Science.gov

Sample records for allelic wrky genes

  1. Genome-wide investigation and transcriptome analysis of the WRKY gene family in Gossypium.

    PubMed

    Ding, Mingquan; Chen, Jiadong; Jiang, Yurong; Lin, Lifeng; Cao, YueFen; Wang, Minhua; Zhang, Yuting; Rong, Junkang; Ye, Wuwei

    2015-02-01

    WRKY transcription factors play important roles in various stress responses in diverse plant species. In cotton, this family has not been well studied, especially in relation to fiber development. Here, the genomes and transcriptomes of Gossypium raimondii and Gossypium arboreum were investigated to identify fiber development related WRKY genes. This represents the first comprehensive comparative study of WRKY transcription factors in both diploid A and D cotton species. In total, 112 G. raimondii and 109 G. arboreum WRKY genes were identified. No significant gene structure or domain alterations were detected between the two species, but many SNPs distributed unequally in exon and intron regions. Physical mapping revealed that the WRKY genes in G. arboreum were not located in the corresponding chromosomes of G. raimondii, suggesting great chromosome rearrangement in the diploid cotton genomes. The cotton WRKY genes, especially subgroups I and II, have expanded through multiple whole genome duplications and tandem duplications compared with other plant species. Sequence comparison showed many functionally divergent sites between WRKY subgroups, while the genes within each group are under strong purifying selection. Transcriptome analysis suggested that many WRKY genes participate in specific fiber development processes such as fiber initiation, elongation and maturation with different expression patterns between species. Complex WRKY gene expression such as differential Dt and At allelic gene expression in G. hirsutum and alternative splicing events were also observed in both diploid and tetraploid cottons during fiber development process. In conclusion, this study provides important information on the evolution and function of WRKY gene family in cotton species.

  2. Alternative Splicing of Rice WRKY62 and WRKY76 Transcription Factor Genes in Pathogen Defense1[OPEN

    PubMed Central

    Chen, Xujun; Zhou, Xiangui; Yang, Fang

    2016-01-01

    The WRKY family of transcription factors (TFs) functions as transcriptional activators or repressors in various signaling pathways. In this study, we discovered that OsWRKY62 and OsWRKY76, two genes of the WRKY IIa subfamily, undergo constitutive and inducible alternative splicing. The full-length OsWRKY62.1 and OsWRKY76.1 proteins formed homocomplexes and heterocomplexes, and the heterocomplex dominates in the nuclei when analyzed in Nicotiana benthamiana leaves. Transgenic overexpression of OsWRKY62.1 and OsWRKY76.1 in rice (Oryza sativa) enhanced plant susceptibility to the blast fungus Magnaporthe oryzae and the leaf blight bacterium Xanthomonas oryzae pv oryzae, whereas RNA interference and loss-of-function knockout plants exhibited elevated resistance. The dsOW62/76 and knockout lines of OsWRKY62 and OsWRKY76 also showed greatly increased expression of defense-related genes and the accumulation of phytoalexins. The ratio of full-length versus truncated transcripts changed in dsOW62/76 plants as well as in response to pathogen infection. The short alternative OsWRKY62.2 and OsWRKY76.2 isoforms could interact with each other and with full-length proteins. OsWRKY62.2 showed a reduced repressor activity in planta, and two sequence determinants required for the repressor activity were identified in the amino terminus of OsWRKY62.1. The amino termini of OsWRKY62 and OsWRKY76 splice variants also showed reduced binding to the canonical W box motif. These results not only enhance our understanding of the DNA-binding property, the repressor sequence motifs, and the negative feedback regulation of the IIa subfamily of WRKYs but also provide evidence for alternative splicing of WRKY TFs during the plant defense response. PMID:27208272

  3. WRKY transcription factor genes in wild rice Oryza nivara.

    PubMed

    Xu, Hengjian; Watanabe, Kenneth A; Zhang, Liyuan; Shen, Qingxi J

    2016-08-01

    The WRKY transcription factor family is one of the largest gene families involved in plant development and stress response. Although many WRKY genes have been studied in cultivated rice (Oryza sativa), the WRKY genes in the wild rice species Oryza nivara, the direct progenitor of O. sativa, have not been studied. O. nivara shows abundant genetic diversity and elite drought and disease resistance features. Herein, a total of 97 O. nivara WRKY (OnWRKY) genes were identified. RNA-sequencing demonstrates that OnWRKY genes were generally expressed at higher levels in the roots of 30-day-old plants. Bioinformatic analyses suggest that most of OnWRKY genes could be induced by salicylic acid, abscisic acid, and drought. Abundant potential MAPK phosphorylation sites in OnWRKYs suggest that activities of most OnWRKYs can be regulated by phosphorylation. Phylogenetic analyses of OnWRKYs support a novel hypothesis that ancient group IIc OnWRKYs were the original ancestors of only some group IIc and group III WRKYs. The analyses also offer strong support that group IIc OnWRKYs containing the HVE sequence in their zinc finger motifs were derived from group Ia WRKYs. This study provides a solid foundation for the study of the evolution and functions of WRKY genes in O. nivara.

  4. WRKY transcription factor genes in wild rice Oryza nivara

    PubMed Central

    Xu, Hengjian; Watanabe, Kenneth A.; Zhang, Liyuan; Shen, Qingxi J.

    2016-01-01

    The WRKY transcription factor family is one of the largest gene families involved in plant development and stress response. Although many WRKY genes have been studied in cultivated rice (Oryza sativa), the WRKY genes in the wild rice species Oryza nivara, the direct progenitor of O. sativa, have not been studied. O. nivara shows abundant genetic diversity and elite drought and disease resistance features. Herein, a total of 97 O. nivara WRKY (OnWRKY) genes were identified. RNA-sequencing demonstrates that OnWRKY genes were generally expressed at higher levels in the roots of 30-day-old plants. Bioinformatic analyses suggest that most of OnWRKY genes could be induced by salicylic acid, abscisic acid, and drought. Abundant potential MAPK phosphorylation sites in OnWRKYs suggest that activities of most OnWRKYs can be regulated by phosphorylation. Phylogenetic analyses of OnWRKYs support a novel hypothesis that ancient group IIc OnWRKYs were the original ancestors of only some group IIc and group III WRKYs. The analyses also offer strong support that group IIc OnWRKYs containing the HVE sequence in their zinc finger motifs were derived from group Ia WRKYs. This study provides a solid foundation for the study of the evolution and functions of WRKY genes in O. nivara. PMID:27345721

  5. Expression of TaWRKY44, a wheat WRKY gene, in transgenic tobacco confers multiple abiotic stress tolerances

    PubMed Central

    Wang, Xiatian; Zeng, Jian; Li, Ying; Rong, Xiaoli; Sun, Jiutong; Sun, Tao; Li, Miao; Wang, Lianzhe; Feng, Ying; Chai, Ruihong; Chen, Mingjie; Chang, Junli; Li, Kexiu; Yang, Guangxiao; He, Guangyuan

    2015-01-01

    The WRKY transcription factors have been reported to be involved in various plant physiological and biochemical processes. In this study, we successfully assembled 10 unigenes from expressed sequence tags (ESTs) of wheat and designated them as TaWRKY44–TaWRKY53, respectively. Among these genes, a subgroup I gene, TaWRKY44, was found to be upregulated by treatments with PEG6000, NaCl, 4°C, abscisic acid (ABA), H2O2 and gibberellin (GA). The TaWRKY44-GFP fusion protein was localized to the nucleus of onion epidermal cells, and TaWRKY44 was able to bind to the core DNA sequences of TTGACC and TTAACC in yeast. The N-terminal of TaWRKY44 showed transcriptional activation activity. Expression of TaWRKY44 in tobacco plants conferred drought and salt tolerance and transgenic tobacco exhibited a higher survival rate, relative water content (RWC), soluble sugar, proline and superoxide dismutase (SOD) content, as well as higher activities of catalase (CAT) and peroxidase (POD), but less ion leakage (IL), lower contents of malondialdehyde (MDA), and H2O2. In addition, expression of TaWRKY44 also increased the seed germination rate in the transgenic lines under osmotic stress conditions while exhibiting a lower H2O2 content and higher SOD, CAT, and POD activities. Expression of TaWRKY44 upregulated the expression of some reactive oxygen species (ROS)-related genes and stress-responsive genes in tobacco under osmotic stresses. These data demonstrate that TaWRKY44 may act as a positive regulator in drought/salt/osmotic stress responses by either efficient ROS elimination through direct or indirect activation of the cellular antioxidant systems or activation of stress-associated gene expression. PMID:26322057

  6. Genome-wide analysis of the WRKY gene family in physic nut (Jatropha curcas L.).

    PubMed

    Xiong, Wangdan; Xu, Xueqin; Zhang, Lin; Wu, Pingzhi; Chen, Yaping; Li, Meiru; Jiang, Huawu; Wu, Guojiang

    2013-07-25

    The WRKY proteins, which contain highly conserved WRKYGQK amino acid sequences and zinc-finger-like motifs, constitute a large family of transcription factors in plants. They participate in diverse physiological and developmental processes. WRKY genes have been identified and characterized in a number of plant species. We identified a total of 58 WRKY genes (JcWRKY) in the genome of the physic nut (Jatropha curcas L.). On the basis of their conserved WRKY domain sequences, all of the JcWRKY proteins could be assigned to one of the previously defined groups, I-III. Phylogenetic analysis of JcWRKY genes with Arabidopsis and rice WRKY genes, and separately with castor bean WRKY genes, revealed no evidence of recent gene duplication in JcWRKY gene family. Analysis of transcript abundance of JcWRKY gene products were tested in different tissues under normal growth condition. In addition, 47 WRKY genes responded to at least one abiotic stress (drought, salinity, phosphate starvation and nitrogen starvation) in individual tissues (leaf, root and/or shoot cortex). Our study provides a useful reference data set as the basis for cloning and functional analysis of physic nut WRKY genes.

  7. Transcription Factor WRKY62 Plays a Role in Pathogen Defense and Hypoxia-Responsive Gene Expression in Rice.

    PubMed

    Fukushima, Setsuko; Mori, Masaki; Sugano, Shoji; Takatsuji, Hiroshi

    2016-12-01

    WRKY62 is a transcriptional repressor regulated downstream of WRKY45, a central transcription factor of the salicylic acid signaling pathway in rice. Previously, WRKY62 was reported to regulate defense negatively. However, our expressional analysis using WRKY62-knockdown rice indicated that WRKY62 positively regulates defense genes, including diterpenoid phytoalexin biosynthetic genes and their transcriptional regulator DPF. Blast and leaf blight resistance tests also showed that WRKY62 is a positive defense regulator. Yeast two-hybrid, co-immunoprecipitation and gel-shift assays showed that WRKY45 and WRKY62 can form a heterodimer, as well as homodimers, that bind to W-boxes in the DPF promoter. In transient assays in rice sheaths, the simultaneous introduction of WRKY45 and WRKY62 as effectors resulted in a strong activation of the DPF promoter:hrLUC reporter gene, whereas the activity declined with excessive WRKY62. Thus, the WRKY45-WRKY62 heterodimer acts as a strong activator, while the WRKY62 homodimer acts as a repressor. While benzothiadiazole induced equivalent numbers of WRKY45 and WRKY62 transcripts, consistent with heterodimer formation and DPF activation, submergence and nitrogen replacement induced only WRKY62 transcripts, consistent with WRKY62 homodimer formation and DPF repression. Moreover, WRKY62 positively regulated hypoxia genes, implying a role forWRKY62 in the modulation of the 'trade-off' between defense and hypoxia responses.

  8. Transcriptomics-based identification of WRKY genes and characterization of a salt and hormone-responsive PgWRKY1 gene in Panax ginseng.

    PubMed

    Nuruzzaman, Mohammed; Cao, Hongzhe; Xiu, Hao; Luo, Tiao; Li, Jijia; Chen, Xianghui; Luo, Junli; Luo, Zhiyong

    2016-02-01

    WRKY proteins belong to a transcription factor (TF) family and play dynamic roles in many plant processes, including plant responses to abiotic and biotic stresses, as well as secondary metabolism. However, no WRKY gene in Panax ginseng C.A. Meyer has been reported to date. In this study, a number of WRKY unigenes from methyl jasmonate (MeJA)-treated adventitious root transcriptome of this species were identified using next-generation sequencing technology. A total of 48 promising WRKY unigenes encoding WRKY proteins were obtained by eliminating wrong and incomplete open reading frame (ORF). Phylogenetic analysis reveals 48 WRKY TFs, including 11 Group I, 36 Group II, and 1 Group III. Moreover, one MeJA-responsive unigene designated as PgWRKY1 was cloned and characterized. It contains an entire ORF of 1077 bp and encodes a polypeptide of 358 amino acid residues. The PgWRKY1 protein contains a single WRKY domain consisting of a conserved amino acid sequence motif WRKYGQK and a C2H2-type zinc-finger motif belonging to WRKY subgroup II-d. Subcellular localization of PgWRKY1-GFP fusion protein in onion and tobacco epidermis cells revealed that PgWRKY1 was exclusively present in the nucleus. Quantitative real-time polymerase chain reaction analysis demonstrated that the expression of PgWRKY1 was relatively higher in roots and lateral roots compared with leaves, stems, and seeds. Importantly, PgWRKY1 expression was significantly induced by salicylic acid, abscisic acid, and NaCl, but downregulated by MeJA treatment. These results suggested that PgWRKY1 might be a multiple stress-inducible gene responding to hormones and salt stresses.

  9. Genome-Wide Identification and Characterization of WRKY Gene Family in Peanut

    PubMed Central

    Song, Hui; Wang, Pengfei; Lin, Jer-Young; Zhao, Chuanzhi; Bi, Yuping; Wang, Xingjun

    2016-01-01

    WRKY, an important transcription factor family, is widely distributed in the plant kingdom. Many reports focused on analysis of phylogenetic relationship and biological function of WRKY protein at the whole genome level in different plant species. However, little is known about WRKY proteins in the genome of Arachis species and their response to salicylic acid (SA) and jasmonic acid (JA) treatment. In this study, we identified 77 and 75 WRKY proteins from the two wild ancestral diploid genomes of cultivated tetraploid peanut, Arachis duranensis and Arachis ipaënsis, using bioinformatics approaches. Most peanut WRKY coding genes were located on A. duranensis chromosome A6 and A. ipaënsis chromosome B3, while the least number of WRKY genes was found in chromosome 9. The WRKY orthologous gene pairs in A. duranensis and A. ipaënsis chromosomes were highly syntenic. Our analysis indicated that segmental duplication events played a major role in AdWRKY and AiWRKY genes, and strong purifying selection was observed in gene duplication pairs. Furthermore, we translate the knowledge gained from the genome-wide analysis result of wild ancestral peanut to cultivated peanut to reveal that gene activities of specific cultivated peanut WRKY gene were changed due to SA and JA treatment. Peanut WRKY7, 8 and 13 genes were down-regulated, whereas WRKY1 and 12 genes were up-regulated with SA and JA treatment. These results could provide valuable information for peanut improvement. PMID:27200012

  10. OsWRKY22, a monocot WRKY gene, plays a role in the resistance response to blast.

    PubMed

    Abbruscato, Pamela; Nepusz, Tamás; Mizzi, Luca; Del Corvo, Marcello; Morandini, Piero; Fumasoni, Irene; Michel, Corinne; Paccanaro, Alberto; Guiderdoni, Emmanuel; Schaffrath, Ulrich; Morel, Jean-Benoît; Piffanelli, Pietro; Faivre-Rampant, Odile

    2012-10-01

    With the aim of identifying novel regulators of host and nonhost resistance to fungi in rice, we carried out a systematic mutant screen of mutagenized lines. Two mutant wrky22 knockout lines revealed clear-cut enhanced susceptibility to both virulent and avirulent Magnaporthe oryzae strains and altered cellular responses to nonhost Magnaporthe grisea and Blumeria graminis fungi. In addition, the analysis of the pathogen responses of 24 overexpressor OsWRKY22 lines revealed enhanced resistance phenotypes on infection with virulent M. oryzae strain, confirming that OsWRKY22 is involved in rice resistance to blast. Bioinformatic analyses determined that the OsWRKY22 gene belongs to a well-defined cluster of monocot-specific WRKYs. The co-regulatory analysis revealed no significant co-regulation of OsWRKY22 with a representative panel of OsWRKYs, supporting its unique role in a series of transcriptional responses. In contrast, inquiring a subset of biotic stress-related Affymetrix data, a large number of resistance and defence-related genes were found to be putatively co-expressed with OsWRKY22. Taken together, all gathered experimental evidence places the monocot-specific OsWRKY22 gene at the convergence point of signal transduction circuits in response to both host and nonhost fungi encountering rice plants.

  11. Genome-wide evolutionary characterization and expression analyses of WRKY family genes in Brachypodium distachyon.

    PubMed

    Wen, Feng; Zhu, Hong; Li, Peng; Jiang, Min; Mao, Wenqing; Ong, Chermaine; Chu, Zhaoqing

    2014-06-01

    Members of plant WRKY gene family are ancient transcription factors that function in plant growth and development and respond to biotic and abiotic stresses. In our present study, we have investigated WRKY family genes in Brachypodium distachyon, a new model plant of family Poaceae. We identified a total of 86 WRKY genes from B. distachyon and explored their chromosomal distribution and evolution, domain alignment, promoter cis-elements, and expression profiles. Combining the analysis of phylogenetic tree of BdWRKY genes and the result of expression profiling, results showed that most of clustered gene pairs had higher similarities in the WRKY domain, suggesting that they might be functionally redundant. Neighbour-joining analysis of 301 WRKY domains from Oryza sativa, Arabidopsis thaliana, and B. distachyon suggested that BdWRKY domains are evolutionarily more closely related to O. sativa WRKY domains than those of A. thaliana. Moreover, tissue-specific expression profile of BdWRKY genes and their responses to phytohormones and several biotic or abiotic stresses were analysed by quantitative real-time PCR. The results showed that the expression of BdWRKY genes was rapidly regulated by stresses and phytohormones, and there was a strong correlation between promoter cis-elements and the phytohormones-induced BdWRKY gene expression.

  12. WRKY domain-encoding genes of a crop legume chickpea (Cicer arietinum): comparative analysis with Medicago truncatula WRKY family and characterization of group-III gene(s)

    PubMed Central

    Kumar, Kamal; Srivastava, Vikas; Purayannur, Savithri; Kaladhar, V. Chandra; Cheruvu, Purnima Jaiswal; Verma, Praveen Kumar

    2016-01-01

    The WRKY genes have been identified as important transcriptional modulators predominantly during the environmental stresses, but they also play critical role at various stages of plant life cycle. We report the identification of WRKY domain (WD)-encoding genes from galegoid clade legumes chickpea (Cicer arietinum L.) and barrel medic (Medicago truncatula). In total, 78 and 98 WD-encoding genes were found in chickpea and barrel medic, respectively. Comparative analysis suggests the presence of both conserved and unique WRKYs, and expansion of WRKY family in M. truncatula primarily by tandem duplication. Exclusively found in galegoid legumes, CaWRKY16 and its orthologues encode for a novel protein having a transmembrane and partial Exo70 domains flanking a group-III WD. Genomic region of galegoids, having CaWRKY16, is more dynamic when compared with millettioids. In onion cells, fused CaWRKY16-EYFP showed punctate fluorescent signals in cytoplasm. The chickpea WRKY group-III genes were further characterized for their transcript level modulation during pathogenic stress and treatments of abscisic acid, jasmonic acid, and salicylic acid (SA) by real-time PCR. Differential regulation of genes was observed during Ascochyta rabiei infection and SA treatment. Characterization of A. rabiei and SA inducible gene CaWRKY50 showed that it localizes to plant nucleus, binds to W-box, and have a C-terminal transactivation domain. Overexpression of CaWRKY50 in tobacco plants resulted in early flowering and senescence. The in-depth comparative account presented here for two legume WRKY genes will be of great utility in hastening functional characterization of crop legume WRKYs and will also help in characterization of Exo70Js. PMID:27060167

  13. WRKY domain-encoding genes of a crop legume chickpea (Cicer arietinum): comparative analysis with Medicago truncatula WRKY family and characterization of group-III gene(s).

    PubMed

    Kumar, Kamal; Srivastava, Vikas; Purayannur, Savithri; Kaladhar, V Chandra; Cheruvu, Purnima Jaiswal; Verma, Praveen Kumar

    2016-06-01

    The WRKY genes have been identified as important transcriptional modulators predominantly during the environmental stresses, but they also play critical role at various stages of plant life cycle. We report the identification of WRKY domain (WD)-encoding genes from galegoid clade legumes chickpea (Cicer arietinum L.) and barrel medic (Medicago truncatula). In total, 78 and 98 WD-encoding genes were found in chickpea and barrel medic, respectively. Comparative analysis suggests the presence of both conserved and unique WRKYs, and expansion of WRKY family in M. truncatula primarily by tandem duplication. Exclusively found in galegoid legumes, CaWRKY16 and its orthologues encode for a novel protein having a transmembrane and partial Exo70 domains flanking a group-III WD. Genomic region of galegoids, having CaWRKY16, is more dynamic when compared with millettioids. In onion cells, fused CaWRKY16-EYFP showed punctate fluorescent signals in cytoplasm. The chickpea WRKY group-III genes were further characterized for their transcript level modulation during pathogenic stress and treatments of abscisic acid, jasmonic acid, and salicylic acid (SA) by real-time PCR. Differential regulation of genes was observed during Ascochyta rabiei infection and SA treatment. Characterization of A. rabiei and SA inducible gene CaWRKY50 showed that it localizes to plant nucleus, binds to W-box, and have a C-terminal transactivation domain. Overexpression of CaWRKY50 in tobacco plants resulted in early flowering and senescence. The in-depth comparative account presented here for two legume WRKY genes will be of great utility in hastening functional characterization of crop legume WRKYs and will also help in characterization of Exo70Js.

  14. [Genome-wide identification and expression analysis of the WRKY gene family in peach].

    PubMed

    Yanbing, Gu; Zhirui, Ji; Fumei, Chi; Zhuang, Qiao; Chengnan, Xu; Junxiang, Zhang; Zongshan, Zhou; Qinglong, Dong

    2016-03-01

    The WRKY transcription factors are one of the largest families of transcriptional regulators and play diverse regulatory roles in biotic and abiotic stresses, plant growth and development processes. In this study, the WRKY DNA-binding domain (Pfam Database number: PF03106) downloaded from Pfam protein families database was exploited to identify WRKY genes from the peach (Prunus persica 'Lovell') genome using HMMER 3.0. The obtained amino acid sequences were analyzed with DNAMAN 5.0, WebLogo 3, MEGA 5.1, MapInspect and MEME bioinformatics softwares. Totally 61 peach WRKY genes were found in the peach genome. Our phylogenetic analysis revealed that peach WRKY genes were classified into three Groups: Ⅰ, Ⅱ and Ⅲ. The WRKY N-terminal and C-terminal domains of Group Ⅰ (group I-N and group I-C) were monophyletic. The Group Ⅱ was sub-divided into five distinct clades (groupⅡ-a, Ⅱ-b, Ⅱ-c, Ⅱ-d and Ⅱ-e). Our domain analysis indicated that the WRKY regions contained a highly conserved heptapeptide stretch WRKYGQK at its N-terminus followed by a zinc-finger motif. The chromosome mapping analysis showed that peach WRKY genes were distributed with different densities over 8 chromosomes. The intron-exon structure analysis revealed that structures of the WRKY gene were highly conserved in the peach. The conserved motif analysis showed that the conserved motifs 1, 2 and 3, which specify the WRKY domain, were observed in all peach WRKY proteins, motif 5 as the unknown domain was observed in group Ⅱ-d, two WRKY domains were assigned to GroupⅠ. SqRT-PCR and qRT-PCR results indicated that 16 PpWRKY genes were expressed in roots, stems, leaves, flowers and fruits at various expression levels. Our analysis thus identified the PpWRKY gene families, and future functional studies are needed to reveal its specific roles.

  15. A WRKY gene from Tamarix hispida, ThWRKY4, mediates abiotic stress responses by modulating reactive oxygen species and expression of stress-responsive genes.

    PubMed

    Zheng, Lei; Liu, Guifeng; Meng, Xiangnan; Liu, Yujia; Ji, Xiaoyu; Li, Yanbang; Nie, Xianguang; Wang, Yucheng

    2013-07-01

    WRKY transcription factors are involved in various biological processes, such as development, metabolism and responses to stress. However, their exact roles in abiotic stress tolerance are largely unknown. Here, we demonstrated a working model for the function of a WRKY gene (ThWRKY4) from Tamarix hispida in the stress response. ThWRKY4 is highly induced by abscisic acid (ABA), salt and drought in the early period of stress (stress for 3, 6, or 9 h), which can be regulated by ABF (ABRE binding factors) and Dof (DNA binding with one finger), and also can be crossregulated by other WRKYs and autoregulated as well. Overexpression of ThWRKY4 conferred tolerance to salt, oxidative and ABA treatment in transgenic plants. ThWRKY4 can improve the tolerance to salt and ABA treatment by improving activities of superoxide dismutase and peroxidase, decreasing levels of O2 (-) and H2O2, reducing electrolyte leakage, keeping the loss of chlorophyll, and protecting cells from death. Microarray analyses showed that overexpression of ThWRKY4 in Arabidopsis leads to 165 and 100 genes significantly up- and downregulated, respectively. Promoter scanning analysis revealed that ThWRKY4 regulates the gene expression via binding to W-box motifs present in their promoter regions. This study shows that ThWRKY4 functions as a transcription factor to positively modulate abiotic stress tolerances, and is involved in modulating reactive oxygen species.

  16. Genome-Wide Identification and Expression Analysis of the WRKY Gene Family in Cassava

    PubMed Central

    Wei, Yunxie; Shi, Haitao; Xia, Zhiqiang; Tie, Weiwei; Ding, Zehong; Yan, Yan; Wang, Wenquan; Hu, Wei; Li, Kaimian

    2016-01-01

    The WRKY family, a large family of transcription factors (TFs) found in higher plants, plays central roles in many aspects of physiological processes and adaption to environment. However, little information is available regarding the WRKY family in cassava (Manihot esculenta). In the present study, 85 WRKY genes were identified from the cassava genome and classified into three groups according to conserved WRKY domains and zinc-finger structure. Conserved motif analysis showed that all of the identified MeWRKYs had the conserved WRKY domain. Gene structure analysis suggested that the number of introns in MeWRKY genes varied from 1 to 5, with the majority of MeWRKY genes containing three exons. Expression profiles of MeWRKY genes in different tissues and in response to drought stress were analyzed using the RNA-seq technique. The results showed that 72 MeWRKY genes had differential expression in their transcript abundance and 78 MeWRKY genes were differentially expressed in response to drought stresses in different accessions, indicating their contribution to plant developmental processes and drought stress resistance in cassava. Finally, the expression of 9 WRKY genes was analyzed by qRT-PCR under osmotic, salt, ABA, H2O2, and cold treatments, indicating that MeWRKYs may be involved in different signaling pathways. Taken together, this systematic analysis identifies some tissue-specific and abiotic stress-responsive candidate MeWRKY genes for further functional assays in planta, and provides a solid foundation for understanding of abiotic stress responses and signal transduction mediated by WRKYs in cassava. PMID:26904033

  17. Evolution and expression analysis of the grape (Vitis vinifera L.) WRKY gene family.

    PubMed

    Guo, Chunlei; Guo, Rongrong; Xu, Xiaozhao; Gao, Min; Li, Xiaoqin; Song, Junyang; Zheng, Yi; Wang, Xiping

    2014-04-01

    WRKY proteins comprise a large family of transcription factors that play important roles in plant defence regulatory networks, including responses to various biotic and abiotic stresses. To date, no large-scale study of WRKY genes has been undertaken in grape (Vitis vinifera L.). In this study, a total of 59 putative grape WRKY genes (VvWRKY) were identified and renamed on the basis of their respective chromosome distribution. A multiple sequence alignment analysis using all predicted grape WRKY genes coding sequences, together with those from Arabidopsis thaliana and tomato (Solanum lycopersicum), indicated that the 59 VvWRKY genes can be classified into three main groups (I-III). An evaluation of the duplication events suggested that several WRKY genes arose before the divergence of the grape and Arabidopsis lineages. Moreover, expression profiles derived from semiquantitative PCR and real-time quantitative PCR analyses showed distinct expression patterns in various tissues and in response to different treatments. Four VvWRKY genes showed a significantly higher expression in roots or leaves, 55 responded to varying degrees to at least one abiotic stress treatment, and the expression of 38 were altered following powdery mildew (Erysiphe necator) infection. Most VvWRKY genes were downregulated in response to abscisic acid or salicylic acid treatments, while the expression of a subset was upregulated by methyl jasmonate or ethylene treatments.

  18. The WRKY Transcription Factor Family in Citrus: Valuable and Useful Candidate Genes for Citrus Breeding.

    PubMed

    Ayadi, M; Hanana, M; Kharrat, N; Merchaoui, H; Marzoug, R Ben; Lauvergeat, V; Rebaï, A; Mzid, R

    2016-10-01

    WRKY transcription factors belong to a large family of plant transcriptional regulators whose members have been reported to be involved in a wide range of biological roles including plant development, adaptation to environmental constraints and response to several diseases. However, little or poor information is available about WRKY's in Citrus. The recent release of completely assembled genomes sequences of Citrus sinensis and Citrus clementina and the availability of ESTs sequences from other citrus species allowed us to perform a genome survey for Citrus WRKY proteins. In the present study, we identified 100 WRKY members from C. sinensis (51), C. clementina (48) and Citrus unshiu (1), and analyzed their chromosomal distribution, gene structure, gene duplication, syntenic relation and phylogenetic analysis. A phylogenetic tree of 100 Citrus WRKY sequences with their orthologs from Arabidopsis has distinguished seven groups. The CsWRKY genes were distributed across all ten sweet orange chromosomes. A comprehensive approach and an integrative analysis of Citrus WRKY gene expression revealed variable profiles of expression within tissues and stress conditions indicating functional diversification. Thus, candidate Citrus WRKY genes have been proposed as potentially involved in fruit acidification, essential oil biosynthesis and abiotic/biotic stress tolerance. Our results provided essential prerequisites for further WRKY genes cloning and functional analysis with an aim of citrus crop improvement.

  19. Genome-Wide Identification and Expression Analysis of WRKY Gene Family in Capsicum annuum L.

    PubMed Central

    Diao, Wei-Ping; Snyder, John C.; Wang, Shu-Bin; Liu, Jin-Bing; Pan, Bao-Gui; Guo, Guang-Jun; Wei, Ge

    2016-01-01

    The WRKY family of transcription factors is one of the most important families of plant transcriptional regulators with members regulating multiple biological processes, especially in regulating defense against biotic and abiotic stresses. However, little information is available about WRKYs in pepper (Capsicum annuum L.). The recent release of completely assembled genome sequences of pepper allowed us to perform a genome-wide investigation for pepper WRKY proteins. In the present study, a total of 71 WRKY genes were identified in the pepper genome. According to structural features of their encoded proteins, the pepper WRKY genes (CaWRKY) were classified into three main groups, with the second group further divided into five subgroups. Genome mapping analysis revealed that CaWRKY were enriched on four chromosomes, especially on chromosome 1, and 15.5% of the family members were tandemly duplicated genes. A phylogenetic tree was constructed depending on WRKY domain' sequences derived from pepper and Arabidopsis. The expression of 21 selected CaWRKY genes in response to seven different biotic and abiotic stresses (salt, heat shock, drought, Phytophtora capsici, SA, MeJA, and ABA) was evaluated by quantitative RT-PCR; Some CaWRKYs were highly expressed and up-regulated by stress treatment. Our results will provide a platform for functional identification and molecular breeding studies of WRKY genes in pepper. PMID:26941768

  20. Genome-wide identification and characterization of WRKY gene family in Salix suchowensis

    PubMed Central

    Ye, Qiaolin; Yin, Tongming

    2016-01-01

    WRKY proteins are the zinc finger transcription factors that were first identified in plants. They can specifically interact with the W-box, which can be found in the promoter region of a large number of plant target genes, to regulate the expressions of downstream target genes. They also participate in diverse physiological and growing processes in plants. Prior to this study, a plenty of WRKY genes have been identified and characterized in herbaceous species, but there is no large-scale study of WRKY genes in willow. With the whole genome sequencing of Salix suchowensis, we have the opportunity to conduct the genome-wide research for willow WRKY gene family. In this study, we identified 85 WRKY genes in the willow genome and renamed them from SsWRKY1 to SsWRKY85 on the basis of their specific distributions on chromosomes. Due to their diverse structural features, the 85 willow WRKY genes could be further classified into three main groups (group I–III), with five subgroups (IIa–IIe) in group II. With the multiple sequence alignment and the manual search, we found three variations of the WRKYGQK heptapeptide: WRKYGRK, WKKYGQK and WRKYGKK, and four variations of the normal zinc finger motif, which might execute some new biological functions. In addition, the SsWRKY genes from the same subgroup share the similar exon–intron structures and conserved motif domains. Further studies of SsWRKY genes revealed that segmental duplication events (SDs) played a more prominent role in the expansion of SsWRKY genes. Distinct expression profiles of SsWRKY genes with RNA sequencing data revealed that diverse expression patterns among five tissues, including tender roots, young leaves, vegetative buds, non-lignified stems and barks. With the analyses of WRKY gene family in willow, it is not only beneficial to complete the functional and annotation information of WRKY genes family in woody plants, but also provide important references to investigate the expansion and evolution

  1. Molecular Cloning and Expression Analysis of Eight PgWRKY Genes in Panax ginseng Responsive to Salt and Hormones.

    PubMed

    Xiu, Hao; Nuruzzaman, Mohammed; Guo, Xiangqian; Cao, Hongzhe; Huang, Jingjia; Chen, Xianghui; Wu, Kunlu; Zhang, Ru; Huang, Yuzhao; Luo, Junli; Luo, Zhiyong

    2016-03-04

    Despite the importance of WRKY genes in plant physiological processes, little is known about their roles in Panax ginseng C.A. Meyer. Forty-eight unigenes on this species were previously reported as WRKY transcripts using the next-generation sequencing (NGS) technology. Subsequently, one gene that encodes PgWRKY1 protein belonging to subgroup II-d was cloned and functionally characterized. In this study, eight WRKY genes from the NGS-based transcriptome sequencing dataset designated as PgWRKY2-9 have been cloned and characterized. The genes encoding WRKY proteins were assigned to WRKY Group II (one subgroup II-c, four subgroup II-d, and three subgroup II-e) based on phylogenetic analysis. The cDNAs of the cloned PgWRKYs encode putative proteins ranging from 194 to 358 amino acid residues, each of which includes one WRKYGQK sequence motif and one C₂H₂-type zinc-finger motif. Quantitative real-time PCR (qRT-PCR) analysis demonstrated that the eight analyzed PgWRKY genes were expressed at different levels in various organs including leaves, roots, adventitious roots, stems, and seeds. Importantly, the transcription responses of these PgWRKYs to methyl jasmonate (MeJA) showed that PgWRKY2, PgWRKY3, PgWRKY4, PgWRKY5, PgWRKY6, and PgWRKY7 were downregulated by MeJA treatment, while PgWRKY8 and PgWRKY9 were upregulated to varying degrees. Moreover, the PgWRKY genes increased or decreased by salicylic acid (SA), abscisic acid (ABA), and NaCl treatments. The results suggest that the PgWRKYs may be multiple stress-inducible genes responding to both salt and hormones.

  2. Overexpression of a cotton (Gossypium hirsutum) WRKY gene, GhWRKY34, in Arabidopsis enhances salt-tolerance of the transgenic plants.

    PubMed

    Zhou, Li; Wang, Na-Na; Gong, Si-Ying; Lu, Rui; Li, Yang; Li, Xue-Bao

    2015-11-01

    Soil salinity is one of the most serious threats in world agriculture, and often influences cotton growth and development, resulting in a significant loss in cotton crop yield. WRKY transcription factors are involved in plant response to high salinity stress, but little is known about the role of WRKY transcription factors in cotton so far. In this study, a member (GhWRKY34) of cotton WRKY family was functionally characterized. This protein containing a WRKY domain and a zinc-finger motif belongs to group III of cotton WRKY family. Subcellular localization assay indicated that GhWRKY34 is localized to the cell nucleus. Overexpression of GhWRKY34 in Arabidopsis enhanced the transgenic plant tolerance to salt stress. Several parameters (such as seed germination, green cotyledons, root length and chlorophyll content) in the GhWRKY34 transgenic lines were significantly higher than those in wild type under NaCl treatment. On the contrary, the GhWRKY34 transgenic plants exhibited a substantially lower ratio of Na(+)/K(+) in leaves and roots dealing with salt stress, compared with wild type. Growth status of the GhWRKY34 transgenic plants was much better than that of wild type under salt stress. Expressions of the stress-related genes were remarkably up-regulated in the transgenic plants under salt stress, compared with those in wild type. Based on the data presented in this study, we hypothesize that GhWRKY34 as a positive transcription regulator may function in plant response to high salinity stress through maintaining the Na(+)/K(+) homeostasis as well as activating the salt stress-related genes in cells.

  3. FcWRKY70, a WRKY protein of Fortunella crassifolia, functions in drought tolerance and modulates putrescine synthesis by regulating arginine decarboxylase gene.

    PubMed

    Gong, Xiaoqing; Zhang, Jingyan; Hu, Jianbing; Wang, Wei; Wu, Hao; Zhang, Qinghua; Liu, Ji-Hong

    2015-11-01

    WRKY comprises a large family of transcription factors in plants, but most WRKY members are still poorly understood. In this study, we report functional characterization of a Group III WRKY gene (FcWRKY70) from Fortunella crassifolia. FcWRKY70 was greatly induced by drought and abscisic acid, but slightly or negligibly by salt and cold. Overexpression of FcWRKY70 in tobacco (Nicotiana nudicaulis) and lemon (Citrus lemon) conferred enhanced tolerance to dehydration and drought stresses. Transgenic tobacco and lemon exhibited higher expression levels of ADC (arginine decarboxylase), and accumulated larger amount of putrescine in comparison with wild type (WT). Treatment with D-arginine, an inhibitor of ADC, caused transgenic tobacco plants more sensitive to dehydration. Knock-down of FcWRKY70 in kumquat down-regulated ADC abundance and decreased putrescine level, accompanied by compromised dehydration tolerance. The promoter region of FcADC contained two W-box elements, which were shown to be interacted with FcWRKY70. Taken together, our data demonstrated that FcWRKY70 functions in drought tolerance by, at least partly, promoting production of putrescine via regulating ADC expression.

  4. Gene Structures, Evolution and Transcriptional Profiling of the WRKY Gene Family in Castor Bean (Ricinus communis L.).

    PubMed

    Zou, Zhi; Yang, Lifu; Wang, Danhua; Huang, Qixing; Mo, Yeyong; Xie, Guishui

    2016-01-01

    WRKY proteins comprise one of the largest transcription factor families in plants and form key regulators of many plant processes. This study presents the characterization of 58 WRKY genes from the castor bean (Ricinus communis L., Euphorbiaceae) genome. Compared with the automatic genome annotation, one more WRKY-encoding locus was identified and 20 out of the 57 predicted gene models were manually corrected. All RcWRKY genes were shown to contain at least one intron in their coding sequences. According to the structural features of the present WRKY domains, the identified RcWRKY genes were assigned to three previously defined groups (I-III). Although castor bean underwent no recent whole-genome duplication event like physic nut (Jatropha curcas L., Euphorbiaceae), comparative genomics analysis indicated that one gene loss, one intron loss and one recent proximal duplication occurred in the RcWRKY gene family. The expression of all 58 RcWRKY genes was supported by ESTs and/or RNA sequencing reads derived from roots, leaves, flowers, seeds and endosperms. Further global expression profiles with RNA sequencing data revealed diverse expression patterns among various tissues. Results obtained from this study not only provide valuable information for future functional analysis and utilization of the castor bean WRKY genes, but also provide a useful reference to investigate the gene family expansion and evolution in Euphorbiaceus plants.

  5. Isolation, Expression, and Promoter Analysis of GbWRKY2: A Novel Transcription Factor Gene from Ginkgo biloba

    PubMed Central

    Liao, Yong-Ling; Shen, Yong-Bao; Chang, Jie; Zhang, Wei-Wei; Cheng, Shui-Yuan; Xu, Feng

    2015-01-01

    WRKY transcription factor is involved in multiple life activities including plant growth and development as well as biotic and abiotic responses. We identified 28 WRKY genes from transcriptome data of Ginkgo biloba according to conserved WRKY domains and zinc finger structure and selected three WRKY genes, which are GbWRKY2, GbWRKY16, and GbWRKY21, for expression pattern analysis. GbWRKY2 was preferentially expressed in flowers and strongly induced by methyl jasmonate. Here, we cloned the full-length cDNA and genomic DNA of GbWRKY2. The full-length cDNA of GbWRKY2 was 1,713 bp containing a 1,014 bp open reading frame encoding a polypeptide of 337 amino acids. The GbWRKY2 genomic DNA had one intron and two exons. The deduced GbWRKY2 contained one WRKY domain and one zinc finger motif. GbWRKY2 was classified into Group II WRKYs. Southern blot analysis revealed that GbWRKY2 was a single copy gene in G. biloba. Many cis-acting elements related to hormone and stress responses were identified in the 1,363 bp-length 5′-flanking sequence of GbWRKY2, including W-box, ABRE-motif, MYBCOREs, and PYRIMIDINE-boxes, revealing the molecular mechanism of upregulated expression of GbWRKY2 by hormone and stress treatments. Further functional characterizations in transiently transformed tobacco leaves allowed us to identify the region that can be considered as the minimal promoter. PMID:26351628

  6. Identification and expression analysis of WRKY family genes under biotic and abiotic stresses in Brassica rapa.

    PubMed

    Kayum, Md Abdul; Jung, Hee-Jeong; Park, Jong-In; Ahmed, Nasar Uddin; Saha, Gopal; Yang, Tae-Jin; Nou, Ill-Sup

    2015-02-01

    WRKY proteins constitute one of the largest transcription factor families in higher plants, and they are involved in multiple biological processes such as plant development, metabolism, and responses to biotic and abiotic stresses. Genes of this family have been well documented in response to many abiotic and biotic stresses in many plant species, but not yet against Pectobacterium carotovorum subsp. carotovorum and Fusarium oxysporum f.sp. conglutinans in any of the plants. Moreover, potentiality of a specific gene may vary depending on stress conditions and genotypes. To identify stress resistance-related potential WRKY genes of Brassica rapa, we analyzed their expressions against above-mentioned pathogens and cold, salt, and drought stresses in B. rapa. Stress resistance-related functions of all Brassica rapa WRKY (BrWRKY) genes were firstly analyzed through homology study with existing biotic and abiotic stress resistance-related WRKY genes of other plant species and found a high degree of homology. We then identified all BrWRKY genes in a Br135K microarray dataset, which was created by applying low-temperature stresses to two contrasting Chinese cabbage doubled haploid (DH) lines, Chiifu and Kenshin, and selected 41 BrWRKY genes with high and differential transcript abundance levels. These selected genes were further investigated under cold, salt, and drought stresses as well as after infection with P. carotovorum subsp. carotovorum and F. oxysporum f.sp. conglutinans in B. rapa. The selected genes showed an organ-specific expression, and 22 BrWRKY genes were differentially expressed in Chiifu compared to Kenshin under cold and drought stresses. Six BrWRKY genes were more responsive in Kenshin compared to Chiffu under salt stress. In addition, eight BrWRKY genes showed differential expression after P. carotovorum subsp. carotovorum infection and five genes after F. oxysporum f.sp. conglutinans infection in B. rapa. Thus, the differentially expressed BrWRKY

  7. The WRKY Transcription Factor WRKY71/EXB1 Controls Shoot Branching by Transcriptionally Regulating RAX Genes in Arabidopsis

    PubMed Central

    Guo, Dongshu; Zhang, Jinzhe; Wang, Xinlei; Han, Xiang; Wei, Baoye; Yu, Hao; Huang, Qingpei

    2015-01-01

    Plant shoot branching is pivotal for developmental plasticity and crop yield. The formation of branch meristems is regulated by several key transcription factors including REGULATOR OF AXILLARY MERISTEMS1 (RAX1), RAX2, and RAX3. However, the regulatory network of shoot branching is still largely unknown. Here, we report the identification of EXCESSIVE BRANCHES1 (EXB1), which affects axillary meristem (AM) initiation and bud activity. Overexpression of EXB1 in the gain-of-function mutant exb1-D leads to severe bushy and dwarf phenotypes, which result from excessive AM initiation and elevated bud activities. EXB1 encodes the WRKY transcription factor WRKY71, which has demonstrated transactivation activities. Disruption of WRKY71/EXB1 by chimeric repressor silencing technology leads to fewer branches, indicating that EXB1 plays important roles in the control of shoot branching. We demonstrate that EXB1 controls AM initiation by positively regulating the transcription of RAX1, RAX2, and RAX3. Disruption of the RAX genes partially rescues the branching phenotype caused by EXB1 overexpression. We further show that EXB1 also regulates auxin homeostasis in control of shoot branching. Our data demonstrate that EXB1 plays pivotal roles in shoot branching by regulating both transcription of RAX genes and auxin pathways. PMID:26578700

  8. The WRKY transcription factor genes in eggplant (Solanum melongena L.) and Turkey Berry (Solanum torvum Sw.).

    PubMed

    Yang, Xu; Deng, Cao; Zhang, Yu; Cheng, Yufu; Huo, Qiuyue; Xue, Linbao

    2015-04-07

    WRKY transcription factors, which play critical roles in stress responses, have not been characterized in eggplant or its wild relative, turkey berry. The recent availability of RNA-sequencing data provides the opportunity to examine WRKY genes from a global perspective. We identified 50 and 62 WRKY genes in eggplant (SmelWRKYs) and turkey berry (StorWRKYs), respectively, all of which could be classified into three groups (I-III) based on the WRKY protein structure. The SmelWRKYs and StorWRKYs contain ~76% and ~95% of the number of WRKYs found in other sequenced asterid species, respectively. Positive selection analysis revealed that different selection constraints could have affected the evolution of these groups. Positively-selected sites were found in Groups IIc and III. Branch-specific selection pressure analysis indicated that most WRKY domains from SmelWRKYs and StorWRKYs are conserved and have evolved at low rates since their divergence. Comparison to homologous WRKY genes in Arabidopsis revealed several potential pathogen resistance-related SmelWRKYs and StorWRKYs, providing possible candidate genetic resources for improving stress tolerance in eggplant and probably other Solanaceae plants. To our knowledge, this is the first report of a genome-wide analyses of the SmelWRKYs and StorWRKYs.

  9. The WRKY Transcription Factor Genes in Eggplant (Solanum melongena L.) and Turkey Berry (Solanum torvum Sw.)

    PubMed Central

    Yang, Xu; Deng, Cao; Zhang, Yu; Cheng, Yufu; Huo, Qiuyue; Xue, Linbao

    2015-01-01

    WRKY transcription factors, which play critical roles in stress responses, have not been characterized in eggplant or its wild relative, turkey berry. The recent availability of RNA-sequencing data provides the opportunity to examine WRKY genes from a global perspective. We identified 50 and 62 WRKY genes in eggplant (SmelWRKYs) and turkey berry (StorWRKYs), respectively, all of which could be classified into three groups (I–III) based on the WRKY protein structure. The SmelWRKYs and StorWRKYs contain ~76% and ~95% of the number of WRKYs found in other sequenced asterid species, respectively. Positive selection analysis revealed that different selection constraints could have affected the evolution of these groups. Positively-selected sites were found in Groups IIc and III. Branch-specific selection pressure analysis indicated that most WRKY domains from SmelWRKYs and StorWRKYs are conserved and have evolved at low rates since their divergence. Comparison to homologous WRKY genes in Arabidopsis revealed several potential pathogen resistance-related SmelWRKYs and StorWRKYs, providing possible candidate genetic resources for improving stress tolerance in eggplant and probably other Solanaceae plants. To our knowledge, this is the first report of a genome-wide analyses of the SmelWRKYs and StorWRKYs. PMID:25853261

  10. In silico identification and characterization of the WRKY gene superfamily in pepper (Capsicum annuum L.).

    PubMed

    Cheng, Y; Yao, Z P; Ruan, M Y; Ye, Q J; Wang, R Q; Zhou, G Z; Luo, J

    2016-09-23

    The WRKY family is one of the most important transcription factor families in plants, involved in the regulation of a broad range of biological roles. The recent releases of whole-genome sequences of pepper (Capsicum annuum L.) allow us to perform a genome-wide identification and characterization of the WRKY family. In this study, 61 CaWRKY proteins were identified in the pepper genome. Based on protein structural and phylogenetic analyses, these proteins were classified into four main groups (I, II, III, and NG), and Group II was further divided into five subgroups (IIa to IIe). Chromosome mapping analysis indicated that CaWRKY genes are distributed across all 12 chromosomes, although the location of four CaWRKYs (CaWRKY58-CaWRKY61) could not be identified. Two pairs of CaWRKYs located on chromosome 01 appear to be tandem duplications. Furthermore, the phylogenetic tree showed a close evolutionary relationship of WRKYs in three species from Solanaceae. In conclusion, this comprehensive analysis of CaWRKYs will provide rich resources for further functional studies in pepper.

  11. Genome-wide identification of WRKY family genes and their response to cold stress in Vitis vinifera

    Technology Transfer Automated Retrieval System (TEKTRAN)

    WRKY transcription factors are one of the largest families of transcriptional regulators in plants. WRKY genes are not only found to play significant roles in biotic and abiotic stress response, but also regulate growth and development. Grapevine (Vitis vinifera) production is largely limited by str...

  12. Transcriptome-wide identification of salt-responsive members of the WRKY gene family in Gossypium aridum.

    PubMed

    Fan, Xinqi; Guo, Qi; Xu, Peng; Gong, YuanYong; Shu, Hongmei; Yang, Yang; Ni, Wanchao; Zhang, Xianggui; Shen, Xinlian

    2015-01-01

    WRKY transcription factors are plant-specific, zinc finger-type transcription factors. The WRKY superfamily is involved in abiotic stress responses in many crops including cotton, a major fiber crop that is widely cultivated and consumed throughout the world. Salinity is an important abiotic stress that results in considerable yield losses. In this study, we identified 109 WRKY genes (GarWRKYs) in a salt-tolerant wild cotton species Gossypium aridum from transcriptome sequencing data to elucidate the roles of these factors in cotton salt tolerance. According to their structural features, the predicted members were divided into three groups (Groups I-III), as previously described for Arabidopsis. Furthermore, 28 salt-responsive GarWRKY genes were identified from digital gene expression data and subjected to real-time quantitative RT-PCR analysis. The expression patterns of most GarWRKY genes revealed by this analysis are in good agreement with those revealed by RNA-Seq analysis. RT-PCR analysis revealed that 27 GarWRKY genes were expressed in roots and one was exclusively expressed in roots. Analysis of gene orthology and motif compositions indicated that WRKY members from Arabidopsis, rice and soybean generally shared the similar motifs within the same subgroup, suggesting they have the similar function. Overexpression-GarWRKY17 and -GarWRKY104 in Arabidopsis revealed that they could positively regulate salt tolerance of transgenic Arabidopsis during different development stages. The comprehensive data generated in this study provide a platform for elucidating the functions of WRKY transcription factors in salt tolerance of G. aridum. In addition, GarWRKYs related to salt tolerance identified in this study will be potential candidates for genetic improvement of cultivated cotton salt stress tolerance.

  13. Comparative Analysis of WRKY Genes Potentially Involved in Salt Stress Responses in Triticum turgidum L. ssp. durum.

    PubMed

    Yousfi, Fatma-Ezzahra; Makhloufi, Emna; Marande, William; Ghorbel, Abdel W; Bouzayen, Mondher; Bergès, Hélène

    2016-01-01

    WRKY transcription factors are involved in multiple aspects of plant growth, development and responses to biotic stresses. Although they have been found to play roles in regulating plant responses to environmental stresses, these roles still need to be explored, especially those pertaining to crops. Durum wheat is the second most widely produced cereal in the world. Complex, large and unsequenced genomes, in addition to a lack of genomic resources, hinder the molecular characterization of tolerance mechanisms. This paper describes the isolation and characterization of five TdWRKY genes from durum wheat (Triticum turgidum L. ssp. durum). A PCR-based screening of a T. turgidum BAC genomic library using primers within the conserved region of WRKY genes resulted in the isolation of five BAC clones. Following sequencing fully the five BACs, fine annotation through Triannot pipeline revealed 74.6% of the entire sequences as transposable elements and a 3.2% gene content with genes organized as islands within oceans of TEs. Each BAC clone harbored a TdWRKY gene. The study showed a very extensive conservation of genomic structure between TdWRKYs and their orthologs from Brachypodium, barley, and T. aestivum. The structural features of TdWRKY proteins suggested that they are novel members of the WRKY family in durum wheat. TdWRKY1/2/4, TdWRKY3, and TdWRKY5 belong to the group Ia, IIa, and IIc, respectively. Enrichment of cis-regulatory elements related to stress responses in the promoters of some TdWRKY genes indicated their potential roles in mediating plant responses to a wide variety of environmental stresses. TdWRKY genes displayed different expression patterns in response to salt stress that distinguishes two durum wheat genotypes with contrasting salt stress tolerance phenotypes. TdWRKY genes tended to react earlier with a down-regulation in sensitive genotype leaves and with an up-regulation in tolerant genotype leaves. The TdWRKY transcripts levels in roots increased

  14. Comparative Analysis of WRKY Genes Potentially Involved in Salt Stress Responses in Triticum turgidum L. ssp. durum

    PubMed Central

    Yousfi, Fatma-Ezzahra; Makhloufi, Emna; Marande, William; Ghorbel, Abdel W.; Bouzayen, Mondher; Bergès, Hélène

    2017-01-01

    WRKY transcription factors are involved in multiple aspects of plant growth, development and responses to biotic stresses. Although they have been found to play roles in regulating plant responses to environmental stresses, these roles still need to be explored, especially those pertaining to crops. Durum wheat is the second most widely produced cereal in the world. Complex, large and unsequenced genomes, in addition to a lack of genomic resources, hinder the molecular characterization of tolerance mechanisms. This paper describes the isolation and characterization of five TdWRKY genes from durum wheat (Triticum turgidum L. ssp. durum). A PCR-based screening of a T. turgidum BAC genomic library using primers within the conserved region of WRKY genes resulted in the isolation of five BAC clones. Following sequencing fully the five BACs, fine annotation through Triannot pipeline revealed 74.6% of the entire sequences as transposable elements and a 3.2% gene content with genes organized as islands within oceans of TEs. Each BAC clone harbored a TdWRKY gene. The study showed a very extensive conservation of genomic structure between TdWRKYs and their orthologs from Brachypodium, barley, and T. aestivum. The structural features of TdWRKY proteins suggested that they are novel members of the WRKY family in durum wheat. TdWRKY1/2/4, TdWRKY3, and TdWRKY5 belong to the group Ia, IIa, and IIc, respectively. Enrichment of cis-regulatory elements related to stress responses in the promoters of some TdWRKY genes indicated their potential roles in mediating plant responses to a wide variety of environmental stresses. TdWRKY genes displayed different expression patterns in response to salt stress that distinguishes two durum wheat genotypes with contrasting salt stress tolerance phenotypes. TdWRKY genes tended to react earlier with a down-regulation in sensitive genotype leaves and with an up-regulation in tolerant genotype leaves. The TdWRKY transcripts levels in roots increased

  15. Cloning and characterization of a novel stress-responsive WRKY transcription factor gene (MusaWRKY71) from Musa spp. cv. Karibale Monthan (ABB group) using transformed banana cells.

