome toxin 1 (tst), and exfoliative toxins (eta and etb) were not detected in a large panel of 48 Staphylococcus sciuri group isolates tested. This strongly suggests that production of the staphylococcal exotoxins by these bacteria is highly unlikely. PMID:16145164

Expression of the Immediate-Early Gene-Encoded Protein Egr-1 ("zif268") during in Vitro Classical Conditioning

ERIC Educational Resources Information Center

Mokin, Maxim; Keifer, Joyce

2005-01-01

Expression of the immediate-early genes (IEGs) has been shown to be induced by activity-dependent synaptic plasticity or behavioral training and is thought to play an important role in long-term memory. In the present study, we examined the induction and expression of the IEG-encoded protein Egr-1 during an in vitro neural correlate of eyeblink…

Unilateral incompatibility gene ui1.1 encodes an S-locus F-box protein expressed in pollen of Solanum species.

PubMed

Li, Wentao; Chetelat, Roger T

2015-04-07

Unilateral interspecific incompatibility (UI) is a postpollination, prezygotic reproductive barrier that prevents hybridization between related species when the female parent is self-incompatible (SI) and the male parent is self-compatible (SC). In tomato and related Solanum species, two genes, ui1.1 and ui6.1, are required for pollen compatibility on pistils of SI species or hybrids. We previously showed that ui6.1 encodes a Cullin1 (CUL1) protein. Here we report that ui1.1 encodes an S-locus F-box (SLF) protein. The ui1.1 gene was mapped to a 0.43-cM, 43.2-Mbp interval at the S-locus on chromosome 1, but positional cloning was hampered by low recombination frequency. We hypothesized that ui1.1 encodes an SLF protein(s) that interacts with CUL1 and Skp1 proteins to form an SCF-type (Skp1, Cullin1, F-box) ubiquitin E3 ligase complex. We identified 23 SLF genes in the S. pennellii genome, of which 19 were also represented in cultivated tomato (S. lycopersicum). Data from recombination events, expression analysis, and sequence annotation highlighted 11 S. pennellii genes as candidates. Genetic transformations demonstrated that one of these, SpSLF-23, is sufficient for ui1.1 function. A survey of cultivated and wild tomato species identified SLF-23 orthologs in each of the SI species, but not in the SC species S. lycopersicum, S. cheesmaniae, and S. galapagense, pollen of which lacks ui1.1 function. These results demonstrate that pollen compatibility in UI is mediated by protein degradation through the ubiquitin-proteasome pathway, a mechanism related to that which controls pollen recognition in SI.

Plasmids encoding PKI(1-31), a specific inhibitor of cAMP-stimulated gene expression, inhibit the basal transcriptional activity of some but not all cAMP-regulated DNA response elements in JEG-3 cells.

PubMed

Grove, J R; Deutsch, P J; Price, D J; Habener, J F; Avruch, J

1989-11-25

Plasmids that encode a bioactive amino-terminal fragment of the heat-stable inhibitor of the cAMP-dependent protein kinase, PKI(1-31), were employed to characterize the role of this protein kinase in the control of transcriptional activity mediated by three DNA regulatory elements in the JEG-3 human placental cell line. The 5'-flanking sequence of the human collagenase gene contains the heptameric sequence, 5'-TGAGTCA-3', previously identified as a "phorbol ester" response element. Reporter genes containing either the intact 1.2-kilobase 5'-flanking sequence from the human collagenase gene or just the 7-base pair (bp) response element, when coupled to an enhancerless promoter, each exhibit both cAMP and phorbol ester-stimulated expression in JEG-3 cells. Cotransfection of either construct with plasmids encoding PKI(1-31) inhibits cAMP-stimulated but not basal- or phorbol ester-stimulated expression. Pretreatment of cells with phorbol ester for 1 or 2 days abrogates completely the response to rechallenge with phorbol ester but does not alter the basal expression of either construct; cAMP-stimulated expression, while modestly inhibited, remains vigorous. The 5'-flanking sequence of the human chorionic gonadotropin-alpha subunit (HCG alpha) gene has two copies of the sequence, 5'-TGACGTCA-3', contained in directly adjacent identical 18-bp segments, previously identified as a cAMP-response element. Reporter genes containing either the intact 1.5 kilobase of 5'-flanking sequence from the HCG alpha gene, or just the 36-bp tandem repeat cAMP response element, when coupled to an enhancerless promoter, both exhibit a vigorous cAMP stimulation of expression but no response to phorbol ester in JEG-3 cells. Cotransfection with plasmids encoding PKI(1-31) inhibits both basal and cAMP-stimulated expression in a parallel fashion. The 5'-flanking sequence of the human enkephalin gene mediates cAMP-stimulated expression of reporter genes in both JEG-3 and CV-1 cells. Plasmids encoding PKI(1-31) inhibit the expression that is stimulated by the addition of cAMP analogs in both cell lines; basal expression, however, is inhibited by PKI(1-31) only in the JEG-3 cell line and not in the CV-1 cells. These observations indicate that, in JEG-3 cells, PKI(1-31) is a specific inhibitor of kinase A-mediated gene transcription, but it does not modify kinase C-directed transcription.(ABSTRACT TRUNCATED AT 400 WORDS)

Discovery of novel cold-induced CISP genes encoding small RNA-binding proteins related to cold adaptation in barley.

PubMed

Ying, Mengchao; Kidou, Shin-Ichiro

2017-07-01

To adapt to cold conditions, barley plants rely on specific mechanisms, which have not been fully understood. In this study, we characterized a novel barley cold-induced gene identified using a PCR-based high coverage gene expression profiling method. The identified gene encodes a small protein that we named CISP1 (Cold-induced Small Protein 1). Homology searches of sequence databases revealed that CISP1 homologs (CISP2 and CISP3) exist in barley genome. Further database analyses showed that the CISP1 homologs were widely distributed in cold-tolerant plants such as wheat and rye. Quantitative reverse transcription PCR analyses indicated that the expression of barley CISP genes was markedly increased in roots exposed to cold conditions. In situ hybridization analyses showed that the CISP1 transcripts were localized in the root tip and lateral root primordium. We also demonstrated that the CISP1 protein bound to RNA. Taken together, these findings indicate that CISP1 and its homologs encoding small RNA-binding proteins may serve as RNA chaperones playing a vital role in the cold adaptation of barley root. This is the first report describing the likely close relationship between root-specific genes and the cold adaptation process, as well as the potential function of the identified genes. Copyright © 2017 Elsevier B.V. All rights reserved.

Long-term outcomes of gene therapy for the treatment of Leber's hereditary optic neuropathy.

PubMed

Yang, Shuo; Ma, Si-Qi; Wan, Xing; He, Heng; Pei, Han; Zhao, Min-Jian; Chen, Chen; Wang, Dao-Wen; Dong, Xiao-Yan; Yuan, Jia-Jia; Li, Bin

2016-08-01

Leber's hereditary optic neuropathy (LHON) is a disease that leads to blindness. Gene therapy has been investigated with some success, and could lead to important advancements in treating LHON. This was a prospective, open-label trial involving 9 LHON patients at Tongji Hospital, Wuhan, China, from August 2011 to December 2015. The purpose of this study was to evaluate the long-term outcomes of gene therapy for LHON. Nine LHON patients voluntarily received an intravitreal injection of rAAV2-ND4. Systemic examinations and visual function tests were performed during the 36-month follow-up period to determine the safety and efficacy of this gene therapy. Based on successful experiments in an animal model of LHON, 1 subject also received an rAAV2-ND4 injection in the second eye 12months after gene therapy was administered in the first eye. Recovery of visual acuity was defined as the primary outcome of this study. Changes in the visual field, visual evoked potential (VEP), optical coherence tomography findings, liver and kidney function, and antibodies against AAV2 were defined as secondary endpoints. Eight patients (Patients 2-9) received unilateral gene therapy and visual function improvement was observed in both treated eyes (Patients 4, 6, 7, and 8) and untreated eyes (Patients 2, 3, 4, 6 and 8). Visual regression fluctuations, defined as changes in visual acuity greater than or equal to 0.3 logMAR, were observed in Patients 2 and 9. Age at disease onset, disease duration, and the amount of remaining optic nerve fibers did not have a significant effect on the visual function improvement. The visual field and pattern reversal VEP also improved. The patient (Patient 1) who received gene therapy in both eyes had improved visual acuity in the injected eye after the first treatment. Unfortunately, visual acuity in this eye decreased 3months after he received gene therapy in the second eye. Animal experiments suggested that ND4 expression remains stable in the contralateral eye after intravitreal injections. No serious safety problem was observed in the 3-year follow-up of the 9 participants enrolled in this virus-based gene therapy. Meanwhile, our results support the use of intravitreal rAAV2-ND4 as an aggressive maneuver in our clinical trial. Further study in additional patients and in these 9 subjects is needed to better understand the effects of rAAV2-ND4 gene therapy on LHON and to increase the applications of this technique. Copyright © 2016 The Ohio State University Wexner Medical Center. Published by Elsevier B.V. All rights reserved.

Three new members of the RNP protein family in Xenopus.

PubMed Central

Good, P J; Rebbert, M L; Dawid, I B

1993-01-01

Many RNP proteins contain one or more copies of the RNA recognition motif (RRM) and are thought to be involved in cellular RNA metabolism. We have previously characterized in Xenopus a nervous system specific gene, nrp1, that is more similar to the hnRNP A/B proteins than to other known proteins (K. Richter, P. J. Good, and I. B. Dawid (1990), New Biol. 2, 556-565). PCR amplification with degenerate primers was used to identify additional cDNAs encoding two RRMs in Xenopus. Three previously uncharacterized genes were identified. Two genes encode hnRNP A/B proteins with two RRMs and a glycine-rich domain. One of these is the Xenopus homolog of the human A2/B1 gene; the other, named hnRNP A3, is similar to both the A1 and A2 hnRNP genes. The Xenopus hnRNP A1, A2 and A3 genes are expressed throughout development and in all adult tissues. Multiple protein isoforms for the hnRNP A2 gene are predicted that differ by the insertion of short peptide sequences in the glycine-rich domain. The third newly isolated gene, named xrp1, encodes a protein that is related by sequence to the nrp1 protein but is expressed ubiquitously. Despite the similarity to nuclear RNP proteins, both the nrp1 and xrp1 proteins are localized to the cytoplasm in the Xenopus oocyte. The xrp1 gene may have a function in all cells that is similar to that executed by nrp1 specifically within the nervous system. Images PMID:8451200

Site-Directed Mutagenesis of HgcA and HgcB Reveals Amino Acid Residues Important for Mercury Methylation

DOE PAGES

Smith, Steven D.; Bridou, Romain; Johs, Alexander; ...

2015-02-27

Methylmercury is a potent neurotoxin that is produced by anaerobic microorganisms from inorganic mercury by a recently discovered pathway. A two-gene cluster, consisting of hgcA and hgcB, encodes two of the proteins essential for this activity. hgcA encodes a corrinoid protein with a strictly conserved cysteine proposed to be the ligand for cobalt in the corrinoid cofactor, whereas hgcB encodes a ferredoxin-like protein thought to be an electron donor to HgcA. Deletion of either gene eliminates mercury methylation by the methylator Desulfovibrio desulfuricans ND132. Here, site-directed mutants of HgcA and HgcB were constructed to determine amino acid residues essential formore » mercury methylation. Mutations of the strictly conserved residue Cys93 in HgcA, the proposed ligand for the corrinoid cobalt, to Ala or Thr completely abolished the methylation capacity, but a His substitution produced measurable methylmercury. Mutations of conserved amino acids near Cys93 had various impacts on the methylation capacity but showed that the structure of the putative “cap helix” region harboring Cys93 is crucial for methylation function. In the ferredoxin-like protein HgcB, only one of two conserved cysteines found at the C terminus was necessary for methylation, but either cysteine sufficed. An additional, strictly conserved cysteine, Cys73, was also determined to be essential for methylation. Ultimately, this study supports the previously predicted importance of Cys93 in HgcA for methylation of mercury and reveals additional residues in HgcA and HgcB that facilitate the production of this neurotoxin.« less

Genetic screening of Wnt signaling factors in advanced retinopathy of prematurity

PubMed Central

Takahashi, Hiroshi; Orimo, Hideo; Hiraoka, Miina; Ogata, Tsutomu; Azuma, Noriyuki

2010-01-01

Purpose To evaluate the possibility of genetic involvement in retinopathy of prematurity (ROP). Although ROP is most often associated with low birthweight and low gestational age, these factors do not necessarily predict the severity of ROP. The possible involvement of other factors, including genetic variants, has been considered. Familial exudative vitreoretinopathy (FEVR) is a hereditary vitreoretinal disorder with clinical manifestations similar to those of ROP. Three genes involving the wingless/int1 (Wnt) receptor signaling pathway—FZD4 for frizzled 4, LRP5 for low-density lipoprotein receptor-related protein 5, and ND for Norrie disease protein—are associated with the development of FEVR. Methods In the present study, 17 Japanese patients with advanced ROP were screened for these three candidate genes of FEVR. Genomic DNA from each patient was subjected to PCR and direct sequencing of the ND, FZD4, and LRP5 genes. Results One patient had a heterozygous mutation in the 5′ untranslated region of the ND gene. Another had a leucine insertion in the signal peptide of LRP5. None showed any mutation in FZD4. Conclusions These findings suggest that genetic changes in the Wnt receptor signaling pathway associate to the development of advanced ROP. PMID:21151595

Two novel mutations in the Norrie disease gene associated with the classical ocular phenotype.

PubMed

Caballero, M; Veske, A; Rodriguez, J J; Lugo, N; Schroeder, B; Hesse, L; Gal, A

1996-12-01

Norrie disease (ND) is a rare X-linked recessive disorder characterized by congenital blindness due to a degenerative and proliferative dysplasia of the neuroretina and, occasionally, by deafness and mental handicap. Here, we report two novel mutations detected in patients with the classical eye features of ND. Both the one-base pair insertion in exon II (544/545 insA) and the two-base pair deletion in the start codon (418delTG) of the ND gene predict a functional 'null allele', i.e. the complete absence of the corresponding gene product.

Prevalence, toxin gene profile, antibiotic resistance, and molecular characterization of Clostridium perfringens from diarrheic and non-diarrheic dogs in Korea.

PubMed

Chon, Jung-Whan; Seo, Kun-Ho; Bae, Dongryeoul; Park, Ji-Hee; Khan, Saeed; Sung, Kidon

2018-05-31

Clostridium perfringens causes diarrhea and other diseases in animals and humans. We investigated the prevalence, toxin gene profiles, and antibiotic resistance of C. perfringens isolated from diarrheic dogs (DD) and non-diarrheic dogs (ND) in two animal hospitals in Seoul, Korea. Fecal samples were collected from clinically DD (n = 49) and ND (n = 34). C. perfringens was isolated from 31 of 49 DD (63.3%) and 21 of 34 ND dogs (61.8%). All C. perfringens strains were positive for the α toxin gene, but not for the β, ε, or ι toxin genes; therefore, all strains were identified as type A C. perfringens . All isolates were cpe -negative, whereas the β2 toxin gene was identified in 83.9% and 61.9% of isolates from DD and ND, respectively. Most isolates were susceptible to ampicillin (94%), chloramphenicol (92%), metronidazole (100%), moxifloxacin (96%), and imipenem (100%). However, 25.0% and 21.2% of isolates were resistant to tetracycline and clindamycin, respectively. Molecular subtyping of the isolated strains was performed by using pulsed-field gel electrophoresis. Fifty-two isolates were classified into 48 pulsotypes based on more than 90% similarity of banding patterns. No notable differences were observed among the isolates from DD and ND.

[Polymorphism in the Serotonin Transporter Gene (SLC6A4) and Emotional Bipolar Disorder in Two Regional Mental Health Centers from the Eje Cafetero (Colombia)].

PubMed

Ramos, Lucero Rengifo; Arias, Duverney Gaviria; Salazar, Liliana Salazar; Vélez, Juan Pablo; Pardo, Stella Lozano

2012-03-01

The indel polymorphisms in the promoting region and the 2(nd) intron polymorphisms in the serotonin transporter gene (SLC6A4) have been associated to bipolar disorder 1 (BD1) in several population studies. The objective was to analyze the genotypic and allelic frequencies in both gene regions in a study of cases and controls with individuals from Risaralda and Quindío (Colombia) so as to establish possible associations to BD1, and compare results with previous and similar studies. 133 patients and 120 controls were studied. L and S indel polymorphisms in the promoting region were analyzed by PCR, together with VNTR STin2.10 and STin 2.12 VNTRs polymorphisms in the 2(nd) intron of the SL-C6A4 gene Genotypic and allelic frequencies for the S and L polymorphisms were similar both in cases and controls. However, the LL genotype was significantly increased both in BD1 population (OR=1.89; CI95%=1.1-3.68), and when discriminated by gender. This particular genotype in general population is OR=2.22; IC95%=1.04-5.66 for women, and OR=1.62; IC 95%=0.71-4.39 for men. No significant genotypic and allelic differences were found for VNTR STin2.10 and STin 2.12. polymorphisms. No association was found between polymorphisms of 5-HTTLPR polymorphisms and the 2(nd) intron of the serotonin transporting gene in general patients with BD1, nor when compared by gender. Our results are similar to those reported for Caucasian populations and differ from those of Asian and Brazilian populations. Copyright © 2012 Asociación Colombiana de Psiquiatría. Publicado por Elsevier España. All rights reserved.

Two Closely Related Genes of Arabidopsis Encode Plastidial Cytidinediphosphate Diacylglycerol Synthases Essential for Photoautotrophic Growth1[C

PubMed Central

Haselier, André; Akbari, Hana; Weth, Agnes; Baumgartner, Werner; Frentzen, Margrit

2010-01-01

Cytidinediphosphate diacylglycerol synthase (CDS) catalyzes the formation of cytidinediphosphate diacylglycerol, an essential precursor of anionic phosphoglycerolipids like phosphatidylglycerol or -inositol. In plant cells, CDS isozymes are located in plastids, mitochondria, and microsomes. Here, we show that these isozymes are encoded by five genes in Arabidopsis (Arabidopsis thaliana). Alternative translation initiation or alternative splicing of CDS2 and CDS4 transcripts can result in up to 10 isoforms. Most of the cDNAs encoding the various plant isoforms were functionally expressed in yeast and rescued the nonviable phenotype of the mutant strain lacking CDS activity. The closely related genes CDS4 and CDS5 were found to encode plastidial isozymes with similar catalytic properties. Inactivation of both genes was required to obtain Arabidopsis mutant lines with a visible phenotype, suggesting that the genes have redundant functions. Analysis of these Arabidopsis mutants provided further independent evidence for the importance of plastidial phosphatidylglycerol for structure and function of thylakoid membranes and, hence, for photoautotrophic growth. PMID:20442275

Identification of a peroxisome proliferator-responsive element upstream of the gene encoding rat peroxisomal enoyl-CoA hydratase/3-hydroxyacyl-CoA dehydrogenase.

PubMed Central

Zhang, B; Marcus, S L; Sajjadi, F G; Alvares, K; Reddy, J K; Subramani, S; Rachubinski, R A; Capone, J P

1992-01-01

Ciprofibrate, a hypolipidemic drug that acts as a peroxisome proliferator, induces the transcription of genes encoding peroxisomal beta-oxidation enzymes. To identify cis-acting promoter elements involved in this induction, 5.8 kilobase pairs of promoter sequence from the gene encoding rat peroxisomal enoyl-CoA hydratase/3-hydroxyacyl-CoA dehydrogenase (EC 4.2.1.17/EC 1.1.1.35) was inserted upstream of a luciferase reporter gene. Transfection of this expression vector into rat hepatoma H4IIEC3 cells in the presence of ciprofibrate resulted in a 5- to 10-fold, cell type-specific increase in luciferase activity as compared to cells transfected in the absence of drug. A peroxisome proliferator-responsive element (PPRE) was localized to a 196-nucleotide region centered at position -2943 from the transcription start site. This PPRE conferred ciprofibrate responsiveness on a heterologous promoter and functioned independently of orientation or position. Gel retardation analysis with nuclear extracts demonstrated that ciprofibrate-treated or untreated H4IIEC3 cells, but not HeLa cells or monkey kidney cells, contained sequence-specific DNA binding factors that interact with the PPRE. These results have implications for understanding the mechanisms of coordinated transcriptional induction of genes encoding peroxisomal proteins by hypolipidemic agents and other peroxisome proliferators. Images PMID:1502166

Generation of α1,3-galactosyltransferase and cytidine monophospho-N-acetylneuraminic acid hydroxylase gene double-knockout pigs

PubMed Central

MIYAGAWA, Shuji; MATSUNARI, Hitomi; WATANABE, Masahito; NAKANO, Kazuaki; UMEYAMA, Kazuhiro; SAKAI, Rieko; TAKAYANAGI, Shuko; TAKEISHI, Toki; FUKUDA, Tooru; YASHIMA, Sayaka; MAEDA, Akira; EGUCHI, Hiroshi; OKUYAMA, Hiroomi; NAGAYA, Masaki; NAGASHIMA, Hiroshi

2015-01-01

Zinc-finger nucleases (ZFNs) and transcription activator-like effector nucleases (TALENs) are new tools for producing gene knockout (KO) animals. The current study reports produced genetically modified pigs, in which two endogenous genes were knocked out. Porcine fibroblast cell lines were derived from homozygous α1,3-galactosyltransferase (GalT) KO pigs. These cells were subjected to an additional KO for the cytidine monophospho-N-acetylneuraminic acid hydroxylase (CMAH) gene. A pair of ZFN-encoding mRNAs targeting exon 8 of the CMAH gene was used to generate the heterozygous CMAH KO cells, from which cloned pigs were produced by somatic cell nuclear transfer (SCNT). One of the cloned pigs obtained was re-cloned after additional KO of the remaining CMAH allele using the same ZFN-encoding mRNAs to generate GalT/CMAH-double homozygous KO pigs. On the other hand, the use of TALEN-encoding mRNAs targeting exon 7 of the CMAH gene resulted in efficient generation of homozygous CMAH KO cells. These cells were used for SCNT to produce cloned pigs homozygous for a double GalT/CMAH KO. These results demonstrate that the combination of TALEN-encoding mRNA, in vitro selection of the nuclear donor cells and SCNT provides a robust method for generating KO pigs. PMID:26227017

Generation of α1,3-galactosyltransferase and cytidine monophospho-N-acetylneuraminic acid hydroxylase gene double-knockout pigs.

PubMed

Miyagawa, Shuji; Matsunari, Hitomi; Watanabe, Masahito; Nakano, Kazuaki; Umeyama, Kazuhiro; Sakai, Rieko; Takayanagi, Shuko; Takeishi, Toki; Fukuda, Tooru; Yashima, Sayaka; Maeda, Akira; Eguchi, Hiroshi; Okuyama, Hiroomi; Nagaya, Masaki; Nagashima, Hiroshi

2015-01-01

Zinc-finger nucleases (ZFNs) and transcription activator-like effector nucleases (TALENs) are new tools for producing gene knockout (KO) animals. The current study reports produced genetically modified pigs, in which two endogenous genes were knocked out. Porcine fibroblast cell lines were derived from homozygous α1,3-galactosyltransferase (GalT) KO pigs. These cells were subjected to an additional KO for the cytidine monophospho-N-acetylneuraminic acid hydroxylase (CMAH) gene. A pair of ZFN-encoding mRNAs targeting exon 8 of the CMAH gene was used to generate the heterozygous CMAH KO cells, from which cloned pigs were produced by somatic cell nuclear transfer (SCNT). One of the cloned pigs obtained was re-cloned after additional KO of the remaining CMAH allele using the same ZFN-encoding mRNAs to generate GalT/CMAH-double homozygous KO pigs. On the other hand, the use of TALEN-encoding mRNAs targeting exon 7 of the CMAH gene resulted in efficient generation of homozygous CMAH KO cells. These cells were used for SCNT to produce cloned pigs homozygous for a double GalT/CMAH KO. These results demonstrate that the combination of TALEN-encoding mRNA, in vitro selection of the nuclear donor cells and SCNT provides a robust method for generating KO pigs.

Genome Enabled Discovery of Carbon Sequestration Genes in Poplar

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

Filichkin, Sergei; Etherington, Elizabeth; Ma, Caiping

2007-02-22

The goals of the S.H. Strauss laboratory portion of 'Genome-enabled discovery of carbon sequestration genes in poplar' are (1) to explore the functions of candidate genes using Populus transformation by inserting genes provided by Oakridge National Laboratory (ORNL) and the University of Florida (UF) into poplar; (2) to expand the poplar transformation toolkit by developing transformation methods for important genotypes; and (3) to allow induced expression, and efficient gene suppression, in roots and other tissues. As part of the transformation improvement effort, OSU developed transformation protocols for Populus trichocarpa 'Nisqually-1' clone and an early flowering P. alba clone, 6K10. Completemore » descriptions of the transformation systems were published (Ma et. al. 2004, Meilan et. al 2004). Twenty-one 'Nisqually-1' and 622 6K10 transgenic plants were generated. To identify root predominant promoters, a set of three promoters were tested for their tissue-specific expression patterns in poplar and in Arabidopsis as a model system. A novel gene, ET304, was identified by analyzing a collection of poplar enhancer trap lines generated at OSU (Filichkin et. al 2006a, 2006b). Other promoters include the pGgMT1 root-predominant promoter from Casuarina glauca and the pAtPIN2 promoter from Arabidopsis root specific PIN2 gene. OSU tested two induction systems, alcohol- and estrogen-inducible, in multiple poplar transgenics. Ethanol proved to be the more efficient when tested in tissue culture and greenhouse conditions. Two estrogen-inducible systems were evaluated in transgenic Populus, neither of which functioned reliably in tissue culture conditions. GATEWAY-compatible plant binary vectors were designed to compare the silencing efficiency of homologous (direct) RNAi vs. heterologous (transitive) RNAi inverted repeats. A set of genes was targeted for post transcriptional silencing in the model Arabidopsis system; these include the floral meristem identity gene (APETALA1 or AP1), auxin response factor gene (ETTIN), the gene encoding transcriptional factor of WD40 family (TRANSPARENTTESTAGLABRA1 or TTG1), and the auxin efflux carrier (PIN-FORMED2 or PIN2) gene. More than 220 transgenic lines of the 1st, 2nd and 3rd generations were analyzed for RNAi suppression phenotypes (Filichkin et. al., manuscript submitted). A total of 108 constructs were supplied by ORNL, UF and OSU and used to generate over 1,881 PCR verified transgenic Populus and over 300 PCR verified transgenic Arabidopsis events. The Populus transgenics alone required Agrobacterium co-cultivations of 124.406 explants.« less

Sequence analysis of mitochondrial ND1 gene can reveal the genetic structure and origin of Bactrocera dorsalis s.s.

PubMed Central

2014-01-01

Background The oriental fruit fly, Bactrocera dorsalis s.s., is one of the most important quarantine pests in many countries, including China. Although the oriental fruit fly has been investigated extensively, its origins and genetic structure remain disputed. In this study, the NADH dehydrogenase subunit 1 (ND1) gene was used as a genetic marker to examine the genetic diversity, population structure, and gene flow of B. dorsalis s.s. throughout its range in China and southeast Asia. Results Haplotype networks and phylogenetic analysis indicated two distinguishable lineages of the fly population but provided no strong support for geographical subdivision in B. philippinensis. Demographic analysis revealed rapid expansion of B. dorsalis s.s. populations in China and Southeast Asia in the recent years. The greatest amount of genetic diversity was observed in Manila, Pattaya, and Bangkok, and asymmetric migration patterns were observed in different parts of China. The data collected here further show that B. dorsalis s.s. in Yunnan, Guangdong, and Fujian Provinces, and in Taiwan might have different origins within southeast Asia. Conclusions Using the mitochondrial ND1 gene, the results of the present study showed B. dorsalis s.s. from different parts of China to have different genetic structures and origins. B. dorsalis s.s. in China and southeast Asia was found to have experienced rapid expansion in recent years. Data further support the existence of two distinguishable lineages of B. dorsalis s.s. in China and indicate genetic diversity and gene flow from multiple origins. The sequences in this paper have been deposited in GenBank/NCBI under accession numbers KC413034–KC413367. PMID:24655832

Pancreatic β-Cell Dysfunction in Diet-Induced Obese Mice: Roles of AMP-Kinase, Protein Kinase Cε, Mitochondrial and Cholesterol Metabolism, and Alterations in Gene Expression

PubMed Central

Pepin, Émilie; Al-Mass, Anfal; Attané, Camille; Zhang, Kezhuo; Lamontagne, Julien; Lussier, Roxane; Madiraju, S. R. Murthy; Joly, Erik; Ruderman, Neil B.; Sladek, Robert; Prentki, Marc; Peyot, Marie-Line

2016-01-01

Diet induced obese (DIO) mice can be stratified according to their weight gain in response to high fat diet as low responders (LDR) and high responders (HDR). This allows the study of β-cell failure and the transitions to prediabetes (LDR) and early diabetes (HDR). C57BL/6N mice were fed for 8 weeks with a normal chow diet (ND) or a high fat diet and stratified as LDR and HDR. Freshly isolated islets from ND, LDR and HDR mice were studied ex-vivo for mitochondrial metabolism, AMPK activity and signalling, the expression and activity of key enzymes of energy metabolism, cholesterol synthesis, and mRNA profiling. Severely compromised glucose-induced insulin secretion in HDR islets, as compared to ND and LDR islets, was associated with suppressed AMP-kinase activity. HDR islets also showed reduced acetyl-CoA carboxylase activity and enhanced activity of 3-hydroxy-3-methylglutaryl-CoA reductase, which led respectively to elevated fatty acid oxidation and increased cholesterol biosynthesis. HDR islets also displayed mitochondrial membrane hyperpolarization and reduced ATP turnover in the presence of elevated glucose. Expression of protein kinase Cε, which reduces both lipolysis and production of signals for insulin secretion, was elevated in DIO islets. Genes whose expression increased or decreased by more than 1.2-fold were minor between LDR and ND islets (17 differentially expressed), but were prominent between HDR and ND islets (1508 differentially expressed). In HDR islets, particularly affected genes were related to cell cycle and proliferation, AMPK signaling, mitochondrial metabolism and cholesterol metabolism. In conclusion, chronically reduced AMPK activity, mitochondrial dysfunction, elevated cholesterol biosynthesis in islets, and substantial alterations in gene expression accompany β-cell failure in HDR islets. The β-cell compensation process in the prediabetic state (LDR) is largely independent of transcriptional adaptive changes, whereas the transition to early diabetes (HDR) is associated with major alterations in gene expression. PMID:27043434

GH51 Arabinofuranosidase and Its Role in the Methylglucuronoarabinoxylan Utilization System in Paenibacillus sp. Strain JDR-2

PubMed Central

Sawhney, Neha

2014-01-01

Methylglucuronoarabinoxylan (MeGAXn) from agricultural residues and energy crops is a significant yet underutilized biomass resource for production of biofuels and chemicals. Mild thermochemical pretreatment of bagasse yields MeGAXn requiring saccharifying enzymes for conversion to fermentable sugars. A xylanolytic bacterium, Paenibacillus sp. strain JDR-2, produces an extracellular cell-associated GH10 endoxylanse (XynA1) which efficiently depolymerizes methylglucuronoxylan (MeGXn) from hardwoods coupled with assimilation of oligosaccharides for further processing by intracellular GH67 α-glucuronidase, GH10 endoxylanase, and GH43 β-xylosidase. This process has been ascribed to genes that comprise a xylan utilization regulon that encodes XynA1 and includes a gene cluster encoding transcriptional regulators, ABC transporters, and intracellular enzymes that convert assimilated oligosaccharides to fermentable sugars. Here we show that Paenibacillus sp. JDR-2 utilized MeGAXn without accumulation of oligosaccharides in the medium. The Paenibacillus sp. JDR-2 growth rate on MeGAXn was 3.1-fold greater than that on oligosaccharides generated from MeGAXn by XynA1. Candidate genes encoding GH51 arabinofuranosidases with potential roles were identified. Following growth on MeGAXn, quantitative reverse transcription-PCR identified a cluster of genes encoding a GH51 arabinofuranosidase (AbfB) and transcriptional regulators which were coordinately expressed along with the genes comprising the xylan utilization regulon. The action of XynA1 on MeGAXn generated arabinoxylobiose, arabinoxylotriose, xylobiose, xylotriose, and methylglucuronoxylotriose. Recombinant AbfB processed arabinoxylooligosaccharides to xylooligosaccharides and arabinose. MeGAXn processing by Paenibacillus sp. JDR-2 may be achieved by extracellular depolymerization by XynA1 coupled to assimilation of oligosaccharides and further processing by intracellular enzymes, including AbfB. Paenibacillus sp. JDR-2 provides a GH10/GH67 system complemented with genes encoding intracellular GH51 arabinofuranosidases for efficient utilization of MeGAXn. PMID:25063665

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

Stols, L.; Donnelly, M.I.; Kulkarni, G.

The malic enzyme gene of Ascaris suum was cloned into the vector pTRC99a in two forms encoding alternative amino-termini. The resulting plasmids, pMEA1 and pMEA2, were introduced into Escherichia coli NZN111, a strain that is unable to grow fermentatively because of inactivation of the genes encoding pyruvate dissimilation. Induction of pMEA1, which encodes the native animoterminus, gave better overexpression of malic enzyme, approx 12-fold compared to uninduced cells. Under the appropriate culture conditions, expression of malic enzyme allowed the fermentative dissimilation of glucose by NZN111. The major fermentation product formed in induced cultures was succinic acid.

Cloning and characterization of the major histone H2A genes completes the cloning and sequencing of known histone genes of Tetrahymena thermophila.

PubMed Central

Liu, X; Gorovsky, M A

1996-01-01

A truncated cDNA clone encoding Tetrahymena thermophila histone H2A2 was isolated using synthetic degenerate oligonucleotide probes derived from H2A protein sequences of Tetrahymena pyriformis. The cDNA clone was used as a homologous probe to isolate a truncated genomic clone encoding H2A1. The remaining regions of the genes for H2A1 (HTA1) and H2A2 (HTA2) were then isolated using inverse PCR on circularized genomic DNA fragments. These partial clones were assembled into intact HTA1 and HTA2 clones. Nucleotide sequences of the two genes were highly homologous within the coding region but not in the noncoding regions. Comparison of the deduced amino acid sequences with protein sequences of T. pyriformis H2As showed only two and three differences respectively, in a total of 137 amino acids for H2A1, and 132 amino acids for H2A2, indicating the two genes arose before the divergence of these two species. The HTA2 gene contains a TAA triplet within the coding region, encoding a glutamine residue. In contrast with the T. thermophila HHO and HTA3 genes, no introns were identified within the two genes. The 5'- and 3'-ends of the histone H2A mRNAs; were determined by RNase protection and by PCR mapping using RACE and RLM-RACE methods. Both genes encode polyadenylated mRNAs and are highly expressed in vegetatively growing cells but only weakly expressed in starved cultures. With the inclusion of these two genes, T. thermophila is the first organism whose entire complement of known core and linker histones, including replication-dependent and basal variants, has been cloned and sequenced. PMID:8760889

Meta-Analyses of ALDH2 and ADH1B with Alcohol Dependence in Asians

ERIC Educational Resources Information Center

Luczak, Susan E.; Glatt, Stephen J.; Wall, Tamara J.

2006-01-01

Meta-analyses were conducted to determine the magnitude of relationships between polymorphisms in 2 genes, ALDH2 and ADH1B, with alcohol dependence in Asians. For each gene, possession of 1 variant [asterisk]2 allele was protective against alcohol dependence, and possession of a 2nd [asterisk]2 allele did not offer significant additional…

[Construction of plant expression plasmid of chimera SBR-CT delta A1].

PubMed

Mai, Sui; Ling, Junqi

2003-08-01

The purpose of this study is to construct plant expression plasmid containing the gene encoding chimera SBR-CT delta A1. The target gene fragment P2, including the gene-encoded chimera SBR-CT delta A1 (3,498-5,378 bp), was obtained by standard PCR amplification. The PCR products were ligated with pGEM-easy vector through TA clone to form plasmid pTSC. The plasmid pTSC and plasmid pPOKII were digested by restricted endonuclease BamHI and KpnI, and the digested products were extracted and purified for recombination. Then the purified P2 and plasmid pPOKII were recombined by T4 DNA ligase to form recombinant plasmid pROSC; inserting bar gene into the plasmid and form pROSB plasmid. The recombined plasmids were isolated and identified by restricted endonuclease cutting and Sanger dideoxy DNA sequencing. P2 gene was linked to pPOKII plasmid and formed recombinant plasmid pROSC. The DNA sequence and orientation were corrected. And bar gene was inserted into pPOSC and form recombinant plasmid pROSB. Plant expression vector pROSC and pROSB containing the gene encoding chimera SBR-CT delta A1, which may provide useful experiment foundation for further study on edible vaccine against caries have been successfully constructed.

Mutations in Mitochondrial DNA From Pancreatic Ductal Adenocarcinomas Associate With Survival Times of Patients and Accumulate as Tumors Progress.

PubMed

Hopkins, Julia F; Denroche, Robert E; Aguiar, Jennifer A; Notta, Faiyaz; Connor, Ashton A; Wilson, Julie M; Stein, Lincoln D; Gallinger, Steven; Boutros, Paul C

2018-05-01

Somatic mutations have been found in the mitochondria in different types of cancer cells, but it is not clear whether these affect tumorigenesis or tumor progression. We analyzed mitochondrial genomes of 268 early-stage, resected pancreatic ductal adenocarcinoma tissues and paired non-tumor tissues. We defined a mitochondrial somatic mutation (mtSNV) as a position where the difference in heteroplasmy fraction between tumor and normal sample was ≥0.2. Our analysis identified 304 mtSNVs, with at least 1 mtSNV in 61% (164 of 268) of tumor samples. The noncoding control region had the greatest proportion of mtSNVs (60 of 304 mutations); this region contains sites that regulate mitochondrial DNA transcription and replication. Frequently mutated genes included ND5, RNR2, and CO1, plus 29 mutations in transfer RNA genes. mtSNVs in 2 separate mitochondrial genes (ND4 and ND6) were associated with shorter overall survival time. This association appeared to depend on the level of mtSNV heteroplasmy. Non-random co-occurrence between mtSNVs and mutations in nuclear genes indicates interactions between nuclear and mitochondrial DNA. In an analysis of primary tumors and metastases from 6 patients, we found tumors to accumulate mitochondrial mutational mutations as they progress. Copyright © 2018 AGA Institute. Published by Elsevier Inc. All rights reserved.

Heterologous expression and transcript analysis of gibberellin biosynthetic genes of grasses reveals novel functionality in the GA3ox family.