    PubMed

    Shekhawat, Upendra K Singh; Ganapathi, Thumballi R; Srinivas, Lingam

    2011-08-01

    WRKY transcription factor proteins play significant roles in plant stress responses. Here, we report the cloning and characterization of a novel WRKY gene, MusaWRKY71 isolated from an edible banana cultivar Musa spp. Karibale Monthan (ABB group). MusaWRKY71, initially identified using in silico approaches from an abiotic stress-related EST library, was later extended towards the 3' end using rapid amplification of cDNA ends technique. The 1299-bp long cDNA of MusaWRKY71 encodes a protein with 280 amino acids and contains a characteristic WRKY domain in the C-terminal half. Although MusaWRKY71 shares good similarity with other monocot WRKY proteins the substantial size difference makes it a unique member of the WRKY family in higher plants. The 918-bp long 5' proximal region determined using thermal asymmetric interlaced-polymerase chain reaction has many putative cis-acting elements and transcription factor binding motifs. Subcellular localization assay of MusaWRKY71 performed using a GFP-fusion platform confirmed its nuclear targeting in transformed banana suspension cells. Importantly, MusaWRKY71 expression in banana plantlets was up-regulated manifold by cold, dehydration, salt, ABA, H2O2, ethylene, salicylic acid and methyl jasmonate treatment indicating its involvement in response to a variety of stress conditions in banana. Further, transient overexpression of MusaWRKY71 in transformed banana cells led to the induction of several genes, homologues of which have been proven to be involved in diverse stress responses in other important plants. The present study is the first report on characterization of a banana stress-related transcription factor using transformed banana cells.

  16. CsWRKY46, a WRKY transcription factor from cucumber, confers cold resistance in transgenic-plant by regulating a set of cold-stress responsive genes in an ABA-dependent manner.

    PubMed

    Zhang, Ying; Yu, Hongjun; Yang, Xueyong; Li, Qiang; Ling, Jian; Wang, Hong; Gu, Xingfang; Huang, Sanwen; Jiang, Weijie

    2016-11-01

    Plant WRKY transcription factors are trans-regulatory proteins that are involved in plant immune responses, development and senescence; however, their roles in abiotic stress are still not well understood, especially in the horticultural crop cucumber. In this study, a novel cucumber WRKY gene, CsWRKY46 was cloned and identified, which was up-regulated in response to cold stress and exogenous abscisic acid (ABA) treatment. CsWRKY46 is belonging to group II of the WRKY family, CsWRKY46 was found exclusively in the nucleus, as indicated by a transient expression assay. Yeast one-hybrid assay shown that CsWRKY46 interact with the W-box in the promoter of ABI5. Transgenic Arabidopsis lines over-expressing CsWRKY46, WRK46-OE1 and WRK46-OE5 had higher seedling survival rates upon freezing treatment compared with that of the wild-type. The above over-expression lines also showed much a higher proline accumulation, less electrolyte leakage and lower malondialdehyde (MDA) levels. Furthermore, the CsWRKY46 overexpression lines were hypersensitive to ABA during seed germination, but the seedlings were not. Quantitative RT-PCR analyses revealed that the expression levels of the ABA-responsive transcription factor ABI5 were higher in the WRKY46-OE lines than in wild-type and that the overexpression of CsWRKY46 increased the expression of stress-inducible genes, including RD29A and COR47. Taken together, our results demonstrated that CsWRKY46 from cucumber conferred cold tolerance to transgenic plants and positively regulated the cold signaling pathway in an ABA-dependent manner.

  17. Arabidopsis flowering locus D influences systemic-acquired-resistance- induced expression and histone modifications of WRKY genes.

    PubMed

    Singh, Vijayata; Roy, Shweta; Singh, Deepjyoti; Nandi, Ashis Kumar

    2014-03-01

    A plant that is in part infected by a pathogen is more resistant throughout its whole body to subsequent infections--a phenomenon known as systemic acquired resistance (SAR). Mobile signals are synthesized at the site of infection and distributed throughout the plant through vascular tissues. Mechanism of SAR development subsequent to reaching the mobile signal in the distal tissue is largely unknown. Recently we showed that flowering locus D (FLD) gene of Arabidopsis thaliana is required in the distal tissue to activate SAR. FLD codes for a homologue of human-lysine-specific histone demethylase. Here we show that FLD function is required for priming (SAR induced elevated expression during challenge inoculation) of WRKY29 and WRKY6 genes. FLD also differentially influences basal and SAR-induced expression of WRKY38, WRKY65 and WRKY53 genes. In addition, we also show that FLD partly localizes in nucleus and influences histone modifications at the promoters of WRKY29 and WRKY6 genes. The results altogether indicate to the possibility of FLD's involvement in epigenetic regulation of SAR.

  18. Phylogenetic and comparative gene expression analysis of barley (Hordeum vulgare)WRKY transcription factor family reveals putatively retained functions betweenmonocots and dicots

    SciTech Connect

    Mangelsen, Elke; Kilian, Joachim; Berendzen, Kenneth W.; Kolukisaoglu, Uner; Harter, Klaus; Jansson, Christer; Wanke, Dierk

    2008-02-01

    WRKY proteins belong to the WRKY-GCM1 superfamily of zinc finger transcription factors that have been subject to a large plant-specific diversification. For the cereal crop barley (Hordeum vulgare), three different WRKY proteins have been characterized so far, as regulators in sucrose signaling, in pathogen defense, and in response to cold and drought, respectively. However, their phylogenetic relationship remained unresolved. In this study, we used the available sequence information to identify a minimum number of 45 barley WRKY transcription factor (HvWRKY) genes. According to their structural features the HvWRKY factors were classified into the previously defined polyphyletic WRKY subgroups 1 to 3. Furthermore, we could assign putative orthologs of the HvWRKY proteins in Arabidopsis and rice. While in most cases clades of orthologous proteins were formed within each group or subgroup, other clades were composed of paralogous proteins for the grasses and Arabidopsis only, which is indicative of specific gene radiation events. To gain insight into their putative functions, we examined expression profiles of WRKY genes from publicly available microarray data resources and found group specific expression patterns. While putative orthologs of the HvWRKY transcription factors have been inferred from phylogenetic sequence analysis, we performed a comparative expression analysis of WRKY genes in Arabidopsis and barley. Indeed, highly correlative expression profiles were found between some of the putative orthologs. HvWRKY genes have not only undergone radiation in monocot or dicot species, but exhibit evolutionary traits specific to grasses. HvWRKY proteins exhibited not only sequence similarities between orthologs with Arabidopsis, but also relatedness in their expression patterns. This correlative expression is indicative for a putative conserved function of related WRKY proteins in mono- and dicot species.

  19. The WRKY45-2 WRKY13 WRKY42 transcriptional regulatory cascade is required for rice resistance to fungal pathogen.

    PubMed

    Cheng, Hongtao; Liu, Hongbo; Deng, Yong; Xiao, Jinghua; Li, Xianghua; Wang, Shiping

    2015-03-01

    Blast caused by fungal Magnaporthe oryzae is a devastating disease of rice (Oryza sativa) worldwide, and this fungus also infects barley (Hordeum vulgare). At least 11 rice WRKY transcription factors have been reported to regulate rice response to M. oryzae either positively or negatively. However, the relationships of these WRKYs in the rice defense signaling pathway against M. oryzae are unknown. Previous studies have revealed that rice WRKY13 (as a transcriptional repressor) and WRKY45-2 enhance resistance to M. oryzae. Here, we show that rice WRKY42, functioning as a transcriptional repressor, suppresses resistance to M. oryzae. WRKY42-RNA interference (RNAi) and WRKY42-overexpressing (oe) plants showed increased resistance and susceptibility to M. oryzae, accompanied by increased or reduced jasmonic acid (JA) content, respectively, compared with wild-type plants. JA pretreatment enhanced the resistance of WRKY42-oe plants to M. oryzae. WRKY13 directly suppressed WRKY42. WRKY45-2, functioning as a transcriptional activator, directly activated WRKY13. In addition, WRKY13 directly suppressed WRKY45-2 by feedback regulation. The WRKY13-RNAi WRKY45-2-oe and WRKY13-oe WRKY42-oe double transgenic lines showed increased susceptibility to M. oryzae compared with WRKY45-2-oe and WRKY13-oe plants, respectively. These results suggest that the three WRKYs form a sequential transcriptional regulatory cascade. WRKY42 may negatively regulate rice response to M. oryzae by suppressing JA signaling-related genes, and WRKY45-2 transcriptionally activates WRKY13, whose encoding protein in turn transcriptionally suppresses WRKY42 to regulate rice resistance to M. oryzae.

  20. Transposon-derived small RNA is responsible for modified function of WRKY45 locus.

    PubMed

    Zhang, Haitao; Tao, Zeng; Hong, Hanming; Chen, Zhihui; Wu, Changyin; Li, Xianghua; Xiao, Jinghua; Wang, Shiping

    2016-02-29

    Transposable elements (TEs) are an important source for generating small interfering RNAs (siRNAs) in plants and animals. Although TE-siRNA-induced silencing of TEs by RNA-directed DNA methylation (RdDM) in the maintenance of genome integrity has been intensively studied, it is unknown whether this type of silencing occurs in suppressing endogenous non-TE genes during host-pathogen interactions. Here we show that a TE-siRNA, TE-siR815, causes opposite functions for the two alleles, WRKY45-1 and WRKY45-2, of the WRKY45 transcription factor in rice resistance to Xanthomonas oryzae pv. oryzae, which causes the most devastating bacterial disease in rice worldwide. Expression of WRKY45-1, but not WRKY45-2, generated TE-siR815, which in turn repressed ST1, an important component in WRKY45-mediated resistance, by RdDM. Suppression of ST1 abolished WRKY45-mediated resistance leading to pathogen susceptibility. These results suggest that TE-siR815 contributes to the natural variation of the WRKY45 locus and TE-siR815-induced suppression of ST1 results in the negative role of WRKY45-1 but positive role of WRKY45-2 in regulating disease resistance.

  1. Overexpression of the pathogen-inducible wheat TaWRKY45 gene confers disease resistance to multiple fungi in transgenic wheat plants.

    PubMed

    Bahrini, Insaf; Ogawa, Taiichi; Kobayashi, Fuminori; Kawahigashi, Hiroyuki; Handa, Hirokazu

    2011-12-01

    Recently we cloned and characterized the gene for the wheat transcription factor TaWRKY45 and showed that TaWRKY45 was upregulated in response to benzothiadiazole (BTH) and Fusarium head blight (FHB) and that its overexpression conferred enhanced resistance against F. graminearum. To characterize the functional role of TaWRKY45 in the disease resistance of wheat, in the present study we conducted expression analyses of TaWRKY45 with inoculations of powdery mildew and leaf rust and evaluated TaWRKY45-overexpressing wheat plants for resistance to these diseases. TaWRKY45 was upregulated in response to infections with Blumeria graminis, a causal fungus for powdery mildew, and Puccinia triticina, a causal fungus for leaf rust. Constitutive overexpression of the TaWRKY45 transgene conferred enhanced resistance against these two fungi on transgenic wheat plants grown under greenhouse conditions. However, the expression of two resistance-related genes, Pm3 and Lr34, was not induced by the inoculation with powdery mildew in TaWRKY45-overexpressing wheat plants. These results suggest that TaWRKY45 is involved in the defense responses for multiple fungal diseases in wheat but that resistance involving TaWRKY45 differs from at least Pm3 and/or Lr34-related resistance. Our present and previous studies indicate that TaWRKY45 may be potentially utilized to improve a wide range of disease resistance in wheat.

  2. Overexpression of the pathogen-inducible wheat TaWRKY45 gene confers disease resistance to multiple fungi in transgenic wheat plants

    PubMed Central

    Bahrini, Insaf; Ogawa, Taiichi; Kobayashi, Fuminori; Kawahigashi, Hiroyuki; Handa, Hirokazu

    2011-01-01

    Recently we cloned and characterized the gene for the wheat transcription factor TaWRKY45 and showed that TaWRKY45 was upregulated in response to benzothiadiazole (BTH) and Fusarium head blight (FHB) and that its overexpression conferred enhanced resistance against F. graminearum. To characterize the functional role of TaWRKY45 in the disease resistance of wheat, in the present study we conducted expression analyses of TaWRKY45 with inoculations of powdery mildew and leaf rust and evaluated TaWRKY45-overexpressing wheat plants for resistance to these diseases. TaWRKY45 was upregulated in response to infections with Blumeria graminis, a causal fungus for powdery mildew, and Puccinia triticina, a causal fungus for leaf rust. Constitutive overexpression of the TaWRKY45 transgene conferred enhanced resistance against these two fungi on transgenic wheat plants grown under greenhouse conditions. However, the expression of two resistance-related genes, Pm3 and Lr34, was not induced by the inoculation with powdery mildew in TaWRKY45-overexpressing wheat plants. These results suggest that TaWRKY45 is involved in the defense responses for multiple fungal diseases in wheat but that resistance involving TaWRKY45 differs from at least Pm3 and/or Lr34-related resistance. Our present and previous studies indicate that TaWRKY45 may be potentially utilized to improve a wide range of disease resistance in wheat. PMID:23136468

  3. Genome-Wide Analysis of the Musa WRKY Gene Family: Evolution and Differential Expression during Development and Stress

    PubMed Central

    Goel, Ridhi; Pandey, Ashutosh; Trivedi, Prabodh K.; Asif, Mehar H.

    2016-01-01

    The WRKY gene family plays an important role in the development and stress responses in plants. As information is not available on the WRKY gene family in Musa species, genome-wide analysis has been carried out in this study using available genomic information from two species, Musa acuminata and Musa balbisiana. Analysis identified 147 and 132 members of the WRKY gene family in M. acuminata and M. balbisiana, respectively. Evolutionary analysis suggests that the WRKY gene family expanded much before the speciation in both the species. Most of the orthologs retained in two species were from the γ duplication event which occurred prior to α and β genome-wide duplication (GWD) events. Analysis also suggests that subtle changes in nucleotide sequences during the course of evolution have led to the development of new motifs which might be involved in neo-functionalization of different WRKY members in two species. Expression and cis-regulatory motif analysis suggest possible involvement of Group II and Group III WRKY members during various stresses and growth/development including fruit ripening process respectively. PMID:27014321

  4. Identification and expression analysis of WRKY transcription factor genes in canola (Brassica napus L.) in response to fungal pathogens and hormone treatments

    PubMed Central

    Yang, Bo; Jiang, Yuanqing; Rahman, Muhammad H; Deyholos, Michael K; Kav, Nat NV

    2009-01-01

    Background Members of plant WRKY transcription factor families are widely implicated in defense responses and various other physiological processes. For canola (Brassica napus L.), no WRKY genes have been described in detail. Because of the economic importance of this crop, and its evolutionary relationship to Arabidopsis thaliana, we sought to characterize a subset of canola WRKY genes in the context of pathogen and hormone responses. Results In this study, we identified 46 WRKY genes from canola by mining the expressed sequence tag (EST) database and cloned cDNA sequences of 38 BnWRKYs. A phylogenetic tree was constructed using the conserved WRKY domain amino acid sequences, which demonstrated that BnWRKYs can be divided into three major groups. We further compared BnWRKYs to the 72 WRKY genes from Arabidopsis and 91 WRKY from rice, and we identified 46 presumptive orthologs of AtWRKY genes. We examined the subcellular localization of four BnWRKY proteins using green fluorescent protein (GFP) and we observed the fluorescent green signals in the nucleus only. The responses of 16 selected BnWRKY genes to two fungal pathogens, Sclerotinia sclerotiorum and Alternaria brassicae, were analyzed by quantitative real time-PCR (qRT-PCR). Transcript abundance of 13 BnWRKY genes changed significantly following pathogen challenge: transcripts of 10 WRKYs increased in abundance, two WRKY transcripts decreased after infection, and one decreased at 12 h post-infection but increased later on (72 h). We also observed that transcript abundance of 13/16 BnWRKY genes was responsive to one or more hormones, including abscisic acid (ABA), and cytokinin (6-benzylaminopurine, BAP) and the defense signaling molecules jasmonic acid (JA), salicylic acid (SA), and ethylene (ET). We compared these transcript expression patterns to those previously described for presumptive orthologs of these genes in Arabidopsis and rice, and observed both similarities and differences in expression patterns

  5. Gene expression analysis of WRKY transcription factors in Arabidopsis thaliana cell cultures during a parabolic flight

    NASA Astrophysics Data System (ADS)

    Babbick, Maren; Barjaktarović, Žarko; Hampp, Ruediger

    Plants sense gravity by specialized cells (statocytes) and adjust growth and development accordingly. It has, however, also been shown that plant cells which are not part of specialized tissues are also able to sense gravitational forces. Therefore we used undifferentiated, homogeneous cell cultures of Arabidopsis thaliana (cv. Columbia) in order to identify early alterations in gene expression as a response to altered gravitational field strengths. In this contribution we report on cell cultures exposed to parabolic flights (approximately 20 sec of microgravity). For this short-term exposure study, we specifically checked for genes at the beginning of signal transduction chains, such as those coding for transcription factors (TFs). TFs are small proteins that regulate expression of their target genes by binding to specific promoter sequences. Our main focus were members of the so-called WRKY TF family. WRKY TFs are known to be involved in various physiological processes like senescence and pathogen defense. By quantifying transcriptional changes of these genes by real-time RT-PCR, we wanted to find out, how gene expression is affected by both hyperand microgravity conditions during a parabolic flight. For this purpose Arabidopsis thaliana callus cultures were metabolically quenched by the injection of RNAlater at the end of the microgravity-phase of each parabola. The data we present will show how fast changes in amounts of transcripts will occur, and to what degree the expression profiles are comparable with data obtained from exposures to hypergravity and simulated microgravity.

  6. Genome-wide identification and comparative expression analysis reveal a rapid expansion and functional divergence of duplicated genes in the WRKY gene family of cabbage, Brassica oleracea var. capitata.

    PubMed

    Yao, Qiu-Yang; Xia, En-Hua; Liu, Fei-Hu; Gao, Li-Zhi

    2015-02-15

    WRKY transcription factors (TFs), one of the ten largest TF families in higher plants, play important roles in regulating plant development and resistance. To date, little is known about the WRKY TF family in Brassica oleracea. Recently, the completed genome sequence of cabbage (B. oleracea var. capitata) allows us to systematically analyze WRKY genes in this species. A total of 148 WRKY genes were characterized and classified into seven subgroups that belong to three major groups. Phylogenetic and synteny analyses revealed that the repertoire of cabbage WRKY genes was derived from a common ancestor shared with Arabidopsis thaliana. The B. oleracea WRKY genes were found to be preferentially retained after the whole-genome triplication (WGT) event in its recent ancestor, suggesting that the WGT event had largely contributed to a rapid expansion of the WRKY gene family in B. oleracea. The analysis of RNA-Seq data from various tissues (i.e., roots, stems, leaves, buds, flowers and siliques) revealed that most of the identified WRKY genes were positively expressed in cabbage, and a large portion of them exhibited patterns of differential and tissue-specific expression, demonstrating that these gene members might play essential roles in plant developmental processes. Comparative analysis of the expression level among duplicated genes showed that gene expression divergence was evidently presented among cabbage WRKY paralogs, indicating functional divergence of these duplicated WRKY genes.

  7. Banana fruit VQ motif-containing protein5 represses cold-responsive transcription factor MaWRKY26 involved in the regulation of JA biosynthetic genes

    PubMed Central

    Ye, Yu-Jie; Xiao, Yun-Yi; Han, Yan-Chao; Shan, Wei; Fan, Zhong-Qi; Xu, Qun-Gang; Kuang, Jian-Fei; Lu, Wang-Jin; Lakshmanan, Prakash; Chen, Jian-Ye

    2016-01-01

    Most harvested fruits and vegetables are stored at low temperature but many of them are highly sensitive to chilling injury. Jasmonic acid (JA), a plant hormone associated with various stress responses, is known to reduce chilling injury in fruits. However, little is known about the transcriptional regulation of JA biosynthesis in relation to cold response of fruits. Here, we show the involvement of a Group I WRKY transcription factor (TF) from banana fruit, MaWRKY26, in regulating JA biosynthesis. MaWRKY26 was found to be nuclear-localized with transcriptional activation property. MaWRKY26 was induced by cold stress or by methyl jasmonate (MeJA), which enhances cold tolerance in banana fruit. More importantly, MaWRKY26 transactivated JA biosynthetic genes MaLOX2, MaAOS3 and MaOPR3 via binding to their promoters. Further, MaWRKY26 physically interacted with a VQ motif-containing protein MaVQ5, and the interaction attenuated MaWRKY26-induced transactivation of JA biosynthetic genes. These results strongly suggest that MaVQ5 might act as a repressor of MaWRKY26 in activating JA biosynthesis. Taken together, our findings provide new insights into the transcriptional regulation of JA biosynthesis in response to cold stress and a better understanding of the molecular aspects of chilling injury in banana fruit. PMID:27004441

  8. The WRKY45-2 WRKY13 WRKY42 Transcriptional Regulatory Cascade Is Required for Rice Resistance to Fungal Pathogen1[OPEN

    PubMed Central

    Cheng, Hongtao; Liu, Hongbo; Deng, Yong; Xiao, Jinghua; Li, Xianghua; Wang, Shiping

    2015-01-01

    Blast caused by fungal Magnaporthe oryzae is a devastating disease of rice (Oryza sativa) worldwide, and this fungus also infects barley (Hordeum vulgare). At least 11 rice WRKY transcription factors have been reported to regulate rice response to M. oryzae either positively or negatively. However, the relationships of these WRKYs in the rice defense signaling pathway against M. oryzae are unknown. Previous studies have revealed that rice WRKY13 (as a transcriptional repressor) and WRKY45-2 enhance resistance to M. oryzae. Here, we show that rice WRKY42, functioning as a transcriptional repressor, suppresses resistance to M. oryzae. WRKY42-RNA interference (RNAi) and WRKY42-overexpressing (oe) plants showed increased resistance and susceptibility to M. oryzae, accompanied by increased or reduced jasmonic acid (JA) content, respectively, compared with wild-type plants. JA pretreatment enhanced the resistance of WRKY42-oe plants to M. oryzae. WRKY13 directly suppressed WRKY42. WRKY45-2, functioning as a transcriptional activator, directly activated WRKY13. In addition, WRKY13 directly suppressed WRKY45-2 by feedback regulation. The WRKY13-RNAi WRKY45-2-oe and WRKY13-oe WRKY42-oe double transgenic lines showed increased susceptibility to M. oryzae compared with WRKY45-2-oe and WRKY13-oe plants, respectively. These results suggest that the three WRKYs form a sequential transcriptional regulatory cascade. WRKY42 may negatively regulate rice response to M. oryzae by suppressing JA signaling-related genes, and WRKY45-2 transcriptionally activates WRKY13, whose encoding protein in turn transcriptionally suppresses WRKY42 to regulate rice resistance to M. oryzae. PMID:25624395

  9. Phylogeny and historical biogeography of the cocosoid palms (Arecaceae, Arecoideae, Cocoseae) inferred from sequences of six WRKY gene family loci

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Arecaceae tribe Cocoseae is the most economically important tribe of palms, including both coconut and African oil palm. It is mostly represented in the Neotropics, with one and two genera endemic to South Africa and Madagascar, respectively. Using primers for six single copy WRKY gene family loci...

  10. Putative WRKYs associated with regulation of fruit ripening revealed by detailed expression analysis of the WRKY gene family in pepper

    PubMed Central

    Cheng, Yuan; JalalAhammed, Golam; Yu, Jiahong; Yao, Zhuping; Ruan, Meiying; Ye, Qingjing; Li, Zhimiao; Wang, Rongqing; Feng, Kun; Zhou, Guozhi; Yang, Yuejian; Diao, Weiping; Wan, Hongjian

    2016-01-01

    WRKY transcription factors play important roles in plant development and stress responses. Here, global expression patterns of pepper CaWRKYs in various tissues as well as response to environmental stresses and plant hormones were systematically analyzed, with an emphasis on fruit ripening. The results showed that most CaWRKYs were expressed in at least two of the tissues tested. Group I, a subfamily of the entire CaWRKY gene family, had a higher expression level in vegetative tissues, whereas groups IIa and III showed relatively lower expression levels. Comparative analysis showed that the constitutively highly expressed WRKY genes were conserved in tomato and pepper, suggesting potential functional similarities. Among the identified 61 CaWRKYs, almost 60% were expressed during pepper fruit maturation, and the group I genes were in higher proportion during the ripening process, indicating an as-yet unknown function of group I in the fruit maturation process. Further analysis suggested that many CaWRKYs expressed during fruit ripening were also regulated by abiotic stresses or plant hormones, indicating that these CaWRKYs play roles in the stress-related signaling pathways during fruit ripening. This study provides new insights to the current research on CaWRKY and contributes to our knowledge about the global regulatory network in pepper fruit ripening. PMID:27991526

  11. Allelic selection of human IL-2 gene.

    PubMed

    Matesanz, F; Delgado, C; Fresno, M; Alcina, A

    2000-12-01

    The allelic expression of mouse IL-2 cannot be definitely extrapolated to what might happen in humans. Therefore, we investigated the regulation of allelic expression of the IL-2 gene in non-genetically manipulated human T lymphocytes by following natural allelic polymorphisms. We found a phenotypically silent punctual change in the human IL-2 at position 114 after the first nucleotide of the initiation codon, which represents a dimorphic polymorphism at the first exon of the IL-2 gene. This allowed the study by single-cell PCR of the regulation of the human IL-2 allelic expression in heterozygous CD4(+) T cells, which was found to be tightly controlled monoallelically. These findings may be used as a suitable marker for monitoring the IL-2 allelic contribution to effector activities and in immune responses against different infections or in pathological situations.

  12. Evolutionary Expansion of WRKY Gene Family in Banana and Its Expression Profile during the Infection of Root Lesion Nematode, Pratylenchus coffeae

    PubMed Central

    Suthanthiram, Backiyarani; Subbaraya, Uma; Marimuthu Somasundram, Saraswathi; Muthu, Mayilvaganan

    2016-01-01

    The WRKY family of transcription factors orchestrate the reprogrammed expression of the complex network of defense genes at various biotic and abiotic stresses. Within the last 96 million years, three rounds of Musa polyploidization events had occurred from selective pressure causing duplication of MusaWRKYs with new activities. Here, we identified a total of 153 WRKY transcription factors available from the DH Pahang genome. Based on their phylogenetic relationship, the MusaWRKYs available with complete gene sequence were classified into the seven common WRKY sub-groups. Synteny analyses data revealed paralogous relationships, with 17 MusaWRKY gene pairs originating from the duplication events that had occurred within the Musa lineage. We also found 15 other MusaWRKY gene pairs originating from much older duplication events that had occurred along Arecales and Poales lineage of commelinids. Based on the synonymous and nonsynonymous substitution rates, the fate of duplicated MusaWRKY genes was predicted to have undergone sub-functionalization in which the duplicated gene copies retain a subset of the ancestral gene function. Also, to understand the regulatory roles of MusaWRKY during a biotic stress, Illumina sequencing was performed on resistant and susceptible cultivars during the infection of root lesion nematode, Pratylenchus coffeae. The differential WRKY gene expression analysis in nematode resistant and susceptible cultivars during challenged and unchallenged conditions had distinguished: 1) MusaWRKYs participating in general banana defense mechanism against P.coffeae common to both susceptible and resistant cultivars, 2) MusaWRKYs that may aid in the pathogen survival as suppressors of plant triggered immunity, 3) MusaWRKYs that may aid in the host defense as activators of plant triggered immunity and 4) cultivar specific MusaWRKY regulation. Mainly, MusaWRKY52, -69 and -92 are found to be P.coffeae specific and can act as activators or repressors in a

  13. Analysis of carotenogenic genes promoters and WRKY transcription factors in response to salt stress in Dunaliella bardawil

    PubMed Central

    Liang, Ming-Hua; Jiang, Jian-Guo

    2017-01-01

    The unicellular alga Dunaliella bardawil is a highly salt-tolerant organism, capable of accumulating glycerol, glycine betaine and β-carotene under salt stress, and has been considered as an excellent model organism to investigate the molecular mechanisms of salt stress responses. In this study, several carotenogenic genes (DbCRTISO, DbZISO, DbLycE and DbChyB), DbBADH genes involved in glycine betaine synthesis and genes encoding probable WRKY transcription factors from D. bardawil were isolated, and promoters of DbCRTISO and DbChyB were cloned. The promoters of DbPSY, DbLycB, DbGGPS, DbCRTISO and DbChyB contained the salt-regulated element (SRE), GT1GMSCAM4, while the DbGGPS promoter has another SRE, DRECRTCOREAT. All promoters of the carotenogenic genes had light-regulated elements and W-box cis-acting elements. Most WRKY transcription factors can bind to the W-box, and play roles in abiotic stress. qRT-PCR analysis showed that salt stress up-regulated both carotenogenic genes and WRKY transcription factors. In contrast, the transcription levels of DbBADH showed minor changes. In D. bardawil, it appears that carotenoid over-accumulation allows for the long-term adaptation to salt stress, while the rapid modulation of glycine betaine biosynthesis provides an initial response. PMID:28128303

  14. Genome-wide identification of WRKY family genes and their response to cold stress in Vitis vinifera

    PubMed Central

    2014-01-01

    Background WRKY transcription factors are one of the largest families of transcriptional regulators in plants. WRKY genes are not only found to play significant roles in biotic and abiotic stress response, but also regulate growth and development. Grapevine (Vitis vinifera) production is largely limited by stressful climate conditions such as cold stress and the role of WRKY genes in the survival of grapevine under these conditions remains unknown. Results We identified a total of 59 VvWRKYs from the V. vinifera genome, belonging to four subgroups according to conserved WRKY domains and zinc-finger structure. The majority of VvWRKYs were expressed in more than one tissue among the 7 tissues examined which included young leaves, mature leaves, tendril, stem apex, root, young fruits and ripe fruits. Publicly available microarray data suggested that a subset of VvWRKYs was activated in response to diverse stresses. Quantitative real-time PCR (qRT-PCR) results demonstrated that the expression levels of 36 VvWRKYs are changed following cold exposure. Comparative analysis was performed on data from publicly available microarray experiments, previous global transcriptome analysis studies, and qRT-PCR. We identified 15 VvWRKYs in at least two of these databases which may relate to cold stress. Among them, the transcription of three genes can be induced by exogenous ABA application, suggesting that they can be involved in an ABA-dependent signaling pathway in response to cold stress. Conclusions We identified 59 VvWRKYs from the V. vinifera genome and 15 of them showed cold stress-induced expression patterns. These genes represented candidate genes for future functional analysis of VvWRKYs involved in the low temperature-related signal pathways in grape. PMID:24755338

  15. Genome-Wide Analysis of the Expression of WRKY Family Genes in Different Developmental Stages of Wild Strawberry (Fragaria vesca) Fruit

    PubMed Central

    Zhang, Qing; Ren, Suyue; Shen, Yuanyue; Qin, Ling; Xing, Yu

    2016-01-01

    WRKY proteins play important regulatory roles in plant developmental processes such as senescence, trichome initiation and embryo morphogenesis. In strawberry, only FaWRKY1 (Fragaria × ananassa) has been characterized, leaving numerous WRKY genes to be identified and their function characterized. The publication of the draft genome sequence of the strawberry genome allowed us to conduct a genome-wide search for WRKY proteins in Fragaria vesca, and to compare the identified proteins with their homologs in model plants. Fifty-nine FvWRKY genes were identified and annotated from the F. vesca genome. Detailed analysis, including gene classification, annotation, phylogenetic evaluation, conserved motif determination and expression profiling, based on RNA-seq data, were performed on all members of the family. Additionally, the expression patterns of the WRKY genes in different fruit developmental stages were further investigated using qRT-PCR, to provide a foundation for further comparative genomics and functional studies of this important class of transcriptional regulators in strawberry. PMID:27138272

  16. WRKY72-type transcription factors contribute to basal immunity in tomato and Arabidopsis as well as gene-for-gene resistance mediated by the tomato R gene Mi-1.

    PubMed

    Bhattarai, Kishor K; Atamian, Hagop S; Kaloshian, Isgouhi; Eulgem, Thomas

    2010-07-01

    WRKY transcription factors play a central role in transcriptional reprogramming associated with plant immune responses. However, due to functional redundancy, typically the contribution of individual members of this family to immunity is only subtle. Using microarray analysis, we found that the paralogous tomato WRKY genes SlWRKY72a and b are transcriptionally up-regulated during disease resistance mediated by the R gene Mi-1. Virus-induced gene silencing of these two genes in tomato resulted in a clear reduction of Mi-1-mediated resistance as well as basal defense against root-knot nematodes (RKN) and potato aphids. Using Arabidopsis T-DNA insertion mutants, we found that their Arabidopsis ortholog, AtWRKY72, is also required for full basal defense against RKN as well as to the oomycete Hyaloperonospora arabidopsidis. Despite their similar roles in basal defense against RKN in both tested plant species, WRKY72-type transcription factors in tomato, but not in Arabidopsis, clearly contributed to basal defense against the bacterial pathogen Pseudomonas syringae. Of the five R genes that we tested in tomato and Arabidopsis, only Mi-1 appeared to be dependent on WRKY72-type transcription factors. Interestingly, AtWRKY72 target genes, identified by microarray analysis of H. arabidopsidis-triggered transcriptional changes, appear to be largely non-responsive to analogs of the defense hormone salicylic acid (SA). Thus, similarly to Mi-1, which in part acts independently of SA, AtWRKY72 appears to utilize SA-independent defense mechanisms. We propose that WRKY72-type transcription factors play a partially conserved role in basal defense in tomato and Arabidopsis, a function that has been recruited to serve Mi-1-dependent immunity.

  17. In silico mining and PCR-based approaches to transcription factor discovery in non-model plants: gene discovery of the WRKY transcription factors in conifers.

    PubMed

    Liu, Jun-Jun; Xiang, Yu

    2011-01-01

    WRKY transcription factors are key regulators of numerous biological processes in plant growth and development, as well as plant responses to abiotic and biotic stresses. Research on biological functions of plant WRKY genes has focused in the past on model plant species or species with largely characterized transcriptomes. However, a variety of non-model plants, such as forest conifers, are essential as feed, biofuel, and wood or for sustainable ecosystems. Identification of WRKY genes in these non-model plants is equally important for understanding the evolutionary and function-adaptive processes of this transcription factor family. Because of limited genomic information, the rarity of regulatory gene mRNAs in transcriptomes, and the sequence divergence to model organism genes, identification of transcription factors in non-model plants using methods similar to those generally used for model plants is difficult. This chapter describes a gene family discovery strategy for identification of WRKY transcription factors in conifers by a combination of in silico-based prediction and PCR-based experimental approaches. Compared to traditional cDNA library screening or EST sequencing at transcriptome scales, this integrated gene discovery strategy provides fast, simple, reliable, and specific methods to unveil the WRKY gene family at both genome and transcriptome levels in non-model plants.

  18. Functional analysis of structurally related soybean GmWRKY58 and GmWRKY76 in plant growth and development.

    PubMed

    Yang, Yan; Chi, Yingjun; Wang, Ze; Zhou, Yuan; Fan, Baofang; Chen, Zhixiang

    2016-08-01

    WRKY transcription factors constitute a large protein superfamily with a predominant role in plant stress responses. In this study we report that two structurally related soybean WRKY proteins, GmWRKY58 and GmWRKY76, play a critical role in plant growth and flowering. GmWRKY58 and GmWRKY76 are both Group III WRKY proteins with a C2HC zinc finger domain and are close homologs of AtWRKY70 and AtWRKY54, two well-characterized Arabidopsis WRKY proteins with an important role in plant responses to biotic and abiotic stresses. GmWRKY58 and GmWRKY76 are both localized to the nucleus, recognize the TTGACC W-box sequence with a high specificity, and function as transcriptional activators in both yeast and plant cells. Expression of GmWRKY58 and GmWRKY76 was detected at low levels in roots, stem, leaves, flowers, and pods. Expression of the two genes in leaves increased substantially during the first 4 weeks after germination but steadily declined thereafter with increased age. To determine their biological functions, transgenic Arabidopsis plants were generated overexpressing GmWRKY58 or GmWRKY76 Unlike AtWRKY70 and AtWRKY54, overexpression of GmWRKY58 or GmWRKY76 had no effect on disease resistance and only small effects on abiotic stress tolerance of the transgenic plants. Significantly, transgenic Arabidopsis plants overexpressing GmWRKY58 or GmWRKY76 flowered substantially earlier than control plants and this early flowering phenotype was associated with increased expression of several flowering-promoting genes, some of which are enriched in W-box sequences in their promoters recognized by GmWRKY58 and GmWRKY76. In addition, virus-induced silencing of GmWRKY58 and GmWRKY76 in soybean resulted in stunted plants with reduced leaf expansion and terminated stem growth. These results provide strong evidence for functional divergence among close structural homologs of WRKY proteins from different plant species.

  19. TaWRKY70 transcription factor in wheat QTL-2DL regulates downstream metabolite biosynthetic genes to resist Fusarium graminearum infection spread within spike

    PubMed Central

    Kage, Udaykumar; Yogendra, Kalenahalli N.; Kushalappa, Ajjamada C.

    2017-01-01

    A semi-comprehensive metabolomics was used to identify the candidate metabolites and genes to decipher mechanisms of resistance in wheat near-isogenic lines (NILs) containing QTL-2DL against Fusarium graminearum (Fg). Metabolites, with high fold-change in abundance, belonging to hydroxycinnamic acid amides (HCAAs): such as coumaroylagmatine, coumaroylputrescine and Fatty acids: phosphatidic acids (PAs) were identified as resistance related induced (RRI) metabolites in rachis of resistant NIL (NIL-R), inoculated with Fg. A WRKY like transcription factor (TF) was identified within the QTL-2DL region, along with three resistance genes that biosynthesized RRI metabolites. Sequencing and in-silico analysis of WRKY confirmed it to be wheat TaWRKY70. Quantitative real time-PCR studies showed a higher expression of TaWRKY70 in NIL-R as compared to NIL-S after Fg inoculation. Further, the functional validation of TaWRKY70 based on virus induced gene silencing (VIGS) in NIL-R, not only confirmed an increased fungal biomass but also decreased expressions of downstream resistance genes: TaACT, TaDGK and TaGLI1, along with decreased abundances of RRI metabolites biosynthesized by them. Among more than 200 FHB resistance QTL identified in wheat, this is the first QTL from which a TF was identified, and its downstream target genes as well as the FHB resistance functions were deciphered. PMID:28198421

  20. OsWRKY74, a WRKY transcription factor, modulates tolerance to phosphate starvation in rice

    PubMed Central

    Dai, Xiaoyan; Wang, Yuanyuan; Zhang, Wen-Hao

    2016-01-01

    The WRKY transcription factor family has 109 members in the rice genome, and has been reported to be involved in the regulation of biotic and abiotic stress in plants. Here, we demonstrated that a rice OsWRKY74 belonging to group III of the WRKY transcription factor family was involved in tolerance to phosphate (Pi) starvation. OsWRKY74 was localized in the nucleus and mainly expressed in roots and leaves. Overexpression of OsWRKY74 significantly enhanced tolerance to Pi starvation, whereas transgenic lines with down-regulation of OsWRKY74 were sensitive to Pi starvation. Root and shoot biomass, and phosphorus (P) concentration in rice OsWRKY74-overexpressing plants were ~16% higher than those of wild-type (WT) plants in Pi-deficient hydroponic solution. In soil pot experiments, >24% increases in tiller number, grain weight and P concentration were observed in rice OsWRKY74-overexpressing plants compared to WT plants when grown in P-deficient medium. Furthermore, Pi starvation-induced changes in root system architecture were more profound in OsWRKY74-overexpressing plants than in WT plants. Expression patterns of a number of Pi-responsive genes were altered in the OsWRKY74-overexpressing and RNA interference lines. In addition, OsWRKY74 may also be involved in the response to deficiencies in iron (Fe) and nitrogen (N) as well as cold stress in rice. In Pi-deficient conditions, OsWRKY74-overexpressing plants exhibited greater accumulation of Fe and up-regulation of the cold-responsive genes than WT plants. These findings highlight the role of OsWRKY74 in modulation of Pi homeostasis and potential crosstalk between P starvation and Fe starvation, and cold stress in rice. PMID:26663563

  1. Phylogenetic Analysis of Seven WRKY Genes across the Palm Subtribe Attaleinae (Arecaceae) Identifies Syagrus as Sister Group of the Coconut

    PubMed Central

    Meerow, Alan W.; Noblick, Larry; Borrone, James W.; Couvreur, Thomas L. P.; Mauro-Herrera, Margarita; Hahn, William J.; Kuhn, David N.; Nakamura, Kyoko; Oleas, Nora H.; Schnell, Raymond J.

    2009-01-01

    Background The Cocoseae is one of 13 tribes of Arecaceae subfam. Arecoideae, and contains a number of palms with significant economic importance, including the monotypic and pantropical Cocos nucifera L., the coconut, the origins of which have been one of the “abominable mysteries” of palm systematics for decades. Previous studies with predominantly plastid genes weakly supported American ancestry for the coconut but ambiguous sister relationships. In this paper, we use multiple single copy nuclear loci to address the phylogeny of the Cocoseae subtribe Attaleinae, and resolve the closest extant relative of the coconut. Methodology/Principal Findings We present the results of combined analysis of DNA sequences of seven WRKY transcription factor loci across 72 samples of Arecaceae tribe Cocoseae subtribe Attaleinae, representing all genera classified within the subtribe, and three outgroup taxa with maximum parsimony, maximum likelihood, and Bayesian approaches, producing highly congruent and well-resolved trees that robustly identify the genus Syagrus as sister to Cocos and resolve novel and well-supported relationships among the other genera of the Attaleinae. We also address incongruence among the gene trees with gene tree reconciliation analysis, and assign estimated ages to the nodes of our tree. Conclusions/Significance This study represents the as yet most extensive phylogenetic analyses of Cocoseae subtribe Attaleinae. We present a well-resolved and supported phylogeny of the subtribe that robustly indicates a sister relationship between Cocos and Syagrus. This is not only of biogeographic interest, but will also open fruitful avenues of inquiry regarding evolution of functional genes useful for crop improvement. Establishment of two major clades of American Attaleinae occurred in the Oligocene (ca. 37 MYBP) in Eastern Brazil. The divergence of Cocos from Syagrus is estimated at 35 MYBP. The biogeographic and morphological congruence that we see for

  2. Global analysis of WRKY transcription factor superfamily in Setaria identifies potential candidates involved in abiotic stress signaling

    PubMed Central

    Muthamilarasan, Mehanathan; Bonthala, Venkata S.; Khandelwal, Rohit; Jaishankar, Jananee; Shweta, Shweta; Nawaz, Kashif; Prasad, Manoj

    2015-01-01

    Transcription factors (TFs) are major players in stress signaling and constitute an integral part of signaling networks. Among the major TFs, WRKY proteins play pivotal roles in regulation of transcriptional reprogramming associated with stress responses. In view of this, genome- and transcriptome-wide identification of WRKY TF family was performed in the C4model plants, Setaria italica (SiWRKY) and S. viridis (SvWRKY), respectively. The study identified 105 SiWRKY and 44 SvWRKY proteins that were computationally analyzed for their physicochemical properties. Sequence alignment and phylogenetic analysis classified these proteins into three major groups, namely I, II, and III with majority of WRKY proteins belonging to group II (53 SiWRKY and 23 SvWRKY), followed by group III (39 SiWRKY and 11 SvWRKY) and group I (10 SiWRKY and 6 SvWRKY). Group II proteins were further classified into 5 subgroups (IIa to IIe) based on their phylogeny. Domain analysis showed the presence of WRKY motif and zinc finger-like structures in these proteins along with additional domains in a few proteins. All SiWRKY genes were physically mapped on the S. italica genome and their duplication analysis revealed that 10 and 8 gene pairs underwent tandem and segmental duplications, respectively. Comparative mapping of SiWRKY and SvWRKY genes in related C4 panicoid genomes demonstrated the orthologous relationships between these genomes. In silico expression analysis of SiWRKY and SvWRKY genes showed their differential expression patterns in different tissues and stress conditions. Expression profiling of candidate SiWRKY genes in response to stress (dehydration and salinity) and hormone treatments (abscisic acid, salicylic acid, and methyl jasmonate) suggested the putative involvement of SiWRKY066 and SiWRKY082 in stress and hormone signaling. These genes could be potential candidates for further characterization to delineate their functional roles in abiotic stress signaling. PMID:26635818

  3. Virus-induced gene silencing of WRKY53 and an inducible phenylalanine ammonia-lyase in wheat reduces aphid resistance.

    PubMed

    Van Eck, Leon; Schultz, Thia; Leach, Jan E; Scofield, Steven R; Peairs, Frank B; Botha, Anna-Maria; Lapitan, Nora L V

    2010-12-01

    Although several wheat genes differentially expressed during the Russian wheat aphid resistance response have recently been identified, their requirement for and specific role in resistance remain unclear. Progress in wheat-aphid interaction research is hampered by inadequate collections of mutant germplasm and difficulty in transforming hexaploid wheat. Virus-induced gene silencing (VIGS) technology is emerging as a viable reverse genetics approach in cereal crops. However, the potential of VIGS for determining aphid defence gene function in wheat has not been evaluated. We report on the use of recombinant barley stripe mosaic virus (BSMV) to target and silence a WRKY53 transcription factor and an inducible phenylalanine ammonia-lyase (PAL) gene, both predicted to contribute to aphid defence in a genetically resistant wheat line. After inoculating resistant wheat with the VIGS constructs, transcript abundance was reduced to levels similar to that observed in susceptible wheat. Notably, the level of PAL expression was also suppressed by the WKRY53 construct, suggesting that these genes operate in the same defence response network. Both knockdowns exhibited a susceptible phenotype upon aphid infestation, and aphids feeding on silenced plants exhibited a significant increase in fitness compared to aphids feeding on control plants. Altered plant phenotype and changes in aphid behaviour after silencing imply that WKRY53 and PAL play key roles in generating a successful resistance response. This study is the first report on the successful use of VIGS to investigate genes involved in wheat-insect interactions.

  4. WRKY13 acts in stem development in Arabidopsis thaliana.

    PubMed

    Li, Wei; Tian, Zhaoxia; Yu, Diqiu

    2015-07-01

    Stems are important for plants to grow erectly. In stems, sclerenchyma cells must develop secondary cell walls to provide plants with physical support. The secondary cell walls are mainly composed of lignin, xylan and cellulose. Deficiency of overall stem development could cause weakened stems. Here we prove that WRKY13 acts in stem development. The wrky13 mutants take on a weaker stem phenotype. The number of sclerenchyma cells, stem diameter and the number of vascular bundles were reduced in wrky13 mutants. Lignin-synthesis-related genes were repressed in wrky13 mutants. Chromatin immunoprecipitation assays proved that WRKY13 could directly bind to the promoter of NST2. Taken together, we proposed that WRKY13 affected the overall development of stem. Identification of the role of WRKY13 may help to resolve agricultural problems caused by weaker stems.

  5. Both JrWRKY2 and JrWRKY7 of Juglans regia mediate responses to abiotic stresses and abscisic acid through formation of homodimers and interaction.

    PubMed

    Yang, G; Zhang, W; Liu, Z; Yi-Maer, A-Y; Zhai, M; Xu, Z

    2017-03-01

    WRKY transcription factors belong to a large protein family that is involved in diverse developmental processes and abiotic stress responses. Currently, there is little understanding of the role of WRKY transcription factors in regulatory mechanisms in plants, especially in the protein-protein interactions that are essential for biological regulatory functions and networks. In the present study, yeast one-hybrid, yeast two-hybrid, transient expression and quantitative RT-PCR were applied to investigate the potential characteristics of two WRKY proteins from Juglans regia, JrWRKY2 (GenBank Accession No. KU057089) and JrWRKY7 (GenBank Accession No. KP784651). JrWRKY2 and JrWRKY7 can form homodimers and interact with each other. JrWRKY2 and JrWRKY7 can bind to W-box motifs. Similarly high levels of transcription were found for JrWRKY2 and JrWRKY7 under NaCl and polyethylene glycol (PEG) stresses, as well as at different developmental stages, e.g., the pistil or terminal leaf. JrWRKY2 and JrWRKY7 were transiently overexpressed in an independent manner in the terminal leaf. Analyses of superoxide dismutase (SOD) and peroxidase (POD) activities, proline and malondialdehyde (MDA) contents, and electrolyte leakage rate showed that JrWRKY2 and JrWRKY7 overexpression improved plant tolerance to NaCl, PEG, abscisic acid, and cold stress. Additionally, JrWRKY2 and JrWRKY7 overexpression elevated transcription of SOD, POD, glutathione peroxidase (GPX), catalase (CAT), ascorbate peroxidase (APX), and MYB genes, but downregulated the expression of NAC. Overall, the results demonstrate that JrWRKY2 and JrWRKY7 are dimeric proteins that can form functional homodimers and interact with each other and that they are involved in abiotic stress responses.

  6. A Wheat WRKY Transcription Factor TaWRKY10 Confers Tolerance to Multiple Abiotic Stresses in Transgenic Tobacco

    PubMed Central

    Chen, Liulin; Wang, Xiatian; Ma, Hui; Hu, Wei; Yao, Ningcong; Feng, Ying; Chai, Ruihong; Yang, Guangxiao; He, Guangyuan

    2013-01-01

    WRKY transcription factors are reported to be involved in defense regulation, stress response and plant growth and development. However, the precise role of WRKY transcription factors in abiotic stress tolerance is not completely understood, especially in crops. In this study, we identified and cloned 10 WRKY genes from genome of wheat (Triticum aestivum L.). TaWRKY10, a gene induced by multiple stresses, was selected for further investigation. TaWRKY10 was upregulated by treatment with polyethylene glycol, NaCl, cold and H2O2. Result of Southern blot indicates that the wheat genome contains three copies of TaWRKY10. The TaWRKY10 protein is localized in the nucleus and functions as a transcriptional activator. Overexpression of TaWRKY10 in tobacco (Nicotiana tabacum L.) resulted in enhanced drought and salt stress tolerance, mainly demonstrated by the transgenic plants exhibiting of increased germination rate, root length, survival rate, and relative water content under these stress conditions. Further investigation showed that transgenic plants also retained higher proline and soluble sugar contents, and lower reactive oxygen species and malonaldehyde contents. Moreover, overexpression of the TaWRKY10 regulated the expression of a series of stress related genes. Taken together, our results indicate that TaWRKY10 functions as a positive factor under drought and salt stresses by regulating the osmotic balance, ROS scavenging and transcription of stress related genes. PMID:23762295

  7. Evidence for a positive regulatory role of strawberry (Fragaria x ananassa) Fa WRKY1 and Arabidopsis At WRKY75 proteins in resistance.

    PubMed

    Encinas-Villarejo, Sonia; Maldonado, Ana M; Amil-Ruiz, Francisco; de los Santos, Berta; Romero, Fernando; Pliego-Alfaro, Fernando; Muñoz-Blanco, Juan; Caballero, José L

    2009-01-01

    Knowledge of the molecular basis of plant resistance to pathogens in species other than Arabidopsis is limited. The function of Fa WRKY1, the first WRKY gene isolated from strawberry (Fragaria x ananassa), an important agronomical fruit crop, has been investigated here. Fa WRKY1 encodes a IIc WRKY transcription factor and is up-regulated in strawberry following Colletotrichum acutatum infection, treatments with elicitors, and wounding. Its Arabidopsis sequence homologue, At WRKY75, has been described as playing a role in regulating phosphate starvation responses. However, using T-DNA insertion mutants, a role for the At WRKY75 and Fa WRKY1 in the activation of basal and R-mediated resistance in Arabidopsis is demonstrated. At wrky75 mutants are more susceptible to virulent and avirulent isolates of Pseudomonas syringae. Overexpression of Fa WRKY1 in At wrky75 mutant and wild type reverts the enhanced susceptible phenotype of the mutant, and even increases resistance to avirulent strains of P. syringae. The resistance phenotype is uncoupled to PATHOGENESIS-RELATED (PR) gene expression, but it is associated with a strong oxidative burst and glutathione-S-transferase (GST) induction. Taken together, these results indicate that At WRKY75 and Fa WRKY1 act as positive regulators of defence during compatible and incompatible interactions in Arabidopsis and, very likely, Fa WRKY1 is an important element mediating defence responses to C. acutatum in strawberry. Moreover, these results provide evidence that Arabidopsis can be a useful model for functional studies in Rosacea species like strawberry.

  8. A gene feature enumeration approach for describing HLA allele polymorphism.

    PubMed

    Mack, Steven J

    2015-12-01

    HLA genotyping via next generation sequencing (NGS) poses challenges for the use of HLA allele names to analyze and discuss sequence polymorphism. NGS will identify many new synonymous and non-coding HLA sequence variants. Allele names identify the types of nucleotide polymorphism that define an allele (non-synonymous, synonymous and non-coding changes), but do not describe how polymorphism is distributed among the individual features (the flanking untranslated regions, exons and introns) of a gene. Further, HLA alleles cannot be named in the absence of antigen-recognition domain (ARD) encoding exons. Here, a system for describing HLA polymorphism in terms of HLA gene features (GFs) is proposed. This system enumerates the unique nucleotide sequences for each GF in an HLA gene, and records these in a GF enumeration notation that allows both more granular dissection of allele-level HLA polymorphism and the discussion and analysis of GFs in the absence of ARD-encoding exon sequences.

  9. PtrWRKY19, a novel WRKY transcription factor, contributes to the regulation of pith secondary wall formation in Populus trichocarpa.

    PubMed

    Yang, Li; Zhao, Xin; Yang, Fan; Fan, Di; Jiang, Yuanzhong; Luo, Keming

    2016-01-28

    WRKY proteins are one of the largest transcription factor families in higher plants and play diverse roles in various biological processes. Previous studies have shown that some WRKY members act as negative regulators of secondary cell wall formation in pith parenchyma cells. However, the regulatory mechanism of pith secondary wall formation in tree species remains largely unknown. In this study, PtrWRKY19 encoding a homolog of Arabidopsis WRKY12 was isolated from Populus trichocarpa. PtrWRKY19 was expressed in all tissues tested, with highest expression in stems, especially in pith. PtrWRKY19 was located in the nucleus and functioned as a transcriptional repressor. Ectopic expression of PtrWRKY19 in an atwrky12 mutant successfully rescued the phenotype in pith cell walls caused by the defect of AtWRKY12, suggesting that PtrWRKY19 had conserved functions for homologous AtWRKY12. Overexpression of PtrWRKY19 in poplar plants led to a significant increase in the number of pith parenchyma cells. qRT-PCR analysis showed that lignin biosynthesis-related genes were repressed in transgenic plants. In transcient reporter assays, PtrWRKY19 was identified to repress transcription from the PtoC4H2 promoter containing the conserved W-box elements. These results indicated that PtrWRKY19 may function as a negative regulator of pith secondary wall formation in poplar.

  10. Observations Suggesting Allelism of the Achondroplasia and Hypochondroplasia Genes

    PubMed Central

    McKusick, Victor A.; Kelly, Thaddeus E.; Dorst, John P.

    1973-01-01

    It is argued that there are at least two alleles at the achondroplasia locus: one responsible for classic achondroplasia and one responsible for hypochondroplasia. Homozygosity for the achondroplasia gene produces a lethal skeletal dysplasia; homozygosity for hypochondroplasia has not been described. We report here a child considered to be a genetic compound for the achondroplasia and hypochondroplasia alleles. Images PMID:4697848

  11. Induced plant defenses in the natural environment: Nicotiana attenuata WRKY3 and WRKY6 coordinate responses to herbivory.

    PubMed

    Skibbe, Melanie; Qu, Nan; Galis, Ivan; Baldwin, Ian T

    2008-07-01

    A plant-specific family of WRKY transcription factors regulates plant responses to pathogens and abiotic stresses. Here, we identify two insect-responsive WRKY genes in the native tobacco Nicotiana attenuata: WRKY3, whose transcripts accumulate in response to wounding, and WRKY6, whose wound responses are significantly amplified when fatty acid-amino acid conjugates (FACs) in larval oral secretions are introduced into wounds during feeding. WRKY3 is required for WRKY6 elicitation, yet neither is elicited by treatment with the phytohormone wound signal jasmonic acid. Silencing either WRKY3 or WRKY6, or both, by stable transformation makes plants highly vulnerable to herbivores under glasshouse conditions and in their native habitat in the Great Basin Desert, Utah, as shown in three field seasons. This susceptibility is associated with impaired jasmonate (JA) accumulation and impairment of the direct (trypsin proteinase inhibitors) and indirect (volatiles) defenses that JA signaling mediates. The response to wounding and herbivore-specific signals (FACs) demonstrates that these WRKYs help plants to differentiate mechanical wounding from herbivore attack, mediating a plant's herbivore-specific defenses. Differences in responses to single and multiple elicitations indicate an important role of WRKY3 and WRKY6 in potentiating and/or sustaining active JA levels during continuous insect attack.

  12. The WRKY transcription factor family in Brachypodium distachyon

    PubMed Central

    2012-01-01

    Background A complete assembled genome sequence of wheat is not yet available. Therefore, model plant systems for wheat are very valuable. Brachypodium distachyon (Brachypodium) is such a system. The WRKY family of transcription factors is one of the most important families of plant transcriptional regulators with members regulating important agronomic traits. Studies of WRKY transcription factors in Brachypodium and wheat therefore promise to lead to new strategies for wheat improvement. Results We have identified and manually curated the WRKY transcription factor family from Brachypodium using a pipeline designed to identify all potential WRKY genes. 86 WRKY transcription factors were found, a total higher than all other current databases. We therefore propose that our numbering system (BdWRKY1-BdWRKY86) becomes the standard nomenclature. In the JGI v1.0 assembly of Brachypodium with the MIPS/JGI v1.0 annotation, nine of the transcription factors have no gene model and eleven gene models are probably incorrectly predicted. In total, twenty WRKY transcription factors (23.3%) do not appear to have accurate gene models. To facilitate use of our data, we have produced The Database of Brachypodium distachyon WRKY Transcription Factors. Each WRKY transcription factor has a gene page that includes predicted protein domains from MEME analyses. These conserved protein domains reflect possible input and output domains in signaling. The database also contains a BLAST search function where a large dataset of WRKY transcription factors, published genes, and an extensive set of wheat ESTs can be searched. We also produced a phylogram containing the WRKY transcription factor families from Brachypodium, rice, Arabidopsis, soybean, and Physcomitrella patens, together with published WRKY transcription factors from wheat. This phylogenetic tree provides evidence for orthologues, co-orthologues, and paralogues of Brachypodium WRKY transcription factors. Conclusions The description

  13. HvWRKY10, HvWRKY19, and HvWRKY28 positively regulate Mla-triggered immunity and basal defense to barley powdery mildew

    Technology Transfer Automated Retrieval System (TEKTRAN)

    WRKY proteins represent a large family of transcription factors (TFs), involved in plant development and defense responses. So far, fifty-five unique barley TFs have been annotated that contain the WRKY domain; twenty-six of these are present on the Barley1 GeneChip. We analyzed time-course expres...

  14. Networks of WRKY transcription factors in defense signaling.

    PubMed

    Eulgem, Thomas; Somssich, Imre E

    2007-08-01

    Members of the complex family of WRKY transcription factors have been implicated in the regulation of transcriptional reprogramming associated with plant immune responses. Recently genetic evidence directly proving their significance as positive and negative regulators of disease resistance has accumulated. WRKY genes were shown to be functionally connected forming a transcriptional network composed of positive and negative feedback loops and feed-forward modules. Within a web of partially redundant elements some WRKY factors hold central positions mediating fast and efficient activation of defense programs. A key mechanism triggering strong immune responses appears to be based on the inactivation of defense-suppressing WRKY proteins.

  15. How the Number of Alleles Influences Gene Expression

    NASA Astrophysics Data System (ADS)

    Hat, Beata; Paszek, Pawel; Kimmel, Marek; Piechor, Kazimierz; Lipniacki, Tomasz

    2007-07-01

    The higher organisms, eukaryotes, are diploid and most of their genes have two homological copies (alleles). However, the number of alleles in a cell is not constant. In the S phase of the cell cycle all the genome is duplicated and then in the G2 phase and mitosis, which together last for several hours, most of the genes have four copies instead of two. Cancer development is, in many cases, associated with a change in allele number. Several genetic diseases are caused by haploinsufficiency: Lack of one of the alleles or its improper functioning. In the paper we consider the stochastic expression of a gene having a variable number of copies. We applied our previously developed method in which the reaction channels are split into slow (connected with change of gene state) and fast (connected with mRNA/protein synthesis/decay), the later being approximated by deterministic reaction rate equations. As a result we represent gene expression as a piecewise deterministic time-continuous Markov process, which is further related with a system of partial differential hyperbolic equations for probability density functions (pdfs) of protein distribution. The stationary pdfs are calculated analytically for haploidal gene or numerically for diploidal and tetraploidal ones. We distinguished nine classes of simultaneous activation of haploid, diploid and tetraploid genes. This allows for analysis of potential consequences of gene duplication or allele loss. We show that when gene activity is autoregulated by a positive feedback, the change in number of gene alleles may have dramatic consequences for its regulation and may not be compensated by the change of efficiency of mRNA synthesis per allele.

  16. Biased gene conversion skews allele frequencies in human populations, increasing the disease burden of recessive alleles.

    PubMed

    Lachance, Joseph; Tishkoff, Sarah A

    2014-10-02

    Gene conversion results in the nonreciprocal transfer of genetic information between two recombining sequences, and there is evidence that this process is biased toward G and C alleles. However, the strength of GC-biased gene conversion (gBGC) in human populations and its effects on hereditary disease have yet to be assessed on a genomic scale. Using high-coverage whole-genome sequences of African hunter-gatherers, agricultural populations, and primate outgroups, we quantified the effects of GC-biased gene conversion on population genomic data sets. We find that genetic distances (FST and population branch statistics) are modified by gBGC. In addition, the site frequency spectrum is left-shifted when ancestral alleles are favored by gBGC and right-shifted when derived alleles are favored by gBGC. Allele frequency shifts due to gBGC mimic the effects of natural selection. As expected, these effects are strongest in high-recombination regions of the human genome. By comparing the relative rates of fixation of unbiased and biased sites, the strength of gene conversion was estimated to be on the order of Nb ≈ 0.05 to 0.09. We also find that derived alleles favored by gBGC are much more likely to be homozygous than derived alleles at unbiased SNPs (+42.2% to 62.8%). This results in a curse of the converted, whereby gBGC causes substantial increases in hereditary disease risks. Taken together, our findings reveal that GC-biased gene conversion has important population genetic and public health implications.

  17. A WRKY Transcription Factor Recruits the SYG1-Like Protein SHB1 to Activate Gene Expression and Seed Cavity Enlargement

    PubMed Central

    Kang, Xiaojun; Li, Wei; Zhou, Yun; Ni, Min

    2013-01-01

    Seed development in Arabidopsis and in many dicots involves an early proliferation of the endosperm to form a large embryo sac or seed cavity close to the size of the mature seed, followed by a second phase during which the embryo grows and replaces the endosperm. SHORT HYPOCOTYL UNDER BLUE1 (SHB1) is a member of the SYG1 protein family in fungi, Caenorhabditis elegans, flies, and mammals. SHB1 gain-of-function enhances endosperm proliferation, increases seed size, and up-regulates the expression of the WRKY transcription factor gene MINISEED3 (MINI3) and the LRR receptor kinase gene HAIKU2 (IKU2). Mutations in either IKU2 or MINI3 retard endosperm proliferation and reduce seed size. However, the molecular mechanisms underlying the establishment of the seed cavity and hence the seed size remain largely unknown. Here, we show that the expression of MINI3 and IKU2 is repressed before fertilization and after 4 days after pollination (DAP), but is activated by SHB1 from 2 to 4 DAP prior to the formation of the seed cavity. SHB1 associates with their promoters but without a recognizable DNA binding motif, and this association is abolished in mini3 mutant. MINI3 binds to W-boxes in, and recruits SHB1 to, its own and IKU2 promoters. Interestingly, SHB1, but not MINI3, activates transcription of pMINI3::GUS or pIKU2::GUS. We reveal a critical developmental switch through the activation of MINI3 expression by SHB1. The recruitment of SHB1 by MINI3 to its own and IKU2 promoters represents a novel two-step amplification to counter the low expression level of IKU2, which is a trigger for endosperm proliferation and seed cavity enlargement. PMID:23505389

  18. Rice WRKY45 plays a crucial role in benzothiadiazole-inducible blast resistance.

    PubMed

    Shimono, Masaki; Sugano, Shoji; Nakayama, Akira; Jiang, Chang-Jie; Ono, Kazuko; Toki, Seiichi; Takatsuji, Hiroshi

    2007-06-01

    Benzothiadiazole (BTH) is a so-called plant activator and protects plants from diseases by activating the salicylic acid (SA) signaling pathway. By microarray screening, we identified BTH- and SA-inducible WRKY transcription factor (TF) genes that were upregulated within 3 h after BTH treatment. Overexpression of one of them, WRKY45, in rice (Oryza sativa) markedly enhanced resistance to rice blast fungus. RNA interference-mediated knockdown of WRKY45 compromised BTH-inducible resistance to blast disease, indicating that it is essential for BTH-induced defense responses. In a transient expression system, WRKY45 activated reporter gene transcription through W-boxes. Epistasis analysis suggested that WRKY45 acts in the SA signaling pathway independently of NH1, a rice ortholog of Arabidopsis thaliana NPR1, which distinguishes WRKY45 from known Arabidopsis WRKY TFs. Two defense-related genes, encoding a glutathione S-transferase and a cytochrome P450, were found to be regulated downstream of WRKY45 but were not regulated by NH1, consistent with the apparent independence of the WRKY45- and NH1-dependent pathways. Defense gene expression in WRKY45-overexpressed rice plants varied with growth conditions, suggesting that some environmental factor(s) acts downstream of WRKY45 transcription. We propose a role for WRKY45 in BTH-induced and SA-mediated defense signaling in rice and its potential utility in improving disease resistance of rice, an importance food resource worldwide.

  19. The WRKY transcription factor family and senescence in switchgrass

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Background: Early aerial senescence in switchgrass (Panicum virgatum) can significantly limit biomass yields. WRKY transcription factors that can regulate senescence could be used to reprogram senescence and enhance biomass yields. Methods: All potential WRKY genes present in the version 1.0 of the...

  20. Analyses of Allele-Specific Gene Expression in Highly Divergent Mouse Crosses Identifies Pervasive Allelic Imbalance

    PubMed Central

    Crowley, James J; Zhabotynsky, Vasyl; Sun, Wei; Huang, Shunping; Pakatci, Isa Kemal; Kim, Yunjung; Wang, Jeremy R; Morgan, Andrew P; Calaway, John D; Aylor, David L; Yun, Zaining; Bell, Timothy A; Buus, Ryan J; Calaway, Mark E; Didion, John P; Gooch, Terry J; Hansen, Stephanie D; Robinson, Nashiya N; Shaw, Ginger D; Spence, Jason S; Quackenbush, Corey R; Barrick, Cordelia J; Nonneman, Randal J.; Kim, Kyungsu; Xenakis, James; Xie, Yuying; Valdar, William; Lenarcic, Alan B; Wang, Wei; Welsh, Catherine E; Fu, Chen-Ping; Zhang, Zhaojun; Holt, James; Guo, Zhishan; Threadgill, David W; Tarantino, Lisa M; Miller, Darla R; Zou, Fei; McMillan, Leonard; Sullivan, Patrick F; de Villena, Fernando Pardo-Manuel

    2015-01-01

    Complex human traits are influenced by variation in regulatory DNA through mechanisms that are not fully understood. Since regulatory elements are conserved between humans and mice, a thorough annotation of cis regulatory variants in mice could aid in this process. Here we provide a detailed portrait of mouse gene expression across multiple tissues in a three-way diallel. Greater than 80% of mouse genes have cis regulatory variation. These effects influence complex traits and usually extend to the human ortholog. Further, we estimate that at least one in every thousand SNPs creates a cis regulatory effect. We also observe two types of parent-of-origin effects, including classical imprinting and a novel, global allelic imbalance in favor of the paternal allele. We conclude that, as with humans, pervasive regulatory variation influences complex genetic traits in mice and provide a new resource toward understanding the genetic control of transcription in mammals. PMID:25730764

  1. WRKY22 Transcription Factor Mediates Dark-Induced Leaf Senescence in Arabidopsis

    PubMed Central

    Zhou, Xiang; Jiang, Zhou; Yu, Diqiu

    2011-01-01

    Arabidopsis WRKY proteins are plant-specific transcrip-tion factors, encoded by a large gene family, which contain the highly conserved amino acid sequence WRKYGQK and the zinc-finger-like motifs, Cys2His2 or Cys2HisCys. They can recognize and bind the TTGAC(C/T) W-box cis-elements found in the promoters of target genes, and are involved in the regulation of gene expression during pathogen defense, wounding, trichome development, and senescence. Here we investigated the physiological function of the Arabidopsis WRKY22 transcription factor during dark-induced senescence. WRKY22 transcription was suppressed by light and promoted by darkness. In addi-tion, AtWRKY22 expression was markedly induced by H2O2. These results indicated that AtWRKY22 was involved in signal pathways in response to abiotic stress. Dark-treated AtWRKY22 over-expression and knockout lines showed accelerated and delayed senescence phenotypes, respectively, and senescence-associated genes exhibited increased and decreased expression levels. Mutual regulation existed between AtWRKY22 and AtWRKY6, AtWR-KY53, and AtWRKY70, respectively. Moreover, AtWRKY22 could influence their relative expression levels by feedback regulation or by other, as yet unknown mechanisms in response to dark. These results prove that AtWRKY22 participates in the dark-induced senescence signal transduction pathway. PMID:21359674

  2. Novel Genomic and Evolutionary Insight of WRKY Transcription Factors in Plant Lineage

    PubMed Central

    Mohanta, Tapan Kumar; Park, Yong-Hwan; Bae, Hanhong

    2016-01-01

    The evolutionarily conserved WRKY transcription factor (TF) regulates different aspects of gene expression in plants, and modulates growth, development, as well as biotic and abiotic stress responses. Therefore, understanding the details regarding WRKY TFs is very important. In this study, large-scale genomic analyses of the WRKY TF gene family from 43 plant species were conducted. The results of our study revealed that WRKY TFs could be grouped and specifically classified as those belonging to the monocot or dicot plant lineage. In this study, we identified several novel WRKY TFs. To our knowledge, this is the first report on a revised grouping system of the WRKY TF gene family in plants. The different forms of novel chimeric forms of WRKY TFs in the plant genome might play a crucial role in their evolution. Tissue-specific gene expression analyses in Glycine max and Phaseolus vulgaris showed that WRKY11-1, WRKY11-2 and WRKY11-3 were ubiquitously expressed in all tissue types, and WRKY15-2 was highly expressed in the stem, root, nodule and pod tissues in G. max and P. vulgaris. PMID:27853303

  3. Tyrosine phosphorylation and protein degradation control the transcriptional activity of WRKY involved in benzylisoquinoline alkaloid biosynthesis

    PubMed Central

    Yamada, Yasuyuki; Sato, Fumihiko

    2016-01-01

    Benzylisoquinoline alkaloids (BIQ) are among the most structurally diverse and pharmaceutically valuable secondary metabolites. A plant-specific WRKY-type transcription factor, CjWRKY1, was isolated from Coptis japonica and identified as a transcriptional activator of BIQ biosynthesis. However, the expression of CjWRKY1 gene alone was not sufficient for the activation of genes encoding biosynthetic enzymes. Here, we report the importance of post-translational regulation of CjWRKY1 in BIQ biosynthesis. First, we detected the differential accumulation of CjWRKY1 protein in two cell lines with similar CjWRKY1 gene expression but different levels of accumulated alkaloids. Further investigation of the WRKY protein identified the phosphorylation of the WRKYGQK core domain at Y115. The CjWRKYY115E phosphorylation-mimic mutant showed loss of nuclear localization, DNA-binding activity, and transactivation activity compared to wild-type CjWRKY1. Rapid degradation of the CjWRKY1 protein was also confirmed following treatment with inhibitors of the 26S proteasome and protease inhibitors. The existence of two independent degradation pathways as well as protein phosphorylation suggests the fine-tuning of CjWRKY1 activities is involved in the regulation of biosynthesis of BIQs. PMID:27552928

  4. Arabidopsis WRKY2 transcription factor mediates seed germination and postgermination arrest of development by abscisic acid

    PubMed Central

    Jiang, Wenbo; Yu, Diqiu

    2009-01-01

    Background Plant WRKY DNA-binding transcription factors are key regulators in certain developmental programs. A number of studies have suggested that WRKY genes may mediate seed germination and postgermination growth. However, it is unclear whether WRKY genes mediate ABA-dependent seed germination and postgermination growth arrest. Results To determine directly the role of Arabidopsis WRKY2 transcription factor during ABA-dependent seed germination and postgermination growth arrest, we isolated T-DNA insertion mutants. Two independent T-DNA insertion mutants for WRKY2 were hypersensitive to ABA responses only during seed germination and postgermination early growth. wrky2 mutants displayed delayed or decreased expression of ABI5 and ABI3, but increased or prolonged expression of Em1 and Em6. wrky2 mutants and wild type showed similar levels of expression for miR159 and its target genes MYB33 and MYB101. Analysis of WRKY2 expression level in ABA-insensitive and ABA-deficient mutants abi5-1, abi3-1, aba2-3 and aba3-1 further indicated that ABA-induced WRKY2 accumulation during germination and postgermination early growth requires ABI5, ABI3, ABA2 and ABA3. Conclusion ABA hypersensitivity of the wrky2 mutants during seed germination and postgermination early seedling establishment is attributable to elevated mRNA levels of ABI5, ABI3 and ABI5-induced Em1 and Em6 in the mutants. WRKY2-mediated ABA responses are independent of miR159 and its target genes MYB33 and MYB101. ABI5, ABI3, ABA2 and ABA3 are important regulators of the transcripts of WRKY2 by ABA treatment. Our results suggest that WRKY2 transcription factor mediates seed germination and postgermination developmental arrest by ABA. PMID:19622176

  5. A WRKY Transcription Factor Regulates Fe Translocation under Fe Deficiency1[OPEN

    PubMed Central

    Yan, Jing Ying; Li, Chun Xiao; Sun, Li; Ren, Jiang Yuan; Li, Gui Xin

    2016-01-01

    Iron (Fe) deficiency affects plant growth and development, leading to reduction of crop yields and quality. Although the regulation of Fe uptake under Fe deficiency has been well studied in the past decade, the regulatory mechanism of Fe translocation inside the plants remains unknown. Here, we show that a WRKY transcription factor WRKY46 is involved in response to Fe deficiency. Lack of WRKY46 (wrky46-1 and wrky46-2 loss-of-function mutants) significantly affects Fe translocation from root to shoot and thus causes obvious chlorosis on the new leaves under Fe deficiency. Gene expression analysis reveals that expression of a nodulin-like gene (VACUOLAR IRON TRANSPORTER1-LIKE1 [VITL1]) is dramatically increased in wrky46-1 mutant. VITL1 expression is inhibited by Fe deficiency, while the expression of WRKY46 is induced in the root stele. Moreover, down-regulation of VITL1 expression can restore the chlorosis phenotype on wrky46-1 under Fe deficiency. Further yeast one-hybrid and chromatin immunoprecipitation experiments indicate that WRKY46 is capable of binding to the specific W-boxes present in the VITL1 promoter. In summary, our results demonstrate that WRKY46 plays an important role in the control of root-to-shoot Fe translocation under Fe deficiency condition via direct regulation of VITL1 transcript levels. PMID:27208259

  6. GhWRKY15, a member of the WRKY transcription factor family identified from cotton (Gossypium hirsutum L.), is involved in disease resistance and plant development

    PubMed Central

    2012-01-01

    Background As a large family of regulatory proteins, WRKY transcription factors play essential roles in the processes of adaptation to diverse environmental stresses and plant growth and development. Although several studies have investigated the role of WRKY transcription factors during these processes, the mechanisms underlying the function of WRKY members need to be further explored, and research focusing on the WRKY family in cotton crops is extremely limited. Results In the present study, a gene encoding a putative WRKY family member, GhWRKY15, was isolated from cotton. GhWRKY15 is present as a single copy gene, and a transient expression analysis indicated that GhWRKY15 was localised to the nucleus. Additionally, a group of cis-acting elements associated with the response to environmental stress and plant growth and development were detected in the promoter. Consistently, northern blot analysis showed that GhWRKY15 expression was significantly induced in cotton seedlings following fungal infection or treatment with salicylic acid, methyl jasmonate or methyl viologen. Furthermore, GhWRKY15-overexpressing tobacco exhibited more resistance to viral and fungal infections compared with wild-type tobacco. The GhWRKY15-overexpressing tobacco also exhibited increased RNA expression of several pathogen-related genes, NONEXPRESSOR OF PR1, and two genes that encode enzymes involved in ET biosynthesis. Importantly, increased activity of the antioxidant enzymes POD and APX during infection and enhanced expression of NtAPX1 and NtGPX in transgenic tobacco following methyl viologen treatment were observed. Moreover, GhWRKY15 transcription was greater in the roots and stems compared with the expression in the cotyledon of cotton, and the stems of transgenic plants displayed faster elongation at the earlier shooting stages compared with wide type tobacco. Additionally, exposure to abiotic stresses, including cold, wounding and drought, resulted in the accumulation of GhWRKY15

  7. CaWRKY6 transcriptionally activates CaWRKY40, regulates Ralstonia solanacearum resistance, and confers high-temperature and high-humidity tolerance in pepper.

    PubMed

    Cai, Hanyang; Yang, Sheng; Yan, Yan; Xiao, Zhuoli; Cheng, Junbin; Wu, Ji; Qiu, Ailian; Lai, Yan; Mou, Shaoliang; Guan, Deyi; Huang, Ronghua; He, Shuilin

    2015-06-01

    High temperature (HT), high humidity (HH), and pathogen infection often co-occur and negatively affect plant growth. However, these stress factors and plant responses are generally studied in isolation. The mechanisms of synergistic responses to combined stresses are poorly understood. We isolated the subgroup IIb WRKY family member CaWRKY6 from Capsicum annuum and performed quantitative real-time PCR analysis. CaWRKY6 expression was upregulated by individual or simultaneous treatment with HT, HH, combined HT and HH (HTHH), and Ralstonia solanacearum inoculation, and responded to exogenous application of jasmonic acid (JA), ethephon, and abscisic acid (ABA). Virus-induced gene silencing of CaWRKY6 enhanced pepper plant susceptibility to R. solanacearum and HTHH, and downregulated the hypersensitive response (HR), JA-, ethylene (ET)-, and ABA-induced marker gene expression, and thermotolerance-associated expression of CaHSP24, ER-small CaSHP, and Chl-small CaHSP. CaWRKY6 overexpression in pepper attenuated the HTHH-induced suppression of resistance to R. solanacearum infection. CaWRKY6 bound to and activated the CaWRKY40 promoter in planta, which is a pepper WRKY that regulates heat-stress tolerance and R. solanacearum resistance. CaWRKY40 silencing significantly blocked HR-induced cell death and reduced transcriptional expression of CaWRKY40. These data suggest that CaWRKY6 is a positive regulator of R. solanacearum resistance and heat-stress tolerance, which occurs in part by activating CaWRKY40.

  8. Gene-based rare allele analysis identified a risk gene of Alzheimer's disease.

    PubMed

    Kim, Jong Hun; Song, Pamela; Lim, Hyunsun; Lee, Jae-Hyung; Lee, Jun Hong; Park, Sun Ah

    2014-01-01

    Alzheimer's disease (AD) has a strong propensity to run in families. However, the known risk genes excluding APOE are not clinically useful. In various complex diseases, gene studies have targeted rare alleles for unsolved heritability. Our study aims to elucidate previously unknown risk genes for AD by targeting rare alleles. We used data from five publicly available genetic studies from the Alzheimer's Disease Neuroimaging Initiative (ADNI) and the database of Genotypes and Phenotypes (dbGaP). A total of 4,171 cases and 9,358 controls were included. The genotype information of rare alleles was imputed using 1,000 genomes. We performed gene-based analysis of rare alleles (minor allele frequency≤3%). The genome-wide significance level was defined as meta P<1.8×10(-6) (0.05/number of genes in human genome = 0.05/28,517). ZNF628, which is located at chromosome 19q13.42, showed a genome-wide significant association with AD. The association of ZNF628 with AD was not dependent on APOE ε4. APOE and TREM2 were also significantly associated with AD, although not at genome-wide significance levels. Other genes identified by targeting common alleles could not be replicated in our gene-based rare allele analysis. We identified that rare variants in ZNF628 are associated with AD. The protein encoded by ZNF628 is known as a transcription factor. Furthermore, the associations of APOE and TREM2 with AD were highly significant, even in gene-based rare allele analysis, which implies that further deep sequencing of these genes is required in AD heritability studies.

  9. Rice WRKY45 plays important roles in fungal and bacterial disease resistance.

    PubMed

    Shimono, Masaki; Koga, Hironori; Akagi, Aya; Hayashi, Nagao; Goto, Shingo; Sawada, Miyuki; Kurihara, Takayuki; Matsushita, Akane; Sugano, Shoji; Jiang, Chang-Jie; Kaku, Hisatoshi; Inoue, Haruhiko; Takatsuji, Hiroshi

    2012-01-01

    Plant 'activators', such as benzothiadiazole (BTH), protect plants from various diseases by priming the plant salicylic acid (SA) signalling pathway. We have reported previously that a transcription factor identified in rice, WRKY45 (OsWRKY45), plays a pivotal role in BTH-induced disease resistance by mediating SA signalling. Here, we report further functional characterization of WRKY45. Different plant activators vary in their action points, either downstream (BTH and tiadinil) or upstream (probenazole) of SA. Rice resistance to Magnaporthe grisea, induced by both types of plant activator, was markedly reduced in WRKY45-knockdown (WRKY45-kd) rice, indicating a universal role for WRKY45 in chemical-induced resistance. Fungal invasion into rice cells was blocked at most attempted invasion sites (pre-invasive defence) in WRKY45-overexpressing (WRKY45-ox) rice. Hydrogen peroxide accumulated within the cell wall underneath invading fungus appressoria or between the cell wall and the cytoplasm, implying a possible role for H(2)O(2) in pre-invasive defence. Moreover, a hypersensitive reaction-like reaction was observed in rice cells, in which fungal growth was inhibited after invasion (post-invasive defence). The two levels of defence mechanism appear to correspond to Type I and II nonhost resistances. The leaf blast resistance of WRKY45-ox rice plants was much higher than that of other known blast-resistant varieties. WRKY45-ox plants also showed strong panicle blast resistance. BTH-induced resistance to Xanthomonas oryzae pv. oryzae was compromised in WRKY45-kd rice, whereas WRKY45-ox plants were highly resistant to this pathogen. However, WRKY45-ox plants were susceptible to Rhizoctonia solani. These results indicate the versatility and limitations of the application of this gene.

  10. GhWRKY68 Reduces Resistance to Salt and Drought in Transgenic Nicotiana benthamiana

    PubMed Central

    Jia, Haihong; Wang, Chen; Wang, Fang; Liu, Shuchang; Li, Guilin; Guo, Xingqi

    2015-01-01

    The WRKY transcription factors modulate numerous physiological processes, including plant growth, development and responses to various environmental stresses. Currently, our understanding of the functions of the majority of the WRKY family members and their possible roles in signalling crosstalk is limited. In particular, very few WRKYs have been identified and characterised from an economically important crop, cotton. In this study, we characterised a novel group IIc WRKY gene, GhWRKY68, which is induced by different abiotic stresses and multiple defence-related signalling molecules. The β-glucuronidase activity driven by the GhWRKY68 promoter was enhanced after exposure to drought, salt, abscisic acid (ABA) and H2O2. The overexpression of GhWRKY68 in Nicotiana benthamiana reduced resistance to drought and salt and affected several physiological indices. GhWRKY68 may mediate salt and drought responses by modulating ABA content and enhancing the transcript levels of ABA-responsive genes. GhWRKY68-overexpressing plants exhibited reduced tolerance to oxidative stress after drought and salt stress treatments, which correlated with the accumulation of reactive oxygen species (ROS), reduced enzyme activities, elevated malondialdehyde (MDA) content and altered ROS-related gene expression. These results indicate that GhWRKY68 is a transcription factor that responds to drought and salt stresses by regulating ABA signalling and modulating cellular ROS. PMID:25793865

  11. An allele of the crm gene blocks cyanobacterial circadian rhythms.

    PubMed

    Boyd, Joseph S; Bordowitz, Juliana R; Bree, Anna C; Golden, Susan S

    2013-08-20

    The SasA-RpaA two-component system constitutes a key output pathway of the cyanobacterial Kai circadian oscillator. To date, rhythm of phycobilisome associated (rpaA) is the only gene other than kaiA, kaiB, and kaiC, which encode the oscillator itself, whose mutation causes completely arrhythmic gene expression. Here we report a unique transposon insertion allele in a small ORF located immediately upstream of rpaA in Synechococcus elongatus PCC 7942 termed crm (for circadian rhythmicity modulator), which results in arrhythmic promoter activity but does not affect steady-state levels of RpaA. The crm ORF complements the defect when expressed in trans, but only if it can be translated, suggesting that crm encodes a small protein. The crm1 insertion allele phenotypes are distinct from those of an rpaA null; crm1 mutants are able to grow in a light:dark cycle and have no detectable oscillations of KaiC phosphorylation, whereas low-amplitude KaiC phosphorylation rhythms persist in the absence of RpaA. Levels of phosphorylated RpaA in vivo measured over time are significantly altered compared with WT in the crm1 mutant as well as in the absence of KaiC. Taken together, these results are consistent with the hypothesis that the Crm polypeptide modulates a circadian-specific activity of RpaA.

  12. Allele Mining Strategies: Principles and Utilisation for Blast Resistance Genes in Rice (Oryza sativa L.).

    PubMed

    Ashkani, Sadegh; Yusop, Mohd Rafii; Shabanimofrad, Mahmoodreza; Azady, Amin; Ghasemzadeh, Ali; Azizi, Parisa; Latif, Mohammad Abdul

    2015-01-01

    Allele mining is a promising way to dissect naturally occurring allelic variants of candidate genes with essential agronomic qualities. With the identification, isolation and characterisation of blast resistance genes in rice, it is now possible to dissect the actual allelic variants of these genes within an array of rice cultivars via allele mining. Multiple alleles from the complex locus serve as a reservoir of variation to generate functional genes. The routine sequence exchange is one of the main mechanisms of R gene evolution and development. Allele mining for resistance genes can be an important method to identify additional resistance alleles and new haplotypes along with the development of allele-specific markers for use in marker-assisted selection. Allele mining can be visualised as a vital link between effective utilisation of genetic and genomic resources in genomics-driven modern plant breeding. This review studies the actual concepts and potential of mining approaches for the discovery of alleles and their utilisation for blast resistance genes in rice. The details provided here will be important to provide the rice breeder with a worthwhile introduction to allele mining and its methodology for breakthrough discovery of fresh alleles hidden in hereditary diversity, which is vital for crop improvement.

  13. The interactome of soybean GmWRKY53 using yeast 2-hybrid library screening to saturation.

    PubMed

    Tripathi, Prateek; Rabara, Roel C; Choudhary, Mani Kant; Miller, Marissa A; Huang, Ying-Sheng; Shen, Qingxi J; Blachon, Stéphanie; Rushton, Paul J

    2015-01-01

    Soybean GmWRKY53 functions in both biotic and abiotic stress signaling. Using GmWRKY53 as a bait yeast 2-hybrid library screening to saturation isolated multiple independent fragments for many interacting proteins, enabling delineation of minimal interacting domains and computation of a confidence score. Multiple independent clones coding for the LATE ELONGATED HYPOCOTYL clock protein GmLCL2 (MYB114) were isolated and the binding site for GmWRKY53 was mapped to 90 amino acids separate from the MYB domain. This suggests a direct input from the clock on GmWRKY53 activity. The GmWRKY53-interacting proteins also included 3 water stress-inducible AP2/ERF transcription factors. One of these (Glyma03g26310) is one of the most strongly water stress induced genes in soybean roots, suggesting that GmWRKY53/ERF complexes regulate water stress responses.

  14. Overexpression of CaWRKY27, a subgroup IIe WRKY transcription factor of Capsicum annuum, positively regulates tobacco resistance to Ralstonia solanacearum infection.

    PubMed

    Dang, Fengfeng; Wang, Yuna; She, Jianju; Lei, Yufen; Liu, Zhiqin; Eulgem, Thomas; Lai, Yan; Lin, Jing; Yu, Lu; Lei, Dan; Guan, Deyi; Li, Xia; Yuan, Qian; He, Shuilin

    2014-03-01

    WRKY proteins are encoded by a large gene family and are linked to many biological processes across a range of plant species. The functions and underlying mechanisms of WRKY proteins have been investigated primarily in model plants such as Arabidopsis and rice. The roles of these transcription factors in non-model plants, including pepper and other Solanaceae, are poorly understood. Here, we characterize the expression and function of a subgroup IIe WRKY protein from pepper (Capsicum annuum), denoted as CaWRKY27. The protein localized to nuclei and activated the transcription of a reporter GUS gene construct driven by the 35S promoter that contained two copies of the W-box in its proximal upstream region. Inoculation of pepper cultivars with Ralstonia solanacearum induced the expression of CaWRKY27 transcript in 76a, a bacterial wilt-resistant pepper cultivar, whereas it downregulated the expression of CaWRKY27 transcript in Gui-1-3, a bacterial wilt-susceptible pepper cultivar. CaWRKY27 transcript levels were also increased by treatments with salicylic acid (SA), methyl jasmonate (MeJA) and ethephon (ETH). Transgenic tobacco plants overexpressing CaWRKY27 exhibited resistance to R. solanacearum infection compared to that of wild-type plants. This resistance was coupled with increased transcript levels in a number of marker genes, including hypersensitive response genes, and SA-, JA- and ET-associated genes. By contrast, virus-induced gene silencing (VIGS) of CaWRKY27 increased the susceptibility of pepper plants to R. solanacearum infection. These results suggest that CaWRKY27 acts as a positive regulator in tobacco resistance responses to R. solanacearum infection through modulation of SA-, JA- and ET-mediated signaling pathways.

  15. Genomic identification of WRKY transcription factors in carrot (Daucus carota) and analysis of evolution and homologous groups for plants

    PubMed Central

    Li, Meng-Yao; Xu, Zhi-Sheng; Tian, Chang; Huang, Ying; Wang, Feng; Xiong, Ai-Sheng

    2016-01-01

    WRKY transcription factors belong to one of the largest transcription factor families. These factors possess functions in plant growth and development, signal transduction, and stress response. Here, we identified 95 DcWRKY genes in carrot based on the carrot genomic and transcriptomic data, and divided them into three groups. Phylogenetic analysis of WRKY proteins from carrot and Arabidopsis divided these proteins into seven subgroups. To elucidate the evolution and distribution of WRKY transcription factors in different species, we constructed a schematic of the phylogenetic tree and compared the WRKY family factors among 22 species, which including plants, slime mold and protozoan. An in-depth study was performed to clarify the homologous factor groups of nine divergent taxa in lower and higher plants. Based on the orthologous factors between carrot and Arabidopsis, 38 DcWRKY proteins were calculated to interact with other proteins in the carrot genome. Yeast two-hybrid assay showed that DcWRKY20 can interact with DcMAPK1 and DcMAPK4. The expression patterns of the selected DcWRKY genes based on transcriptome data and qRT-PCR suggested that those selected DcWRKY genes are involved in root development, biotic and abiotic stress response. This comprehensive analysis provides a basis for investigating the evolution and function of WRKY genes. PMID:26975939

  16. SlWRKY70 is required for Mi-1-mediated resistance to aphids and nematodes in tomato.

    PubMed

    Atamian, Hagop S; Eulgem, Thomas; Kaloshian, Isgouhi

    2012-02-01

    Plant resistance (R) gene-mediated defense responses against biotic stresses include vast transcriptional reprogramming. In several plant-pathogen systems, members of the WRKY family of transcription factors have been demonstrated to act as both positive and negative regulators of plant defense transcriptional networks. To identify the possible roles of tomato (Solanum lycopersicum) WRKY transcription factors in defense mediated by the R gene Mi-1 against potato aphid, Macrosiphum euphorbiae, and root-knot nematode (RKN), Meloidogyne javanica, we used tobacco rattle virus (TRV)-based virus-induced gene silencing and transcriptionally suppressed SlWRKY70, a tomato ortholog of the Arabidopsis thaliana WRKY70 gene. Silencing SlWRKY70 attenuated Mi-1-mediated resistance against both potato aphid and RKN showing that SlWRKY70 is required for Mi-1 function. Furthermore, we found SlWRKY70 transcripts to be inducible in response to aphid infestation and RKN inoculation. Mi-1-mediated recognition of these pests modulates this transcriptional response. As previously described for AtWRKY70, we found SlWRKY70 transcript levels to be up-regulated by salicylic acid and suppressed by methyl jasmonate. This indicates that some aspects of WRKY70 regulation are conserved among distantly related eudicots.

  17. Transcriptome-wide identification of Camellia sinensis WRKY transcription factors in response to temperature stress.

    PubMed

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

    2016-02-01

    Tea plant [Camellia sinensis (L.) O. Kuntze] is a leaf-type healthy non-alcoholic beverage crop, which has been widely introduced worldwide. Tea is rich in various secondary metabolites, which are important for human health. However, varied climate and complex geography have posed challenges for tea plant survival. The WRKY gene family in plants is a large transcription factor family that is involved in biological processes related to stress defenses, development, and metabolite synthesis. Therefore, identification and analysis of WRKY family transcription factors in tea plant have a profound significance. In the present study, 50 putative C. sinensis WRKY proteins (CsWRKYs) with complete WRKY domain were identified and divided into three Groups (Group I-III) on the basis of phylogenetic analysis results. The distribution of WRKY family transcription factors among plantae, fungi, and protozoa showed that the number of WRKY genes increased in higher plant, whereas the number of these genes did not correspond to the evolutionary relationships of different species. Structural feature and annotation analysis results showed that CsWRKY proteins contained WRKYGQK/WRKYGKK domains and C2H2/C2HC-type zinc-finger structure: D-X18-R-X1-Y-X2-C-X4-7-C-X23-H motif; CsWRKY proteins may be associated with the biological processes of abiotic and biotic stresses, tissue development, and hormone and secondary metabolite biosynthesis. Temperature stresses suggested that the candidate CsWRKY genes were involved in responses to extreme temperatures. The current study established an extensive overview of the WRKY family transcription factors in tea plant. This study also provided a global survey of CsWRKY transcription factors and a foundation of future functional identification and molecular breeding.

  18. Populus euphratica HSF binds the promoter of WRKY1 to enhance salt tolerance.

    PubMed

    Shen, Zedan; Yao, Jun; Sun, Jian; Chang, Liwei; Wang, Shaojie; Ding, Mingquan; Qian, Zeyong; Zhang, Huilong; Zhao, Nan; Sa, Gang; Hou, Peichen; Lang, Tao; Wang, Feifei; Zhao, Rui; Shen, Xin; Chen, Shaoliang

    2015-06-01

    Poplar species increase expressions of transcription factors to deal with salt environments. We assessed the salt-induced transcriptional responses of heat-shock transcription factor (HSF) and WRKY1 in Populus euphratica, and their roles in salt tolerance. High NaCl (200mM) induced PeHSF and PeWRKY1 expressions in P. euphratica, with a rapid rise in roots than in leaves. Moreover, the salt-elicited PeHSF reached its peak level 6h earlier than PeWRKY1 in leaves. PeWRKY1 was down-regulated in salinized P. euphratica when PeHSF was silenced by tobacco rattle virus-based gene silencing. Subcellular assays in onion epidermal cells and Arabidopsis protoplasts revealed that PeHSF and PeWRKY1 were restricted to the nucleus. Transgenic tobacco plants overexpressing PeWRKY1 showed improved salt tolerance in terms of survival rate, root growth, photosynthesis, and ion fluxes. We further isolated an 1182-bp promoter fragment upstream of the translational start of PeWRKY1 from P. euphratica. Promoter sequence analysis revealed that PeWRKY1 harbours four tandem repeats of heat shock element (HSE) in the upstream regulatory region. Yeast one-hybrid assay showed that PeHSF directly binds the cis-acting HSE. To determine whether the HSE cluster was important for salt-induced PeWRKY1 expression, the promoter-reporter construct PeWRKY1-pro::GUS was transferred to tobacco plants. β-glucuronidase activities increased in root, leaf, and stem tissues under salt stress. Therefore, we conclude that salinity increased PeHSF transcription in P. euphratica, and that PeHSF binds the cis-acting HSE of the PeWRKY1 promoter, thus activating PeWRKY1 expression.

  19. Nuclear ubiquitin proteasome degradation affects WRKY45 function in the rice defense program.

    PubMed

    Matsushita, Akane; Inoue, Haruhiko; Goto, Shingo; Nakayama, Akira; Sugano, Shoji; Hayashi, Nagao; Takatsuji, Hiroshi

    2013-01-01

    The transcriptional activator WRKY45 plays a major role in the salicylic acid/benzothiadiazole-induced defense program in rice. Here, we show that the nuclear ubiquitin-proteasome system (UPS) plays a role in regulating the function of WRKY45. Proteasome inhibitors induced accumulation of polyubiquitinated WRKY45 and transient up-regulation of WRKY45 target genes in rice cells, suggesting that WRKY45 is constantly degraded by the UPS to suppress defense responses in the absence of defense signals. Mutational analysis of the nuclear localization signal indicated that UPS-dependent WRKY45 degradation occurs in the nuclei. Interestingly, the transcriptional activity of WRKY45 after salicylic acid treatment was impaired by proteasome inhibition. The same C-terminal region in WRKY45 was essential for both transcriptional activity and UPS-dependent degradation. These results suggest that UPS regulation also plays a role in the transcriptional activity of WRKY45. It has been reported that AtNPR1, the central regulator of the salicylic acid pathway in Arabidopsis, is regulated by the UPS. We found that OsNPR1/NH1, the rice counterpart of NPR1, was not stabilized by proteasome inhibition under uninfected conditions. We discuss the differences in post-translational regulation of salicylic acid pathway components between rice and Arabidopsis.

  20. The WRKY transcription factors in the diploid woodland strawberry Fragaria vesca: Identification and expression analysis under biotic and abiotic stresses.

    PubMed

    Wei, Wei; Hu, Yang; Han, Yong-Tao; Zhang, Kai; Zhao, Feng-Li; Feng, Jia-Yue

    2016-08-01

    WRKY proteins comprise a large family of transcription factors that play important roles in response to biotic and abiotic stresses and in plant growth and development. To date, little is known about the WRKY gene family in strawberry. In this study, we identified 62 WRKY genes (FvWRKYs) in the wild diploid woodland strawberry (Fragaria vesca, 2n = 2x = 14) accession Heilongjiang-3. According to the phylogenetic analysis and structural features, these identified strawberry FvWRKY genes were classified into three main groups. In addition, eight FvWRKY-GFP fusion proteins showed distinct subcellular localizations in Arabidopsis mesophyll protoplasts. Furthermore, we examined the expression of the 62 FvWRKY genes in 'Heilongjiang-3' under various conditions, including biotic stress (Podosphaera aphanis), abiotic stresses (drought, salt, cold, and heat), and hormone treatments (abscisic acid, ethephon, methyl jasmonate, and salicylic acid). The expression levels of 33 FvWRKY genes were upregulated, while 12 FvWRKY genes were downregulated during powdery mildew infection. FvWRKY genes responded to drought and salt treatment to a greater extent than to temperature stress. Expression profiles derived from quantitative real-time PCR suggested that 11 FvWRKY genes responded dramatically to various stimuli at the transcriptional level, indicating versatile roles in responses to biotic and abiotic stresses. Interaction networks revealed that the crucial pathways controlled by WRKY proteins may be involved in the differential response to biotic stress. Taken together, the present work may provide the basis for future studies of the genetic modification of WRKY genes for pathogen resistance and stress tolerance in strawberry.

  1. Extensive allelic variation in gene expression in populus F1 hybrids.

    PubMed

    Zhuang, Yan; Adams, Keith L

    2007-12-01

    Hybridization between plant species can induce speciation as well as phenotypic novelty and heterosis. Hybrids also can show genome rearrangements and gene expression changes compared with their parents. Here we determined the allelic variation in gene expression in Populus trichocarpa x Populus deltoides F(1) hybrids. Among 30 genes analyzed in four independently formed hybrids, 17 showed >1.5-fold expression biases for one of the two alleles, and there was monoallelic expression of one gene. Expression ratios of the alleles differed between leaves and stems for 10 genes. The results suggest differential regulation of the two parental alleles in the hybrids. To determine if the allelic expression biases were caused by hybridization we compared the ratios of species-specific transcripts between an F(1) hybrid and its parents. Thirteen of 19 genes showed allelic expression ratios in the hybrid that were significantly different from the ratios of the parental species. The P. deltoides allele of one gene was silenced in the hybrid. Modes of gene regulation were inferred from the hybrid-parent comparisons. Cis-regulation was inferred for 6 genes, trans-regulation for 1 gene, and combined cis- and trans-regulation for 9 genes. The results from this study indicate that hybridization between plant species can have extensive effects on allelic expression patterns, some of which might lead to phenotypic changes.

  2. Transcription factors WRKY70 and WRKY11 served as regulators in rhizobacterium Bacillus cereus AR156-induced systemic resistance to Pseudomonas syringae pv. tomato DC3000 in Arabidopsis.

    PubMed

    Jiang, Chun-Hao; Huang, Zi-Yang; Xie, Ping; Gu, Chun; Li, Ke; Wang, Da-Chen; Yu, Yi-Yang; Fan, Zhi-Hang; Wang, Chun-Juan; Wang, Yun-Peng; Guo, Ya-Hui; Guo, Jian-Hua

    2016-01-01

    The activation of both the SA and JA/ETsignalling pathways may lead to more efficient general and broad resistance to Pst DC3000 by non-pathogenic rhizobacteria. However, the mechanisms that govern this simultaneous activation are unclear. Using Arabidopsis as a model system, two transcription factors, WRKY11 and WRKY70, were identified as important regulators involved in Induced Systemic Resistance (ISR) triggered by Bacillus cereus AR156. The results revealed that AR156 treatment significantly stimulated the transcription of WRKY70, but suppressed that of WRKY11 in Arabidopsis leaves. Furthermore, they were shown to be required for AR156 enhancing the activation of cellular defence responses and the transcription level of the plant defence response gene. Overexpression of the two transcription factors in Arabidopsis also showed that they were essential for AR156 to elicit ISR. AR156-triggered ISR was completely abolished in the double mutant of the two transcription factors, but still partially retained in the single mutants, indicating that the regulation of the two transcription factors depend on two different pathways. The target genes of the two transcription factors and epistasis analysis suggested that WRKY11 regulated AR156-triggered ISR through activating the JA signalling pathway, and WRKY70 regulated the ISR through activating the SA signalling pathway. In addition, both WRKY11 and WRKY70 modulated AR156-triggered ISR in a NPR1-dependent manner. In conclusion, WRKY11 and WRKY70 played an important role in regulating the signalling transduction pathways involved in AR156-triggered ISR. This study is the first to illustrate the mechanism by which a single rhizobacterium elicits ISR by simultaneously activating both the SA and JA/ET signalling pathways.

  3. Ectopic expression of a WRKY homolog from Glycine soja alters flowering time in Arabidopsis.

    PubMed

    Luo, Xiao; Sun, Xiaoli; Liu, Baohui; Zhu, Dan; Bai, Xi; Cai, Hua; Ji, Wei; Cao, Lei; Wu, Jing; Wang, Mingchao; Ding, Xiaodong; Zhu, Yanming

    2013-01-01

    Flowering is a critical event in the life cycle of plants; the WRKY-type transcription factors are reported to be involved in many developmental processes sunch as trichome development and epicuticular wax loading, but whether they are involved in flowering time regulation is still unknown. Within this study, we provide clear evidence that GsWRKY20, a member of WRKY gene family from wild soybean, is involved in controlling plant flowering time. Expression of GsWRKY20 was abundant in the shoot tips and inflorescence meristems of wild soybean. Phenotypic analysis showed that GsWRKY20 over-expression lines flowered earlier than the wild-type plants under all conditions: long-day and short-day photoperiods, vernalization, or exogenous GA3 application, indicating that GsWRKY20 may mainly be involved in an autonomous flowering pathway. Further analyses by qRT-PCR and microarray suggests that GsWRKY20 accelerating plant flowering might primarily be through the regulation of flowering-related genes (i.e., FLC, FT, SOC1 and CO) and floral meristem identity genes (i.e., AP1, SEP3, AP3, PI and AG). Our results provide the evidence demonstrating the effectiveness of manipulating GsWRKY20 for altering plant flowering time.

  4. WRKY41 controls Arabidopsis seed dormancy via direct regulation of ABI3 transcript levels not downstream of ABA.

    PubMed

    Ding, Zhong Jie; Yan, Jing Ying; Li, Gui Xin; Wu, Zhong Chang; Zhang, Shu Qun; Zheng, Shao Jian

    2014-09-01

    Although seed dormancy is an important agronomic trait, its molecular basis is poorly understood. ABSCISIC ACID INSENSITIVE 3 (ABI3) plays an essential role in the establishment of seed dormancy. Here, we show that the lack of a seed-expressed WRKY transcription factor, WRKY41, confers reduced primary seed dormancy and thermoinhibition, phenotypes resembling those for a lack of ABI3. Loss-of-function abi3-17 and wrky41 alleles also both confer reduced sensitivity to ABA during germination and early seedling growth. Absence of WRKY41 decreases ABI3 transcript abundance in maturing and imbibed seeds, whereas transgenically overexpressing WRKY41 increases ABI3 expression. Moreover, transgenic overexpression of ABI3 completely restores seed dormancy phenotypes on wrky41. ChIP-qPCR and EMSA reveal that WRKY41 binds directly to the ABI3 promoter through three adjacent W-boxes, and a transactivation assay indicates that these W-boxes are essential for ABI3 expression. Whilst RT-qPCR analysis shows that the regulation of ABI3 by WRKY41 is not through ABA and other factors known to promote ABI3 transcription during seed maturation and germination, we also show that high concentrations of ABA might promote negative feedback regulation of WRKY41 expression. Finally, analysis of the wrky41 aba2 double mutant confirms that WRKY41 and ABA collaboratively regulate ABI3 expression and seed dormancy. In summary, our results demonstrate that WRKY41 is an important regulator of ABI3 expression, and hence of seed dormancy.

  5. Overexpression of GhWRKY27a reduces tolerance to drought stress and resistance to Rhizoctonia solani infection in transgenic Nicotiana benthamiana

    PubMed Central

    Yan, Yan; Jia, Haihong; Wang, Fang; Wang, Chen; Liu, Shuchang; Guo, Xingqi

    2015-01-01

    WRKY proteins constitute transcriptional regulators involved in various biological processes, especially in coping with diverse biotic and abiotic stresses. However, in contrast to other well-characterized WRKY groups, the functions of group III WRKY transcription factors are poorly understood in the economically important crop cotton (Gossypium hirsutum). In this study, a group III WRKY gene from cotton, GhWRKY27a, was isolated and characterized. Our data indicated that GhWRKY27a localized to the nucleus and that GhWRKY27a expression could be strongly induced by abiotic stresses, pathogen infection, and multiple defense-related signaling molecules. Virus-induced gene silencing (VIGS) of GhWRKY27a enhanced tolerance to drought stress in cotton. In contrast, GhWRKY27a overexpression in Nicotiana benthamiana markedly reduced plant tolerance to drought stress, as determined through physiological analyses of leaf water loss, survival rates, and the stomatal aperture. This susceptibility was coupled with reduced stomatal closure in response to abscisic acid and decreased expression of stress-related genes. In addition, GhWRKY27a-overexpressing plants exhibited reduced resistance to Rhizoctonia solani infection, mainly demonstrated by the transgenic lines exhibiting more severe disease symptoms, accompanied by attenuated expression of defense-related genes in N. benthamiana. Taken together, these findings indicated that GhWRKY27a functions in negative responses to drought tolerance and in resistance to R. solani infection. PMID:26483697

  6. Single-strand conformation polymorphism analysis of candidate genes for reliable identification of alleles by capillary array electrophoresis.

    PubMed

    Kuhn, David N; Borrone, James; Meerow, Alan W; Motamayor, Juan C; Brown, J Steven; Schnell, Raymond J

    2005-01-01

    We investigated the reliability of capillary array electrophoresis-single strand conformation polymorphism (CAE-SSCP) to determine if it can be used to identify novel alleles of candidate genes in a germplasm collection. Both strands of three different size fragments (160, 245 and 437 bp) that differed by one or more nucleotides in sequence were analyzed at four different temperatures (18 degrees C, 25 degrees C, 30 degrees C, and 35 degrees C). Mixtures of amplified fragments of either the intron interrupting the C-terminal WRKY domain of the Tc10 locus or the NBS domain of the TcRGH1 locus of Theobroma cacao were electroinjected into all 16 capillaries of an ABI 3100 Genetic Analyzer and analyzed three times at each temperature. Multiplexing of samples of different size range is possible, as intermediate and large fragments were analyzed simultaneously in these experiments. A statistical analysis of the means of the fragment mobilities demonstrated that single-stranded conformers of the fragments could be reliably identified by their mobility at all temperatures and size classes. The order of elution of fragments was not consistent over strands or temperatures for the intermediate and large fragments. If samples are only run once at a single temperature, small fragments could be identified from a single strand at a single temperature. A combination of data from both strands of a single run was needed to identify correctly all four of the intermediate fragments and no combination of data from strands or temperatures would allow the correct identification of two large fragments that differed by only a single single-nucleotide polymorphism (SNP) from a single run. Thus, to adequately assess alleles at a candidate gene locus using SSCP on a capillary array, fragments should be < or =250 bp, samples should be analyzed at two different temperatures between 18 degrees C and 30 degrees C to reduce the variability introduced by the capillaries, data should be combined

  7. The Transcription Factor OsWRKY45 Negatively Modulates the Resistance of Rice to the Brown Planthopper Nilaparvata lugens

    PubMed Central

    Huangfu, Jiayi; Li, Jiancai; Li, Ran; Ye, Meng; Kuai, Peng; Zhang, Tongfang; Lou, Yonggen

    2016-01-01

    WRKY transcription factors play a central role not only in plant growth and development but also in plant stress responses. However, the role of WRKY transcription factors in herbivore-induced plant defenses and their underlying mechanisms, especially in rice, remains largely unclear. Here, we cloned a rice WRKY gene OsWRKY45, whose expression was induced by mechanical wounding, by infestation of the brown planthopper (BPH, Nilaparvata lugens) and by treatment with jasmonic acid (JA) or salicylic acid (SA). The antisense expression of OsWRKY45 (as-wrky) enhanced BPH-induced levels of H2O2 and ethylene, reduced feeding and oviposition preference as well as the survival rate of BPH, and delayed the development of BPH nymphs. Consistently, lower population densities of BPH on as-wrky lines, compared to those on wild-type (WT) plants, were observed in field experiments. On the other hand, as-wrky lines in the field had lower susceptibility to sheath blight (caused by Rhizoctonia solani) but higher susceptibility to rice blast (caused by Magnaporthe oryzae) than did WT plants. These findings suggest that OsWRKY45 plays important but contrasting roles in regulating the resistance of rice to pathogens and herbivores, and attention should be paid if OsWRKY45 is used to develop disease or herbivore-resistant rice. PMID:27258255

  8. The Transcription Factor OsWRKY45 Negatively Modulates the Resistance of Rice to the Brown Planthopper Nilaparvata lugens.