PubMed

Pearce, Stephen; Huttly, Alison K; Prosser, Ian M; Li, Yi-dan; Vaughan, Simon P; Gallova, Barbora; Patil, Archana; Coghill, Jane A; Dubcovsky, Jorge; Hedden, Peter; Phillips, Andrew L

2015-06-05

The gibberellin (GA) pathway plays a central role in the regulation of plant development, with the 2-oxoglutarate-dependent dioxygenases (2-ODDs: GA20ox, GA3ox, GA2ox) that catalyse the later steps in the biosynthetic pathway of particularly importance in regulating bioactive GA levels. Although GA has important impacts on crop yield and quality, our understanding of the regulation of GA biosynthesis during wheat and barley development remains limited. In this study we identified or assembled genes encoding the GA 2-ODDs of wheat, barley and Brachypodium distachyon and characterised the wheat genes by heterologous expression and transcript analysis. The wheat, barley and Brachypodium genomes each contain orthologous copies of the GA20ox, GA3ox and GA2ox genes identified in rice, with the exception of OsGA3ox1 and OsGA2ox5 which are absent in these species. Some additional paralogs of 2-ODD genes were identified: notably, a novel gene in the wheat B genome related to GA3ox2 was shown to encode a GA 1-oxidase, named as TaGA1ox-B1. This enzyme is likely to be responsible for the abundant 1β-hydroxylated GAs present in developing wheat grains. We also identified a related gene in barley, located in a syntenic position to TaGA1ox-B1, that encodes a GA 3,18-dihydroxylase which similarly accounts for the accumulation of unusual GAs in barley grains. Transcript analysis showed that some paralogs of the different classes of 2-ODD were expressed mainly in a single tissue or at specific developmental stages. In particular, TaGA20ox3, TaGA1ox1, TaGA3ox3 and TaGA2ox7 were predominantly expressed in developing grain. More detailed analysis of grain-specific gene expression showed that while the transcripts of biosynthetic genes were most abundant in the endosperm, genes encoding inactivation and signalling components were more highly expressed in the seed coat and pericarp. The comprehensive expression and functional characterisation of the multigene families encoding the 2-ODD enzymes of the GA pathway in wheat and barley will provide the basis for a better understanding of GA-regulated development in these species. This analysis revealed the existence of a novel, endosperm-specific GA 1-oxidase in wheat and a related GA 3,18-dihydroxylase enzyme in barley that may play important roles during grain expansion and development.

Deletion of a Single-Copy Trna Affects Microtubule Function in Saccharomyces Cerevisiae

PubMed Central

Reijo, R. A.; Cho, D. S.; Huffaker, T. C.

1993-01-01

rts1-1 was identified as an extragenic suppressor of tub2-104, a cold-sensitive allele of the sole gene encoding β-tubulin in the yeast, Saccharomyces cerevisiae. In addition, rts1-1 cells are heat sensitive and resistant to the microtubule-destabilizing drug, benomyl. The rts1-1 mutation is a deletion of approximately 5 kb of genomic DNA on chromosome X that includes one open reading frame and three tRNA genes. Dissection of this region shows that heat sensitivity is due to deletion of the open reading frame (HIT1). Suppression and benomyl resistance are caused by deletion of the gene encoding a tRNA(AGG)(Arg) (HSX1). Northern analysis of rts1-1 cells indicates that HSX1 is the only gene encoding this tRNA. Deletion of HSX1 does not suppress the tub2-104 mutation by misreading at the AGG codons in TUB2. It also does not suppress by interfering with the protein arginylation that targets certain proteins for degradation. These results leave open the prospect that this tRNA(AGG)(Arg) plays a novel role in the cell. PMID:8307335

Targeted knock-out of a gene encoding sulfite reductase in the moss Physcomitrella patens affects gametophytic and sporophytic development.

PubMed

Wiedemann, Gertrud; Hermsen, Corinna; Melzer, Michael; Büttner-Mainik, Annette; Rennenberg, Heinz; Reski, Ralf; Kopriva, Stanislav

2010-06-03

A key step in sulfate assimilation into cysteine is the reduction of sulfite to sulfide by sulfite reductase (SiR). This enzyme is encoded by three genes in the moss Physcomitrella patens. To obtain a first insight into the roles of the individual isoforms, we deleted the gene encoding the SiR1 isoform in P. patens by homologous recombination and subsequently analysed the DeltaSiR1 mutants. While DeltaSiR1 mutants showed no obvious alteration in sulfur metabolism, their regeneration from protoplasts and their ability to produce mature spores was significantly affected, highlighting an unexpected link between moss sulfate assimilation and development, that is yet to be characterized. Copyright 2010 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

Viral repression of fungal pheromone precursor gene expression.

PubMed

Zhang, L; Baasiri, R A; Van Alfen, N K

1998-02-01

Biological control of chestnut blight caused by the filamentous ascomycete Cryphonectria parasitica can be achieved with a virus that infects this fungus. This hypovirus causes a perturbation of fungal development that results in low virulence (hypovirulence), poor asexual sporulation, and female infertility without affecting fungal growth in culture. At the molecular level, the virus is known to affect the transcription of a number of fungal genes. Two of these genes, Vir1 and Vir2, produce abundant transcripts in noninfected strains of the fungus, but the transcripts are not detectable in virus-infected strains. We report here that these two genes encode the pheromone precursors of the Mat-2 mating type of the fungus; consequently, these genes have been renamed Mf2/1 and Mf2/2. To determine if the virus affects the mating systems of both mating types of this fungus, the pheromone precursor gene, Mf1/1, of a Mat-1 strain was cloned and likewise was found to be repressed in virus-infected strains. The suppression of transcription of the pheromone precursor genes of this fungus could be the cause of the mating defect of infected strains of the fungus. Although published reports suggest that a G alpha(i) subunit may be involved in this regulation, our results do not support this hypothesis. The prepropheromone encoded by Mf1/1 is structurally similar to that of the prepro-p-factor of Schizosaccharomyces pombe. This is the first description of the complete set of pheromone precursor genes encoded by a filamentous ascomycete.

Expression profiles of genes for mitochondrial respiratory energy-dissipating systems and antioxidant enzymes in wheat leaves during de-etiolation.

PubMed

Garmash, Elena V; Velegzhaninov, Ilya O; Grabelnych, Olga I; Borovik, Olga A; Silina, Ekaterina V; Voinikov, Victor K; Golovko, Tamara K

2017-08-01

Mitochondrial respiratory components participate in the maintenance of chloroplast functional activity. This study investigates the effects 48h de-etiolation of spring wheat seedlings (Triticum aestivum L., var. Irgina) on the expression of genes that encode energy-dissipating respiratory components and antioxidant enzymes under continuous light conditions. The expression of AOX1a following the prolonged darkness exhibited a pattern indicating a prominent dependence on light. The expression of other respiratory genes, including NDA2, NDB2, and UCP1b, increased during de-etiolation and dark-to-light transition; however, changes in the expression of these genes occurred later than those in AOX1a expression. A high expression of NDA1 was detected after 12h of de-etiolation. The suppression of AOX1a, NDA2, NDB2, and UCP1b was observed 24h after de-etiolation when the photosynthetic apparatus and its defence systems against excess light were completely developed. The expression patterns of the respiratory genes and several genes encoding antioxidant enzymes (MnSOD, Cu-ZnSOD, t-APX, GR, and GRX) were quite similar. Our data indicate that the induction of nuclear genes encoding respiratory and antioxidant enzymes allow the plants to control reactive oxygen species (ROS) production and avoid oxidative stress during de-etiolation. Copyright © 2017 Elsevier GmbH. All rights reserved.

Meta-omic signatures of microbial metal and nitrogen cycling in marine oxygen minimum zones

PubMed Central

Glass, Jennifer B.; Kretz, Cecilia B.; Ganesh, Sangita; Ranjan, Piyush; Seston, Sherry L.; Buck, Kristen N.; Landing, William M.; Morton, Peter L.; Moffett, James W.; Giovannoni, Stephen J.; Vergin, Kevin L.; Stewart, Frank J.

2015-01-01

Iron (Fe) and copper (Cu) are essential cofactors for microbial metalloenzymes, but little is known about the metalloenyzme inventory of anaerobic marine microbial communities despite their importance to the nitrogen cycle. We compared dissolved O2, NO3−, NO2−, Fe and Cu concentrations with nucleic acid sequences encoding Fe and Cu-binding proteins in 21 metagenomes and 9 metatranscriptomes from Eastern Tropical North and South Pacific oxygen minimum zones and 7 metagenomes from the Bermuda Atlantic Time-series Station. Dissolved Fe concentrations increased sharply at upper oxic-anoxic transition zones, with the highest Fe:Cu molar ratio (1.8) occurring at the anoxic core of the Eastern Tropical North Pacific oxygen minimum zone and matching the predicted maximum ratio based on data from diverse ocean sites. The relative abundance of genes encoding Fe-binding proteins was negatively correlated with O2, driven by significant increases in genes encoding Fe-proteins involved in dissimilatory nitrogen metabolisms under anoxia. Transcripts encoding cytochrome c oxidase, the Fe- and Cu-containing terminal reductase in aerobic respiration, were positively correlated with O2 content. A comparison of the taxonomy of genes encoding Fe- and Cu-binding vs. bulk proteins in OMZs revealed that Planctomycetes represented a higher percentage of Fe genes while Thaumarchaeota represented a higher percentage of Cu genes, particularly at oxyclines. These results are broadly consistent with higher relative abundance of genes encoding Fe-proteins in the genome of a marine planctomycete vs. higher relative abundance of genes encoding Cu-proteins in the genome of a marine thaumarchaeote. These findings highlight the importance of metalloenzymes for microbial processes in oxygen minimum zones and suggest preferential Cu use in oxic habitats with Cu > Fe vs. preferential Fe use in anoxic niches with Fe > Cu. PMID:26441925

Meta-omic signatures of microbial metal and nitrogen cycling in marine oxygen minimum zones.

PubMed

Glass, Jennifer B; Kretz, Cecilia B; Ganesh, Sangita; Ranjan, Piyush; Seston, Sherry L; Buck, Kristen N; Landing, William M; Morton, Peter L; Moffett, James W; Giovannoni, Stephen J; Vergin, Kevin L; Stewart, Frank J

2015-01-01

Iron (Fe) and copper (Cu) are essential cofactors for microbial metalloenzymes, but little is known about the metalloenyzme inventory of anaerobic marine microbial communities despite their importance to the nitrogen cycle. We compared dissolved O2, NO[Formula: see text], NO[Formula: see text], Fe and Cu concentrations with nucleic acid sequences encoding Fe and Cu-binding proteins in 21 metagenomes and 9 metatranscriptomes from Eastern Tropical North and South Pacific oxygen minimum zones and 7 metagenomes from the Bermuda Atlantic Time-series Station. Dissolved Fe concentrations increased sharply at upper oxic-anoxic transition zones, with the highest Fe:Cu molar ratio (1.8) occurring at the anoxic core of the Eastern Tropical North Pacific oxygen minimum zone and matching the predicted maximum ratio based on data from diverse ocean sites. The relative abundance of genes encoding Fe-binding proteins was negatively correlated with O2, driven by significant increases in genes encoding Fe-proteins involved in dissimilatory nitrogen metabolisms under anoxia. Transcripts encoding cytochrome c oxidase, the Fe- and Cu-containing terminal reductase in aerobic respiration, were positively correlated with O2 content. A comparison of the taxonomy of genes encoding Fe- and Cu-binding vs. bulk proteins in OMZs revealed that Planctomycetes represented a higher percentage of Fe genes while Thaumarchaeota represented a higher percentage of Cu genes, particularly at oxyclines. These results are broadly consistent with higher relative abundance of genes encoding Fe-proteins in the genome of a marine planctomycete vs. higher relative abundance of genes encoding Cu-proteins in the genome of a marine thaumarchaeote. These findings highlight the importance of metalloenzymes for microbial processes in oxygen minimum zones and suggest preferential Cu use in oxic habitats with Cu > Fe vs. preferential Fe use in anoxic niches with Fe > Cu.

Glycogen Metabolic Genes Are Involved in Trehalose-6-Phosphate Synthase-Mediated Regulation of Pathogenicity by the Rice Blast Fungus Magnaporthe oryzae

PubMed Central

Wilson, Richard A.; Wang, Zheng-Yi; Kershaw, Michael J.; Talbot, Nicholas J.

2013-01-01

The filamentous fungus Magnaporthe oryzae is the causal agent of rice blast disease. Here we show that glycogen metabolic genes play an important role in plant infection by M. oryzae. Targeted deletion of AGL1 and GPH1, which encode amyloglucosidase and glycogen phosphorylase, respectively, prevented mobilisation of glycogen stores during appressorium development and caused a significant reduction in the ability of M. oryzae to cause rice blast disease. By contrast, targeted mutation of GSN1, which encodes glycogen synthase, significantly reduced the synthesis of intracellular glycogen, but had no effect on fungal pathogenicity. We found that loss of AGL1 and GPH1 led to a reduction in expression of TPS1 and TPS3, which encode components of the trehalose-6-phosphate synthase complex, that acts as a genetic switch in M. oryzae. Tps1 responds to glucose-6-phosphate levels and the balance of NADP/NADPH to regulate virulence-associated gene expression, in association with Nmr transcriptional inhibitors. We show that deletion of the NMR3 transcriptional inhibitor gene partially restores virulence to a Δagl1Δgph1 mutant, suggesting that glycogen metabolic genes are necessary for operation of the NADPH-dependent genetic switch in M. oryzae. PMID:24098112

Delay-Encoded Harmonic Imaging (DE-HI) in Multiplane-Wave Compounding.

PubMed

Gong, Ping; Song, Pengfei; Chen, Shigao

2017-04-01

The development of ultrafast ultrasound imaging brings great opportunities to improve imaging technologies such as shear wave elastography and ultrafast Doppler imaging. In ultrafast imaging, several tilted plane or diverging wave images are coherently combined to form a compounded image, leading to trade-offs among image signal-to-noise ratio (SNR), resolution, and post-compounded frame rate. Multiplane wave (MW) imaging is proposed to solve this trade-off by encoding multiple plane waves with Hadamard matrix during one transmission event (i.e. pulse-echo event), to improve image SNR without sacrificing the resolution or frame rate. However, it suffers from stronger reverberation artifacts in B-mode images compared to standard plane wave compounding due to longer transmitted pulses. If harmonic imaging can be combined with MW imaging, the reverberation artifacts and other clutter noises such as sidelobes and multipath scattering clutters should be suppressed. The challenge, however, is that the Hadamard codes used in MW imaging cannot encode the 2 nd harmonic component by inversing the pulse polarity. In this paper, we propose a delay-encoded harmonic imaging (DE-HI) technique to encode the 2 nd harmonic with a one quarter period delay calculated at the transmit center frequency, rather than reversing the pulse polarity during multiplane wave emissions. Received DE-HI signals can then be decoded in the frequency domain to recover the signals as in single plane wave emissions, but mainly with improved SNR at the 2 nd harmonic component instead of the fundamental component. DE-HI was tested experimentally with a point target, a B-mode imaging phantom, and in-vivo human liver imaging. Improvements in image contrast-to-noise ratio (CNR), spatial resolution, and lesion-signal-to-noise ratio ( l SNR) have been achieved compared to standard plane wave compounding, MW imaging, and standard harmonic imaging (maximal improvement of 116% on CNR and 115% on l SNR as compared to standard HI around 55 mm depth in the B-mode imaging phantom study). The potential high frame rate and the stability of encoding and decoding processes of DE-HI were also demonstrated, which made DE-HI promising for a wide spectrum of imaging applications.

Next-generation sequencing reveals a novel NDP gene mutation in a Chinese family with Norrie disease.

PubMed

Huang, Xiaoyan; Tian, Mao; Li, Jiankang; Cui, Ling; Li, Min; Zhang, Jianguo

2017-11-01

Norrie disease (ND) is a rare X-linked genetic disorder, the main symptoms of which are congenital blindness and white pupils. It has been reported that ND is caused by mutations in the NDP gene. Although many mutations in NDP have been reported, the genetic cause for many patients remains unknown. In this study, the aim is to investigate the genetic defect in a five-generation family with typical symptoms of ND. To identify the causative gene, next-generation sequencing based target capture sequencing was performed. Segregation analysis of the candidate variant was performed in additional family members using Sanger sequencing. We identified a novel missense variant (c.314C>A) located within the NDP gene. The mutation cosegregated within all affected individuals in the family and was not found in unaffected members. By happenstance, in this family, we also detected a known pathogenic variant of retinitis pigmentosa in a healthy individual. c.314C>A mutation of NDP gene is a novel mutation and broadens the genetic spectrum of ND.

Next-generation sequencing reveals a novel NDP gene mutation in a Chinese family with Norrie disease

PubMed Central

Huang, Xiaoyan; Tian, Mao; Li, Jiankang; Cui, Ling; Li, Min; Zhang, Jianguo

2017-01-01

Purpose: Norrie disease (ND) is a rare X-linked genetic disorder, the main symptoms of which are congenital blindness and white pupils. It has been reported that ND is caused by mutations in the NDP gene. Although many mutations in NDP have been reported, the genetic cause for many patients remains unknown. In this study, the aim is to investigate the genetic defect in a five-generation family with typical symptoms of ND. Methods: To identify the causative gene, next-generation sequencing based target capture sequencing was performed. Segregation analysis of the candidate variant was performed in additional family members using Sanger sequencing. Results: We identified a novel missense variant (c.314C>A) located within the NDP gene. The mutation cosegregated within all affected individuals in the family and was not found in unaffected members. By happenstance, in this family, we also detected a known pathogenic variant of retinitis pigmentosa in a healthy individual. Conclusion: c.314C>A mutation of NDP gene is a novel mutation and broadens the genetic spectrum of ND. PMID:29133643

Oxalate-Metabolising Genes of the White-Rot Fungus Dichomitus squalens Are Differentially Induced on Wood and at High Proton Concentration

PubMed Central

de Vries, Ronald P.; Timonen, Sari; Hildén, Kristiina

2014-01-01

Oxalic acid is a prevalent fungal metabolite with versatile roles in growth and nutrition, including degradation of plant biomass. However, the toxicity of oxalic acid makes regulation of its intra- and extracellular concentration crucial. To increase the knowledge of fungal oxalate metabolism, a transcriptional level study on oxalate-catabolising genes was performed with an effective lignin-degrading white-rot fungus Dichomitus squalens, which has demonstrated particular abilities in production and degradation of oxalic acid. The expression of oxalic-acid decomposing oxalate decarboxylase (ODC) and formic-acid decomposing formate dehydrogenase (FDH) encoding genes was followed during the growth of D. squalens on its natural spruce wood substrate. The effect of high proton concentration on the regulation of the oxalate-catabolising genes was determined after addition of organic acid (oxalic acid) and inorganic acid (hydrochloric acid) to the liquid cultures of D. squalens. In order to evaluate the co-expression of oxalate-catabolising and manganese peroxidase (MnP) encoding genes, the expression of one MnP encoding gene, mnp1, of D. squalens was also surveyed in the solid state and liquid cultures. Sequential action of ODC and FDH encoding genes was detected in the studied cultivations. The odc1, fdh2 and fdh3 genes of D. squalens showed constitutive expression, whereas ODC2 and FHD1 most likely are the main responsible enzymes for detoxification of high concentrations of oxalic and formic acids. The results also confirmed the central role of ODC1 when D. squalens grows on coniferous wood. Phylogenetic analysis revealed that fungal ODCs have evolved from at least two gene copies whereas FDHs have a single ancestral gene. As a conclusion, the multiplicity of oxalate-catabolising genes and their differential regulation on wood and in acid-amended cultures of D. squalens point to divergent physiological roles for the corresponding enzymes. PMID:24505339

Ti plasmid-encoded genes responsible for catabolism of the crown gall opine mannopine by Agrobacterium tumefaciens are homologs of the T-region genes responsible for synthesis of this opine by the plant tumor.

PubMed

Kim, K S; Farrand, S K

1996-06-01

Agrobacterium tumefaciens NT1 harboring pSaB4, which contains the 14-kb BamHI fragment 4 from the octopine/mannityl opine-type Ti plasmid pTi15955, grew well with agropine (AGR) but slowly with mannopine (MOP) as the sole carbon source. When a second plasmid encoding a dedicated transport system for MOP was introduced, these cells grew well with both AGR and MOP. Transposon insertion mutagenesis and subcloning identified a 5.7-kb region of BamHI fragment 4 that encodes functions required for the degradation of MOP. DNA sequence analysis revealed seven putative genes in this region: mocD (moc for mannityl opine catabolism) and mocE, oriented from right to left, and mocRCBAS, oriented from left to right. Significant identities exist at the nucleotide and derived amino acid sequence levels between these moc genes and the mas genes that are responsible for opine biosynthesis in crown gall tumors. MocD is a homolog of Mas2, the anabolic conjugase encoded by mas2'. MocE and MocC are related to the amino half and the carboxyl half, respectively, of Mas1 (MOP reductase), the second enzyme for MOP biosynthesis. These results indicate that the moc and mas genes evolved from a common origin. MocR and MocS are related to each other and to a putative repressor for the AGR degradation system encoded by the rhizogenic plasmid pRiA4. MocB and MocA are homologs of 6-phosphogluconate dehydratase and glucose-6-phosphate dehydrogenase, respectively. Mutations in mocD and mocE, but not mocC, are suppressed by functions encoded by the chromosome or the 450-kb megaplasmid present in many Agrobacterium isolates. We propose that moc genes derived from genes located elsewhere in the bacterial genome and that the tumor-expressed mas genes evolved from the bacterial moc genes.

Ti plasmid-encoded genes responsible for catabolism of the crown gall opine mannopine by Agrobacterium tumefaciens are homologs of the T-region genes responsible for synthesis of this opine by the plant tumor.

PubMed Central

Kim, K S; Farrand, S K

1996-01-01

Agrobacterium tumefaciens NT1 harboring pSaB4, which contains the 14-kb BamHI fragment 4 from the octopine/mannityl opine-type Ti plasmid pTi15955, grew well with agropine (AGR) but slowly with mannopine (MOP) as the sole carbon source. When a second plasmid encoding a dedicated transport system for MOP was introduced, these cells grew well with both AGR and MOP. Transposon insertion mutagenesis and subcloning identified a 5.7-kb region of BamHI fragment 4 that encodes functions required for the degradation of MOP. DNA sequence analysis revealed seven putative genes in this region: mocD (moc for mannityl opine catabolism) and mocE, oriented from right to left, and mocRCBAS, oriented from left to right. Significant identities exist at the nucleotide and derived amino acid sequence levels between these moc genes and the mas genes that are responsible for opine biosynthesis in crown gall tumors. MocD is a homolog of Mas2, the anabolic conjugase encoded by mas2'. MocE and MocC are related to the amino half and the carboxyl half, respectively, of Mas1 (MOP reductase), the second enzyme for MOP biosynthesis. These results indicate that the moc and mas genes evolved from a common origin. MocR and MocS are related to each other and to a putative repressor for the AGR degradation system encoded by the rhizogenic plasmid pRiA4. MocB and MocA are homologs of 6-phosphogluconate dehydratase and glucose-6-phosphate dehydrogenase, respectively. Mutations in mocD and mocE, but not mocC, are suppressed by functions encoded by the chromosome or the 450-kb megaplasmid present in many Agrobacterium isolates. We propose that moc genes derived from genes located elsewhere in the bacterial genome and that the tumor-expressed mas genes evolved from the bacterial moc genes. PMID:8655509

Epstein–Barr virus latent genes

PubMed Central

Kang, Myung-Soo; Kieff, Elliott

2015-01-01

Latent Epstein–Barr virus (EBV) infection has a substantial role in causing many human disorders. The persistence of these viral genomes in all malignant cells, yet with the expression of limited latent genes, is consistent with the notion that EBV latent genes are important for malignant cell growth. While the EBV-encoded nuclear antigen-1 (EBNA-1) and latent membrane protein-2A (LMP-2A) are critical, the EBNA-leader proteins, EBNA-2, EBNA-3A, EBNA-3C and LMP-1, are individually essential for in vitro transformation of primary B cells to lymphoblastoid cell lines. EBV-encoded RNAs and EBNA-3Bs are dispensable. In this review, the roles of EBV latent genes are summarized. PMID:25613728

DNA Sequence Analysis of a Complementary DNA for Cold-Regulated Arabidopsis Gene cor15 and Characterization of the COR 15 Polypeptide 1

PubMed Central

Lin, Chentao; Thomashow, Michael F.

1992-01-01

Previous studies have indicated that changes in gene expression occur in Arabidopsis thaliana L. (Heyn) during cold acclimation and that certain of the cor (cold-regulated) genes encode polypeptides that share the unusual property of remaining soluble upon boiling in aqueous solution. Here, we identify a cDNA clone for a cold-regulated gene encoding one of the “boiling-stable” polypeptides, COR15. DNA sequence analysis indicated that the gene, designated cor15, encodes a 14.7-kilodalton hydrophilic polypeptide having an N-terminal amino acid sequence that closely resembles transit peptides that target proteins to the stromal compartment of chloroplasts. Immunological studies indicated that COR15 is processed in vivo and that the mature polypeptide, COR 15m, is present in the soluble fraction of chloroplasts. Possible functions of COR 15m are discussed. ImagesFigure 1Figure 4Figure 5Figure 6Figure 7 PMID:16668917

Effect of long-term actual spaceflight on the expression of key genes encoding serotonin and dopamine system

NASA Astrophysics Data System (ADS)

Popova, Nina; Shenkman, Boris; Naumenko, Vladimir; Kulikov, Alexander; Kondaurova, Elena; Tsybko, Anton; Kulikova, Elisabeth; Krasnov, I. B.; Bazhenova, Ekaterina; Sinyakova, Nadezhda

The effect of long-term spaceflight on the central nervous system represents important but yet undeveloped problem. The aim of our work was to study the effect of 30-days spaceflight of mice on Russian biosatellite BION-M1 on the expression in the brain regions of key genes of a) serotonin (5-HT) system (main enzymes in 5-HT metabolism - tryptophan hydroxylase-2 (TPH-2), monoamine oxydase A (MAO A), 5-HT1A, 5-HT2A and 5-HT3 receptors); b) pivotal enzymes in DA metabolism (tyrosine hydroxylase, COMT, MAO A, MAO B) and D1, D2 receptors. Decreased expression of genes encoding the 5-HT catabolism (MAO A) and 5-HT2A receptor in some brain regions was shown. There were no differences between “spaceflight” and control mice in the expression of TPH-2 and 5-HT1A, 5-HT3 receptor genes. Significant changes were found in genetic control of DA system. Long-term spaceflight decreased the expression of genes encoding the enzyme in DA synthesis (tyrosine hydroxylase in s.nigra), DA metabolism (MAO B in the midbrain and COMT in the striatum), and D1 receptor in hypothalamus. These data suggested that 1) microgravity affected genetic control of 5-HT and especially the nigrostriatal DA system implicated in the central regulation of muscular tonus and movement, 2) the decrease in the expression of genes encoding key enzyme in DA synthesis, DA degradation and D1 receptor contributes to the movement impairment and dyskinesia produced by the spaceflight. The study was supported by Russian Foundation for Basic Research grant No. 14-04-00173.

Prevalence and Characterization of Carbapenem-Resistant Enterobacteriaceae Isolated from Mulago National Referral Hospital, Uganda

PubMed Central

Okoche, Deogratius; Asiimwe, Benon B.; Katabazi, Fred Ashaba; Kato, Laban; Najjuka, Christine F.

2015-01-01

Introduction Carbapenemases have increasingly been reported in enterobacteriaceae worldwide. Most carbapenemases are plasmid encoded hence resistance can easily spread. Carbapenem-resistant enterobacteriaceae are reported to cause mortality in up to 50% of patients who acquire bloodstream infections. We set out to determine the burden of carbapenem resistance as well as establish genes encoding for carbapenemases in enterobacteriaceae clinical isolates obtained from Mulago National Referral Hospital, Uganda. Methods This was a cross-sectional study with a total of 196 clinical isolates previously collected from pus swabs, urine, blood, sputum, tracheal aspirates, cervical swabs, endomentrial aspirates, rectal swabs, Vaginal swabs, ear swabs, products of conception, wound biopsy and amniotic fluid. All isolates were subjected to phenotypic carbapenemase screening using Boronic acid-based inhibition, Modified Hodge and EDTA double combined disk test. In addition, all the isolates were subjected to PCR assay to confirm presence of carbapenemase encoding genes. Results The study found carbapenemase prevalence of 22.4% (44/196) in the isolates using phenotypic tests, with the genotypic prevalence slightly higher at 28.6% (56/196). Over all, the most prevalent gene was blaVIM (21,10.7%), followed by blaOXA-48 (19, 9.7%), blaIMP (12, 6.1%), blaKPC (10, 5.1%) and blaNDM-1 (5, 2.6%). Among 56 isolates positive for 67 carbapenemase encoding genes, Klebsiella pneumonia was the species with the highest number (52.2%). Most 32/67(47.7%) of these resistance genes were in bacteria isolated from pus swabs. Conclusion There is a high prevalence of carbapenemases and carbapenem-resistance encoding genes among third generation cephalosporins resistant Enterobacteriaceae in Uganda, indicating a danger of limited treatment options in this setting in the near future. PMID:26284519

Mutations in a gene encoding the. cap alpha. subunit of a Saccharomyces cerevisiae G protein indicate a role in mating pheromone signaling

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

Jahng, K.Y.; Ferguson, J.; Reed, S.I.

1988-06-01

Mutations which allowed conjugation by Saccharomyces cerevisiae cells lacking a mating pheromone receptor gene were selected. One of the genes defined by such mutations was isolated from a yeast genomic library by complementation of a temperature-sensitive mutation and is identically to the gene GPA1 (also known as SCG1), recently shown to be highly homologous to gene encoding the ..cap alpha.. subunits of mammalian G proteins. Physiological analysis of temperature-sensitive gpal mutations suggests that the encoded G protein is involved in signaling in response to mating pheromones. Mutational disruption of G-protein activity causes cell-cycle arrest in G/sub 1/, deposition of mating-specificmore » cell surface aggultinins, and induction of pheromone-specific mRNa, all of which are responses to pheromone in wild-type cells. In addition, mutants can conjugate without the benefit of mating pheromone or pheromone receptor. A model is presented where the activated G protein has a negative impact on a constitutive signal which normally keeps the pheromone response repressed.« less

Role of protein farnesylation events in the ABA-mediated regulation of the Pinoresinol-Lariciresinol Reductase 1 (LuPLR1) gene expression and lignan biosynthesis in flax (Linum usitatissimum L.).

PubMed

Corbin, Cyrielle; Decourtil, Cédric; Marosevic, Djurdjica; Bailly, Marlène; Lopez, Tatiana; Renouard, Sullivan; Doussot, Joël; Dutilleul, Christelle; Auguin, Daniel; Giglioli-Guivarc'h, Nathalie; Lainé, Eric; Lamblin, Frédéric; Hano, Christophe

2013-11-01

A Linum usitatissimum LuERA1 gene encoding a putative ortholog of the ERA1 (Enhanced Response to ABA 1) gene of Arabidopsis thaliana (encoding the beta subunit of a farnesyltransferase) was analyzed in silico and for its expression in flax. The gene and the protein sequences are highly similar to other sequences already characterized in plants and all the features of a farnesyltransferase were detected. Molecular modeling of LuERA1 protein confirmed its farnesyltransferase nature. LuERA1 is expressed in the vegetative organs and also in the outer seedcoat of the flaxseed, where it could modulate the previously observed regulation operated by ABA on lignan synthesis. This effect could be mediated by the regulation of the transcription of a key gene for lignan synthesis in flax, the LuPLR1 gene, encoding a pinoresinol lariciresinol reductase. The positive effect of manumycin A, a specific inhibitor of farnesyltransferase, on lignan biosynthesis in flax cell suspension systems supports the hypothesis of the involvement of such an enzyme in the negative regulation of ABA action. In Arabidopsis, ERA1 is able to negatively regulate the ABA effects and the mutant era1 has an enhanced sensitivity to ABA. When expressed in an Arabidopsis cell suspension (heterologous system) LuERA1 is able to reverse the effect of the era1 mutation. RNAi experiments in flax targeting the farnesyltransferase β-subunit encoded by the LuERA1 gene led to an increase LuPLR1 expression level associated with an increased content of lignan in transgenic calli. Altogether these results strongly suggest a role of the product of this LuERA1 gene in the ABA-mediated upregulation of lignan biosynthesis in flax cells through the activation of LuPLR1 promoter. This ABA signaling pathway involving ERA1 probably acts through the ABRE box found in the promoter sequence of LuPLR1, a key gene for lignan synthesis in flax, as demonstrated by LuPLR1 gene promoter-reporter experiments in flax cells using wild type and mutated promoter sequences. Copyright © 2013 Elsevier Masson SAS. All rights reserved.

CD8 T cell response and evolutionary pressure to HIV-1 cryptic epitopes derived from antisense transcription

PubMed Central

Carlson, Jonathan; Yan, Jiyu; Akinsiku, Olusimidele T.; Schaefer, Malinda; Sabbaj, Steffanie; Bet, Anne; Levy, David N.; Heath, Sonya; Tang, Jianming; Kaslow, Richard A.; Walker, Bruce D.; Ndung’u, Thumbi; Goulder, Philip J.; Heckerman, David; Hunter, Eric; Goepfert, Paul A.

2010-01-01

Retroviruses pack multiple genes into relatively small genomes by encoding several genes in the same genomic region with overlapping reading frames. Both sense and antisense HIV-1 transcripts contain open reading frames for known functional proteins as well as numerous alternative reading frames (ARFs). At least some ARFs have the potential to encode proteins of unknown function, and their antigenic properties can be considered as cryptic epitopes (CEs). To examine the extent of active immune response to virally encoded CEs, we analyzed human leukocyte antigen class I–associated polymorphisms in HIV-1 gag, pol, and nef genes from a large cohort of South Africans with chronic infection. In all, 391 CEs and 168 conventional epitopes were predicted, with the majority (307; 79%) of CEs derived from antisense transcripts. In further evaluation of CD8 T cell responses to a subset of the predicted CEs in patients with primary or chronic infection, both sense- and antisense-encoded CEs were immunogenic at both stages of infection. In addition, CEs often mutated during the first year of infection, which was consistent with immune selection for escape variants. These findings indicate that the HIV-1 genome might encode and deploy a large potential repertoire of unconventional epitopes to enhance vaccine-induced antiviral immunity. PMID:20065064

Molecular cloning and nucleotide sequences of the genes for two essential proteins constituting a novel enzyme system for heptaprenyl diphosphate synthesis.

PubMed

Koike-Takeshita, A; Koyama, T; Obata, S; Ogura, K

1995-08-04

The genes encoding two dissociable components essential for Bacillus stearothermophilus heptaprenyl diphosphate synthase (all-trans-hexparenyl-diphosphate:isopentenyl-diphosphate hexaprenyl-trans-transferase, EC 2.5.1.30) were cloned, and their nucleotide sequences were determined. Sequence analyses revealed the presence of three open reading frames within 2,350 base pairs, designated as ORF-1, ORF-2, and ORF-3 in order of nucleotide sequence, which encode proteins of 220, 234, and 323 amino acids, respectively. Deletion experiments have shown that expression of the enzymatic activity requires the presence of ORF-1 and ORF-3, but ORF-2 is not essential. As a result, this enzyme was proved genetically to consist of two different protein compounds with molecular masses of 25 kDa (Component I) and 36 kDa (Component II), encoded by two of the three tandem genes. The protein encoded by ORF-1 has no similarity to any protein so far registered. However, the protein encoded by ORF-3 shows a 32% similarity to the farnesyl diphosphate synthase of the same bacterium and has seven highly conserved regions that have been shown typical in prenyltransferases (Koyama, T., Obata, S., Osabe, M., Takeshita, A., Yokoyama, K., Uchida, M., Nishino, T., and Ogura, K. (1993) J. Biochem. (Tokyo) 113, 355-363).

Molecular cloning of low-temperature-inducible ribosomal proteins from soybean.

PubMed

Kim, Kee-Young; Park, Seong-Whan; Chung, Young-Soo; Chung, Chung-Han; Kim, Jung-In; Lee, Jai-Heon

2004-05-01

Three ribosomal protein genes induced by low-temperature treatment were isolated from soybean. GmRPS13 (742 bp) encodes a 17.1 kDa protein which has 95% identity with the 40S ribosomal protein S13 of Panax ginseng (AB043974). GmRPS6 (925 bp) encodes a 28.1 kDa protein which has 94% identity with the 40S ribosomal protein S6 of Asparagus officinalis (AJ277533). GmRPL37 (494 bp) encodes a 10.7 kDa protein which has 85% identity with the 60S ribosomal protein L37 of Arabidopsis thaliana (AF370216). The expression of these ribosomal protein genes started to increase 3 d after low-temperature treatment, whereas the cold-stress protein src1 was highly induced from the first day. Such late response of these ribosomal protein genes may be due to secondary signals during cold adaptation. The induction of ribosomal protein genes might enhance the translation process or help proper ribosome functioning under low-temperature conditions.

Another face of the Treacher Collins syndrome (TCOF1) gene: identification of additional exons.

PubMed

So, Rolando B; Gonzales, Bianca; Henning, Dale; Dixon, Jill; Dixon, Michael J; Valdez, Benigno C

2004-03-17

Treacher Collins syndrome (TCS) is characterized by an abnormality in craniofacial development during early embryogenesis. TCS is caused by mutations in the gene TCOF1, which encodes the nucleolar phosphoprotein treacle. Genetic and proteomic characterizations of TCS/treacle are based on the previously reported 26 exons of TCOF1. Here, we report the identification of 231-nucleotide (nt) exon 6A (between exons 6 and 7) and 108-nt exon 16A (between exons 16 and 17). Isoforms with exon 6A are up to 3.7-fold more abundant than alternatively spliced variants without exon 6A, but only minor isoforms contain exon 16A. Exon 6A encodes a peptide sequence containing basic and acidic domains similar to 10 other exons of TCOF1. Unlike the other exons, exon 6A encodes a nuclear localization signal (NLS) which does not, however, alter the nucleolar localization of full-length treacle. The discovery of exons 6A and 16A is relevant to mutational analysis of the TCOF1 gene in TCS patients, and to functional analysis of its gene product.

Metabolic engineering and adaptive evolution for efficient production of D-lactic acid in Saccharomyces cerevisiae.

PubMed

Baek, Seung-Ho; Kwon, Eunice Y; Kim, Yong Hwan; Hahn, Ji-Sook

2016-03-01

There is an increasing demand for microbial production of lactic acid (LA) as a monomer of biodegradable poly lactic acid (PLA). Both optical isomers, D-LA and L-LA, are required to produce stereocomplex PLA with improved properties. In this study, we developed Saccharomyces cerevisiae strains for efficient production of D-LA. D-LA production was achieved by expressing highly stereospecific D-lactate dehydrogenase gene (ldhA, LEUM_1756) from Leuconostoc mesenteroides subsp. mesenteroides ATCC 8293 in S. cerevisiae lacking natural LA production activity. D-LA consumption after glucose depletion was inhibited by deleting DLD1 encoding D-lactate dehydrogenase and JEN1 encoding monocarboxylate transporter. In addition, ethanol production was reduced by deleting PDC1 and ADH1 genes encoding major pyruvate decarboxylase and alcohol dehydrogenase, respectively, and glycerol production was eliminated by deleting GPD1 and GPD2 genes encoding glycerol-3-phosphate dehydrogenase. LA tolerance of the engineered D-LA-producing strain was enhanced by adaptive evolution and overexpression of HAA1 encoding a transcriptional activator involved in weak acid stress response, resulting in effective D-LA production up to 48.9 g/L without neutralization. In a flask fed-batch fermentation under neutralizing condition, our evolved strain produced 112.0 g/L D-LA with a yield of 0.80 g/g glucose and a productivity of 2.2 g/(L · h).