    PubMed

    Huangfu, Jiayi; Li, Jiancai; Li, Ran; Ye, Meng; Kuai, Peng; Zhang, Tongfang; Lou, Yonggen

    2016-05-31

    WRKY transcription factors play a central role not only in plant growth and development but also in plant stress responses. However, the role of WRKY transcription factors in herbivore-induced plant defenses and their underlying mechanisms, especially in rice, remains largely unclear. Here, we cloned a rice WRKY gene OsWRKY45, whose expression was induced by mechanical wounding, by infestation of the brown planthopper (BPH, Nilaparvata lugens) and by treatment with jasmonic acid (JA) or salicylic acid (SA). The antisense expression of OsWRKY45 (as-wrky) enhanced BPH-induced levels of H₂O₂ and ethylene, reduced feeding and oviposition preference as well as the survival rate of BPH, and delayed the development of BPH nymphs. Consistently, lower population densities of BPH on as-wrky lines, compared to those on wild-type (WT) plants, were observed in field experiments. On the other hand, as-wrky lines in the field had lower susceptibility to sheath blight (caused by Rhizoctonia solani) but higher susceptibility to rice blast (caused by Magnaporthe oryzae) than did WT plants. These findings suggest that OsWRKY45 plays important but contrasting roles in regulating the resistance of rice to pathogens and herbivores, and attention should be paid if OsWRKY45 is used to develop disease or herbivore-resistant rice.

  9. Involvement of CmWRKY10 in Drought Tolerance of Chrysanthemum through the ABA-Signaling Pathway.

    PubMed

    Jaffar, Muhammad Abuzar; Song, Aiping; Faheem, Muhammad; Chen, Sumei; Jiang, Jiafu; Liu, Chen; Fan, Qingqing; Chen, Fadi

    2016-05-11

    Drought is one of the important abiotic factors that adversely affects plant growth and production. The WRKY transcription factor plays a pivotal role in plant growth and development, as well as in the elevation of many abiotic stresses. Among three major groups of the WRKY family, the group IIe WRKY has been the least studied in floral crops. Here, we report functional aspects of group IIe WRKY member, i.e., CmWRKY10 in chrysanthemum involved in drought tolerance. The transactivation assay showed that CmWRKY10 had transcriptional activity in yeast cells and subcellular localization demonstrated that it was localized in nucleus. Our previous study showed that CmWRKY10 could be induced by drought in chrysanthemum. Moreover, the overexpression of CmWRKY10 in transgenic chrysanthemum plants improved tolerance to drought stress compared to wild-type (WT). High expression of DREB1A, DREB2A, CuZnSOD, NCED3A, and NCED3B transcripts in overexpressed plants provided strong evidence that drought tolerance mechanism was associated with abscisic acid (ABA) pathway. In addition, lower accumulation of reactive oxygen species (ROS) and higher enzymatic activity of peroxidase, superoxide dismutase and catalase in CmWRKY10 overexpressed lines than that of WT demonstrates its role in drought tolerance. Together, these findings reveal that CmWRKY10 works as a positive regulator in drought stress by regulating stress-related genes.

  10. WRKY76 is a rice transcriptional repressor playing opposite roles in blast disease resistance and cold stress tolerance.

    PubMed

    Yokotani, Naoki; Sato, Yuko; Tanabe, Shigeru; Chujo, Tetsuya; Shimizu, Takafumi; Okada, Kazunori; Yamane, Hisakazu; Shimono, Masaki; Sugano, Shoji; Takatsuji, Hiroshi; Kaku, Hisatoshi; Minami, Eiichi; Nishizawa, Yoko

    2013-11-01

    OsWRKY76 encodes a group IIa WRKY transcription factor of rice. The expression of OsWRKY76 was induced within 48h after inoculation with rice blast fungus (Magnaporthe oryzae), and by wounding, low temperature, benzothiadiazole, and abscisic acid. Green fluorescent protein-fused OsWRKY76 localized to the nuclei in rice epidermal cells. OsWRKY76 showed sequence-specific DNA binding to the W-box element in vitro and exhibited W-box-mediated transcriptional repressor activity in cultured rice cells. Overexpression of OsWRKY76 in rice plants resulted in drastically increased susceptibility to M. oryzae, but improved tolerance to cold stress. Microarray analysis revealed that overexpression of OsWRKY76 suppresses the induction of a specific set of PR genes and of genes involved in phytoalexin synthesis after inoculation with blast fungus, consistent with the observation that the levels of phytoalexins in the transgenic rice plants remained significantly lower than those in non-transformed control plants. Furthermore, overexpression of OsWRKY76 led to the increased expression of abiotic stress-associated genes such as peroxidase and lipid metabolism genes. These results strongly suggest that OsWRKY76 plays dual and opposing roles in blast disease resistance and cold tolerance.

  11. Genome-wide identification and characterization of the Populus WRKY transcription factor family and analysis of their expression in response to biotic and abiotic stresses.

    PubMed

    Jiang, Yuanzhong; Duan, Yanjiao; Yin, Jia; Ye, Shenglong; Zhu, Jingru; Zhang, Faqi; Lu, Wanxiang; Fan, Di; Luo, Keming

    2014-12-01

    WRKY proteins are a large family of regulators involved in various developmental and physiological processes, especially in coping with diverse biotic and abiotic stresses. In this study, 100 putative PtrWRKY genes encoded the proteins contained in the complete WRKY domain in Populus. Phylogenetic analysis revealed that the members of this superfamily among poplar, Arabidopsis, and other species were divided into three groups with several subgroups based on the structures of the WRKY protein sequences. Various cis-acting elements related to stress and defence responses were found in the promoter regions of PtrWRKY genes by promoter analysis. High-throughput transcriptomic analyses identified that 61 of the PtrWRKY genes were induced by biotic and abiotic treatments, such as Marssonina brunnea, salicylic acid (SA), methyl jasmonate (MeJA), wounding, cold, and salinity. Among these PtrWRKY genes, transcripts of 46 selected genes were observed in different tissues, including roots, stems, and leaves. Quantitative RT-PCR analysis further confirmed the induced expression of 18 PtrWRKY genes by one or more stress treatments. The overexpression of an SA-inducible gene, PtrWRKY89, accelerated expression of PR protein genes and improved resistance to pathogens in transgenic poplar, suggesting that PtrWRKY89 is a regulator of an SA-dependent defence-signalling pathway in poplar. Taken together, our results provided significant information for improving the resistance and stress tolerance of woody plants.

  12. Genome-wide analysis of WRKY transcription factors in Solanum lycopersicum.

    PubMed

    Huang, Shengxiong; Gao, Yongfeng; Liu, Jikai; Peng, Xiaoli; Niu, Xiangli; Fei, Zhangjun; Cao, Shuqing; Liu, Yongsheng

    2012-06-01

    The WRKY transcription factors have been implicated in multiple biological processes in plants, especially in regulating defense against biotic and abiotic stresses. However, little information is available about the WRKYs in tomato (Solanum lycopersicum). The recent release of the whole-genome sequence of tomato allowed us to perform a genome-wide investigation for tomato WRKY proteins, and to compare these positively identified proteins with their orthologs in model plants, such as Arabidopsis and rice. In the present study, based on the recently released tomato whole-genome sequences, we identified 81 SlWRKY genes that were classified into three main groups, with the second group further divided into five subgroups. Depending on WRKY domains' sequences derived from tomato, Arabidopsis and rice, construction of a phylogenetic tree demonstrated distinct clustering and unique gene expansion of WRKY genes among the three species. Genome mapping analysis revealed that tomato WRKY genes were enriched on several chromosomes, especially on chromosome 5, and 16 % of the family members were tandemly duplicated genes. The tomato WRKYs from each group were shown to share similar motif compositions. Furthermore, tomato WRKY genes showed distinct temporal and spatial expression patterns in different developmental processes and in response to various biotic and abiotic stresses. The expression of 18 selected tomato WRKY genes in response to drought and salt stresses and Pseudomonas syringae invasion, respectively, was validated by quantitative RT-PCR. Our results will provide a platform for functional identification and molecular breeding study of WRKY genes in tomato and probably other Solanaceae plants.

  13. AtWRKY22 promotes susceptibility to aphids and modulates salicylic acid and jasmonic acid signalling

    PubMed Central

    Kloth, Karen J.; Wiegers, Gerrie L.; Busscher-Lange, Jacqueline; van Haarst, Jan C.; Kruijer, Willem; Bouwmeester, Harro J.; Dicke, Marcel; Jongsma, Maarten A.

    2016-01-01

    Aphids induce many transcriptional perturbations in their host plants, but the signalling cascades responsible and the effects on plant resistance are largely unknown. Through a genome-wide association (GWA) mapping study in Arabidopsis thaliana, we identified WRKY22 as a candidate gene associated with feeding behaviour of the green peach aphid, Myzus persicae. The transcription factor WRKY22 is known to be involved in pathogen-triggered immunity, and WRKY22 gene expression has been shown to be induced by aphids. Assessment of aphid population development and feeding behaviour on knockout mutants and overexpression lines showed that WRKY22 increases susceptibility to M. persicae via a mesophyll-located mechanism. mRNA sequencing analysis of aphid-infested wrky22 knockout plants revealed the up-regulation of genes involved in salicylic acid (SA) signalling and down-regulation of genes involved in plant growth and cell-wall loosening. In addition, mechanostimulation of knockout plants by clip cages up-regulated jasmonic acid (JA)-responsive genes, resulting in substantial negative JA–SA crosstalk. Based on this and previous studies, WRKY22 is considered to modulate the interplay between the SA and JA pathways in response to a wide range of biotic and abiotic stimuli. Its induction by aphids and its role in suppressing SA and JA signalling make WRKY22 a potential target for aphids to manipulate host plant defences. PMID:27107291

  14. Transcription factor StWRKY1 regulates phenylpropanoid metabolites conferring late blight resistance in potato.

    PubMed

    Yogendra, Kalenahalli N; Kumar, Arun; Sarkar, Kobir; Li, Yunliang; Pushpa, Doddaraju; Mosa, Kareem A; Duggavathi, Raj; Kushalappa, Ajjamada C

    2015-12-01

    Quantitative resistance is polygenically controlled and durable, but the underlying molecular and biochemical mechanisms are poorly understood. Secondary cell wall thickening is a critical process in quantitative resistance, regulated by transcriptional networks. This paper provides compelling evidence on the functionality of StWRKY1 transcription factor, in a compatible interaction of potato-Phytophthora infestans, to extend our knowledge on the regulation of the metabolic pathway genes leading to strengthening the secondary cell wall. A metabolomics approach was used to identify resistance-related metabolites belonging to the phenylpropanoid pathway and their biosynthetic genes regulated by StWRKY1. The StWRKY1 gene in resistant potato was silenced to decipher its role in the regulation of phenylpropanoid pathway genes to strengthen the secondary cell wall. Sequencing of the promoter region of StWRKY1 in susceptible genotypes revealed the absence of heat shock elements (HSEs). Simultaneous induction of both the heat shock protein (sHSP17.8) and StWRKY1 following pathogen invasion enables functioning of the latter to interact with the HSE present in the resistant StWRKY1 promoter region. EMSA and luciferase transient expression assays further revealed direct binding of StWRKY1 to promoters of hydroxycinnamic acid amide (HCAA) biosynthetic genes encoding 4-coumarate:CoA ligase and tyramine hydroxycinnamoyl transferase. Silencing of the StWRKY1 gene was associated with signs of reduced late blight resistance by significantly increasing the pathogen biomass and decreasing the abundance of HCAAs. This study provides convincing evidence on the role of StWRKY1 in the regulation of downstream genes to biosynthesize HCAAs, which are deposited to reinforce secondary cell walls.

  15. Transcription factor StWRKY1 regulates phenylpropanoid metabolites conferring late blight resistance in potato

    PubMed Central

    Yogendra, Kalenahalli N.; Kumar, Arun; Sarkar, Kobir; Li, Yunliang; Pushpa, Doddaraju; Mosa, Kareem A.; Duggavathi, Raj; Kushalappa, Ajjamada C.

    2015-01-01

    Quantitative resistance is polygenically controlled and durable, but the underlying molecular and biochemical mechanisms are poorly understood. Secondary cell wall thickening is a critical process in quantitative resistance, regulated by transcriptional networks. This paper provides compelling evidence on the functionality of StWRKY1 transcription factor, in a compatible interaction of potato–Phytophthora infestans, to extend our knowledge on the regulation of the metabolic pathway genes leading to strengthening the secondary cell wall. A metabolomics approach was used to identify resistance-related metabolites belonging to the phenylpropanoid pathway and their biosynthetic genes regulated by StWRKY1. The StWRKY1 gene in resistant potato was silenced to decipher its role in the regulation of phenylpropanoid pathway genes to strengthen the secondary cell wall. Sequencing of the promoter region of StWRKY1 in susceptible genotypes revealed the absence of heat shock elements (HSEs). Simultaneous induction of both the heat shock protein (sHSP17.8) and StWRKY1 following pathogen invasion enables functioning of the latter to interact with the HSE present in the resistant StWRKY1 promoter region. EMSA and luciferase transient expression assays further revealed direct binding of StWRKY1 to promoters of hydroxycinnamic acid amide (HCAA) biosynthetic genes encoding 4-coumarate:CoA ligase and tyramine hydroxycinnamoyl transferase. Silencing of the StWRKY1 gene was associated with signs of reduced late blight resistance by significantly increasing the pathogen biomass and decreasing the abundance of HCAAs. This study provides convincing evidence on the role of StWRKY1 in the regulation of downstream genes to biosynthesize HCAAs, which are deposited to reinforce secondary cell walls. PMID:26417019

  16. A Novel Dominant Transformer Allele of the Sex-Determining Gene Her-1 of Caenorhabditis Elegans

    PubMed Central

    Trent, C.; Wood, W. B.; Horvitz, H. R.

    1988-01-01

    We have characterized a novel dominant allele of the sex-determining gene her-1 of Caenorhabditis elegans. This allele, called n695, results in the incomplete transformation of XX animals into phenotypic males. Previously characterized recessive her-1 alleles transform XO animals into phenotypic hermaphrodites. We have identified five new recessive her-1 mutations as intragenic suppressors of n695. Three of these suppressors are weak, temperature-sensitive alleles. We show that the recessive her-1 mutations are loss-of-function alleles, and that the her-1(n695) mutation results in a gain-of-function at the her-1 locus. The existence of dominant and recessive alleles that cause opposite phenotypic transformations demonstrates that the her-1 gene acts to control sexual identity in C. elegans. PMID:3220248

  17. Arabidopsis WRKY70 is required for full RPP4-mediated disease resistance and basal defense against Hyaloperonospora parasitica.

    PubMed

    Knoth, Colleen; Ringler, Jon; Dangl, Jeffery L; Eulgem, Thomas

    2007-02-01

    AtWRKY70, encoding a WRKY transcription factor, is co-expressed with a set of Arabidopsis genes that share a pattern of RPP4- and RPP7-dependent late upregulation in response to Hyaloperonospora parasitica infection (LURP) genes. We show that AtWRKY70 is required for both full RPP4-mediated resistance and basal defense against H. parasitica. These two defense pathways are related to each other, because they require PAD4 and salicylic acid (SA). RPP7 function, which is independent from PAD4 and SA, is not affected by insertions in AtWRKY70. Although AtWRKY70 is required for RPP4-resistance, it appears not to contribute significantly to RPP4-triggered cell death. Furthermore, our data indicate that AtWRKY70 functions downstream of defense-associated reactive oxygen intermediates and SA. Constitutive and RPP4-induced transcript levels of two other LURP genes are reduced in AtWRKY70 T-DNA mutants, indicating a direct or indirect role for AtWRKY70 in their regulation. We propose that AtWRKY70 is a component of a basal defense mechanism that is boosted by engagement of either RPP4 or RPP7 and is required for RPP4-mediated resistance.

  18. An Updated Collection of Sequence Barcoded Temperature-Sensitive Alleles of Yeast Essential Genes.

    PubMed

    Kofoed, Megan; Milbury, Karissa L; Chiang, Jennifer H; Sinha, Sunita; Ben-Aroya, Shay; Giaever, Guri; Nislow, Corey; Hieter, Philip; Stirling, Peter C

    2015-07-14

    Systematic analyses of essential gene function using mutant collections in Saccharomyces cerevisiae have been conducted using collections of heterozygous diploids, promoter shut-off alleles, through alleles with destabilized mRNA, destabilized protein, or bearing mutations that lead to a temperature-sensitive (ts) phenotype. We previously described a method for construction of barcoded ts alleles in a systematic fashion. Here we report the completion of this collection of alleles covering 600 essential yeast genes. This resource covers a larger gene repertoire than previous collections and provides a complementary set of strains suitable for single gene and genomic analyses. We use deep sequencing to characterize the amino acid changes leading to the ts phenotype in half of the alleles. We also use high-throughput approaches to describe the relative ts behavior of the alleles. Finally, we demonstrate the experimental usefulness of the collection in a high-content, functional genomic screen for ts alleles that increase spontaneous P-body formation. By increasing the number of alleles and improving the annotation, this ts collection will serve as a community resource for probing new aspects of biology for essential yeast genes.

  19. An Updated Collection of Sequence Barcoded Temperature-Sensitive Alleles of Yeast Essential Genes

    PubMed Central

    Kofoed, Megan; Milbury, Karissa L.; Chiang, Jennifer H.; Sinha, Sunita; Ben-Aroya, Shay; Giaever, Guri; Nislow, Corey; Hieter, Philip; Stirling, Peter C.

    2015-01-01

    Systematic analyses of essential gene function using mutant collections in Saccharomyces cerevisiae have been conducted using collections of heterozygous diploids, promoter shut-off alleles, through alleles with destabilized mRNA, destabilized protein, or bearing mutations that lead to a temperature-sensitive (ts) phenotype. We previously described a method for construction of barcoded ts alleles in a systematic fashion. Here we report the completion of this collection of alleles covering 600 essential yeast genes. This resource covers a larger gene repertoire than previous collections and provides a complementary set of strains suitable for single gene and genomic analyses. We use deep sequencing to characterize the amino acid changes leading to the ts phenotype in half of the alleles. We also use high-throughput approaches to describe the relative ts behavior of the alleles. Finally, we demonstrate the experimental usefulness of the collection in a high-content, functional genomic screen for ts alleles that increase spontaneous P-body formation. By increasing the number of alleles and improving the annotation, this ts collection will serve as a community resource for probing new aspects of biology for essential yeast genes. PMID:26175450

  20. WRKY45-dependent priming of diterpenoid phytoalexin biosynthesis in rice and the role of cytokinin in triggering the reaction.

    PubMed

    Akagi, Aya; Fukushima, Setsuko; Okada, Kazunori; Jiang, Chang-Jie; Yoshida, Riichiro; Nakayama, Akira; Shimono, Masaki; Sugano, Shoji; Yamane, Hisakazu; Takatsuji, Hiroshi

    2014-09-01

    Plant activators such as benzothiadiazole (BTH) protect plants against diseases by priming the salicylic acid (SA) signaling pathway. In rice, the transcription factor WRKY45 plays a central role in this process. To investigate the mechanism involved in defense-priming by BTH and the role of WRKY45 in this process, we analyzed the transcripts of biosynthetic genes for diterpenoid phytoalexins (DPs) during the rice-Magnaporthe oryzae interaction. The DP biosynthetic genes were barely upregulated in BTH-treated rice plants, but were induced rapidly after M. oryzae infection in a WRKY45-dependent manner. These results indicate that the DP biosynthetic genes were primed by BTH through WRKY45. Rapid induction of the DP biosynthetic genes was also observed after M. oryzae infection to WRKY45-overexpressing (WRKY45-ox) plants. The changes in gene transcription resulted in accumulation of DPs in WRKY45-ox and BTH-pretreated rice after M. oryzae infection. Previously, we reported that cytokinins (CKs), especially isopentenyladenines, accumulated in M. oryzae-infected rice. Here, we show that DP biosynthetic genes are regulated by the SA/CK synergism in a WRKY45-dependent manner. Together, we propose that CK plays a role in mediating the signal of M. oryzae infection to trigger the induction of DP biosynthetic genes in BTH-primed plants.

  1. A wheat salinity-induced WRKY transcription factor TaWRKY93 confers multiple abiotic stress tolerance in Arabidopsis thaliana.

    PubMed

    Qin, Yuxiang; Tian, Yanchen; Liu, Xiuzhi

    2015-08-21

    Wheat is an important crop in the world. But most of the cultivars are salt sensitive, and often adversely affected by salt stress. WRKY transcription factors play a major role in plant responses to salt stress, but the effective salinity regulatory WRKYs identified in bread wheat are limited and the mechanism of salt stress tolerance is also not well explored. Here, we identified a salt (NaCl) induced class II WRKY transcription factor TaWRKY93. Its transcript level was strongly induced by salt (NaCl) and exogenous abscisic acid (ABA). Over-expression of TaWRKY93 in Arabidopsis thaliana enhanced salt (NaCl), drought, low temperature and osmotic (mannitol) stress tolerance, mainly demonstrated by transgenic plants forming longer primary roots or more lateral roots on MS plates supplemented with NaCl and mannitol individually, higher survival rate under drought and low temperature stress. Further, transgenic plants maintained a more proline content, higher relative water content and less electrolyte leakage than the wild type plants. The transcript abundance of a series of abiotic stress-related genes was up-regulated in the TaWRKY93 transgenic plants. In summary, TaWRKY93 is a new positive regulator of abiotic stress, it may increase salinity, drought and low temperature stress tolerance through enhancing osmotic adjustment, maintaining membrane stability and increasing transcription of stress related genes, and contribute to the superior agricultural traits of SR3 through promoting root development. It can be used as a candidate gene for wheat transgenic engineering breeding against abiotic stress.

  2. Characterization of WRKY co-regulatory networks in rice and Arabidopsis

    PubMed Central

    Berri, Stefano; Abbruscato, Pamela; Faivre-Rampant, Odile; Brasileiro, Ana CM; Fumasoni, Irene; Satoh, Kouji; Kikuchi, Shoshi; Mizzi, Luca; Morandini, Piero; Pè, Mario Enrico; Piffanelli, Pietro

    2009-01-01

    Background The WRKY transcription factor gene family has a very ancient origin and has undergone extensive duplications in the plant kingdom. Several studies have pointed out their involvement in a range of biological processes, revealing that a large number of WRKY genes are transcriptionally regulated under conditions of biotic and/or abiotic stress. To investigate the existence of WRKY co-regulatory networks in plants, a whole gene family WRKYs expression study was carried out in rice (Oryza sativa). This analysis was extended to Arabidopsis thaliana taking advantage of an extensive repository of gene expression data. Results The presented results suggested that 24 members of the rice WRKY gene family (22% of the total) were differentially-regulated in response to at least one of the stress conditions tested. We defined the existence of nine OsWRKY gene clusters comprising both phylogenetically related and unrelated genes that were significantly co-expressed, suggesting that specific sets of WRKY genes might act in co-regulatory networks. This hypothesis was tested by Pearson Correlation Coefficient analysis of the Arabidopsis WRKY gene family in a large set of Affymetrix microarray experiments. AtWRKYs were found to belong to two main co-regulatory networks (COR-A, COR-B) and two smaller ones (COR-C and COR-D), all including genes belonging to distinct phylogenetic groups. The COR-A network contained several AtWRKY genes known to be involved mostly in response to pathogens, whose physical and/or genetic interaction was experimentally proven. We also showed that specific co-regulatory networks were conserved between the two model species by identifying Arabidopsis orthologs of the co-expressed OsWRKY genes. Conclusion In this work we identified sets of co-expressed WRKY genes in both rice and Arabidopsis that are functionally likely to cooperate in the same signal transduction pathways. We propose that, making use of data from co-regulatory networks, it is

  3. Genome-Wide Identification and Expression Analysis of WRKY Transcription Factors under Multiple Stresses in Brassica napus

    PubMed Central

    He, Yajun; Mao, Shaoshuai; Gao, Yulong; Zhu, Liying; Wu, Daoming; Cui, Yixin; Li, Jiana; Qian, Wei

    2016-01-01

    WRKY transcription factors play important roles in responses to environmental stress stimuli. Using a genome-wide domain analysis, we identified 287 WRKY genes with 343 WRKY domains in the sequenced genome of Brassica napus, 139 in the A sub-genome and 148 in the C sub-genome. These genes were classified into eight groups based on phylogenetic analysis. In the 343 WRKY domains, a total of 26 members showed divergence in the WRKY domain, and 21 belonged to group I. This finding suggested that WRKY genes in group I are more active and variable compared with genes in other groups. Using genome-wide identification and analysis of the WRKY gene family in Brassica napus, we observed genome duplication, chromosomal/segmental duplications and tandem duplication. All of these duplications contributed to the expansion of the WRKY gene family. The duplicate segments that were detected indicated that genome duplication events occurred in the two diploid progenitors B. rapa and B. olearecea before they combined to form B. napus. Analysis of the public microarray database and EST database for B. napus indicated that 74 WRKY genes were induced or preferentially expressed under stress conditions. According to the public QTL data, we identified 77 WRKY genes in 31 QTL regions related to various stress tolerance. We further evaluated the expression of 26 BnaWRKY genes under multiple stresses by qRT-PCR. Most of the genes were induced by low temperature, salinity and drought stress, indicating that the WRKYs play important roles in B. napus stress responses. Further, three BnaWRKY genes were strongly responsive to the three multiple stresses simultaneously, which suggests that these 3 WRKY may have multi-functional roles in stress tolerance and can potentially be used in breeding new rapeseed cultivars. We also found six tandem repeat pairs exhibiting similar expression profiles under the various stress conditions, and three pairs were mapped in the stress related QTL regions

  4. Genome-wide detection of allelic gene expression in hepatocellular carcinoma cells using a human exome SNP chip.

    PubMed

    Park, Yon Mi; Cheong, Hyun Sub; Lee, Jong-Keuk

    2014-11-10

    Allelic variations in gene expression influence many biological responses and cause phenotypic variations in humans. In this study, Illumina Human Exome BeadChips containing more than 240,000 single nucleotide polymorphisms (SNPs) were used to identify changes in allelic gene expression in hepatocellular carcinoma cells following lipopolysaccharide (LPS) stimulation. We found 17 monoallelically expressed genes, 58 allelic imbalanced genes, and 7 genes showing allele substitution. In addition, we also detected 33 differentially expressed genes following LPS treatment in vitro using these human exome SNP chips. However, alterations in allelic gene expression following LPS treatment were detected in only three genes (MLXIPL, TNC, and MX2), which were observed in one cell line sample only, indicating that changes in allelic gene expression following LPS stimulation of liver cells are rare events. Among a total of 75 genes showing allelic expression in hepatocellular carcinoma cells, either monoallelic or imbalanced, 43 genes (57.33%) had expression quantitative trait loci (eQTL) data, indicating that high-density exome SNP chips are useful and reliable for studying allelic gene expression. Furthermore, most genes showing allelic expression were regulated by cis-acting mechanisms and were also significantly associated with several human diseases. Overall, our study provides a better understanding of allele-specific gene expression in hepatocellular carcinoma cells with and without LPS stimulation and potential clues for the cause of human disease due to alterations in allelic gene expression.

  5. WRKY Proteins: Signaling and Regulation of Expression during Abiotic Stress Responses

    PubMed Central

    Banerjee, Aditya

    2015-01-01

    WRKY proteins are emerging players in plant signaling and have been thoroughly reported to play important roles in plants under biotic stress like pathogen attack. However, recent advances in this field do reveal the enormous significance of these proteins in eliciting responses induced by abiotic stresses. WRKY proteins act as major transcription factors, either as positive or negative regulators. Specific WRKY factors which help in the expression of a cluster of stress-responsive genes are being targeted and genetically modified to induce improved abiotic stress tolerance in plants. The knowledge regarding the signaling cascade leading to the activation of the WRKY proteins, their interaction with other proteins of the signaling pathway, and the downstream genes activated by them are altogether vital for justified targeting of the WRKY genes. WRKY proteins have also been considered to generate tolerance against multiple abiotic stresses with possible roles in mediating a cross talk between abiotic and biotic stress responses. In this review, we have reckoned the diverse signaling pattern and biological functions of WRKY proteins throughout the plant kingdom along with the growing prospects in this field of research. PMID:25879071

  6. Large-scale profiling and identification of potential regulatory mechanisms for allelic gene expression in colorectal cancer cells.

    PubMed

    Lee, Robin Dong-Woo; Song, Min-Young; Lee, Jong-Keuk

    2013-01-01

    Allelic variation in gene expression is common in humans and this variation is associated with phenotypic variation. In this study, we employed high-density single nucleotide polymorphism (SNP) chips containing 13,900 exonic SNPs to identify genes with allelic gene expression in cells from colorectal cancer cell lines. We found 2 monoallelically expressed genes (ERAP2 and MYLK4), 32 genes with an allelic imbalance in their expression, and 13 genes showing allele substitution by RNA editing. Among a total of 34 allelically expressed genes in colorectal cancer cells, 15 genes (44.1%) were associated with cis-acting eQTL, indicating that large portions of allelically expressed genes are regulated by cis-acting mechanisms of gene expression. In addition, potential regulatory variants present in the proximal promoter regions of genes showing either monoallelic expression or allelic imbalance were not tightly linked with coding SNPs, which were detected with allelic gene expression. These results suggest that multiple rare variants could be involved in the cis-acting regulatory mechanism of allelic gene expression. In the comparison with allelic gene expression data from Centre d'Etude du Polymorphisme Humain (CEPH) family B cells, 12 genes showed B-cell specific allelic imbalance and 1 noncoding SNP showed colorectal cancer cell-specific allelic imbalance. In addition, different patterns of allele substitution were observed between B cells and colorectal cancer cells. Overall, our study not only indicates that allelic gene expression is common in colorectal cancer cells, but our study also provides a better understanding of allele-specific gene expression in colorectal cancer cells.

  7. The cotton WRKY transcription factor GhWRKY17 functions in drought and salt stress in transgenic Nicotiana benthamiana through ABA signaling and the modulation of reactive oxygen species production.

    PubMed

    Yan, Huiru; Jia, Haihong; Chen, Xiaobo; Hao, Lili; An, Hailong; Guo, Xingqi

    2014-12-01

    Drought and high salinity are two major environmental factors that significantly limit the productivity of agricultural crops worldwide. WRKY transcription factors play essential roles in the adaptation of plants to abiotic stresses. However, WRKY genes involved in drought and salt tolerance in cotton (Gossypium hirsutum) are largely unknown. Here, a group IId WRKY gene, GhWRKY17, was isolated and characterized. GhWRKY17 was found to be induced after exposure to drought, salt, H2O2 and ABA. The constitutive expression of GhWRKY17 in Nicotiana benthamiana remarkably reduced plant tolerance to drought and salt stress, as determined through physiological analyses of the germination rate, root growth, survival rate, leaf water loss and Chl content. GhWRKY17 transgenic plants were observed to be more sensitive to ABA-mediated seed germination and root growth. However, overexpressing GhWRKY17 in N. benthamiana impaired ABA-induced stomatal closure. Furthermore, we found that GhWRKY17 modulated the increased sensitivity of plants to drought by reducing the level of ABA, and transcript levels of ABA-inducible genes, including AREB, DREB, NCED, ERD and LEA, were clearly repressed under drought and salt stress conditions. Consistent with the accumulation of reactive oxygen species (ROS), reduced proline contents and enzyme activities, elevated electrolyte leakage and malondialdehyde, and lower expression of ROS-scavenging genes, including APX, CAT and SOD, the GhWRKY17 transgenic plants exhibited reduced tolerance to oxidative stress compared with wild-type plants. These results therefore indicate that GhWRKY17 responds to drought and salt stress through ABA signaling and the regulation of cellular ROS production in plants.

  8. Molecular definition of an allelic series of mutations disrupting the mouse Lmx1a (dreher) gene.

    PubMed

    Chizhikov, Victor; Steshina, Ekaterina; Roberts, Richard; Ilkin, Yesim; Washburn, Linda; Millen, Kathleen J

    2006-10-01

    Mice homozygous for the dreher (dr) mutation are characterized by pigmentation and skeletal abnormalities and striking behavioral phenotypes, including ataxia, vestibular deficits, and hyperactivity. The ataxia is associated with a cerebellar malformation that is remarkably similar to human Dandy-Walker malformation. Previously, positional cloning identified mutations in LIM homeobox transcription factor 1 alpha gene (Lmx1a) in three dr alleles. Two of these alleles, however, are extinct and unavailable for further analysis. In this article we report a new spontaneous dr allele and describe the Lmx1a mutations in this and six additional dr alleles. Strikingly, deletion null, missense, and frameshift mutations in these alleles all cause similar cerebellar malformations, suggesting that all dr mutations analyzed to date are null alleles.

  9. SNP-based large-scale identification of allele-specific gene expression in human B cells.

    PubMed

    Song, Min-Young; Kim, Hye-Eun; Kim, Sun; Choi, Ick-Hwa; Lee, Jong-Keuk

    2012-02-10

    Polymorphism and variations in gene expression provide the genetic basis for human variation. Allelic variation of gene expression, in particular, may play a crucial role in phenotypic variation and disease susceptibility. To identify genes with allelic expression in human cells, we genotyped genomic DNA and cDNA isolated from 31 immortalized B cell lines from three Centre d'Etude du Polymorphisme Humain (CEPH) families using high-density single-nucleotide polymorphism (SNP) chips containing 13,900 exonic SNPs. We identified seven SNPs in five genes with monoallelic expression, 146 SNPs in 125 genes with allelic imbalance in expression with preferentially higher expression of one allele in a heterozygous individual. The monoallelically expressed genes (ERAP2, MDGA1, LOC644422, SDCCAG3P1 and CLTCL1) were regulated by cis-acting, non-imprinted differential allelic control. In addition, all monoallelic gene expression patterns and allelic imbalances in gene expression in B cells were transmitted from parents to offspring in the pedigree, indicating genetic transmission of allelic gene expression. Furthermore, frequent allele substitution, probably due to RNA editing, was also observed in 21 genes in 23 SNPs as well as in 48 SNPs located in regions containing no known genes. In this study, we demonstrated that allelic gene expression is frequently observed in human B cells, and SNP chips are very useful tools for detecting allelic gene expression. Overall, our data provide a valuable framework for better understanding allelic gene expression in human B cells.

  10. Phylogenetic analysis of seven WRKY genes across the palm subtribe Attaleinae (Areceaceae) identifies Syagrus as sister to the coconut

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The origins of the coconut (Cocos nucifera) have been one of the "abominable mysteries" of palm systematics for decades. Previous studies with predominantly plastid genes have indicated an American ancestry for the coconut but with weak support and ambiguous sister relationships. We used primers d...

  11. Virus-induced gene silencing of WRKY53 and an inducible phenylalanine ammonia-lyase in wheat reduces aphid resistance

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Although several wheat genes differentially expressed during the Russian wheat aphid resistance response have recently been identified, their requirement for and specific role in resistance remain unclear. Progress in wheat-aphid interaction research is hampered by inadequate collections of mutant g...

  12. Ectopic Expression of the Wild Grape WRKY Transcription Factor VqWRKY52 in Arabidopsis thaliana Enhances Resistance to the Biotrophic Pathogen Powdery Mildew But Not to the Necrotrophic Pathogen Botrytis cinerea

    PubMed Central

    Wang, Xianhang; Guo, Rongrong; Tu, Mingxing; Wang, Dejun; Guo, Chunlei; Wan, Ran; Li, Zhi; Wang, Xiping

    2017-01-01

    WRKY transcription factors are known to play important roles in plant responses to biotic stresses. We previously showed that the expression of the WRKY gene, VqWRKY52, from Chinese wild Vitis quinquangularis was strongly induced 24 h post inoculation with powdery mildew. In this study, we analyzed the expression levels of VqWRKY52 following treatment with the defense related hormones salicylic acid (SA) and methyl jasmonate, revealing that VqWRKY52 was strongly induced by SA but not JA. We characterized the VqWRKY52 gene, which encodes a WRKY III gene family member, and found that ectopic expression in Arabidopsis thaliana enhanced resistance to powdery mildew and Pseudomonas syringae pv. tomato DC3000, but increased susceptibility to Botrytis cinerea, compared with wild type (WT) plants. The transgenic A. thaliana lines displayed strong cell death induced by the biotrophic powdery mildew pathogen, the hemibiotrophic P. syringe pathogen and the necrotrophic pathogen B. cinerea. In addition, the relative expression levels of various defense-related genes were compared between the transgenic A. thaliana lines and WT plants following the infection by different pathogens. Collectively, the results indicated that VqWRKY52 plays essential roles in the SA dependent signal transduction pathway and that it can enhance the hypersensitive response cell death triggered by microbial pathogens. PMID:28197166

  13. Ectopic Expression of the Wild Grape WRKY Transcription Factor VqWRKY52 in Arabidopsis thaliana Enhances Resistance to the Biotrophic Pathogen Powdery Mildew But Not to the Necrotrophic Pathogen Botrytis cinerea.

    PubMed

    Wang, Xianhang; Guo, Rongrong; Tu, Mingxing; Wang, Dejun; Guo, Chunlei; Wan, Ran; Li, Zhi; Wang, Xiping

    2017-01-01

    WRKY transcription factors are known to play important roles in plant responses to biotic stresses. We previously showed that the expression of the WRKY gene, VqWRKY52, from Chinese wild Vitis quinquangularis was strongly induced 24 h post inoculation with powdery mildew. In this study, we analyzed the expression levels of VqWRKY52 following treatment with the defense related hormones salicylic acid (SA) and methyl jasmonate, revealing that VqWRKY52 was strongly induced by SA but not JA. We characterized the VqWRKY52 gene, which encodes a WRKY III gene family member, and found that ectopic expression in Arabidopsis thaliana enhanced resistance to powdery mildew and Pseudomonas syringae pv. tomato DC3000, but increased susceptibility to Botrytis cinerea, compared with wild type (WT) plants. The transgenic A. thaliana lines displayed strong cell death induced by the biotrophic powdery mildew pathogen, the hemibiotrophic P. syringe pathogen and the necrotrophic pathogen B. cinerea. In addition, the relative expression levels of various defense-related genes were compared between the transgenic A. thaliana lines and WT plants following the infection by different pathogens. Collectively, the results indicated that VqWRKY52 plays essential roles in the SA dependent signal transduction pathway and that it can enhance the hypersensitive response cell death triggered by microbial pathogens.

  14. Identification of novel alleles of the rice blast resistance gene Pi54

    NASA Astrophysics Data System (ADS)

    Vasudevan, Kumar; Gruissem, Wilhelm; Bhullar, Navreet K.

    2015-10-01

    Rice blast is one of the most devastating rice diseases and continuous resistance breeding is required to control the disease. The rice blast resistance gene Pi54 initially identified in an Indian cultivar confers broad-spectrum resistance in India. We explored the allelic diversity of the Pi54 gene among 885 Indian rice genotypes that were found resistant in our screening against field mixture of naturally existing M. oryzae strains as well as against five unique strains. These genotypes are also annotated as rice blast resistant in the International Rice Genebank database. Sequence-based allele mining was used to amplify and clone the Pi54 allelic variants. Nine new alleles of Pi54 were identified based on the nucleotide sequence comparison to the Pi54 reference sequence as well as to already known Pi54 alleles. DNA sequence analysis of the newly identified Pi54 alleles revealed several single polymorphic sites, three double deletions and an eight base pair deletion. A SNP-rich region was found between a tyrosine kinase phosphorylation site and the nucleotide binding site (NBS) domain. Together, the newly identified Pi54 alleles expand the allelic series and are candidates for rice blast resistance breeding programs.

  15. Identification of novel alleles of the rice blast resistance gene Pi54.

    PubMed

    Vasudevan, Kumar; Gruissem, Wilhelm; Bhullar, Navreet K

    2015-10-26

    Rice blast is one of the most devastating rice diseases and continuous resistance breeding is required to control the disease. The rice blast resistance gene Pi54 initially identified in an Indian cultivar confers broad-spectrum resistance in India. We explored the allelic diversity of the Pi54 gene among 885 Indian rice genotypes that were found resistant in our screening against field mixture of naturally existing M. oryzae strains as well as against five unique strains. These genotypes are also annotated as rice blast resistant in the International Rice Genebank database. Sequence-based allele mining was used to amplify and clone the Pi54 allelic variants. Nine new alleles of Pi54 were identified based on the nucleotide sequence comparison to the Pi54 reference sequence as well as to already known Pi54 alleles. DNA sequence analysis of the newly identified Pi54 alleles revealed several single polymorphic sites, three double deletions and an eight base pair deletion. A SNP-rich region was found between a tyrosine kinase phosphorylation site and the nucleotide binding site (NBS) domain. Together, the newly identified Pi54 alleles expand the allelic series and are candidates for rice blast resistance breeding programs.

  16. Expression and Functional Analysis of WRKY Transcription Factors in Chinese Wild Hazel, Corylus heterophylla Fisch

    PubMed Central

    Liang, Li-Song; Ma, Qing-Hua; Chen, Xin; Zong, Jian-Wei; Wang, Gui-Xi

    2015-01-01

    Plant WRKY transcription factors are known to regulate various biotic and abiotic stress responses. In this study we identified a total of 30 putative WRKY unigenes in a transcriptome dataset of the Chinese wild Hazel, Corylus heterophylla, a species that is noted for its cold tolerance. Thirteen full-length of these ChWRKY genes were cloned and found to encode complete protein sequences, and they were divided into three groups, based on the number of WRKY domains and the pattern of zinc finger structures. Representatives of each of the groups, Unigene25835 (group I), Unigene37641 (group II) and Unigene20441 (group III), were transiently expressed as fusion proteins with yellow fluorescent fusion protein in Nicotiana benthamiana, where they were observed to accumulate in the nucleus, in accordance with their predicted roles as transcriptional activators. An analysis of the expression patterns of all 30 WRKY genes revealed differences in transcript abundance profiles following exposure to cold, drought and high salinity conditions. Among the stress-inducible genes, 23 were up-regulated by all three abiotic stresses and the WRKY genes collectively exhibited four different patterns of expression in flower buds during the overwintering period from November to April. The organ/tissue related expression analysis showed that 18 WRKY genes were highly expressed in stem but only 2 (Unigene9262 and Unigene43101) were greatest in male anthotaxies. The expression of Unigene37641, a member of the group II WRKY genes, was substantially up-regulated by cold, drought and salinity treatments, and its overexpression in Arabidopsis thaliana resulted in better seedling growth, compared with wild type plants, under cold treatment conditions. The transgenic lines also had exhibited higher soluble protein content, superoxide dismutase and peroxidase activiety and lower levels of malondialdehyde, which collectively suggets that Unigene37641 expression promotes cold tolerance. PMID

  17. Expression and Functional Analysis of WRKY Transcription Factors in Chinese Wild Hazel, Corylus heterophylla Fisch.

    PubMed

    Zhao, Tian-Tian; Zhang, Jin; Liang, Li-Song; Ma, Qing-Hua; Chen, Xin; Zong, Jian-Wei; Wang, Gui-Xi

    2015-01-01

    Plant WRKY transcription factors are known to regulate various biotic and abiotic stress responses. In this study we identified a total of 30 putative WRKY unigenes in a transcriptome dataset of the Chinese wild Hazel, Corylus heterophylla, a species that is noted for its cold tolerance. Thirteen full-length of these ChWRKY genes were cloned and found to encode complete protein sequences, and they were divided into three groups, based on the number of WRKY domains and the pattern of zinc finger structures. Representatives of each of the groups, Unigene25835 (group I), Unigene37641 (group II) and Unigene20441 (group III), were transiently expressed as fusion proteins with yellow fluorescent fusion protein in Nicotiana benthamiana, where they were observed to accumulate in the nucleus, in accordance with their predicted roles as transcriptional activators. An analysis of the expression patterns of all 30 WRKY genes revealed differences in transcript abundance profiles following exposure to cold, drought and high salinity conditions. Among the stress-inducible genes, 23 were up-regulated by all three abiotic stresses and the WRKY genes collectively exhibited four different patterns of expression in flower buds during the overwintering period from November to April. The organ/tissue related expression analysis showed that 18 WRKY genes were highly expressed in stem but only 2 (Unigene9262 and Unigene43101) were greatest in male anthotaxies. The expression of Unigene37641, a member of the group II WRKY genes, was substantially up-regulated by cold, drought and salinity treatments, and its overexpression in Arabidopsis thaliana resulted in better seedling growth, compared with wild type plants, under cold treatment conditions. The transgenic lines also had exhibited higher soluble protein content, superoxide dismutase and peroxidase activiety and lower levels of malondialdehyde, which collectively suggets that Unigene37641 expression promotes cold tolerance.

  18. Allelic inclusion in a pre-B-cell line that generates immunoglobulin heavy chain genes in vitro.

    PubMed Central

    Beck-Engeser, G; Jäck, H M; Wabl, M

    1987-01-01

    In a pre-B-cell line that rearranges its heavy chain gene segments in vitro, we found that the rate of productive rearrangement on one allele was not influenced by the presence of heavy chain protein encoded by the other allele. This shows that allelic exclusion of heavy chain genes is not regulated at the genetic level. Images PMID:3103122

  19. Allelic polymorphism in transcriptional regulatory regions of HLA-DQB genes

    PubMed Central

    1991-01-01

    Class II genes of the human major histocompatibility complex (MHC) are highly polymorphic. Allelic variation of structural genes provides diversity in immune cell interactions, contributing to the formation of the T cell repertoire and to susceptibility to certain autoimmune diseases. We now report that allelic polymorphism also exists in the promoter and upstream regulatory regions (URR) of human histocompatibility leukocyte antigen (HLA) class II genes. Nucleotide sequencing of these regulatory regions of seven alleles of the DQB locus reveals a number of allele-specific polymorphisms, some of which lie in functionally critical consensus regions thought to be highly conserved in class II promoters. These sequence differences also correspond to allelic differences in binding of nuclear proteins to the URR. Fragments of the URR of two DQB alleles were analyzed for binding to nuclear proteins extracted from human B lymphoblastoid cell lines (B- LCL). Gel retardation assays showed substantially different banding patterns to the two promoters, including prominent variation in nuclear protein binding to the partially conserved X box regions and a novel upstream polymorphic sequence element. Comparison of these two polymorphic alleles in a transient expression system demonstrated a marked difference in their promoter strengths determined by relative abilities to initiate transcription of the chloramphenicol acetyltransferase reporter gene in human B-LCL. Shuttling of URR sequences between alleles showed that functional variation corresponded to both the X box and upstream sequence polymorphic sites. These findings identify an important source of MHC class II diversity, and suggest the possibility that such regulatory region polymorphisms may confer allelic differences in expression, inducibility, and/or tissue specificity of class II molecules. PMID:1985121

  20. Regulation of WRKY46 Transcription Factor Function by Mitogen-Activated Protein Kinases in Arabidopsis thaliana

    PubMed Central

    Sheikh, Arsheed H.; Eschen-Lippold, Lennart; Pecher, Pascal; Hoehenwarter, Wolfgang; Sinha, Alok K.; Scheel, Dierk; Lee, Justin

    2016-01-01

    Mitogen-activated protein kinase (MAPK) cascades are central signaling pathways activated in plants after sensing internal developmental and external stress cues. Knowledge about the downstream substrate proteins of MAPKs is still limited in plants. We screened Arabidopsis WRKY transcription factors as potential targets downstream of MAPKs, and concentrated on characterizing WRKY46 as a substrate of the MAPK, MPK3. Mass spectrometry revealed in vitro phosphorylation of WRKY46 at amino acid position S168 by MPK3. However, mutagenesis studies showed that a second phosphosite, S250, can also be phosphorylated. Elicitation with pathogen-associated molecular patterns (PAMPs), such as the bacterial flagellin-derived flg22 peptide led to in vivo destabilization of WRKY46 in Arabidopsis protoplasts. Mutation of either phosphorylation site reduced the PAMP-induced degradation of WRKY46. Furthermore, the protein for the double phosphosite mutant is expressed at higher levels compared to wild-type proteins or single phosphosite mutants. In line with its nuclear localization and predicted function as a transcriptional activator, overexpression of WRKY46 in protoplasts raised basal plant defense as reflected by the increase in promoter activity of the PAMP-responsive gene, NHL10, in a MAPK-dependent manner. Thus, MAPK-mediated regulation of WRKY46 is a mechanism to control plant defense. PMID:26870073

  1. Allelic gene expression imbalance of bovine IGF2, LEP and CCL2 genes in liver, kidney and pituitary.

    PubMed

    Olbromski, R; Siadkowska, E; Zelazowska, B; Zwierzchowski, L

    2013-02-01

    Allelic expression imbalance (AEI) is an important genetic factor being the cause of differences in phenotypic traits that can be heritable. Studying AEI can be useful in searching for factors that modulate gene expression and help to understand molecular mechanisms underlying phenotypic changes. Although it was commonly recognized in many species and we know many genes show allelic expression imbalance, this phenomena was not studied on a larger scale in cattle. Using the pyrosequencing method we analyzed a set of 29 bovine genes in order to find those that have preferential allelic expression. The study was conducted in three tissues: liver, pituitary and kindey. Out of the studied group of genes 3 of them-LEP (leptin), IGF2 (insulin-like growth factor 2), CCL2 (chemokine C-C motif ligand 2) showed allelic expression imbalance.

  2. The transcriptional network of WRKY53 in cereals links oxidative responses to biotic and abiotic stress inputs.

    PubMed

    Van Eck, Leon; Davidson, Rebecca M; Wu, Shuchi; Zhao, Bingyu Y; Botha, Anna-Maria; Leach, Jan E; Lapitan, Nora L V

    2014-06-01

    The transcription factor WRKY53 is expressed during biotic and abiotic stress responses in cereals, but little is currently known about its regulation, structure and downstream targets. We sequenced the wheat ortholog TaWRKY53 and its promoter region, which revealed extensive similarity in gene architecture and cis-acting regulatory elements to the rice ortholog OsWRKY53, including the presence of stress-responsive abscisic acid-responsive elements (ABRE) motifs and GCC-boxes. Four proteins interacted with the WRKY53 promoter in yeast one-hybrid assays, suggesting that this gene can receive inputs from diverse stress-related pathways such as calcium signalling and senescence, and environmental cues such as drought and ultraviolet radiation. The Ser/Thr receptor kinase ORK10/LRK10 and the apoplastic peroxidase POC1 are two downstream targets for regulation by the WRKY53 transcription factor, predicted based on the presence of W-box motifs in their promoters and coregulation with WRKY53, and verified by electrophoretic mobility shift assay (EMSA). Both ORK10/LRK10 and POC1 are upregulated during cereal responses to pathogens and aphids and important components of the oxidative burst during the hypersensitive response. Taken with our yeast two-hybrid assay which identified a strong protein-protein interaction between microsomal glutathione S-transferase 3 and WRKY53, this implies that the WRKY53 transcriptional network regulates oxidative responses to a wide array of stresses.

  3. Characterization of the gene and protein of the common alpha 1-antitrypsin normal M2 allele.

    PubMed Central

    Nukiwa, T; Brantly, M L; Ogushi, F; Fells, G A; Crystal, R G

    1988-01-01

    The normal M2 variant of alpha 1-antitrypsin (alpha 1AT) was cloned from a genomic DNA library of an individual homozygous for this allele. Sequencing of all coding exons of the M2 gene revealed it was identical to the common M1(Val213) gene except for two bases (M1(Val213) CGT Arg101, M2 CAT His101; M1(Val213) GAA Glu376 M2 GAC Asp376). Analysis of the sequence of the M1(Val213) and M2 genes around residue 101 revealed the M1 Arg101----M2 His101 caused a loss of the cutting site for the restriction endonuclease RsaI. Using this enzyme, as well as 19-mer oligonucleotides probes centered at residues 101 and 376, evaluation of genomic DNA from 22 M1 alleles and 14 M2 alleles revealed that residue 101 was Arg in all M1 alleles and His in all M2 alleles, while residue 376 was Glu in all M1 alleles and Asp in all M2 alleles. Despite the differences in sequence at two amino acids, the M1(Val213) and M2 proteins function similarly as assessed by quantification of the association rate constant of each for their natural substrate neutrophil elastase. In the context that there are two mutations separating the M1(Val213) and M2 alleles, it is likely that there is another alpha 1AT variant that was an intermediate in the evolution of these genes. Images Figure 2 Figure 4 Figure 1 Figure 3 PMID:2901226

  4. No evidence for allelic association between bipolar disorder and monoamine oxidase A gene polymorphisms

    SciTech Connect

    Craddock, N.; Daniels, J.; Roberts, E.

    1995-08-14

    We have tested the hypothesis that DNA markers in the MAOA gene show allelic association with bipolar affective disorder. Eighty-four unrelated Caucasian patients with DSM III-R bipolar disorder and 84 Caucasian controls were typed for three markers in MAOA: a dinucleotide repeat in intron 2, a VNTR in intron 1, and an Fnu4HI RFLP in exon 8. No evidence for allelic association was observed between any of the markers and bipolar disorder. 9 refs., 1 tab.

  5. [Analysis of allelic content of genes responsible for baking properties in allocytoplasmic wheat hybrids].

    PubMed

    Klimushina, M V; Divashuk, M G; Mukhammed, T A K; Semenov, O G; Karlov, G I

    2013-05-01

    A collection comprised of allocytoplasmic hybrids of mild wheat (ACPH) was screened for the allelic state of genes responsible for baking properties (high-molecular glutenins, puroindolines, and Waxy). The possibility of the introgression of the Waxy gene of T. timopheevii into the mild wheat genome was demonstrated in several ACPH samples using the set of molecular markers. Allelic gene variants responsible for the baking properties were revealed for 22 ACPH samples, which make it possible to detect the most challenging samples for both molecular-genetic research and applied science.

  6. The Arabidopsis Mitochondrial Protease FtSH4 Is Involved in Leaf Senescence via Regulation of WRKY-Dependent Salicylic Acid Accumulation and Signaling.

    PubMed

    Zhang, Shengchun; Li, Cui; Wang, Rui; Chen, Yaxue; Shu, Si; Huang, Ruihua; Zhang, Daowei; Li, Jian; Xiao, Shi; Yao, Nan; Yang, Chengwei

    2017-04-01

    Mitochondria and autophagy play important roles in the networks that regulate plant leaf senescence and cell death. However, the molecular mechanisms underlying the interactions between mitochondrial signaling and autophagy are currently not well understood. This study characterized the function of the Arabidopsis (Arabidopsis thaliana) mitochondrial AAA-protease gene FtSH4 in regulating autophagy and senescence, finding that FtSH4 mediates WRKY-dependent salicylic acid (SA) accumulation and signaling. Knockout of FtSH4 in the ftsh4-4 mutant resulted in severe leaf senescence, cell death, and high autophagy levels. The level of SA increased dramatically in the ftsh4-4 mutant. Expression of nahG in the ftsh4-4 mutant led to decreased SA levels and suppressed the leaf senescence and cell death phenotypes. The transcript levels of several SA synthesis and signaling genes, including SALICYLIC ACIDINDUCTION DEFICIENT2 (SID2), NON-RACE-SPECIFIC DISEASE RESISTANCE1 (NDR1), and NONEXPRESSOR OF PATHOGENESIS-RELATED PROTEINS1 (NPR1), increased significantly in the ftsh4-4 mutants compared with the wild type. Loss of function of SID2, NDR1, or NPR1 in the ftsh4-4 mutant reversed the ftsh4-4 senescence and autophagy phenotypes. Furthermore, ftsh4-4 mutants had elevated levels of transcripts of several WRKY genes, including WRKY40, WRKY46, WRKY51, WRKY60, WRKY63, and WRKY75; all of these WRKY proteins can bind to the promoter of SID2 Loss of function of WRKY75 in the ftsh4-4 mutants decreased the levels of SA and reversed the senescence phenotype. Taken together, these results suggest that the mitochondrial ATP-dependent protease FtSH4 may regulate the expression of WRKY genes by modifying the level of reactive oxygen species and the WRKY transcription factors that control SA synthesis and signaling in autophagy and senescence.

  7. Patterns of variation among distinct alleles of the Flag silk gene from Nephila clavipes.

    PubMed

    Higgins, Linden E; White, Sheryl; Nuñez-Farfán, Juan; Vargas, Jesus

    2007-02-20

    Spider silk proteins and their genes are very attractive to researchers in a wide range of disciplines because they permit linking many levels of organization. However, hypotheses of silk gene evolution have been built primarily upon single sequences of each gene each species, and little is known about allelic variation within a species. Silk genes are known for their repeat structure with high levels of homogenization of nucleotide and amino acid sequence among repeated units. One common explanation for this homogeneity is gene convergence. To test this model, we sequenced multiple alleles of one intron-exon segment from the Flag gene from four populations of the spider Nephila clavipes and compared the new sequences to a published sequence. Our analysis revealed very high levels of heterozygosity in this gene, with no pattern of population differentiation. There was no evidence of gene convergence within any of these alleles, with high levels of nucleotide and amino acid substitution among the repeating motifs. Our data suggest that minimally, there is relaxed selection on mutations in this gene and that there may actually be positive selection for heterozygosity.

  8. Genome-Wide Investigation of WRKY Transcription Factors Involved in Terminal Drought Stress Response in Common Bean

    PubMed Central

    Wu, Jing; Chen, Jibao; Wang, Lanfen; Wang, Shumin

    2017-01-01

    WRKY transcription factor plays a key role in drought stress. However, the characteristics of the WRKY gene family in the common bean (Phaseolus vulgaris L.) are unknown. In this study, we identified 88 complete WRKY proteins from the draft genome sequence of the “G19833” common bean. The predicted genes were non-randomly distributed in all chromosomes. Basic information, amino acid motifs, phylogenetic tree and the expression patterns of PvWRKY genes were analyzed, and the proteins were classified into groups 1, 2, and 3. Group 2 was further divided into five subgroups: 2a, 2b, 2c, 2d, and 2e. Finally, we detected 19 WRKY genes that were responsive to drought stress using qRT-PCR; 11 were down-regulated, and 8 were up-regulated under drought stress. This study comprehensively examines WRKY proteins in the common bean, a model food legume, and it provides a foundation for the functional characterization of the WRKY family and opportunities for understanding the mechanisms of drought stress tolerance in this plant. PMID:28386267

  9. StWRKY8 transcription factor regulates benzylisoquinoline alkaloid pathway in potato conferring resistance to late blight.

    PubMed

    Yogendra, Kalenahalli N; Dhokane, Dhananjay; Kushalappa, Ajjamada C; Sarmiento, Felipe; Rodriguez, Ernesto; Mosquera, Teresa

    2017-03-01

    The resistance to late blight is either qualitative or quantitative in nature. Quantitative resistance is durable, but challenging due to polygenic inheritance. In the present study, the diploid potato genotypes resistant and susceptible to late blight, were profiled for metabolites. Tissue specific metabolite analysis of benzylisoquinoline alkaloids (BIAs) in response to pathogen infection revealed increased accumulation of morphinone, codeine-6-glucuronide and morphine-3-glucuronides. These BIAs are antimicrobial compounds and possibly involved in cell wall reinforcement, especially through cross-linking cell wall pectins. Quantitative reverse transcription-PCR studies revealed higher expressions of TyDC, NCS, COR-2 and StWRKY8 transcription factor genes, in resistant genotypes than in susceptible genotype, following pathogen inoculation. A luciferase transient expression assay confirmed the binding of the StWRKY8 TF to promoters of downstream genes, elucidating a direct regulatory role on BIAs biosynthetic genes. Sequence analysis of StWRKY8 in potato genotypes revealed polymorphism in the WRKY DNA binding domain in the susceptible genotype, which is important for the regulatory function of this gene. A complementation assay of StWRKY8 in Arabidopsis wrky33 mutant background was associated with decreased fungal biomass. In conclusion, StWRKY8 regulates the biosynthesis of BIAs that are both antimicrobial and reinforce cell walls to contain the pathogen to initial infection.

  10. Genome-Wide Investigation of WRKY Transcription Factors Involved in Terminal Drought Stress Response in Common Bean.

    PubMed

    Wu, Jing; Chen, Jibao; Wang, Lanfen; Wang, Shumin

    2017-01-01

    WRKY transcription factor plays a key role in drought stress. However, the characteristics of the WRKY gene family in the common bean (Phaseolus vulgaris L.) are unknown. In this study, we identified 88 complete WRKY proteins from the draft genome sequence of the "G19833" common bean. The predicted genes were non-randomly distributed in all chromosomes. Basic information, amino acid motifs, phylogenetic tree and the expression patterns of PvWRKY genes were analyzed, and the proteins were classified into groups 1, 2, and 3. Group 2 was further divided into five subgroups: 2a, 2b, 2c, 2d, and 2e. Finally, we detected 19 WRKY genes that were responsive to drought stress using qRT-PCR; 11 were down-regulated, and 8 were up-regulated under drought stress. This study comprehensively examines WRKY proteins in the common bean, a model food legume, and it provides a foundation for the functional characterization of the WRKY family and opportunities for understanding the mechanisms of drought stress tolerance in this plant.

  11. Deletions of recessive disease genes: CNV contribution to carrier states and disease-causing alleles.

    PubMed

    Boone, Philip M; Campbell, Ian M; Baggett, Brett C; Soens, Zachry T; Rao, Mitchell M; Hixson, Patricia M; Patel, Ankita; Bi, Weimin; Cheung, Sau Wai; Lalani, Seema R; Beaudet, Arthur L; Stankiewicz, Pawel; Shaw, Chad A; Lupski, James R

    2013-09-01

    Over 1200 recessive disease genes have been described in humans. The prevalence, allelic architecture, and per-genome load of pathogenic alleles in these genes remain to be fully elucidated, as does the contribution of DNA copy-number variants (CNVs) to carrier status and recessive disease. We mined CNV data from 21,470 individuals obtained by array-comparative genomic hybridization in a clinical diagnostic setting to identify deletions encompassing or disrupting recessive disease genes. We identified 3212 heterozygous potential carrier deletions affecting 419 unique recessive disease genes. Deletion frequency of these genes ranged from one occurrence to 1.5%. When compared with recessive disease genes never deleted in our cohort, the 419 recessive disease genes affected by at least one carrier deletion were longer and located farther from known dominant disease genes, suggesting that the formation and/or prevalence of carrier CNVs may be affected by both local and adjacent genomic features and by selection. Some subjects had multiple carrier CNVs (307 subjects) and/or carrier deletions encompassing more than one recessive disease gene (206 deletions). Heterozygous deletions spanning multiple recessive disease genes may confer carrier status for multiple single-gene disorders, for complex syndromes resulting from the combination of two or more recessive conditions, or may potentially cause clinical phenotypes due to a multiply heterozygous state. In addition to carrier mutations, we identified homozygous and hemizygous deletions potentially causative for recessive disease. We provide further evidence that CNVs contribute to the allelic architecture of both carrier and recessive disease-causing mutations. Thus, a complete recessive carrier screening method or diagnostic test should detect CNV alleles.

  12. Allelic association of human dopamine D sub 2 receptor gene in alcoholism

    SciTech Connect

    Blum, K.; Sheridan, P.J.; Montgomery, A.; Jagadeeswaran, P.; Nogami, H.; Briggs, A.H. ); Noble, E.P.; Ritchie, T.; Cohn, J.B. )

    1990-04-18

    In a blinded experiment, the authors report the first allelic association of the dopamine D{sub 2} receptor gene in alcoholism. From 70 brain samples of alcoholics and nonalcoholics, DNA was digested with restriction endonucleases and probed with a clone that contained the entire 3{prime} coding exon, the polyadenylation signal, and approximately 16.4 kilobases of noncoding 3{prime} sequence of the human dopamine D{sub 2} receptor gene ({lambda}hD2G1). In the present samples, the presence of A1 allele of the dopamine D{sub 2} receptor gene correctly classified 77% of alcoholics, and its absence classified 72% of nonalcoholics. The polymorphic pattern of this receptor gene suggests that a gene that confers susceptibility to at least one form of alcoholism is located on the q22-q23 region of chromosome 11.

  13. Differential allelic expression of a fibrillin gene (FBNI) in patients with Marfan syndrome

    SciTech Connect

    Hewett, D.; Lynch, J.; Sykes, B.; Firth, H.; Child, A.

    1994-09-01

    Marfan syndrome is a connective-tissue disorder affecting cardiovascular, skeletal, and ocular systems. The major Marfan locus has been identified as the FBN1 gene on chromosome 15; this codes for the extracellular-matrix protein fibrillin, a 350-kD constituent of the 8-10-nm elastin-associated microfibrils. The authors identified five MFS patients who were heterozygous for an RsaI restriction-site dimorphism in the 3{prime} UTR of the FBN1 gene. This expressed variation was used to distinguish the mRNA output from each of the two FBN1 alleles in fibroblast cultures from these five patients. Three of the patients were shown to produce <5% of the normal level of FBN1 transcripts from one of their alleles. This null-allele phenotype was not observed in 10 nonmarfanoid fibroblast cell lines. 26 refs., 4 figs.

  14. Is the Ala12 variant of the PPARG gene an "unthrifty allele"?

    PubMed Central

    Ruiz-Narvaez, E

    2005-01-01

    Background: The thrifty genotype hypothesis proposes that genetic susceptibility to type 2 diabetes results from the positive selection of "thrifty" alleles in the past. A corollary of this hypothesis is that genetic variants protecting against the development of diabetes are "unthrifty" and thus subject to negative selection during human evolution. Methods: It was assessed whether age estimates of the diabetes protective PPARG Ala12 allele indicate effects of natural selection. Based on published data from four populations, the date of origin of the diabetes protective PPARG Ala12 variant was estimated using both allele frequency and linkage disequilibrium (LD) with the C1431T single nucleotide polymorphism in exon 6 of the PPARG gene. Results: The best LD based estimate of the age of the Ala12 allele gave an average of ∼32 000 years with a maximum upper bound of ∼58 000 years. Assuming a population with a growth rate of r = 0.01 per generation, the frequency based estimate of the age of the Ala12 variant gave an average of ∼27 000 years with a maximum upper bound of ∼42 000 years. Discussion: The similarity of both time estimates is consistent with selective equivalence of the diabetes protective PPARG Ala12 allele and the diabetes susceptible PPARG Pro12 allele. PMID:15994875

  15. The Beet Cyst Nematode Heterodera schachtii Modulates the Expression of WRKY Transcription Factors in Syncytia to Favour Its Development in Arabidopsis Roots

    PubMed Central

    Ali, Muhammad Amjad; Wieczorek, Krzysztof; Kreil, David P.; Bohlmann, Holger

    2014-01-01

    Cyst nematodes invade the roots of their host plants as second stage juveniles and induce a syncytium which is the only source of nutrients throughout their life. A recent transcriptome analysis of syncytia induced by the beet cyst nematode Heterodera schachtii in Arabidopsis roots has shown that thousands of genes are up-regulated or down-regulated in syncytia as compared to root segments from uninfected plants. Among the down-regulated genes are many which code for WRKY transcription factors. Arabidopsis contains 66 WRKY genes with 59 represented by the ATH1 GeneChip. Of these, 28 were significantly down-regulated and 6 up-regulated in syncytia as compared to control root segments. We have studied here the down-regulated genes WRKY6, WRKY11, WRKY17 and WRKY33 in detail. We confirmed the down-regulation in syncytia with promoter::GUS lines. Using various overexpression lines and mutants it was shown that the down-regulation of these WRKY genes is important for nematode development, probably through interfering with plant defense reactions. In case of WRKY33, this might involve the production of the phytoalexin camalexin. PMID:25033038

  16. Overexpression of the Brassica rapa transcription factor WRKY12 results in reduced soft rot symptoms caused by Pectobacterium carotovorum in Arabidopsis and Chinese cabbage.

    PubMed

    Kim, H S; Park, Y H; Nam, H; Lee, Y M; Song, K; Choi, C; Ahn, I; Park, S R; Lee, Y H; Hwang, D J

    2014-09-01

    Chinese cabbage (Brassica rapa L. ssp. pekinensis), an important vegetable crop, can succumb to diseases such as bacterial soft rot, resulting in significant loss of crop productivity and quality. Pectobacterium carotovorum ssp. carotovorum (Pcc) causes soft rot disease in various plants, including Chinese cabbage. To overcome crop loss caused by bacterial soft rot, a gene from Chinese cabbage was isolated and characterised in this study. We isolated the BrWRKY12 gene from Chinese cabbage, which is a group II member of the WRKY transcription factor superfamily. The 645-bp coding sequence of BrWRKY12 translates to a protein with a molecular mass of approximately 24.4 kDa, and BrWRKY12 was exclusively localised in the nucleus. Transcripts of BrWRKY12 were induced by Pcc infection in Brassica. Heterologous expression of BrWRKY12 resulted in reduced susceptibility to Pcc but not to Pseudomonas syringae pv. tomato in Arabidopsis. Defence-associated genes, such as AtPDF1.2 and AtPGIP2, were constitutively expressed in transgenic lines overexpressing BrWRKY12. The expression of AtWKRY12, which is the closest orthologue of BrWRKY12, was down-regulated by Pcc in Arabidopsis. However, the Atwrky12-2 mutants did not show any difference in response to Pcc, pointing to a difference in function of WRKY12 in Brassica and Arabidopsis. Furthermore, BrWRKY12 in Chinese cabbage also exhibited enhanced resistance to bacterial soft rot and increased the expression of defence-associated genes. In summary, BrWRKY12 confers enhanced resistance to Pcc through transcriptional activation of defence-related genes.

  17. Allele specific-PCR and melting curve analysis showed relatively high frequency of β-casein gene A1 allele in Iranian Holstein, Simmental and native cows.

    PubMed

    Gholami, M; Hafezian, S H; Rahimi, G; Farhadi, A; Rahimi, Z; Kahrizi, D; Kiani, S; Karim, H; Vaziri, S; Muhammadi, S; Veisi, F; Ghadiri, K; Shetabi, H; Zargooshi, J

    2016-10-31

    There are two allelic forms of A1 and A2 of β-casein gene in dairy cattle. Proteolytic digestion of bovine β-casein A1 type produces bioactive peptide of β-casomorphin-7 known as milk devil. β-casomorphin-7 causes many diseases, including type 1 diabetes, cardiovascular disease syndrome, sudden death and madness. The aim of the present study was to determine the different allelic forms of β-casein gene in Iranian Holstein, Simmental and native cattle in order to identify A1 and A2 variants. The blood samples were collected randomly and DNA was extracted using modified salting out method. An 854 bp fragment including part of exon 7 and part of intron 6 of β-casein gene was amplified by allele specific polymerase chain reaction (AS-PCR). Also, the accuracy of AS-PCR genotyping has been confirmed by melting temperature curve analysis using Real-time PCR machinery. The comparison of observed allele and genotype frequency among the studied breeds was performed using the Fisher exact and Chi-squared test, respectively by SAS program. Obtained results showed the A1 allele frequencies of 50, 51.57, 54.5, 49.4 and 46.6% in Holstein, Simmental, Sistani, Taleshi and Mazandarani cattle populations, respectively. The chi-square test was shown that no any populations were in Hardy-Weinberg equilibrium for studied marker locus. Comparison and analysis of the test results for allelic frequency showed no any significant differences between breeds (P>0.05). The frequency of observed genotypes only differs significantly between Holstein and Taleshi breeds but no any statistically significant differences were found for other breeds (P>0.05). A relatively high frequency of β-casein A1 allele was observed in Iranian native cattle. Therefore, determine the genotypes and preference alleles A2 in these native and commercial cattle is recommended.

  18. Regulatory Divergence between Parental Alleles Determines Gene Expression Patterns in Hybrids

    PubMed Central

    Combes, Marie-Christine; Hueber, Yann; Dereeper, Alexis; Rialle, Stéphanie; Herrera, Juan-Carlos; Lashermes, Philippe

    2015-01-01

    Both hybridization and allopolyploidization generate novel phenotypes by conciliating divergent genomes and regulatory networks in the same cellular context. To understand the rewiring of gene expression in hybrids, the total expression of 21,025 genes and the allele-specific expression of over 11,000 genes were quantified in interspecific hybrids and their parental species, Coffea canephora and Coffea eugenioides using RNA-seq technology. Between parental species, cis- and trans-regulatory divergences affected around 32% and 35% of analyzed genes, respectively, with nearly 17% of them showing both. The relative importance of trans-regulatory divergences between both species could be related to their low genetic divergence and perennial habit. In hybrids, among divergently expressed genes between parental species and hybrids, 77% was expressed like one parent (expression level dominance), including 65% like C. eugenioides. Gene expression was shown to result from the expression of both alleles affected by intertwined parental trans-regulatory factors. A strong impact of C. eugenioides trans-regulatory factors on the upregulation of C. canephora alleles was revealed. The gene expression patterns appeared determined by complex combinations of cis- and trans-regulatory divergences. In particular, the observed biased expression level dominance seemed to be derived from the asymmetric effects of trans-regulatory parental factors on regulation of alleles. More generally, this study illustrates the effects of divergent trans-regulatory parental factors on the gene expression pattern in hybrids. The characteristics of the transcriptional response to hybridization appear to be determined by the compatibility of gene regulatory networks and therefore depend on genetic divergences between the parental species and their evolutionary history. PMID:25819221

  19. Association between Age and the 7 Repeat Allele of the Dopamine D4 Receptor Gene

    PubMed Central

    Szekely, Anna; Bircher, Julianna; Vereczkei, Andrea; Balota, David A.; Sasvari-Szekely, Maria; Ronai, Zsolt

    2016-01-01

    Longevity is in part (25%) inherited, and genetic studies aim to uncover allelic variants that play an important role in prolonging life span. Results to date confirm only a few gene variants associated with longevity, while others show inconsistent results. However, GWAS studies concentrate on single nucleotide polymorphisms, and there are only a handful of studies investigating variable number of tandem repeat variations related to longevity. Recently, Grady and colleagues (2013) reported a remarkable (66%) accumulation of those carrying the 7 repeat allele of the dopamine D4 receptor gene in a large population of 90–109 years old Californian centenarians, as compared to an ancestry-matched young population. In the present study we demonstrate the same association using continuous age groups in an 18–97 years old Caucasian sample (N = 1801, p = 0.007). We found a continuous pattern of increase from 18–75, however frequency of allele 7 carriers decreased in our oldest age groups. Possible role of gene-environment interaction effects driven by historical events are discussed. In accordance with previous findings, we observed association preferentially in females (p = 0.003). Our results underlie the importance of investigating non-disease related genetic variants as inherited components of longevity, and confirm, that the 7-repeat allele of the dopamine D4 receptor gene is a longevity enabling genetic factor, accumulating in the elderly female population. PMID:27992450

  20. Association study of human VN1R1 pheromone receptor gene alleles and gender.

    PubMed

    Mitropoulos, Constantinos; Papachatzopoulou, Adamantia; Menounos, Panagiotis G; Kolonelou, Christina; Pappa, Magda; Bertolis, George; Gerou, Spiros; Patrinos, George P

    2007-01-01

    Pheromones are water-soluble chemicals that elicit neuroendocrine and physiological changes, while they also provide information about gender within individuals of the same species. VN1R1 is the only functional pheromone receptor in humans. We have undertaken a large mutation screening approach in 425 adult individuals from the Hellenic population to investigate whether the allelic differences, namely alleles 1a and 1b present in the human VN1R1 gene, are gender specific. Here we show that both VN1R1 1a and 1b alleles are found in chromosomes of both male and female subjects at frequency of 26.35% and 73.65%, respectively. Given the fact that those allelic differences potentially cause minor changes in the protein conformation and its transmembrane domains, as simulated by the TMHMM software, our data suggest that the allelic differences in the human VN1R1 gene are unlikely to be associated with gender and hence to contribute to distinct gender-specific behavior.

  1. The allelic relationship of genes giving resistance to mungbean yellow mosaic virus in blackgram.

    PubMed

    Verma, R P; Singh, D P

    1986-09-01

    The allelic relationship of resistance genes for MYMV was studied in blackgram (V. mungo (L.) Hepper). The resistant donors to MYMV - 'Pant U84' and 'UPU 2', and their F1, F2 and F3 generations - were inoculated artificially using an insect vector, whitefly (Bemisia tabaci Genn.). The two recessive genes previously reported for resistance were found to be the same in both donors.

  2. Microarray analysis of Arabidopsis WRKY33 mutants in response to the necrotrophic fungus Botrytis cinerea

    PubMed Central

    Sham, Arjun; Moustafa, Khaled; Al-Shamisi, Shamma; Alyan, Sofyan; Iratni, Rabah

    2017-01-01

    The WRKY33 transcription factor was reported for resistance to the necrotrophic fungus Botrytis cinerea. Using microarray-based analysis, we compared Arabidopsis WRKY33 overexpressing lines and wrky33 mutant that showed altered susceptibility to B. cinerea with their corresponding wild-type plants. In the wild-type, about 1660 genes (7% of the transcriptome) were induced and 1054 genes (5% of the transcriptome) were repressed at least twofold at early stages of inoculation with B. cinerea, confirming previous data of the contribution of these genes in B. cinerea resistance. In Arabidopsis wild-type plant infected with B. cinerea, the expressions of the differentially expressed genes encoding for proteins and metabolites involved in pathogen defense and non-defense responses, seem to be dependent on a functional WRKY33 gene. The expression profile of 12-oxo-phytodienoic acid- and phytoprostane A1-treated Arabidopsis plants in response to B. cinerea revealed that cyclopentenones can also modulate WRKY33 regulation upon inoculation with B. cinerea. These results support the role of electrophilic oxylipins in mediating plant responses to B. cinerea infection through the TGA transcription factor. Future directions toward the identification of the molecular components in cyclopentenone signaling will elucidate the novel oxylipin signal transduction pathways in plant defense. PMID:28207847

  3. MAP kinases phosphorylate rice WRKY45.

    PubMed

    Ueno, Yoshihisa; Yoshida, Riichiro; Kishi-Kaboshi, Mitsuko; Matsushita, Akane; Jiang, Chang-Jie; Goto, Shingo; Takahashi, Akira; Hirochika, Hirohiko; Takatsuji, Hiroshi

    2013-06-01

    WRKY45 transcription factor is a central regulator of disease resistance mediated by the salicylic acid (SA) signaling pathway in rice. SA-activated WRKY45 protein induces the accumulation of its own mRNA. However, the mechanism underlying this regulation is still unknown. Here, we report three lines of evidence showing that a mitogen-activated protein kinase (MAPK) cascade is involved in this regulation. An inhibitor of MAPK kinase (MAPKK) suppressed the increase in WRKY45 transcript level in response to SA. Two MAPKs, OsMPK4 and OsMPK6, phosphorylated WRKY45 protein in vitro. The activity of OsMPK6 was rapidly upregulated by SA treatment in rice cells. These results suggest that WRKY45 is regulated by MAPK-dependent phosphorylation in the SA pathway.

  4. [Association of allelic polymorphisms of genes matrix Gla-protein system with ischemic atherothrombotic stroke].

    PubMed

    Garbuzova, V Yu; Stroy, D A; Dosenko, V E; Dubovyk, Ye I; Borodenko, A O; Shimko, K A; Obukhova, O A; Ataman, O V

    2015-01-01

    There are results of the determination of 10 polymorphisms of matrix Gla-protein system (gene MGP-T(-138)-->C (rs1800802), G(-7)-->A (rs1800801), Thr83-->Ala (rs4236), gene VDR-FokI (rs2228570), BsmI (rs1544410), ApaI (rs7975232), TaqI (rs731236), gene GGCX-Arg325-->Gln (rs699664), gene VKORS1-T(2255)-->C (rs2359612), gene BMP-2-Ser37-->Ala (rs2273073)) into 170 patients with ischemic atherothrombotic stroke (IATS) and 124 healthy individual is (control group). It is established that there is a connection between the IATS and polymorphic variants of genes MGP (G(-7)-->A) and VKORC1 (T(2255)-->C). The risk of IATS in carriers of minor allele A/A (G(-7)-->A polymorphism) in 2.6 times higher than in carriers of the major allele (G/A + G/G), and C/C genotype (T(2255)-->C polymorphism) in 2.2 times higher than the homozygotes of major allele. The coincidence of patients T/C and G/G, C/C and G/A genotypes, and A/A genotype (G(-7)-->A polymorphism) with any genotype T(2255)-->C polymorphism are increases the risk of IATS.

  5. Constitutive expression of a salinity-induced wheat WRKY transcription factor enhances salinity and ionic stress tolerance in transgenic Arabidopsis thaliana

    SciTech Connect

    Qin, Yuxiang; Tian, Yanchen; Han, Lu; Yang, Xinchao

    2013-11-15

    Highlights: •A class II WRKY transcription factor, TaWRKY79 was isolated and characterized. •TaWRKY79 was induced by NaCl or abscisic acid. •843 bp regulatory segment was sufficient to respond to ABA or NaCl treatment. •TaWRKY79 enhanced salinity and ionic tolerance while reduced sensitivity to ABA. •TaWRKY79 increased salinity and ionic tolerance in an ABA-dependent pathway. -- Abstract: The isolation and characterization of TaWRKY79, a wheat class II WRKY transcription factor, is described. Its 1297 bp coding region includes a 987 bp long open reading frame. TaWRKY79 was induced by stressing seedlings with either NaCl or abscisic acid (ABA). When a fusion between an 843 bp segment upstream of the TaWRKY79 coding sequence and GUS was introduced into Arabidopsis thaliana, GUS staining indicated that this upstream segment captured the sequence(s) required to respond to ABA or NaCl treatment. When TaWRKY79 was constitutively expressed as a transgene in A. thaliana, the transgenic plants showed an improved capacity to extend their primary root in the presence of either 100 mM NaCl, 10 mM LiCl or 2 μM ABA. The inference was that TaWRKY79 enhanced the level of tolerance to both salinity and ionic stress, while reducing the level of sensitivity to ABA. The ABA-related genes ABA1, ABA2 ABI1 and ABI5 were all up-regulated in the TaWRKY79 transgenic plants, suggesting that the transcription factor operates in an ABA-dependent pathway.

  6. Disagreement in genotyping results of drug resistance alleles of the Plasmodium falciparum dihydrofolate reductase (Pfdhfr) gene by allele-specific PCR (ASPCR) assays and Sanger sequencing.

    PubMed

    Sharma, Divya; Lather, Manila; Dykes, Cherry L; Dang, Amita S; Adak, Tridibes; Singh, Om P

    2016-01-01

    The rapid spread of antimalarial drug resistance in Plasmodium falciparum over the past few decades has necessitated intensive monitoring of such resistance for an effective malaria control strategy. P. falciparum dihydropteroate synthase (Pfdhps) and P. falciparum dihydrofolate reductase (Pfdhfr) genes act as molecular markers for resistance against the antimalarial drugs sulphadoxine and pyrimethamine, respectively. Resistance to pyrimethamine which is used as a partner drug in artemisinin combination therapy (ACT) is associated with several mutations in the Pfdhfr gene, namely A16V, N51I, C59R, S108N/T and I164L. Therefore, routine monitoring of Pfdhfr-drug-resistant alleles in a population may help in effective drug resistance management. Allele-specific PCR (ASPCR) is one of the commonly used methods for molecular genotyping of these alleles. In this study, we genotyped 55 samples of P. falciparum for allele discrimination at four codons of Pfdhfr (N51, C59, S108 and I164) by ASPCR using published methods and by Sanger's DNA sequencing method. We found that the ASPCR identified a significantly higher number of mutant alleles as compared to the DNA sequencing method. Such discrepancies arise due to the non-specificity of some of the allele-specific primer sets and due to the lack of sensitivity of Sanger's DNA sequencing method to detect minor alleles present in multiple clone infections. This study reveals the need of a highly specific and sensitive method for genotyping and detecting minor drug-resistant alleles present in multiple clonal infections.

  7. Distribution of the mutated delta 32 allele of the CCR5 gene in a Sicilian population.

    PubMed

    Sidoti, A; D'Angelo, R; Rinaldi, C; De Luca, G; Pino, F; Salpietro, C; Giunta, D E; Saltalamacchia, F; Amato, A

    2005-06-01

    The CCR5 gene encodes a cell-surface chemokine receptor molecule that serves as a co-receptor for macrophage-tropic strains of human immunodeficiency virus type 1 (HIV-1). A mutation in this gene may alter the expression or the function of the protein product, thereby altering chemokine binding and/or signalling or HIV-1 infection of cells that normally express CCR5 protein. Individuals homozygous for a 32-bp deletion allele of CCR5 (CCR5 delta32), heritable as a Mendelian trait, are relatively resistant to HIV-1 infection. The CCR5 delta32 mutation is present in the Caucasian population at different frequencies. The aim of this study was to investigate the frequency of truncated alleles of the CCR5 delta32 gene in a Sicilian population, as the interpopulation variation in CCR5 delta32 frequency may be a significant factor in the prediction of AIDS endemicity in future studies. We examined 901 healthy individuals from several Sicilian provinces. We found a mean (+/- standard deviation) delta32 allele frequency (fr) of 0.04 +/- 0.012. The highest value was observed in the province of Messina, with a mean delta32 allele frequency of 0.06 +/- 0.024, where we collected samples from a cohort of 114 HIV-1-infected individuals. The observed frequency amongst these patients was quite low (fr = 0.03 +/- 0.031) compared to the healthy population, although the difference was not statistically significant.

  8. Allelic association of the D2 dopamine receptor gene with receptor-binding characteristics in alcoholism

    SciTech Connect

    Noble, E.P.; Blum, K.; Ritchie, T.; Montgomery, A.; Sheridan, P.J. )

    1991-07-01

    The allelic association of the human D2 dopamine receptor gene with the binding characteristics of the D2 dopamine receptor was determined in 66 brains of alcoholic and non-alcoholic subjects. In a blinded experiment, DNA from the cerebral cortex was treated with the restriction endonuclease Taql and probed with a 1.5-kilobase (kb) digest of a clone (lambda hD2G1) of the human D2 dopamine receptor gene. The binding characteristics (Kd (binding affinity) and Bmax (number of binding sites)) of the D2 dopamine receptor were determined in the caudate nuclei of these brains using tritiated spiperone as the ligand. The adjusted Kd was significantly lower in alcoholic than in nonalcoholic subjects. In subjects with the A1 allele, in whom a high association with alcoholism was found, the Bmax was significantly reduced compared with the Bmax of subjects with the A2 allele. Moreover, a progressively reduced Bmax was found in subjects with A2/A2, A1/A2, and A1/A1 alleles, with subjects with A2/A2 having the highest mean values, and subjects with A1/A1, the lowest. The polymorphic pattern of the D2 dopamine receptor gene and its differential expression of receptors suggests the involvement of the dopaminergic system in conferring susceptibility to at least one subtype of severe alcoholism.

  9. Genome-wide transcript analysis of maize hybrids: allelic additive gene expression and yield heterosis.

    PubMed

    Guo, Mei; Rupe, Mary A; Yang, Xiaofeng; Crasta, Oswald; Zinselmeier, Christopher; Smith, Oscar S; Bowen, Ben

    2006-09-01

    Heterosis, or hybrid vigor, has been widely exploited in plant breeding for many decades, but the molecular mechanisms underlying the phenomenon remain unknown. In this study, we applied genome-wide transcript profiling to gain a global picture of the ways in which a large proportion of genes are expressed in the immature ear tissues of a series of 16 maize hybrids that vary in their degree of heterosis. Key observations include: (1) the proportion of allelic additively expressed genes is positively associated with hybrid yield and heterosis; (2) the proportion of genes that exhibit a bias towards the expression level of the paternal parent is negatively correlated with hybrid yield and heterosis; and (3) there is no correlation between the over- or under-expression of specific genes in maize hybrids with either yield or heterosis. The relationship of the expression patterns with hybrid performance is substantiated by analysis of a genetically improved modern hybrid (Pioneer hybrid 3394) versus a less improved older hybrid (Pioneer hybrid 3306) grown at different levels of plant density stress. The proportion of allelic additively expressed genes is positively associated with the modern high yielding hybrid, heterosis and high yielding environments, whereas the converse is true for the paternally biased gene expression. The dynamic changes of gene expression in hybrids responding to genotype and environment may result from differential regulation of the two parental alleles. Our findings suggest that differential allele regulation may play an important role in hybrid yield or heterosis, and provide a new insight to the molecular understanding of the underlying mechanisms of heterosis.

  10. How species evolve collectively: implications of gene flow and selection for the spread of advantageous alleles

    PubMed Central

    MORJAN, CARRIE L.; RIESEBERG, LOREN H.

    2008-01-01

    The traditional view that species are held together through gene flow has been challenged by observations that migration is too restricted among populations of many species to prevent local divergence. However, only very low levels of gene flow are necessary to permit the spread of highly advantageous alleles, providing an alternative means by which low-migration species might be held together. We re-evaluate these arguments given the recent and wide availability of indirect estimates of gene flow. Our literature review of FST values for a broad range of taxa suggests that gene flow in many taxa is considerably greater than suspected from earlier studies and often is sufficiently high to homogenize even neutral alleles. However, there are numerous species from essentially all organismal groups that lack sufficient gene flow to prevent divergence. Crude estimates on the strength of selection on phenotypic traits and effect sizes of quantitative trait loci (QTL) suggest that selection coefficients for leading QTL underlying phenotypic traits may be high enough to permit their rapid spread across populations. Thus, species may evolve collectively at major loci through the spread of favourable alleles, while simultaneously differentiating at other loci due to drift and local selection. PMID:15140081

  11. Ribosomal protein genes are highly enriched among genes with allele-specific expression in the interspecific F1 hybrid catfish.

    PubMed

    Chen, Ailu; Wang, Ruijia; Liu, Shikai; Peatman, Eric; Sun, Luyang; Bao, Lisui; Jiang, Chen; Li, Chao; Li, Yun; Zeng, Qifan; Liu, Zhanjiang

    2016-06-01

    Interspecific hybrids provide a rich source for the analysis of allele-specific expression (ASE). In this work, we analyzed ASE in F1 hybrid catfish using RNA-Seq datasets. While the vast majority of genes were expressed with both alleles, 7-8 % SNPs exhibited significant differences in allele ratios of expression. Of the 66,251 and 177,841 SNPs identified from the datasets of the liver and gill, 5420 (8.2 %) and 13,390 (7.5 %) SNPs were identified as significant ASE-SNPs, respectively. With these SNPs, a total of 1519 and 3075 ASE-genes were identified. Gene Ontology analysis revealed that genes encoding cytoplasmic ribosomal proteins (RP) were highly enriched among ASE genes. Parent-of-origin was determined for 27 and 30 ASE RP genes in the liver and gill, respectively. The results indicated that genes from both channel catfish and blue catfish were involved in ASE. However, each RP gene appeared to be almost exclusively expressed from only one parent, indicating that ribosomes in the hybrid catfish were in the "hybrid" form. Overall representation of RP transcripts among the transcriptome appeared lower in the F1 hybrid catfish than in channel catfish or blue catfish, suggesting that the "hybrid" ribosomes may work more efficiently for translation in the F1 hybrid catfish.

  12. A pseudodeficiency allele (D152N) of the human beta-glucuronidase gene.

    PubMed

    Vervoort, R; Islam, M R; Sly, W; Chabas, A; Wevers, R; de Jong, J; Liebaers, I; Lissens, W

    1995-10-01

    We present evidence that a 480G-->A transition in the coding region of the beta-glucuronidase gene, which results in an aspartic-acid-to-asparagine substitution at amino acid position 152 (D152N), produces a pseudodeficiency allele (GUSBp) that leads to greatly reduced levels of beta-glucuronidase activity without apparent deleterious consequences. The 480G-->A mutation was found initially in the pseudodeficient mother of a child with mucopolysaccharidosis VII (MPSVII), but it was not on her disease-causing allele, which carried the L176F mutation. The 480G-->A change was also present in an unrelated individual with another MPSVII allele who had unusually low beta-glucuronidase activity, but whose clinical symptoms were probably unrelated to beta-glucuronidase deficiency. This individual also had an R357X mutation, probably on his second allele. We screened 100 unrelated normal individuals for the 480G-->A mutation with a PCR method and detected one carrier. Reduced beta-glucuronidase activity following transfection of COS cells with the D152N cDNA supported the causal relationship between the D152N allele and pseudodeficiency. The mutation reduced the fraction of expressed enzyme that was secreted. Pulse-chase experiments indicated that the reduced activity in COS cells was due to accelerated intracellular turnover of the D152N enzyme. They also suggested that a potential glycosylation site created by the mutation is utilized in approximately 50% of the enzyme expressed.

  13. Differential alleleic expression of the type II collagen gene (COL2A2) in osteoarthritic cartilage

    SciTech Connect

    Loughlin, J.; Irven, C.; Sykes, B.; Athanasou, N.; Carr, A.

    1995-05-01

    Osteoarthritis (OA) is a common debilitating disease resulting from the degeneration of articular cartilage. The major protein of cartilage is type II collagen, which is encoded by the COL2A1 gene. Mutations at this locus have been discovered in several individuals with inherited disorders of cartilage. We have identified 27 primary OA patients who are heterozygous for sequence dimorphisms located in the coding region of COL2A1. These dimorphisms were used to distinguish the mRNA output from each of the two COL2A1 alleles in articular cartilage obtained from each patient. Three patients demonstrated differential allelic expression and produced <12% of the normal level of mRNA from one of their COL2A1 alleles. The same allele shows reduced expression in a well-defined OA population than in a control group, suggesting the possible existence of a rare COL2A1 allele that predisposes to OA. 31 refs., 4 figs., 3 tabs.

  14. Null alleles of the aldolase B gene in patients with hereditary fructose intolerance.

    PubMed

    Ali, M; Tunçman, G; Cross, N C; Vidailhet, M; Bökesoy, I; Gitzelmann, R; Cox, T M

    1994-06-01

    We report three new mutations in the gene for aldolase B that are associated with hereditary fructose intolerance (HFI). Two nonsense mutations create opal termination codons: R3op (C-->T, Arg3-->ter, exon 2) was found in homozygous form in four affected members of a large consanguineous Turkish pedigree and R59op (C-->T, Arg59-->ter, exon 3) was found on one allele in a woman of Austrian origin known to harbour one copy of the east European mutation, N334K (Asn334-->Lys). The third mutation occurred in a French HFI patient known to be heterozygous for the widespread mutation, A174D (Ala174-->Asp): a single mutation, G-->A, in the consensus acceptor site 3' of intron 6 was found on the remaining allele. These mutations are predicted to abrogate synthesis of functional protein and thus represent null alleles of aldolase B. The mutant alleles can be readily detected in the amplification refractory mutation system (ARMS) or (for R59op and 3' intron 6) by digestion of amplified genomic fragments with DdeI or A1wNI, respectively, to facilitate direct diagnosis of HFI by molecular analysis of aldolase B genes.

  15. Allelic expression of mammalian imprinted genes in a matrotrophic lizard, Pseudemoia entrecasteauxii.

    PubMed

    Griffith, Oliver W; Brandley, Matthew C; Belov, Katherine; Thompson, Michael B

    2016-03-01

    Genomic imprinting is a process that results in the differential expression of genes depending on their parent of origin. It occurs in both plants and live-bearing mammals, with imprinted genes typically regulating the ability of an embryo to manipulate the maternal provision of nutrients. Genomic imprinting increases the potential for selection to act separately on paternally and maternally expressed genes, which increases the number of opportunities that selection can facilitate embryonic control over maternal nutrient provision. By looking for imprinting in an independent matrotrophic lineage, the viviparous lizard Pseudemoia entrecasteauxii (Scincidae), we test the hypothesis that genomic imprinting facilitates the evolution of substantial placental nutrient transport to embryos (matrotrophy). We sequenced transcriptomes from the embryonic component of lizard placentae to determine whether there are parent-of-origin differences in expression of genes that are imprinted in mammals. Of these genes, 19 had sufficiently high expression in the lizard to identify polymorphisms in transcribed sequences. We identified bi-allelic expression in 17 genes (including insulin-like growth factor 2), indicating that neither allele was imprinted. These data suggest that either genomic imprinting has not evolved in this matrotrophic skink or, if it has, it has evolved in different genes to mammals. We outline how these hypotheses can be tested. This study highlights important differences between mammalian and reptile pregnancy and the absence of any shared imprinting genes reflects fundamental differences in the way that pregnancy has evolved in these two lineages.

  16. Development of disease-resistant rice by optimized expression of WRKY45.

    PubMed

    Goto, Shingo; Sasakura-Shimoda, Fuyuko; Suetsugu, Mai; Selvaraj, Michael Gomez; Hayashi, Nagao; Yamazaki, Muneo; Ishitani, Manabu; Shimono, Masaki; Sugano, Shoji; Matsushita, Akane; Tanabata, Takanari; Takatsuji, Hiroshi

    2015-08-01

    The rice transcription factor WRKY45 plays a central role in the salicylic acid signalling pathway and mediates chemical-induced resistance to multiple pathogens, including Magnaporthe oryzae and Xanthomonas oryzae pv. oryzae. Previously, we reported that rice transformants overexpressing WRKY45 driven by the maize ubiquitin promoter were strongly resistant to both pathogens; however, their growth and yield were negatively affected because of the trade-off between the two conflicting traits. Also, some unknown environmental factor(s) exacerbated this problem. Here, we report the development of transgenic rice lines resistant to both pathogens and with agronomic traits almost comparable to those of wild-type rice. This was achieved by optimizing the promoter driving WRKY45 expression. We isolated 16 constitutive promoters from rice genomic DNA and tested their ability to drive WRKY45 expression. Comparisons among different transformant lines showed that, overall, the strength of WRKY45 expression was positively correlated with disease resistance and negatively correlated with agronomic traits. We conducted field trials to evaluate the growth of transgenic and control lines. The agronomic traits of two lines expressing WRKY45 driven by the OsUbi7 promoter (PO sUbi7 lines) were nearly comparable to those of untransformed rice, and both lines were pathogen resistant. Interestingly, excessive WRKY45 expression rendered rice plants sensitive to low temperature and salinity, and stress sensitivity was correlated with the induction of defence genes by these stresses. These negative effects were barely observed in the PO sUbi7 lines. Moreover, their patterns of defence gene expression were similar to those in plants primed by chemical defence inducers.

  17. Gene Deletion by Fluorescence-Reported Allelic Exchange Mutagenesis in Chlamydia trachomatis

    PubMed Central

    Mueller, Konrad E.; Wolf, Katerina

    2016-01-01

    ABSTRACT Although progress in Chlamydia genetics has been rapid, genomic modification has previously been limited to point mutations and group II intron insertions which truncate protein products. The bacterium has thus far been intractable to gene deletion or more-complex genomic integrations such as allelic exchange. Herein, we present a novel suicide vector dependent on inducible expression of a chlamydial gene that renders Chlamydia trachomatis fully genetically tractable and permits rapid reverse genetics by fluorescence-reported allelic exchange mutagenesis (FRAEM). We describe the first available system of targeting chlamydial genes for deletion or allelic exchange as well as curing plasmids from C. trachomatis serovar L2. Furthermore, this approach permits the monitoring of mutagenesis by fluorescence microscopy without disturbing bacterial growth, a significant asset when manipulating obligate intracellular organisms. As proof of principle, trpA was successfully deleted and replaced with a sequence encoding both green fluorescent protein (GFP) and β-lactamase. The trpA-deficient strain was unable to grow in indole-containing medium, and this phenotype was reversed by complementation with trpA expressed in trans. To assess reproducibility at alternate sites, FRAEM was repeated for genes encoding type III secretion effectors CTL0063, CTL0064, and CTL0065. In all four cases, stable mutants were recovered one passage after the observation of transformants, and allelic exchange was limited to the specific target gene, as confirmed by whole-genome sequencing. Deleted sequences were not detected by quantitative real-time PCR (qPCR) from isogenic mutant populations. We demonstrate that utilization of the chlamydial suicide vector with FRAEM renders C. trachomatis highly amenable to versatile and efficient genetic manipulation. PMID:26787828

  18. Populus simonii × Populus nigra WRKY70 is involved in salt stress and leaf blight disease responses.

    PubMed

    Zhao, Hui; Jiang, Jing; Li, Kailong; Liu, Guifeng; Tsai, Chung-Jui

    2017-03-22

    WRKY transcription factors (TFs) are important regulators in the complex stress response signaling networks in plants, but the detailed mechanisms underlying these regulatory networks have not been fully characterized. In the present study, we identified a Group III WRKY gene (PsnWRKY70, Potri.016G137900) from Populussimonii × Populusnigra and explored its function under salt and pathogen stresses. The promoter sequence that is located 2471-bp upstream from the start codon (SC) of PsnWRKY70 contained many stress-responsive cis-elements. Yeast one-hybrid assay suggested the upstream regulators, PsnWRKY70, PsnNAM (Potri.009G141600), PsnMYB (Potri.006G000800) and PsnGT1 (Potri.010G055000), probably modulate the expression of the PsnWRKY70 gene by specifically binding to the W-box or GT1GMSCAM4 (GT1) element. Yeast two-hybrid assay and transcriptome analysis revealed that HP1 (Potri.004G092100), RRM (Potri.008G146700), Ulp1 (Potri.002G105700) and some mitogen-activated protein kinase cascade members probably interact with PsnWRKY70 TF to response to salt stress. Compared with non-transgenic (NT) plants, PsnWRKY70-overexpressing (OEX) plants exhibited improved leaf blight disease resistance, while PsnWRKY70-repressing (REX) plants displayed enhanced salt stress tolerance. PsnWRKY70, PsnNAM, PsnMYB and PsnGT1 exhibited similar expression patterns in NT under salt and leaf blight disease stresses. The differentially expressed genes (DEGs) from NT vs OEX1 and the DEGs from NT vs REX1 exhibited considerable diversification. Most of the DEGs between NT and OEX1 were involved in aromatic amino acid biosynthesis, secondary metabolism, programmed cell death, peroxisomes and disease resistance. Most of the DEGs between NT and REX1 were related to desiccation response, urea transmembrane transport, abscisic acid response, calcium ion transport and hydrogen peroxide transmembrane transport. Our findings not only revealed the salt stress response signal transduction pathway of PsnWRKY

  19. The heterogeneous allelic repertoire of human toll-like receptor (TLR) genes.

    PubMed

    Georgel, Philippe; Macquin, Cécile; Bahram, Seiamak

    2009-11-17

    Toll-Like Receptors (TLR) are critical elements of the innate arm of the vertebrate immune system. They constitute a multigenic family of receptors which collectively bind a diverse array of--exogeneous as well as endogeneous--ligands. An exponential burst of knowledge has defined their biological role in fight against infections and generation/modulation of auto-immune disorders. Hence, they could at least be conceptually recognized--despite being structurally unrelated - as innate counterparts to Major Histocompatibility Complex (MHC) molecules--equally recognizing antigenic ligands (albeit structurally more homogeneous i.e., peptides), again derived from self and/or non-self sources--preeminent this time in adaptive immunity. Our great disparities in face of infections and/or susceptibility to auto-immune diseases have provoked an intense search for genetic explanations, in part satisfied by the extraordinary MHC allelic repertoire. An equally in-depth and systematic analysis of TLR diversity is lacking despite numerous independent reports of a growing number of SNPs within these loci. The work described here aims at providing a preliminary picture of the allelic repertoire--and not purely SNPs--of all 10 human TLR coding sequences (with exception of TLR3) within a single cohort of up to 100 individuals. It appears from our work that TLR are unequally polymorphic: TLR2 (DNA alleles: 7/protein alleles: 3), 4 (4/3), 7 (6/3), 8 (9/2) and 9 (8/3) being comparatively least diverse whereas TLR1 (11/10), 5 (14/12), 6 (10/8) and 10 (15/10) show a substantial number of alleles. In addition to allelic assignment of a large number of SNPs, 10 new polymorphic positions were hereby identified. Hence this work depicts a first overview of the diversity of almost all human TLR genes, a prelude for large-scale population genetics as well as genetic association studies.

  20. Identification of genes escaping X inactivation by allelic expression analysis in a novel hybrid mouse model.

    PubMed

    Berletch, Joel B; Ma, Wenxiu; Yang, Fan; Shendure, Jay; Noble, William S; Disteche, Christine M; Deng, Xinxian

    2015-12-01

    X chromosome inactivation (XCI) is a female-specific mechanism that serves to balance gene dosage between the sexes whereby one X chromosome in females is inactivated during early development. Despite this silencing, a small portion of genes escape inactivation and remain expressed from the inactive X (Xi). Little is known about the distribution of escape from XCI in different tissues in vivo and about the mechanisms that control tissue-specific differences. Using a new binomial model in conjunction with a mouse model with identifiable alleles and skewed X inactivation we are able to survey genes that escape XCI in vivo. We show that escape from X inactivation can be a common feature of some genes, whereas others escape in a tissue specific manner. Furthermore, we characterize the chromatin environment of escape genes and show that expression from the Xi correlates with factors associated with open chromatin and that CTCF co-localizes with escape genes. Here, we provide a detailed description of the experimental design and data analysis pipeline we used to assay allele-specific expression and epigenetic characteristics of genes escaping X inactivation. The data is publicly available through the GEO database under ascension numbers GSM1014171, GSE44255, and GSE59779. Interpretation and discussion of these data are included in a previously published study (Berletch et al., 2015) [1].

  1. IDP-ASE: haplotyping and quantifying allele-specific expression at the gene and gene isoform level by hybrid sequencing.

    PubMed

    Deonovic, Benjamin; Wang, Yunhao; Weirather, Jason; Wang, Xiu-Jie; Au, Kin Fai

    2016-11-28

    Allele-specific expression (ASE) is a fundamental problem in studying gene regulation and diploid transcriptome profiles, with two key challenges: (i) haplotyping and (ii) estimation of ASE at the gene isoform level. Existing ASE analysis methods are limited by a dependence on haplotyping from laborious experiments or extra genome/family trio data. In addition, there is a lack of methods for gene isoform level ASE analysis. We developed a tool, IDP-ASE, for full ASE analysis. By innovative integration of Third Generation Sequencing (TGS) long reads with Second Generation Sequencing (SGS) short reads, the accuracy of haplotyping and ASE quantification at the gene and gene isoform level was greatly improved as demonstrated by the gold standard data GM12878 data and semi-simulation data. In addition to methodology development, applications of IDP-ASE to human embryonic stem cells and breast cancer cells indicate that the imbalance of ASE and non-uniformity of gene isoform ASE is widespread, including tumorigenesis relevant genes and pluripotency markers. These results show that gene isoform expression and allele-specific expression cooperate to provide high diversity and complexity of gene regulation and expression, highlighting the importance of studying ASE at the gene isoform level. Our study provides a robust bioinformatics solution to understand ASE using RNA sequencing data only.

  2. Quantification of the paternal allele bias for new germline mutations in the retinoblastoma gene

    SciTech Connect

    Morrow, J.F.; Rapaport, J.M.; Dryia, T.P.

    1994-09-01

    New germline mutations in the human retinoblastoma gene preferentially arise on a paternally derived allele. In nonhereditary retinoblastoma, the initial somatic mutation seems to have no such bias. The few previous reports of these phenomena included relatively few cases (less than a dozen new germline or initial somatic mutations), so that the magnitude of the paternal allele bias for new germline mutations is not known. Knowledge of the magnitude of the bias is valuable for genetic counseling, since, for example, patients with new germline mutations who reproduce transmit risk for retinoblastoma according to the risk that the transmitted allele has a germline mutation. We sought to quantitate the paternal allele bias and to determine whether paternal age is a factor possibly accounting for it. We studied 311 families with retinoblastoma (261 simplex, 50 multiplex) that underwent clinical genetic testing and 5 informative families recruited from earlier research. Using RFLPs and polymorphic microsatellites in the retinoblastoma gene, we could determine the parental origin of 45 new germline mutations and 44 probable initial somatic mutations. Thirty-seven of the 45 new germline mutations, or 82%, arose on a paternal allele while only 24 of the 44 initial somatic mutations (55%) did so. Increased paternal age does not appear to account for the excess of new paternal germline mutations, since the average age of fathers of children with new germline mutations (29.4 years, n=26, incomplete records on 11) was not significantly different from the average age of fathers of children with maternal germline mutations or somatic initial mutations (29.8 years, n=35, incomplete records on 17).

  3. Epidemiological and Evolutionary Outcomes in Gene-for-Gene and Matching Allele Models

    PubMed Central

    Thrall, Peter H.; Barrett, Luke G.; Dodds, Peter N.; Burdon, Jeremy J.