Tanscriptomic Study of the Soybean-Fusarium virguliforme Interaction Revealed a Novel Ankyrin-Repeat Containing Defense Gene, Expression of Whose during Infection Led to Enhanced Resistance to the Fungal Pathogen in Transgenic Soybean Plants

PubMed Central

Ngaki, Micheline N.; Wang, Bing; Sahu, Binod B.; Srivastava, Subodh K.; Farooqi, Mohammad S.; Kambakam, Sekhar; Swaminathan, Sivakumar

2016-01-01

Fusarium virguliforme causes the serious disease sudden death syndrome (SDS) in soybean. Host resistance to this pathogen is partial and is encoded by a large number of quantitative trait loci, each conditioning small effects. Breeding SDS resistance is therefore challenging and identification of single-gene encoded novel resistance mechanisms is becoming a priority to fight this devastating this fungal pathogen. In this transcriptomic study we identified a few putative soybean defense genes, expression of which is suppressed during F. virguliforme infection. The F. virguliforme infection-suppressed genes were broadly classified into four major classes. The steady state transcript levels of many of these genes were suppressed to undetectable levels immediately following F. virguliforme infection. One of these classes contains two novel genes encoding ankyrin repeat-containing proteins. Expression of one of these genes, GmARP1, during F. virguliforme infection enhances SDS resistance among the transgenic soybean plants. Our data suggest that GmARP1 is a novel defense gene and the pathogen presumably suppress its expression to establish compatible interaction. PMID:27760122

White collar-1, a central regulator of blue light responses in Neurospora, is a zinc finger protein.

PubMed Central

Ballario, P; Vittorioso, P; Magrelli, A; Talora, C; Cabibbo, A; Macino, G

1996-01-01

The Neurospora crassa blind mutant white collar-1 (wc-1) is pleiotropically defective in all blue light-induced phenomena, establishing a role for the wc-1 gene product in the signal transduction pathway. We report the cloning of the wc-1 gene isolated by chromosome walking and mutant complementation. The elucidation of the wc-1 gene product provides a key piece of the blue light signal transduction puzzle. The wc-1 gene encodes a 125 kDa protein whose encoded motifs include a single class four, zinc finger DNA binding domain and a glutamine-rich putative transcription activation domain. We demonstrate that the wc-1 zinc finger domain, expressed in Escherichia coli, is able to bind specifically to the promoter of a blue light-regulated gene of Neurospora using an in vitro gel retardation assay. Furthermore, we show that wc-1 gene expression is autoregulated and is transcriptionally induced by blue light irradiation. Images PMID:8612589

Mutations in extracellular matrix genes NID1 and LAMC1 cause autosomal dominant Dandy-Walker malformation and occipital cephaloceles

PubMed Central

Darbro, Benjamin W.; Mahajan, Vinit B.; Gakhar, Lokesh; Skeie, Jessica M.; Campbell, Elizabeth; Wu, Shu; Bing, Xinyu; Millen, Kathleen J.; Dobyns, William B.; Kessler, John A.; Jalali, Ali; Cremer, James; Segre, Alberto; Manak, J. Robert; Aldinger, Kimerbly A.; Suzuki, Satoshi; Natsume, Nagato; Ono, Maya; Hai, Huynh Dai; Viet, Le Thi; Loddo, Sara; Valente, Enza M.; Bernardini, Laura; Ghonge, Nitin; Ferguson, Polly J.; Bassuk, Alexander G.

2013-01-01

We performed whole-exome sequencing of a family with autosomal dominant Dandy-Walker malformation and occipital cephaloceles (ADDWOC) and detected a mutation in the extracellular matrix protein encoding gene NID1. In a second family, protein interaction network analysis identified a mutation in LAMC1, which encodes a NID1 binding partner. Structural modeling the NID1-LAMC1 complex demonstrated that each mutation disrupts the interaction. These findings implicate the extracellular matrix in the pathogenesis of Dandy-Walker spectrum disorders. PMID:23674478

Plasmid-dependent methylotrophy in thermotolerant Bacillus methanolicus.

PubMed

Brautaset, Trygve; Jakobsen M, Øyvind M; Flickinger, Michael C; Valla, Svein; Ellingsen, Trond E

2004-03-01

Bacillus methanolicus can efficiently utilize methanol as a sole carbon source and has an optimum growth temperature of 50 degrees C. With the exception of mannitol, no sugars have been reported to support rapid growth of this organism, which is classified as a restrictive methylotroph. Here we describe the DNA sequence and characterization of a 19,167-bp circular plasmid, designated pBM19, isolated from B. methanolicus MGA3. Sequence analysis of pBM19 demonstrated the presence of the methanol dehydrogenase gene, mdh, which is crucial for methanol consumption in this bacterium. In addition, five genes (pfk, encoding phosphofructokinase; rpe, encoding ribulose-5-phosphate 3-epimerase; tkt, encoding transketolase; glpX, encoding fructose-1,6-bisphosphatase; and fba, encoding fructose-1,6-bisphosphate aldolase) with deduced roles in methanol assimilation via the ribulose monophosphate pathway are encoded by pBM19. A shuttle vector, pTB1.9, harboring the pBM19 minimal replicon (repB and ori) was constructed and used to transform MGA3. Analysis of the resulting recombinant strain demonstrated that it was cured of pBM19 and was not able to grow on methanol. A pTB1.9 derivative harboring the complete mdh gene could not restore growth on methanol when it was introduced into the pBM19-cured strain, suggesting that additional pBM19 genes are required for consumption of this carbon source. Screening of 13 thermotolerant B. methanolicus wild-type strains showed that they all harbor plasmids similar to pBM19, and this is the first report describing plasmid-linked methylotrophy in any microorganism. Our findings should have an effect on future genetic manipulations of this organism, and they contribute to a new understanding of the biology of methylotrophs.

Plasmid-Dependent Methylotrophy in Thermotolerant Bacillus methanolicus

PubMed Central

Brautaset, Trygve; Jakobsen, Øyvind M.; Flickinger, Michael C.; Valla, Svein; Ellingsen, Trond E.

2004-01-01

Bacillus methanolicus can efficiently utilize methanol as a sole carbon source and has an optimum growth temperature of 50°C. With the exception of mannitol, no sugars have been reported to support rapid growth of this organism, which is classified as a restrictive methylotroph. Here we describe the DNA sequence and characterization of a 19,167-bp circular plasmid, designated pBM19, isolated from B. methanolicus MGA3. Sequence analysis of pBM19 demonstrated the presence of the methanol dehydrogenase gene, mdh, which is crucial for methanol consumption in this bacterium. In addition, five genes (pfk, encoding phosphofructokinase; rpe, encoding ribulose-5-phosphate 3-epimerase; tkt, encoding transketolase; glpX, encoding fructose-1,6-bisphosphatase; and fba, encoding fructose-1,6-bisphosphate aldolase) with deduced roles in methanol assimilation via the ribulose monophosphate pathway are encoded by pBM19. A shuttle vector, pTB1.9, harboring the pBM19 minimal replicon (repB and ori) was constructed and used to transform MGA3. Analysis of the resulting recombinant strain demonstrated that it was cured of pBM19 and was not able to grow on methanol. A pTB1.9 derivative harboring the complete mdh gene could not restore growth on methanol when it was introduced into the pBM19-cured strain, suggesting that additional pBM19 genes are required for consumption of this carbon source. Screening of 13 thermotolerant B. methanolicus wild-type strains showed that they all harbor plasmids similar to pBM19, and this is the first report describing plasmid-linked methylotrophy in any microorganism. Our findings should have an effect on future genetic manipulations of this organism, and they contribute to a new understanding of the biology of methylotrophs. PMID:14973041

Effect of hypoxia on the expression of nuclear genes encoding mitochondrial proteins in U87 glioma cells.

PubMed

Minchenko, O H; Riabovol, O O; Tsymbal, D O; Minchenko, D O; Ratushna, O O

2016-01-01

We have studied the effect of hypoxia on the expression of nuclear genes encoding mitochondrial proteins in U87 glioma cells under the inhibition of IRE1 (inositol requiring enzyme-1), which controls cell proliferation and tumor growth as a central mediator of endoplasmic reticulum stress. It was shown that hypoxia down-regulated gene expression of malate dehydrogenase 2 (MDH2), malic enzyme 2 (ME2), mitochondrial aspartate aminotransferase (GOT2), and subunit B of succinate dehydrogenase (SDHB) in control (transfected by empty vector) glioma cells in a gene specific manner. At the same time, the expression level of mitochondrial NADP+-dependent isocitrate dehydrogenase 2 (IDH2) and subunit D of succinate dehydrogenase (SDHD) genes in these cells does not significantly change in hypoxic conditions. It was also shown that the inhibition of ІRE1 signaling enzyme function in U87 glioma cells decreases the effect of hypoxia on the expression of ME2, GOT2, and SDHB genes and introduces the sensitivity of IDH2 gene to hypoxia. Furthermore, the expression of all studied genes depends on IRE1-mediated endoplasmic reticulum stress signaling in gene specific manner, because ІRE1 knockdown significantly decreases their expression in normoxic conditions, except for IDH2 gene, which expression level is strongly up-regulated. Therefore, changes in the expression level of nuclear genes encoding ME2, MDH2, IDH2, SDHB, SDHD, and GOT2 proteins possibly reflect metabolic reprogramming of mitochondria by hypoxia and IRE1-mediated endoplasmic reticulum stress signaling and correlate with suppression of glioma cell proliferation under inhibition of the IRE1 enzyme function.

Cloning of the canine gene encoding transcription factor Pit-1 and its exclusion as candidate gene in a canine model of pituitary dwarfism.

PubMed

Lantinga-van Leeuwen, I S; Mol, J A; Kooistra, H S; Rijnberk, A; Breen, M; Renier, C; van Oost, B A

2000-01-01

Combined pituitary hormone deficiency (CPHD) is an autosomal recessive inherited disease of German shepherd dogs characterized primarily by dwarfism. In mice and humans a similar genetic disorder has been described that results from an alteration in the gene encoding the transcription factor Pit-1. In this study we characterized the canine Pit-1 gene, determined the chromosomal localization of the Pit-1 gene, and screened dwarf German shepherd dogs for the presence of mutations in this gene. The full-length canine Pit-1 cDNA contained an open reading frame encoding 291 amino acids, 92 bp of 5'-untranslated region, and 1959 bp of 3'-untranslated region. The deduced amino acid sequence was highly homologous with Pit-1 of other mammalian species. Using a Pit-1 BAC clone as probe, the Pit-1 gene was mapped by FISH to canine Chromosome (Chr) 31. In dwarf German shepherd dogs a C to A transversion was detected, causing a Phe (TTC) to Leu (TTA) substitution at codon 81. This alteration was present neither in other canine breeds analyzed nor in other mammalian species. However, healthy German shepherd dogs were also homozygous for the mutant allele, indicating that it is not the primary disease-causing mutation. In addition, linkage analysis of polymorphic DNA markers flanking the Pit-1 gene, 41K19 and 52L05, revealed no co-segregation between the Pit-1 locus and the CPHD phenotype. These findings suggest that a gene other than Pit-1 is responsible for the pituitary anomaly in dwarf German shepherd dogs.

Multiple conversion between the genes encoding bacterial class-I release factors

PubMed Central

Ishikawa, Sohta A.; Kamikawa, Ryoma; Inagaki, Yuji

2015-01-01

Bacteria require two class-I release factors, RF1 and RF2, that recognize stop codons and promote peptide release from the ribosome. RF1 and RF2 were most likely established through gene duplication followed by altering their stop codon specificities in the common ancestor of extant bacteria. This scenario expects that the two RF gene families have taken independent evolutionary trajectories after the ancestral gene duplication event. However, we here report two independent cases of conversion between RF1 and RF2 genes (RF1-RF2 gene conversion), which were severely examined by procedures incorporating the maximum-likelihood phylogenetic method. In both cases, RF1-RF2 gene conversion was predicted to occur in the region encoding nearly entire domain 3, of which functions are common between RF paralogues. Nevertheless, the ‘direction’ of gene conversion appeared to be opposite from one another—from RF2 gene to RF1 gene in one case, while from RF1 gene to RF2 gene in the other. The two cases of RF1-RF2 gene conversion prompt us to propose two novel aspects in the evolution of bacterial class-I release factors: (i) domain 3 is interchangeable between RF paralogues, and (ii) RF1-RF2 gene conversion have occurred frequently in bacterial genome evolution. PMID:26257102

Influence of culture conditions and preconditioning on survival of Lactobacillus delbrueckii subspecies bulgaricus ND02 during lyophilization.

PubMed

Shao, Yuyu; Gao, Shuran; Guo, Huiling; Zhang, Heping

2014-03-01

The cryotolerance of Lactobacillus delbrueckii ssp. bulgaricus is weak during vacuum freeze-drying. Many factors affect cryoresistance of these bacteria, such as cryoprotectant composition, the lyophilization technology used, and the intrinsic characteristics of the bacteria. In this research, we explored the fermentation technology and other preconditioning treatments of cells in improving the cryoresistance of Lactobacillus delbrueckii ssp. bulgaricus strains during lyophilization. The addition of yeast extract in the propagation medium exerted a negative effect on the cryotolerance of these bacteria and decreased survival during lyophilization. The count of the freeze-dried cells from medium containing a high level (4%) of yeast extract was only 4.1 × 10(9) cfu/g, indicating a death rate as high as 88%, compared with the culture medium without yeast extract, with a lower death rate of 44.7%. When Lactobacillus delbrueckii ssp. bulgaricus ND02 was propagated in yeast extract-free de Man, Rogosa, and Sharpe broth at a set pH value of 5.1, the cells showed unexpectedly higher survival after freeze-drying. Viable counts of the lyophilized cell of strain ND02 cultivated at pH 5.1 could reach 1.05 × 10(11)cfu/g and survival of the freeze-drying process was 68.3%, whereas at pH 5.7, survival was only 51.2%. We also examined the effects of pretreatment of cells on survival of the bacteria after vacuum freeze-drying. By analyzing the effect of pretreatment conditions on the expression of cold- and heat-shock genes, we established 2 pretreatments that improved survival of cells after lyophilization. Optimal fermentation conditions and pretreatment of the cell-cryoprotectant mixture at 10°C for 2h or 37°C for 30 min improved the cryoresistance of 4 strains of Lactobacillus delbrueckii ssp. bulgaricus to varying degrees. Cells of IMAU20269 and IMAU20291 that were pretreated showed enhanced survival of 16.06 and 16.82%, respectively, after lyophilization. Expression of cold- and heat-shock genes for pretreated strains ND02, IMAU80423, IMAU20269, and IMAU20291 was analyzed by using quantitative PCR. From the expression of 2 cold shock-induced genes (cspA and cspB) and 6 heat shock-induced genes (groES, hsp, hsp20, hsp40, hsp60, and hsp70), strain ND02 showed a higher relative quantity of gene expression and displayed superior resistance to cold-induced stress during the freeze-drying process. Copyright © 2014 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Complete mitochondrial genome of the monogonont rotifer, Brachionus koreanus (Rotifera, Brachionidae).

PubMed

Hwang, Dae-Sik; Suga, Koushirou; Sakakura, Yoshitaka; Park, Heum Gi; Hagiwara, Atsushi; Rhee, Jae-Sung; Lee, Jae-Seong

2014-02-01

The complete mitochondrial genome was obtained from the assembled genome data sequenced by next generation sequencing (NGS) technology from the monogonont rotifer Brachionus koreanus. The mitochondrial genome of B. koreanus was composed of two circular chromosomes designated as mtDNA-I (10,421 bp) and mtDNA-II (11,923 bp). The gene contents of B. koreanus were identical with previously reported B. plicatilis mitochondrial genomes. However, gene orders of B. koreanus showed one rearrangement between the two species. Of 12 protein-coding genes (PCGs), 3 genes (ATP6, ND1, and ND3) had an incomplete stop codon. The A + T base composition of B. koreanus mitochondrial genome was high (68.81%). They also showed anti-G bias (12.03% and 10.97%) on the second and third position of PCGs as well as slight anti-C bias (15.96% and 14.31%) on the first and third position of PCGs.

Mitochondrial gene arrangement of the horseshoe crab Limulus polyphemus L.: conservation of major features among arthropod classes

NASA Technical Reports Server (NTRS)

Staton, J. L.; Daehler, L. L.; Brown, W. M.; Jacobs, D. K. (Principal Investigator)

1997-01-01

Numerous complete mitochondrial DNA sequences have been determined for species within two arthropod groups, insects and crustaceans, but there are none for a third, the chelicerates. Most mitochondrial gene arrangements reported for crustaceans and insect species are identical or nearly identical to that of Drosophila yakuba. Sequences across 36 of the gene boundaries in the mitochondrial DNA (mtDNA) of a representative chelicerate. Limulus polyphemus L., also reveal an arrangement like that of Drosophila yakuba. Only the position of the tRNA(LEU)(UUR) gene differs; in Limulus it is between the genes for tRNA(LEU)(CUN) and ND1. This positioning is also found in onychophorans, mollusks, and annelids, but not in insects and crustaceans, and indicates that tRNA(LEU)(CUN)-tRNA(LEU)(UUR)-ND1 was the ancestral gene arrangement for these groups, as suggested earlier. There are no differences in the relative arrangements of protein-coding and ribosomal RNA genes between Limulus and Drosophila, and none have been observed within arthropods. The high degree of similarity of mitochondrial gene arrangements within arthropods is striking, since some taxa last shared a common ancestor before the Cambrian, and contrasts with the extensive mtDNA rearrangements occasionally observed within some other metazoan phyla (e.g., mollusks and nematodes).

Gene-breaking: A new paradigm for human retrotransposon-mediated gene evolution

PubMed Central

Wheelan, Sarah J.; Aizawa, Yasunori; Han, Jeffrey S.; Boeke, Jef D.

2005-01-01

The L1 retrotransposon is the most highly successful autonomous retrotransposon in mammals. This prolific genome parasite may on occasion benefit its host through genome rearrangements or adjustments of host gene expression. In examining possible effects of L1 elements on host gene expression, we investigated whether a full-length L1 element inserted in the antisense orientation into an intron of a cellular gene may actually split the gene's transcript into two smaller transcripts: (1) a transcript containing the upstream exons and terminating in the major antisense polyadenylation site (MAPS) of the L1, and (2) a transcript derived from the L1 antisense promoter (ASP) that includes the downstream exons of the gene. Bioinformatic analysis and experimental follow-up provide evidence for this L1 “gene-breaking” hypothesis. We identified three human genes apparently “broken” by L1 elements, as well as 12 more candidate genes. Most of the inserted L1 elements in our 15 candidate genes predate the human/chimp divergence. If indeed split, the transcripts of these genes may in at least one case encode potentially interacting proteins, and in another case may encode novel proteins. Gene-breaking represents a new mechanism through which L1 elements remodel mammalian genomes. PMID:16024818

Identification of Fasciola flukes in Thailand based on their spermatogenesis and nuclear ribosomal DNA, and their intraspecific relationships based on mitochondrial DNA.

PubMed

Chaichanasak, Pannigan; Ichikawa, Madoka; Sobhon, Prasert; Itagaki, Tadashi

2012-12-01

We analyzed 147 Fasciola flukes obtained from cattle in Thailand based on their spermatogenetic ability, and nuclear ribosomal internal transcribed spacer 1 (ITS1) and mitochondrial nicotiamide adenine dinucleotide dehydrogenase subunit 1 (ND1) genes as molecular markers. One hundred twenty-eight flukes, which had abundant sperm in their seminal vesicles (spermic) and showed the PCR-RFLP pattern of F. gigantica in the ITS1, were accurately identified as F. gigantica. The other 19 flukes that had no sperm in their seminal vesicles were aspermic Fasciola sp. with the RFLP patterns identical to that of F. gigantica. Twenty-nine ND1 haplotypes (Fg-ND1-Thai 2-30) were distinguished in the 128 F. gigantica flukes and were divided into haplotypes unique to Thailand and those common to other countries, suggesting the possibility that ancestral haplotypes were introduced into Thailand. Three haplotypes (Fg-ND1-Thai 7, 9 and 27) appeared to be the major haplotypes found in F. gigantica from Thailand. Only one haplotype (Fg-ND1-Thai 1) was found in the 19 aspermic Fasciola sp. flukes obtained from geographical regions, and the nucleotide sequence of Fg-ND1-Thai 1 was identical to that of the aspermic Fasciola sp. from Japan, Korea, China, Vietnam and Myanmar, suggesting that they were descendants with a common provenance and expanded to these countries in the relatively recent past. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

Gene cloning and overexpression of two conjugated polyketone reductases, novel aldo-keto reductase family enzymes, of Candida parapsilosis.

PubMed

Kataoka, M; Delacruz-Hidalgo, A-R G; Akond, M A; Sakuradani, E; Kita, K; Shimizu, S

2004-04-01

The genes encoding two conjugated polyketone reductases (CPR-C1, CPR-C2) of Candida parapsilosis IFO 0708 were cloned and sequenced. The genes encoded a total of 304 and 307 amino acid residues for CPR-C1 and CPR-C2, respectively. The deduced amino acid sequences of the two enzymes showed high similarity to each other and to several proteins of the aldo-keto reductase (AKR) superfamily. However, several amino acid residues in putative active sites of AKRs were not conserved in CPR-C1 and CPR-C2. The two CPR genes were overexpressed in Escherichia coli. The E. coli transformant bearing the CPR-C2 gene almost stoichiometrically reduced 30 mg ketopantoyl lactone/ml to D-pantoyl lactone.

Prevalence of ESBL and MBL encoding genes in Acinetobacter baumannii strains isolated from patients of intensive care units (ICU)

PubMed Central

Safari, Marzieh; Mozaffari Nejad, Amir Sasan; Bahador, Abas; Jafari, Rasool; Alikhani, Mohammad Yousef

2015-01-01

The aim of this study was to investigate the prevalence of ESBL and MBL encoding genes among A. baumannii isolates. In this cross sectional study, 100 A. baumannii strains were isolated from ICU wards of 3 educational hospitals of Hamadan City, Iran in 2011. Phenotypic identification of the production of ESBLs and MBLs has been carried out by using E-test and DDST methods, respectively. PCR technique was used for amplification of the ESBL and MBL encoding genes, namely: CTX-M, SHV, TEM, OXA-51, VIM-Family, IMP-Family, SPM-1, SIM-1, and GIM-1. Eighty seven (87%), 95 (95%), 98 (98%) and 95 (95%) out of 100 A. baumannii isolates were resistant to imipenem, meropenem, ceftazidime and cefotaxime, respectively. Also, 99% and 7% of the isolates were MBLs and ESBLs produced phenotypically. Thirty (30%), 20 (20%) and 58 (58%) out of 100 A. baumannii isolates have been confirmed to harbor the blaVIM-family, TEM and SHV genes, respectively. Our results show no significant relationship between the detected gens with production of MBLs and ESBLs in spite of high prevalence of MBL encoding and drug resistant A. baumannii. Probably some other genes rather than what we studied are involved in phenotypic production of MBLs and ESBLs and subsequent drug resistance in Hamadan area, Iran. PMID:26150748

The 2p21 deletion syndrome: characterization of the transcription content.

PubMed

Parvari, Ruti; Gonen, Yael; Alshafee, Ismael; Buriakovsky, Sophia; Regev, Kfir; Hershkovitz, Eli

2005-08-01

The vast majority of small-deletion syndromes are caused by haploinsufficiency of one or several genes and are transmitted as dominant traits. We have previously identified a homozygous deletion of 179,311 bp on chromosome 2p21 as the cause of a unique syndrome, inherited in a recessive mode, consisting of cystinuria, neonatal seizures, hypotonia, severe somatic and developmental delay, facial dysmorphism, and reduced activity of all the respiratory chain enzymatic complexes that are encoded in the mitochondria. We now present the transcription content of this region: Multiple splicing variants of the genes protein phosphatase 1B (formerly 2C) magnesium-dependent, beta isoform (PPM1B), SLC3A1, and KIAA0436 (approved gene symbol PREPL) were identified and their patterns of expression analyzed. The spliced variants are predicted to have additional functions compared to the known variants and their patterns of expression fit the tissues affected by the syndrome. The first exon of an additional gene (C2orf34) is encoded in the deleted region and the gene is not expressed in the patients. In addition several transcripts with very short open reading frames are also encoded in the deletion. The identification of all transcripts encoded in the region deleted in the patients is the first step in the study of the genotype-phenotype correlation of the 2p21 patients.

Analysis and Manipulation of Aspartate Pathway Genes for l-Lysine Overproduction from Methanol by Bacillus methanolicus▿

PubMed Central

Nærdal, Ingemar; Netzer, Roman; Ellingsen, Trond E.; Brautaset, Trygve

2011-01-01

We investigated the regulation and roles of six aspartate pathway genes in l-lysine overproduction in Bacillus methanolicus: dapG, encoding aspartokinase I (AKI); lysC, encoding AKII; yclM, encoding AKIII; asd, encoding aspartate semialdehyde dehydrogenase; dapA, encoding dihydrodipicolinate synthase; and lysA, encoding meso-diaminopimelate decarboxylase. Analysis of the wild-type strain revealed that in vivo lysC transcription was repressed 5-fold by l-lysine and induced 2-fold by dl-methionine added to the growth medium. Surprisingly, yclM transcription was repressed 5-fold by dl-methionine, while the dapG, asd, dapA, and lysA genes were not significantly repressed by any of the aspartate pathway amino acids. We show that the l-lysine-overproducing classical B. methanolicus mutant NOA2#13A52-8A66 has—in addition to a hom-1 mutation—chromosomal mutations in the dapG coding region and in the lysA promoter region. No mutations were found in its dapA, lysC, asd, and yclM genes. The mutant dapG gene product had abolished feedback inhibition by meso-diaminopimelate in vitro, and the lysA mutation was accompanied by an elevated (6-fold) lysA transcription level in vivo. Moreover, yclM transcription was increased 16-fold in mutant strain NOA2#13A52-8A66 compared to the wild-type strain. Overexpression of wild-type and mutant aspartate pathway genes demonstrated that all six genes are important for l-lysine overproduction as tested in shake flasks, and the effects were dependent on the genetic background tested. Coupled overexpression of up to three genes resulted in additive (above 80-fold) increased l-lysine production levels. PMID:21724876

Analysis and manipulation of aspartate pathway genes for L-lysine overproduction from methanol by Bacillus methanolicus.

PubMed

Nærdal, Ingemar; Netzer, Roman; Ellingsen, Trond E; Brautaset, Trygve

2011-09-01

We investigated the regulation and roles of six aspartate pathway genes in L-lysine overproduction in Bacillus methanolicus: dapG, encoding aspartokinase I (AKI); lysC, encoding AKII; yclM, encoding AKIII; asd, encoding aspartate semialdehyde dehydrogenase; dapA, encoding dihydrodipicolinate synthase; and lysA, encoding meso-diaminopimelate decarboxylase. Analysis of the wild-type strain revealed that in vivo lysC transcription was repressed 5-fold by L-lysine and induced 2-fold by dl-methionine added to the growth medium. Surprisingly, yclM transcription was repressed 5-fold by dl-methionine, while the dapG, asd, dapA, and lysA genes were not significantly repressed by any of the aspartate pathway amino acids. We show that the L-lysine-overproducing classical B. methanolicus mutant NOA2#13A52-8A66 has-in addition to a hom-1 mutation-chromosomal mutations in the dapG coding region and in the lysA promoter region. No mutations were found in its dapA, lysC, asd, and yclM genes. The mutant dapG gene product had abolished feedback inhibition by meso-diaminopimelate in vitro, and the lysA mutation was accompanied by an elevated (6-fold) lysA transcription level in vivo. Moreover, yclM transcription was increased 16-fold in mutant strain NOA2#13A52-8A66 compared to the wild-type strain. Overexpression of wild-type and mutant aspartate pathway genes demonstrated that all six genes are important for L-lysine overproduction as tested in shake flasks, and the effects were dependent on the genetic background tested. Coupled overexpression of up to three genes resulted in additive (above 80-fold) increased L-lysine production levels.

'Laminopathies': A wide spectrum of human diseases

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

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

2007-06-10

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

FZD4S, a splicing variant of frizzled-4, encodes a soluble-type positive regulator of the WNT signaling pathway.

PubMed

Sagara, N; Kirikoshi, H; Terasaki, H; Yasuhiko, Y; Toda, G; Shiokawa, K; Katoh, M

2001-04-06

Frizzled-1 (FZD1)-FZD10 are seven-transmembrane-type WNT receptors, and SFRP1-SFRP5 are soluble-type WNT antagonists. These molecules are encoded by mutually distinct genes. We have previously isolated and characterized the 7.7-kb FZD4 mRNA, encoding a seven-transmembrane receptor with the extracellular cysteine-rich domain (CRD). Here, we have cloned and characterized FZD4S, a splicing variant of the FZD4 gene. FZD4S, corresponding to the 10.0-kb FZD4 mRNA, consisted of exon 1, intron 1, and exon 2 of the FZD4 gene. FZD4S encoded a soluble-type polypeptide with the N-terminal part of CRD, and was expressed in human fetal kidney. Injection of synthetic FZD4S mRNA into the ventral marginal zone of Xenopus embryos at the 4-cell stage did not induce axis duplication by itself, but augmented the axis duplication potential of coinjected Xwnt-8 mRNA. These results indicate that the FZD4 gene gives rise to soluble-type FZD4S as well as seven-transmembrane-type FZD4 due to alternative splicing, and strongly suggest that FZD4S plays a role as a positive regulator of the WNT signaling pathway. Copyright 2001 Academic Press.

Molecular basis of the allelic inheritance of rabbit immunoglobulin VH allotypes: implications for the generation of antibody diversity.

PubMed

Knight, K L; Becker, R S

1990-03-23

Rabbits are unique in that their immunoglobulin VH regions bear allotypic markers encoded by allelic genes. The presence of these markers on most serum immunoglobulins is difficult to explain, as the germline contains several hundred VH genes. We cloned VH genes from normal rabbits of the VHa allotypes a1, a2, and a3 and from a mutant a2 rabbit, Alicia, which expresses almost no a2 allotype. The D-proximal VH gene VH1 of normal rabbits encoded prototype a1, a2, or a3 allotype VH regions in a1, a2, or a3 rabbits, respectively; VH1 was shown to be preferentially utilized in leukemic rabbit B cells. This VH1 gene was deleted from the germline of the Alicia rabbit. These data suggest that the allelic inheritance of a allotypes results from preferential utilization of VH1 in VDJ rearrangements. We suggest that antibody diversity in rabbit primarily results from somatic hypermutation and gene conversion.

Plasmid-Encoded Tetracycline Efflux Pump Protein Alters Bacterial Stress Responses and Ecological Fitness of Acinetobacter oleivorans

PubMed Central

Hong, Hyerim; Jung, Jaejoon; Park, Woojun

2014-01-01

Acquisition of the extracellular tetracycline (TC) resistance plasmid pAST2 affected host gene expression and phenotype in the oil-degrading soil bacterium, Acinetobacter oleivorans DR1. Whole-transcriptome profiling of DR1 cells harboring pAST2 revealed that all the plasmid genes were highly expressed under TC conditions, and the expression levels of many host chromosomal genes were modulated by the presence of pAST2. The host energy burden imposed by replication of pAST2 led to (i) lowered ATP concentrations, (ii) downregulated expression of many genes involved in cellular growth, and (iii) reduced growth rate. Interestingly, some phenotypes were restored by deleting the plasmid-encoded efflux pump gene tetH, suggesting that the membrane integrity changes resulting from the incorporation of efflux pump proteins also resulted in altered host response under the tested conditions. Alteration of membrane integrity by tetH deletion was shown by measuring permeability of fluorescent probe and membrane hydrophobicity. The presence of the plasmid conferred peroxide and superoxide resistance to cells, but only peroxide resistance was diminished by tetH gene deletion, suggesting that the plasmid-encoded membrane-bound efflux pump protein provided peroxide resistance. The downregulation of fimbriae-related genes presumably led to reduced swimming motility, but this phenotype was recovered by tetH gene deletion. Our data suggest that not only the plasmid replication burden, but also its encoded efflux pump protein altered host chromosomal gene expression and phenotype, which also alters the ecological fitness of the host in the environment. PMID:25229538

Plasmid-encoded tetracycline efflux pump protein alters bacterial stress responses and ecological fitness of Acinetobacter oleivorans.

PubMed

Hong, Hyerim; Jung, Jaejoon; Park, Woojun

2014-01-01

Acquisition of the extracellular tetracycline (TC) resistance plasmid pAST2 affected host gene expression and phenotype in the oil-degrading soil bacterium, Acinetobacter oleivorans DR1. Whole-transcriptome profiling of DR1 cells harboring pAST2 revealed that all the plasmid genes were highly expressed under TC conditions, and the expression levels of many host chromosomal genes were modulated by the presence of pAST2. The host energy burden imposed by replication of pAST2 led to (i) lowered ATP concentrations, (ii) downregulated expression of many genes involved in cellular growth, and (iii) reduced growth rate. Interestingly, some phenotypes were restored by deleting the plasmid-encoded efflux pump gene tetH, suggesting that the membrane integrity changes resulting from the incorporation of efflux pump proteins also resulted in altered host response under the tested conditions. Alteration of membrane integrity by tetH deletion was shown by measuring permeability of fluorescent probe and membrane hydrophobicity. The presence of the plasmid conferred peroxide and superoxide resistance to cells, but only peroxide resistance was diminished by tetH gene deletion, suggesting that the plasmid-encoded membrane-bound efflux pump protein provided peroxide resistance. The downregulation of fimbriae-related genes presumably led to reduced swimming motility, but this phenotype was recovered by tetH gene deletion. Our data suggest that not only the plasmid replication burden, but also its encoded efflux pump protein altered host chromosomal gene expression and phenotype, which also alters the ecological fitness of the host in the environment.

The switch from fermentation to respiration in Saccharomyces cerevisiae is regulated by the Ert1 transcriptional activator/repressor.

PubMed

Gasmi, Najla; Jacques, Pierre-Etienne; Klimova, Natalia; Guo, Xiao; Ricciardi, Alessandra; Robert, François; Turcotte, Bernard

2014-10-01

In the yeast Saccharomyces cerevisiae, fermentation is the major pathway for energy production, even under aerobic conditions. However, when glucose becomes scarce, ethanol produced during fermentation is used as a carbon source, requiring a shift to respiration. This adaptation results in massive reprogramming of gene expression. Increased expression of genes for gluconeogenesis and the glyoxylate cycle is observed upon a shift to ethanol and, conversely, expression of some fermentation genes is reduced. The zinc cluster proteins Cat8, Sip4, and Rds2, as well as Adr1, have been shown to mediate this reprogramming of gene expression. In this study, we have characterized the gene YBR239C encoding a putative zinc cluster protein and it was named ERT1 (ethanol regulated transcription factor 1). ChIP-chip analysis showed that Ert1 binds to a limited number of targets in the presence of glucose. The strongest enrichment was observed at the promoter of PCK1 encoding an important gluconeogenic enzyme. With ethanol as the carbon source, enrichment was observed with many additional genes involved in gluconeogenesis and mitochondrial function. Use of lacZ reporters and quantitative RT-PCR analyses demonstrated that Ert1 regulates expression of its target genes in a manner that is highly redundant with other regulators of gluconeogenesis. Interestingly, in the presence of ethanol, Ert1 is a repressor of PDC1 encoding an important enzyme for fermentation. We also show that Ert1 binds directly to the PCK1 and PDC1 promoters. In summary, Ert1 is a novel factor involved in the regulation of gluconeogenesis as well as a key fermentation gene. Copyright © 2014 by the Genetics Society of America.

Sequence analysis and expression of the M1 and M2 matrix protein genes of hirame rhabdovirus (HIRRV)

USGS Publications Warehouse

Nishizawa, T.; Kurath, G.; Winton, J.R.

1997-01-01

We have cloned and sequenced a 2318 nucleotide region of the genomic RNA of hirame rhabdovirus (HIRRV), an important viral pathogen of Japanese flounder Paralichthys olivaceus. This region comprises approximately two-thirds of the 3' end of the nucleocapsid protein (N) gene and the complete matrix protein (M1 and M2) genes with the associated intergenic regions. The partial N gene sequence was 812 nucleotides in length with an open reading frame (ORF) that encoded the carboxyl-terminal 250 amino acids of the N protein. The M1 and M2 genes were 771 and 700 nucleotides in length, respectively, with ORFs encoding proteins of 227 and 193 amino acids. The M1 gene sequence contained an additional small ORF that could encode a highly basic, arginine-rich protein of 25 amino acids. Comparisons of the N, M1, and M2 gene sequences of HIRRV with the corresponding sequences of the fish rhabdoviruses, infectious hematopoietic necrosis virus (IHNV) or viral hemorrhagic septicemia virus (VHSV) indicated that HIRRV was more closely related to IHNV than to VHSV, but was clearly distinct from either. The putative consensus gene termination sequence for IHNV and VHSV, AGAYAG(A)(7), was present in the N-M1, M1-M2, and M2-G intergenic regions of HIRRV as were the putative transcription initiation sequences YGGCAC and AACA. An Escherichia coli expression system was used to produce recombinant proteins from the M1 and M2 genes of HIRRV. These were the same size as the authentic M1 and M2 proteins and reacted with anti-HIRRV rabbit serum in western blots. These reagents can be used for further study of the fish immune response and to test novel control methods.

Gene Expression by the Sulfate-Reducing Bacterium Desulfovibrio vulgaris Hildenborough Grown on an Iron Electrode under Cathodic Protection Conditions▿ †

PubMed Central

Caffrey, Sean M.; Park, Hyung Soo; Been, Jenny; Gordon, Paul; Sensen, Christoph W.; Voordouw, Gerrit

2008-01-01

The genome sequence of the sulfate-reducing bacterium Desulfovibrio vulgaris Hildenborough was reanalyzed to design unique 70-mer oligonucleotide probes against 2,824 probable protein-coding regions. These included three genes not previously annotated, including one that encodes a c-type cytochrome. Using microarrays printed with these 70-mer probes, we analyzed the gene expression profile of wild-type D. vulgaris grown on cathodic hydrogen, generated at an iron electrode surface with an imposed negative potential of −1.1 V (cathodic protection conditions). The gene expression profile of cells grown on cathodic hydrogen was compared to that of cells grown with gaseous hydrogen bubbling through the culture. Relative to the latter, the electrode-grown cells overexpressed two hydrogenases, the hyn-1 genes for [NiFe] hydrogenase 1 and the hyd genes, encoding [Fe] hydrogenase. The hmc genes for the high-molecular-weight cytochrome complex, which allows electron flow from the hydrogenases across the cytoplasmic membrane, were also overexpressed. In contrast, cells grown on gaseous hydrogen overexpressed the hys genes for [NiFeSe] hydrogenase. Cells growing on the electrode also overexpressed genes encoding proteins which promote biofilm formation. Although the gene expression profiles for these two modes of growth were distinct, they were more closely related to each other than to that for cells grown in a lactate- and sulfate-containing medium. Electrochemically measured corrosion rates were lower for iron electrodes covered with hyn-1, hyd, and hmc mutant biofilms than for wild-type biofilms. This confirms the importance, suggested by the gene expression studies, of the corresponding gene products in D. vulgaris-mediated iron corrosion. PMID:18310429

Novel Type V Staphylococcal Cassette Chromosome mec Driven by a Novel Cassette Chromosome Recombinase, ccrC

PubMed Central

Ito, Teruyo; Ma, Xiao Xue; Takeuchi, Fumihiko; Okuma, Keiko; Yuzawa, Harumi; Hiramatsu, Keiichi

2004-01-01

Staphylococcal cassette chromosome mec (SCCmec) is a mobile genetic element composed of the mec gene complex, which encodes methicillin resistance, and the ccr gene complex, which encodes the recombinases responsible for its mobility. The mec gene complex has been classified into four classes, and the ccr gene complex has been classified into three allotypes. Different combinations of mec gene complex classes and ccr gene complex types have so far defined four types of SCCmec elements. Now we introduce the fifth allotype of SCCmec, which was found on the chromosome of a community-acquired methicillin-resistant Staphylococcus aureus strain (strain WIS [WBG8318]) isolated in Australia. The element shared the same chromosomal integration site with the four extant types of SCCmec and the characteristic nucleotide sequences at the chromosome-SCCmec junction regions. The novel SCCmec carried mecA bracketed by IS431 (IS431-mecA-ΔmecR1-IS431), which is designated the class C2 mec gene complex; and instead of ccrA and ccrB genes, it carried a single copy of a gene homologue that encoded cassette chromosome recombinase. Since the open reading frame (ORF) was found to encode an enzyme which catalyzes the precise excision as well as site- and orientation-specific integration of the element, we designated the ORF cassette chromosome recombinase C (ccrC), and we designated the element type V SCCmec. Type V SCCmec is a small SCCmec element (28 kb) and does not carry any antibiotic resistance genes besides mecA. Unlike the extant SCCmec types, it carries a set of foreign genes encoding a restriction-modification system that might play a role in the stabilization of the element on the chromosome. PMID:15215121

Structure and genomic organization of the human B1 receptor gene for kinins (BDKRB1).