    2016-01-01

    Gene-for-gene (GFG) and matching-allele (MA) models are qualitatively different paradigms for describing the outcome of genetic interactions between hosts and pathogens. The GFG paradigm was largely built on the foundations of Flor’s early work on the flax–flax rust interaction and is based on the concept of genetic recognition leading to incompatible disease outcomes, typical of host immune recognition. In contrast, the MA model is based on the assumption that genetic recognition leads to compatible interactions, which can result when pathogens require specific host factors to cause infection. Results from classical MA and GFG models have led to important predictions regarding various coevolutionary phenomena, including the role of fitness costs associated with resistance and infectivity, the distribution of resistance genes in wild populations, patterns of local adaptation and the evolution and maintenance of sexual reproduction. Empirical evidence (which we review briefly here), particularly from recent molecular advances in understanding of the mechanisms that determine the outcome of host–pathogen encounters, suggests considerable variation in specific details of the functioning of interactions between hosts and pathogens, which may contain elements of both models. In this regard, GFG and MA scenarios likely represent endpoints of a continuum of potentially more complex interactions that occur in nature. Increasingly, this has been recognized in theoretical studies of coevolutionary processes in plant host–pathogen and animal host-parasite associations (e.g., departures from strict GFG/MA assumptions, diploid genetics, multi-step infection processes). However, few studies have explored how different genetic assumptions about host resistance and pathogen infectivity might impact on disease epidemiology or pathogen persistence within and among populations. Here, we use spatially explicit simulations of the basic MA and GFG scenarios to highlight

  4. Blast resistance of CC-NB-LRR protein Pb1 is mediated by WRKY45 through protein-protein interaction.

    PubMed

    Inoue, Haruhiko; Hayashi, Nagao; Matsushita, Akane; Xinqiong, Liu; Nakayama, Akira; Sugano, Shoji; Jiang, Chang-Jie; Takatsuji, Hiroshi

    2013-06-04

    Panicle blast 1 (Pb1) is a panicle blast resistance gene derived from the indica rice cultivar "Modan." Pb1 encodes a coiled-coil-nucleotide-binding site-leucine-rich repeat (CC-NB-LRR) protein and confers durable, broad-spectrum resistance to Magnaporthe oryzae races. Here, we investigated the molecular mechanisms underlying Pb1-mediated blast resistance. The Pb1 protein interacted with WRKY45, a transcription factor involved in induced resistance via the salicylic acid signaling pathway that is regulated by the ubiquitin proteasome system. Pb1-mediated panicle blast resistance was largely compromised when WRKY45 was knocked down in a Pb1-containing rice cultivar. Leaf-blast resistance by Pb1 overexpression (Pb1-ox) was also compromised in WRKY45 knockdown/Pb1-ox rice. Blast infection induced higher accumulation of WRKY45 in Pb1-ox than in control Nipponbare rice. Overexpression of Pb1-Quad, a coiled-coil domain mutant that had weak interaction with WRKY45, resulted in significantly weaker blast resistance than that of wild-type Pb1. Overexpression of Pb1 with a nuclear export sequence failed to confer blast resistance to rice. These results suggest that the blast resistance of Pb1 depends on its interaction with WRKY45 in the nucleus. In a transient system using rice protoplasts, coexpression of Pb1 enhanced WRKY45 accumulation and increased WRKY45-dependent transactivation activity, suggesting that protection of WRKY45 from ubiquitin proteasome system degradation is possibly involved in Pb1-dependent blast resistance.

  5. Distribution of allelic variants of the chromosomal gene bla OXA-114-like in Achromobacter xylosoxidans clinical isolates.

    PubMed

    Traglia, German Matías; Almuzara, Marisa; Merkier, Andrea Karina; Papalia, Mariana; Galanternik, Laura; Radice, Marcela; Vay, Carlos; Centrón, Daniela; Ramírez, María Soledad

    2013-11-01

    Achromobacter xylosoxidans is increasingly being documented in cystic fibrosis patients. The bla(OXA-114) gene has been recognized as a naturally occurring chromosomal gene, exhibiting different allelic variants. In the population under study, the bla(OXA-114)-like gene was found in 19/19 non-epidemiological-related clinical isolates of A. xylosoxidans with ten different alleles including 1 novel OXA-114 variant.

  6. Detection, Validation, and Downstream Analysis of Allelic Variation in Gene Expression

    PubMed Central

    Ciobanu, Daniel C.; Lu, Lu; Mozhui, Khyobeni; Wang, Xusheng; Jagalur, Manjunatha; Morris, John A.; Taylor, William L.; Dietz, Klaus; Simon, Perikles; Williams, Robert W.

    2010-01-01

    Common sequence variants within a gene often generate important differences in expression of corresponding mRNAs. This high level of local (allelic) control—or cis modulation—rivals that produced by gene targeting, but expression is titrated finely over a range of levels. We are interested in exploiting this allelic variation to study gene function and downstream consequences of differences in expression dosage. We have used several bioinformatics and molecular approaches to estimate error rates in the discovery of cis modulation and to analyze some of the biological and technical confounds that contribute to the variation in gene expression profiling. Our analysis of SNPs and alternative transcripts, combined with eQTL maps and selective gene resequencing, revealed that between 17 and 25% of apparent cis modulation is caused by SNPs that overlap probes rather than by genuine quantitative differences in mRNA levels. This estimate climbs to 40–50% when qualitative differences between isoform variants are included. We have developed an analytical approach to filter differences in expression and improve the yield of genuine cis-modulated transcripts to ∼80%. This improvement is important because the resulting variation can be successfully used to study downstream consequences of altered expression on higher-order phenotypes. Using a systems genetics approach we show that two validated cis-modulated genes, Stk25 and Rasd2, are likely to control expression of downstream targets and affect disease susceptibility. PMID:19884314

  7. A systems biology perspective on the role of WRKY transcription factors in drought responses in plants.

    PubMed

    Tripathi, Prateek; Rabara, Roel C; Rushton, Paul J

    2014-02-01

    Drought is one of the major challenges affecting crop productivity and yield. However, water stress responses are notoriously multigenic and quantitative with strong environmental effects on phenotypes. It is also clear that water stress often does not occur alone under field conditions but rather in conjunction with other abiotic stresses such as high temperature and high light intensities. A multidisciplinary approach with successful integration of a whole range of -omics technologies will not only define the system, but also provide new gene targets for both transgenic approaches and marker-assisted selection. Transcription factors are major players in water stress signaling and some constitute major hubs in the signaling webs. The main transcription factors in this network include MYB, bHLH, bZIP, ERF, NAC, and WRKY transcription factors. The role of WRKY transcription factors in abiotic stress signaling networks is just becoming apparent and systems biology approaches are starting to define their places in the signaling network. Using systems biology approaches, there are now many transcriptomic analyses and promoter analyses that concern WRKY transcription factors. In addition, reports on nuclear proteomics have identified WRKY proteins that are up-regulated at the protein level by water stress. Interactomics has started to identify different classes of WRKY-interacting proteins. What are often lacking are connections between metabolomics, WRKY transcription factors, promoters, biosynthetic pathways, fluxes and downstream responses. As more levels of the system are characterized, a more detailed understanding of the roles of WRKY transcription factors in drought responses in crops will be obtained.

  8. The transcription factor VvWRKY33 is involved in the regulation of grapevine (Vitis vinifera) defense against the oomycete pathogen Plasmopara viticola.

    PubMed

    Merz, Patrick R; Moser, Tina; Höll, Janine; Kortekamp, Andreas; Buchholz, Günther; Zyprian, Eva; Bogs, Jochen

    2015-03-01

    Grapevine (Vitis vinifera ssp. vinifera) is one of the most important fruit species; however, it is highly susceptible to various pathogens, which can cause severe crop losses in viticulture. It has been shown that several WRKY class transcription factors (TFs) are part of the signal transduction cascade, which leads to the activation of plant defense reactions against various pathogens. In the present investigation, a full-length cDNA was isolated from V. vinifera leaf tissue encoding a predicted protein, designated VvWRKY33, which shows the characteristics of group I WRKY protein family. VvWRKY33 induction correlates with the expression of VvPR10.1 (pathogenesis-related 10.1) gene in the leaves of the resistant cultivar 'Regent' after infection with Plasmopara viticola, whereas in the susceptible cultivar 'Lemberger' VvWRKY33 and VvPR10.1 are not induced. Corresponding expression of the TF and VvPR10.1 was even obtained in uninfected ripening berries. In planta, analysis of VvWRKY33 has been performed by ectopic expression of VvWRKY33 in grapevine leaves of greenhouse plants mediated via Agrobacterium tumefaciens transformation. In consequence, VvWRKY33 strongly increases resistance to P. viticola in the susceptible cultivar 'Shiraz' and reduces pathogen sporulation of about 50-70%, indicating a functional role for resistance in grapevine. Complementation of the resistance-deficient Arabidopsis thaliana Columbia-0 (Col-0) mutant line wrky33-1 by constitutive expression of VvWRKY33 restores resistance against Botrytis cinerea to wild-type level and in some complemented mutant lines even exceeds the resistance level of the parental line Col-0. Our results support the involvement of VvWRKY33 in the defense reaction of grapevine against different pathogens.

  9. Patterns of human genetic variation inferred from comparative analysis of allelic mutations in blood group antigen genes.

    PubMed

    Patnaik, Santosh Kumar; Blumenfeld, Olga O

    2011-03-01

    Comparative analysis of allelic variation of a gene sheds light on the pattern and process of its diversification at the population level. Gene families for which a large number of allelic forms have been verified by sequencing provide a useful resource for such studies. In this regard, human blood group-encoding genes are unique in that differences of cell surface traits among individuals and populations can be readily detected by serological screening, and correlation between the variant cell surface phenotype and the genotype is, in most cases, unequivocal. Here, we perform a comprehensive analysis of allelic forms, compiled in the Blood Group Antigen Gene Mutation database, of ABO, RHD/CE, GYPA/B/E and FUT1/2 gene families that encode the ABO, RH, MNS, and H/h blood group system antigens, respectively. These genes are excellent illustrative examples showing distinct mutational patterns among the alleles, and leading to speculation on how their origin may have been driven by recurrent but different molecular mechanisms. We illustrate how alignment of alleles of a gene may provide an additional insight into the DNA variation process and its pathways, and how this approach may serve to catalog alleles of a gene, simplifying the task and content of mutation databases.

  10. Allelic diversity in an NLR gene BPH9 enables rice to combat planthopper variation

    PubMed Central

    Zhao, Yan; Huang, Jin; Wang, Zhizheng; Jing, Shengli; Wang, Yang; Ouyang, Yidan; Cai, Baodong; Xin, Xiu-Fang; Liu, Xin; Zhang, Chunxiao; Pan, Yufang; Ma, Rui; Li, Qiaofeng; Jiang, Weihua; Zeng, Ya; Shangguan, Xinxin; Wang, Huiying; Du, Bo; Zhu, Lili; Xu, Xun; Feng, Yu-Qi; He, Sheng Yang; Chen, Rongzhi; Zhang, Qifa; He, Guangcun

    2016-01-01

    Brown planthopper (BPH), Nilaparvata lugens Stål, is one of the most devastating insect pests of rice (Oryza sativa L.). Currently, 30 BPH-resistance genes have been genetically defined, most of which are clustered on specific chromosome regions. Here, we describe molecular cloning and characterization of a BPH-resistance gene, BPH9, mapped on the long arm of rice chromosome 12 (12L). BPH9 encodes a rare type of nucleotide-binding and leucine-rich repeat (NLR)-containing protein that localizes to the endomembrane system and causes a cell death phenotype. BPH9 activates salicylic acid- and jasmonic acid-signaling pathways in rice plants and confers both antixenosis and antibiosis to BPH. We further demonstrated that the eight BPH-resistance genes that are clustered on chromosome 12L, including the widely used BPH1, are allelic with each other. To honor the priority in the literature, we thus designated this locus as BPH1/9. These eight genes can be classified into four allelotypes, BPH1/9-1, -2, -7, and -9. These allelotypes confer varying levels of resistance to different biotypes of BPH. The coding region of BPH1/9 shows a high level of diversity in rice germplasm. Homologous fragments of the nucleotide-binding (NB) and leucine-rich repeat (LRR) domains exist, which might have served as a repository for generating allele diversity. Our findings reveal a rice plant strategy for modifying the genetic information to gain the upper hand in the struggle against insect herbivores. Further exploration of natural allelic variation and artificial shuffling within this gene may allow breeding to be tailored to control emerging biotypes of BPH. PMID:27791169

  11. Characterization of 12 silent alleles of the human butyrylcholinesterase (BCHE) gene.

    PubMed Central

    Primo-Parmo, S. L.; Bartels, C. F.; Wiersema, B.; van der Spek, A. F.; Innis, J. W.; La Du, B. N.

    1996-01-01

    The silent phenotype of human butyrylcholinesterase (BChE), present in most human populations in frequencies of approximately 1/100,000, is characterized by the complete absence of BChE activity or by activity <10% of the average levels of the usual phenotype. Heterogeneity in this phenotype has been well established at the phenotypic level, but only a few silent BCHE alleles have been characterized at the DNA level. Twelve silent alleles of the human butyrylcholinesterase gene (BCHE) have been identified in 17 apparently unrelated patients who were selected by their increased sensitivity to the muscle relaxant succinylcholine. All of these alleles are characterized by single nucleotide substitutions or deletions leading to distinct changes in the structure of the BChE enzyme molecule. Nine of the nucleotide substitutions result in the replacement of single amino acid residues. Three of these variants, BCHE*33C, BCHE*198G, and BCHE*201T, produce normal amounts of immunoreactive but enzymatically inactive BChE protein in the plasma. The other six amino acid substitutions, encoded by BCHE*37S, BCHE*125F, BCHE*170E, BCHE*471R, and BCHE*518L, seem to cause reduced expression of BChE protein, and their role in determining the silent phenotype was confirmed by expression in cell culture. The other four silent alleles, BCHE*271STOP, BCHE*500STOP, BCHE*FS6, and BCHE*I2E3-8G, encode BChES truncated at their C-terminus because of premature stop codons caused by nucleotide substitutions, a frame shift, or altered splicing. The large number of different silent BCHE alleles found within a relatively small number of patients shows that the heterogeneity of the silent BChE phenotype is high. The characterization of silent BChE variants will be useful in the study of the structure/function relationship for this and other closely related enzymes. Images Figure 2 PMID:8554068

  12. Hybrid male sterility in rice controlled by interaction between divergent alleles of two adjacent genes.

    PubMed

    Long, Yunming; Zhao, Lifeng; Niu, Baixiao; Su, Jing; Wu, Hao; Chen, Yuanling; Zhang, Qunyu; Guo, Jingxin; Zhuang, Chuxiong; Mei, Mantong; Xia, Jixing; Wang, Lan; Wu, Haibin; Liu, Yao-Guang

    2008-12-02

    Sterility is common in hybrids between divergent populations, such as the indica and japonica subspecies of Asian cultivated rice (Oryza sativa). Although multiple loci for plant hybrid sterility have been identified, it remains unknown how alleles of the loci interact at the molecular level. Here we show that a locus for indica-japonica hybrid male sterility, Sa, comprises two adjacent genes, SaM and SaF, encoding a small ubiquitin-like modifier E3 ligase-like protein and an F-box protein, respectively. Most indica cultivars contain a haplotype SaM(+)SaF(+), whereas all japonica cultivars have SaM(-)SaF(-) that diverged by nucleotide variations in wild rice. Male semi-sterility in this heterozygous complex locus is caused by abortion of pollen carrying SaM(-). This allele-specific gamete elimination results from a selective interaction of SaF(+) with SaM(-), a truncated protein, but not with SaM(+) because of the presence of an inhibitory domain, although SaM(+) is required for this male sterility. Lack of any one of the three alleles in recombinant plants does not produce male sterility. We propose a two-gene/three-component interaction model for this hybrid male sterility system. The findings have implications for overcoming male sterility in inter-subspecific hybrid rice breeding.

  13. Ancient allelism at the cytosolic chaperonin-alpha-encoding gene of the zebrafish.

    PubMed Central

    Takami, K; Figueroa, F; Mayer, W E; Klein, J

    2000-01-01

    The T-complex protein 1, TCP1, gene codes for the CCT-alpha subunit of the group II chaperonins. The gene was first described in the house mouse, in which it is closely linked to the T locus at a distance of approximately 11 cM from the Mhc. In the zebrafish, Danio rerio, in which the T homolog is linked to the class I Mhc loci, the TCP1 locus segregates independently of both the T and the Mhc loci. Despite its conservation between species, the zebrafish TCP1 locus is highly polymorphic. In a sample of 15 individuals and the screening of a cDNA library, 12 different alleles were found, and some of the allelic pairs were found to differ by up to nine nucleotides in a 275-bp-long stretch of sequence. The substitutions occur in both translated and untranslated regions, but in the former they occur predominantly at synonymous codon sites. Phylogenetically, the alleles fall into two groups distinguished also by the presence or absence of a 10-bp insertion/deletion in the 3' untranslated region. The two groups may have diverged as long as 3.5 mya, and the polymorphic differences may have accumulated by genetic drift in geographically isolated populations. PMID:10628990

  14. GmWRKY27 interacts with GmMYB174 to reduce expression of GmNAC29 for stress tolerance in soybean plants.

    PubMed

    Wang, Fang; Chen, Hao-Wei; Li, Qing-Tian; Wei, Wei; Li, Wei; Zhang, Wan-Ke; Ma, Biao; Bi, Ying-Dong; Lai, Yong-Cai; Liu, Xin-Lei; Man, Wei-Qun; Zhang, Jin-Song; Chen, Shou-Yi

    2015-07-01

    Soybean (Glycine max) is an important crop for oil and protein resources worldwide. The molecular mechanism of the abiotic stress response in soybean is largely unclear. We previously identified multiple stress-responsive WRKY genes from soybean. Here, we further characterized the roles of one of these genes, GmWRKY27, in abiotic stress tolerance using a transgenic hairy root assay. GmWRKY27 expression was increased by various abiotic stresses. Over-expression and RNAi analysis demonstrated that GmWRKY27 improves salt and drought tolerance in transgenic soybean hairy roots. Measurement of physiological parameters, including reactive oxygen species and proline contents, supported this conclusion. GmWRKY27 inhibits expression of a downstream gene GmNAC29 by binding to the W-boxes in its promoter region. The GmNAC29 is a negative factor of stress tolerance as indicated by the performance of transgenic hairy roots under stress. GmWRKY27 interacts with GmMYB174, which also suppresses GmNAC29 expression and enhances drought stress tolerance. The GmWRKY27 and GmMYB174 may have evolved to bind to neighbouring cis elements in the GmNAC29 promoter to co-reduce promoter activity and gene expression. Our study discloses a valuable mechanism in soybean for regulation of the stress response by two associated transcription factors. Manipulation of these genes should facilitate improvements in stress tolerance in soybean and other crops.

  15. Systematic morphological profiling of human gene and allele function via Cell Painting.

    PubMed

    Rohban, Mohammad Hossein; Singh, Shantanu; Wu, Xiaoyun; Berthet, Julia B; Bray, Mark-Anthony; Shrestha, Yashaswi; Varelas, Xaralabos; Boehm, Jesse S; Carpenter, Anne E

    2017-03-18

    We hypothesized that human genes and disease-associated alleles might be systematically functionally annotated using morphological profiling of cDNA constructs, via a microscopy-based Cell Painting assay. Indeed, 50% of the 220 tested genes yielded detectable morphological profiles, which grouped into biologically meaningful gene clusters consistent with known functional annotation (e.g., the RAS-RAF-MEK-ERK cascade). We used novel subpopulation-based visualization methods to interpret the morphological changes for specific clusters. This unbiased morphologic map of gene function revealed TRAF2/c-REL negative regulation of YAP1/WWTR1-responsive pathways. We confirmed this discovery of functional connectivity between the NF-κB pathway and Hippo pathway effectors at the transcriptional level, thereby expanding knowledge of these two signaling pathways that critically regulate tumor initiation and progression. We make the images and raw data publicly available, providing an initial morphological map of major biological pathways for future study.

  16. The Arg160Trp allele of melanocortin-1 receptor gene might protect against vitiligo.

    PubMed

    Széll, Márta; Baltás, Eszter; Bodai, László; Bata-Csörgo, Zsuzsanna; Nagy, Nikoletta; Dallos, Attila; Pourfarzi, Reza; Simics, Eniko; Kondorosi, Ildikó; Szalai, Zsuzsanna; Tóth, Gábor K; Hunyadi, János; Dobozy, Attila; Kemény, Lajos

    2008-01-01

    Melanocortin-1 receptor (MC1R) and agouti signaling protein (ASIP) play pivotal roles in the regulation of human pigmentation. We aimed to study whether single nucleotide polymorphisms (SNPs) of the MC1R and ASIP genes contribute to the pathogenesis of the polygenic pigment skin disorder, vitiligo. The PCR-amplified, full-length MC1R gene was studied with sequence analysis, and the 3' untranslated region (3' UTR) SNP of ASIP was detected using restriction fragment length polymorphism. The allele frequency of the ASIP SNP did not show any difference between the skin type, hair color and eye color-matched 97 vitiligo patients and the 59 healthy control individuals. As one of the MC1R polymorphisms showed significantly higher incidence among fair-skinned individuals (Fitzpatrick I+II, n=140) than among dark-skinned individuals (Fitzpatrick III+IV, n=90), both vitiligo patients and controls were divided into two groups and the frequency of the MC1R alleles was studied separately in fair-skinned and dark-skinned subgroups of diseased and healthy groups. C478T, one of the MC1R SNPs studied in 108 fair-skinned vitiligo patients and in 70 fair-skinned healthy control individuals, showed a significant difference (P=0.0262, odds ratio [95% confidence interval]=3.6 [0.0046-0.1003]) in allele frequency between the two groups: the allele frequency was higher in the control group, suggesting protection against vitiligo. Computer prediction of antigenicity has revealed that the Arg160Trp amino acid change caused by this SNP results in a decrease in antigenicity of the affected peptide epitope.

  17. Efficient CRISPR-rAAV engineering of endogenous genes to study protein function by allele-specific RNAi.

    PubMed

    Kaulich, Manuel; Lee, Yeon J; Lönn, Peter; Springer, Aaron D; Meade, Bryan R; Dowdy, Steven F

    2015-04-20

    Gene knockout strategies, RNAi and rescue experiments are all employed to study mammalian gene function. However, the disadvantages of these approaches include: loss of function adaptation, reduced viability and gene overexpression that rarely matches endogenous levels. Here, we developed an endogenous gene knockdown/rescue strategy that combines RNAi selectivity with a highly efficient CRISPR directed recombinant Adeno-Associated Virus (rAAV) mediated gene targeting approach to introduce allele-specific mutations plus an allele-selective siRNA Sensitive (siSN) site that allows for studying gene mutations while maintaining endogenous expression and regulation of the gene of interest. CRISPR/Cas9 plus rAAV targeted gene-replacement and introduction of allele-specific RNAi sensitivity mutations in the CDK2 and CDK1 genes resulted in a >85% site-specific recombination of Neo-resistant clones versus ∼8% for rAAV alone. RNAi knockdown of wild type (WT) Cdk2 with siWT in heterozygotic knockin cells resulted in the mutant Cdk2 phenotype cell cycle arrest, whereas allele specific knockdown of mutant CDK2 with siSN resulted in a wild type phenotype. Together, these observations demonstrate the ability of CRISPR plus rAAV to efficiently recombine a genomic locus and tag it with a selective siRNA sequence that allows for allele-selective phenotypic assays of the gene of interest while it remains expressed and regulated under endogenous control mechanisms.

  18. Meiotic recombination generates rich diversity in NK cell receptor genes, alleles, and haplotypes

    PubMed Central

    Norman, Paul J.; Abi-Rached, Laurent; Gendzekhadze, Ketevan; Hammond, John A.; Moesta, Achim K.; Sharma, Deepti; Graef, Thorsten; McQueen, Karina L.; Guethlein, Lisbeth A.; Carrington, Christine V.F.; Chandanayingyong, Dasdayanee; Chang, Yih-Hsin; Crespí, Catalina; Saruhan-Direskeneli, Güher; Hameed, Kamran; Kamkamidze, Giorgi; Koram, Kwadwo A.; Layrisse, Zulay; Matamoros, Nuria; Milà, Joan; Park, Myoung Hee; Pitchappan, Ramasamy M.; Ramdath, D. Dan; Shiau, Ming-Yuh; Stephens, Henry A.F.; Struik, Siske; Tyan, Dolly; Verity, David H.; Vaughan, Robert W.; Davis, Ronald W.; Fraser, Patricia A.; Riley, Eleanor M.; Ronaghi, Mostafa; Parham, Peter

    2009-01-01

    Natural killer (NK) cells contribute to the essential functions of innate immunity and reproduction. Various genes encode NK cell receptors that recognize the major histocompatibility complex (MHC) Class I molecules expressed by other cells. For primate NK cells, the killer-cell immunoglobulin-like receptors (KIR) are a variable and rapidly evolving family of MHC Class I receptors. Studied here is KIR3DL1/S1, which encodes receptors for highly polymorphic human HLA-A and -B and comprises three ancient allelic lineages that have been preserved by balancing selection throughout human evolution. While the 3DS1 lineage of activating receptors has been conserved, the two 3DL1 lineages of inhibitory receptors were diversified through inter-lineage recombination with each other and with 3DS1. Prominent targets for recombination were D0-domain polymorphisms, which modulate enhancer function, and dimorphism at position 283 in the D2 domain, which influences inhibitory function. In African populations, unequal crossing over between the 3DL1 and 3DL2 genes produced a deleted KIR haplotype in which the telomeric “half” was reduced to a single fusion gene with functional properties distinct from its 3DL1 and 3DL2 parents. Conversely, in Eurasian populations, duplication of the KIR3DL1/S1 locus by unequal crossing over has enabled individuals to carry and express alleles of all three KIR3DL1/S1 lineages. These results demonstrate how meiotic recombination combines with an ancient, preserved diversity to create new KIR phenotypes upon which natural selection acts. A consequence of such recombination is to blur the distinction between alleles and loci in the rapidly evolving human KIR gene family. PMID:19411600

  19. Allelic switching of the imprinted IGF2R gene in cloned bovine fetuses and calves.

    PubMed

    Suteevun-Phermthai, T; Curchoe, C L; Evans, A C; Boland, E; Rizos, D; Fair, T; Duffy, P; Sung, L Y; Du, F; Chaubal, S; Xu, J; Wechayant, T; Yang, X; Lonergan, P; Parnpai, R; Tian, X C

    2009-11-01

    Cloned animals often suffer from loss of development to term and abnormalities, typically classified under the umbrella term of Large Offspring Syndrome (LOS). Cattle are an interesting species to study because of the relatively greater success rate of nuclear transfer in this species compared with all species cloned to date. The imprinted insulin-like growth factor receptor (IGF2R; mannose-6-phosphate) gene was chosen to investigate aspects of fetal growth and development in cloned cattle in the present study. IGF2R gene expression patterns in identical genetic clones of several age groups were assessed in day 25, day 45, and day 75 fetuses as well as spontaneously aborted fetuses, calves that died shortly after birth and healthy cloned calves using single stranded conformational polymorphism gel electrophoresis. A variable pattern of IGF2R allelic expression in major organs such as the brain, cotyledon, heart, liver, lung, spleen, kidney and intercotyledon was observed using a G/A transition in the 3'UTR of IGF2R. IGF2R gene expression was also assessed by real time RT-PCR and found to be highly variable among the clone groups. Proper IGF2R gene expression is necessary for survival to term, but is most likely not a cause of early fetal lethality or an indicator of postnatal fitness. Contrary to previous reports of the transmission of imprinting patterns from somatic donor cells to cloned animals within organs in the same cloned animal the paternal allele of IGF2R can be imprinted in one tissue while the maternal allele is imprinted in another tissue. This observation has never been reported in any species in which imprinting has been studied.

  20. Selective Retention of an Inactive Allele of the DKK2 Tumor Suppressor Gene in Hepatocellular Carcinoma

    PubMed Central

    Lin, Yung-Feng; Li, Ling-Hui; Lin, Chih-Hung; Tsou, Mei-Hua; Chuang, Ming-Tai Kiffer; Wu, Keh-Ming; Liao, Tsai-Lien; Li, Jian-Chiuan; Wang, Wei-Jie; Tomita, Angela; Tomita, Beverly; Huang, Shiu-Feng; Tsai, Shih-Feng

    2016-01-01

    In an effort to identify the functional alleles associated with hepatocellular carcinoma (HCC), we investigated 152 genes found in the 4q21-25 region that exhibited loss of heterozygosity (LOH). A total of 2,293 pairs of primers were designed for 1,449 exonic and upstream promoter regions to amplify and sequence 76.8–114 Mb on human chromosome 4. Based on the results from analyzing 12 HCC patients and 12 healthy human controls, we discovered 1,574 sequence variations. Among the 99 variants associated with HCC (p < 0.05), four are from the Dickkopf 2 (DKK2) gene: three in the promoter region (g.-967A>T, g.-923C>A, and g.-441T>G) and one in the 5’UTR (c.550T>C). To verify the results, we expanded the subject cohort to 47 HCC cases and 88 healthy controls for conducting haplotype analysis. Eight haplotypes were detected in the non-tumor liver tissue samples, but one major haplotype (TAGC) was found in the tumor tissue samples. Using a reporter assay, this HCC-associated allele registered the lowest level of promoter activity among all the tested haplotype sequences. Retention of this allele in LOH was associated with reduced DKK2 transcription in the HCC tumor tissues. In HuH-7 cells, DKK2 functioned in the Wnt/β-catenin signaling pathway, as an antagonist of Wnt3a, in a dose-dependent manner that inhibited Wnt3a-induced cell proliferation. Taken together, the genotyping and functional findings are consistent with the hypothesis that DKK2 is a tumor suppressor; by selectively retaining a transcriptionally inactive DKK2 allele, the reduction of DKK2 function results in unchecked Wnt/β-catenin signaling, contributing to HCC oncogenesis. Thus our study reveals a new mechanism through which a tumor suppressor gene in a LOH region loses its function by allelic selection. PMID:27203079

  1. A note on the change in gene frequency of a selected allele in partial full-sib mating populations

    SciTech Connect

    Caballero, A.

    1996-02-01

    The change in gene frequency of a selected allele in partial full-sib mating populations was analyzed. The implications of these papers is important in terms of the fixation probability of genes because, for the same equilibrium inbreeding coefficient, fixation rates of mutant genes would be larger for partial full-sib mating than for partial selfing. 4 refs.

  2. Identification of Alleles of Puroindoline Genes and Their Effect on Wheat (Triticum aestivum L.) Grain Texture

    PubMed Central

    Štiasna, Klára; Vyhnánek, Tomáš; Trojan, Václav; Mrkvicová, Eva; Hřivna, Luděk; Havel, Ladislav

    2016-01-01

    Summary Grain hardness is one of the most important quality characteristics of wheat (Triticum aestivum L.). It is a significant property of wheat grains and relates to milling quality and end product quality. Grain hardness is caused by the presence of puroindoline genes (Pina and Pinb). A collection of 25 genotypes of wheat with unusual grain colour (blue aleurone, purple and white pericarp, yellow endosperm) was studied by polymerase chain reaction (PCR) for the diversity within Pina and Pinb (alleles: Pina-D1a, Pina-D1b, Pinb-D1a, Pinb- -D1b, Pinb-D1c and Pinb-D1d). The endosperm structure was determined by a non-destructive method using light transflectance meter and grain hardness by a texture analyser. Genotype Novosibirskaya 67 and isogenic ANK lines revealed hitherto unknown alleles at the locus for the annealing of primers of Pinb-D1. Allele Pinb-D1c was found to be absent from each genotype. The mealy endosperm ranged from 0 to 100% and grain hardness from 15.10 to 26.87 N per sample. PMID:27904399

  3. Structure and expression of wild-type and suppressible alleles of the Drosophila purple gene

    SciTech Connect

    Kim, Nacksung |; Park, Dongkook; Yim, John

    1996-04-01

    Viable mutant alleles of purple (pr), such as pr{sup bw}, exhibit mutant eye colors. This reflects low 6-pyruvoyl tetrahydropterin (PTP) synthase activity required for pigment synthesis. PTP synthase is also required for synthesis of the enzyme cofactor biopterin; presumably this is why some pr alleles are lethal. The pr{sup bw} eye color phenotype is suppressed by suppressor of sable [su(s)] mutations. The pr gene was cloned to explore the mechanism of this suppression. pr produces two PTP synthase mRNAs: one constitutively from a distal promoter and one in late pupae and young adult heads from a proximal promoter. The latter presumably supports eye pigment synthesis. The pr{sup bw} allele has a 412 retrotransposon in an intron spliced from both mRNAs. However, the head-specific mRNA is reduced > 10-fold in pr{sup bw} and is restored by a su(s) mutation, while the constitutive transcript is barely affected. The Su(s) protein probably alters processing of RNA containing 412. Because the intron containing 412 is the first in the head-specific mRNA and the second in the constitutive mRNA, binding of splicing machinery to nascent transcripts before the 412 insertion is transcribed may preclude the effects of Su(s) protein. 43 refs., 9 figs.

  4. Genetic influences on bone density: Physiological correlates of vitamin D receptor gene alleles in premonopausal women

    SciTech Connect

    Howard, G.; Nguyen, T.; Morrison, N.

    1995-09-01

    Common vitamin D receptor (VDR) gene alleles have recently been shown to contribute to the genetic variability in bone mass and bone turnover; however, the physiological mechanisms involved are unknown. To examine this, the response to 7 days of 2 {mu}g oral 1,25-dihydroxyvitamin D[1,25-(OH){sub 2}D] (calcitrol) stimulation was assessed in 21 premenopausal women, homozygous for one or other of the common VDR alleles (bb, N = 11; BB, n = 10). Indices of bone turnover and calcium homeostasis were measured during 2 weeks. Baseline osteocalcin, 1,25-(OH){sub 2}D, type I collagen carboxyterminal telopeptide, and inorganic phosphate levels were significantly higher and spinal bone mineral density was significantly lower in the BB allelic group. After calcitrol administration, similar levels of 1,25-(OH){sub 2}D were attained throughout the study in both genotypic groups. The increase in serum osteocalcin levels in the BB group was significantly less than that in the bb group (11% vs. 32%, P = 0.01). The genotype-related baseline difference in osteocalcin levels was not apparent at the similar serum 1,25-(OH){sub 2}D levels. By contrast, the baseline differences in phosphate and type I collagen carboxyterminal telopeptide persisted throughout the study. Serum ionized calcium levels did not differ between genotypes, nor did it move out of normal range values. However, parathyroid hormone was less suppressed in the low bone density group (38% vs. 11%, P = 0.01). These data indicate that the VDR alleles are associated with differences in the vitamin D endocrine system and may have important implications in relation to the pathophysiology of osteoporosis. 35 refs., 2 figs., 1 tab.

  5. Comprehensive analysis of imprinted genes in maize reveals allelic variation for imprinting and limited conservation with other species.

    PubMed

    Waters, Amanda J; Bilinski, Paul; Eichten, Steven R; Vaughn, Matthew W; Ross-Ibarra, Jeffrey; Gehring, Mary; Springer, Nathan M

    2013-11-26

    In plants, a subset of genes exhibit imprinting in endosperm tissue such that expression is primarily from the maternal or paternal allele. Imprinting may arise as a consequence of mechanisms for silencing of transposons during reproduction, and in some cases imprinted expression of particular genes may provide a selective advantage such that it is conserved across species. Separate mechanisms for the origin of imprinted expression patterns and maintenance of these patterns may result in substantial variation in the targets of imprinting in different species. Here we present deep sequencing of RNAs isolated from reciprocal crosses of four diverse maize genotypes, providing a comprehensive analysis that allows evaluation of imprinting at more than 95% of endosperm-expressed genes. We find that over 500 genes exhibit statistically significant parent-of-origin effects in maize endosperm tissue, but focused our analyses on a subset of these genes that had >90% expression from the maternal allele (69 genes) or from the paternal allele (108 genes) in at least one reciprocal cross. Over 10% of imprinted genes show evidence of allelic variation for imprinting. A comparison of imprinting in maize and rice reveals that 13% of genes with syntenic orthologs in both species exhibit conserved imprinting. Genes that exhibit conserved imprinting between maize and rice have elevated nonsynonymous to synonymous substitution ratios compared with other imprinted genes, suggesting a history of more rapid evolution. Together, these data suggest that imprinting only has functional relevance at a subset of loci that currently exhibit imprinting in maize.

  6. Effect of metallothionein 2A gene polymorphism on allele-specific gene expression and metal content in prostate cancer

    SciTech Connect

    Krześlak, Anna; Forma, Ewa; Jóźwiak, Paweł; Szymczyk, Agnieszka; Bryś, Magdalena

    2013-05-01

    Metallothioneins (MTs) are highly conserved, small molecular weight, cysteine rich proteins. The major physiological functions of metallothioneins include homeostasis of essential metals Zn and Cu and protection against cytotoxicity of heavy metals. The aim of this study was to determine whether there is an association between the − 5 A/G single nucleotide polymorphism (SNP; rs28366003) in core promoter region and expression of metallothionein 2A (MT2A) gene and metal concentration in prostate cancer tissues. MT2A polymorphism was determined by the polymerase chain reaction–restriction fragment length polymorphism technique (PCR–RFLP) using 412 prostate cancer tissue samples. MT2A gene expression analysis was performed by real-time RT-PCR method. A significant association between rs28366003 genotype and MT2A expression level was found. The average mRNA level was found to be lower among minor allele carriers (the risk allele) than average expression among homozygotes for the major allele. Metal levels were analyzed by flamed atomic absorption spectrometer system. Highly statistically significant associations were detected between the SNP and Cd, Zn, Cu and Pb levels. The results of Spearman's rank correlation showed that the expressions of MT2A and Cu, Pb and Ni concentrations were negatively correlated. On the basis of the results obtained in this study, we suggest that SNP polymorphism may affect the MT2A gene expression in prostate and this is associated with some metal accumulation. - Highlights: • MT2A gene expression and metal content in prostate cancer tissues • Association between SNP (rs28366003) and expression of MT2A • Significant associations between the SNP and Cd, Zn, Cu and Pb levels • Negative correlation between MT2A gene expression and Cu, Pb and Ni levels.

  7. The HLA-B*83:01 allele is generated by a gene conversion event including whole of exon 2 and partial introns 1 and 2 between B*44 and B*56 alleles.

    PubMed

    Cervera, I; Herraiz, M A; Vidart, J A; Peñaloza, J; Martinez-Laso, J

    2011-02-01

    Several studies have indicated the gene conversion as the most important mechanism about the MHC polymorphism generation when intron sequences are studied. The data obtained confirm that the B*83:01 allele is generated by gene conversion event including exon 2 and partial intron 1 and 2 between B*44 and B*56 alleles.

  8. Genome-wide screen of genes imprinted in sorghum endosperm, and the roles of allelic differential cytosine methylation.

    PubMed

    Zhang, Meishan; Li, Ning; He, Wenan; Zhang, Huakun; Yang, Wei; Liu, Bao

    2016-02-01

    Imprinting is an epigenetic phenomenon referring to allele-biased expression of certain genes depending on their parent of origin. Accumulated evidence suggests that, while imprinting is a conserved mechanism across kingdoms, the identities of the imprinted genes are largely species-specific. Using deep RNA sequencing of endosperm 14 days after pollination in sorghum, 5683 genes (29.27% of the total 19 418 expressed genes) were found to harbor diagnostic single nucleotide polymorphisms between two parental lines. The analysis of parent-of-origin expression patterns in the endosperm of a pair of reciprocal F1 hybrids between the two sorghum lines led to identification of 101 genes with ≥ fivefold allelic expression difference in both hybrids, including 85 maternal expressed genes (MEGs) and 16 paternal expressed genes (PEGs). Thirty of these genes were previously identified as imprinted in endosperm of maize (Zea mays), rice (Oryza sativa) or Arabidopsis, while the remaining 71 genes are sorghum-specific imprinted genes relative to these three plant species. Allele-biased expression of virtually all of the 14 tested imprinted genes (nine MEGs and five PEGs) was validated by pyrosequencing using independent sources of RNA from various developmental stages and dissected parts of endosperm. Forty-six imprinted genes (30 MEGs and 16 PEGs) were assayed by quantitative RT-PCR, and the majority of them showed endosperm-specific or preferential expression relative to embryo and other tissues. DNA methylation analysis of the 5' upstream region and gene body for seven imprinted genes indicated that, while three of the four PEGs were associated with hypomethylation of maternal alleles, no MEG was associated with allele-differential methylation.

  9. Self-recognition in social amoebae is mediated by allelic pairs of tiger genes.

    PubMed

    Hirose, Shigenori; Benabentos, Rocio; Ho, Hsing-I; Kuspa, Adam; Shaulsky, Gad

    2011-07-22

    Free-living cells of the social amoebae Dictyostelium discoideum can aggregate and develop into multicellular fruiting bodies in which many die altruistically as they become stalk cells that support the surviving spores. Dictyostelium cells exhibit kin discrimination--a potential defense against cheaters, which sporulate without contributing to the stalk. Kin discrimination depends on strain relatedness, and the polymorphic genes tgrB1 and tgrC1 are potential components of that mechanism. Here, we demonstrate a direct role for these genes in kin discrimination. We show that a matching pair of tgrB1 and tgrC1 alleles is necessary and sufficient for attractive self-recognition, which is mediated by differential cell-cell adhesion. We propose that TgrB1 and TgrC1 proteins mediate this adhesion through direct binding. This system is a genetically tractable ancient model of eukaryotic self-recognition.

  10. Geographically Distinct and Domain-Specific Sequence Variations in the Alleles of Rice Blast Resistance Gene Pib

    PubMed Central

    Vasudevan, Kumar; Vera Cruz, Casiana M.; Gruissem, Wilhelm; Bhullar, Navreet K.

    2016-01-01

    Rice blast is caused by Magnaporthe oryzae, which is the most destructive fungal pathogen affecting rice growing regions worldwide. The rice blast resistance gene Pib confers broad-spectrum resistance against Southeast Asian M. oryzae races. We investigated the allelic diversity of Pib in rice germplasm originating from 12 major rice growing countries. Twenty-five new Pib alleles were identified that have unique single nucleotide polymorphisms (SNPs), insertions and/or deletions, in addition to the polymorphic nucleotides that are shared between the different alleles. These partially or completely shared polymorphic nucleotides indicate frequent sequence exchange events between the Pib alleles. In some of the new Pib alleles, nucleotide diversity is high in the LRR domain, whereas, in others it is distributed among the NB-ARC and LRR domains. Most of the polymorphic amino acids in LRR and NB-ARC2 domains are predicted as solvent-exposed. Several of the alleles and the unique SNPs are country specific, suggesting a diversifying selection of alleles in various geographical locations in response to the locally prevalent M. oryzae population. Together, the new Pib alleles are an important genetic resource for rice blast resistance breeding programs and provide new information on rice-M. oryzae interactions at the molecular level. PMID:27446145

  11. Geographically Distinct and Domain-Specific Sequence Variations in the Alleles of Rice Blast Resistance Gene Pib.

    PubMed

    Vasudevan, Kumar; Vera Cruz, Casiana M; Gruissem, Wilhelm; Bhullar, Navreet K

    2016-01-01

    Rice blast is caused by Magnaporthe oryzae, which is the most destructive fungal pathogen affecting rice growing regions worldwide. The rice blast resistance gene Pib confers broad-spectrum resistance against Southeast Asian M. oryzae races. We investigated the allelic diversity of Pib in rice germplasm originating from 12 major rice growing countries. Twenty-five new Pib alleles were identified that have unique single nucleotide polymorphisms (SNPs), insertions and/or deletions, in addition to the polymorphic nucleotides that are shared between the different alleles. These partially or completely shared polymorphic nucleotides indicate frequent sequence exchange events between the Pib alleles. In some of the new Pib alleles, nucleotide diversity is high in the LRR domain, whereas, in others it is distributed among the NB-ARC and LRR domains. Most of the polymorphic amino acids in LRR and NB-ARC2 domains are predicted as solvent-exposed. Several of the alleles and the unique SNPs are country specific, suggesting a diversifying selection of alleles in various geographical locations in response to the locally prevalent M. oryzae population. Together, the new Pib alleles are an important genetic resource for rice blast resistance breeding programs and provide new information on rice-M. oryzae interactions at the molecular level.

  12. Molecular characterization of an allelic series of mutations in the mouse Nox3 gene.

    PubMed

    Flaherty, John P; Fairfield, Heather E; Spruce, Catrina A; McCarty, Christopher M; Bergstrom, David E

    2011-04-01

    The inner ear consists of the cochlea (the organ of hearing) and the vestibular system (the organs of balance). Within the vestibular system, linear acceleration and gravity are detected by the saccule and utricle. Resting above the neurosensory epithelia of these organs are otoconia, minute proteinaceous and crystalline (calcite) inertial masses that shift under the physical forces imparted by linear movements and gravity. It is the transduction and sensation of these movements and their integration with vision and proprioceptive inputs that contribute to the sensation of balance. It has been proposed that a reactive oxygen species- (ROS-) generating NADPH oxidase comprising the gene products of the Nox3, Noxo1, and Cyba genes plays a critical and constructive role in the process of inner-ear development, specifically, the deposition of otoconia. Inactivation in mouse of any of the NADPH oxidase components encoded by the Nox3, Noxo1, or Cyba gene results in the complete congenital absence of otoconia and profound vestibular dysfunction. Here we describe our use of PCR, reverse transcription-PCR (RT-PCR), and rapid amplification of cDNA ends (RACE) with traditional and high-throughput (HTP) sequencing technologies to extend and complete the molecular characterization of an allelic series of seven mutations in the Nox3 gene. Collectively, the mutation spectrum includes an endogenous retrovirus insertion, two missense mutations, a splice donor mutation, a splice acceptor mutation, premature translational termination, and a small duplication. Together, these alleles provide tools to investigate the mechanisms of otoconial deposition over development, throughout aging, and in various disease states.

  13. Molecular characterization of an allelic series of mutations in the mouse Nox3 gene

    PubMed Central

    Flaherty, John P.; Fairfield, Heather E.; Spruce, Catrina A.; McCarty, Christopher M.; Bergstrom, David E.

    2011-01-01

    The inner ear consists of the cochlea (the organ of hearing) and the vestibular system (the organs of balance). Within the vestibular system, linear acceleration and gravity are detected by the saccule and utricle. Resting above the neurosensory epithelia of these organs are otoconia, minute proteinaceous and crystalline (calcite) inertial masses that shift under the physical forces imparted by linear movements and gravity. It is the transduction and sensation of these movements and their integration with vision and proprioceptive inputs that contribute to the sensation of balance. It has been proposed that a reactive oxygen species- (ROS-) generating NADPH oxidase comprising the gene products of the Nox3, Noxo1, and Cyba genes plays a critical and constructive role in the process of inner-ear development, specifically, the deposition of otoconia. Inactivation in mouse of any of the NADPH oxidase components encoded by the Nox3, Noxo1, or Cyba gene results in the complete congenital absence of otoconia and profound vestibular dysfunction. Here we describe our use of PCR, reverse transcription-PCR (RT-PCR), and rapid amplification of cDNA ends (RACE) with traditional and high-throughput (HTP) sequencing technologies to extend and complete the molecular characterization of an allelic series of seven mutations in the Nox3 gene. Collectively, the mutation spectrum includes an endogenous retrovirus insertion, two missense mutations, a splice donor mutation, a splice acceptor mutation, premature translational termination, and a small duplication. Together, these alleles provide tools to investigate the mechanisms of otoconial deposition over development, throughout aging, and in various disease states. PMID:21161235

  14. Analysis of a Larger SNP Dataset from the HapMap Project Confirmed That the Modern Human A Allele of the ABO Blood Group Genes Is a Descendant of a Recombinant between B and O Alleles.

    PubMed

    Itou, Masaya; Sato, Mitsuharu; Kitano, Takashi

    2013-01-01

    The human ABO blood group gene consists of three main alleles (A, B, and O) that encode a glycosyltransferase. The A and B alleles differ by two critical amino acids in exon 7, and the major O allele has a single nucleotide deletion (Δ261) in exon 6. Previous evolutionary studies have revealed that the A allele is the most ancient, B allele diverged from the A allele with two critical amino acid substitutions in exon 7, and the major O allele diverged from the A allele with Δ261 in exon 6. However, a recent phylogenetic network analysis study showed that the A allele of humans emerged through a recombination between the B and O alleles. In the previous study, a restricted dataset from only two populations was used. In this study, therefore, we used a large single nucleotide polymorphism (SNP) dataset from the HapMap Project. The results indicated that the A101-A201-O09 haplogroup was a recombinant lineage between the B and O haplotypes, containing the intact exon 6 from the B allele and the two critical A type sites in exon 7 from the major O allele. Its recombination point was assumed to be located just behind Δ261 in exon 6.

  15. Rice WRKY4 acts as a transcriptional activator mediating defense responses toward Rhizoctonia solani, the causing agent of rice sheath blight.

    PubMed

    Wang, Haihua; Meng, Jiao; Peng, Xixu; Tang, Xinke; Zhou, Pinglan; Xiang, Jianhua; Deng, Xiaobo

    2015-09-01

    WRKY transcription factors have been implicated in the regulation of transcriptional reprogramming associated with various plant processes but most notably with plant defense responses to pathogens. Here we demonstrate that expression of rice WRKY4 gene (OsWRKY4) was rapidly and strongly induced upon infection of Rhizoctonia solani, the causing agent of rice sheath blight, and exogenous jasmonic acid (JA) and ethylene (ET). OsWRKY4 is localized to the nucleus of plant cells and possesses transcriptional activation ability. Modulation of OsWRKY4 transcript levels by constitutive overexpression increases resistance to the necrotrophic sheath blight fungus, concomitant with elevated expression of JA- and ET-responsive pathogenesis-related (PR) genes such as PR1a, PR1b, PR5 and PR10/PBZ1. Suppression by RNA interference (RNAi), on the other hand, compromises resistance to the fungal pathogen. Yeast one-hybrid assay and transient expression in tobacco cells reveal that OsWRKY4 specifically binds to the promoter regions of PR1b and PR5 which contain W-box (TTGAC[C/T]), or W-box like (TGAC[C/T]) cis-elements. In conclusion, we propose that OsWRKY4 functions as an important positive regulator that is implicated in the defense responses to rice sheath blight via JA/ET-dependent signal pathway.

  16. Phylogenetic analysis of six WRKY transcription factor loci across the spiny cocosoid palm subtribes Bactridinae and Elaeidinae (Areceaceae, Cocoseae),and comparison of several gene tree/species tree reconciliation approaches

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The Cocoseae is one of 13 tribes of Arecaceae subfamily Arecoideae, and contains a number of palms with significant economic importance, including the monotypic and pantropical Cocos nucifera, the coconut, and African oil palm (Elaeis guineensis). Using seven single copy WRKY transcription factor g...

  17. A hypervariable STR polymorphism in the CFI gene: southern origin of East Asian-specific group H alleles.

    PubMed

    Yuasa, Isao; Jin, Feng; Harihara, Shinji; Matsusue, Aya; Fujihara, Junko; Takeshita, Haruo; Akane, Atsushi; Umetsu, Kazuo; Saitou, Naruya; Chattopadhyay, Prasanta K

    2013-09-01

    Previous studies of four populations revealed that a hypervariable short tandem repeat (iSTR) in intron 7 of the human complement factor I (CFI) gene on chromosome 4q was unique, with 17 possible East Asian-specific group H alleles observed at relatively high frequencies. To develop a deeper anthropological and forensic understanding of iSTR, 1161 additional individuals from 11 Asian populations were investigated. Group H alleles of iSTR and c.1217A allele of a SNP in exon 11 of the CFI gene were associated with each other and were almost entirely confined to East Asian populations. Han Chinese in Changsha, southern China, showed the highest frequency for East Asian-specific group H alleles (0.201) among 15 populations. Group H alleles were observed to decrease gradually from south to north in 11 East Asian populations. This expansion of group H alleles provides evidence that southern China and Southeast Asia are a hotspot of Asian diversity and a genetic reservoir of Asians after they entered East Asia. The expected heterozygosity values of iSTR ranged from 0.927 in Thais to 0.874 in Oroqens, higher than those of an STR in the fibrinogen alpha chain (FGA) gene on chromosome 4q. Thus, iSTR is a useful marker for anthropological and forensic genetics.

  18. Allelic diversity of a beer haze active protein gene in cultivated and Tibetan wild barley and development of allelic specific markers.

    PubMed

    Ye, Lingzhen; Dai, Fei; Qiu, Long; Sun, Dongfa; Zhang, Guoping

    2011-07-13

    The formation of haze is a serious quality problem in beer production. It has been shown that the use of silica elute (SE)-ve malt (absence of molecular weight (MW) ∼14000 Da) for brewing can improve haze stability in the resultant beer, and the protein was identified as a barley trypsin inhibitor of the chloroform/methanol type (BTI-CMe). The objectives of this study were to determine (1) the allelic diversity of the gene controlling BTI-CMe in cultivated and Tibetan wild barley and (2) allele-specific (AS) markers for screening SE protein type. A survey of 172 Tibetan annual wild barley accessions and 71 cultivated barley genotypes was conducted, and 104 wild accessions and 35 cultivated genotypes were identified as SE+ve and 68 wild accessions and 36 cultivated genotypes as SE-ve. The allelic diversity of the gene controlling BTI-CMe was investigated by cloning, alignment, and association analysis. It was found that there were significant differences between the SE+ve and SE-ve types in single-nucleotide polymorphisms at 234 (SNP(234)), SNP(313), and SNP(385.) Furthermore, two sets of AS markers were developed to screen SE protein type based on SNP(313). AS-PCR had results very similar to those obtained by immunoblot method. Mapping analysis showed that the gene controlling the MW∼14 kDa band was located on the short arm of chromosome 3H, at the position of marker BPB-0527 (33.302 cM) in the Franklin/Yerong DH population.

  19. Allelic polymorphism in IL-1 beta and IL-1 receptor antagonist (IL-1Ra) genes in inflammatory bowel disease.

    PubMed Central

    Bioque, G; Crusius, J B; Koutroubakis, I; Bouma, G; Kostense, P J; Meuwissen, S G; Peña, A S

    1995-01-01

    Recent reports have shown that allele 2 of the IL-1 receptor antagonist (IL-1Ra) gene is over-represented in ulcerative colitis (UC). Healthy individuals carrying allele 2 of this gene have increased production of IL-1Ra protein. Since the final outcome of the biological effects of IL-1 beta may depend on the relative proportion of these two cytokines, we have studied if a TaqI polymorphism in the IL-1 beta gene, which is relevant to IL-1 beta protein production, may be involved in the genetic susceptibility to UC and Crohn's disease (CD), in association with the established IL-1Ra gene polymorphism. Polymorphisms in the closely linked genes for IL-1 beta and IL-1Ra were typed in 100 unrelated Dutch patients with UC, 79 with CD, and 71 healthy controls. The polymorphic regions in exon 5 of the IL-1 beta gene and in intron 2 of the IL-1Ra gene, were studied by polymerase chain reaction (PCR)-based methods. The IL-1 beta allele frequencies in UC and CD patients did not differ from those in healthy controls. In order to study if the IL-1 beta gene polymorphism might participate synergistically with the IL-1Ra gene polymorphism in susceptibility to UC and CD, individuals were distributed into carriers and non-carriers of allele 2 of the genes encoding IL-1 beta and IL-1Ra, in each of the patient groups and controls. Results indicated a significant association of this pair of genes, estimated by the odds ratio (OR) after performing Fisher's exact test, in the UC group (P = 0.023, OR = 2.81), as well as in the CD group (P = 0.01, OR = 3.79). Thus, non-carriers of IL-1 beta allele 2 were more often present in the subgroup of patients carrying the IL-1Ra allele 2. By contrast, no association of these alleles was detected in the group of healthy controls (P = 1.00, OR = 0.92). These results suggest that the IL-1 beta/IL-1Ra allelic cluster may participate in defining the biological basis of predisposition to chronic inflammatory bowel diseases. PMID:7586694

  20. Alleles of the maize P gene with distinct tissue specificities encode Myb-homologous proteins with C-terminal replacements.