PubMed

Bachvarov, D R; Hess, J F; Menke, J G; Larrivée, J F; Marceau, F

1996-05-01

Two subtypes of mammalian bradykinin receptors, B1 and B2 (BDKRB1 and BDKRB2), have been defined based on their pharmacological properties. The B1 type kinin receptors have weak affinity for intact BK or Lys-BK but strong affinity for kinin metabolites without the C-terminal arginine (e.g., des-Arg9-BK and Lys-des-Arg9-BK, also called des-Arg10-kallidin), which are generated by kininase I. The B1 receptor expression is up-regulated following tissue injury and inflammation (hyperemia, exudation, hyperalgesia, etc.). In the present study, we have cloned and sequenced the gene encoding human B1 receptor from a human genomic library. The human B1 receptor gene contains three exons separated by two introns. The first and the second exon are noncoding, while the coding region and the 3'-flanking region are located entirely on the third exon. The exon-intron arrangement of the human B1 receptor gene shows significant similarity with the genes encoding the B2 receptor subtype in human, mouse, and rat. Sequence analysis of the 5'-flanking region revealed the presence of a consensus TATA box and of numerous candidate transcription factor binding sequences. Primer extension experiments have shown the existence of multiple transcription initiation sites situated downstream and upstream from the consensus TATA box. Genomic Southern blot analysis indicated that the human B1 receptor is encoded by a single-copy gene.

Multi-functional acetyl-CoA carboxylase from Brassica napus is encoded by a multi-gene family: indication for plastidic localization of at least one isoform.

PubMed

Schulte, W; Töpfer, R; Stracke, R; Schell, J; Martini, N

1997-04-01

Three genes coding for different multifunctional acetyl-CoA carboxylase (ACCase; EC 6.4.1.2) isoenzymes from Brassica napus were isolated and divided into two major classes according to structural features in their 5' regions: class I comprises two genes with an additional coding exon of approximately 300 bp at the 5' end, and class II is represented by one gene carrying an intron of 586 bp in its 5' untranslated region. Fusion of the peptide sequence encoded by the additional first exon of a class I ACCase gene to the jellyfish Aequorea victoria green fluorescent protein (GFP) and transient expression in tobacco protoplasts targeted GFP to the chloroplasts. In contrast to the deduced primary structure of the biotin carboxylase domain encoded by the class I gene, the corresponding amino acid sequence of the class II ACCase shows higher identity with that of the Arabidopsis ACCase, both lacking a transit peptide. The Arabidopsis ACCase has been proposed to be a cytosolic isoenzyme. These observations indicate that the two classes of ACCase genes encode plastidic and cytosolic isoforms of multi-functional, eukaryotic type, respectively, and that B. napus contains at least one multi-functional ACCase besides the multi-subunit, prokaryotic type located in plastids. Southern blot analysis of genomic DNA from B. napus, Brassica rapa, and Brassica oleracea, the ancestors of amphidiploid rapeseed, using a fragment of a multi-functional ACCase gene as a probe revealed that ACCase is encoded by a multi-gene family of at least five members.

The predominant WT1 isoform (+KTS) encodes a DNA-binding protein targeting the planar cell polarity gene Scribble in renal podocytes.

PubMed

Wells, Julie; Rivera, Miguel N; Kim, Woo Jae; Starbuck, Kristen; Haber, Daniel A

2010-07-01

WT1 encodes a tumor suppressor first identified by its inactivation in Wilms' Tumor. Although one WT1 splicing variant encodes a well-characterized zinc finger transcription factor, little is known about the function of the most prevalent WT1 isoform, whose DNA binding domain is disrupted by a three-amino acid (KTS) insertion. Using cells that conditionally express WT1(+KTS), we undertook a genome-wide chromatin immunoprecipitation and cloning analysis to identify candidate WT1(+KTS)-regulated promoters. We identified the planar cell polarity gene Scribble (SCRB) as the first WT1(+KTS) target gene in podocytes of the kidney. WT1 and SCRB expression patterns overlap precisely in developing renal glomeruli of mice, and WT1(+KTS) binds to a 33-nucleotide region within the Scribble promoter in mouse and human cell lines and kidneys. Together, our results support a role for the predominant WT1(+KTS) isoform in transcriptional regulation and suggest a link between the WT1-dependent tumor suppressor pathway and a key component of the planar cell polarity pathway.

Viral Repression of Fungal Pheromone Precursor Gene Expression

PubMed Central

Zhang, Lei; Baasiri, Rudeina A.; Van Alfen, Neal K.

1998-01-01

Biological control of chestnut blight caused by the filamentous ascomycete Cryphonectria parasitica can be achieved with a virus that infects this fungus. This hypovirus causes a perturbation of fungal development that results in low virulence (hypovirulence), poor asexual sporulation, and female infertility without affecting fungal growth in culture. At the molecular level, the virus is known to affect the transcription of a number of fungal genes. Two of these genes, Vir1 and Vir2, produce abundant transcripts in noninfected strains of the fungus, but the transcripts are not detectable in virus-infected strains. We report here that these two genes encode the pheromone precursors of the Mat-2 mating type of the fungus; consequently, these genes have been renamed Mf2/1 and Mf2/2. To determine if the virus affects the mating systems of both mating types of this fungus, the pheromone precursor gene, Mf1/1, of a Mat-1 strain was cloned and likewise was found to be repressed in virus-infected strains. The suppression of transcription of the pheromone precursor genes of this fungus could be the cause of the mating defect of infected strains of the fungus. Although published reports suggest that a Gαi subunit may be involved in this regulation, our results do not support this hypothesis. The prepropheromone encoded by Mf1/1 is structurally similar to that of the prepro-p-factor of Schizosaccharomyces pombe. This is the first description of the complete set of pheromone precursor genes encoded by a filamentous ascomycete. PMID:9447992

Developmentally distinct MYB genes encode functionally equivalent proteins in Arabidopsis.

PubMed

Lee, M M; Schiefelbein, J

2001-05-01

The duplication and divergence of developmental control genes is thought to have driven morphological diversification during the evolution of multicellular organisms. To examine the molecular basis of this process, we analyzed the functional relationship between two paralogous MYB transcription factor genes, WEREWOLF (WER) and GLABROUS1 (GL1), in Arabidopsis. The WER and GL1 genes specify distinct cell types and exhibit non-overlapping expression patterns during Arabidopsis development. Nevertheless, reciprocal complementation experiments with a series of gene fusions showed that WER and GL1 encode functionally equivalent proteins, and their unique roles in plant development are entirely due to differences in their cis-regulatory sequences. Similar experiments with a distantly related MYB gene (MYB2) showed that its product cannot functionally substitute for WER or GL1. Furthermore, an analysis of the WER and GL1 proteins shows that conserved sequences correspond to specific functional domains. These results provide new insights into the evolution of the MYB gene family in Arabidopsis, and, more generally, they demonstrate that novel developmental gene function may arise solely by the modification of cis-regulatory sequences.

Evolution and Variation of Renin Genes in Mice

PubMed Central

Dickinson, Douglas P.; Gross, Kenneth W.; Piccini, Nina; Wilson, Carol M.

1984-01-01

Inbred strains of mice carry Ren-1, a gene encoding the thermostable Renin-1 isozyme. Ren-1 is expressed at relatively low levels in mouse submandibular gland and kidney. Some strains also carry Ren-2, a gene encoding the thermolabile Renin-2 isozyme. Ren-2 is expressed at high levels in the mouse submandibular gland and at very low levels, if at all, in the kidney. Ren-1 and Ren-2 are closely linked on mouse chromosome 1, show extensive homology in coding and noncoding regions and provide a model for studying the regulation of gene expression. An investigation of renin genes and enzymatic activity in wild-derived mice identified several restriction site polymorphisms as well as putative variants in renin gene expression and protein structure. The number of renin genes carried by different subpopulations of wild-derived mice is consistent with the occurrence of a gene duplication event prior to the divergence of M. spretus (2.75–5.5 million yr ago). This conclusion is in agreement with a prior estimate based upon comparative sequence analysis of Ren-1 and Ren-2 from inbred laboratory mice. PMID:6389258

The mitochondrial gene encoding ribosomal protein S12 has been translocated to the nuclear genome in Oenothera.

PubMed Central

Grohmann, L; Brennicke, A; Schuster, W

1992-01-01

The Oenothera mitochondrial genome contains only a gene fragment for ribosomal protein S12 (rps12), while other plants encode a functional gene in the mitochondrion. The complete Oenothera rps12 gene is located in the nucleus. The transit sequence necessary to target this protein to the mitochondrion is encoded by a 5'-extension of the open reading frame. Comparison of the amino acid sequence encoded by the nuclear gene with the polypeptides encoded by edited mitochondrial cDNA and genomic sequences of other plants suggests that gene transfer between mitochondrion and nucleus started from edited mitochondrial RNA molecules. Mechanisms and requirements of gene transfer and activation are discussed. Images PMID:1454526

moxFG region encodes four polypeptides in the methanol-oxidizing bacterium Methylobacterium sp. strain AM1

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

Anderson, D.J.; Lidstrom, M.E.

The polypeptides encoded by a putative methanol oxidation (mox) operon of Methylobacterium sp. strain AM1 were expressed in Escherichia coli, using a coupled in vivo T7 RNA polymerase/promoter gene expression system. Two mox genes had been previously mapped to this region: moxF, the gene encoding the methanol dehydrogenase (MeDH) polypeptide; and moxG, a gene believed to encode a soluble type c cytochrome, cytochrome c/sub L/. In this study, four polypeptides of M/sub r/, 60,000, 30,000, 20,000, and 12,000 were found to be encoded by the moxFG region and were tentatively designated moxF, -J, -G, and -I, respectively. The arrangement ofmore » the genes (5' to 3') was found to be moxFJGI. The identities of three of the four polypeptides were determined by protein immunoblot analysis. The product of moxF, the M/sub r/-60,000 polypeptide, was confirmed to be the MeDH polypeptide. The product of moxG, the M/sub r/-20,000 polypeptide, was identified as mature cytochrome c/sub L/, and the product of moxI, the M/sub r/-12,000 polypeptide, was identified as a MeDH-associated polypeptide that copurifies with the holoenzyme. The identity of the M/sub r/-30,000 polypeptide (the moxJ gene product) could not be determined. The function of the M/sub r/-12,000 MeDH-associated polypeptide is not yet clear. However, it is not present in mutants that lack the M/sub r/-60,000 MeDH subunit, and it appears that the stability of the MeDH-associated polypeptide is dependent on the presence of the M/sub r/-60,000 MeDH polypeptide. Our data suggest that both the M/sub r/-30,000 and -12,000 polypeptides are involved in methanol oxidation, which would bring to 12 the number of mox genes in Methylobacterium sp. strain AM1.« less

Identification of a transcriptional activation domain in yeast repressor activator protein 1 (Rap1) using an altered DNA-binding specificity variant

PubMed Central

Johnson, Amanda N.; Weil, P. Anthony

2017-01-01

Repressor activator protein 1 (Rap1) performs multiple vital cellular functions in the budding yeast Saccharomyces cerevisiae. These include regulation of telomere length, transcriptional repression of both telomere-proximal genes and the silent mating type loci, and transcriptional activation of hundreds of mRNA-encoding genes, including the highly transcribed ribosomal protein- and glycolytic enzyme-encoding genes. Studies of the contributions of Rap1 to telomere length regulation and transcriptional repression have yielded significant mechanistic insights. However, the mechanism of Rap1 transcriptional activation remains poorly understood because Rap1 is encoded by a single copy essential gene and is involved in many disparate and essential cellular functions, preventing easy interpretation of attempts to directly dissect Rap1 structure-function relationships. Moreover, conflicting reports on the ability of Rap1-heterologous DNA-binding domain fusion proteins to serve as chimeric transcriptional activators challenge use of this approach to study Rap1. Described here is the development of an altered DNA-binding specificity variant of Rap1 (Rap1AS). We used Rap1AS to map and characterize a 41-amino acid activation domain (AD) within the Rap1 C terminus. We found that this AD is required for transcription of both chimeric reporter genes and authentic chromosomal Rap1 enhancer-containing target genes. Finally, as predicted for a bona fide AD, mutation of this newly identified AD reduced the efficiency of Rap1 binding to a known transcriptional coactivator TFIID-binding target, Taf5. In summary, we show here that Rap1 contains an AD required for Rap1-dependent gene transcription. The Rap1AS variant will likely also be useful for studies of the functions of Rap1 in other biological pathways. PMID:28196871

X-linked juvenile retinoschisis: mutations at the retinoschisis and Norrie disease gene loci?

PubMed

Hiraoka, M; Rossi, F; Trese, M T; Shastry, B S

2001-01-01

Juvenile retinoschisis (RS) and Norrie disease (ND) are X-linked recessive retinal disorders. Both disorders, in the majority of cases, are monogenic and are caused by mutations in the RS and ND genes, respectively. Here we report the identification of a family in which mutations in both the RS and ND genes are segregating with RS pathology. Although the mutations identified in this report were not functionally characterized with regard to their pathogenicity, it is likely that both of them are involved in RS pathology in the family analyzed. This suggests the complexity and digenic nature of monogenic human disorders in some cases. If this proves to be a widespread problem, it will complicate the strategies used to identify the genes involved in diseases and to develop methods for intervention.

Disruption of the Crithidia fasciculata RNH1 gene results in the loss of two active forms of ribonuclease H.

PubMed Central

Ray, D S; Hines, J C

1995-01-01

Both prokaryotic and eukaryotic cells contain multiple forms of ribonuclease H, a ribonuclease that specifically degrades the RNA strand of RNA-DNA hybrids and which has been implicated in the processing of initiator RNAs and in the removal of RNA primers from Okazaki fragments. The Crithidia fasciculata RNH1 gene encodes an RNase H and was shown to be a single-copy gene in this diploid trypanosomatid. The RNH1 gene has been disrupted by targeted gene disruption using hygromycin or G418 drug-resistance cassettes. Major active forms of RNase H (38 and 45 kDa) were observed on activity gels of extracts of wild-type cells or cells in which one allele of RNH1 was disrupted. Both the 38 and 45 kDa activities were absent in extracts of cells in which both alleles of RNH1 were disrupted indicating that both forms of the C.fasciculata RNase H are encoded by the RNH1 gene. Images PMID:7630731

Alternative intronic promoters in development and disease.

PubMed

Vacik, Tomas; Raska, Ivan

2017-05-01

Approximately 20,000 mammalian genes are estimated to encode between 250 thousand and 1 million different proteins. This enormous diversity of the mammalian proteome is caused by the ability of a single-gene locus to encode multiple protein isoforms. Protein isoforms encoded by one gene locus can be functionally distinct, and they can even have antagonistic functions. One of the mechanisms involved in creating this proteome complexity is alternative promoter usage. Alternative intronic promoters are located downstream from their canonical counterparts and drive the expression of alternative RNA isoforms that lack upstream exons. These upstream exons can encode some important functional domains, and proteins encoded by alternative mRNA isoforms can be thus functionally distinct from the full-length protein encoded by canonical mRNA isoforms. Since any misbalance of functionally distinct protein isoforms is likely to have detrimental consequences for the cell and the whole organism, their expression must be precisely regulated. Misregulation of alternative intronic promoters is frequently associated with various developmental defects and diseases including cancer, and it is becoming increasingly clear that this phenomenon deserves more attention.

Coordinated Rates of Evolution between Interacting Plastid and Nuclear Genes in Geraniaceae

PubMed Central

Zhang, Jin; Ruhlman, Tracey A.; Sabir, Jamal; Blazier, J. Chris; Jansen, Robert K.

2015-01-01

Although gene coevolution has been widely observed within individuals and between different organisms, rarely has this phenomenon been investigated within a phylogenetic framework. The Geraniaceae is an attractive system in which to study plastid-nuclear genome coevolution due to the highly elevated evolutionary rates in plastid genomes. In plants, the plastid-encoded RNA polymerase (PEP) is a protein complex composed of subunits encoded by both plastid (rpoA, rpoB, rpoC1, and rpoC2) and nuclear genes (sig1-6). We used transcriptome and genomic data for 27 species of Geraniales in a systematic evaluation of coevolution between genes encoding subunits of the PEP holoenzyme. We detected strong correlations of dN (nonsynonymous substitutions) but not dS (synonymous substitutions) within rpoB/sig1 and rpoC2/sig2, but not for other plastid/nuclear gene pairs, and identified the correlation of dN/dS ratio between rpoB/C1/C2 and sig1/5/6, rpoC1/C2 and sig2, and rpoB/C2 and sig3 genes. Correlated rates between interacting plastid and nuclear sequences across the Geraniales could result from plastid-nuclear genome coevolution. Analyses of coevolved amino acid positions suggest that structurally mediated coevolution is not the major driver of plastid-nuclear coevolution. The detection of strong correlation of evolutionary rates between SIG and RNAP genes suggests a plausible explanation for plastome-genome incompatibility in Geraniaceae. PMID:25724640

LAMP-1-chimeric DNA vaccines enhance the antibody response in Japanese flounder, Paralichthys olivaceus.

PubMed

Rondón-Barragán, Iang; Nozaki, Reiko; Hirono, Ikuo; Kondo, Hidehiro

2017-08-01

DNA vaccination is one method to protect farmed fish from viral and bacterial diseases. Chimeric antigens encoded by DNA vaccines have been shown to increase the resistance to viral diseases. Here, we sequenced the gene encoding lysosome-associated membrane protein-1 from Japanese flounder, Paralichthys olivaceus, (JfLAMP-1) and assessed its use in a chimeric DNA vaccine fused with the major capsule protein (MCP) from red seabream iridovirus (RSIV). JfLAMP-1 cDNA has a length of 1248 bp encoding 415 aa, which contains transmembrane and cytoplasmic domains. JfLAMP-1 is constitutively expressed in several tissues and its expression in spleen was upregulated following injection of formalin-killed cells (FKC) of Edwardsiella tarda. Immunofluorescence analysis showed that JfLAMP-1 is distributed in the small and large granules in the cytoplasm and groups close to the nucleus. The DNA encoding the luminal domain of JfLAMP-1 was replaced with the gene for the RSIV MCP, and the construct was cloned in an expression vector (pCIneo). Fish vaccinated with pCLAMP-MCP had significantly higher antibody levels than fish vaccinated with pCIneo vector harboring the MCP gene (p < 0.05) at day 30 post-vaccination. Copyright © 2017 Elsevier Ltd. All rights reserved.

Bacteriophage-based vectors for site-specific insertion of DNA in the chromosome of Corynebacteria.

PubMed

Oram, Mark; Woolston, Joelle E; Jacobson, Andrew D; Holmes, Randall K; Oram, Diana M

2007-04-15

In Corynebacterium diphtheriae, diphtheria toxin is encoded by the tox gene of some temperate corynephages such as beta. beta-like corynephages are capable of inserting into the C. diphtheriae chromosome at two specific sites, attB1 and attB2. Transcription of the phage-encoded tox gene, and many chromosomally encoded genes, is regulated by the DtxR protein in response to Fe(2+) levels. Characterizing DtxR-dependent gene regulation is pivotal in understanding diphtheria pathogenesis and mechanisms of iron-dependent gene expression; although this has been hampered by a lack of molecular genetic tools in C. diphtheriae and related Coryneform species. To expand the systems for genetic manipulation of C. diphtheriae, we constructed plasmid vectors capable of integrating into the chromosome. These plasmids contain the beta-encoded attP site and the DIP0182 integrase gene of C. diphtheriae NCTC13129. When these vectors were delivered to the cytoplasm of non-lysogenic C. diphtheriae, they integrated into either the attB1 or attB2 sites with comparable frequency. Lysogens were also transformed with these vectors, by virtue of the second attB site. An integrated vector carrying an intact dtxR gene complemented the mutant phenotypes of a C. diphtheriae DeltadtxR strain. Additionally, strains of beta-susceptible C. ulcerans, and C. glutamicum, a species non-permissive for beta, were each transformed with these vectors. This work significantly extends the tools available for targeted transformation of both pathogenic and non-pathogenic Corynebacterium species.

Regulation of the expression of plant resistance gene SNC1 by a protein with a conserved BAT2 domain.

PubMed

Li, Yingzhong; Tessaro, Mark J; Li, Xin; Zhang, Yuelin

2010-07-01

Plant Resistance (R) genes encode immune receptors that recognize pathogens and activate defense responses. Because of fitness costs associated with maintaining R protein-mediated resistance, expression levels of R genes have to be tightly regulated. However, mechanisms on how R-gene expression is regulated are poorly understood. Here we show that MODIFIER OF snc1, 1 (MOS1) regulates the expression of SUPPRESSOR OF npr1-1, CONSTITUTIVE1 (SNC1), which encodes a Toll/interleukin receptor-nucleotide binding site-leucine-rich repeat type of R protein in Arabidopsis (Arabidopsis thaliana). In the mos1 loss-of-function mutant plants, snc1 expression is repressed and constitutive resistance responses mediated by snc1 are lost. The repression of snc1 expression in mos1 is released by knocking out DECREASE IN DNA METHYLATION1. In mos1 mutants, DNA methylation in a region upstream of SNC1 is altered. Furthermore, expression of snc1 transgenes using the native promoter does not require MOS1, indicating that regulation of SNC1 expression by MOS1 is at the chromatin level. Map-based cloning of MOS1 revealed that it encodes a novel protein with a HLA-B ASSOCIATED TRANSCRIPT2 (BAT2) domain that is conserved in plants and animals. Our study on MOS1 suggests that BAT2 domain-containing proteins may function in regulation of gene expression at chromatin level.

Gene ercA, encoding a putative iron-containing alcohol dehydrogenase, is involved in regulation of ethanol utilization in Pseudomonas aeruginosa.

PubMed

Hempel, Niels; Görisch, Helmut; Mern, Demissew S

2013-09-01

Several two-component regulatory systems are known to be involved in the signal transduction pathway of the ethanol oxidation system in Pseudomonas aeruginosa ATCC 17933. These sensor kinases and response regulators are organized in a hierarchical manner. In addition, a cytoplasmic putative iron-containing alcohol dehydrogenase (Fe-ADH) encoded by ercA (PA1991) has been identified to play an essential role in this regulatory network. The gene ercA (PA1991) is located next to ercS, which encodes a sensor kinase. Inactivation of ercA (PA1991) by insertion of a kanamycin resistance cassette created mutant NH1. NH1 showed poor growth on various alcohols. On ethanol, NH1 grew only with an extremely extended lag phase. During the induction period on ethanol, transcription of structural genes exa and pqqABCDEH, encoding components of initial ethanol oxidation in P. aeruginosa, was drastically reduced in NH1, which indicates the regulatory function of ercA (PA1991). However, transcription in the extremely delayed logarithmic growth phase was comparable to that in the wild type. To date, the involvement of an Fe-ADH in signal transduction processes has not been reported.

Gene ercA, Encoding a Putative Iron-Containing Alcohol Dehydrogenase, Is Involved in Regulation of Ethanol Utilization in Pseudomonas aeruginosa

PubMed Central

Hempel, Niels; Görisch, Helmut

2013-01-01

Several two-component regulatory systems are known to be involved in the signal transduction pathway of the ethanol oxidation system in Pseudomonas aeruginosa ATCC 17933. These sensor kinases and response regulators are organized in a hierarchical manner. In addition, a cytoplasmic putative iron-containing alcohol dehydrogenase (Fe-ADH) encoded by ercA (PA1991) has been identified to play an essential role in this regulatory network. The gene ercA (PA1991) is located next to ercS, which encodes a sensor kinase. Inactivation of ercA (PA1991) by insertion of a kanamycin resistance cassette created mutant NH1. NH1 showed poor growth on various alcohols. On ethanol, NH1 grew only with an extremely extended lag phase. During the induction period on ethanol, transcription of structural genes exa and pqqABCDEH, encoding components of initial ethanol oxidation in P. aeruginosa, was drastically reduced in NH1, which indicates the regulatory function of ercA (PA1991). However, transcription in the extremely delayed logarithmic growth phase was comparable to that in the wild type. To date, the involvement of an Fe-ADH in signal transduction processes has not been reported. PMID:23813731

Identification of novel serine proteinase gene transcripts in the midguts of two tropical insect pests, Scirpophaga incertulas (Wk.) and Helicoverpa armigera (Hb.).

PubMed

Mazumdar-Leighton, S; Babu, C R; Bennett, J

2000-01-01

We have used RT PCR and 3'RACE to identify diverse serine proteinase genes expressed in the midguts of the rice yellow stem borer (Scirpophaga incertulas) and Asian corn borer (Helicoverpa armigera). The RT-PCR primers encoded the conserved regions around the active site histidine57 and serine195 of Drosophila melanogaster alpha trypsin, including aspartate189 of the specificity pocket. These primers amplified three transcripts (SiP1-3) from midguts of S. incertulas, and two transcripts (HaP1-2) from midguts of H. armigera. The five RT PCR products were sequenced to permit design of gene-specific forward primers for use with anchored oligo dT primers in 3'RACE. Sequencing of the 3'RACE products indicated that SiP1, SiP2 and HaP1 encoded trypsin-like serine proteinases, while HaP2 encoded a chymotrypsin-like serine proteinases. The SiP3 transcript proved to be an abundant 960 nt mRNA encoding a trypsin-like protein in which the active site serine195 was replaced by aspartate. The possible functions of this unusual protein are discussed.

Molecular characterization of genes encoding inward rectifier potassium (Kir) channels in the bed bug (Cimex lectularius).

PubMed

Mamidala, Praveen; Mittapelly, Priyanka; Jones, Susan C; Piermarini, Peter M; Mittapalli, Omprakash

2013-04-01

The molecular genetics of inward-rectifier potassium (Kir) channels in insects is poorly understood. To date, Kir channel genes have been characterized only from a few representative dipterans (i.e., fruit flies and mosquitoes). The goal of the present study was to characterize Kir channel cDNAs in a hemipteran, the bed bug (Cimex lectularius). Using our previously reported bed bug transcriptome (RNA-seq), we identified two cDNAs that encode putative Kir channels. One was a full-length cDNA that encodes a protein belonging to the insect 'Kir3' clade, which we designate as 'ClKir3'. The other was a partial cDNA that encodes a protein with similarity to both the insect 'Kir1' and 'Kir2' clades, which we designate as 'ClKir1/2'. Quantitative real-time PCR analysis revealed that ClKir1/2 and ClKir3 exhibited peak expression levels in late-instar nymphs and early-instar nymphs, respectively. Furthermore, ClKir3, but not ClKir1/2, showed tissue-specific expression in Malpighian tubules of adult bed bugs. Lastly, using an improved procedure for delivering double-stranded RNA (dsRNA) to male and female bed bugs (via the cervical membrane) we demonstrate rapid and systemic knockdown of ClKir3 transcripts. In conclusion, we demonstrate that the bed bug possesses at least two genes encoding Kir channels, and that RNAi is possible for at least Kir3, thereby offering a potential approach for elucidating the roles of Kir channel genes in bed bug physiology. Copyright © 2013 Elsevier Inc. All rights reserved.

Gene coding for the E1 endoglucanase

DOEpatents

Thomas, Steven R.; Laymon, Robert A.; Himmel, Michael E.

1996-01-01

The gene encoding Acidothermus cellulolyticus E1 endoglucanase is cloned and expressed in heterologous microorganisms. A new modified E1 endoglucanase enzyme is produced along with variants of the gene and enzyme. The E1 endoglucanase is useful for hydrolyzing cellulose to sugars for simultaneous or later fermentation into alcohol.

Gene coding for the E1 endoglucanase

DOEpatents

Thomas, S.R.; Laymon, R.A.; Himmel, M.E.

1996-07-16

The gene encoding Acidothermus cellulolyticus E1 endoglucanase is cloned and expressed in heterologous microorganisms. A new modified E1 endoglucanase enzyme is produced along with variants of the gene and enzyme. The E1 endoglucanase is useful for hydrolyzing cellulose to sugars for simultaneous or later fermentation into alcohol. 6 figs.

The genome of the insecticidal Chromobacterium subtsugae PRAA4-1 and its comparison with that of Chromobacterium violaceum ATCC 12472.

PubMed

Blackburn, Michael B; Sparks, Michael E; Gundersen-Rindal, Dawn E

2016-12-01

The genome of Chromobacterium subtsugae strain PRAA4-1, a betaproteobacterium producing insecticidal compounds, was sequenced and compared with the genome of C. violaceum ATCC 12472. The genome of C. subtsugae displayed a reduction in genes devoted to capsular and extracellular polysaccharide, possessed no genes encoding nitrate reductases, and exhibited many more phage-related sequences than were observed for C. violaceum. The genomes of both species possess a number of gene clusters predicted to encode biosynthetic complexes for secondary metabolites; these clusters suggest they produce overlapping, but distinct assortments of metabolites.

TaFROG encodes a Pooideae orphan protein that interacts with SnRK1 and enhances resistance to the mycotoxigenic fungus fusarium graminearum.

USDA-ARS?s Scientific Manuscript database

All genomes encode taxonomically restricted ‘orphan’ genes, most of which are of unknown function. We report the functional characterization of the orphan gene TaFROG as a component of the wheat resistance to the globally important Fusarium head blight (FHB) disease. TaFROG is taxonomically restrict...

Analysis of gene expression provides insights into the mechanism of cadmium tolerance in Acidithiobacillus ferrooxidans.

PubMed

Chen, Minjie; Li, Yanjun; Zhang, Li; Wang, Jianying; Zheng, Chunli; Zhang, Xuefeng

2015-02-01

Acidithiobacillus ferrooxidans plays a critical role in metal solubilization in the biomining industry, and occupies an ecological niche characterized by high acidity and high concentrations of toxic heavy metal ions. In order to investigate the possible metal resistance mechanism, the cellular distribution of cadmium was tested. The result indicated that Cd(2+) entered the cells upon initial exposure resulting in increased intracellular concentrations, followed by its excretion from the cells during subsequent growth and adaptation. Sequence homology analyses were used to identify 10 genes predicted to participate in heavy metal homeostasis, and the expression of these genes was investigated in cells cultured in the presence of increasing concentrations of toxic divalent cadmium (Cd(2+)). The results suggested that one gene (cmtR A.f ) encoded a putative Cd(2+)/Pb(2+)-responsive transcriptional regulator; four genes (czcA1 A.f , czcA2 A.f , czcB1 A.f ; and czcC1 A.f ) encoded heavy metal efflux proteins for Cd(2+); two genes (cadA1 A.f and cadB1 A.f ) encoded putative cation channel proteins related to the transport of Cd(2+). No significant enhancement of gene expression was observed at low concentrations of Cd(2+) (5 mM) and most of the putative metal resistance genes were up-regulated except cmtR A.f , cadB3 A.f ; and czcB1 A.f at higher concentrations (15 and 30 mM) according to real-time polymerase chain reaction. A model was developed for the mechanism of resistance to cadmium ions based on homology analyses of the predicted genes, the transcription of putative Cd(2+) resistance genes, and previous work.

LacI Transcriptional Regulatory Networks in Clostridium thermocellum DSM1313

DOE PAGES

Wilson, Charlotte M.; Klingeman, Dawn M.; Schlachter, Caleb; ...

2016-12-21

Organisms regulate gene expression in response to the environment to coordinate metabolic reactions.Clostridium thermocellumexpresses enzymes for both lignocellulose solubilization and its fermentation to produce ethanol. In one LacI regulator termed GlyR3 inC. thermocellumATCC 27405 we identified a repressor of neighboring genes with repression relieved by laminaribiose (a β-1,3 disaccharide). To better understand the threeC. thermocellumLacI regulons, deletion mutants were constructed using the genetically tractable DSM1313 strain. DSM1313lacIgenes Clo1313_2023, Clo1313_0089, and Clo1313_0396 encode homologs of GlyR1, GlyR2, and GlyR3 from strain ATCC 27405, respectively. Furthermore, growth on cellobiose or pretreated switchgrass was unaffected by any of the gene deletions under controlled-pHmore » fermentations. Global gene expression patterns from time course analyses identified glycoside hydrolase genes encoding hemicellulases, including cellulosomal enzymes, that were highly upregulated (5- to 100-fold) in the absence of each LacI regulator, suggesting that these were repressed under wild-type conditions and that relatively few genes were controlled by each regulator under the conditions tested. Clo1313_2022, encoding lichenase enzyme LicB, was derepressed in a ΔglyR1strain. Higher expression of Clo1313_1398, which encodes the Man5A mannanase, was observed in a ΔglyR2strain, and α-mannobiose was identified as a probable inducer for GlyR2-regulated genes. For the ΔglyR3strain, upregulation of the two genes adjacent toglyR3in thecelC-glyR3-licAoperon was consistent with earlier studies. Electrophoretic mobility shift assays have confirmed LacI transcription factor binding to specific regions of gene promoters. IMPORTANCEUnderstandingC. thermocellumgene regulation is of importance for improved fundamental knowledge of this industrially relevant bacterium. Most LacI transcription factors regulate local genomic regions; however, a small number of those genes encode global regulatory proteins with extensive regulons. This study indicates that there are small specificC. thermocellumLacI regulons. Finally, the identification of LacI repressor activity for hemicellulase gene expression is a key result of this work and will add to the small body of existing literature on the area of gene regulation inC. thermocellum.« less

Retinal vascular abnormalities and dragged maculae in a carrier with a new NDP mutation (c.268delC) that caused severe Norrie disease in the proband.

PubMed

Lin, Phoebe; Shankar, Suma P; Duncan, Jacque; Slavotinek, Anne; Stone, Edwin M; Rutar, Tina

2010-02-01

Norrie disease (ND) is caused by mutations in the ND pseudoglioma (NDP) gene (MIM 300658) located at chromosome Xp11.4-p11.3. ND is characterized by abnormal retinal vascular development and vitreoretinal disorganization presenting at birth. Systemic manifestations include sensorineural deafness, progressive mental disorder, behavioral and psychological problems, growth failure, and seizures. Other vitreoretinopathies that are associated with NDP gene mutations include X-linked familial exudative vitreoretinopathy, Coats disease, persistent fetal vasculature, and retinopathy of prematurity. Phenotypic variability associated with NDP gene mutations has been well documented in affected male patients. However, there are limited data on signs in female carriers, with mild peripheral retinal abnormalities reported in both carrier and noncarrier females of families with NDP gene mutations. Here, we report a family harboring a single base-pair deletion, c.268delC, in the NDP gene causing a severe ND phenotype in the male proband and peripheral retinal vascular abnormalities with dragged maculae similar to those observed in familial exudative vitreoretinopathy in his carrier mother. Copyright (c) 2010 American Association for Pediatric Ophthalmology and Strabismus. Published by Mosby, Inc. All rights reserved.

Expression profiles of key phenylpropanoid genes during Vanilla planifolia pod development reveal a positive correlation between PAL gene expression and vanillin biosynthesis.

PubMed

Fock-Bastide, Isabelle; Palama, Tony Lionel; Bory, Séverine; Lécolier, Aurélie; Noirot, Michel; Joët, Thierry

2014-01-01

In Vanilla planifolia pods, development of flavor precursors is dependent on the phenylpropanoid pathway. The distinctive vanilla aroma is produced by numerous phenolic compounds of which vanillin is the most important. Because of the economic importance of vanilla, vanillin biosynthetic pathways have been extensively studied but agreement has not yet been reached on the processes leading to its accumulation. In order to explore the transcriptional control exerted on these pathways, five key phenylpropanoid genes expressed during pod development were identified and their mRNA accumulation profiles were evaluated during pod development and maturation using quantitative real-time PCR. As a prerequisite for expression analysis using qRT-PCR, five potential reference genes were tested, and two genes encoding Actin and EF1 were shown to be the most stable reference genes for accurate normalization during pod development. For the first time, genes encoding a phenylalanine ammonia-lyase (VpPAL1) and a cinnamate 4-hydroxylase (VpC4H1) were identified in vanilla pods and studied during maturation. Among phenylpropanoid genes, differential regulation was observed from 3 to 8 months after pollination. VpPAL1 was gradually up-regulated, reaching the maximum expression level at maturity. In contrast, genes encoding 4HBS, C4H, OMT2 and OMT3 did not show significant increase in expression levels after the fourth month post-pollination. Expression profiling of these key phenylpropanoid genes is also discussed in light of accumulation patterns for key phenolic compounds. Interestingly, VpPAL1 gene expression was shown to be positively correlated to maturation and vanillin accumulation. Copyright © 2013 Elsevier Masson SAS. All rights reserved.

CHLORELLA VIRUSES

PubMed Central

Yamada, Takashi; Onimatsu, Hideki; Van Etten, James L.

2007-01-01

Chlorella viruses or chloroviruses are large, icosahedral, plaque‐forming, double‐stranded‐DNA—containing viruses that replicate in certain strains of the unicellular green alga Chlorella. DNA sequence analysis of the 330‐kbp genome of Paramecium bursaria chlorella virus 1 (PBCV‐1), the prototype of this virus family (Phycodnaviridae), predict ∼366 protein‐encoding genes and 11 tRNA genes. The predicted gene products of ∼50% of these genes resemble proteins of known function, including many that are completely unexpected for a virus. In addition, the chlorella viruses have several features and encode many gene products that distinguish them from most viruses. These products include: (1) multiple DNA methyltransferases and DNA site‐specific endonucleases, (2) the enzymes required to glycosylate their proteins and synthesize polysaccharides such as hyaluronan and chitin, (3) a virus‐encoded K+ channel (called Kcv) located in the internal membrane of the virions, (4) a SET domain containing protein (referred to as vSET) that dimethylates Lys27 in histone 3, and (5) PBCV‐1 has three types of introns; a self‐splicing intron, a spliceosomal processed intron, and a small tRNA intron. Accumulating evidence indicates that the chlorella viruses have a very long evolutionary history. This review mainly deals with research on the virion structure, genome rearrangements, gene expression, cell wall degradation, polysaccharide synthesis, and evolution of PBCV‐1 as well as other related viruses. PMID:16877063

Molecular and Mutational Analysis of a Gelsolin-Family Member Encoded by the Flightless I Gene of Drosophila Melanogaster

PubMed Central

de-Couet, H. G.; Fong, KSK.; Weeds, A. G.; McLaughlin, P. J.; Miklos, GLG.