    PubMed Central

    Chopra, S; Athma, P; Peterson, T

    1996-01-01

    The maize P gene is a transcriptional regulator of genes encoding enzymes for flavonoid biosynthesis in the pathway leading to the production of a red phlobaphene pigment. Multiple alleles of the P gene confer distinct patterns of pigmentation to specific floral organs, such as the kernel pericarp and cob tissues. To determine the basis of allele-specific pigmentation, we have characterized the gene products and transcript accumulation patterns of the P-wr allele, which specifies colorless pericarps and red cob tissues. RNA transcripts of P-wr are present in colorless pericarps as well as in the colored cob tissues; however, the expression of P-wr in pericarp does not induce the accumulation of transcripts from the C2 and A1 genes, which encode enzymes for flavonoid pigment biosynthesis. The coding sequences of P-wr were compared with the P-rr allele, which specifies red pericarp and red cob. The P-wr and P-rr cDNA sequences are very similar in their 5' regions. There are only two nucleotide changes that result in amino acid differences; both are outside of the Myb-homologous DNA binding domain. In contrast, the 3' coding region of P-rr is replaced by a unique 210-bp sequence in P-wr. The predicted P-wr protein has a C-terminal sequence resembling a cysteine-containing metal binding domain that is not present in the P-rr protein. These results indicate that the differential pericarp pigmentation specified by the P-rr and P-wr alleles does not result from an absence of P-wr transcripts in pericarps. Rather, the allele-specific patterns of P-rr and P-wr pigmentation may be associated with structural differences in the proteins encoded by each allele. PMID:8768374

  1. Enrichment of pathogenic alleles in the brittle cornea gene, ZNF469, in keratoconus.

    PubMed

    Lechner, Judith; Porter, Louise F; Rice, Aine; Vitart, Veronique; Armstrong, David J; Schorderet, Daniel F; Munier, Francis L; Wright, Alan F; Inglehearn, Chris F; Black, Graeme C; Simpson, David A; Manson, Forbes; Willoughby, Colin E

    2014-10-15

    Keratoconus, a common inherited ocular disorder resulting in progressive corneal thinning, is the leading indication for corneal transplantation in the developed world. Genome-wide association studies have identified common SNPs 100 kb upstream of ZNF469 strongly associated with corneal thickness. Homozygous mutations in ZNF469 and PR domain-containing protein 5 (PRDM5) genes result in brittle cornea syndrome (BCS) Types 1 and 2, respectively. BCS is an autosomal recessive generalized connective tissue disorder associated with extreme corneal thinning and a high risk of corneal rupture. Some individuals with heterozygous PRDM5 mutations demonstrate a carrier ocular phenotype, which includes a mildly reduced corneal thickness, keratoconus and blue sclera. We hypothesized that heterozygous variants in PRDM5 and ZNF469 predispose to the development of isolated keratoconus. We found a significant enrichment of potentially pathologic heterozygous alleles in ZNF469 associated with the development of keratoconus (P = 0.00102) resulting in a relative risk of 12.0. This enrichment of rare potentially pathogenic alleles in ZNF469 in 12.5% of keratoconus patients represents a significant mutational load and highlights ZNF469 as the most significant genetic factor responsible for keratoconus identified to date.

  2. Enrichment of pathogenic alleles in the brittle cornea gene, ZNF469, in keratoconus

    PubMed Central

    Lechner, Judith; Porter, Louise F.; Rice, Aine; Vitart, Veronique; Armstrong, David J.; Schorderet, Daniel F.; Munier, Francis L.; Wright, Alan F.; Inglehearn, Chris F.; Black, Graeme C.; Simpson, David A.; Manson, Forbes; Willoughby, Colin E.

    2014-01-01

    Keratoconus, a common inherited ocular disorder resulting in progressive corneal thinning, is the leading indication for corneal transplantation in the developed world. Genome-wide association studies have identified common SNPs 100 kb upstream of ZNF469 strongly associated with corneal thickness. Homozygous mutations in ZNF469 and PR domain-containing protein 5 (PRDM5) genes result in brittle cornea syndrome (BCS) Types 1 and 2, respectively. BCS is an autosomal recessive generalized connective tissue disorder associated with extreme corneal thinning and a high risk of corneal rupture. Some individuals with heterozygous PRDM5 mutations demonstrate a carrier ocular phenotype, which includes a mildly reduced corneal thickness, keratoconus and blue sclera. We hypothesized that heterozygous variants in PRDM5 and ZNF469 predispose to the development of isolated keratoconus. We found a significant enrichment of potentially pathologic heterozygous alleles in ZNF469 associated with the development of keratoconus (P = 0.00102) resulting in a relative risk of 12.0. This enrichment of rare potentially pathogenic alleles in ZNF469 in 12.5% of keratoconus patients represents a significant mutational load and highlights ZNF469 as the most significant genetic factor responsible for keratoconus identified to date. PMID:24895405

  3. Systematic morphological profiling of human gene and allele function via Cell Painting

    PubMed Central

    Rohban, Mohammad Hossein; Singh, Shantanu; Wu, Xiaoyun; Berthet, Julia B; Bray, Mark-Anthony; Shrestha, Yashaswi; Varelas, Xaralabos; Boehm, Jesse S; Carpenter, Anne E

    2017-01-01

    We hypothesized that human genes and disease-associated alleles might be systematically functionally annotated using morphological profiling of cDNA constructs, via a microscopy-based Cell Painting assay. Indeed, 50% of the 220 tested genes yielded detectable morphological profiles, which grouped into biologically meaningful gene clusters consistent with known functional annotation (e.g., the RAS-RAF-MEK-ERK cascade). We used novel subpopulation-based visualization methods to interpret the morphological changes for specific clusters. This unbiased morphologic map of gene function revealed TRAF2/c-REL negative regulation of YAP1/WWTR1-responsive pathways. We confirmed this discovery of functional connectivity between the NF-κB pathway and Hippo pathway effectors at the transcriptional level, thereby expanding knowledge of these two signaling pathways that critically regulate tumor initiation and progression. We make the images and raw data publicly available, providing an initial morphological map of major biological pathways for future study. DOI: http://dx.doi.org/10.7554/eLife.24060.001 PMID:28315521

  4. Identification of Bradyrhizobium elkanii Genes Involved in Incompatibility with Soybean Plants Carrying the Rj4 Allele

    PubMed Central

    Faruque, Omar M.; Miwa, Hiroki; Yasuda, Michiko; Fujii, Yoshiharu; Kaneko, Takakazu; Sato, Shusei

    2015-01-01

    Symbioses between leguminous plants and soil bacteria known as rhizobia are of great importance to agricultural production and nitrogen cycling. While these mutualistic symbioses can involve a wide range of rhizobia, some legumes exhibit incompatibility with specific strains, resulting in ineffective nodulation. The formation of nodules in soybean plants (Glycine max) is controlled by several host genes, which are referred to as Rj genes. The soybean cultivar BARC2 carries the Rj4 gene, which restricts nodulation by specific strains, including Bradyrhizobium elkanii USDA61. Here we employed transposon mutagenesis to identify the genetic locus in USDA61 that determines incompatibility with soybean varieties carrying the Rj4 allele. Introduction of the Tn5 transposon into USDA61 resulted in the formation of nitrogen fixation nodules on the roots of soybean cultivar BARC2 (Rj4 Rj4). Sequencing analysis of the sequence flanking the Tn5 insertion revealed that six genes encoding a putative histidine kinase, transcriptional regulator, DNA-binding transcriptional activator, helix-turn-helix-type transcriptional regulator, phage shock protein, and cysteine protease were disrupted. The cysteine protease mutant had a high degree of similarity with the type 3 effector protein XopD of Xanthomonas campestris. Our findings shed light on the diverse and complicated mechanisms that underlie these highly host-specific interactions and indicate the involvement of a type 3 effector in Rj4 nodulation restriction, suggesting that Rj4 incompatibility is partly mediated by effector-triggered immunity. PMID:26187957

  5. Rice WRKY13 regulates cross talk between abiotic and biotic stress signaling pathways by selective binding to different cis-elements.

    PubMed

    Xiao, Jun; Cheng, Hongtao; Li, Xianghua; Xiao, Jinghua; Xu, Caiguo; Wang, Shiping

    2013-12-01

    Plants use a complex signal transduction network to regulate their adaptation to the ever-changing environment. Rice (Oryza sativa) WRKY13 plays a vital role in the cross talk between abiotic and biotic stress signaling pathways by suppressing abiotic stress resistance and activating disease resistance. However, it is not clear how WRKY13 directly regulates this cross talk. Here, we show that WRKY13 is a transcriptional repressor. During the rice responses to drought stress and bacterial infection, WRKY13 selectively bound to certain site- and sequence-specific cis-elements on the promoters of SNAC1 (for STRESS RESPONSIVE NO APICAL MERISTEM, ARABIDOPSIS TRANSCRIPTION ACTIVATION FACTOR1/2, CUP-SHAPED COTYLEDON), the overexpression of which increases drought resistance, and WRKY45-1, the knockout of which increases both bacterial disease and drought resistance. WRKY13 also bound to two cis-elements of its native promoter to autoregulate the balance of its gene expression in different physiological activities. WRKY13 was induced in leaf vascular tissue, where bacteria proliferate, during infection, and in guard cells, where the transcriptional factor SNAC1 enhances drought resistance, during both bacterial infection and drought stress. These results suggest that WRKY13 regulates the antagonistic cross talk between drought and disease resistance pathways by directly suppressing SNAC1 and WRKY45-1 and autoregulating its own expression via site- and sequence-specific cis-elements on the promoters of these genes in vascular tissue where bacteria proliferate and guard cells where the transcriptional factor SNAC1 mediates drought resistance by promoting stomatal closure.

  6. Ectopic Expression of JcWRKY Transcription Factor Confers Salinity Tolerance via Salicylic Acid Signaling

    PubMed Central

    Agarwal, Parinita; Dabi, Mitali; Sapara, Komal K.; Joshi, Priyanka S.; Agarwal, Pradeep K.

    2016-01-01

    Plants, being sessile, have developed intricate signaling network to specifically respond to the diverse environmental stress. The plant-specific WRKY TFs form one of the largest TF family and are involved in diverse plant processes, involving growth, development and stress signaling through auto and cross regulation with different genes and TFs. Here, we report the functional characterization of a salicylic acid -inducible JcWRKY TF. The JcWRKY overexpression confers salinity tolerance in transgenic tobacco, as was evident by increased chlorophyll content and seed germination potential. The transgenic plants showed increased soluble sugar, membrane stability, reduced electrolyte leakage and generation of reactive oxygen species (H2O2 and O2•-) as compared to the wild type. Furthermore, the low SA treatment along with salinity improved the tolerance potential of the transgenics by maintaining ROS homeostasis and high K+/Na+ ratio. The transcript expression of SA biosynthetic gene ICS1 and antioxidative enzymes (CAT and SOD) showed upregulation during stress. Thus, the present study reflects that JcWRKY is working in co-ordination with SA signaling to orchestrate the different biochemical and molecular pathways to maneuvre salt stress tolerance of the transgenic plants. PMID:27799936

  7. Allele-Dependent Differences in Quorum-Sensing Dynamics Result in Variant Expression of Virulence Genes in Staphylococcus aureus

    PubMed Central

    Geisinger, Edward; Chen, John

    2012-01-01

    Agr is an autoinducing, quorum-sensing system that functions in many Gram-positive species and is best characterized in the pathogen Staphylococcus aureus, in which it is a global regulator of virulence gene expression. Allelic variations in the agr genes have resulted in the emergence of four quorum-sensing specificity groups in S. aureus, which correlate with different strain pathotypes. The basis for these predilections is unclear but is hypothesized to involve the phenomenon of quorum-sensing interference between strains of different agr groups, which may drive S. aureus strain isolation and divergence. Whether properties intrinsic to each agr allele directly influence virulence phenotypes within S. aureus is unknown. In this study, we examined group-specific differences in agr autoinduction and virulence gene regulation by utilizing congenic strains, each harboring a unique S. aureus agr allele, enabling a dissection of agr locus-dependent versus genotype-dependent effects on quorum-sensing dynamics and virulence factor production. Employing a reporter fusion to the principal agr promoter, P3, we observed allele-dependent differences in the timing and magnitude of agr activation. These differences were mediated by polymorphisms within the agrBDCA genes and translated to significant variations in the expression of a key transcriptional regulator, Rot, and of several important exoproteins and surface factors involved in pathogenesis. This work uncovers the contribution of divergent quorum-sensing alleles to variant expression of virulence determinants within a bacterial species. PMID:22467783

  8. Construction and application of a Korean reference panel for imputing classical alleles and amino acids of human leukocyte antigen genes.

    PubMed

    Kim, Kwangwoo; Bang, So-Young; Lee, Hye-Soon; Bae, Sang-Cheol

    2014-01-01

    Genetic variations of human leukocyte antigen (HLA) genes within the major histocompatibility complex (MHC) locus are strongly associated with disease susceptibility and prognosis for many diseases, including many autoimmune diseases. In this study, we developed a Korean HLA reference panel for imputing classical alleles and amino acid residues of several HLA genes. An HLA reference panel has potential for use in identifying and fine-mapping disease associations with the MHC locus in East Asian populations, including Koreans. A total of 413 unrelated Korean subjects were analyzed for single nucleotide polymorphisms (SNPs) at the MHC locus and six HLA genes, including HLA-A, -B, -C, -DRB1, -DPB1, and -DQB1. The HLA reference panel was constructed by phasing the 5,858 MHC SNPs, 233 classical HLA alleles, and 1,387 amino acid residue markers from 1,025 amino acid positions as binary variables. The imputation accuracy of the HLA reference panel was assessed by measuring concordance rates between imputed and genotyped alleles of the HLA genes from a subset of the study subjects and East Asian HapMap individuals. Average concordance rates were 95.6% and 91.1% at 2-digit and 4-digit allele resolutions, respectively. The imputation accuracy was minimally affected by SNP density of a test dataset for imputation. In conclusion, the Korean HLA reference panel we developed was highly suitable for imputing HLA alleles and amino acids from MHC SNPs in East Asians, including Koreans.

  9. Spelt-specific alleles in HMW glutenin genes from modern and historical European spelt ( Triticum spelta L.).

    PubMed

    Blatter, Robert H. E.; Jacomet, Stefanie; Schlumbaum, Angela

    2002-02-01

    A partial promoter region of the high-molecular weight (HMW) glutenin genes was studied in two wheat specimens, a 300 year-old spelt ( Triticum spelta L.) and an approximately 250 year-old bread wheat ( Triticum aestivum L.) from Switzerland. Sequences were compared to a recent Swiss landrace T. spelta'Oberkulmer.' The alleles from the historical bread wheat were most similar to those of modern T. aestivumcultivars, whereas in the historical and the recent spelt specific alleles were detected. Pairwise genetic distances up to 0.03 within 200 bp from the HMW Glu-A1-2, Glu-B1-1 and Glu-B1-2 alleles in spelt to the most-similar alleles from bread wheat suggest a polyphyletic origin. The spelt Glu-B1-1 allele, which was unlike the corresponding alleles in bread wheat, was closer related to an allele found in tetraploid wheat cultivars. The results are discussed in context of the origin of European spelt.

  10. No evidence for strong recent positive selection favoring the 7 repeat allele of VNTR in the DRD4 gene.

    PubMed

    Naka, Izumi; Nishida, Nao; Ohashi, Jun

    2011-01-01

    The human dopamine receptor D4 (DRD4) gene contains a 48-bp variable number of tandem repeat (VNTR) in exon 3, encoding the third intracellular loop of this dopamine receptor. The DRD4 7R allele, which seems to have a single origin, is commonly observed in various human populations and the nucleotide diversity of the DRD4 7R haplotype at the DRD4 locus is reduced compared to the most common DRD4 4R haplotype. Based on these observations, previous studies have hypothesized that positive selection has acted on the DRD4 7R allele. However, the degrees of linkage disequilibrium (LD) of the DRD4 7R allele with single nucleotide polymorphisms (SNPs) outside the DRD4 locus have not been evaluated. In this study, to re-examine the possibility of recent positive selection favoring the DRD4 7R allele, we genotyped HapMap subjects for DRD4 VNTR, and conducted several neutrality tests including long range haplotype test and iHS test based on the extended haplotype homozygosity. Our results indicated that LD of the DRD4 7R allele was not extended compared to SNP alleles with the similar frequency. Thus, we conclude that the DRD4 7R allele has not been subjected to strong recent positive selection.

  11. The Rice Transcription Factor WRKY53 Suppresses Herbivore-Induced Defenses by Acting as a Negative Feedback Modulator of Mitogen-Activated Protein Kinase Activity1

    PubMed Central

    Hu, Lingfei; Ye, Meng; Zhang, Tongfang; Zhou, Guoxin; Wang, Qi; Lu, Jing

    2015-01-01

    The mechanisms by which herbivore-attacked plants activate their defenses are well studied. By contrast, little is known about the regulatory mechanisms that allow them to control their defensive investment and avoid a defensive overshoot. We characterized a rice (Oryza sativa) WRKY gene, OsWRKY53, whose expression is rapidly induced upon wounding and induced in a delayed fashion upon attack by the striped stem borer (SSB) Chilo suppressalis. The transcript levels of OsWRKY53 are independent of endogenous jasmonic acid but positively regulated by the mitogen-activated protein kinases OsMPK3/OsMPK6. OsWRKY53 physically interacts with OsMPK3/OsMPK6 and suppresses their activity in vitro. By consequence, it modulates the expression of defensive, MPK-regulated WRKYs and thereby reduces jasmonic acid, jasmonoyl-isoleucine, and ethylene induction. This phytohormonal reconfiguration is associated with a reduction in trypsin protease inhibitor activity and improved SSB performance. OsWRKY53 is also shown to be a negative regulator of plant growth. Taken together, these results show that OsWRKY53 functions as a negative feedback modulator of MPK3/MPK6 and thereby acts as an early suppressor of induced defenses. OsWRKY53 therefore enables rice plants to control the magnitude of their defensive investment during early signaling. PMID:26453434

  12. Identification and expression of the WRKY transcription factors of Carica papaya in response to abiotic and biotic stresses.

    PubMed

    Pan, Lin-Jie; Jiang, Ling

    2014-03-01

    The WRKY transcription factor (TF) plays a very important role in the response of plants to various abiotic and biotic stresses. A local papaya database was built according to the GenBank expressed sequence tag database using the BioEdit software. Fifty-two coding sequences of Carica papaya WRKY TFs were predicted using the tBLASTn tool. The phylogenetic tree of the WRKY proteins was classified. The expression profiles of 13 selected C. papaya WRKY TF genes under stress induction were constructed by quantitative real-time polymerase chain reaction. The expression levels of these WRKY genes in response to 3 abiotic and 2 biotic stresses were evaluated. TF807.3 and TF72.14 are upregulated by low temperature; TF807.3, TF43.76, TF12.199 and TF12.62 are involved in the response to drought stress; TF9.35, TF18.51, TF72.14 and TF12.199 is involved in response to wound; TF12.199, TF807.3, TF21.156 and TF18.51 was induced by PRSV pathogen; TF72.14 and TF43.76 are upregulated by SA. The regulated expression levels of above eight genes normalized against housekeeping gene actin were significant at probability of 0.01 levels. These WRKY TFs could be related to corresponding stress resistance and selected as the candidate genes, especially, the two genes TF807.3 and TF12.199, which were regulated notably by four stresses respectively. This study may provide useful information and candidate genes for the development of transgenic stress tolerant papaya varieties.

  13. The fear gene stathmin alleles generated heterosis on feed efficiency parameters in Peking ducks.

    PubMed

    Wang, Yan Hong; Zhang, Hai Yan; Zhang, Chun Lei; Chen, Hong; Fang, Xing Tang; Zhang, Yun Sheng; Hou, Shui Sheng

    2012-01-01

    Stathmin is an inhibitor of microtubule formation, as highly expressed in the lateral nucleus (LA) of the amygdala as well as in the thalamic and cortical structures that send information to the LA about the learned and innate fear. So we assume that STMN1 genetic variation may also affect the physical activity so as to influence the Residual Feed Intake (RFI) of duck. The Single Nucleotide Polymorphisms (SNPs) in duck Stathmin gene were screened by sequencing and genotyped by restriction endonuclease Msp I, EcoR I, Xho I, Taq I, EcoR II. A total of five SNPs (c.187 -15G > A, c.187 -110T > C, c.379 -95G > A, c.379 -318C > T, c.426 C > T) were detected in duck STMN1 gene. The c.187 -15G > A is near the 3' splice site of intron 2, which has a putative effect on the STMN1 pre-mRNA secondary structures. The c.187 -15G > A genotypes had significant effect on RFI of Peking drakes (P < 0.01). Individuals with heterozygous genotypes were more productive than that with homozygous genotypes, which suggested a molecular heterosis in c.187 -15 alleles on RFI. The current study is the first step to confirm the relationship between STMN1 gene polymorphisms and RFI. Supplemental material is available for this article. Go to the publisher's online edition of Animal Biotechnology for a figure of linkage disequilibrium between SNPs and table about frequencies of haploype.

  14. Association of BLV infection profiles with alleles of the BoLA-DRB3.2 gene.

    PubMed

    Juliarena, M A; Poli, M; Sala, L; Ceriani, C; Gutierrez, S; Dolcini, G; Rodríguez, E M; Mariño, B; Rodríguez-Dubra, C; Esteban, E N

    2008-08-01

    Bovine leukaemia virus (BLV) causes lymphosarcoma and persistent lymphocytosis (PL). Some MHC class II gene polymorphisms have been associated with resistance and susceptibility to the development of lymphosarcoma and PL, as well as with a reduced number of circulating BLV-infected lymphocytes. Previously, 230 BLV-infected Holstein cattle were classified into two infection profiles characterized by low and high proviral loads (LPL and HPL respectively). Here, the influence of the polymorphism at the BoLA-DRB3.2* gene of these animals was examined. After genotyping, the association between the BoLA-DRB3.2* alleles and the BLV infection profile was determined as the odds ratio (OR). Two subtypes of allele *11 were identified (ISAG*0901 and *0902). Allele ISAG*0902 showed a stronger association with the LPL profile (OR = 8.24; P < 0.0001) than allele *11 itself (OR = 5.82; P < 0.0001). Allele ISAG*1701 (*12) also showed significant association with the LPL profile (OR = 3.46; P < 0.0055). Only one allele, ISAG*1501 or 03 (*16), showed significant association with HPL (OR = 0.36; P < 0.0005). The DRB3.2* alleles were assigned to three categories: resistant (R), susceptible (S) and neutral (N). Based on their DRB3 genotypes, cattle were classified as homozygous or heterozygous. The RR and RN genotypes were associated with the LPL profile, while the SS and NS genotypes were associated with the HPL profile. The RS genotype could not be associated with any particular profile. Our results show that allele ISAG*0902 appears to be the best BLV resistance marker in Holstein cattle.

  15. The human gene connectome as a map of short cuts for morbid allele discovery.

    PubMed

    Itan, Yuval; Zhang, Shen-Ying; Vogt, Guillaume; Abhyankar, Avinash; Herman, Melina; Nitschke, Patrick; Fried, Dror; Quintana-Murci, Lluis; Abel, Laurent; Casanova, Jean-Laurent

    2013-04-02

    High-throughput genomic data reveal thousands of gene variants per patient, and it is often difficult to determine which of these variants underlies disease in a given individual. However, at the population level, there may be some degree of phenotypic homogeneity, with alterations of specific physiological pathways underlying the pathogenesis of a particular disease. We describe here the human gene connectome (HGC) as a unique approach for human mendelian genetic research, facilitating the interpretation of abundant genetic data from patients with the same disease, and guiding subsequent experimental investigations. We first defined the set of the shortest plausible biological distances, routes, and degrees of separation between all pairs of human genes by applying a shortest distance algorithm to the full human gene network. We then designed a hypothesis-driven application of the HGC, in which we generated a Toll-like receptor 3-specific connectome useful for the genetic dissection of inborn errors of Toll-like receptor 3 immunity. In addition, we developed a functional genomic alignment approach from the HGC. In functional genomic alignment, the genes are clustered according to biological distance (rather than the traditional molecular evolutionary genetic distance), as estimated from the HGC. Finally, we compared the HGC with three state-of-the-art methods: String, FunCoup, and HumanNet. We demonstrated that the existing methods are more suitable for polygenic studies, whereas HGC approaches are more suitable for monogenic studies. The HGC and functional genomic alignment data and computer programs are freely available to noncommercial users from http://lab.rockefeller.edu/casanova/HGC and should facilitate the genome-wide selection of disease-causing candidate alleles for experimental validation.

  16. The human gene connectome as a map of short cuts for morbid allele discovery

    PubMed Central

    Itan, Yuval; Zhang, Shen-Ying; Vogt, Guillaume; Abhyankar, Avinash; Herman, Melina; Nitschke, Patrick; Fried, Dror; Quintana-Murci, Lluis; Abel, Laurent; Casanova, Jean-Laurent

    2013-01-01

    High-throughput genomic data reveal thousands of gene variants per patient, and it is often difficult to determine which of these variants underlies disease in a given individual. However, at the population level, there may be some degree of phenotypic homogeneity, with alterations of specific physiological pathways underlying the pathogenesis of a particular disease. We describe here the human gene connectome (HGC) as a unique approach for human Mendelian genetic research, facilitating the interpretation of abundant genetic data from patients with the same disease, and guiding subsequent experimental investigations. We first defined the set of the shortest plausible biological distances, routes, and degrees of separation between all pairs of human genes by applying a shortest distance algorithm to the full human gene network. We then designed a hypothesis-driven application of the HGC, in which we generated a Toll-like receptor 3-specific connectome useful for the genetic dissection of inborn errors of Toll-like receptor 3 immunity. In addition, we developed a functional genomic alignment approach from the HGC. In functional genomic alignment, the genes are clustered according to biological distance (rather than the traditional molecular evolutionary genetic distance), as estimated from the HGC. Finally, we compared the HGC with three state-of-the-art methods: String, FunCoup, and HumanNet. We demonstrated that the existing methods are more suitable for polygenic studies, whereas HGC approaches are more suitable for monogenic studies. The HGC and functional genomic alignment data and computer programs are freely available to noncommercial users from http://lab.rockefeller.edu/casanova/HGC and should facilitate the genome-wide selection of disease-causing candidate alleles for experimental validation. PMID:23509278

  17. Vitamin D receptor alleles: Cloning and characterization of the VDR gene and RT-PCR of VDR cDNA

    SciTech Connect

    Javed, A.A.; Huang, Y.; Bombard, A.T.

    1994-09-01

    Vitamin D{sub 3} receptors (VDR) function as regulators through the action of the ligand 1{alpha}, 25-dihydroxy vitamin D{sub 3}. The receptor specifically finds its ligand and exerts it effect on the regulation of the expression of target genes. It has been shown that mutations in the VDR gene affect the function of the receptors and cause a corresponding disorder state. Recently, it has been reported that common allelic variations found normally in the Caucasian (Australian) population pose varying degrees of risk for osteoporosis. We present here the cloning of the VDR gene and RT-PCR of VDR cDNA. Studies are in progress to establish allele frequency in the Black, Hispanic and Caucasian populations to systematically study the influence of allele types and to develop a risk profile for osteoporosis. The present method for detection of various alleles is based on RFLP analysis. We are developing PCR-based methods for the rapid detection and typing of alleles.

  18. Allelic Diversity and Geographical Distribution of the Gene Encoding Plasmodium falciparum Merozoite Surface Protein-3 in Thailand.

    PubMed

    Sawaswong, Vorthon; Simpalipan, Phumin; Siripoon, Napaporn; Harnyuttanakorn, Pongchai; Pattaradilokrat, Sittiporn

    2015-04-01

    Merozoite surface proteins (MSPs) of malaria parasites play critical roles during the erythrocyte invasion and so are potential candidates for malaria vaccine development. However, because MSPs are often under strong immune selection, they can exhibit extensive genetic diversity. The gene encoding the merozoite surface protein-3 (MSP-3) of Plasmodium falciparum displays 2 allelic types, K1 and 3D7. In Thailand, the allelic frequency of the P. falciparum msp-3 gene was evaluated in a single P. falciparum population in Tak at the Thailand and Myanmar border. However, no study has yet looked at the extent of genetic diversity of the msp-3 gene in P. falciparum populations in other localities. Here, we genotyped the msp-3 alleles of 63 P. falciparum samples collected from 5 geographical populations along the borders of Thailand with 3 neighboring countries (Myanmar, Laos, and Cambodia). Our study indicated that the K1 and 3D7 alleles coexisted, but at different proportions in different Thai P. falciparum populations. K1 was more prevalent in populations at the Thailand-Myanmar and Thailand-Cambodia borders, whilst 3D7 was more prevalent at the Thailand-Laos border. Global analysis of the msp-3 allele frequencies revealed that proportions of K1 and 3D7 alleles of msp-3 also varied in different continents, suggesting the divergence of malaria parasite populations. In conclusion, the variation in the msp-3 allelic patterns of P. falciparum in Thailand provides fundamental knowledge for inferring the P. falciparum population structure and for the best design of msp-3 based malaria vaccines.

  19. Natural variation in the Pto pathogen resistance gene within species of wild tomato (Lycopersicon). I. Functional analysis of Pto alleles.

    PubMed

    Rose, Laura E; Langley, Charles H; Bernal, Adriana J; Michelmore, Richard W

    2005-09-01

    Disease resistance to the bacterial pathogen Pseudomonas syringae pv. tomato (Pst) in the cultivated tomato, Lycopersicon esculentum, and the closely related L. pimpinellifolium is triggered by the physical interaction between plant disease resistance protein, Pto, and the pathogen avirulence protein, AvrPto. To investigate the extent to which variation in the Pto gene is responsible for naturally occurring variation in resistance to Pst, we determined the resistance phenotype of 51 accessions from seven species of Lycopersicon to isogenic strains of Pst differing in the presence of avrPto. One-third of the plants displayed resistance specifically when the pathogen expressed AvrPto, consistent with a gene-for-gene interaction. To test whether this resistance in these species was conferred specifically by the Pto gene, alleles of Pto were amplified and sequenced from 49 individuals and a subset (16) of these alleles was tested in planta using Agrobacterium-mediated transient assays. Eleven alleles conferred a hypersensitive resistance response (HR) in the presence of AvrPto, while 5 did not. Ten amino acid substitutions associated with the absence of AvrPto recognition and HR were identified, none of which had been identified in previous structure-function studies. Additionally, 3 alleles encoding putative pseudogenes of Pto were isolated from two species of Lycopersicon. Therefore, a large proportion, but not all, of the natural variation in the reaction to strains of Pst expressing AvrPto can be attributed to sequence variation in the Pto gene.

  20. Natural Variation in the Pto Pathogen Resistance Gene Within Species of Wild Tomato (Lycopersicon). I. Functional Analysis of Pto Alleles

    PubMed Central

    Rose, Laura E.; Langley, Charles H.; Bernal, Adriana J.; Michelmore, Richard W.

    2005-01-01

    Disease resistance to the bacterial pathogen Pseudomonas syringae pv. tomato (Pst) in the cultivated tomato, Lycopersicon esculentum, and the closely related L. pimpinellifolium is triggered by the physical interaction between plant disease resistance protein, Pto, and the pathogen avirulence protein, AvrPto. To investigate the extent to which variation in the Pto gene is responsible for naturally occurring variation in resistance to Pst, we determined the resistance phenotype of 51 accessions from seven species of Lycopersicon to isogenic strains of Pst differing in the presence of avrPto. One-third of the plants displayed resistance specifically when the pathogen expressed AvrPto, consistent with a gene-for-gene interaction. To test whether this resistance in these species was conferred specifically by the Pto gene, alleles of Pto were amplified and sequenced from 49 individuals and a subset (16) of these alleles was tested in planta using Agrobacterium-mediated transient assays. Eleven alleles conferred a hypersensitive resistance response (HR) in the presence of AvrPto, while 5 did not. Ten amino acid substitutions associated with the absence of AvrPto recognition and HR were identified, none of which had been identified in previous structure-function studies. Additionally, 3 alleles encoding putative pseudogenes of Pto were isolated from two species of Lycopersicon. Therefore, a large proportion, but not all, of the natural variation in the reaction to strains of Pst expressing AvrPto can be attributed to sequence variation in the Pto gene. PMID:15944360

  1. Heterologous Expression of AtWRKY57 Confers Drought Tolerance in Oryza sativa.

    PubMed

    Jiang, Yanjuan; Qiu, Yuping; Hu, Yanru; Yu, Diqiu

    2016-01-01

    Drought stress is a severe environmental factor that greatly restricts plant distribution and crop production. Recently, we have found that overexpressing AtWRKY57 enhanced drought tolerance in Arabidopsis thaliana. In this study, we further reported that the Arabidopsis WRKY57 transcription factor was able to confer drought tolerance to transgenic rice (Oryza sativa) plants. The enhanced drought tolerance of transgenic rice was resulted from the lower water loss rates, cell death, malondialdehyde contents and relative electrolyte leakage while a higher proline content and reactive oxygen species-scavenging enzyme activities was observed during stress conditions. Moreover, further investigation revealed that the expression levels of several stress-responsive genes were up-regulated in drought-tolerant transgenic rice plants, compared with those in wild-type plants. In addition to the drought tolerance, the AtWRKY57 over-expressing plants also had enhanced salt and PEG stress tolerances. Taken together, our study indicates that over-expressing AtWRKY57 in rice improved not only drought tolerance but also salt and PEG tolerance, demonstrating its potential role in crop improvement.

  2. Heterologous Expression of AtWRKY57 Confers Drought Tolerance in Oryza sativa

    PubMed Central

    Jiang, Yanjuan; Qiu, Yuping; Hu, Yanru; Yu, Diqiu

    2016-01-01

    Drought stress is a severe environmental factor that greatly restricts plant distribution and crop production. Recently, we have found that overexpressing AtWRKY57 enhanced drought tolerance in Arabidopsis thaliana. In this study, we further reported that the Arabidopsis WRKY57 transcription factor was able to confer drought tolerance to transgenic rice (Oryza sativa) plants. The enhanced drought tolerance of transgenic rice was resulted from the lower water loss rates, cell death, malondialdehyde contents and relative electrolyte leakage while a higher proline content and reactive oxygen species-scavenging enzyme activities was observed during stress conditions. Moreover, further investigation revealed that the expression levels of several stress-responsive genes were up-regulated in drought-tolerant transgenic rice plants, compared with those in wild-type plants. In addition to the drought tolerance, the AtWRKY57 over-expressing plants also had enhanced salt and PEG stress tolerances. Taken together, our study indicates that over-expressing AtWRKY57 in rice improved not only drought tolerance but also salt and PEG tolerance, demonstrating its potential role in crop improvement. PMID:26904091

  3. Environmental Stability of Seed Carbohydrate Profiles in Soybeans Containing Different Alleles of the Raffinose Synthase 2 (RS2) Gene.

    PubMed

    Bilyeu, Kristin D; Wiebold, William J

    2016-02-10

    Soybean [Glycine max (L.) Merr.] is important for the high protein meal used for livestock feed formulations. Carbohydrates contribute positively or negatively to the potential metabolizable energy in soybean meal. The positive carbohydrate present in soybean meal consists primarily of sucrose, whereas the negative carbohydrate components are the raffinose family of oligosaccharides (RFOs), raffinose and stachyose. Increasing sucrose and decreasing raffinose and stachyose are critical targets to improve soybean. In three recently characterized lines, variant alleles of the soybean raffinose synthase 2 (RS2) gene were associated with increased sucrose and decreased RFOs. The objective of this research was to compare the environmental stability of seed carbohydrates in soybean lines containing wild-type or variant alleles of RS2 utilizing a field location study and a date of planting study. The results define the carbohydrate variation in distinct regional and temporal environments using soybean lines with different alleles of the RS2 gene.

  4. Generation and Characterization of Mice Carrying a Conditional Allele of the Wwox Tumor Suppressor Gene

    PubMed Central

    Ludes-Meyers, John H.; Kil, Hyunsuk; Parker-Thornburg, Jan; Kusewitt, Donna F.; Bedford, Mark T.; Aldaz, C. Marcelo

    2009-01-01

    WWOX, the gene that spans the second most common human chromosomal fragile site, FRA16D, is inactivated in multiple human cancers and behaves as a suppressor of tumor growth. Since we are interested in understanding WWOX function in both normal and cancer tissues we generated mice harboring a conditional Wwox allele by flanking Exon 1 of the Wwox gene with LoxP sites. Wwox knockout (KO) mice were developed by breeding with transgenic mice carrying the Cre-recombinase gene under the control of the adenovirus EIIA promoter. We found that Wwox KO mice suffered from severe metabolic defect(s) resulting in growth retardation and all mice died by 3 wk of age. All Wwox KO mice displayed significant hypocapnia suggesting a state of metabolic acidosis. This finding and the known high expression of Wwox in kidney tubules suggest a role for Wwox in acid/base balance. Importantly, Wwox KO mice displayed histopathological and hematological signs of impaired hematopoeisis, leukopenia, and splenic atrophy. Impaired hematopoeisis can also be a contributing factor to metabolic acidosis and death. Hypoglycemia and hypocalcemia was also observed affecting the KO mice. In addition, bone metabolic defects were evident in Wwox KO mice. Bones were smaller and thinner having reduced bone volume as a consequence of a defect in mineralization. No evidence of spontaneous neoplasia was observed in Wwox KO mice. We have generated a new mouse model to inactivate the Wwox tumor suppressor gene conditionally. This will greatly facilitate the functional analysis of Wwox in adult mice and will allow investigating neoplastic transformation in specific target tissues. PMID:19936220

  5. Gene therapy by allele selection in a mouse model of beta-thalassemia.

    PubMed

    Eckardt, Sigrid; Leu, N Adrian; Yanchik, Ashley; Hatada, Seigo; Kyba, Michael; McLaughlin, K John

    2011-02-01

    To be of therapeutic use, autologous stem cells derived from patients with inherited genetic disorders require genetic modification via gene repair or insertion. Here, we present proof of principle that, for diseases associated with dominant alleles (gain-of-function or haploinsufficient loss-of-function), disease allele–free ES cells can be derived from afflicted individuals without genome manipulation. This approach capitalizes on the derivation of uniparental cells, such as parthenogenetic (PG) ES cell lines from disease allele–free gametes. Diploid mammalian uniparental embryos with only maternally (oocyte-) or paternally (sperm-)derived genomes fail early in development due to the nonequivalence of parental genomes caused by genomic imprinting. However, these uniparental embryos develop to the blastocyst stage, allowing the derivation of ES cell lines. Using a mouse model for dominant beta-thalassemia, we developed disease allele–free PG ES cell lines from the oocytes of affected animals. Phenotype correction was obtained in donor-genotype recipients after transplantation of in vitro hematopoietic ES cell derivatives. This genetic correction strategy without gene targeting is potentially applicable to any dominant disease. It could also be the sole approach for larger or more complex mutations that cannot be corrected by homologous recombination.

  6. MspI allelic pattern of bovine growth hormone gene in Indian zebu cattle (Bos indicus) breeds.

    PubMed

    Sodhi, M; Mukesh, M; Prakash, B; Mishra, B P; Sobti, R C; Singh, Karn P; Singh, Satbir; Ahlawat, S P S

    2007-02-01

    The MspI allelic variation in intron III of the bovine growth hormone (bGH) gene was explored using PCR-RFLP in 750 animals belonging to 17 well-recognized breeds of Indian zebu cattle (Bos indicus) reared in different geographic locations of the country. Restriction digestion analysis of a 329-bp PCR fragment of the bGH intron III region with MspI restriction enzyme revealed two alleles (MspI- and MspI+) and two genotypes (-/- and +/-) across the 17 cattle breeds studied. The allelic frequency varied from 0.67 to 0.94 for MspI (-) and from 0.06 to 0.33 for MspI (+) across the 17 breeds, with a combined average frequency of 0.87 and 0.13, respectively. No animal with +/+ genotype was detected across the samples analyzed. The chi-square test showed that the difference in MspI allelic frequency was not significant (p > 0.05), regardless of the geographic origin, coat color, or utility of the cattle breed. The high MspI (-) allele frequencies obtained for Indian zebu cattle in this study are in sharp contrast to those reported for taurine breeds from northern Europe, Mediterranean countries, and America. Findings of this study further substantiate the hypothesis that the MspI (-) allele has an Indian origin.

  7. Multiple heterologies increase mitotic double-strand break-induced allelic gene conversion tract lengths in yeast.

    PubMed Central

    Nickoloff, J A; Sweetser, D B; Clikeman, J A; Khalsa, G J; Wheeler, S L

    1999-01-01

    Spontaneous and double-strand break (DSB)-induced allelic recombination in yeast was investigated in crosses between ura3 heteroalleles inactivated by an HO site and a +1 frameshift mutation, with flanking markers defining a 3.4-kbp interval. In some crosses, nine additional phenotypically silent RFLP mutations were present at approximately 100-bp intervals. Increasing heterology from 0.2 to 1% in this interval reduced spontaneous, but not DSB-induced, recombination. For DSB-induced events, 75% were continuous tract gene conversions without a crossover in this interval; discontinuous tracts and conversions associated with a crossover each comprised approximately 7% of events, and 10% also converted markers in unbroken alleles. Loss of heterozygosity was seen for all markers centromere distal to the HO site in 50% of products; such loss could reflect gene conversion, break-induced replication, chromosome loss, or G2 crossovers. Using telomere-marked strains we determined that nearly all allelic DSB repair occurs by gene conversion. We further show that most allelic conversion results from mismatch repair of heteroduplex DNA. Interestingly, markers shared between the sparsely and densely marked interval converted at higher rates in the densely marked interval. Thus, the extra markers increased gene conversion tract lengths, which may reflect mismatch repair-induced recombination, or a shift from restoration- to conversion-type repair. PMID:10511547

  8. BGMUT: NCBI dbRBC database of allelic variations of genes encoding antigens of blood group systems.

    PubMed

    Patnaik, Santosh Kumar; Helmberg, Wolfgang; Blumenfeld, Olga O

    2012-01-01

    Analogous to human leukocyte antigens, blood group antigens are surface markers on the erythrocyte cell membrane whose structures differ among individuals and which can be serologically identified. The Blood Group Antigen Gene Mutation Database (BGMUT) is an online repository of allelic variations in genes that determine the antigens of various human blood group systems. The database is manually curated with allelic information collated from scientific literature and from direct submissions from research laboratories. Currently, the database documents sequence variations of a total of 1251 alleles of all 40 gene loci that together are known to affect antigens of 30 human blood group systems. When available, information on the geographic or ethnic prevalence of an allele is also provided. The BGMUT website also has general information on the human blood group systems and the genes responsible for them. BGMUT is a part of the dbRBC resource of the National Center for Biotechnology Information, USA, and is available online at http://www.ncbi.nlm.nih.gov/projects/gv/rbc/xslcgi.fcgi?cmd=bgmut. The database should be of use to members of the transfusion medicine community, those interested in studies of genetic variation and related topics such as human migrations, and students as well as members of the general public.

  9. Single strand conformation polymorphism analysis of candidate genes for reliable identification of alleles by capillary array electrophoresis.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    We investigated the reliability of capillary array electrophoresis-SSCP to determine if it can be used to identify novel alleles of candidate genes in a germplasm collection. Both strands of three different size fragments (160 bp, 245 pb and 437 bp) that differed by one or more nucleotides in sequen...

  10. Argument within a Scientific Debate: The Case of the DRD2 A1 Allele as a Gene for Alcoholism.

    ERIC Educational Resources Information Center

    Wastyn, Ronald O.; Wastyn, M. Linda

    1997-01-01

    Investigates how opposing parties advanced arguments to the scientific community about the validity of DRD2 A1 allele as a gene causing alcoholism. Demonstrates to what extent scientists debate each other in journals by advancing opposing viewpoints with rigor and insight. Reveals what it means when scientists label a discovery in terms of finding…

  11. The 2-repeat allele of the MAOA gene confers an increased risk for shooting and stabbing behaviors.

    PubMed

    Beaver, Kevin M; Barnes, J C; Boutwell, Brian B

    2014-09-01

    There has been a great deal of research examining the link between a polymorphism in the promoter region of the MAOA gene and antisocial phenotypes. The results of these studies have consistently revealed that low activity MAOA alleles are related to antisocial behaviors for males who were maltreated as children. Recently, though, some evidence has emerged indicating that a rare allele of the MAOA gene-that is, the 2-repeat allele-may have effects on violence that are independent of the environment. The current study builds on this research and examines the association between the 2-repeat allele and shooting and stabbing behaviors in a sample of males drawn from the National Longitudinal Study of Adolescent Health. Analyses revealed that African-American males who carry the 2-repeat allele are significantly more likely than all other genotypes to engage in shooting and stabbing behaviors and to report having multiple shooting and stabbing victims. The limitations of the study are discussed and suggestions for future research are offered.

  12. Allele frequencies of three factor VIII gene polymorphisms in Iranian populations and their application in hemophilia A carrier detection.

    PubMed

    Azimifar, S Babak; Seyedna, S Yoosef; Zeinali, Sirous

    2006-05-01

    Hemophilia A is an X-linked recessive bleeding disorder caused by a quantitative or qualitative deficiency of blood coagulation factor VIII (FVIII). ARMS (amplification refractory mutation system) primers were designed to determine allele frequencies of three FVIII gene linked markers, IVS7 nt 27 G/A SNP, BclI/intron 18, and HindIII/intron 19 among 85 normal Iranian women from unrelated families. Then same method was applied to perform carrier detection for hemophilia A families. The allele frequencies of IVS7 nt 27 "G"/"A" allele, BclI "T"/"A" allele, and HindIII "C"/"T" allele among normal women were 0.88/0.12, 0.52/0.48, and 0.48/0.52, respectively. The three polymorphisms were found to be in strong linkage disequilibrium, which decreased the overall heterozygosity to 51%. Twenty-one women from 15 unrelated hemophilia A families were referred to us for hemophilia A carrier detection. Taking advantage of these three biallelic polymorphisms in conjunction with multiallelic St14 VNTR (locus DXS52), IVS13 (CA)n STR, and IVS22 (CA)n STR, carrier status was determined in 16 women (16/21 or 76% of the at-risk women) from 11 families (11/15 or 73% of the families). The used ARMS methods are rapid and can easily be applied in conjunction with other FVIII gene linked polymorphisms for indirect mutation detection of hemophilia A where they are informative.

  13. Allelic Diversity and Population Structure in Oenococcus oeni as Determined from Sequence Analysis of Housekeeping Genes

    PubMed Central

    de las Rivas, Blanca; Marcobal, Ángela; Muñoz, Rosario

    2004-01-01

    Oenococcus oeni is the organism of choice for promoting malolactic fermentation in wine. The population biology of O. oeni is poorly understood and remains unclear. For a better understanding of the mode of genetic variation within this species, we investigated by using multilocus sequence typing (MLST) with the gyrB, pgm, ddl, recP, and mleA genes the genetic diversity and genetic relationships among 18 O. oeni strains isolated in various years from wines of the United States, France, Germany, Spain, and Italy. These strains have also been characterized by ribotyping and restriction fragment length polymorphism (RFLP) analysis of the PCR-amplified 16S-23S rRNA gene intergenic spacer region (ISR). Ribotyping grouped the strains into two groups; however, the RFLP analysis of the ISRs showed no differences in the strains analyzed. In contrast, MLST in oenococci had a good discriminatory ability, and we have found a higher genetic diversity than indicated by ribotyping analysis. All sequence types were represented by a single strain, and all the strains could be distinguished from each other because they had unique combinations of alleles. Strains assumed to be identical showed the same sequence type. Phylogenetic analyses indicated a panmictic population structure in O. oeni. Sequences were analyzed for evidence of recombination by split decomposition analysis and analysis of clustered polymorphisms. All results indicated that recombination plays a major role in creating the genetic heterogeneity of O. oeni. A low standardized index of association value indicated that the O. oeni genes analyzed are close to linkage equilibrium. This study constitutes the first step in the development of an MLST method for O. oeni and the first example of the application of MLST to a nonpathogenic food production bacteria. PMID:15574919

  14. Regulation of Specialized Metabolism by WRKY Transcription Factors

    PubMed Central

    Schluttenhofer, Craig; Yuan, Ling

    2015-01-01

    WRKY transcription factors (TFs) are well known for regulating plant abiotic and biotic stress tolerance. However, much less is known about how WRKY TFs affect plant-specialized metabolism. Analysis of WRKY TFs regulating the production of specialized metabolites emphasizes the values of the family outside of traditionally accepted roles in stress tolerance. WRKYs with conserved roles across plant species seem to be essential in regulating specialized metabolism. Overall, the WRKY family plays an essential role in regulating the biosynthesis of important pharmaceutical, aromatherapy, biofuel, and industrial components, warranting considerable attention in the forthcoming years. PMID:25501946

  15. Efficient bi-allelic gene knockout and site-specific knock-in mediated by TALENs in pigs.

    PubMed

    Yao, Jing; Huang, Jiaojiao; Hai, Tang; Wang, Xianlong; Qin, Guosong; Zhang, Hongyong; Wu, Rong; Cao, Chunwei; Xi, Jianzhong Jeff; Yuan, Zengqiang; Zhao, Jianguo

    2014-11-05

    Pigs are ideal organ donors for xenotransplantation and an excellent model for studying human diseases, such as neurodegenerative disease. Transcription activator-like effector nucleases (TALENs) are used widely for gene targeting in various model animals. Here, we developed a strategy using TALENs to target the GGTA1, Parkin and DJ-1 genes in the porcine genome using Large White porcine fibroblast cells without any foreign gene integration. In total, 5% (2/40), 2.5% (2/80), and 22% (11/50) of the obtained colonies of fibroblast cells were mutated for GGTA1, Parkin, and DJ-1, respectively. Among these mutant colonies, over 1/3 were bi-allelic knockouts (KO), and no off-target cleavage was detected. We also successfully used single-strand oligodeoxynucleotides to introduce a short sequence into the DJ-1 locus. Mixed DJ-1 mutant colonies were used as donor cells for somatic cell nuclear transfer (SCNT), and three female piglets were obtained (two were bi-allelically mutated, and one was mono-allelically mutated). Western blot analysis showed that the expression of the DJ-1 protein was disrupted in KO piglets. These results imply that a combination of TALENs technology with SCNT can efficiently generate bi-allelic KO pigs without the integration of exogenous DNA. These DJ-1 KO pigs will provide valuable information for studying Parkinson's disease.

  16. Genotypic and allelic frequencies of gene polymorphisms associated with meat tenderness in Nellore beef cattle.

    PubMed

    Carvalho, M E; Eler, J P; Bonin, M N; Rezende, F M; Biase, F H; Meirelles, F V; Regitano, L C A; Coutinho, L L; Balieiro, J C C; Ferraz, J B S

    2017-02-16

    The objectives of this study were to characterize the allelic and genotypic frequencies of polymorphisms in the µ-calpain and calpastatin genes, and to assess their association with meat tenderness and animal growth in Nellore cattle. We evaluated 605 Nellore animals at 24 months of age, on average, at slaughter. The polymorphisms were determined for the molecular markers CAPN316, CAPN530, CAPN4751, CAPN4753, and UOGACAST1. Analyses of meat tenderness at 7, 14, and 21 days of maturation were performed in samples of longissimus thoracis obtained between the 12th and 13th rib and sheared using a Warner Bratzler Shear Force. Significant effects were observed for meat tenderness at days 7, 14, and 21 of maturation for the marker CAPN4751, at day 21 for the marker CAPN4753, and at days 14 and 21 for the marker UOGCAST1. For genotypic combinations of markers, the results were significant for the combination CAPN4751/UOGCAST1 in the three maturation periods and CAPN4753/UOGCAST1 at days 14 and 21 of maturation.

  17. Increased Atherosclerosis and Endothelial Dysfunction in Mice Bearing Constitutively Deacetylated Alleles of Foxo1 Gene*

    PubMed Central

    Qiang, Li; Tsuchiya, Kyoichiro; Kim-Muller, Ja-Young; Lin, Hua V.; Welch, Carrie; Accili, Domenico

    2012-01-01

    Complications of atherosclerosis are the leading cause of death of patients with type 2 (insulin-resistant) diabetes. Understanding the mechanisms by which insulin resistance and hyperglycemia contribute to atherogenesis in key target tissues (liver, vessel wall, hematopoietic cells) can assist in the design of therapeutic approaches. We have shown that hyperglycemia induces FoxO1 deacetylation and that targeted knock-in of alleles encoding constitutively deacetylated FoxO1 in mice (Foxo1KR/KR) improves hepatic lipid metabolism and decreases macrophage inflammation, setting the stage for a potential anti-atherogenic effect of this mutation. Surprisingly, we report here that when Foxo1KR/KR mice are intercrossed with low density lipoprotein receptor knock-out mice (Ldlr−/−), they develop larger aortic root atherosclerotic lesions than Ldlr−/− controls despite lower plasma cholesterol and triglyceride levels. The phenotype is unaffected by transplanting bone marrow from Ldlr−/− mice into Foxo1KR/KR mice, indicating that it is independent of hematopoietic cells and suggesting that the primary lesion in Foxo1KR/KR mice occurs in the vessel wall. Experiments in isolated endothelial cells from Foxo1KR/KR mice indicate that deacetylation favors FoxO1 nuclear accumulation and exerts target gene-specific effects, resulting in higher Icam1 and Tnfα expression and increased monocyte adhesion. The data indicate that FoxO1 deacetylation can promote vascular endothelial changes conducive to atherosclerotic plaque formation. PMID:22389493

  18. Neural correlate of autistic-like traits and a common allele in the oxytocin receptor gene

    PubMed Central

    Saito, Yuki; Suga, Motomu; Tochigi, Mamoru; Abe, Osamu; Yahata, Noriaki; Kawakubo, Yuki; Liu, Xiaoxi; Kawamura, Yoshiya; Sasaki, Tsukasa; Kasai, Kiyoto

    2014-01-01

    Sub-clinical autistic-like traits (ALTs) are continuously distributed in the general population and genetically linked to autism. Although identifying the neurogenetic backgrounds of ALTs might enhance our ability to identify those of autism, they are largely unstudied. Here, we have examined the neuroanatomical basis of ALTs and their association with the oxytocin receptor gene (OXTR) rs2254298A, a known risk allele for autism in Asian populations which has also been implicated in limbic–paralimbic brain structures. First, we extracted a four-factor structure of ALTs, as measured using the Autism-Spectrum Quotient, including ‘prosociality’, ‘communication’, ‘details/patterns’ and ‘imagination’ in 135 neurotypical adults (79 men, 56 women) to reduce the genetic heterogeneity of ALTs. Then, in the same population, voxel-based morphometry revealed that lower ‘prosociality’, which indicates strong ALTs, was significantly correlated to smaller regional grey matter volume in the right insula in males. Males with lower ‘prosociality’ also had less interregional structural coupling between the right insula and the ventral anterior cingulate cortex. Furthermore, males with OXTR rs2254298A had significantly smaller grey matter volume in the right insula. These results show that decreased volume of the insula is a neuroanatomical correlate of ALTs and a potential intermediate phenotype linking ALTs with OXTR in male subjects. PMID:23946005

  19. Allelic Variation in Developmental Genes and Effects on Winter Wheat Heading Date in the U.S. Great Plains.

    PubMed

    Grogan, Sarah M; Brown-Guedira, Gina; Haley, Scott D; McMaster, Gregory S; Reid, Scott D; Smith, Jared; Byrne, Patrick F

    2016-01-01

    Heading date in wheat (Triticum aestivum L.) and other small grain cereals is affected by the vernalization and photoperiod pathways. The reduced-height loci also have an effect on growth and development. Heading date, which occurs just prior to anthesis, was evaluated in a population of 299 hard winter wheat entries representative of the U.S. Great Plains region, grown in nine environments during 2011-2012 and 2012-2013. The germplasm was evaluated for candidate genes at vernalization (Vrn-A1, Vrn-B1, and Vrn-D1), photoperiod (Ppd-A1, Ppd-B1 and Ppd-D1), and reduced-height (Rht-B1 and Rht-D1) loci using polymerase chain reaction (PCR) and Kompetitive Allele Specific PCR (KASP) assays. Our objectives were to determine allelic variants known to affect flowering time, assess the effect of allelic variants on heading date, and investigate changes in the geographic and temporal distribution of alleles and haplotypes. Our analyses enhanced understanding of the roles developmental genes have on the timing of heading date in wheat under varying environmental conditions, which could be used by breeding programs to improve breeding strategies under current and future climate scenarios. The significant main effects and two-way interactions between the candidate genes explained an average of 44% of variability in heading date at each environment. Among the loci we evaluated, most of the variation in heading date was explained by Ppd-D1, Ppd-B1, and their interaction. The prevalence of the photoperiod sensitive alleles Ppd-A1b, Ppd-B1b, and Ppd-D1b has gradually decreased in U.S. Great Plains germplasm over the past century. There is also geographic variation for photoperiod sensitive and reduced-height alleles, with germplasm from breeding programs in the northern Great Plains having greater incidences of the photoperiod sensitive alleles and lower incidence of the semi-dwarf alleles than germplasm from breeding programs in the central or southern plains.