1995-01-01

The flightless locus of Drosophila melanogaster has been analyzed at the genetic, molecular, ultrastructural and comparative crystallographic levels. The gene encodes a single transcript encoding a protein consisting of a leucine-rich amino terminal half and a carboxyterminal half with high sequence similarity to gelsolin. We determined the genomic sequence of the flightless landscape, the breakpoints of four chromosomal rearrangements, and the molecular lesions in two lethal and two viable alleles of the gene. The two alleles that lead to flight muscle abnormalities encode mutant proteins exhibiting amino acid replacements within the S1-like domain of their gelsolin-like region. Furthermore, the deduced intronexon structure of the D. melanogaster gene has been compared with that of the Caenorhabditis elegans homologue. Furthermore, the sequence similarities of the flightless protein with gelsolin allow it to be evaluated in the context of the published crystallographic structure of the S1 domain of gelsolin. Amino acids considered essential for the structural integrity of the core are found to be highly conserved in the predicted flightless protein. Some of the residues considered essential for actin and calcium binding in gelsolin S1 and villin V1 are also well conserved. These data are discussed in light of the phenotypic characteristics of the mutants and the putative functions of the protein. PMID:8582612

Characterisation of Four LIM Protein-Encoding Genes Involved in Infection-Related Development and Pathogenicity by the Rice Blast Fungus Magnaporthe oryzae

PubMed Central

Li, Ya; Yue, Xiaofeng; Que, Yawei; Yan, Xia; Ma, Zhonghua; Talbot, Nicholas J.; Wang, Zhengyi

2014-01-01

LIM domain proteins contain contiguous double-zinc finger domains and play important roles in cytoskeletal re-organisation and organ development in multi-cellular eukaryotes. Here, we report the characterization of four genes encoding LIM proteins in the rice blast fungus Magnaporthe oryzae. Targeted gene replacement of either the paxillin-encoding gene, PAX1, or LRG1 resulted in a significant reduction in hyphal growth and loss of pathogenicity, while deletion of RGA1 caused defects in conidiogenesis and appressorium development. A fourth LIM domain gene, LDP1, was not required for infection-associated development by M. oryzae. Live cell imaging revealed that Lrg1-GFP and Rga1-GFP both localize to septal pores, while Pax1-GFP is present in the cytoplasm. To explore the function of individual LIM domains, we carried out systematic deletion of each LIM domain, which revealed the importance of the Lrg1-LIM2 and Lrg1-RhoGAP domains for Lrg1 function and overlapping functions of the three LIM domains of Pax1. Interestingly, deletion of either PAX1 or LRG1 led to decreased sensitivity to cell wall-perturbing agents, such as Congo Red and SDS (sodium dodecyl sulfate). qRT-PCR analysis demonstrated the importance of both Lrg1 and Pax1 to regulation of genes associated with cell wall biogenesis. When considered together, our results indicate that LIM domain proteins are key regulators of infection-associated morphogenesis by the rice blast fungus. PMID:24505448

Mechanisms of macroevolution: polyphagous plasticity in butterfly larvae revealed by RNA-Seq.

PubMed

de la Paz Celorio-Mancera, Maria; Wheat, Christopher W; Vogel, Heiko; Söderlind, Lina; Janz, Niklas; Nylin, Sören

2013-10-01

Transcriptome studies of insect herbivory are still rare, yet studies in model systems have uncovered patterns of transcript regulation that appear to provide insights into how insect herbivores attain polyphagy, such as a general increase in expression breadth and regulation of ribosomal, digestion- and detoxification-related genes. We investigated the potential generality of these emerging patterns, in the Swedish comma, Polygonia c-album, which is a polyphagous, widely-distributed butterfly. Urtica dioica and Ribes uva-crispa are hosts of P. c-album, but Ribes represents a recent evolutionary shift onto a very divergent host. Utilizing the assembled transcriptome for read mapping, we assessed gene expression finding that caterpillar life-history (i.e. 2nd vs. 4th-instar regulation) had a limited influence on gene expression plasticity. In contrast, differential expression in response to host-plant identified genes encoding serine-type endopeptidases, membrane-associated proteins and transporters. Differential regulation of genes involved in nucleic acid binding was also observed suggesting that polyphagy involves large scale transcriptional changes. Additionally, transcripts coding for structural constituents of the cuticle were differentially expressed in caterpillars in response to their diet indicating that the insect cuticle may be a target for plant defence. Our results state that emerging patterns of transcript regulation from model species appear relevant in species when placed in an evolutionary context. © 2013 John Wiley & Sons Ltd.

Genotype-phenotype variations in five Spanish families with Norrie disease or X-linked FEVR.

PubMed

Riveiro-Alvarez, Rosa; Trujillo-Tiebas, Maria José; Gimenez-Pardo, Ascension; Garcia-Hoyos, Maria; Cantalapiedra, Diego; Lorda-Sanchez, Isabel; Rodriguez de Alba, Marta; Ramos, Carmen; Ayuso, Carmen

2005-09-02

Norrie disease (OMIM 310600) is a rare X-linked disorder characterized by congenital blindness in males. Approximately 40 to 50% of the cases develop deafness and mental retardation. X-linked familial exudative vitreoretinopathy (XL-FEVR) is a hereditary ocular disorder characterized by a failure of peripheral retinal vascularization. Both X-linked disorders are due to mutations in the NDP gene, which encodes a 133 amino acid protein called Norrin, but autosomal recessive (AR) and autosomal dominant (AD) forms of FEVR have also been described. In this study, we report the molecular findings and the related phenotype in five Spanish families affected with Norrie disease or XL-FEVR due to mutations of the NDP gene. The study was conducted in 45 subjects from five Spanish families. These families were clinically diagnosed with Norrie disease or similar conditions. The three exons of the NDP gene were analyzed by automatic DNA sequencing. Haplotype analyses were also performed. Two new nonsense mutations, apart from other mutations previously described in the NDP gene, were found in those patients affected with ND or X-linked FEVR. An important genotype-phenotype variation was found in relation to the different mutations of the NDP gene. In fact, the same mutation may be responsible for different phenotypes. We speculate that there might be other molecular factors that interact in the retina with Norrin, which contribute to the resultant phenotypes.

Toward a comprehensive and systematic methylome signature in colorectal cancers.

PubMed

Ashktorab, Hassan; Rahi, Hamed; Wansley, Daniel; Varma, Sudhir; Shokrani, Babak; Lee, Edward; Daremipouran, Mohammad; Laiyemo, Adeyinka; Goel, Ajay; Carethers, John M; Brim, Hassan

2013-08-01

CpG Island Methylator Phenotype (CIMP) is one of the underlying mechanisms in colorectal cancer (CRC). This study aimed to define a methylome signature in CRC through a methylation microarray analysis and a compilation of promising CIMP markers from the literature. Illumina HumanMethylation27 (IHM27) array data was generated and analyzed based on statistical differences in methylation data (1st approach) or based on overall differences in methylation percentages using lower 95% CI (2nd approach). Pyrosequencing was performed for the validation of nine genes. A meta-analysis was used to identify CIMP and non-CIMP markers that were hypermethylated in CRC but did not yet make it to the CIMP genes' list. Our 1st approach for array data analysis demonstrated the limitations in selecting genes for further validation, highlighting the need for the 2nd bioinformatics approach to adequately select genes with differential aberrant methylation. A more comprehensive list, which included non-CIMP genes, such as APC, EVL, CD109, PTEN, TWIST1, DCC, PTPRD, SFRP1, ICAM5, RASSF1A, EYA4, 30ST2, LAMA1, KCNQ5, ADHEF1, and TFPI2, was established. Array data are useful to categorize and cluster colonic lesions based on their global methylation profiles; however, its usefulness in identifying robust methylation markers is limited and rely on the data analysis method. We have identified 16 non-CIMP-panel genes for which we provide rationale for inclusion in a more comprehensive characterization of CIMP+ CRCs. The identification of a definitive list for methylome specific genes in CRC will contribute to better clinical management of CRC patients.

Method for increasing thermostability in cellulase ennzymes

DOEpatents

Adney, William S.; Thomas, Steven R.; Baker, John O.; Himmel, Michael E.; Chou, Yat-Chen

1998-01-01

The gene encoding Acidothermus cellulolyticus E1 endoglucanase is cloned and expressed in Pichia pastoris. A new modified E1 endoglucanase enzyme comprising the catalytic domain of the full size E1 enzyme demonstrates enhanced thermostability and is produced by two methods. The first method of producing the new modified E1 is proteolytic cleavage to remove the cellulose binding domain and linker peptide of the full size E1. The second method of producing the new modified E1 is genetic truncation of the gene encoding the full size E1 so that the catalytic domain is expressed in the expression product.

Mutations in extracellular matrix genes NID1 and LAMC1 cause autosomal dominant Dandy-Walker malformation and occipital cephaloceles.

PubMed

Darbro, Benjamin W; Mahajan, Vinit B; Gakhar, Lokesh; Skeie, Jessica M; Campbell, Elizabeth; Wu, Shu; Bing, Xinyu; Millen, Kathleen J; Dobyns, William B; Kessler, John A; Jalali, Ali; Cremer, James; Segre, Alberto; Manak, J Robert; Aldinger, Kimerbly A; Suzuki, Satoshi; Natsume, Nagato; Ono, Maya; Hai, Huynh Dai; Viet, Le Thi; Loddo, Sara; Valente, Enza M; Bernardini, Laura; Ghonge, Nitin; Ferguson, Polly J; Bassuk, Alexander G

2013-08-01

We performed whole-exome sequencing of a family with autosomal dominant Dandy-Walker malformation and occipital cephaloceles and detected a mutation in the extracellular matrix (ECM) protein-encoding gene NID1. In a second family, protein interaction network analysis identified a mutation in LAMC1, which encodes a NID1-binding partner. Structural modeling of the NID1-LAMC1 complex demonstrated that each mutation disrupts the interaction. These findings implicate the ECM in the pathogenesis of Dandy-Walker spectrum disorders. © 2013 WILEY PERIODICALS, INC.

Norrie disease pedigree carrying the novel mutation C65Y, predicted to disrupt the cystine knot growth factor motif, analyzed by RasI restriction digestion

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

Strasberg, P.M.; Liede, H.A.; Stein, T.

1994-09-01

Norrie disease (MIM 310600; ND) is an X-linked (Xp11.2-11.3) neurodevelopmental disorder characterized by congenital blindness, retinal dysplasia with pseudoglioma formation, and often associated with progressive mental retardation and deafness. The ND gene, comprised of 3 exons, codes for an evolutionarily conserved protein of 133 amino acids. We have analyzed 8 pedigrees segregating Norrie disease. Although microdeletions have been detected in several typical ND patients, Southern blot analysis with probes L1.28, MAO-A, MAO-B, TIMP-3.9X, pTak8, and M27{beta} failed to detect such deletions in these 8 ND pedigrees. With the cloning of the ND gene, PCR analysis of all 3 exons likewisemore » did not reveal any insertions or deletions. SSCP analysis ({sup 35}S-dNTP PCR) on PCR products of exon 3 showed a band shift for 1 patient. Repeat `cold` SSCP on minigels (3 inches x 4 inches) followed by liver staining was confirmatory. Direct sequencing revealed a G{r_arrow}A transition at nucleotide 610 corresponding to amino acid 65, changing Cys to Tyr. The mutation created an RsaI site, such that the uncut, normal, and mutant PCR products (using the same PCR primers) were 297 bp, 243 and 54 bp, and 177, 72 and 54 bp respectively. Affected males in the relevant pedigree had restricted PCR products of 177, 72 and 54 bp, carrier mothers 243, 177, 72, and 54 bp, and normals, including 30 unrelated individuals, 243 and 54 bp. Recent evidence indicates that the ND gene has a C-terminal domain homologous to that of TGF{beta}, thus identifying it as putative peptide growth factor, providing a monogenic disease model for the family of cystine knot growth factors. This is the first report of a mutation in Cys 2, critical for crosslinking to Cys 5 forming a disulphide bridge which holds the cystine knot growth factor tertiary structure together.« less

Differences in virulence genes and genome patterns of mastitis-associated Staphylococcus aureus among goat, cow, and human isolates in Taiwan.

PubMed

Chu, Chishih; Wei, Yajiun; Chuang, Shih-Te; Yu, Changyou; Changchien, Chih-Hsuan; Su, Yaochi

2013-03-01

A total of 117 mastitis-associated Staphylococcus aureus isolates from cow, goat, and human patients were analyzed for differences in antibiotic susceptibility, virulence genes, and genotypes using accessory gene regulator (agr) typing, pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). Multidrug-resistant (MDR) S. aureus were commonly found in all sources, though they were predominantly found in human and goat isolates. Almost 70% of the isolates were resistant to ampicillin and penicillin. Host-associated virulence genes were identified as follows: tst, a gene encoding toxic shock syndrome toxin, was found in goat isolates; lukED and lukM, genes encoding leukocidin, found in cow isolates; lukPV, a gene encoding leukocidin, found in human isolates; and eta, a gene encoding for exfoliative toxin, found in both human and cow isolates. All four types of hemolysin, α, β, γ, and δ, were identified in human isolates, three types (α, γ, and δ), were identified in cow isolates, and two types (α and δ) were identified in goat isolates. Agr-typing determined agr1 to be the main subtype in human and cow isolates. PFGE and MLST analysis revealed the presence of diverse genotypes among the three sources. In conclusion, mastitis-associated, genetically diverse strains of MDR S. aureus differed in virulence genes among human, cow, and goat isolates.

The Tomato Terpene Synthase Gene Family1[W][OA

PubMed Central

Falara, Vasiliki; Akhtar, Tariq A.; Nguyen, Thuong T.H.; Spyropoulou, Eleni A.; Bleeker, Petra M.; Schauvinhold, Ines; Matsuba, Yuki; Bonini, Megan E.; Schilmiller, Anthony L.; Last, Robert L.; Schuurink, Robert C.; Pichersky, Eran

2011-01-01

Compounds of the terpenoid class play numerous roles in the interactions of plants with their environment, such as attracting pollinators and defending the plant against pests. We show here that the genome of cultivated tomato (Solanum lycopersicum) contains 44 terpene synthase (TPS) genes, including 29 that are functional or potentially functional. Of these 29 TPS genes, 26 were expressed in at least some organs or tissues of the plant. The enzymatic functions of eight of the TPS proteins were previously reported, and here we report the specific in vitro catalytic activity of 10 additional tomato terpene synthases. Many of the tomato TPS genes are found in clusters, notably on chromosomes 1, 2, 6, 8, and 10. All TPS family clades previously identified in angiosperms are also present in tomato. The largest clade of functional TPS genes found in tomato, with 12 members, is the TPS-a clade, and it appears to encode only sesquiterpene synthases, one of which is localized to the mitochondria, while the rest are likely cytosolic. A few additional sesquiterpene synthases are encoded by TPS-b clade genes. Some of the tomato sesquiterpene synthases use z,z-farnesyl diphosphate in vitro as well, or more efficiently than, the e,e-farnesyl diphosphate substrate. Genes encoding monoterpene synthases are also prevalent, and they fall into three clades: TPS-b, TPS-g, and TPS-e/f. With the exception of two enzymes involved in the synthesis of ent-kaurene, the precursor of gibberellins, no other tomato TPS genes could be demonstrated to encode diterpene synthases so far. PMID:21813655

The development of bactericidal yeast strains by expressing the Pediococcus acidilactici pediocin gene (pedA) in Saccharomyces cerevisiae.

PubMed

Schoeman, H; Vivier, M A; Du Toit, M; Dicks, L M; Pretorius, I S

1999-06-15

The excessive use of sulphur dioxide and other chemical preservatives in wine, beer and other fermented food and beverage products to prevent the growth of unwanted microbes holds various disadvantages for the quality of the end-products and is confronted by mounting consumer resistance. The objective of this study was to investigate the feasibility of controlling spoilage bacteria during yeast-based fermentations by engineering bactericidal strains of Saccharomyces cerevisiae. To test this novel concept, we have successfully expressed a bacteriocin gene in yeast. The pediocin operon of Pediococcus acidilactici PAC1.0 consists of four clustered genes, namely pedA (encoding a 62 amino acid precursor of the PA-1 pediocin), pedB (encoding an immunity factor), pedC (encoding a PA-1 transport protein) and pedD (encoding a protein involved in the transport and processing of PA-1). The pedA gene was inserted into a yeast expression/secretion cassette and introduced as a multicopy episomal plasmid into a laboratory strain (Y294) of S. cerevisiae. Northern blot analysis confirmed that the pedA structural gene in this construct (ADH1P-MFa1S-pedA-ADH1T, designated PED1), was efficiently expressed under the control of the yeast alcohol dehydrogenase I gene promoter (ADH1P) and terminator (ADH1T). Secretion of the PED1-encoded pediocin PA-1 was directed by the yeast mating pheromone alpha-factor's secretion signal (MFa1S). The presence of biologically active antimicrobial peptides produced by the yeast transformants was indicated by agar diffusion assays against sensitive indicator bacteria (e.g. Listeria monocytogenes B73). Protein analysis indicated the secreted heterologous peptide to be approximately 4.6 kDa, which conforms to the expected size. The heterologous peptide was present at relatively low levels in the yeast supernatant but pediocin activity was readily detected when intact yeast colonies were used in sensitive strain overlays. This study could lead to the development of bactericidal yeast strains where S. cerevisiae starter cultures not only conduct the fermentations in the wine, brewing and baking industries but also act as biological control agents to inhibit the growth of spoilage bacteria.

Positive Darwinian Selection in the Piston That Powers Proton Pumps in Complex I of the Mitochondria of Pacific Salmon

PubMed Central

Garvin, Michael R.; Bielawski, Joseph P.; Gharrett, Anthony J.

2011-01-01

The mechanism of oxidative phosphorylation is well understood, but evolution of the proteins involved is not. We combined phylogenetic, genomic, and structural biology analyses to examine the evolution of twelve mitochondrial encoded proteins of closely related, yet phenotypically diverse, Pacific salmon. Two separate analyses identified the same seven positively selected sites in ND5. A strong signal was also detected at three sites of ND2. An energetic coupling analysis revealed several structures in the ND5 protein that may have co-evolved with the selected sites. These data implicate Complex I, specifically the piston arm of ND5 where it connects the proton pumps, as important in the evolution of Pacific salmon. Lastly, the lineage to Chinook experienced rapid evolution at the piston arm. PMID:21969854

Positive Darwinian selection in the piston that powers proton pumps in complex I of the mitochondria of Pacific salmon.

PubMed

Garvin, Michael R; Bielawski, Joseph P; Gharrett, Anthony J

2011-01-01

The mechanism of oxidative phosphorylation is well understood, but evolution of the proteins involved is not. We combined phylogenetic, genomic, and structural biology analyses to examine the evolution of twelve mitochondrial encoded proteins of closely related, yet phenotypically diverse, Pacific salmon. Two separate analyses identified the same seven positively selected sites in ND5. A strong signal was also detected at three sites of ND2. An energetic coupling analysis revealed several structures in the ND5 protein that may have co-evolved with the selected sites. These data implicate Complex I, specifically the piston arm of ND5 where it connects the proton pumps, as important in the evolution of Pacific salmon. Lastly, the lineage to Chinook experienced rapid evolution at the piston arm.

Trichome-Related Mutants Provide a New Perspective on Multicellular Trichome Initiation and Development in Cucumber (Cucumis sativus L)

PubMed Central

Liu, Xingwang; Bartholomew, Ezra; Cai, Yanling; Ren, Huazhong

2016-01-01

Trichomes are specialized epidermal cells located in aerial parts of plants that function in plant defense against biotic and abiotic stresses. The simple unicellular trichomes of Arabidopsis serve as an excellent model to study the molecular mechanism of cell differentiation and pattern formation in plants. Loss-of-function mutations in Arabidopsis thaliana have suggested that the core genes GL1 (which encodes a MYB transcription factor) and TTG1 (which encodes a WD40 repeat-containing protein) are important for the initiation and spacing of leaf trichomes, while for normal trichome initiation, the genes GL3, and EGL3 (which encode a bHLH protein) are needed. However, the positive regulatory genes involved in multicellular trichrome development in cucumber remain unclear. This review focuses on the phenotype of mutants (csgl3, tril, tbh, mict, and csgl1) with disturbed trichomes in cucumber and then infers which gene(s) play key roles in trichome initiation and development in those mutants. Evidence indicates that MICT, TBH, and CsGL1 are allelic with alternative splicing. CsGL3 and TRIL are allelic and override the effect of TBH, MICT, and CsGL1 on the regulation of multicellular trichome development; and affect trichome initiation. CsGL3, TRIL, MICT, TBH, and CsGL1 encode HD-Zip proteins with different subfamilies. Genetic and molecular analyses have revealed that CsGL3, TRIL, MICT, TBH, and CsGL1 are responsible for the differentiation of epidermal cells and the development of trichomes. Based on current knowledge, a positive regulator pathway model for trichome development in cucumber was proposed and compared to a model in Arabidopsis. These data suggest that trichome development in cucumber may differ from that in Arabidopsis. PMID:27559338

Expression of the nuclear gene TaF(A)d is under mitochondrial retrograde regulation in anthers of male sterile wheat plants with timopheevii cytoplasm.

PubMed

Xu, Pei; Yang, Yuwen; Zhang, Zhengzhi; Chen, Weihua; Zhang, Caiqin; Zhang, Lixia; Zou, Sixiang; Ma, Zhengqiang

2008-01-01

Alterations of mitochondrial-encoded subunits of the F(o)F(1)-ATP synthase are frequently associated with cytoplasmic male sterility (CMS) in plants; however, little is known about the relationship of the nuclear encoded subunits of this enzyme with CMS. In the present study, the full cDNA of the gene TaF(A)d that encodes the putative F(A)d subunit of the F(o)F(1)-ATP synthase was isolated from the wheat (Triticum aestivum) fertility restorer '2114' for timopheevii cytoplasm-based CMS. The deduced 238 amino acid polypeptide is highly similar to its counterparts in dicots and other monocots but has low homology to its mammalian equivalents. TaF(A)d is a single copy gene in wheat and maps to the short arm of the group 6 chromosomes. Transient expression of the TaF(A)d-GFP fusion in onion epidermal cells demonstrated TaF(A)d's mitochondrial location. TaF(A)d was expressed abundantly in stem, leaf, anther, and ovary tissues of 2114. Nevertheless, its expression was repressed in anthers of CMS plants with timopheevii cytoplasm. Genic male sterility did not affect its expression in anthers. The expression of the nuclear gene encoding the 20 kDa subunit of F(o) was down-regulated in a manner similar to TaF(A)d in the T-CMS anthers while that of genes encoding the 6 kDa subunit of F(o) and the gamma subunit of F(1) was unaffected. These observations implied that TaF(A)d is under mitochondrial retrograde regulation in the anthers of CMS plants with timopheevii cytoplasm.

Producing human ceramide-NS by metabolic engineering using yeast Saccharomyces cerevisiae.

PubMed

Murakami, Suguru; Shimamoto, Toshi; Nagano, Hideaki; Tsuruno, Masahiro; Okuhara, Hiroaki; Hatanaka, Haruyo; Tojo, Hiromasa; Kodama, Yukiko; Funato, Kouichi

2015-11-17

Ceramide is one of the most important intercellular components responsible for the barrier and moisture retention functions of the skin. Because of the risks involved with using products of animal origin and the low productivity of plants, the availability of ceramides is currently limited. In this study, we successfully developed a system that produces sphingosine-containing human ceramide-NS in the yeast Saccharomyces cerevisiae by eliminating the genes for yeast sphingolipid hydroxylases (encoded by SUR2 and SCS7) and introducing the gene for a human sphingolipid desaturase (encoded by DES1). The inactivation of the ceramidase gene YDC1, overexpression of the inositol phosphosphingolipid phospholipase C gene ISC1, and endoplasmic reticulum localization of the DES1 gene product resulted in enhanced production of ceramide-NS. The engineered yeast strains can serve as hosts not only for providing a sustainable source of ceramide-NS but also for developing further systems to produce sphingosine-containing sphingolipids.

The relationship between mitochondrial DNA copy number and stallion sperm function.

PubMed

Darr, Christa R; Moraes, Luis E; Connon, Richard E; Love, Charles C; Teague, Sheila; Varner, Dickson D; Meyers, Stuart A

2017-05-01

Mitochondrial DNA (mtDNA) copy number has been utilized as a measure of sperm quality in several species including mice, dogs, and humans, and has been suggested as a potential biomarker of fertility in stallion sperm. The results of the present study extend this recent discovery using sperm samples from American Quarter Horse stallions of varying age. By determining copy number of three mitochondrial genes, cytochrome b (CYTB), NADH dehydrogenase 1 (ND1) and NADH dehydrogenase 4 (ND4), instead of a single gene, we demonstrate an improved understanding of mtDNA fate in stallion sperm mitochondria following spermatogenesis. Sperm samples from 37 stallions ranging from 3 to 24 years old were collected at four breeding ranches in north and central Texas during the 2015 breeding season. Samples were analyzed for sperm motion characteristics, nuclear DNA denaturability and mtDNA copy number. Mitochondrial DNA content in individual sperm was determined by real-time qPCR and normalized with a single copy nuclear gene, Beta actin. Exploratory correlation analysis revealed that total motility was negatively correlated with CYTB copy number and sperm chromatin structure. Stallion age did not have a significant effect on copy number for any of the genes. Copy number differences existed between the three genes with CYTB having the greatest number of copies (20.6 ± 1.2 copies, range: 6.0 to 41.1) followed by ND4 (15.5 ± 0.8 copies, range: 6.7 to 27.8) and finally ND1 (12.0 ± 1.0 copies, range: 0.4 to 26.6) (P < 0.05). Varying copy number across mitochondrial genes is likely to be a result of mtDNA fragmentation and degradation since downregulation of sperm mtDNA occurs during spermatogenesis and may be important for normal sperm function. Beta regression analysis suggested that for every unit increase in mtDNA copy number of CYTB, there was a 4% decrease in the odds of sperm movement (P = 0.001). Influential analysis suggested that results are robust and not highly influenced by data from individual stallions despite the low number of stallions sampled with low sperm motility. Further genome sequencing is necessary to investigate if mutations or deletions are the underlying causes of inconsistent copy numbers across mitochondrial genes. In conclusion, we show, for the first time, that increased mtDNA copy number is associated with decreased total sperm motility in stallions. We therefore suggest that mtDNA copy number may be an indicator of defective spermatogenesis in stallions. Copyright © 2017 Elsevier Inc. All rights reserved.

The impact of common dopamine D2 receptor gene polymorphisms on D2/3 receptor availability: C957T as a key determinant in putamen and ventral striatum.

PubMed

Smith, C T; Dang, L C; Buckholtz, J W; Tetreault, A M; Cowan, R L; Kessler, R M; Zald, D H

2017-04-11

Dopamine function is broadly implicated in multiple neuropsychiatric conditions believed to have a genetic basis. Although a few positron emission tomography (PET) studies have investigated the impact of single-nucleotide polymorphisms (SNPs) in the dopamine D2 receptor gene (DRD2) on D2/3 receptor availability (binding potential, BP ND ), these studies have often been limited by small sample size. Furthermore, the most commonly studied SNP in D2/3 BP ND (Taq1A) is not located in the DRD2 gene itself, suggesting that its linkage with other DRD2 SNPs may explain previous PET findings. Here, in the largest PET genetic study to date (n=84), we tested for effects of the C957T and -141C Ins/Del SNPs (located within DRD2) as well as Taq1A on BP ND of the high-affinity D2 receptor tracer 18 F-Fallypride. In a whole-brain voxelwise analysis, we found a positive linear effect of C957T T allele status on striatal BP ND bilaterally. The multilocus genetic scores containing C957T and one or both of the other SNPs produced qualitatively similar striatal results to C957T alone. The number of C957T T alleles predicted BP ND in anatomically defined putamen and ventral striatum (but not caudate) regions of interest, suggesting some regional specificity of effects in the striatum. By contrast, no significant effects arose in cortical regions. Taken together, our data support the critical role of C957T in striatal D2/3 receptor availability. This work has implications for a number of psychiatric conditions in which dopamine signaling and variation in C957T status have been implicated, including schizophrenia and substance use disorders.

Cloning and characterization of an inulinase gene from the marine yeast Candida membranifaciens subsp. flavinogenie W14-3 and its expression in Saccharomyces sp. W0 for ethanol production.

PubMed

Zhang, Lin-Lin; Tan, Mei-Juan; Liu, Guang-Lei; Chi, Zhe; Wang, Guang-Yuan; Chi, Zhen-Ming

2015-04-01

The INU1 gene encoding an exo-inulinase from the marine-derived yeast Candida membranifaciens subsp. flavinogenie W14-3 was cloned and characterized. It had an open reading frame of 1,536 bp long encoding an inulinase. The coding region of it was not interrupted by any intron. The cloned gene encoded 512 amino acid residues of a protein with a putative signal peptide of 23 amino acids and a calculated molecular mass of 57.8 kDa. The protein sequence deduced from the inulinase gene contained the inulinase consensus sequences (WMNDPNGL), (RDP), ECP FS and Q. The protein also had six conserved putative N-glycosylation sites. The deduced inulinase from the yeast strain W14-3 was found to be closely related to that from Candida kutaonensis sp. nov. KRF1, Kluyveromyces marxianus, and Cryptococcus aureus G7a. The inulinase gene with its signal peptide encoding sequence was subcloned into the pMIRSC11 expression vector and expressed in Saccharomyces sp. W0. The recombinant yeast strain W14-3-INU-112 obtained could produce 16.8 U/ml of inulinase activity and 12.5 % (v/v) ethanol from 250 g/l of inulin within 168 h. The monosaccharides were detected after the hydrolysis of inulin with the crude inulinase (the yeast culture). All the results indicated that the cloned gene and the recombinant yeast strain W14-3-INU-112 had potential applications in biotechnology.

Open reading frames in a 4556 nucleotide sequence within MDV-1 BamHI-D DNA fragment: evidence for splicing of mRNA from a new viral glycoprotein gene.

PubMed

Becker, Y; Asher, Y; Tabor, E; Davidson, I; Malkinson, M

1994-01-01

A DNA segment of the MDV-1 BamHI-D fragment was sequenced, and the open reading frames (ORFs) present in the 4556 nucleotide fragment were analyzed by computer programs. Computer analysis identified 19 putative ORFs in the sequence ranging from a coding capacity of 37 amino acids (aa) (ORF-1a) to 684aa (ORF-1). The special properties of four ORFs (1a, 1, 2, and 3) were investigated. Two adjacent ORFs, ORF-1a and ORF-1, were found by computer analysis to have the properties of two introns encoding a glycoprotein: ORF-1a encodes an aa sequence with the properties of a signal peptide, and ORF-1 encodes a polypeptide with a membrane anchor domain and putative N-glycosylation sites in the aa sequence. ORF-1a and ORF-1 were found to be transcribed in MDV-1-infected cells. Two RNA transcripts were detected: a precursor RNA and its spliced form. Both are transcribed from a promoter located 5' to ORF-1a, and splice donor and acceptor sites are used to splice the mRNA after cleavage of a 71-nucleotide sequence. This finding suggest that ORF-1a and ORF-1 are two introns of a new MDV-1 glycoprotein gene. The DNA sequence containing ORF-1 was transiently expressed in COS-1 cells, and the viral protein produced in these cells was found to react with anti-MDV serotype-1 Antigen B-specific monoclonal antibodies. These studies indicate that the protein encoded by ORF-1 has antigenic properties resembling Antigen B of MDV-1. A gene homologous to ORF-1 was detected in the genome of both MDV-2(SB1) and MDV-3(HVT), which serve as commercial vaccine strains. Two additional ORFs were noted in the 4556 nucleotide sequence: ORF-2, which encodes a 333 aa polypeptide initiating in the UL and terminating in the TRL prior to the putative origin of replication, and ORF-3, which encodes a 155 aa polypeptide that is partly homologous to the phosphoprotein pp38 encoded by the BamHI-H sequence. The 65 N-terminal aa of the two gene products are identical, both being derived from the nucleotide sequences in the TRL and IRL, respectively. Additional homologous aa sequences are the hydrophobic aa domain in the middle of both proteins. The functions of ORF-2, ORF-3, and additional ORFs are under study.

Localization and physical mapping of genes encoding the A+U-rich element RNA-binding protein AUF1 to human chromosomes 4 and X

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

Wagner, B.J.; Long, L.; Pettenati, M.J.

Messenger RNAs encoding many oncoproteins and cytokines are relatively unstable. Their instability, which ensures appropriate levels and timing of expression, is controlled in part by proteins that bind to A + U-rich instability elements (AREs) present in the 3{prime}-untranslated regions of the mRNAs. cDNAs encoding the AUF1 family of ARE-binding proteins were cloned from human and murine cDNA libraries. In the present study monochromosomal somatic cell hybrids were used to localize two AUF1 loci to human chromosomes 4 and X. In situ hybridization analyses using P1 clones as probes identified the 4q21.1-q21.2 and Xq12 regions as the locations of themore » AUF1 genes. 10 refs., 2 figs.« less

Identification of the Gene Encoding Isoprimeverose-producing Oligoxyloglucan Hydrolase in Aspergillus oryzae*

PubMed Central

Matsuzawa, Tomohiko; Mitsuishi, Yasushi; Kameyama, Akihiko

2016-01-01

Aspergillus oryzae produces a unique β-glucosidase, isoprimeverose-producing oligoxyloglucan hydrolase (IPase), that recognizes and releases isoprimeverose (α-d-xylopyranose-(1→6)-d-glucopyranose) units from the non-reducing ends of oligoxyloglucans. A gene encoding A. oryzae IPase, termed ipeA, was identified and expressed in Pichia pastoris. With the exception of cellobiose, IpeA hydrolyzes a variety of oligoxyloglucans and is a member of the glycoside hydrolase family 3. Xylopyranosyl branching at the non-reducing ends was vital for IPase activity, and galactosylation at a α-1,6-linked xylopyranosyl side chain completely abolished IpeA activity. Hepta-oligoxyloglucan saccharide (Xyl3Glc4) substrate was preferred over tri- (Xyl1Glc2) and tetra- (Xyl2Glc2) oligoxyloglucan saccharides substrates. IpeA transferred isoprimeverose units to other saccharides, indicating transglycosylation activity. The ipeA gene was expressed in xylose and xyloglucan media and was strongly induced in the presence of xyloglucan endo-xyloglucanase-hydrolyzed products. This is the first study to report the identification of a gene encoding IPase in eukaryotes. PMID:26755723

Identification and characterization of a gene product that regulates type 1 piliation in Escherichia coli.

PubMed Central

Orndorff, P E; Falkow, S

1984-01-01

The recombinant plasmid pSH2 confers type 1 piliation (Pil+) on a nonpiliated (Pil-) strain of Escherichia coli K-12. At least four plasmid-encoded gene products are involved in pilus biosynthesis and expression. We present evidence which indicates that one gene encodes an inhibitor of piliation. Hyperpiliated (Hyp) mutants were isolated after Tn5 insertion mutagenesis of pSH2 and introduction of the plasmid DNA into a Pil- strain of E. coli as unique small, compact colonies. Also, Hyp mutants clumped during growth in static broth and were piliated under several cultural conditions that normally suppressed piliation. Electron microscopic examination of Hyp mutants associated an observed 40-fold increase in pilin antigen with an increase in the number and length of pili per cell. All Hyp mutants examined failed to produce a 23-kilodalton protein that was encoded by a gene adjacent to the structural (pilin) gene for type 1 pili, and all Tn5 insertion mutations that produced the Hyp phenotype mapped in this region (hyp). Piliation in Hyp mutants could be reduced to near parental levels by introducing a second plasmid containing a parental hyp gene. Thus the 23-kilodalton (hyp) protein appears to act in trans to regulate the level of piliation. Images PMID:6148338

Resurrecting KIR2DP1: A Key Intermediate in the Evolution of Human Inhibitory NK Cell Receptors That Recognize HLA-C.

PubMed

Hilton, Hugo G; Blokhuis, Jeroen H; Guethlein, Lisbeth A; Norman, Paul J; Parham, Peter

2017-03-01

KIR2DP1 is an inactive member of the human lineage III KIR family, which includes all HLA-C-specific receptor genes. The lethal, and only, defect in KIR2DP1 is a nucleotide deletion in codon 88. Fixed in modern humans, the deletion is also in archaic human genomes. KIR2DP1 is polymorphic, with dimorphism at specificity-determining position 44. By repairing the deletion, we resurrected 11 alleles of KIR2DP1 F , the functional antecedent of KIR2DP1 We demonstrate how K44-KIR2DP1 F with lysine 44 recognized C1 + HLA-C, whereas T44-KIR2DP1 F recognized C2 + HLA-C. Dimorphisms at 12 other KIR2DP1 F residues modulate receptor avidity or signaling. KIR2DP1 and KIR2DL1 are neighbors in the centromeric KIR region and are in tight linkage disequilibrium. Like KIR2DL1 , KIR2DP1 contributed to CenA and CenB KIR haplotype differences. Encoded on CenA , C1-specific K44-KIR2DP1 F were stronger receptors than the attenuated C2-specific T44-KIR2DP1 F encoded on CenB The last common ancestor of humans and chimpanzees had diverse lineage III KIR that passed on to chimpanzees but not to humans. Early humans inherited activating KIR2DS4 and an inhibitory lineage III KIR , likely encoding a C1-specific receptor. The latter spawned the modern family of HLA-C receptors. KIR2DP1 F has properties consistent with KIR2DP1 F having been the founder gene. The first KIR2DP1 F alleles encoded K44-C1 receptors; subsequently KIR2DP1 F alleles encoding T44-C2 receptors evolved. The emergence of dedicated KIR2DL2/3 and KIR2DL1 genes encoding C1 and C2 receptors, respectively, could have led to obsolescence of KIR2DP1 F Alternatively, pathogen subversion caused its demise. Preservation of KIR2DP1 F functional polymorphism was a side effect of fixation of the deletion in KIR2DP1 F by micro gene conversion. Copyright © 2017 by The American Association of Immunologists, Inc.

[Cloning and characterization of Caveolin-1 gene in pigeon, Columba livia domestica].

PubMed

Zhang, Ying; Yu, Jian-Feng; Yang, Li; Wang, Xing-Guo; Gu, Zhi-Liang

2010-10-01

Caveolins, a class of principal proteins forming the structure of caveolae in plasmalemma, were encoded by caveolins gene family. Caveolin-1 gene is a member of caveolins gene family. In the present study, a full-length of 2605 bp caveolin-1 cDNA sequence in Columba livia domestica, which included a 537 bp complete ORF encoding a 178 amino acids long putative peptide, were obtained by using RT-PCR and RACE technique. The Columba livia domestica caveolin-1 CDS shared 80.1% - 93.4% homology with Bos taurus, Canis lupus familiaris, Gallus gallus and Rattus norvegicus. Meanwhile, the putative amino acid sequence of Columba livia domestica caveolin-1 shared 85.4% - 97.2% homology with the above species. The semi-quantity RT-PCR revealed that Caveolin-1 expressions were detectable in all the Columba livia domestica tissues and the expressional level of caveolin-1 gene was high in adipose, medium in various muscles, low in liver. These results demonstrated that Caveolin-1 gene was potentially involved in some metabolic pathways in adipose and muscle.

vanC Cluster of Vancomycin-Resistant Enterococcus gallinarum BM4174

PubMed Central

Arias, Cesar A.; Courvalin, Patrice; Reynolds, Peter E.

2000-01-01

Glycopeptide-resistant enterococci of the VanC type synthesize UDP-muramyl-pentapeptide[d-Ser] for cell wall assembly and prevent synthesis of peptidoglycan precursors ending in d-Ala. The vanC cluster of Enterococcus gallinarum BM4174 consists of five genes: vanC-1, vanXYC, vanT, vanRC, and vanSC. Three genes are sufficient for resistance: vanC-1 encodes a ligase that synthesizes the dipeptide d-Ala-d-Ser for addition to UDP-MurNAc-tripeptide, vanXYC encodes a d,d-dipeptidase–carboxypeptidase that hydrolyzes d-Ala-d-Ala and removes d-Ala from UDP-MurNAc-pentapeptide[d-Ala], and vanT encodes a membrane-bound serine racemase that provides d-Ser for the synthetic pathway. The three genes are clustered: the start codons of vanXYC and vanT overlap the termination codons of vanC-1 and vanXYC, respectively. Two genes which encode proteins with homology to the VanS-VanR two-component regulatory system were present downstream from the resistance genes. The predicted amino acid sequence of VanRC exhibited 50% identity to VanR and 33% identity to VanRB. VanSC had 40% identity to VanS over a region of 308 amino acids and 24% identity to VanSB over a region of 285 amino acids. All residues with important functions in response regulators and histidine kinases were conserved in VanRC and VanSC, respectively. Induction experiments based on the determination of d,d-carboxypeptidase activity in cytoplasmic extracts confirmed that the genes were expressed constitutively. Using a promoter-probing vector, regions upstream from the resistance and regulatory genes were identified that have promoter activity. PMID:10817725

Structure of genes for dermaseptins B, antimicrobial peptides from frog skin. Exon 1-encoded prepropeptide is conserved in genes for peptides of highly different structures and activities.

PubMed

Vouille, V; Amiche, M; Nicolas, P

1997-09-01

We cloned the genes of two members of the dermaseptin family, broad-spectrum antimicrobial peptides isolated from the skin of the arboreal frog Phyllomedusa bicolor. The dermaseptin gene Drg2 has a 2-exon coding structure interrupted by a small 137-bp intron, wherein exon 1 encoded a 22-residue hydrophobic signal peptide and the first three amino acids of the acidic propiece; exon 2 contained the 18 additional acidic residues of the propiece plus a typical prohormone processing signal Lys-Arg and a 32-residue dermaseptin progenitor sequence. The dermaseptin genes Drg2 and Drg1g2 have conserved sequences at both untranslated ends and in the first and second coding exons. In contrast, Drg1g2 comprises a third coding exon for a short version of the acidic propiece and a second dermaseptin progenitor sequence. Structural conservation between the two genes suggests that Drg1g2 arose recently from an ancestral Drg2-like gene through amplification of part of the second coding exon and 3'-untranslated region. Analysis of the cDNAs coding precursors for several frog skin peptides of highly different structures and activities demonstrates that the signal peptides and part of the acidic propieces are encoded by conserved nucleotides encompassed by the first coding exon of the dermaseptin genes. The organization of the genes that belong to this family, with the signal peptide and the progenitor sequence on separate exons, permits strikingly different peptides to be directed into the secretory pathway. The recruitment of such a homologous 'secretory' exon by otherwise non-homologous genes may have been an early event in the evolution of amphibian.

Recombinational inactivation of the gene encoding nitrate reductase in Aspergillus parasiticus.

PubMed Central

Wu, T S; Linz, J E

1993-01-01

Functional disruption of the gene encoding nitrate reductase (niaD) in Aspergillus parasiticus was conducted by two strategies, one-step gene replacement and the integrative disruption. Plasmid pPN-1, in which an internal DNA fragment of the niaD gene was replaced by a functional gene encoding orotidine monophosphate decarboxylase (pyrG), was constructed. Plasmid pPN-1 was introduced in linear form into A. parasiticus CS10 (ver-1 wh-1 pyrG) by transformation. Approximately 25% of the uridine prototrophic transformants (pyrG+) were chlorate resistant (Chlr), demonstrating their inability to utilize nitrate as a sole nitrogen source. The genetic block in nitrate utilization was confirmed to occur in the niaD gene by the absence of growth of the A. parasiticus CS10 transformants on medium containing nitrate as the sole nitrogen source and the ability to grow on several alternative nitrogen sources. Southern hybridization analysis of Chlr transformants demonstrated that the resident niaD locus was replaced by the nonfunctional allele in pPN-1. To generate an integrative disruption vector (pSKPYRG), an internal fragment of the niaD gene was subcloned into a plasmid containing the pyrG gene as a selectable marker. Circular pSKPYRG was transformed into A. parasiticus CS10. Chlr pyrG+ transformants were screened for nitrate utilization and by Southern hybridization analysis. Integrative disruption of the genomic niaD gene occurred in less than 2% of the transformants. Three gene replacement disruption transformants and two integrative disruption transformants were tested for mitotic stability after growth under nonselective conditions. All five transformants were found to stably retain the Chlr phenotype after growth on nonselective medium.(ABSTRACT TRUNCATED AT 250 WORDS) Images PMID:8215371

Sweet Taste Receptor Gene Variation and Aspartame Taste in Primates and Other Species

PubMed Central

Li, Xia; Bachmanov, Alexander A.; Maehashi, Kenji; Li, Weihua; Lim, Raymond; Brand, Joseph G.; Beauchamp, Gary K.; Reed, Danielle R.; Thai, Chloe

2011-01-01

Aspartame is a sweetener added to foods and beverages as a low-calorie sugar replacement. Unlike sugars, which are apparently perceived as sweet and desirable by a range of mammals, the ability to taste aspartame varies, with humans, apes, and Old World monkeys perceiving aspartame as sweet but not other primate species. To investigate whether the ability to perceive the sweetness of aspartame correlates with variations in the DNA sequence of the genes encoding sweet taste receptor proteins, T1R2 and T1R3, we sequenced these genes in 9 aspartame taster and nontaster primate species. We then compared these sequences with sequences of their orthologs in 4 other nontasters species. We identified 9 variant sites in the gene encoding T1R2 and 32 variant sites in the gene encoding T1R3 that distinguish aspartame tasters and nontasters. Molecular docking of aspartame to computer-generated models of the T1R2 + T1R3 receptor dimer suggests that species variation at a secondary, allosteric binding site in the T1R2 protein is the most likely origin of differences in perception of the sweetness of aspartame. These results identified a previously unknown site of aspartame interaction with the sweet receptor and suggest that the ability to taste aspartame might have developed during evolution to exploit a specialized food niche. PMID:21414996

Sweet taste receptor gene variation and aspartame taste in primates and other species.

PubMed

Li, Xia; Bachmanov, Alexander A; Maehashi, Kenji; Li, Weihua; Lim, Raymond; Brand, Joseph G; Beauchamp, Gary K; Reed, Danielle R; Thai, Chloe; Floriano, Wely B

2011-06-01

Aspartame is a sweetener added to foods and beverages as a low-calorie sugar replacement. Unlike sugars, which are apparently perceived as sweet and desirable by a range of mammals, the ability to taste aspartame varies, with humans, apes, and Old World monkeys perceiving aspartame as sweet but not other primate species. To investigate whether the ability to perceive the sweetness of aspartame correlates with variations in the DNA sequence of the genes encoding sweet taste receptor proteins, T1R2 and T1R3, we sequenced these genes in 9 aspartame taster and nontaster primate species. We then compared these sequences with sequences of their orthologs in 4 other nontasters species. We identified 9 variant sites in the gene encoding T1R2 and 32 variant sites in the gene encoding T1R3 that distinguish aspartame tasters and nontasters. Molecular docking of aspartame to computer-generated models of the T1R2 + T1R3 receptor dimer suggests that species variation at a secondary, allosteric binding site in the T1R2 protein is the most likely origin of differences in perception of the sweetness of aspartame. These results identified a previously unknown site of aspartame interaction with the sweet receptor and suggest that the ability to taste aspartame might have developed during evolution to exploit a specialized food niche.

The Chlamydomonas Dhc1 gene encodes a dynein heavy chain subunit required for assembly of the I1 inner arm complex.

PubMed Central

Myster, S H; Knott, J A; O'Toole, E; Porter, M E

1997-01-01

Multiple members of the dynein heavy chain (Dhc) gene family have been recovered in several organisms, but the relationships between these sequences and the Dhc isoforms that they encode are largely unknown. To identify Dhc loci and determine the specific functions of the individual Dhc isoforms, we have screened a collection of motility mutants generated by insertional mutagenesis in Chlamydomonas. In this report, we characterize one strain, pf9-3, in which the insertion event was accompanied by a deletion of approximately 13 kb of genomic DNA within the transcription unit of the Dhc1 gene. Northern blot analysis confirms that pf9-3 is a null mutation. Biochemical and structural studies of isolated axonemes demonstrate that the pf9-3 mutant fails to assemble the I1 inner arm complex, a two-headed dynein isoform composed of two Dhcs (1 alpha and 1 beta) and three intermediate chains. To determine if the Dhc1 gene product corresponds to one of the Dhcs of the I1 complex, antibodies were generated against a Dhc1-specific peptide sequence. Immunoblot analysis reveals that the Dhc1 gene encodes the 1 alpha Dhc subunit. These studies thus, identify the first inner arm Dhc locus to be described in any organism and further demonstrate that the 1 alpha Dhc subunit plays an essential role in the assembly of the I1 inner arm complex. Images PMID:9247642

Pseudomonas cremoricolorata Strain ND07 Produces N-acyl Homoserine Lactones as Quorum Sensing Molecules

PubMed Central

Yunos, Nina Yusrina Muhamad; Tan, Wen-Si; Koh, Chong-Lek; Sam, Choon-Kook; Mohamad, Nur Izzati; Tan, Pui-Wan; Adrian, Tan-Guan-Sheng; Yin, Wai-Fong; Chan, Kok-Gan

2014-01-01

Quorum sensing (QS) is a bacterial cell-to-cell communication system controlling QS-mediated genes which is synchronized with the population density. The regulation of specific gene activity is dependent on the signaling molecules produced, namely N-acyl homoserine lactones (AHLs). We report here the identification and characterization of AHLs produced by bacterial strain ND07 isolated from a Malaysian fresh water sample. Molecular identification showed that strain ND07 is clustered closely to Pseudomonas cremoricolorata. Spent culture supernatant extract of P. cremoricolorata strain ND07 activated the AHL biosensor Chromobacterium violaceum CV026. Using high resolution triple quadrupole liquid chromatography-mass spectrometry, it was confirmed that P. cremoricolorata strain ND07 produced N-octanoyl-l-homoserine lactone (C8-HSL) and N-decanoyl-l-homoserine lactone (C10-HSL). To the best of our knowledge, this is the first documentation on the production of C10-HSL in P. cremoricolorata strain ND07. PMID:24984061

Identification of a novel fatty acid elongase with a wide substrate specificity from arachidonic acid-producing fungus Mortierella alpina 1S-4.

PubMed

Sakuradani, Eiji; Nojiri, Masutoshi; Suzuki, Haruna; Shimizu, Sakayu

2009-09-01

The isolation and characterization of a gene (MALCE1) that encodes a fatty acid elongase from arachidonic acid-producing fungus Mortierella alpina 1S-4 are described. MALCE1 was confirmed to encode a fatty acid elongase by its expression in yeast Saccharomyces cerevisiae, resulting in the accumulation of 18-, 19-, and 20-carbon monounsaturated fatty acids and eicosanoic acid. Furthermore, the MALCE1 yeast transformant efficiently elongated exogenous 9-hexadecenoic acid, 9,12-octadecadienoic acid, and 9,12,15-octadecatrienoic acid. The MALCE1 gene-silenced strain obtained from M. alpina 1S-4 exhibited a low content of octadecanoic acid and a high content of hexadecanoic acid, compared with those in the wild strain. The enzyme encoded by MALCE1 was demonstrated to be involved in the conversion of hexadecanoic acid to octadecanoic acid, its main role in M. alpina 1S-4.

Both a PKS and a PPTase are involved in melanin biosynthesis and regulation of Aureobasidium melanogenum XJ5-1 isolated from the Taklimakan desert.

PubMed

Jiang, Hong; Liu, Guang-Lei; Chi, Zhe; Wang, Jian-Ming; Zhang, Ly-Ly; Chi, Zhen-Ming

2017-02-20

A PKS1 gene responsible for the melanin biosynthesis and a NPG1 gene in Aureobasidium melanogenum XJ5-1 were cloned and characterized. An ORF of the PKS1 gene encoding a protein with 2165 amino acids contained 6495bp while an ORF of the NPG1 gene encoding a protein with 340 amino acids had 1076bp. After analysis of their promoters, it was found that expression of both the PKS1 gene and the NPG1 gene was repressed by nitrogen sources and glucose, respectively. The PKS deduced from the cloned gene consisted of one ketosynthase, one acyl transferase, two acyl carrier proteins, one thioesterase and one cyclase while the PPTase belonged to the family Sfp-type. After disruption of the PKS1 gene and the NPG1 gene, expression of the PKS1 gene and the NPG1 gene and the melanin biosynthesis in the disruptants K5 and DP107 disappeared and expression of the PKS1 gene in the disruptant DP107 was also negatively influenced. However, after the NPG1 gene was complemented in the disruptant DP107, the melanin biosynthesis in the complementary strain BP17 was restored and expression of the PKS1 gene and the NPG1 gene was greatly enhanced, suggesting that the PKS was indeed activated and regulated by the PPTase and expression of the PKS1 gene and the NPG1 gene had a coordinate regulation. Copyright © 2016 Elsevier B.V. All rights reserved.

The possible role of maternal bonding style and CHRNB2 gene polymorphisms in nicotine dependence and related depressive phenotype.

PubMed

Csala, Iren; Egervari, Luca; Dome, Peter; Faludi, Gabor; Dome, Balazs; Lazary, Judit

2015-06-03

Neuronal nicotinic acetylcholinergic receptors (nAChR) and especially α4β2 nAChRs are the major targets for cessation medications and also for some promising antidepressant agents. Furthermore, depressive symptoms pose multifacet difficulties during cessation therapy. However, gene encoding for the β2 subunit of nAChRs has been poorly investigated in association with depression. Since both nicotine dependence (ND) and depressive phenotype are complex disorders, we investigated the effects of a significant early life experience, maternal bonding style (MB) and CHRNB2 gene SNPs on smoking-related depression. We recruited two hundred and thirty-two treatment-seeking smokers in our study. Phenotypic variants were evaluated using the Fagerstrom Test for Nicotine Dependence (FTND), the Zung Self-Rating Depression Scale (ZSDS) and the Parental Bonding Instrument (PBI). Besides the total score (TS) of ZSDS, impulsivity (ZSDS-I) and suicidal ideation (ZSDS-S) were distinguished as phenotypic variable. DNAs were extracted from buccal mucosa samples and one SNP in promoter and two SNPs in 3' UTR of CHRNB2 gene were genotyped. GLM and ANOVA tests were performed for genotype associations and interaction analyses. Maternal bonding had a significant impact on depressive phenotypes. Low care, high protection and affectionless control (ALC) were associated with ZSDS-TS and all subphenotypes of ZSDS. One SNP, the rs2072660 in 3' UTR, had a significant effect on the FTND score (p=0.010). Direct association of CHRNB2 variants and depressive phenotypes were not significant. However, in interaction with ALC, rs2072660 was significantly associated with ZSDS-S (p=0.005). MB had no significant effect on smoking-related phenotype. Our results highlight the important role of 3' UTR in the CHRNB2 gene in the shared molecular background of ND and depressive phenotype. Parental bonding style can be suggested as a significant environmental factor in further GxE studies of depression. The presented significant GxE interaction on smoking-related suicidal subphenotype may help establish further investigations on development of more effective and safer smoking cessation and antidepressant agents. Copyright © 2015 Elsevier Inc. All rights reserved.

Characterization of a new multigene family encoding isomaltases in the yeast Saccharomyces cerevisiae, the IMA family.

PubMed

Teste, Marie-Ange; François, Jean Marie; Parrou, Jean-Luc

2010-08-27

It has been known for a long time that the yeast Saccharomyces cerevisiae can assimilate alpha-methylglucopyranoside and isomaltose. We here report the identification of 5 genes (YGR287c, YIL172c, YJL216c, YJL221c and YOL157c), which, similar to the SUCx, MALx, or HXTx multigene families, are located in the subtelomeric regions of different chromosomes. They share high nucleotide sequence identities between themselves (66-100%) and with the MALx2 genes (63-74%). Comparison of their amino acid sequences underlined a substitution of threonine by valine in region II, one of the four highly conserved regions of the alpha-glucosidase family. This change was previously shown to be sufficient to discriminate alpha-1,4- to alpha-1,6-glucosidase activity in YGR287c (Yamamoto, K., Nakayama, A., Yamamoto, Y., and Tabata, S. (2004) Eur. J. Biochem. 271, 3414-3420). We showed that each of these five genes encodes a protein with alpha-glucosidase activity on isomaltose, and we therefore renamed these genes IMA1 to IMA5 for IsoMAltase. Our results also illustrated that sequence polymorphisms among this family led to interesting variability of gene expression patterns and of catalytic efficiencies on different substrates, which altogether should account for the absence of functional redundancy for growth on isomaltose. Indeed, deletion studies revealed that IMA1/YGR287c encodes the major isomaltase and that growth on isomaltose required the presence of AGT1, which encodes an alpha-glucoside transporter. Expressions of IMA1 and IMA5/YJL216c were strongly induced by maltose, isomaltose, and alpha-methylglucopyranoside, in accordance with their regulation by the Malx3p-transcription system. The physiological relevance of this IMAx multigene family in S. cerevisiae is discussed.

Global gene expression analysis in a mouse model for Norrie disease: late involvement of photoreceptor cells.

PubMed

Lenzner, Steffen; Prietz, Sandra; Feil, Silke; Nuber, Ulrike A; Ropers, H-Hilger; Berger, Wolfgang

2002-09-01

Mutations in the NDP gene give rise to a variety of eye diseases, including classic Norrie disease (ND), X-linked exudative vitreoretinopathy (EVRX), retinal telangiectasis (Coats disease), and advanced retinopathy of prematurity (ROP). The gene product is a cystine-knot-containing extracellular signaling molecule of unknown function. In the current study, gene expression was determined in a mouse model of ND, to unravel disease-associated mechanisms at the molecular level. Gene transcription in the eyes of 2-year-old Ndp knockout mice was compared with that in the eyes of age-matched wild-type control animals, by means of cDNA subtraction and microarrays. Clones (n = 3072) from the cDNA subtraction libraries were spotted onto glass slides and hybridized with fluorescently labeled RNA-derived targets. More than 230 differentially expressed clones were sequenced, and their expression patterns were verified by virtual Northern blot analysis. Numerous gene transcripts that are absent or downregulated in the eye of Ndp knockout mice are photoreceptor cell specific. In younger Ndp knockout mice (up to 1 year old), however, all these transcripts were found to be expressed at normal levels. The identification of numerous photoreceptor cell-specific transcripts with a reduced expression in 2-year-old, but not in young, Ndp knockout mice indicates that normal gene expression in these light-sensitive cells of mutant mice is established and maintained over a long period and that rods and cones are affected relatively late in the mouse model of ND. Obviously, the absence of the Ndp gene product is not compatible with long-term survival of photoreceptor cells in the mouse.

The L83L ORF of African swine fever virus strain Georgia encodes for a non-essential gene that interacts with host protein IL-1ß

USDA-ARS?s Scientific Manuscript database

African swine fever virus (ASFV) causes a contagious and frequently lethal disease of pigs that produces significant economic consequences to the swine industry. ASFV genome encodes for more than 150 genes, but only a few of them have been studied in detail. Here we report the characterization of op...

Hepatic transcriptional changes in critical genes for gluconeogenesis following castration of bulls

PubMed Central

Fassah, Dilla Mareistia; Jeong, Jin Young

2018-01-01

Objective This study was performed to understand transcriptional changes in the genes involved in gluconeogenesis and glycolysis pathways following castration of bulls. Methods Twenty Korean bulls were weaned at average 3 months of age, and castrated at 6 months. Liver tissues were collected from bulls (n = 10) and steers (n = 10) of Korean cattle, and hepatic gene expression levels were measured using quantitative real-time polymerase chain reaction. We examined hepatic transcription levels of genes encoding enzymes for irreversible reactions in both gluconeogenesis and glycolysis as well as genes encoding enzymes for the utilization of several glucogenic substrates. Correlations between hepatic gene expression and carcass characteristics were performed to understand their associations. Results Castration increased the mRNA (3.6 fold; p<0.01) and protein levels (1.4 fold; p< 0.05) of pyruvate carboxylase and mitochondrial phosphoenolpyruvate carboxykinase genes (1.7 fold; p<0.05). Hepatic mRNA levels of genes encoding the glycolysis enzymes were not changed by castration. Castration increased mRNA levels of both lactate dehydrogenase A (1.5 fold; p<0.05) and lactate dehydrogenase B (2.2 fold; p<0.01) genes for lactate utilization. Castration increased mRNA levels of glycerol kinase (2.7 fold; p<0.05) and glycerol-3-phosphate dehydrogenase 1 (1.5 fold; p<0.05) genes for glycerol utilization. Castration also increased mRNA levels of propionyl-CoA carboxylase beta (mitochondrial) (3.5 fold; p<0.01) and acyl-CoA synthetase short chain family member 3 (1.3 fold; p = 0.06) genes for propionate incorporation. Conclusion Castration increases transcription levels of critical genes coding for enzymes involved in irreversible gluconeogenesis reactions from pyruvate to glucose and enzymes responsible for incorporation of glucogenic substrates including lactate, glycerol, and propionate. Hepatic gluconeogenic gene expression levels were associated with intramuscular fat deposition. PMID:29502393

Hepatic transcriptional changes in critical genes for gluconeogenesis following castration of bulls.

PubMed

Fassah, Dilla Mareistia; Jeong, Jin Young; Baik, Myunggi

2018-04-01

This study was performed to understand transcriptional changes in the genes involved in gluconeogenesis and glycolysis pathways following castration of bulls. Twenty Korean bulls were weaned at average 3 months of age, and castrated at 6 months. Liver tissues were collected from bulls (n = 10) and steers (n = 10) of Korean cattle, and hepatic gene expression levels were measured using quantitative real-time polymerase chain reaction. We examined hepatic transcription levels of genes encoding enzymes for irreversible reactions in both gluconeogenesis and glycolysis as well as genes encoding enzymes for the utilization of several glucogenic substrates. Correlations between hepatic gene expression and carcass characteristics were performed to understand their associations. Castration increased the mRNA (3.6 fold; p<0.01) and protein levels (1.4 fold; p< 0.05) of pyruvate carboxylase and mitochondrial phosphoenolpyruvate carboxykinase genes (1.7 fold; p<0.05). Hepatic mRNA levels of genes encoding the glycolysis enzymes were not changed by castration. Castration increased mRNA levels of both lactate dehydrogenase A (1.5 fold; p<0.05) and lactate dehydrogenase B (2.2 fold; p<0.01) genes for lactate utilization. Castration increased mRNA levels of glycerol kinase (2.7 fold; p<0.05) and glycerol-3-phosphate dehydrogenase 1 (1.5 fold; p<0.05) genes for glycerol utilization. Castration also increased mRNA levels of propionyl-CoA carboxylase beta (mitochondrial) (3.5 fold; p<0.01) and acyl-CoA synthetase short chain family member 3 (1.3 fold; p = 0.06) genes for propionate incorporation. Castration increases transcription levels of critical genes coding for enzymes involved in irreversible gluconeogenesis reactions from pyruvate to glucose and enzymes responsible for incorporation of glucogenic substrates including lactate, glycerol, and propionate. Hepatic gluconeogenic gene expression levels were associated with intramuscular fat deposition.

A novel gene encoding a TIG multiple domain protein is a positional candidate for autosomal recessive polycystic kidney disease.

PubMed

Xiong, Huaqi; Chen, Yongxiong; Yi, Yajun; Tsuchiya, Karen; Moeckel, Gilbert; Cheung, Joseph; Liang, Dan; Tham, Kyi; Xu, Xiaohu; Chen, Xing-Zhen; Pei, York; Zhao, Zhizhuang Jeo; Wu, Guanqing

2002-07-01

Autosomal recessive polycystic kidney disease (ARPKD) is a common hereditary renal cystic disease in infants and children. By genetic linkage analyses, the gene responsible for this disease, termed polycystic kidney and hepatic disease 1 (PKHD1), was mapped on human chromosome 6p21.1-p12, and has been further localized to a 1-cM genetic interval flanked by the D6S1714/D6S243 (telomeric) and D6S1024 (centromeric) markers. We recently identified a novel gene in this genetic interval from kidney cDNA, using cloning strategies. The gene PKHD1 (PKHD1-tentative) encodes a novel 3396-amino-acid protein with no apparent homology with any known proteins. We named its gene product "tigmin" because it contains multiple TIG domains, which usually are seen in proteins containing immunoglobulin-like folds. PKHD1 encodes an 11.6-kb transcript and is composed of 61 exons spanning an approximately 365-kb genomic region on chromosome 6p12-p11.2 adjacent to the marker D6S1714. Northern blot analyses demonstrated that the gene has discrete bands with one peak signal at approximately 11 kb, indicating that PKHD1 is likely to have multiple alternative transcripts. PKHD1 is highly expressed in adult and infant kidneys and weakly expressed in liver in northern blot analysis. This expression pattern parallels the tissue involvement observed in ARPKD. In situ hybridization analysis further revealed that the expression of PKHD1 in the kidney is mainly localized to the epithelial cells of the collecting duct, the specific tubular segment involved in cyst formation in ARPKD. These features of PKHD1 make it a strong positional candidate gene for ARPKD.

The alpha1-fetoprotein locus is activated by a nuclear receptor of the Drosophila FTZ-F1 family.

PubMed

Galarneau, L; Paré, J F; Allard, D; Hamel, D; Levesque, L; Tugwood, J D; Green, S; Bélanger, L

1996-07-01

The alpha1-fetoprotein (AFP) gene is located between the albumin and alpha-albumin genes and is activated by transcription factor FTF (fetoprotein transcription factor), presumed to transduce early developmental signals to the albumin gene cluster. We have identified FTF as an orphan nuclear receptor of the Drosophila FTZ-F1 family. FTF recognizes the DNA sequence 5'-TCAAGGTCA-3', the canonical recognition motif for FTZ-F1 receptors. cDNA sequence homologies indicate that rat FTF is the ortholog of mouse LRH-1 and Xenopus xFF1rA. Rodent FTF is encoded by a single-copy gene, related to the gene encoding steroidogenic factor 1 (SF-1). The 5.2-kb FTF transcript is translated from several in-frame initiator codons into FTF isoforms (54 to 64 kDa) which appear to bind DNA as monomers, with no need for a specific ligand, similar KdS (approximately equal 3 x 10(-10) M), and similar transcriptional effects. FTF activates the AFP promoter without the use of an amino-terminal activation domain; carboxy-terminus-truncated FTF exerts strong dominant negative effects. In the AFP promoter, FTF recruits an accessory trans-activator which imparts glucocorticoid reactivity upon the AFP gene. FTF binding sites are found in the promoters of other liver-expressed genes, some encoding liver transcription factors; FTF, liver alpha1-antitrypsin promoter factor LFB2, and HNF-3beta promoter factor UF2-H3beta are probably the same factor. FTF is also abundantly expressed in the pancreas and may exert differentiation functions in endodermal sublineages, similar to SF-1 in steroidogenic tissues. HepG2 hepatoma cells seem to express a mutated form of FTF.

GmHs1-1, encoding a calcineurin-like protein, controls hard-seededness in soybean

USDA-ARS?s Scientific Manuscript database

Loss of seed-coat impermeability was an essential step towards domestication of many leguminous crops for production of their highly nutritious seeds. Here we show that seed-coat impermeability in wild soybean is controlled by a single gene, Hard seededness 1 (Hs1), which encodes a calcineurin-like ...

Neurotactin functions in concert with other identified CAMs in growth cone guidance in Drosophila.

PubMed

Speicher, S; García-Alonso, L; Carmena, A; Martín-Bermudo, M D; de la Escalera, S; Jiménez, F

1998-02-01

We have isolated and characterized mutations in Drosophila neurotactin, a gene that encodes a cell adhesion protein widely expressed during neural development. Analysis of both loss and gain of gene function conditions during embryonic and postembryonic development revealed specific requirements for neurotactin during axon outgrowth, fasciculation, and guidance. Furthermore, embryos of some double mutant combinations of neurotactin and other genes encoding adhesion/signaling molecules, including neuroglian, derailed, and kekkon1, displayed phenotypic synergy. This result provides evidence for functional cooperativity in vivo between the adhesion and signaling pathways controlled by neurotactin and the other three genes.

Nucleotide sequence analysis reveals linked N-acetyl hydrolase, thioesterase, transport, and regulatory genes encoded by the bialaphos biosynthetic gene cluster of Streptomyces hygroscopicus.

PubMed Central

Raibaud, A; Zalacain, M; Holt, T G; Tizard, R; Thompson, C J

1991-01-01

Nucleotide sequence analysis of a 5,000-bp region of the bialaphos antibiotic production (bap) gene cluster defined five open reading frames (ORFs) which predicted structural genes in the order bah, ORF1, ORF2, and ORF3 followed by the regulatory gene, brpA (H. Anzai, T. Murakami, S. Imai, A. Satoh, K. Nagaoka, and C.J. Thompson, J. Bacteriol. 169:3482-3488, 1987). The four structural genes were translationally coupled and apparently cotranscribed from an undefined promoter(s) under the positive control of the brpA gene product. S1 mapping experiments indicated that brpA was transcribed by two promoters (brpAp1 and brpAp2) which initiate transcription 150 and 157 bp upstream of brp A within an intergenic region and at least one promoter further upstream within the bap gene cluster (brpAp3). All three transcripts were present at low levels during exponential growth and increased just before the stationary phase. The levels of the brpAp3 band continued to increase at the onset of stationary phase, whereas brpAp1-and brpAp2-protected fragments showed no further change. BrpA contained a possible helix-turn-helix motif at its C terminus which was similar to the C-terminal regulatory motif found in the receiver component of a family of two-component transcriptional activator proteins. This motif was not associated with the N-terminal domain conserved in other members of the family. The structural gene cluster sequenced began with bah, encoding a bialaphos acetylhydrolase which removes the N-acetyl group from bialaphos as one of the final steps in the biosynthetic pathway. The observation that Bah was similar to a rat and to a bacterial (Acinetobacter calcoaceticus) lipase probably reflects the fact that the ester bonds of triglycerides and the amide bond linking acetate to phosphinothricin are similar and hydrolysis is catalyzed by structurally related enzymes. This was followed by two regions encoding ORF1 and ORF2 which were similar to each other (48% nucleotide identity, 31% amino acid identity), as well as to GrsT, a protein encoded by a gene located adjacent to gramicidin S synthetase in Bacillus brevis, and to vertebrate (mallard duck and rat) thioesterases. The amino acid sequence and hydrophobicity profile of ORF3 indicated that it was related to a family of membrane transport proteins. It was strikingly similar to the citrate uptake protein encoded by the transposon Tn3411. Images PMID:2066341

Molecular Characterization of Human Respiratory Syncytial Virus in the Philippines, 2012-2013.

PubMed

Malasao, Rungnapa; Okamoto, Michiko; Chaimongkol, Natthawan; Imamura, Tadatsugu; Tohma, Kentaro; Dapat, Isolde; Dapat, Clyde; Suzuki, Akira; Saito, Mayuko; Saito, Mariko; Tamaki, Raita; Pedrera-Rico, Gay Anne Granada; Aniceto, Rapunzel; Quicho, Reynaldo Frederick Negosa; Segubre-Mercado, Edelwisa; Lupisan, Socorro; Oshitani, Hitoshi

2015-01-01

Human respiratory syncytial virus (HRSV) is a major cause of acute lower respiratory tract infections in infants and children worldwide. We performed molecular analysis of HRSV among infants and children with clinical diagnosis of severe pneumonia in four study sites in the Philippines, including Biliran, Leyte, Palawan, and Metro Manila from June 2012 to July 2013. Nasopharyngeal swabs were collected and screened for HRSV using real-time polymerase chain reaction (PCR). Positive samples were tested by conventional PCR and sequenced for the second hypervariable region (2nd HVR) of the G gene. Among a total of 1,505 samples, 423 samples were positive for HRSV (28.1%), of which 305 (72.1%) and 118 (27.9%) were identified as HRSV-A and HRSV-B, respectively. Two genotypes of HRSV-A, NA1 and ON1, were identified during the study period. The novel ON1 genotype with a 72-nucleotide duplication in 2nd HVR of the G gene increased rapidly and finally became the predominant genotype in 2013 with an evolutionary rate higher than the NA1 genotype. Moreover, in the ON1 genotype, we found positive selection at amino acid position 274 (p<0.05) and massive O- and N-glycosylation in the 2nd HVR of the G gene. Among HRSV-B, BA9 was the predominant genotype circulating in the Philippines. However, two sporadic cases of GB2 genotype were found, which might share a common ancestor with other Asian strains. These findings suggest that HRSV is an important cause of severe acute respiratory infection among children in the Philippines and revealed the emergence and subsequent predominance of the ON1 genotype and the sporadic detection of the GB2 genotype. Both genotypes were detected for the first time in the Philippines.

Molecular Characterization of Human Respiratory Syncytial Virus in the Philippines, 2012-2013

PubMed Central

Malasao, Rungnapa; Okamoto, Michiko; Chaimongkol, Natthawan; Imamura, Tadatsugu; Tohma, Kentaro; Dapat, Isolde; Dapat, Clyde; Suzuki, Akira; Saito, Mayuko; Saito, Mariko; Tamaki, Raita; Pedrera-Rico, Gay Anne Granada; Aniceto, Rapunzel; Quicho, Reynaldo Frederick Negosa; Segubre-Mercado, Edelwisa; Lupisan, Socorro; Oshitani, Hitoshi

2015-01-01

Human respiratory syncytial virus (HRSV) is a major cause of acute lower respiratory tract infections in infants and children worldwide. We performed molecular analysis of HRSV among infants and children with clinical diagnosis of severe pneumonia in four study sites in the Philippines, including Biliran, Leyte, Palawan, and Metro Manila from June 2012 to July 2013. Nasopharyngeal swabs were collected and screened for HRSV using real-time polymerase chain reaction (PCR). Positive samples were tested by conventional PCR and sequenced for the second hypervariable region (2nd HVR) of the G gene. Among a total of 1,505 samples, 423 samples were positive for HRSV (28.1%), of which 305 (72.1%) and 118 (27.9%) were identified as HRSV-A and HRSV-B, respectively. Two genotypes of HRSV-A, NA1 and ON1, were identified during the study period. The novel ON1 genotype with a 72-nucleotide duplication in 2nd HVR of the G gene increased rapidly and finally became the predominant genotype in 2013 with an evolutionary rate higher than the NA1 genotype. Moreover, in the ON1 genotype, we found positive selection at amino acid position 274 (p<0.05) and massive O- and N-glycosylation in the 2nd HVR of the G gene. Among HRSV-B, BA9 was the predominant genotype circulating in the Philippines. However, two sporadic cases of GB2 genotype were found, which might share a common ancestor with other Asian strains. These findings suggest that HRSV is an important cause of severe acute respiratory infection among children in the Philippines and revealed the emergence and subsequent predominance of the ON1 genotype and the sporadic detection of the GB2 genotype. Both genotypes were detected for the first time in the Philippines. PMID:26540236

Stannous Fluoride Effects on Gene Expression of Streptococcus mutans and Actinomyces viscosus.

PubMed

Shi, Y; Li, R; White, D J; Biesbrock, A R

2018-02-01

A genome-wide transcriptional analysis was performed to elucidate the bacterial cellular response of Streptococcus mutans and Actinomyces viscosus to NaF and SnF 2 . The minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) of SnF 2 were predetermined before microarray study. Gene expression profiling microarray experiments were carried out in the absence (control) and presence (experimental) of 10 ppm and 100 ppm Sn 2+ (in the form of SnF 2 ) and fluoride controls for 10-min exposures (4 biological replicates/treatment). These Sn 2+ levels and treatment time were chosen because they have been shown to slow bacterial growth of S. mutans (10 ppm) and A. viscosus (100 ppm) without affecting cell viability. All data generated by microarray experiments were analyzed with bioinformatics tools by applying the following criteria: 1) a q value should be ≤0.05, and 2) an absolute fold change in transcript level should be ≥1.5. Microarray results showed SnF 2 significantly inhibited several genes encoding enzymes of the galactose pathway upon a 10-min exposure versus a negative control: lacA and lacB (A and B subunits of the galactose-6-P isomerase), lacC (tagatose-6-P kinase), lacD (tagatose-1,6-bP adolase), galK (galactokinase), galT (galactose-1-phosphate uridylyltransferase), and galE (UDP-glucose 4-epimerase). A gene fruK encoding fructose-1-phosphate kinase in the fructose pathway was also significantly inhibited. Several genes encoding fructose/mannose-specific enzyme IIABC components in the phosphotransferase system (PTS) were also downregulated, as was ldh encoding lactate dehydrogenase, a key enzyme involved in lactic acid synthesis. SnF 2 downregulated the transcription of most key enzyme genes involved in the galactose pathway and also suppressed several key genes involved in the PTS, which transports sugars into the cell in the first step of glycolysis.

Characterization of key triacylglycerol biosynthesis processes in rhodococci

DOE PAGES

Amara, Sawsan; Seghezzi, Nicolas; Otani, Hiroshi; ...

2016-04-29

In this study, oleaginous microorganisms have considerable potential for biofuel and commodity chemical production. Under nitrogen-limitation, Rhodococcus jostii RHA1 grown on benzoate, an analog of lignin depolymerization products, accumulated triacylglycerols (TAGs) to 55% of its dry weight during transition to stationary phase, with the predominant fatty acids being C16:0 and C17:0. Transcriptomic analyses of RHA1 grown under conditions of N-limitation and N-excess revealed 1,826 dysregulated genes. Genes whose transcripts were more abundant under N-limitation included those involved in ammonium assimilation, benzoate catabolism, fatty acid biosynthesis and the methylmalonyl-CoA pathway. Of the 16 atf genes potentially encoding diacylglycerol O-acyltransferases, atf8 transcriptsmore » were the most abundant during N-limitation (~50-fold more abundant than during N-excess). Consistent with Atf8 being a physiological determinant of TAG accumulation, a Δ atf8 mutant accumulated 70% less TAG than wild-type RHA1 while atf8 overexpression increased TAG accumulation 20%. Genes encoding type-2 phosphatidic acid phosphatases were not significantly expressed. By contrast, three genes potentially encoding phosphatases of the haloacid dehalogenase superfamily and that cluster with, or are fused with other Kennedy pathway genes were dysregulated. Overall, these findings advance our understanding of TAG metabolism in mycolic acid-containing bacteria and provide a framework to engineer strains for increased TAG production.« less

Characterization of key triacylglycerol biosynthesis processes in rhodococci

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

Amara, Sawsan; Seghezzi, Nicolas; Otani, Hiroshi

In this study, oleaginous microorganisms have considerable potential for biofuel and commodity chemical production. Under nitrogen-limitation, Rhodococcus jostii RHA1 grown on benzoate, an analog of lignin depolymerization products, accumulated triacylglycerols (TAGs) to 55% of its dry weight during transition to stationary phase, with the predominant fatty acids being C16:0 and C17:0. Transcriptomic analyses of RHA1 grown under conditions of N-limitation and N-excess revealed 1,826 dysregulated genes. Genes whose transcripts were more abundant under N-limitation included those involved in ammonium assimilation, benzoate catabolism, fatty acid biosynthesis and the methylmalonyl-CoA pathway. Of the 16 atf genes potentially encoding diacylglycerol O-acyltransferases, atf8 transcriptsmore » were the most abundant during N-limitation (~50-fold more abundant than during N-excess). Consistent with Atf8 being a physiological determinant of TAG accumulation, a Δ atf8 mutant accumulated 70% less TAG than wild-type RHA1 while atf8 overexpression increased TAG accumulation 20%. Genes encoding type-2 phosphatidic acid phosphatases were not significantly expressed. By contrast, three genes potentially encoding phosphatases of the haloacid dehalogenase superfamily and that cluster with, or are fused with other Kennedy pathway genes were dysregulated. Overall, these findings advance our understanding of TAG metabolism in mycolic acid-containing bacteria and provide a framework to engineer strains for increased TAG production.« less

Cloning and expression analysis of FaPR-1 gene in strawberry

NASA Astrophysics Data System (ADS)

Mo, Fan; Luo, Ya; Ge, Cong; Mo, Qin; Ling, Yajie; Luo, Shu; Tang, Haoru

2018-04-01

The FaPR-1 gene was cloned by RT-PCR from `Benihoppe' strawberry and its bioinformatics analysis was conducted. The results showed that the open reading frame was 483 bp encoding encoding l60 amino acids which protein molecular weight and theoretical isoelectricity were 17854.17 and 8.72 respectively. Subcellular localization prediction shows that this gene is located extracellularly. By comparing strawberry FaPR-l and other plant Pathogenesis-related protein, homology and phylogenetic tree construction showed that the homology with grapes, peach is relatively close. In the treatments of ABA, sucrose and the mixture of the two, the expression of FaPR-1 in strawberry fruit were significantly increased.

Characterization of a Gene Cluster Involved in 4-Chlorocatechol Degradation by Pseudomonas reinekei MT1▿

PubMed Central

Cámara, Beatriz; Nikodem, Patricia; Bielecki, Piotr; Bobadilla, Roberto; Junca, Howard; Pieper, Dietmar H.

2009-01-01

Pseudomonas reinekei MT1 has previously been reported to degrade 4- and 5-chlorosalicylate by a pathway with 4-chlorocatechol, 3-chloromuconate, 4-chloromuconolactone, and maleylacetate as intermediates, and a gene cluster channeling various salicylates into an intradiol cleavage route has been reported. We now report that during growth on 5-chlorosalicylate, besides a novel (chloro)catechol 1,2-dioxygenase, C12OccaA, a novel (chloro)muconate cycloisomerase, MCIccaB, which showed features not yet reported, was induced. This cycloisomerase, which was practically inactive with muconate, evolved for the turnover of 3-substituted muconates and transforms 3-chloromuconate into equal amounts of cis-dienelactone and protoanemonin, suggesting that it is a functional intermediate between chloromuconate cycloisomerases and muconate cycloisomerases. The corresponding genes, ccaA (C12OccaA) and ccaB (MCIccaB), were located in a 5.1-kb genomic region clustered with genes encoding trans-dienelactone hydrolase (ccaC) and maleylacetate reductase (ccaD) and a putative regulatory gene, ccaR, homologous to regulators of the IclR-type family. Thus, this region includes genes sufficient to enable MT1 to transform 4-chlorocatechol to 3-oxoadipate. Phylogenetic analysis showed that C12OccaA and MCIccaB are only distantly related to previously described catechol 1,2-dioxygenases and muconate cycloisomerases. Kinetic analysis indicated that MCIccaB and the previously identified C12OsalD, rather than C12OccaA, are crucial for 5-chlorosalicylate degradation. Thus, MT1 uses enzymes encoded by a completely novel gene cluster for degradation of chlorosalicylates, which, together with a gene cluster encoding enzymes for channeling salicylates into the ortho-cleavage pathway, form an effective pathway for 4- and 5-chlorosalicylate mineralization. PMID:19465655

Crystal structures of OrfX2 and P47 from a Botulinum neurotoxin OrfX-type gene cluster.

PubMed

Gustafsson, Robert; Berntsson, Ronnie P-A; Martínez-Carranza, Markel; El Tekle, Geniver; Odegrip, Richard; Johnson, Eric A; Stenmark, Pål

2017-11-01

Botulinum neurotoxins are highly toxic substances and are all encoded together with one of two alternative gene clusters, the HA or the OrfX gene cluster. Very little is known about the function and structure of the proteins encoded in the OrfX gene cluster, which in addition to the toxin contains five proteins (OrfX1, OrfX2, OrfX3, P47, and NTNH). We here present the structures of OrfX2 and P47, solved to 2.1 and 1.8 Å, respectively. We show that they belong to the TULIP protein superfamily, which are often involved in lipid binding. OrfX1 and OrfX2 were both found to bind phosphatidylinositol lipids. © 2017 Federation of European Biochemical Societies.

Evolutionary Characteristics of Missing Proteins: Insights into the Evolution of Human Chromosomes Related to Missing-Protein-Encoding Genes.

PubMed

Xu, Aishi; Li, Guang; Yang, Dong; Wu, Songfeng; Ouyang, Hongsheng; Xu, Ping; He, Fuchu

2015-12-04

Although the "missing protein" is a temporary concept in C-HPP, the biological information for their "missing" could be an important clue in evolutionary studies. Here we classified missing-protein-encoding genes into two groups, the genes encoding PE2 proteins (with transcript evidence) and the genes encoding PE3/4 proteins (with no transcript evidence). These missing-protein-encoding genes distribute unevenly among different chromosomes, chromosomal regions, or gene clusters. In the view of evolutionary features, PE3/4 genes tend to be young, spreading at the nonhomology chromosomal regions and evolving at higher rates. Interestingly, there is a higher proportion of singletons in PE3/4 genes than the proportion of singletons in all genes (background) and OTCSGs (organ, tissue, cell type-specific genes). More importantly, most of the paralogous PE3/4 genes belong to the newly duplicated members of the paralogous gene groups, which mainly contribute to special biological functions, such as "smell perception". These functions are heavily restricted into specific type of cells, tissues, or specific developmental stages, acting as the new functional requirements that facilitated the emergence of the missing-protein-encoding genes during evolution. In addition, the criteria for the extremely special physical-chemical proteins were first set up based on the properties of PE2 proteins, and the evolutionary characteristics of those proteins were explored. Overall, the evolutionary analyses of missing-protein-encoding genes are expected to be highly instructive for proteomics and functional studies in the future.

Gene family encoding the major toxins of lethal Amanita mushrooms

PubMed Central

Hallen, Heather E.; Luo, Hong; Scott-Craig, John S.; Walton, Jonathan D.

2007-01-01

Amatoxins, the lethal constituents of poisonous mushrooms in the genus Amanita, are bicyclic octapeptides. Two genes in A. bisporigera, AMA1 and PHA1, directly encode α-amanitin, an amatoxin, and the related bicyclic heptapeptide phallacidin, a phallotoxin, indicating that these compounds are synthesized on ribosomes and not by nonribosomal peptide synthetases. α-Amanitin and phallacidin are synthesized as proproteins of 35 and 34 amino acids, respectively, from which they are predicted to be cleaved by a prolyl oligopeptidase. AMA1 and PHA1 are present in other toxic species of Amanita section Phalloidae but are absent from nontoxic species in other sections. The genomes of A. bisporigera and A. phalloides contain multiple sequences related to AMA1 and PHA1. The predicted protein products of this family of genes are characterized by a hypervariable “toxin” region capable of encoding a wide variety of peptides of 7–10 amino acids flanked by conserved sequences. Our results suggest that these fungi have a broad capacity to synthesize cyclic peptides on ribosomes. PMID:18025465

Mutations in the gene encoding the Sigma 2 subunit of the adaptor protein 1 complex, AP1S2, cause X-linked mental retardation.

PubMed

Tarpey, Patrick S; Stevens, Claire; Teague, Jon; Edkins, Sarah; O'Meara, Sarah; Avis, Tim; Barthorpe, Syd; Buck, Gemma; Butler, Adam; Cole, Jennifer; Dicks, Ed; Gray, Kristian; Halliday, Kelly; Harrison, Rachel; Hills, Katy; Hinton, Jonathon; Jones, David; Menzies, Andrew; Mironenko, Tatiana; Perry, Janet; Raine, Keiran; Richardson, David; Shepherd, Rebecca; Small, Alexandra; Tofts, Calli; Varian, Jennifer; West, Sofie; Widaa, Sara; Yates, Andy; Catford, Rachael; Butler, Julia; Mallya, Uma; Moon, Jenny; Luo, Ying; Dorkins, Huw; Thompson, Deborah; Easton, Douglas F; Wooster, Richard; Bobrow, Martin; Carpenter, Nancy; Simensen, Richard J; Schwartz, Charles E; Stevenson, Roger E; Turner, Gillian; Partington, Michael; Gecz, Jozef; Stratton, Michael R; Futreal, P Andrew; Raymond, F Lucy

2006-12-01

In a systematic sequencing screen of the coding exons of the X chromosome in 250 families with X-linked mental retardation (XLMR), we identified two nonsense mutations and one consensus splice-site mutation in the AP1S2 gene on Xp22 in three families. Affected individuals in these families showed mild-to-profound mental retardation. Other features included hypotonia early in life and delay in walking. AP1S2 encodes an adaptin protein that constitutes part of the adaptor protein complex found at the cytoplasmic face of coated vesicles located at the Golgi complex. The complex mediates the recruitment of clathrin to the vesicle membrane. Aberrant endocytic processing through disruption of adaptor protein complexes is likely to result from the AP1S2 mutations identified in the three XLMR-affected families, and such defects may plausibly cause abnormal synaptic development and function. AP1S2 is the first reported XLMR gene that encodes a protein directly involved in the assembly of endocytic vesicles.

Widespread unidirectional transfer of mitochondrial DNA: a case in western Palaearctic water frogs.

PubMed

Plötner, J; Uzzell, T; Beerli, P; Spolsky, C; Ohst, T; Litvinchuk, S N; Guex, G-D; Reyer, H-U; Hotz, H

2008-05-01

Interspecies transfer of mitochondrial (mt) DNA is a common phenomenon in plants, invertebrates and vertebrates, normally linked with hybridization of closely related species in zones of sympatry or parapatry. In central Europe, in an area north of 48 degrees N latitude and between 8 degrees and 22 degrees E longitude, western Palaearctic water frogs show massive unidirectional introgression of mtDNA: 33.7% of 407 Rana ridibunda possessed mtDNA specific for Rana lessonae. By contrast, no R. lessonae with R. ridibunda mtDNA was observed. That R. ridibunda with introgressed mitochondrial genomes were found exclusively within the range of the hybrid Rana esculenta and that most hybrids had lessonae mtDNA (90.4% of 335 individuals investigated) is evidence that R. esculenta serves as a vehicle for transfer of lessonae mtDNA into R. ridibunda. Such introgression has occurred several times independently. The abundance and wide distribution of individuals with introgressed mitochondrial genomes show that R. lessonae mt genomes work successfully in a R. ridibunda chromosomal background despite their high sequence divergence from R. ridibunda mtDNAs (14.2-15.2% in the ND2/ND3 genes). Greater effectiveness of enzymes encoded by R. lessonae mtDNA may be advantageous to individuals of R. ridibunda and probably R. esculenta in the northern parts of their ranges.

A Novel KCNJ11 Mutation Associated with Transient Neonatal Diabetes

PubMed Central

Gole, Evangelia; Oikonomou, Stavroula; Ellard, Sian; De Franco, Elisa; Karavanaki, Kyriaki

2018-01-01

Neonatal diabetes mellitus (NDM) is a rare type of monogenic diabetes that presents in the first 6 months of life. Activating mutations in the KCNJ11 gene encoding for the Kir6.2 subunit of the ATP-sensitive potassium (KATP ) channel can lead to transient NDM (TNDM) or to permanent NDM (PNDM). A female infant presented on the 22nd day of life with severe hyperglycemia and ketoacidosis (glucose: 907mg/dL, blood gas pH: 6.84, HCO3: 6 mmol/L). She was initially managed with intravenous (IV) fluids and IV insulin. Ketoacidosis resolved within 48 hours and she was started on subcutaneous insulin injections with intermediate acting insulin NPH twice daily requiring initially 0.75-1.35 IU/kg/d. Pre-prandial C-peptide levels were 0.51 ng/mL (normal: 1.77-4.68). Insulin requirements were gradually reduced and insulin administration was discontinued at the age of 10 months with subsequent normal glucose and HbA1c levels. C-peptide levels normalized (pre-prandial: 1.6 ng/mL, postprandial: 2 ng/mL). Genetic analysis identified a novel missense mutation (p.Pro254Gln) in the KCNJ11 gene. We report a novel KCNJ11 mutation in a patient who presented in the first month of life with a phenotype of NDM that subsided at the age of 10 months. It is likely that the novel p.P254Q mutation results in mild impairment of the KATP channel function leading to TNDM. PMID:28943514

Identification of a Novel GLA Gene Mutation, p.Ile239Met, in Fabry Disease With a Predominant Cardiac Phenotype.

PubMed

Csányi, Beáta; Hategan, Lidia; Nagy, Viktória; Obál, Izabella; Varga, Edina T; Borbás, János; Tringer, Annamária; Eichler, Sabrina; Forster, Tamás; Rolfs, Arndt; Sepp, Róbert

2017-05-31

Fabry disease (FD) is an X-linked inherited lysosomal storage disorder caused by mutations in the GLA gene, encoding for the enzyme α-galactosidase A. Although hundreds of mutations in the GLA gene have been described, many of them are variants of unknown significance. Here we report a novel GLA mutation, p.Ile239Met, identified in a large Hungarian three-generation family with FD. A 69 year-old female index patient with a clinical history of renal failure, hypertrophic cardiomyopathy, and 2nd degree AV block was screened for mutation in the GLA gene. Genetic screening identified a previously unreported heterozygous mutation in exon 5 of the GLA gene (c.717A>G; p.Ile239Met). Family screening indicated that altogether 6 family members carried the mutation (5 females, 1 male, average age: 55 ± 16 years). Three family members, including the index patient, manifested the cardiac phenotype of hypertrophic cardiomyopathy, while two other family members were diagnosed with left ventricular hypertrophy. Taking affection status as the presence of hypertrophic cardiomyopathy, left ventricular hypertrophy or elevated lyso-Gb3 levels, all affected family members carried the mutation. Linkage analysis of the family gave a two-point LOD score of 2.01 between the affection status and the p.Ile239Met GLA mutation. Lyso-Gb3 levels were elevated in all carrier family members (range: 2.4-13.8 ng/mL; upper limit of normal +2STD: ≤ 1.8 ng/mL). The GLA enzyme level was markedly reduced in the affected male family member (< 0.2 µmol/L/hour; upper limit of normal ± 2STD: ≥ 2.6 µmol/L/hour). We conclude that the p. Ile239Met GLA mutation is a pathogenic mutation for FD associated with predominant cardiac phenotype.

[Genetic instability of probiotic characteristics in the Bifidobacterium longum subsp. longum B379M strain during cultivation and maintenance].

PubMed

Averina, O V; Nezametdinova, V Z; Alekseeva, M G; Danilenko, V N

2012-11-01

The stability of inheriting several genes in the Russian commercial strain Bifidobacterium longum subsp. longum B379M during cultivation and maintenance under laboratory conditions has been studied. The examined genes code for probiotic characteristics, such as utilization of several sugars (lacA2 gene, encoding beta-galactosidase; ara gene, encoding arabinosidase; and galA gene, encoding arabinogalactan endo-beta-galactosidase); synthesis of bacteriocins (lans gene, encoding lanthionine synthetase); and mobile gene tet(W), conferring resistance to the antibiotic tetracycline. The other gene families studied include the genes responsible for signal transduction and adaptation to stress conditions in the majority of bacteria (serine/threonine protein kinases and the toxin-antitoxin systems of MazEF and RelBE types) and transcription regulators (genes encoding WhiB family proteins). Genomic DNA was analyzed by PCR using specially selected primers. A loss of the genes galA and tet(W) has been shown. It is proposed to expand the requirements on probiotic strains, namely, to control retention of the key probiotic genes using molecular biological methods.

Isolated gene encoding an enzyme with UDP-glucose pyrophosphorylase and phosphoglucomutase activities from Cyclotella cryptica

DOEpatents

Jarvis, Eric E.; Roessler, Paul G.

1999-01-01

The present invention relates to a cloned gene which encodes an enzyme, the purified enzyme, and the applications and products resulting from the use of the gene and enzyme. The gene, isolated from Cyclotella cryptica, encodes a multifunctional enzyme that has both UDP-glucose pyrophosphorylase and phosphoglucomutase activities.

Human Genomic Signatures of Brain Oscillations During Memory Encoding.

PubMed

Berto, Stefano; Wang, Guang-Zhong; Germi, James; Lega, Bradley C; Konopka, Genevieve

2018-05-01

Memory encoding is an essential step for all learning. However, the genetic and molecular mechanisms underlying human memory encoding remain poorly understood, and how this molecular framework permits the emergence of specific patterns of brain oscillations observed during mnemonic processing is unknown. Here, we directly compare intracranial electroencephalography recordings from the neocortex in individuals performing an episodic memory task with human gene expression from the same areas. We identify genes correlated with oscillatory memory effects across 6 frequency bands. These genes are enriched for autism-related genes and have preferential expression in neurons, in particular genes encoding synaptic proteins and ion channels, supporting the idea that the genes regulating voltage gradients are involved in the modulation of oscillatory patterns during successful memory encoding across brain areas. Memory-related genes are distinct from those correlated with other forms of cognitive processing and resting state fMRI. These data are the first to identify correlations between gene expression and active human brain states as well as provide a molecular window into memory encoding oscillations in the human brain.

Combined molecular and morphological phylogenetic analyses of the New Zealand wolf spider genus Anoteropsis (Araneae: Lycosidae).

PubMed

Vink, Cor J; Paterson, Adrian M

2003-09-01

Datasets from the mitochondrial gene regions NADH dehydrogenase subunit I (ND1) and cytochrome c oxidase subunit I (COI) of the 20 species in the New Zealand wolf spider (Lycosidae) genus Anoteropsis were generated. Sequence data were phylogenetically analysed using parsimony and maximum likelihood analyses. The phylogenies generated from the ND1 and COI sequence data and a previously generated morphological dataset were significantly congruent (p<0.001). Sequence data were combined with morphological data and phylogenetically analysed using parsimony. The ND1 region sequenced included part of tRNA(Leu(CUN)), which appears to have an unstable amino-acyl arm and no TpsiC arm in lycosids. Analyses supported the existence of five species groups within Anoteropsis and the monophyly of species represented by multiple samples. A radiation of Anoteropsis species within the last five million years is inferred from the ND1 and COI likelihood phylograms, habitat and geological data, which also indicates that Anoteropsis arrived in New Zealand some time after it separated from Gondwana.

Coordinated rates of evolution between interacting plastid and nuclear genes in Geraniaceae.

PubMed

Zhang, Jin; Ruhlman, Tracey A; Sabir, Jamal; Blazier, J Chris; Jansen, Robert K

2015-03-01

Although gene coevolution has been widely observed within individuals and between different organisms, rarely has this phenomenon been investigated within a phylogenetic framework. The Geraniaceae is an attractive system in which to study plastid-nuclear genome coevolution due to the highly elevated evolutionary rates in plastid genomes. In plants, the plastid-encoded RNA polymerase (PEP) is a protein complex composed of subunits encoded by both plastid (rpoA, rpoB, rpoC1, and rpoC2) and nuclear genes (sig1-6). We used transcriptome and genomic data for 27 species of Geraniales in a systematic evaluation of coevolution between genes encoding subunits of the PEP holoenzyme. We detected strong correlations of dN (nonsynonymous substitutions) but not dS (synonymous substitutions) within rpoB/sig1 and rpoC2/sig2, but not for other plastid/nuclear gene pairs, and identified the correlation of dN/dS ratio between rpoB/C1/C2 and sig1/5/6, rpoC1/C2 and sig2, and rpoB/C2 and sig3 genes. Correlated rates between interacting plastid and nuclear sequences across the Geraniales could result from plastid-nuclear genome coevolution. Analyses of coevolved amino acid positions suggest that structurally mediated coevolution is not the major driver of plastid-nuclear coevolution. The detection of strong correlation of evolutionary rates between SIG and RNAP genes suggests a plausible explanation for plastome-genome incompatibility in Geraniaceae. © 2015 American Society of Plant Biologists. All rights reserved.

Functional Analysis of the Lactobacillus casei BL23 Sortases

PubMed Central

Muñoz-Provencio, Diego; Rodríguez-Díaz, Jesús; Collado, María Carmen; Langella, Philippe; Bermúdez-Humarán, Luis G.

2012-01-01

Sortases are a class of enzymes that anchor surface proteins to the cell wall of Gram-positive bacteria. Lactobacillus casei BL23 harbors four sortase genes, two belonging to class A (srtA1 and srtA2) and two belonging to class C (srtC1 and srtC2). Class C sortases were clustered with genes encoding their putative substrates that were homologous to the SpaEFG and SpaCBA proteins that encode mucus adhesive pili in Lactobacillus rhamnosus GG. Twenty-three genes encoding putative sortase substrates were identified in the L. casei BL23 genome with unknown (35%), enzymatic (30%), or adhesion-related (35%) functions. Strains disrupted in srtA1, srtA2, srtC1, and srtC2 and an srtA1 srtA2 double mutant were constructed. The transcription of all four sortase encoding genes was detected, but only the mutation of srtA1 resulted in a decrease in bacterial surface hydrophobicity. The β-N-acetyl-glucosaminidase and cell wall proteinase activities of whole cells diminished in the srtA1 mutant and, to a greater extent, in the srtA1 srtA2 double mutant. Cell wall anchoring of the staphylococcal NucA reporter protein fused to a cell wall sorting sequence was also affected in the srtA mutants, and the percentages of adhesion to Caco-2 and HT-29 intestinal epithelial cells were reduced for the srtA1 srtA2 strain. Mutations in srtC1 or srtC2 result in an undetectable phenotype. Together, these results suggest that SrtA1 is the housekeeping sortase in L. casei BL23 and SrtA2 would carry out redundant or complementary functions that become evident when SrtA1 activity is absent. PMID:23042174

Bacteriophage-based Vectors for Site-specific Insertion of DNA in the Chromosome of Corynebacteria

PubMed Central

Oram, Mark; Woolston, Joelle E.; Jacobson, Andrew D.; Holmes, Randall K.; Oram, Diana M.

2007-01-01

In Corynebacterium diphtheriae, diphtheria toxin is encoded by the tox gene of some temperate corynephages such as β. β-like corynephages are capable of inserting into the C. diphtheriae chromosome at two specific sites, attB1 and attB2. Transcription of the phage-encoded tox gene, and many chromosomally-encoded genes, is regulated by the DtxR protein in response to Fe2+ levels. Characterizing DtxR-dependent gene regulation is pivotal in understanding diphtheria pathogenesis and mechanisms of iron-dependent gene expression; although this has been hampered by a lack of molecular genetic tools in C. diphtheriae and related Coryneform species. To expand the systems for genetic manipulation of C. diphtheriae, we constructed plasmid vectors capable of integrating into the chromosome. These plasmids contain the β-encoded attP site and the DIP0182 integrase gene of C. diphtheriae NCTC13129. When these vectors were delivered to the cytoplasm of non-lysogenic C. diphtheriae, they integrated into either the attB1 or attB2 sites with comparable frequency. Lysogens were also transformed with these vectors, by virtue of the second attB site. An integrated vector carrying an intact dtxR gene complemented the mutant phenotypes of a C. diphtheriae ΔdtxR strain. Additionally, strains of β-susceptible C. ulcerans, and C. glutamicum, a species non-permissive for β, were each transformed with these vectors. This work significantly extends the tools available for targeted transformation of both pathogenic and non-pathogenic Corynebacterium species. PMID:17275217

Colon and Endometrial Cancers with Mismatch Repair Deficiency can Arise from Somatic, Rather Than Germline, Mutations

PubMed Central

Haraldsdottir, Sigurdis; Hampel, Heather; Tomsic, Jerneja; Frankel, Wendy L.; Pearlman, Rachel; de la Chapelle, Albert; Pritchard, Colin C.

2014-01-01

Background & Aims Patients with Lynch syndrome carry germline mutations in single alleles of genes encoding the MMR proteins MLH1, MSH2, MSH6 and PMS2; when the second allele becomes mutated, cancer can develop. Increased screening for Lynch syndrome has identified patients with tumors that have deficiency in MMR, but no germline mutations in genes encoding MMR proteins. We investigated whether tumors with deficient MMR had acquired somatic mutations in patients without germline mutations in MMR genes using next-generation sequencing. Methods We analyzed blood and tumor samples from 32 patients with colorectal or endometrial cancer who participated in Lynch syndrome screening studies in Ohio and were found to have tumors with MMR deficiency (based on microsatellite instability and/or absence of MMR proteins in immunohistochemical analysis, without hypermethylation of MLH1), but no germline mutations in MMR genes. Tumor DNA was sequenced for MLH1, MSH2, MSH6, PMS2, EPCAM, POLE and POLD1 with ColoSeq and mutation frequencies were established. Results Twenty-two of 32 patients (69%) were found to have two somatic (tumor) mutations in MMR genes encoding proteins that were lost from tumor samples, based on immunohistochemistry. Of the 10 tumors without somatic mutations in MMR genes, 3 had somatic mutations with possible loss of heterozygosity that could lead to MMR deficiency, 6 were found to be false-positive results (19%), and 1 had no mutations known to be associated with MMR deficiency. All of the tumors found to have somatic MMR mutations were of the hypermutated phenotype (>12 mutations/Mb); 6 had mutation frequencies >200 per Mb, and 5 of these had somatic mutations in POLE, which encodes a DNA polymerase. Conclusions Some patients are found to have tumors with MMR deficiency during screening for Lynch syndrome, yet have no identifiable germline mutations in MMR genes. We found that almost 70% of these patients acquire somatic mutations in MMR genes, leading to a hypermutated phenotype of tumor cells. Patients with colon or endometrial cancers with MMR deficiency not explained by germline mutations might undergo analysis for tumor mutations in MMR genes, to guide future surveillance guidelines. PMID:25194673

Leber's hereditary optic neuropathy is associated with mitochondrial ND6 T14502C mutation

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

Zhao, Fuxin; Zhejiang Provincial Key Laboratory of Medical Genetics, School of Life Sciences, Wenzhou Medical College, Wenzhou, Zhejiang 325003; Guan, Minqiang

2009-11-20

We report here the clinical, genetic, and molecular characterization of three Chinese families with Leber's hereditary optic neuropathy (LHON). There were variable severity and age of onset in visual impairment among these families. Strikingly, there were extremely low penetrances of visual impairment in these Chinese families. Sequence analysis of complete mitochondrial genomes in these pedigrees showed the homoplasmic T14502C (I58V) mutation, which localized at a highly conserved isoleucine at position 58 of ND6, and distinct sets of mtDNA polymorphisms belonging to haplogroups M10a, F1a1, and H2. The occurrence of T14502C mutation in these several genetically unrelated subjects affected by visualmore » impairment strongly indicates that this mutation is involved in the pathogenesis of visual impairment. Here, mtDNA variants I187T in the ND1, A122V in CO1, S99A in the A6, and V254I in CO3 exhibited an evolutionary conservation, indicating a potential modifying role in the development of visual impairment associated with T14502C mutation in those families. Furthermore, nuclear modifier gene(s) or environmental factor(s) may play a role in the phenotypic manifestation of the LHON-associated T14502C mutation in these Chinese families.« less

Bone Collagen: New Clues to its Mineralization Mechanism From Recessive Osteogenesis Imperfecta

PubMed Central

Eyre, David R.; Ann Weis, Mary

2013-01-01

Until 2006 the only mutations known to cause osteogenesis imperfecta (OI) were in the two genes coding for type I collagen chains. These dominant mutations affecting the expression or primary sequence of collagen α1(I) and α2(I) chains account for over 90% of OI cases. Since then a growing list of mutant genes causing the 5–10% of recessive cases has rapidly emerged. They include CRTAP, LEPRE1 and PPIB, which encode three proteins forming the prolyl 3-hydroxylase complex; PLOD2 and FKBP10, which encode respectively lysyl hydroxylase 2 and a foldase required for its activity in forming mature cross-links in bone collagen; SERPIN H1, which encodes the collagen chaperone HSP47; SERPIN F1, which encodes pigment epithelium-derived factor required for osteoid mineralization; and BMP1, which encodes the type I procollagen C-propeptidase. All cause fragile bone in infancy, which can include over-mineralization or under-mineralization defects as well as abnormal collagen post-translational modifications. Consistently both dominant and recessive variants lead to abnormal cross-linking chemistry in bone collagen. These recent discoveries strengthen the potential for a common pathogenic mechanism of misassembled collagen fibrils. Of the new genes identified, eight encode proteins required for collagen post-translational modification, chaperoning of newly synthesized collagen chains into native molecules or transport through the endoplasmic reticulum and Golgi for polymerization, cross-linking and mineralization. In reviewing these findings, we conclude that a common theme is emerging in the pathogenesis of brittle bone disease of mishandled collagen assembly with important insights on post-translational features of bone collagen that have evolved to optimize it as a biomineral template. PMID:23508630

Extensive diversification of IgD-, IgY-, and truncated IgY(δFc)-encoding genes in the red-eared turtle (Trachemys scripta elegans).

PubMed

Li, Lingxiao; Wang, Tao; Sun, Yi; Cheng, Gang; Yang, Hui; Wei, Zhiguo; Wang, Ping; Hu, Xiaoxiang; Ren, Liming; Meng, Qingyong; Zhang, Ran; Guo, Ying; Hammarström, Lennart; Li, Ning; Zhao, Yaofeng

2012-10-15

IgY(ΔFc), containing only CH1 and CH2 domains, is expressed in the serum of some birds and reptiles, such as ducks and turtles. The duck IgY(ΔFc) is produced by the same υ gene that expresses the intact IgY form (CH1-4) using different transcriptional termination sites. In this study, we show that intact IgY and IgY(ΔFc) are encoded by distinct genes in the red-eared turtle (Trachemys scripta elegans). At least eight IgY and five IgY(ΔFc) transcripts were found in a single turtle. Together with Southern blotting, our data suggest that multiple genes encoding both IgY forms are present in the turtle genome. Both of the IgY forms were detected in the serum using rabbit polyclonal Abs. In addition, we show that multiple copies of the turtle δ gene are present in the genome and that alternative splicing is extensively involved in the generation of both the secretory and membrane-bound forms of the IgD H chain transcripts. Although a single μ gene was identified, the α gene was not identified in this species.

Cloning and sequence analysis of the invertase gene INV 1 from the yeast Pichia anomala.

PubMed

Pérez, J A; Rodríguez, J; Rodríguez, L; Ruiz, T

1996-02-01

A genomic library from the yeast Pichia anomala has been constructed and employed to clone the gene encoding the sucrose-hydrolysing enzyme invertase by complementation of a sucrose non-fermenting mutant of Saccharomyces cerevisiae. The cloned gene, INV1, was sequenced and found to encode a polypeptide of 550 amino acids which contained a 22 amino-acid signal sequence and ten potential glycosylation sites. The amino-acid sequence shows significant identity with other yeast invertases and also with Kluyveromyces marxianus inulinase, a yeast beta-fructofuranosidase which has a different substrate specificity. The nucleotide sequences of the 5' and 3' non-coding regions were found to contain several consensus motifs probably involved in the initiation and termination of gene transcription.

Genomes of ubiquitous marine and hypersaline Hydrogenovibrio, Thiomicrorhabdus and Thiomicrospira spp. encode a diversity of mechanisms to sustain chemolithoautotrophy in heterogeneous environments.

PubMed

Scott, Kathleen M; Williams, John; Porter, Cody M B; Russel, Sydney; Harmer, Tara L; Paul, John H; Antonen, Kirsten M; Bridges, Megan K; Camper, Gary J; Campla, Christie K; Casella, Leila G; Chase, Eva; Conrad, James W; Cruz, Mercedez C; Dunlap, Darren S; Duran, Laura; Fahsbender, Elizabeth M; Goldsmith, Dawn B; Keeley, Ryan F; Kondoff, Matthew R; Kussy, Breanna I; Lane, Marannda K; Lawler, Stephanie; Leigh, Brittany A; Lewis, Courtney; Lostal, Lygia M; Marking, Devon; Mancera, Paola A; McClenthan, Evan C; McIntyre, Emily A; Mine, Jessica A; Modi, Swapnil; Moore, Brittney D; Morgan, William A; Nelson, Kaleigh M; Nguyen, Kimmy N; Ogburn, Nicholas; Parrino, David G; Pedapudi, Anangamanjari D; Pelham, Rebecca P; Preece, Amanda M; Rampersad, Elizabeth A; Richardson, Jason C; Rodgers, Christina M; Schaffer, Brent L; Sheridan, Nancy E; Solone, Michael R; Staley, Zachery R; Tabuchi, Maki; Waide, Ramond J; Wanjugi, Pauline W; Young, Suzanne; Clum, Alicia; Daum, Chris; Huntemann, Marcel; Ivanova, Natalia; Kyrpides, Nikos; Mikhailova, Natalia; Palaniappan, Krishnaveni; Pillay, Manoj; Reddy, T B K; Shapiro, Nicole; Stamatis, Dimitrios; Varghese, Neha; Woyke, Tanja; Boden, Rich; Freyermuth, Sharyn K; Kerfeld, Cheryl A

2018-03-09

Chemolithoautotrophic bacteria from the genera Hydrogenovibrio, Thiomicrorhabdus and Thiomicrospira are common, sometimes dominant, isolates from sulfidic habitats including hydrothermal vents, soda and salt lakes and marine sediments. Their genome sequences confirm their membership in a deeply branching clade of the Gammaproteobacteria. Several adaptations to heterogeneous habitats are apparent. Their genomes include large numbers of genes for sensing and responding to their environment (EAL- and GGDEF-domain proteins and methyl-accepting chemotaxis proteins) despite their small sizes (2.1-3.1 Mbp). An array of sulfur-oxidizing complexes are encoded, likely to facilitate these organisms' use of multiple forms of reduced sulfur as electron donors. Hydrogenase genes are present in some taxa, including group 1d and 2b hydrogenases in Hydrogenovibrio marinus and H. thermophilus MA2-6, acquired via horizontal gene transfer. In addition to high-affinity cbb 3 cytochrome c oxidase, some also encode cytochrome bd-type quinol oxidase or ba 3 -type cytochrome c oxidase, which could facilitate growth under different oxygen tensions, or maintain redox balance. Carboxysome operons are present in most, with genes downstream encoding transporters from four evolutionarily distinct families, which may act with the carboxysomes to form CO 2 concentrating mechanisms. These adaptations to habitat variability likely contribute to the cosmopolitan distribution of these organisms. © 2018 Society for Applied Microbiology and John Wiley & Sons Ltd.

DNA types of aspermic Fasciola species in Japan.

PubMed

Ichikawa, Madoka; Iwata, Noriyuki; Itagaki, Tadashi

2010-10-01

In order to reveal DNA types of aspermic Fasciola forms in Japan, Fasciola specimens obtained from eight prefectures that had not been previously reported were analyzed for DNA of ribosomal internal transcribed spacer 1 (ITS1) and mitochondrial NADH dehydrogenase 1 (ND1) gene. Five combinations in DNA types of both ITS1 and ND1 were revealed from the results of this study and previous studies. The DNA type Fsp2, which is identical to that of F. gigantica in both ITS1 and ND1, was the most predominant in Japan, followed by Fsp1, which is the same DNA type as that of F. hepatica. Fasciola forms with Fsp1 mainly occurred in the northern region of Japan and those with Fsp2 were mainly in the western region. The founder effect related to migration of definitive host and susceptibility of intermediate host snail might play an important role in both geographical distribution and frequency of DNA types in Japanese Fasciola specimens.

Regulation of HSD17B1 and SRD5A1 in lymphocytes.

PubMed

Zhou, Z; Speiser, P W

1999-11-01

We previously reported lymphocyte expression of genes encoding enzymes required for steroid metabolism; however, only 17beta-HSD and 5alpha-reductase showed significant enzyme activity. We now investigate regulation of lymphocyte expression for genes encoding 17beta-HSD and 5alpha-reductase. Cultured human T and B lymphoid cell lines and peripheral blood mononuclear cells were treated with known regulators of steroidogenic gene expression including forskolin, PMA, ionomycin, various steroids, interleukin (IL)-4, and IL-6. Treatment with 10 or 50 microM forskolin resulted in a 20-60% reduction of expression for HSD17B1 (encoding 17beta-HSD I) in T and B lymphoid cell lines and peripheral blood mononuclear cells, although such a change was not observed in the expression of SRD5A1 (encoding 5alpha-reductase I). No significant changes were found when cells were treated for 24 h with various concentrations of PMA or ionomycin. Incubation with 10(-9) to 10(-7) M androstenedione or estradiol increased expression of HSD17B1, while testosterone decreased the expression of this gene. SRD5A1 expression was increased in the presence of 5alpha-DHT although no consistent changes were observed when the cells were treated with testosterone. Other steroids, including dexamethasone, progesterone, and 6-hydroxypregnanolone, produced no effects on expression of either HSD17B1 or SRD5A1. Treatment with 0.1-10 ng/ml of IL-4 or IL-6 also did not effect significant changes in gene expression. These data implicate the involvement of the cAMP-protein kinase signal transduction pathway in regulating lymphocyte expression of HSD17B1. Furthermore, it appears that lymphocyte HSD17B1 and SRD5A1 are regulated to some extent by specific steroids. Copyright 1999 Academic Press.

Phylogenetic analysis of the Pacific cutthroat trout (Oncorhynchus clarki ssp.: Salmonidae) based on partial mtDNA ND4 sequences: a closer look at the highly fragmented inland species.

PubMed

Wilson, Wade D; Turner, Thomas F

2009-08-01

The genus Oncorhynchus includes Pacific salmon and trout (anadromous and land-locked) species of the western United States and Mexico. All species and subspecies in this group are threatened, endangered, sensitive, or species of conservation concern in portions of their native ranges. To examine the relationships of the species within Oncorhynchus we sequenced a 768 bp fragment of the protein-encoding ND4 mtDNA region. We included all six recognized subspecies of O. clarki (cutthroat trout), O. gilaegilae (Gila trout) and O. g. apache (Apache trout). Gene trees from likelihood and Bayesian phylogenetic analyses revealed that Salvelinus was the sister group to Oncorhynchus, and as expected based on previous studies, O. clarki was sister to a clade that consisted of O. mykiss plus O. g. gilae and O. g. apache. Within the cutthroat clade (O. clarki), the coastal form O. c. clarki was basal with the Rio Grande cutthroat (O. c. virginalis) most derived. Divergence dating based on a fossil calibration molecular clock showed the oldest clade (mean node age) was O. masou ssp., which diverged roughly 7.6 MYA. Highest probability density intervals for divergence of O. masou overlapped with divergence (6.3 MYA) of Pacific salmon clades ((O. gorbuscha + O. nerka) and (O. tshawytscha + O. kisutch)). The Pacific trout clade ((O. mykiss + O. gilae ssp.) + (O. clarki ssp.)) diverged from the Pacific salmon around 6.3 MYA, with most of the diversification within the O. clarki clade occurring in the last 1 MY.

The predominant WT1 isoform (+KTS) encodes a DNA binding protein targeting the planar cell polarity gene Scribble in renal podocytes

PubMed Central

Wells, Julie; Rivera, Miguel N.; Kim, Woo Jae; Starbuck, Kristen; Haber, Daniel A.

2010-01-01

WT1 encodes a tumor suppressor, first identified by its inactivation in Wilms Tumor. While one WT1 splicing variant encodes a well-characterized zinc finger transcription factor, little is known about the function of the most prevalent WT1 isoform, whose DNA binding domain is disrupted by a three amino acid (KTS) insertion. Using cells which conditionally express WT1(+KTS), we undertook a genome-wide chromatin immunoprecipitation and cloning (ChIP-cloning) analysis to identify candidate WT1(+KTS) regulated promoters. We identified the planar cell polarity (PCP) gene Scribble (SCRB) as the first WT1(+KTS) target gene in podocytes of the kidney. WT1 and SCRB expression patterns overlap precisely in developing renal glomeruli of mice, and WT1(+KTS) binds to a 33 nucleotide region within the Scribble promoter in both mouse and human cell lines and kidneys. Together, our results support a role for the predominant WT1(+KTS) isoform in transcriptional regulation and suggest a link between the WT1-dependent tumor suppressor pathway and a key component of the planar cell polarity pathway. PMID:20571064

CARB-9, a Carbenicillinase Encoded in the VCR Region of Vibrio cholerae Non-O1, Non-O139 Belongs to a Family of Cassette-Encoded β-Lactamases†

PubMed Central

Petroni, Alejandro; Melano, Roberto G.; Saka, Héctor A.; Garutti, Alicia; Mange, Laura; Pasterán, Fernando; Rapoport, Melina; Miranda, Mariana; Faccone, Diego; Rossi, Alicia; Hoffman, Paul S.; Galas, Marcelo F.

2004-01-01

The gene blaCARB-9 was located in the Vibrio cholerae super-integron, but in a different location relative to blaCARB-7. CARB-9 (pI 5.2) conferred β-lactam MICs four to eight times lower than those conferred by CARB-7, differing at Ambler's positions V97I, L124F, and T228K. Comparison of the genetic environments of all reported blaCARB genes indicated that the CARB enzymes constitute a family of cassette-encoded β-lactamases. PMID:15388476

Physiological roles of pyruvate ferredoxin oxidoreductase and pyruvate formate-lyase in Thermoanaerobacterium saccharolyticum JW/SL-YS485

DOE PAGES

Zhou, Jilai; Olson, Daniel G.; Lanahan, Anthony A.; ...

2015-09-15

We report that Thermoanaerobacter saccharolyticum is a thermophilic microorganism that has been engineered to produce ethanol at high titer (30–70 g/L) and greater than 90 % theoretical yield. However, few genes involved in pyruvate to ethanol production pathway have been unambiguously identified. In T. saccharolyticum, the products of six putative pfor gene clusters and one pfl gene may be responsible for the conversion of pyruvate to acetyl-CoA. To gain insights into the physiological roles of PFOR and PFL, we studied the effect of deletions of several genes thought to encode these activities. We found that that pyruvate ferredoxin oxidoreductase enzymemore » (PFOR) is encoded by the pforA gene and plays a key role in pyruvate dissimilation. We further demonstrated that pyruvate formate-lyase activity (PFL) is encoded by the pfl gene. Although the pfl gene is normally expressed at low levels, it is crucial for biosynthesis in T. saccharolyticum. In pforA deletion strains, pfl expression increased and was able to partially compensate for the loss of PFOR activity. Deletion of both pforA and pfl resulted in a strain that required acetate and formate for growth and produced lactate as the primary fermentation product, achieving 88 % theoretical lactate yield. PFOR encoded by Tsac_0046 and PFL encoded by Tsac_0628 are only two routes for converting pyruvate to acetyl-CoA in T. saccharolyticum. The physiological role of PFOR is pyruvate dissimilation, whereas that of PFL is supplying C1 units for biosynthesis.« less

Use of essential gene, encoding prophobilinogen deaminase from extreme psychrophilic Colwellia sp. C1, to generate temperature-sensitive strain of Francisella novicida.

PubMed

Pankowski, J A

2016-08-01

Previously, several essential genes from psychrophilic bacteria have been substituted for their homologues in mesophilic bacterial pathogens to make the latter temperature sensitive. It has been noted that an essential ligA gene from an extreme psychrophile, Colwellia sp. C1, yielded a gene product that is inactivated at 27°C, the lowest that has been observed for any psychrophilic enzyme, and hypothesized that other essential proteins of that strain would also have low inactivation temperatures. This work describes the partial sequencing of the genome of Colwellia sp. C1 strain and the identification of 24 open reading frames encoding homologues of highly conserved bacterial essential genes. The gene encoding porphobilinogen deaminase (hemC), which is involved in the pathway of haem synthesis, has been tested for its ability to convert Francisella novicida into a temperature-sensitive strain. The hybrid strain carrying the C1-derived hemC gene exhibited a temperature-sensitive phenotype with a restrictive temperature of 36°C. These results support the conclusion that Colwellia sp. C1 is a rich source of heat-labile enzymes. The issue of biosafety is often raised when it comes to work with pathogenic organisms. The main concern is caused by the risk of researchers being exposed to infectious doses of dangerous microbes. This paper analyses essential genes identified in partial genomic sequence of the psychrophilic bacterium Collwelia sp. C1. These sequences can be used as a mean of generating temperature-sensitive strains of pathogenic bacteria. Such strains are incapable of surviving at the temperature of human body. This means they could be applied as vaccines or for safer work with dangerous organisms. © 2016 The Society for Applied Microbiology.

Identification and characterization of a second CD4-like gene in teleost fish.

PubMed

Dijkstra, Johannes Martinus; Somamoto, Tomonori; Moore, Lindsey; Hordvik, Ivar; Ototake, Mitsuru; Fischer, Uwe

2006-02-01

In fish, T cell subdivision is not well studied, although CD8 and CD4 homologues have been reported. This study describes a second teleost CD4-like gene, CD4-like 2 (CD4L-2). Two rainbow trout copies of this gene were found, -2a and -2b, encoding molecules sharing 81% aa identity. The 2a/2b duplication may be related to tetraploid ancestry of salmonid fishes. In the Fugu genome CD4L-2 lies head to tail with an earlier reported, very different CD4-like gene [Suetake, H., Araki, K., Suzuki, Y., 2004. Cloning, expression, and characterization of fugu CD4, the first ectothermic animal CD4. Immunogenetics 56, 368-374], which was designated CD4L-1 in the present article. The flanking genes of the Fugu CD4L-1 and CD4L-2 are reminiscent of the genes surrounding CD4 and LAG-3 in mammals. However, neither synteny nor phylogenetic analysis could decide between CD4 and LAG-3 identity for the fish CD4L genes. CD4L-1 and CD4L-2 share a tyrosine protein kinase p56(lck) binding motif in the cytoplasmic tail with CD4 but not with LAG-3. Trout CD4L-2 expression is highest in the thymus, similar to mammalian and chicken CD4, whereas Fugu CD4L-1 expression was highest in the spleen. However, CD4L-2 encodes only two IG-like domains, whereas CD4L-1, CD4 and LAG-3 encode four. The CD4-like genes 1 and 2 in fish apparently went through an evolution different from that of LAG-3 and CD4 in higher vertebrates.

Characterization of two chitin synthase genes of the red flour beetle, Tribolium castaneum, and alternate exon usage in one of the genes during development.

PubMed

Arakane, Yasuyuki; Hogenkamp, David G; Zhu, Yu Cheng; Kramer, Karl J; Specht, Charles A; Beeman, Richard W; Kanost, Michael R; Muthukrishnan, Subbaratnam

2004-03-01

Two chitin synthase (CHS) genes of the red flour beetle, Tribolium castaneum, were sequenced and their transcription patterns during development examined. By screening a BAC library of genomic DNA from T. castaneum (Tc) with a DNA probe encoding the catalytic domain of a putative Tribolium CHS, several clones that contained CHS genes were identified. Two distinct PCR products were amplified from these BAC clones and confirmed to be highly similar to CHS genes from other insects, nematodes and fungi. The DNA sequences of these genes, TcCHS1 and TcCHS2, were determined by amplification of overlapping PCR fragments from two of the BAC DNAs and mapped to different linkage groups. Each ORF was identified and full-length cDNAs were also amplified, cloned and sequenced. TcCHS1 and TcCHS2 encode transmembrane proteins of 1558 and 1464 amino acids, respectively. The TcCHS1 gene was found to use alternate exons, each encoding 59 amino acids, a feature not found in the TcCHS2 gene. During development, Tribolium expressed TcCHS1 predominantly in the embryonic and pupal stages, whereas TcCHS2 was prevalent in the late larval and adult stages. The alternate exon 8a of TcCHS1 was utilized over a much broader range of development than exon 8b. We propose that the two isoforms of the TcCHS1 enzyme are used predominantly for the formation of chitin in embryonic and pupal cuticles, whereas TcCHS2 is utilized primarily for the synthesis of peritrophic membrane-associated chitin in the midgut.

Method for increasing thermostability in cellulase ennzymes

DOEpatents

Adney, W.S.; Thomas, S.R.; Baker, J.O.; Himmel, M.E.; Chou, Y.C.

1998-01-27

The gene encoding Acidothermus cellulolyticus E1 endoglucanase is cloned and expressed in Pichia pastoris. A new modified E1 endoglucanase enzyme comprising the catalytic domain of the full size E1 enzyme demonstrates enhanced thermostability and is produced by two methods. The first method of producing the new modified E1 is proteolytic cleavage to remove the cellulose binding domain and linker peptide of the full size E1. The second method of producing the new modified E1 is genetic truncation of the gene encoding the full size E1 so that the catalytic domain is expressed in the expression product. 8 figs.

Characterization, genetic diversity, and evolutionary link of Cucumber mosaic virus strain New Delhi from India.

PubMed

Koundal, Vikas; Haq, Qazi Mohd Rizwanul; Praveen, Shelly

2011-02-01

The genome of Cucumber mosaic virus New Delhi strain (CMV-ND) from India, obtained from tomato, was completely sequenced and compared with full genome sequences of 14 known CMV strains from subgroups I and II, for their genetic diversity. Sequence analysis suggests CMV-ND shares maximum sequence identity at the nucleotide level with a CMV strain from Taiwan. Among all 15 strains of CMV, the encoded protein 2b is least conserved, whereas the coat protein (CP) is most conserved. Sequence identity values and phylogram results indicate that CMV-ND belongs to subgroup I. Based on the recombination detection program result, it appears that CMV is prone to recombination, and different RNA components of CMV-ND have evolved differently. Recombinational analysis of all 15 CMV strains detected maximum recombination breakpoints in RNA2; CP showed the least recombination sites.

Transposon mutagenesis and cloning analysis of the pathways for degradation of 2,4-dichlorophenoxyacetic acid and 3-chlorobenzoate in Alcaligenes eutrophus JMP134(pJP4).

PubMed Central

Don, R H; Weightman, A J; Knackmuss, H J; Timmis, K N

1985-01-01

Plasmid pJP4 permits its host bacterium, strain JMP134, to degrade and utilize as sole sources of carbon and energy 3-chlorobenzoate and 2,4-dichlorophenoxyacetic acid (R. H. Don and J. M. Pemberton, J. Bacteriol. 145:681-686, 1981). Mutagenesis of pJP4 by transposons Tn5 and Tn1771 enabled localization of five genes for enzymes involved in these catabolic pathways. Four of the genes, tfdB, tfdC, tfdD, and tfdE, encoded 2,4-dichlorophenol hydroxylase, dichlorocatechol 1,2-dioxygenase, chloromuconate cycloisomerase, and chlorodienelactone hydrolase, respectively. No function has been assigned to the fifth gene, tfdF, although it may encode a trans-chlorodiene-lactone isomerase. Inactivation of genes tfdC, tfdD, and tfdE, which encode the transformation of dichlorocatechol to chloromaleylacetic acid, prevented host strain JMP134 from degrading both 3-chlorobenzoate and 2,4-dichlorophenoxyacetic acid, which indicates that the pathways for these two substrates utilize common enzymes for the dissimilation of chlorocatechols. Studies with cloned catabolic genes from pJP4 indicated that whereas all essential steps in the degradation of 2,4-dichlorophenoxyacetic acid are plasmid encoded, the conversion of 3-chlorobenzoate to chlorocatechol is specified by chromosomal genes. PMID:2981813

A novel missense mutation in the NDP gene in a child with Norrie disease and severe neurological involvement including infantile spasms.

PubMed

Lev, Dorit; Weigl, Yuval; Hasan, Mariana; Gak, Eva; Davidovich, Michael; Vinkler, Chana; Leshinsky-Silver, Esther; Lerman-Sagie, Tally; Watemberg, Nathan

2007-05-01

Norrie disease (ND) is a rare X-linked recessive disorder characterized by congenital blindness and in some cases, mental retardation and deafness. Other neurological complications, particularly epilepsy, are rare. We report on a novel mutation identified in a patient with ND and profound mental retardation. The patient was diagnosed at the age of 6 months due to congenital blindness. At the age of 8 months he developed infantile spasms, which were diagnosed at 11 months as his EEG demonstrated hypsarrhythmia. Mutation analysis of the ND gene (NDP) of the affected child and his mother revealed a novel missense mutation at position c.134T > A resulting in amino acid change at codon V45E. To the best of our knowledge, such severe neurological involvement has not been previously reported in ND patients. The severity of the phenotype may suggest the functional importance of this site of the NDP gene.

Molecular and functional characterization of novel fructosyltransferases and invertases from Agave tequilana.

PubMed

Cortés-Romero, Celso; Martínez-Hernández, Aída; Mellado-Mojica, Erika; López, Mercedes G; Simpson, June

2012-01-01

Fructans are the main storage polysaccharides found in Agave species. The synthesis of these complex carbohydrates relies on the activities of specific fructosyltransferase enzymes closely related to the hydrolytic invertases. Analysis of Agave tequilana transcriptome data led to the identification of ESTs encoding putative fructosyltransferases and invertases. Based on sequence alignments and structure/function relationships, two different genes were predicted to encode 1-SST and 6G-FFT type fructosyltransferases, in addition, 4 genes encoding putative cell wall invertases and 4 genes encoding putative vacuolar invertases were also identified. Probable functions for each gene, were assigned based on conserved amino acid sequences and confirmed for 2 fructosyltransferases and one invertase by analyzing the enzymatic activity of recombinant Agave protein s expressed and purified from Pichia pastoris. The genome organization of the fructosyltransferase/invertase genes, for which the corresponding cDNA contained the complete open reading frame, was found to be well conserved since all genes were shown to carry a 9 bp mini-exon and all showed a similar structure of 8 exons/7 introns with the exception of a cell wall invertase gene which has 7 exons and 6 introns. Fructosyltransferase genes were strongly expressed in the storage organs of the plants, especially in vegetative stages of development and to lower levels in photosynthetic tissues, in contrast to the invertase genes where higher levels of expression were observed in leaf tissues and in mature plants.

Molecular and Functional Characterization of Novel Fructosyltransferases and Invertases from Agave tequilana

PubMed Central

Cortés-Romero, Celso; Martínez-Hernández, Aída; Mellado-Mojica, Erika; López, Mercedes G.; Simpson, June

2012-01-01

Fructans are the main storage polysaccharides found in Agave species. The synthesis of these complex carbohydrates relies on the activities of specific fructosyltransferase enzymes closely related to the hydrolytic invertases. Analysis of Agave tequilana transcriptome data led to the identification of ESTs encoding putative fructosyltransferases and invertases. Based on sequence alignments and structure/function relationships, two different genes were predicted to encode 1-SST and 6G-FFT type fructosyltransferases, in addition, 4 genes encoding putative cell wall invertases and 4 genes encoding putative vacuolar invertases were also identified. Probable functions for each gene, were assigned based on conserved amino acid sequences and confirmed for 2 fructosyltransferases and one invertase by analyzing the enzymatic activity of recombinant Agave protein s expressed and purified from Pichia pastoris. The genome organization of the fructosyltransferase/invertase genes, for which the corresponding cDNA contained the complete open reading frame, was found to be well conserved since all genes were shown to carry a 9 bp mini-exon and all showed a similar structure of 8 exons/7 introns with the exception of a cell wall invertase gene which has 7 exons and 6 introns. Fructosyltransferase genes were strongly expressed in the storage organs of the plants, especially in vegetative stages of development and to lower levels in photosynthetic tissues, in contrast to the invertase genes where higher levels of expression were observed in leaf tissues and in mature plants. PMID:22558253

Mice mutant for both Hoxa1 and Hoxb1 show extensive remodeling of the hindbrain and defects in craniofacial development.

PubMed

Rossel, M; Capecchi, M R

1999-11-01

The analysis of mice mutant for both Hoxa1 and Hoxb1 suggests that these two genes function together to pattern the hindbrain. Separately, mutations in Hoxa1 and Hoxb1 have profoundly different effects on hindbrain development. Hoxa1 mutations disrupt the rhombomeric organization of the hindbrain, whereas Hoxb1 mutations do not alter the rhombomeric pattern, but instead influence the fate of cells originating in rhombomere 4. We suggest that these differences are not the consequences of different functional roles for these gene products, but rather reflect differences in the kinetics of Hoxa1 and Hoxb1 gene expression. In strong support of the idea that Hoxa1 and Hoxb1 have overlapping functions, Hoxa1/Hoxb1 double mutant homozygotes exhibit a plethora of defects either not seen, or seen only in a very mild form, in mice mutant for only Hoxa1 or Hoxb1. Examples include: the loss of both rhombomeres 4 and 5, the selective loss of the 2(nd) branchial arch, and the loss of most, but not all, 2(nd) branchial arch-derived tissues. We suggest that the early role for both of these genes in hindbrain development is specification of rhombomere identities and that the aberrant development of the hindbrain in Hoxa1/Hoxb1 double mutants proceeds through two phases, the misspecification of rhombomeres within the hindbrain, followed subsequently by size regulation of the misspecified hindbrain through induction of apoptosis.

Cloning and Expression of the Erwinia carotovora subsp. carotovora Gene Encoding the Low-Molecular-Weight Bacteriocin Carocin S1▿

PubMed Central

Chuang, Duen-yau; Chien, Yung-chei; Wu, Huang-Pin

2007-01-01

The purpose of this study was to clone the carocin S1 gene and express it in a non-carocin-producing strain of Erwinia carotovora. A mutant, TH22-10, which produced a high-molecular-weight bacteriocin but not a low-molecular-weight bacteriocin, was obtained by Tn5 insertional mutagenesis using H-rif-8-2 (a spontaneous rifampin-resistant mutant of Erwinia carotovora subsp. carotovora 89-H-4). Using thermal asymmetric interlaced PCR, the DNA sequence from the Tn5 insertion site and the DNA sequence of the contiguous 2,280-bp region were determined. Two complete open reading frames (ORF), designated ORF2 and ORF3, were identified within the sequence fragment. ORF2 and ORF3 were identified with the carocin S1 genes, caroS1K (ORF2) and caroS1I (ORF3), which, respectively, encode a killing protein (CaroS1K) and an immunity protein (CaroS1I). These genes were homologous to the pyocin S3 gene and the pyocin AP41 gene. Carocin S1 was expressed in E. carotovora subsp. carotovora Ea1068 and replicated in TH22-10 but could not be expressed in Escherichia coli (JM101) because a consensus sequence resembling an SOS box was absent. A putative sequence similar to the consensus sequence for the E. coli cyclic AMP receptor protein binding site (−312 bp) was found upstream of the start codon. Production of this bacteriocin was also induced by glucose and lactose. The homology search results indicated that the carocin S1 gene (between bp 1078 and bp 1704) was homologous to the pyocin S3 and pyocin AP41 genes in Pseudomonas aeruginosa. These genes encode proteins with nuclease activity (domain 4). This study found that carocin S1 also has nuclease activity. PMID:17071754

Characterization and functional analysis of the MAL and MPH Loci for maltose utilization in some ale and lager yeast strains.

PubMed

Vidgren, Virve; Ruohonen, Laura; Londesborough, John

2005-12-01

Maltose and maltotriose are the major sugars in brewer's wort. Brewer's yeasts contain multiple genes for maltose transporters. It is not known which of these express functional transporters. We correlated maltose transport kinetics with the genotypes of some ale and lager yeasts. Maltose transport by two ale strains was strongly inhibited by other alpha-glucosides, suggesting the use of broad substrate specificity transporters, such as Agt1p. Maltose transport by three lager strains was weakly inhibited by other alpha-glucosides, suggesting the use of narrow substrate specificity transporters. Hybridization studies showed that all five strains contained complete MAL1, MAL2, MAL3, and MAL4 loci, except for one ale strain, which lacked a MAL2 locus. All five strains also contained both AGT1 (coding a broad specificity alpha-glucoside transporter) and MAL11 alleles. MPH genes (maltose permease homologues) were present in the lager but not in the ale strains. During growth on maltose, the lager strains expressed AGT1 at low levels and MALx1 genes at high levels, whereas the ale strains expressed AGT1 at high levels and MALx1 genes at low levels. MPHx expression was negligible in all strains. The AGT1 sequences from the ale strains encoded full-length (616 amino acid) polypeptides, but those from both sequenced lager strains encoded truncated (394 amino acid) polypeptides that are unlikely to be functional transporters. Thus, despite the apparently similar genotypes of these ale and lager strains revealed by hybridization, maltose is predominantly carried by AGT1-encoded transporters in the ale strains and by MALx1-encoded transporters in the lager strains.

MACF1 gene structure: a hybrid of plectin and dystrophin.

PubMed

Gong, T W; Besirli, C G; Lomax, M I

2001-11-01

Mammalian MACF1 (Macrophin1; previously named ACF7) is a giant cytoskeletal linker protein with three known isoforms that arise by alternative splicing. We isolated a 19.1-kb cDNA encoding a fourth isoform (MACF1-4) with a unique N-terminus. Instead of an N-terminal actin-binding domain found in the other three isoforms, MACF1-4 has eight plectin repeats. The MACF1 gene is located on human Chr 1p32, contains at least 102 exons, spans over 270 kb, and gives rise to four major isoforms with different N-termini. The genomic organization of the actin-binding domain is highly conserved in mammalian genes for both plectin and BPAG1. All eight plectin repeats are encoded by one large exon; this feature is similar to the genomic structure of plectin. The intron positions within spectrin repeats in MACF1 are very similar to those in the dystrophin gene. This demonstrates that MACF1 has characteristic features of genes for two classes of cytoskeletal proteins, i.e., plectin and dystrophin.

Systems-level analysis of risk genes reveals the modular nature of schizophrenia.

PubMed

Liu, Jiewei; Li, Ming; Luo, Xiong-Jian; Su, Bing

2018-05-19

Schizophrenia (SCZ) is a complex mental disorder with high heritability. Genetic studies (especially recent genome-wide association studies) have identified many risk genes for schizophrenia. However, the physical interactions among the proteins encoded by schizophrenia risk genes remain elusive and it is not known whether the identified risk genes converge on common molecular networks or pathways. Here we systematically investigated the network characteristics of schizophrenia risk genes using the high-confidence protein-protein interactions (PPI) from the human interactome. We found that schizophrenia risk genes encode a densely interconnected PPI network (P = 4.15 × 10 -31 ). Compared with the background genes, the schizophrenia risk genes in the interactome have significantly higher degree (P = 5.39 × 10 -11 ), closeness centrality (P = 7.56 × 10 -11 ), betweeness centrality (P = 1.29 × 10 -11 ), clustering coefficient (P = 2.22 × 10 -2 ), and shorter average shortest path length (P = 7.56 × 10 -11 ). Based on the densely interconnected PPI network, we identified 48 hub genes and 4 modules formed by highly interconnected schizophrenia genes. We showed that the proteins encoded by schizophrenia hub genes have significantly more direct physical interactions. Gene ontology (GO) analysis revealed that cell adhesion, cell cycle, immune system response, and GABR-receptor complex categories were enriched in the modules formed by highly interconnected schizophrenia risk genes. Our study reveals that schizophrenia risk genes encode a densely interconnected molecular network and demonstrates the modular nature of schizophrenia. Copyright © 2018 Elsevier B.V. All rights reserved.

Increased enzyme production under liquid culture conditions in the industrial fungus Aspergillus oryzae by disruption of the genes encoding cell wall α-1,3-glucan synthase.

PubMed

Miyazawa, Ken; Yoshimi, Akira; Zhang, Silai; Sano, Motoaki; Nakayama, Mayumi; Gomi, Katsuya; Abe, Keietsu

2016-09-01

Under liquid culture conditions, the hyphae of filamentous fungi aggregate to form pellets, which reduces cell density and fermentation productivity. Previously, we found that loss of α-1,3-glucan in the cell wall of the fungus Aspergillus nidulans increased hyphal dispersion. Therefore, here we constructed a mutant of the industrial fungus A. oryzae in which the three genes encoding α-1,3-glucan synthase were disrupted (tripleΔ). Although the hyphae of the tripleΔ mutant were not fully dispersed, the mutant strain did form smaller pellets than the wild-type strain. We next examined enzyme productivity under liquid culture conditions by transforming the cutinase-encoding gene cutL1 into A. oryzae wild-type and the tripleΔ mutant (i.e. wild-type-cutL1, tripleΔ-cutL1). A. oryzae tripleΔ-cutL1 formed smaller hyphal pellets and showed both greater biomass and increased CutL1 productivity compared with wild-type-cutL1, which might be attributable to a decrease in the number of tripleΔ-cutL1 cells under anaerobic conditions.

Characterization of Chicken Spleen Transcriptome after Infection with Salmonella enterica Serovar Enteritidis

PubMed Central

Matulova, Marta; Rajova, Jana; Vlasatikova, Lenka; Volf, Jiri; Stepanova, Hana; Havlickova, Hana; Sisak, Frantisek; Rychlik, Ivan

2012-01-01

In this study we were interested in identification of new markers of chicken response to Salmonella Enteritidis infection. To reach this aim, gene expression in the spleens of naive chickens and those intravenously infected with S. Enteritidis with or without previous oral vaccination was determined by 454 pyrosequencing of splenic mRNA/cDNA. Forty genes with increased expression at the level of transcription were identified. The most inducible genes encoded avidin (AVD), extracellular fatty acid binding protein (EXFABP), immune responsive gene 1 (IRG1), chemokine ah221 (AH221), trappin-6-like protein (TRAP6) and serum amyloid A (SAA). Using cDNA from sorted splenic B-lymphocytes, macrophages, CD4, CD8 and γδ T-lymphocytes, we found that the above mentioned genes were preferentially expressed in macrophages. AVD, EXFABP, IRG1, AH221, TRAP6 and SAA were induced also in the cecum of chickens orally infected with S. Enteritidis on day 1 of life or day 42 of life. Unusual results were obtained for the immunoglobulin encoding transcripts. Prior to the infection, transcripts coding for the constant parts of IgM, IgY, IgA and Ig light chain were detected in B-lymphocytes. However, after the infection, immunoglobulin encoding transcripts were expressed also by T-lymphocytes and macrophages. Expression of AVD, EXFABP, IRG1, AH221, TRAP6, SAA and all immunoglobulin genes can be therefore used for the characterization of the course of S. Enteritidis infection in chickens. PMID:23094107

Isolated gene encoding an enzyme with UDP-glucose pyrophosphorylase and phosphoglucomutase activities from Cyclotella cryptica

DOEpatents

Jarvis, E.E.; Roessler, P.G.

1999-07-27

The present invention relates to a cloned gene which encodes an enzyme, the purified enzyme, and the applications and products resulting from the use of the gene and enzyme. The gene, isolated from Cyclotella cryptica, encodes a multifunctional enzyme that has both UDP-glucose pyrophosphorylase and phosphoglucomutase activities. 8 figs.

Microarray Analyses of Gene Expression during Adventitious Root Development in Pinus contorta1[w

PubMed Central

Brinker, Monika; van Zyl, Leonel; Liu, Wenbin; Craig, Deborah; Sederoff, Ronald R.; Clapham, David H.; von Arnold, Sara

2004-01-01

In order to investigate the gene expression pattern during adventitious root development, RNA of Pinus contorta hypocotyls, pulse-treated with the auxin indole-3-butyric acid and harvested at distinct developmental time points of root development, was hybridized to microarrays containing 2,178 cDNAs from Pinus taeda. Over the period of observation of root development, the transcript levels of 220 genes changed significantly. During the root initiation phase, genes involved in cell replication and cell wall weakening and a transcript encoding a PINHEAD/ZWILLE-like protein were up-regulated, while genes related to auxin transport, photosynthesis, and cell wall synthesis were down-regulated. In addition, there were changes in transcript abundance of genes related to water stress. During the root meristem formation phase the transcript abundances of genes involved in auxin transport, auxin responsive transcription, and cell wall synthesis, and of a gene encoding a B-box zinc finger-like protein, increased, while those encoding proteins involved in cell wall weakening decreased. Changes of transcript abundance of genes related to water stress during the root meristem formation and root formation phase indicate that the plant roots had become functional in water transport. Simultaneously, genes involved in auxin transport were up-regulated, while genes related to cell wall modification were down-regulated. Finally, during the root elongation phase down-regulation of transcripts encoding proteins involved in cell replication and stress occurred. Based on the observed changes in transcript abundances, we suggest hypotheses about the relative importance of various physiological processes during the auxin-induced development of roots in P. contorta. PMID:15247392

Growth phase-dependent control of R27 conjugation is mediated by the interplay between the plasmid-encoded regulatory circuit TrhR/TrhY-HtdA and the cAMP regulon.

PubMed

Gibert, Marta; Paytubi, Sonia; Beltrán, Sergi; Juárez, Antonio; Balsalobre, Carlos; Madrid, Cristina

2016-12-01

Plasmids of the incompatibility group HI1 (IncHI1) have been isolated from several Gram-negative pathogens and are associated with the spread of multidrug resistance. Their conjugation is tightly regulated and it is inhibited at temperatures higher than 30°C, indicating that conjugation occurs outside warm-blooded hosts. Using R27, the prototype of IncHI1 plasmids, we report that plasmid transfer efficiency in E. coli strongly depends on the physiological state of the donor cells. Conjugation frequency is high when cells are actively growing, dropping sharply when cells enter the stationary phase of growth. Accordingly, our transcriptomic assays show significant downregulation of numerous R27 genes during the stationary phase, including several tra (transfer) genes. Growth phase-dependent regulation of tra genes transcription is independent of H-NS, a silencer of horizontal gene transfer, and ppGpp and RpoS, regulators of the stationary phase, but highly dependent on the plasmid-encoded regulatory circuit TrhR/TrhY-HtdA. The metabolic sensor cAMP, whose synthesis is chromosomally encoded, is also involved in the growth phase regulation of R27 conjugation by modulating htdA expression. Our data suggest that the involvement of regulators encoded by both chromosome and plasmid are required for efficient physiological control of IncHI1 plasmid conjugation. © 2016 Society for Applied Microbiology and John Wiley & Sons Ltd.

The VPH1 gene encodes a 95-kDa integral membrane polypeptide required for in vivo assembly and activity of the yeast vacuolar H(+)-ATPase.

PubMed

Manolson, M F; Proteau, D; Preston, R A; Stenbit, A; Roberts, B T; Hoyt, M A; Preuss, D; Mulholland, J; Botstein, D; Jones, E W

1992-07-15

Yeast vacuolar acidification-defective (vph) mutants were identified using the pH-sensitive fluorescence of 6-carboxyfluorescein diacetate (Preston, R. A., Murphy, R. F., and Jones, E. W. (1989) Proc. Natl. Acad. Sci. U.S.A. 86, 7027-7031). Vacuoles purified from yeast bearing the vph1-1 mutation had no detectable bafilomycin-sensitive ATPase activity or ATP-dependent proton pumping. The peripherally bound nucleotide-binding subunits of the vacuolar H(+)-ATPase (60 and 69 kDa) were no longer associated with vacuolar membranes yet were present in wild type levels in yeast whole cell extracts. The VPH1 gene was cloned by complementation of the vph1-1 mutation and independently cloned by screening a lambda gt11 expression library with antibodies directed against a 95-kDa vacuolar integral membrane protein. Deletion disruption of the VPH1 gene revealed that the VPH1 gene is not essential for viability but is required for vacuolar H(+)-ATPase assembly and vacuolar acidification. VPH1 encodes a predicted polypeptide of 840 amino acid residues (molecular mass 95.6 kDa) and contains six putative membrane-spanning regions. Cell fractionation and immunodetection demonstrate that Vph1p is a vacuolar integral membrane protein that co-purifies with vacuolar H(+)-ATPase activity. Multiple sequence alignments show extensive homology over the entire lengths of the following four polypeptides: Vph1p, the 116-kDa polypeptide of the rat clathrin-coated vesicles/synaptic vesicle proton pump, the predicted polypeptide encoded by the yeast gene STV1 (Similar To VPH1, identified as an open reading frame next to the BUB2 gene), and the TJ6 mouse immune suppressor factor.

Glutathione S-transferase-encoding gene as a potential probe for environmental bacterial isolates capable of degrading polycyclic aromatic hydrocarbons.

PubMed Central

Lloyd-Jones, G; Lau, P C

1997-01-01

Homologs of the glutathione S-transferase (GST)-encoding gene were identified in a collection of aromatic hydrocarbon-degrading Sphingomonas spp. isolated from New Zealand, Antarctica, and the United States by using PCR primers designed from the GST-encoding gene of Sphingomonas paucimobilis EPA505. Sequence analysis of PCR fragments generated from these isolates and of the GST gene amplified from DNA extracted from polycyclic aromatic hydrocarbon (PAH)-contaminated soil revealed a high degree of conservation, which may make the GST-encoding gene a potentially useful marker for PAH-degrading bacteria. PMID:9251217

Enterotoxin-encoding genes in Staphylococcus spp. from bulk goat milk.

PubMed

Lyra, Daniele G; Sousa, Francisca G C; Borges, Maria F; Givisiez, Patrícia E N; Queiroga, Rita C R E; Souza, Evandro L; Gebreyes, Wondwossen A; Oliveira, Celso J B

2013-02-01

Although Staphylococcus aureus has been implicated as the main Staphylococcus species causing human food poisoning, recent studies have shown that coagulase-negative Staphylococcus could also harbor enterotoxin-encoding genes. Such organisms are often present in goat milk and are the most important mastitis-causing agents. Therefore, this study aimed to investigate the occurrence of enterotoxin-encoding genes among coagulase-positive (CoPS) and coagulase-negative (CoNS) staphylococci isolated from raw goat milk produced in the semi-arid region of Paraiba, the most important region for goat milk production in Brazil. Enterotoxin-encoding genes were screened in 74 staphylococci isolates (30 CoPS and 44 CoNS) by polymerase chain reaction targeting the genes sea, seb, sec, sed, see, seg, seh, and sei. Enterotoxin-encoding genes were found in nine (12.2%) isolates, and four different genes (sea, sec, seg, and sei) were identified amongst the isolates. The most frequent genes were seg and sei, which were often found simultaneously in 44.5% of the isolates. The gene sec was the most frequent among the classical genes, and sea was found only in one isolate. All CoPS isolates (n=7) harboring enterotoxigenic genes were identified as S. aureus. The two coagulase-negative isolates were S. haemolyticus and S. hominis subsp. hominis and they harbored sei and sec genes, respectively. A higher frequency of enterotoxin-encoding genes was observed amongst CoPS (23.3%) than CoNS (4.5%) isolates (p<0.05), reinforcing the importance of S. aureus as a potential foodborne agent. However, the potential risk posed by CoNS in goat milk should not be ignored because it has a higher occurrence in goat milk and enterotoxin-encoding genes were detected in some isolates.

KNQ1, a Kluyveromyces lactis gene encoding a transmembrane protein, may be involved in iron homeostasis.

PubMed

Marchi, Emmanuela; Lodi, Tiziana; Donnini, Claudia

2007-08-01

The original purpose of the experiments described in this article was to identify, in the biotechnologically important yeast Kluyveromyces lactis, gene(s) that are potentially involved in oxidative protein folding within the endoplasmic reticulum (ER), which often represents a bottleneck for heterologous protein production. Because treatment with the membrane-permeable reducing agent dithiothreitol inhibits disulfide bond formation and mimics the reducing effect that the normal transit of folding proteins has in the ER environment, the strategy was to search for genes that conferred higher levels of resistance to dithiothreitol when present in multiple copies. We identified a gene (KNQ1) encoding a drug efflux permease for several toxic compounds that in multiple copies conferred increased dithiothreitol resistance. However, the KNQ1 product is not involved in the excretion of dithiothreitol or in recombinant protein secretion. We generated a knq1 null mutant, and showed that both overexpression and deletion of the KNQ1 gene resulted in increased resistance to dithiothreitol. KNQ1 amplification and deletion resulted in enhanced transcription of iron transport genes, suggesting, for the membrane-associated protein Knq1p, a new, unexpected role in iron homeostasis on which dithiothreitol tolerance may depend.

Expression analysis of the N-Myc downstream-regulated gene 1 indicates that myelinating Schwann cells are the primary disease target in hereditary motor and sensory neuropathy-Lom.

PubMed

Berger, Philipp; Sirkowski, Erich E; Scherer, Steven S; Suter, Ueli

2004-11-01

Mutations in the gene encoding N-myc downstream-regulated gene-1 (NDRG1) lead to truncations of the encoded protein and are associated with an autosomal recessive demyelinating neuropathy--hereditary motor and sensory neuropathy-Lom. NDRG1 protein is highly expressed in peripheral nerve and is localized in the cytoplasm of myelinating Schwann cells, including the paranodes and Schmidt-Lanterman incisures. In contrast, sensory and motor neurons as well as their axons lack NDRG1. NDRG1 mRNA levels in developing and injured adult sciatic nerves parallel those of myelin-related genes, indicating that the expression of NDRG1 in myelinating Schwann cells is regulated by axonal interactions. Oligodendrocytes also express NDRG1, and the subtle CNS deficits of affected patients may result from a lack of NDRG1 in these cells. Our data predict that the loss of NDRG1 leads to a Schwann cell autonomous phenotype resulting in demyelination, with secondary axonal loss.

CLA1, a novel gene required for chloroplast development, is highly conserved in evolution.

PubMed

Mandel, M A; Feldmann, K A; Herrera-Estrella, L; Rocha-Sosa, M; León, P

1996-05-01

An albino mutant designated cla1-1 (for "cloroplastos alterados', or "altered chloroplasts') has been isolated from a T-DNA-generated library of Arabidopsis thaliana. In cla1-1 plants, chloroplast development is arrested at an early stage. cla1-1 plants behave like wild-type in their capacity to etiolate and produce anthocyanins indicating that the light signal transduction pathway seems to be unaffected. Genetic and molecular analyses show that the disruption of a single gene, CLA1, by the T-DNA insertion is responsible for the mutant phenotype. RNA expression patterns indicate that CLA1 is positively regulated by light and that it has different effects on the steady-state RNA levels of some nuclear- and chloroplast-encoded photosynthetic genes. Although the specific function of the CLA1 gene is still unknown, it encodes a novel protein conserved in evolution between photosynthetic bacteria and plants which is essential for chloroplast development in Arabidopsis.