  20. Allelic Variation in Developmental Genes and Effects on Winter Wheat Heading Date in the U.S. Great Plains

    PubMed Central

    Brown-Guedira, Gina; Haley, Scott D.; McMaster, Gregory S.; Reid, Scott D.; Smith, Jared; Byrne, Patrick F.

    2016-01-01

    Heading date in wheat (Triticum aestivum L.) and other small grain cereals is affected by the vernalization and photoperiod pathways. The reduced-height loci also have an effect on growth and development. Heading date, which occurs just prior to anthesis, was evaluated in a population of 299 hard winter wheat entries representative of the U.S. Great Plains region, grown in nine environments during 2011–2012 and 2012–2013. The germplasm was evaluated for candidate genes at vernalization (Vrn-A1, Vrn-B1, and Vrn-D1), photoperiod (Ppd-A1, Ppd-B1 and Ppd-D1), and reduced-height (Rht-B1 and Rht-D1) loci using polymerase chain reaction (PCR) and Kompetitive Allele Specific PCR (KASP) assays. Our objectives were to determine allelic variants known to affect flowering time, assess the effect of allelic variants on heading date, and investigate changes in the geographic and temporal distribution of alleles and haplotypes. Our analyses enhanced understanding of the roles developmental genes have on the timing of heading date in wheat under varying environmental conditions, which could be used by breeding programs to improve breeding strategies under current and future climate scenarios. The significant main effects and two-way interactions between the candidate genes explained an average of 44% of variability in heading date at each environment. Among the loci we evaluated, most of the variation in heading date was explained by Ppd-D1, Ppd-B1, and their interaction. The prevalence of the photoperiod sensitive alleles Ppd-A1b, Ppd-B1b, and Ppd-D1b has gradually decreased in U.S. Great Plains germplasm over the past century. There is also geographic variation for photoperiod sensitive and reduced-height alleles, with germplasm from breeding programs in the northern Great Plains having greater incidences of the photoperiod sensitive alleles and lower incidence of the semi-dwarf alleles than germplasm from breeding programs in the central or southern plains. PMID:27058239

  1. Integrating the genetic and physical maps of Arabidopsis thaliana: identification of mapped alleles of cloned essential (EMB) genes.

    PubMed

    Meinke, David; Sweeney, Colleen; Muralla, Rosanna

    2009-10-08

    The classical genetic map of Arabidopsis includes more than 130 genes with an embryo-defective (emb) mutant phenotype. Many of these essential genes remain to be cloned. Hundreds of additional EMB genes have been cloned and catalogued (www.seedgenes.org) but not mapped. To facilitate EMB gene identification and assess the current level of saturation, we updated the classical map, compared the physical and genetic locations of mapped loci, and performed allelism tests between mapped (but not cloned) and cloned (but not mapped) emb mutants with similar chromosome locations. Two hundred pairwise combinations of genes located on chromosomes 1 and 5 were tested and more than 1100 total crosses were screened. Sixteen of 51 mapped emb mutants examined were found to be disrupted in a known EMB gene. Alleles of a wide range of published EMB genes (YDA, GLA1, TIL1, AtASP38, AtDEK1, EMB506, DG1, OEP80) were discovered. Two EMS mutants isolated 30 years ago, T-DNA mutants with complex insertion sites, and a mutant with an atypical, embryo-specific phenotype were resolved. The frequency of allelism encountered was consistent with past estimates of 500 to 1000 EMB loci. New EMB genes identified among mapped T-DNA insertion mutants included CHC1, which is required for chromatin remodeling, and SHS1/AtBT1, which encodes a plastidial nucleotide transporter similar to the maize Brittle1 protein required for normal endosperm development. Two classical genetic markers (PY, ALB1) were identified based on similar map locations of known genes required for thiamine (THIC) and chlorophyll (PDE166) biosynthesis. The alignment of genetic and physical maps presented here should facilitate the continued analysis of essential genes in Arabidopsis and further characterization of a broad spectrum of mutant phenotypes in a model plant.

  2. Wide allelic heterogeneity with predominance of large IDS gene complex rearrangements in a sample of Mexican patients with Hunter syndrome.

    PubMed

    Alcántara-Ortigoza, M A; García-de Teresa, B; González-Del Angel, A; Berumen, J; Guardado-Estrada, M; Fernández-Hernández, L; Navarrete-Martínez, J I; Maza-Morales, M; Rius-Domínguez, R

    2016-05-01

    Hunter syndrome or mucopolysaccharidosis type II (MPSII) is caused by pathogenic variants in the IDS gene. This is the first study that examines the mutational spectrum in 25 unrelated Mexican MPSII families. The responsible genotype was identified in 96% of the families (24/25) with 10 novel pathogenic variants: c.133G>C, c.1003C>T, c.1025A>C, c.463_464delinsCCGTATAGCTGG, c.754_767del, c.1132_1133del, c.1463del, c.508-1G>C, c.1006+1G>T and c.(-217_103del). Extensive IDS gene deletions were identified in four patients; using DNA microarray analysis two patients showed the loss of the entire AFF2 gene, and epilepsy developed in only one of them. Wide allelic heterogeneity was noted, with large gene alterations (e.g. IDS/IDSP1 gene inversions, partial to extensive IDS deletions, and one chimeric IDS-IDSP1 allele) that occurred at higher frequencies than previously reported (36% vs 18.9-29%). The frequency of carrier mothers (80%) is consistent with previous descriptions (>70%). Carrier assignment allowed molecular prenatal diagnoses. Notably, somatic and germline mosaicism was identified in one family, and two patients presented thrombocytopenic purpura and pancytopenia after idursulfase enzyme replacement treatment. Our findings suggest a wide allelic heterogeneity in Mexican MPSII patients; DNA microarray analysis contributes to further delineation of the resulting phenotype for IDS and neighboring loci deletions.

  3. Organ-specific gene expression in maize: The P-wr allele. Final report, August 15, 1993--August 14, 1996

    SciTech Connect

    Peterson, T.A.

    1997-06-01

    The ultimate aim of our work is to understand how a regulatory gene produces a specific pattern of gene expression during plant development. Our model is the P-wr gene of maize, which produces a distinctive pattern of pigmentation of maize floral organs. We are investigating this system using a combination of classical genetic and molecular approaches. Mechanisms of organ-specific gene expression are a subject of intense research interest, as it is the operation of these mechanisms during eukaryotic development which determine the characteristics of each organism Allele-specific expression has been characterized in only a few other plant genes. In maize, organ-specific pigmentation regulated by the R, B, and Pl genes is achieved by differential transcription of functionally conserved protein coding sequences. Our studies point to a strikingly different mechanism of organ-specific gene expression, involving post-transcriptional regulation of the regulatory P gene. The novel pigmentation pattern of the P-wr allele is associated with differences in the encoded protein. Furthermore, the P-wr gene itself is present as a unique tandemly amplified structure, which may affect its transcriptional regulation.

  4. Serum lipid levels and M/L55 allele distribution of HDL paraoxonase gene in Saami and Finnish men.

    PubMed

    Malin, R; Lehtinen, S; Luoma, P; Näyhä, S; Hassi, J; Koivula, T; Lehtimäki, T

    2001-01-01

    Paraoxonase (PON) is an antioxidative enzyme, which eliminates lipid peroxides. The mutation in codon 55 of PON1 gene causes a change of methionine (M-allele) to leucine (L-allele) and influences PON activity. The Saami are a population living in the northern part of Fennoscandia. In previous studies their death rate from coronary artery disease (CAD) was found to be low. We compared PON M/L55 allele frequencies of 68 Saami and 68 Finnish men and related the PON genotypes to plasma lipid levels and to the levels of autoantibodies against oxidized LDL. The M/L55 genotypes were determined by PCR and restriction enzyme digestion. ELISA was used to measure antibodies against oxidized LDL. The L- and M-allele frequencies were 64% and 36% in Saami population and 64% and 36% in Finnish men, respectively (p = NS, Fisher's exact test). There were also no significant differences in plasma lipid levels or in antibody levels against oxidized LDL between PON genotypes or between Saami and Finnish men. Our results indicate that the PON M/L55 genotype is not associated with plasma lipid levels or the levels of autoantibodies against oxidized LDL in these populations. The Saami men have the same PON M/L55 allele distribution as the Finnish men and the PON genotype might thus not be one factor protecting Saami against CAD.

  5. WRKY6 Transcription Factor Restricts Arsenate Uptake and Transposon Activation in Arabidopsis[W

    PubMed Central

    Castrillo, Gabriel; Sánchez-Bermejo, Eduardo; de Lorenzo, Laura; Crevillén, Pedro; Fraile-Escanciano, Ana; TC, Mohan; Mouriz, Alfonso; Catarecha, Pablo; Sobrino-Plata, Juan; Olsson, Sanna; Leo del Puerto, Yolanda; Mateos, Isabel; Rojo, Enrique; Hernández, Luis E.; Jarillo, Jose A.; Piñeiro, Manuel; Paz-Ares, Javier; Leyva, Antonio

    2013-01-01

    Stress constantly challenges plant adaptation to the environment. Of all stress types, arsenic was a major threat during the early evolution of plants. The most prevalent chemical form of arsenic is arsenate, whose similarity to phosphate renders it easily incorporated into cells via the phosphate transporters. Here, we found that arsenate stress provokes a notable transposon burst in plants, in coordination with arsenate/phosphate transporter repression, which immediately restricts arsenate uptake. This repression was accompanied by delocalization of the phosphate transporter from the plasma membrane. When arsenate was removed, the system rapidly restored transcriptional expression and membrane localization of the transporter. We identify WRKY6 as an arsenate-responsive transcription factor that mediates arsenate/phosphate transporter gene expression and restricts arsenate-induced transposon activation. Plants therefore have a dual WRKY-dependent signaling mechanism that modulates arsenate uptake and transposon expression, providing a coordinated strategy for arsenate tolerance and transposon gene silencing. PMID:23922208

  6. Wound induced tanscriptional regulation of benzylisoquinoline pathway and characterization of wound inducible PsWRKY transcription factor from Papaver somniferum.

    PubMed

    Mishra, Sonal; Triptahi, Vineeta; Singh, Seema; Phukan, Ujjal J; Gupta, M M; Shanker, Karuna; Shukla, Rakesh Kumar

    2013-01-01

    Wounding is required to be made in the walls of the green seed pod of Opium poppy prior exudation of latex. To withstand this kind of trauma plants regulate expression of some metabolites through an induced transcript level. 167 unique wound-inducible ESTs were identified by a repetitive round of cDNA subtraction after 5 hours of wounding in Papaver somniferum seedlings. Further repetitive reverse northern analysis of these ESTs revealed 80 transcripts showing more than two fold induction, validated through semi-quantitative RT-PCR & real time expression analysis. One of the major classified categories among identified ESTs belonged to benzylisoquinoline transcripts. Tissue specific metabolite analysis of benzylisoquinoline alkaloids (BIAs) in response to wounding revealed increased accumulation of narcotine and papaverine. Promoter analysis of seven transcripts of BIAs pathway showed the presence of W-box cis-element with the consensus sequence of TGAC, which is the proposed binding site for WRKY type transcription factors. One of the Wound inducible 'WRKY' EST isolated from our subtracted library was made full-length and named as 'PsWRKY'. Bacterially expressed PsWRKY interacted with the W-box element having consensus sequence TTGACT/C present in the promoter region of BIAs biosynthetic pathway genes. PsWRKY further activated the TYDC promoter in yeast and transiently in tobacco BY2 cells. Preferential expression of PsWRKY in straw and capsule and its interaction with consensus W-box element present in BIAs pathway gene transcripts suggest its possible involvement in the wound induced regulation of BIAs pathway.

  7. Genome-wide transcriptomic analysis reveals correlation between higher WRKY61 expression and reduced symptom severity in Turnip crinkle virus infected Arabidopsis thaliana

    PubMed Central

    Gao, Ruimin; Liu, Peng; Yong, Yuhan; Wong, Sek-Man

    2016-01-01

    Turnip crinkle virus (TCV) is a carmovirus that infects many Arabidopsis ecotypes. Most studies mainly focused on discovery of resistance genes against TCV infection, and there is no Next Generation Sequencing based comparative genome wide transcriptome analysis reported. In this study, RNA-seq based transcriptome analysis revealed that 238 (155 up-regulated and 83 down-regulated) significant differentially expressed genes with at least 15-fold change were determined. Fifteen genes (including upregulated, unchanged and downregulated) were selected for RNA-seq data validation using quantitative real-time PCR, which showed consistencies between these two sets of data. GO enrichment analysis showed that numerous terms such as stress, immunity, defence and chemical stimulus were affected in TCV-infected plants. One putative plant defence related gene named WRKY61 was selected for further investigation. It showed that WRKY61 overexpression plants displayed reduced symptoms and less virus accumulation, as compared to wild type (WT) and WRKY61 deficient lines, suggesting that higher WRKY61 expression level reduced TCV viral accumulation. In conclusion, our transcriptome analysis showed that global gene expression was detected in TCV-infected Arabidopsis thaliana. WRKY61 gene was shown to be negatively correlated with TCV infection and viral symptoms, which may be connected to plant immunity pathways. PMID:27086702

  8. Evidence for a genetic association between alleles of monoamine oxidase A gene and bipolar affective disorder

    SciTech Connect

    Lim, L.C.C.; Sham, P.; Castle, D.

    1995-08-14

    We present evidence of a genetic association between bipolar disorder and alleles at 3 monoamine oxidase A (MAOA) markers, but not with alleles of a monoamine oxidase B (MAOB) polymorphism. The 3 MAOA markers, including one associated with low MAOA activity, show strong allelic association with each other but surprisingly not with MAOB. Our results are significantly only for females, though the number of males in our sample is too small to draw any definite conclusions. Our data is consistent with recent reports of reduced MAOA activity in patients with abnormal behavioral phenotypes. The strength of the association is weak, but significant, which suggests that alleles at the MAOA locus contribute to susceptibility to bipolar disorder rather than being a major determinant. 58 refs., 1 fig., 3 tabs.

  9. Allele Mining and Selective Patterns of Pi9 Gene in a Set of Rice Landraces from India

    PubMed Central

    Imam, Jahangir; Mandal, Nimai P.; Variar, Mukund; Shukla, Pratyoosh

    2016-01-01

    Allelic variants of the broad-spectrum blast resistance gene, Pi9 (nucleotide binding site-leucine-rich repeat region) have been analyzed in Indian rice landraces. They were selected from the list of 338 rice landraces phenotyped in the rice blast nursery at central Rainfed Upland Rice Research Station, Hazaribag. Six of them were further selected on the basis of their resistance and susceptible pattern for virulence analysis and selective pattern study of Pi9 gene. The sequence analysis and phylogenetic study illustrated that such sequences are vastly homologous and clustered into two groups. All the blast resistance Pi9 alleles were grouped into one cluster, whereas Pi9 alleles of susceptible landraces formed another cluster even though these landraces have a low level of DNA polymorphisms. A total number of 136 polymorphic sites comprising of transitions, transversions, and insertion and deletions (InDels) were identified in the 2.9 kb sequence of Pi9 alleles. Lower variation in the form of mutations (77) (Transition + Transversion), and InDels (59) were observed in the Pi9 alleles isolated from rice landraces studied. The results showed that the Pi9 alleles of the selected rice landraces were less variable, suggesting that the rice landraces would have been exposed to less number of pathotypes across the country. The positive Tajima’s D (0.33580), P > 0.10 (not significant) was observed among the seven rice landraces, which suggests the balancing selection of Pi9 alleles. The value of synonymous substitution (-0.43337) was less than the non-synonymous substitution (0.78808). The greater non-synonymous substitution than the synonymous means that the coding region, mainly the leucine-rich repeat domain was under diversified selection. In this study, the Pi9 gene has been subjected to balancing selection with low nucleotide diversity which is different from the earlier reports, this may be because of the closeness of the rice landraces, cultivated in the same

  10. Variant alleles of the CYP1B1 gene are associated with colorectal cancer susceptibility

    PubMed Central

    2010-01-01

    Background CYP1B1 is a P450 enzyme which is involved in the activation of pro-carcinogens to carcinogens as well as sex hormone metabolism. Because differences in the activity of the enzyme have been correlated with variant alleles of single nucleotide polymorphisms (SNPs), it represents an attractive candidate gene for studies into colorectal cancer susceptibility. Methods We genotyped 597 cancer patients and 597controls for three CYP1B1 SNPs, which have previously been shown to be associated with altered enzymatic activity. Using the three SNPs, eight different haplotypes were constructed. The haplotype frequencies were estimated in cases and controls and then compared. The odds ratio for each tumour type, associated with each haplotype was estimated, with reference to the most common haplotype observed in the controls. Results The three SNPs rs10012, rs1056827 and rs1056836 alone did not provide any significant evidence of association with colorectal cancer risk. Haplotypes of rs1056827 and rs10012 or rs1056827 and rs1056836 revealed an association with colorectal cancer which was significantly stronger in the homozygous carriers. One haplotype was under represented in the colorectal cancer patient group compared to the control population suggesting a protective effect. Conclusion Genetic variants within the CYP1B1 that are associated with altered function appear to influence susceptibility to a colorectal cancer in Poland. Three haplotypes were associated with altered cancer risk; one conferred protection and two were associated with an increased risk of disease. These observations should be confirmed in other populations. PMID:20701755

  11. Epigenetic programming via histone methylation at WRKY53 controls leaf senescence in Arabidopsis thaliana.

    PubMed

    Ay, Nicole; Irmler, Kristina; Fischer, Andreas; Uhlemann, Ria; Reuter, Gunter; Humbeck, Klaus

    2009-04-01

    Leaf senescence, the final step of leaf development, involves extensive reprogramming of gene expression. Here, we show that these processes include discrete changes of epigenetic indexing, as well as global alterations in chromatin organization. During leaf senescence, the interphase nuclei show a decondensation of chromocenter heterochromatin, and changes in the nuclear distribution of the H3K4me2, H3K4me3, and the H3K27me2 and H3K27me3 histone modification marks that index active and inactive chromatin, respectively. Locus-specific epigenetic indexing was studied at the WRKY53 key regulator of leaf senescence. During senescence, when the locus becomes activated, H3K4me2 and H3K4me3 are significantly increased at the 5' end and at coding regions. Impairment of these processes is observed in plants overexpressing the SUVH2 histone methyltransferase, which causes ectopic heterochromatization. In these plants the transcriptional initiation of WRKY53 and of the senescence-associated genes SIRK, SAG101, ANAC083, SAG12 and SAG24 is inhibited, resulting in a delay of leaf senescence. In SUVH2 overexpression plants, significant levels of H3K27me2 and H3K27me3 are detected at the 5'-end region of WRKY53, resulting in its transcriptional repression. Furthermore, SUVH2 overexpression inhibits senescence-associated global changes in chromatin organization. Our data suggest that complex epigenetic processes control the senescence-specific gene expression pattern.

  12. The GM2 gangliosidoses databases: allelic variation at the HEXA, HEXB, and GM2A gene loci.

    PubMed

    Cordeiro, P; Hechtman, P; Kaplan, F

    2000-01-01

    The GM2 gangliosidoses are a group of recessive disorders characterized by accumulation of GM2 ganglioside in neuronal cells. The genes responsible for these disorders are HEXA (Tay-Sachs disease and variants), HEXB (Sandhoff disease and variants), and GM2A (AB variant of GM2 gangliosidosis). We report the establishment of three relational locus-specific databases recording allelic variation at the HEXA, HEXB, and GM2A genes and accessed at the GM2 gangliosidoses home page (http://data.mch.mcgill.ca/gm2-gangliosidoses). Submission forms are available for the addition of new mutations to the databases. The databases are available online for users to search and retrieve information about specific alleles by a number of fields describing mutations, phenotypes, or author(s).

  13. The WRKY45-Dependent Signaling Pathway Is Required For Resistance against Striga hermonthica Parasitism.

    PubMed

    Mutuku, J Musembi; Yoshida, Satoko; Shimizu, Takafumi; Ichihashi, Yasunori; Wakatake, Takanori; Takahashi, Akira; Seo, Mitsunori; Shirasu, Ken

    2015-07-01

    The root hemiparasite witchweed (Striga spp.) is a devastating agricultural pest that causes losses of up to $1 billion US annually in sub-Saharan Africa. Development of resistant crops is one of the cost-effective ways to address this problem. However, the molecular mechanisms underlying resistance are not well understood. To understand molecular events upon Striga spp. infection, we conducted genome-scale RNA sequencing expression analysis using Striga hermonthica-infected rice (Oryza sativa) roots. We found that transcripts grouped under the Gene Ontology term defense response were significantly enriched in up-regulated differentially expressed genes. In particular, we found that both jasmonic acid (JA) and salicylic acid (SA) pathways were induced, but the induction of the JA pathway preceded that of the SA pathway. Foliar application of JA resulted in higher resistance. The hebiba mutant plants, which lack the JA biosynthesis gene allene oxide cyclase, exhibited severe S. hermonthica susceptibility. The resistant phenotype was recovered by application of JA. By contrast, the SA-deficient NahG rice plants were resistant against S. hermonthica, indicating that endogenous SA is not required for resistance. However, knocking down WRKY45, a regulator of the SA/benzothiadiazole pathway, resulted in enhanced susceptibility. Interestingly, NahG plants induced the JA pathway, which was down-regulated in WRKY45-knockdown plants, linking the resistant and susceptible phenotypes to the JA pathway. Consistently, the susceptibility phenotype in the WRKY45-knockdown plants was recovered by foliar JA application. These results point to a model in which WRKY45 modulates a cross talk in resistance against S. hermonthica by positively regulating both SA/benzothiadiazole and JA pathways.

  14. The WRKY45-Dependent Signaling Pathway Is Required For Resistance against Striga hermonthica Parasitism1[OPEN

    PubMed Central

    Yoshida, Satoko; Takahashi, Akira; Seo, Mitsunori

    2015-01-01

    The root hemiparasite witchweed (Striga spp.) is a devastating agricultural pest that causes losses of up to $1 billion US annually in sub-Saharan Africa. Development of resistant crops is one of the cost-effective ways to address this problem. However, the molecular mechanisms underlying resistance are not well understood. To understand molecular events upon Striga spp. infection, we conducted genome-scale RNA sequencing expression analysis using Striga hermonthica-infected rice (Oryza sativa) roots. We found that transcripts grouped under the Gene Ontology term defense response were significantly enriched in up-regulated differentially expressed genes. In particular, we found that both jasmonic acid (JA) and salicylic acid (SA) pathways were induced, but the induction of the JA pathway preceded that of the SA pathway. Foliar application of JA resulted in higher resistance. The hebiba mutant plants, which lack the JA biosynthesis gene ALLENE OXIDE CYCLASE, exhibited severe S. hermonthica susceptibility. The resistant phenotype was recovered by application of JA. By contrast, the SA-deficient NahG rice plants were resistant against S. hermonthica, indicating that endogenous SA is not required for resistance. However, knocking down WRKY45, a regulator of the SA/benzothiadiazole pathway, resulted in enhanced susceptibility. Interestingly, NahG plants induced the JA pathway, which was down-regulated in WRKY45-knockdown plants, linking the resistant and susceptible phenotypes to the JA pathway. Consistently, the susceptibility phenotype in the WRKY45-knockdown plants was recovered by foliar JA application. These results point to a model in which WRKY45 modulates a cross talk in resistance against S. hermonthica by positively regulating both SA/benzothiadiazole and JA pathways. PMID:26025049

  15. Polymorphisms and allele frequencies of glutathione S-transferases A1 and P1 genes in the Polish population.

    PubMed

    Skrzypczak-Zielinska, M; Zakerska-Banaszak, O; Tamowicz, B; Sobieraj, I; Drweska-Matelska, N; Szalata, M; Slomski, R; Mikstacki, A

    2015-03-31

    Glutathione S-transferases (GST) A1 and P1 are crucial enzymes involved in the biotransformation of drugs, carcinogens, and toxins, and their activity may influence drug response, susceptibility to diseases, and carcinogenesis. The genes encoding these enzymes, GSTA1 and GSTP1, have been examined in many studies because of their genetic variability, which may affect enzymatic activity. The goal of this study was to determine the distribution of the alleles GSTA1*A/*B and GSTP1*A, *B, and *C in the Polish population. A total of 160 subjects from the Polish population were genotyped for 2 polymorphisms (I105V and A114V) in the GSTP1 gene using pyrosequencing. The promoter region of the GSTA1 gene was screened using sequencing. The detected variants were subjected to haplotype analysis. We found that the distribution of the alleles GSTA1*A/*B and GSTP1*A, *B, and *C in the Polish population correspond to the results of studies in Caucasians. Furthermore, we identified additional single nucleotide polymorphisms, excluding 3 well-known changes (G-52A, C-69T, T-567G), which are linked to alleles GSTA1*A/*B, that affect enzyme activity. A total of 4 haplotypes were identified in 160 Polish individuals.

  16. Sequenced Alleles of the Caenorhabditis Elegans Sex-Determining Gene Her-1 Include a Novel Class of Conditional Promoter Mutations

    PubMed Central

    Perry, M. D.; Trent, C.; Robertson, B.; Chamblin, C.; Wood, W. B.

    1994-01-01

    In the control of Caenorhabditis elegans sex determination, the her-1 gene must normally be activated to allow male development of XO animals and deactivated to allow hermaphrodite development of XX animals. The gene is regulated at the transcriptional level and has two nested male-specific transcripts. The larger of these encodes a small, novel, cysteine-rich protein responsible for masculinizing activity. Of the 32 extant mutant alleles, 30 cause partial or complete loss of masculinizing function (lf), while 2 are gain-of-function (gf) alleles resulting in abnormal masculinization of XX animals. We have identified the DNA sequence changes in each of these 32 alleles. Most affect the protein coding functions of the gene, but six are in the promoter region, including the two gf mutations. These two mutations may define a binding site for negative regulators of her-1. Three of the four remaining promoter mutations are single base changes that cause, surprisingly, temperature-sensitive loss of her-1 function. Such conditional promoter mutations have previously not been found among either prokaryotic or eukaryotic mutants analyzed at the molecular level. PMID:7828816

  17. Characterization of Opsin Gene Alleles Affecting Color Vision in a Wild Population of Titi Monkeys (Callicebus brunneus)

    PubMed Central

    Bunce, John A.; Isbell, Lynne A.; Neitz, Maureen; Bonci, Daniela; Surridge, Alison K.; Jacobs, Gerald H.; Smith, David Glenn

    2011-01-01

    The color vision of most platyrrhine primates is determined by alleles at the polymorphic X-linked locus coding for the opsin responsible for the middle- to long-wavelength (M/L) cone photopigment. Females who are heterozygous at the locus have trichromatic vision while homozygous females and all males are dichromatic. This study characterized the opsin alleles in a wild population of the socially monogamous platyrrhine monkey Callicebus brunneus (the brown titi monkey), a primate that an earlier study suggests may possess an unusual number of alleles at this locus and thus may be a subject of special interest in the study of primate color vision. Direct sequencing of regions of the M/L opsin gene using feces-, blood-, and saliva-derived DNA obtained from 14 individuals yielded evidence for the presence of three functionally distinct alleles, corresponding to the most common M/L photopigment variants inferred from a physiological study of cone spectral sensitivity in captive Callicebus. PMID:20938927

  18. CaCDPK15 positively regulates pepper responses to Ralstonia solanacearum inoculation and forms a positive-feedback loop with CaWRKY40 to amplify defense signaling

    PubMed Central

    Shen, Lei; Yang, Sheng; Yang, Tong; Liang, Jiaqi; Cheng, Wei; Wen, Jiayu; Liu, Yanyan; Li, Jiazhi; Shi, Lanping; Tang, Qian; Shi, Wei; Hu, Jiong; Liu, Cailing; Zhang, Yangwen; Mou, Shaoliang; Liu, Zhiqin; Cai, Hanyang; He, Li; Guan, Deyi; Wu, Yang; He, Shuilin

    2016-01-01

    CaWRKY40 is a positive regulator of pepper (Capsicum annum) response to Ralstonia solanacearum inoculation (RSI), but the underlying mechanism remains largely unknown. Here, we functionally characterize CaCDPK15 in the defense signaling mediated by CaWRKY40. Pathogen-responsive TGA, W, and ERE boxes were identified in the CaCDPK15 promoter (pCaCDPK15), and pCaCDPK15-driven GUS expression was significantly enhanced in response to RSI and exogenously applied salicylic acid, methyl jasmonate, abscisic acid, and ethephon. Virus-induced gene silencing (VIGS) of CaCDPK15 significantly increased the susceptibility of pepper to RSI and downregulated the immunity-associated markers CaNPR1, CaPR1, and CaDEF1. By contrast, transient CaCDPK15 overexpression significantly activated hypersensitive response associated cell death, upregulated the immunity-associated marker genes, upregulated CaWRKY40 expression, and enriched CaWRKY40 at the promoters of its targets genes. Although CaCDPK15 failed to interact with CaWRKY40, the direct binding of CaWRKY40 to pCaCDPK15 was detected by chromatin immunoprecipitation, which was significantly potentiated by RSI in pepper plants. These combined results suggest that RSI in pepper induces CaCDPK15 and indirectly activates downstream CaWRKY40, which in turn potentiates CaCDPK15 expression. This positive-feedback loop would amplify defense signaling against RSI and efficiently activate strong plant immunity. PMID:26928570

  19. Allele-specific gene expression patterns in primary leukemic cells reveal regulation of gene expression by CpG site methylation

    PubMed Central

    Milani, Lili; Lundmark, Anders; Nordlund, Jessica; Kiialainen, Anna; Flaegstad, Trond; Jonmundsson, Gudmundur; Kanerva, Jukka; Schmiegelow, Kjeld; Gunderson, Kevin L.; Lönnerholm, Gudmar; Syvänen, Ann-Christine

    2009-01-01

    To identify genes that are regulated by cis-acting functional elements in acute lymphoblastic leukemia (ALL) we determined the allele-specific expression (ASE) levels of 2529 genes by genotyping a genome-wide panel of single nucleotide polymorphisms in RNA and DNA from bone marrow and blood samples of 197 children with ALL. Using a reproducible, quantitative genotyping method and stringent criteria for scoring ASE, we found that 16% of the analyzed genes display ASE in multiple ALL cell samples. For most of the genes, the level of ASE varied largely between the samples, from 1.4-fold overexpression of one allele to apparent monoallelic expression. For genes exhibiting ASE, 55% displayed bidirectional ASE in which overexpression of either of the two SNP alleles occurred. For bidirectional ASE we also observed overall higher levels of ASE and correlation with the methylation level of these sites. Our results demonstrate that CpG site methylation is one of the factors that regulates gene expression in ALL cells. PMID:18997001

  20. Genetic basis for the hierarchical interaction between Tobamovirus spp. and L resistance gene alleles from different pepper species.

    PubMed

    Tomita, Reiko; Sekine, Ken-Taro; Mizumoto, Hiroyuki; Sakamoto, Masaru; Murai, Jun; Kiba, Akinori; Hikichi, Yasufumi; Suzuki, Kazumi; Kobayashi, Kappei

    2011-01-01

    The pepper L gene conditions the plant's resistance to Tobamovirus spp. Alleles L(1), L(2), L(3), and L(4) confer a broadening spectra of resistance to different virus pathotypes. In this study, we report the genetic basis for the hierarchical interaction between L genes and Tobamovirus pathotypes. We cloned L(3) using map-based methods, and L(1), L(1a), L(1c), L(2), L(2b), and L(4) using a homology-based method. L gene alleles encode coiled-coil, nucleotide-binding, leucine-rich repeat (LRR)-type resistance proteins with the ability to induce resistance response to the viral coat protein (CP) avirulence effectors by themselves. Their different recognition spectra in original pepper species were reproduced in an Agrobacterium tumefaciens-mediated transient expression system in Nicotiana benthamiana. Chimera analysis with L(1), which showed the narrowest recognition spectrum, indicates that the broader recognition spectra conferred by L(2), L(2b), L(3), and L(4) require different subregions of the LRR domain. We identified a critical amino acid residue for the determination of recognition spectra but other regions also influenced the L genes' resistance spectra. The results suggest that the hierarchical interactions between L genes and Tobamovirus spp. are determined by the interaction of multiple subregions of the LRR domain of L proteins with different viral CP themselves or some protein complexes including them.

  1. Several different lactase persistence associated alleles and high diversity of the lactase gene in the admixed Brazilian population.

    PubMed

    Friedrich, Deise C; Santos, Sidney E B; Ribeiro-dos-Santos, Ândrea K C; Hutz, Mara H

    2012-01-01

    Adult-type hypolactasia is a common phenotype caused by the lactase enzyme deficiency. The -13910 C>T polymorphism, located 14 Kb upstream of the lactase gene (LCT) in the MCM6 gene was associated with lactase persistence (LP) in Europeans. This polymorphism is rare in Africa but several other variants associated with lactase persistence were observed in Africans. The aims of this study were to identify polymorphisms in the MCM6 region associated with the lactase persistence phenotype and to determine the distribution of LCT gene haplotypes in 981 individuals from North, Northeast and South Brazil. These polymorphisms were genotyped by PCR based methods and sequencing. The -13779*C,-13910*T, -13937*A, -14010*C, -14011*T LP alleles previously described in the MCM6 gene region that acts as an enhancer for the LCT gene were identified in Brazilians. The most common LP allele was -13910*T. Its frequency was highly correlated with European ancestry in the Brazilian populations investigated. The -13910*T was higher (0.295) in southern Brazilians of European ancestry and lower (0.175) in the Northern admixed population. LCT haplotypes were derived from the 10 LCT SNPs genotyped. Overall twenty six haplotypes previously described were identified in the four Brazilian populations studied. The Multidimensional Scaling analysis showed that Belém, in the north, was closer to Amerindians. Northeastern and southern Afro-descendants were more related with Bantu-speaking South Africans whereas the Southern population with European ancestry grouped with Southern and Northern Europeans. This study shows a high variability considering the number of LCT haplotypes observed. Due to the highly admixed nature of the Brazilian populations, the diagnosis of hypolactasia in Brazil, based only in the investigation of the -13910*T allele is an oversimplification.

  2. Two bi-allelic single nucleotide polymorphisms within the promoter region of the horse tumour necrosis factor alpha gene.

    PubMed

    Matiasovic, J; Lukeszová, L; Horín, P

    2002-08-01

    Primers based on GenBank sequences within the 5' untranslated region (UTR) of the human and horse tumour necrosis factor alpha (TNF-alpha) genes were designed and used to amplify a 522-bp product. Sequencing of five clones derived from five independent PCRs obtained from three different animals of three different breeds (Old Kladruber, Akhal-Teke and Shetland Pony) revealed a high level of sequence identity to the TNF-alpha promoter regions of other species. The existing GenBank horse sequences were confirmed and extended upstream by 230 nucleotides. Based on the sequence obtained, a new horse-specific forward primer was designed to amplify a 213-bp PCR product, which was screened for polymorphism using single-strand conformation polymorphism (SSCP). Three allelic variants of the horse TNF-alpha gene were identified and sequenced (GenBank accession numbers ADF 349558-60). Two single nucleotide polymorphisms explained the existence of the three SSCP alleles detected: C/T and T/C single base pair substitutions at positions 137 and 147, respectively. Differences in allelic frequencies between Old Kladruber and Akhal-Teke breeds were observed.

  3. What phylogeny and gene genealogy analyses reveal about homoplasy in citrus microsatellite alleles

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Sixty-five microsatellite alleles from three Simple Sequence Repeat (SSR) loci (cAGG9, CCT01 and GT03) of various Citrus, Fortunella or Poncirus accessions were cloned and sequenced to determine their mode of evolution. This data was used to assess sequence variation by calculating the average numb...

  4. Human-specific derived alleles of CD33 and other genes protect against postreproductive cognitive decline

    PubMed Central

    Schwarz, Flavio; Springer, Stevan A.; Altheide, Tasha K.; Varki, Nissi M.; Gagneux, Pascal; Varki, Ajit

    2016-01-01

    The individuals of most vertebrate species die when they can no longer reproduce. Humans are a rare exception, having evolved a prolonged postreproductive lifespan. Elders contribute to cooperative offspring care, assist in foraging, and communicate important ecological and cultural knowledge, increasing the survival of younger individuals. Age-related deterioration of cognitive capacity in humans compromises these benefits and also burdens the group with socially costly members. We investigated the contribution of the immunoregulatory receptor CD33 to a uniquely human postreproductive disease, Alzheimer’s dementia. Surprisingly, even though selection at advanced age is expected to be weak, a CD33 allele protective against Alzheimer’s disease is derived and unique to humans and favors a functional molecular state of CD33 resembling that of the chimpanzee. Thus, derived alleles may be compensatory and restore interactions altered as a consequence of human-specific brain evolution. We found several other examples of derived alleles at other human loci that protect against age-related cognitive deterioration arising from neurodegenerative disease or cerebrovascular insufficiency. Selection by inclusive fitness may be strong enough to favor alleles protecting specifically against cognitive decline in postreproductive humans. Such selection would operate by maximizing the contributions of postreproductive individuals to the fitness of younger kin. PMID:26621708

  5. Characterization of allele-specific expression of the X-linked gene MAO-A in trophectoderm cells of bovine embryos produced by somatic cell nuclear transfer.

    PubMed

    Ferreira, A R; Aguiar Filho, L F C; Sousa, R V; Sartori, R; Franco, M M

    2015-10-05

    Somatic cell nuclear transfer (SCNT) may affect epigenetic mechanisms and alter the expression of genes related to embryo development and X chromosome inactivation (XCI). We characterized allele-specific expression of the X-linked gene monoamine oxidase type A (MAO-A) in the trophectoderm (TF) of embryos produced by SCNT. Total RNA was isolated from individual biopsies (N = 25), and the allele-specific expression assessed by reverse transcription-polymerase chain reaction-restriction fragment length polymorphism. Both paternal and maternal alleles were expressed in the trophectoderm. However, a higher frequency of the mono-allelic expression of a specific allele was observed (N = 17; 68%), with the remaining samples showing the presence of mRNA from both alleles (N = 8; 32%). Considering that MAO-A is subject to XCI in bovine, our results suggest that SCNT may influence XCI because neither an imprinted (mono-allelic expression in all samples) nor a random (presence of mRNA from both alleles in all samples) pattern of XCI was observed in TF. Due to the importance of XCI in mammalian embryo development and its sensitivity to in vitro conditions, X-linked genes subject to XCI are candidates for use in the development of embryo quality molecular markers for assisted reproduction.

  6. Increased brain activity to unpleasant stimuli in individuals with the 7R allele of the DRD4 gene.

    PubMed

    Gehricke, Jean-G; Swanson, James M; Duong, Sophie; Nguyen, Jenny; Wigal, Timothy L; Fallon, James; Caburian, Cyrus; Muftuler, Lutfi Tugan; Moyzis, Robert K

    2015-01-30

    The aim of the study was to examine functional brain activity in response to unpleasant images in individuals with the 7-repeat (7R) allele compared to individuals with the 4-repeat (4R) allele of the dopamine receptor D4 (DRD4) gene (VNTR in exon 3). Based on the response ready hypothesis, individuals with the DRD4-4R/7R genotype were expected to show greater functional brain activity in response to unpleasant compared to neutral stimuli in specific regions of the frontal, temporal, parietal and limbic lobes, which form the networks involved in attentional, emotional, and preparatory responses. Functional Magnetic Resonance Imaging activity was studied in 26 young adults (13 with the DRD4-4R/7R genotype and 13 with the DRD4-4R/4R genotype). Participants were asked to look at and subjectively rate unpleasant and neutral images. Results showed increased brain activity in response to unpleasant images compared to neutral images in the right temporal lobe in participants with the DRD4-4R/7R genotype versus participants with the DRD4-4R/4R genotype. The increase in right temporal lobe activity in individuals with DRD4-4R/7R suggests greater involvement in processing negative emotional stimuli. Intriguingly, no differences were found between the two genotypes in the subjective ratings of the images. The findings corroborate the response ready hypothesis, which suggests that individuals with the 7R allele are more responsive to negative emotional stimuli compared to individuals with the 4R allele of the DRD4 gene.

  7. The role of allelic variation in estrogen receptor genes and major depression in the Nurses Health Study

    PubMed Central

    Keyes, K.; Agnew-Blais, J.; Roberts, A.; Hamilton, A.; De Vivo, I.; Ranu, H.; Koenen, K.

    2015-01-01

    Purpose The role of exogenous and endogenous sex hormones in the etiology of depression remains elusive, in part because sex hormone variation is often correlated with behaviors, life stage changes, and other factors that may influence depression. Estrogen receptor alpha (ESR1) and beta (ESR2) are known to regulate gene expression and estrogen response in areas of the brain associated with major depression and are unlikely to be correlated with exogenous factors that may influence depression. Methods We examined whether functional polymorphisms in these genes are associated with lifetime major depression and chronic major depression among a sample of women from the Nurses’ Health Study II (N=2,576). DSM-IV depressive disorder symptoms were assessed by structured interview in 2007. Genotyping was performed on DNA extracted from blood using Taq-man. Results Women with the AA alleles of ESR2 RS4986938 had the higher prevalence of lifetime major depression than women with other allele frequencies (36.7% for those with AA versus 28.5% with GA and 29.1% with GG, p=0.02) and chronic major depression (14.7% for those with AA versus 9.3% with GA and 9.1 % with GG, p=0.01). History of post-menopausal hormone (PMH) use modified the association of ESR1 polymorphism RS2234693 with any lifetime depression; specifically, those with the TT allele had the highest risk of lifetime depression among PMH users, and the lowest risk of depression among non-PMH users (p-value for interaction=0.02). Further, carriers of the AA alleles in ESR1 polymorphism RS9340799 had increased prevalence of lifetime major depression only among lifetime PMH-users (p=0.007). Conclusions Our findings support the hypothesis that estrogen receptor polymorphisms influence risk for major depression; the role of estrogen receptors and other sex steroid-related genetic factors may provide unique insights into etiology. PMID:26169989

  8. Allele variations in the OCA2 gene (pink-eyed-dilution locus) are associated with genetic susceptibility to melanoma.

    PubMed

    Jannot, Anne-Sophie; Meziani, Roubila; Bertrand, Guylene; Gérard, Benedicte; Descamps, Vincent; Archimbaud, Alain; Picard, Catherine; Ollivaud, Laurence; Basset-Seguin, Nicole; Kerob, Delphine; Lanternier, Guy; Lebbe, Celeste; Saiag, P; Crickx, Beatrice; Clerget-Darpoux, Françoise; Grandchamp, Bernard; Soufir, Nadem; Melan-Cohort

    2005-08-01

    The occuloalbinism 2 (OCA2) gene, localized at 15q11, encodes a melanosomal transmembrane protein that is involved in the most common form of human occulo-cutaneous albinism, a human genetic disorder characterized by fair pigmentation and susceptibility to skin cancer. We wondered whether allele variations at this locus could influence susceptibility to malignant melanoma (MM). In all, 10 intragenic single-nucleotide polymorphisms (SNPs) were genotyped in 113 patients with melanomas and in 105 Caucasian control subjects with no personal or family history of skin cancer. By comparing allelic distribution between cases and controls, we show that MM and OCA2 are associated (p value=0.030 after correction for multiple testing). Then, a recently developed strategy, the 'combination test' enabled us to show that a combination formed by two SNPs was most strongly associated to MM, suggesting a possible interaction between intragenic SNPs. In addition, the role of OCA2 on MM risk was also detected using a logistic model taking into account the presence of variants of the melanocortin 1 receptor gene (MC1R, a key pigmentation gene) and all pigmentation characteristics as melanoma risk factors. Our data demonstrate that a second pigmentation gene, in addition to MC1R, is involved in genetic susceptibility to melanoma.

  9. Nucleotide sequence of the wild-type RAD4 gene of Saccharomyces cerevisiae and characterization of mutant rad4 alleles.

    PubMed Central

    Couto, L B; Friedberg, E C

    1989-01-01

    Shuttle plasmids carrying the wild-type RAD4 gene of Saccharomyces cerevisiae cannot be propagated in Escherichia coli (R. Fleer, W. Siede, and E. C. Friedberg, J. Bacteriol. 169:4884-4892, 1987). In order to determine the nucleotide sequence of the cloned gene, we used a plasmid carrying a mutant allele that allows plasmid propagation in E. coli. The wild-type sequence in the region of this mutation was determined from a second plasmid carrying a different mutant rad4 allele. We established the locations and characteristics of a number of spontaneously generated plasmid-borne RAD4 mutations that alleviate the toxicity of the wild-type gene in E. coli and of several mutagen-induced chromosomal mutations that inactivate the excision repair function of RAD4. These mutations are situated in very close proximity to each other, and all are expected to result in the expression of truncated polypeptides missing the carboxy-terminal one-third of the Rad4 polypeptide. This region of the gene may be important both for the toxic effect of the Rad4 protein in E. coli and for its role in DNA repair in S. cerevisiae. PMID:2649477

  10. Powerful Identification of Cis-regulatory SNPs in Human Primary Monocytes Using Allele-Specific Gene Expression

    PubMed Central

    Almlöf, Jonas Carlsson; Lundmark, Per; Lundmark, Anders; Ge, Bing; Maouche, Seraya; Göring, Harald H. H.; Liljedahl, Ulrika; Enström, Camilla; Brocheton, Jessy; Proust, Carole; Godefroy, Tiphaine; Sambrook, Jennifer G.; Jolley, Jennifer; Crisp-Hihn, Abigail; Foad, Nicola; Lloyd-Jones, Heather; Stephens, Jonathan; Gwilliam, Rhian; Rice, Catherine M.; Hengstenberg, Christian; Samani, Nilesh J.; Erdmann, Jeanette; Schunkert, Heribert; Pastinen, Tomi; Deloukas, Panos; Goodall, Alison H.; Ouwehand, Willem H.; Cambien, François; Syvänen, Ann-Christine

    2012-01-01

    A large number of genome-wide association studies have been performed during the past five years to identify associations between SNPs and human complex diseases and traits. The assignment of a functional role for the identified disease-associated SNP is not straight-forward. Genome-wide expression quantitative trait locus (eQTL) analysis is frequently used as the initial step to define a function while allele-specific gene expression (ASE) analysis has not yet gained a wide-spread use in disease mapping studies. We compared the power to identify cis-acting regulatory SNPs (cis-rSNPs) by genome-wide allele-specific gene expression (ASE) analysis with that of traditional expression quantitative trait locus (eQTL) mapping. Our study included 395 healthy blood donors for whom global gene expression profiles in circulating monocytes were determined by Illumina BeadArrays. ASE was assessed in a subset of these monocytes from 188 donors by quantitative genotyping of mRNA using a genome-wide panel of SNP markers. The performance of the two methods for detecting cis-rSNPs was evaluated by comparing associations between SNP genotypes and gene expression levels in sample sets of varying size. We found that up to 8-fold more samples are required for eQTL mapping to reach the same statistical power as that obtained by ASE analysis for the same rSNPs. The performance of ASE is insensitive to SNPs with low minor allele frequencies and detects a larger number of significantly associated rSNPs using the same sample size as eQTL mapping. An unequivocal conclusion from our comparison is that ASE analysis is more sensitive for detecting cis-rSNPs than standard eQTL mapping. Our study shows the potential of ASE mapping in tissue samples and primary cells which are difficult to obtain in large numbers. PMID:23300628

  11. Incorporating allelic variation for reconstructing the evolutionary history of organisms from multiple genes: An example from Rosa in North America.

    PubMed

    Joly, Simon; Bruneau, Anne

    2006-08-01

    Allelic variation within individuals holds information regarding the relationships of organisms, which is expected to be particularly important for reconstructing the evolutionary history of closely related taxa. However, little effort has been committed to incorporate such information for reconstructing the phylogeny of organisms. Haplotype trees represent a solution when one nonrecombinant marker is considered, but there is no satisfying method when multiple genes are to be combined. In this paper, we propose an algorithm that converts a distance matrix of alleles to a distance matrix among organisms. This algorithm allows the incorporation of allelic variation for reconstructing the phylogeny of organisms from one or more genes. The method is applied to reconstruct the phylogeny of the seven native diploid species of Rosa sect. Cinnamomeae in North America. The glyceralgehyde 3-phosphate dehydrogenase (GAPDH), the triose phosphate isomerase (TPI), and the malate synthase (MS) genes were sequenced for 40 individuals from these species. The three genes had little genetic variation, and most species showed incomplete lineage sorting, suggesting these species have a recent origin. Despite these difficulties, the networks (NeighborNet) of organisms reconstructed from the matrix obtained with the algorithm recovered groups that more closely match taxonomic boundaries than did the haplotype trees. The combined network of individuals shows that species west of the Rocky Mountains, Rosa gymnocarpa and R. pisocarpa, form exclusive groups and that together they are distinct from eastern species. In the east, three groups were found to be exclusive: R. nitida-R. palustris, R. foliolosa, and R. blanda-R. woodsii. These groups are congruent with the morphology and the ecology of species. The method is also useful for representing hybrid individuals when the relationships are reconstructed using a phylogenetic network.

  12. The "putative" role of transcription factors from HlWRKY family in the regulation of the final steps of prenylflavonid and bitter acids biosynthesis in hop (Humulus lupulus L.).

    PubMed

    Matoušek, Jaroslav; Kocábek, Tomáš; Patzak, Josef; Bříza, Jindřich; Siglová, Kristýna; Mishra, Ajay Kumar; Duraisamy, Ganesh Selvaraj; Týcová, Anna; Ono, Eiichiro; Krofta, Karel

    2016-10-01

    Lupulin glands localized in female hop (Humulus lupulus L.) cones are valuable source of bitter acids, essential oils and polyphenols. These compounds are used in brewing industry and are important for biomedical applications. In this study we describe the potential effect of transcription factors from WRKY family in the activation of the final steps of lupulin biosynthesis. In particular, lupulin gland-specific transcription factor HlWRKY1 that shows significant similarity to AtWRKY75, has ability to activate the set of promoters driving key genes of xanthohumol and bitter acids biosynthesis such as chalcone synthase H1, valerophenone synthase, prenyltransferase 1, 1L and 2 and O-methyltransferase-1. When combined with co-factor HlWDR1 and silencing suppressor p19, HlWRKY1 is able to enhance transient expression of gus gene driven by Omt1 and Chs_H1 promoters to significant level as compared to 35S promoter of CaMV in Nicotiana. benthamiana. Transformation of hop with dual Agrobacterium vector bearing HlWRKY1/HlWDR1 led to ectopic overexpression of these transgenes and further activation of lupulin-specific genes expression in hop leaves. It was further showed that (1) HlWRKY1 is endowed with promoter autoactivation; (2) It is regulated by post-transcriptional gene silencing (PTGS) mechanism; (3) It is stimulated by kinase co-expression. Since HlWRKY1 promotes expression of lupulin-specific HlMyb3 gene therefore it can constitute a significant component in hop lupulin regulation network. Putative involvement of HlWRKY1 in the regulation of lupulin biosynthesis may suggest the original physiological function of lupulin components in hop as flower and seed protective compounds.

  13. Advancing allele group-specific amplification of the complete HLA-C gene--isolation of novel alleles from three allele groups (C*04, C*07 and C*08).

    PubMed

    Cisneros, E; Martínez-Pomar, N; Vilches, M; Martín, P; de Pablo, R; Nuñez Del Prado, N; Nieto, A; Matamoros, N; Moraru, M; Vilches, C

    2013-10-01

    A variety of strategies have been designed for sequence-based HLA typing (SBT) and for the isolation of new human leucocyte antigen (HLA) alleles, but unambiguous characterization of complete genomic sequences remains a challenge. We recently reported a simple method for the group-specific amplification (GSA) and sequencing of a full-length C*04 genomic sequence in isolation from the accompanying allele. Here we build on this strategy and present homologous methods that enable the isolation of HLA-C alleles belonging to another two allele groups. Using this approach, which can be applied to sequence-based typing in some clinical settings, we have successfully characterized three novel HLA-C alleles (C*04:128, C*07:01:01:02, and C*08:62).

  14. Increased root hair density by loss of WRKY6 in Arabidopsis thaliana

    PubMed Central

    Benz, Martin

    2017-01-01

    Root hairs are unicellular elongations of certain rhizodermal cells that improve the uptake of sparingly soluble and immobile soil nutrients. Among different Arabidopsis thaliana genotypes, root hair density, length and the local acclimation to low inorganic phosphate (Pi) differs considerably, when analyzed on split agar plates. Here, genome-wide association fine mapping identified significant single nucleotide polymorphisms associated with the increased root hair density in the absence of local phosphate on chromosome 1. A loss-of-functionmutant of the candidate transcription factor gene WRKY6, which is involved in the acclimation of plants to low phosphorus, had increased root hair density. This is partially explained by a reduced cortical cell diameter in wrky6-3, reducing the rhizodermal cell numbers adjacent to the cortical cells. As a consequence, rhizodermal cells in positions that are in contact with two cortical cells are found more often, leading to higher hair density. Distinct cortical cell diameters and epidermal cell lengths distinguish other Arabidopsis accessions with distinct root hair density and −Pi response from diploid Col-0, while tetraploid Col-0 had generally larger root cell sizes, which explain longer hairs. A distinct radial root morphology within Arabidopsis accessions and wrky6-3explains some, but not all, differences in the root hair acclimation to –Pi. PMID:28149680

  15. Characterization of the dwg mutations: dwg and dwg(Bayer) are new mutant alleles of the Ggt1 gene.

    PubMed

    Tsuji, Takehito; Yamada, Kaoru; Kunieda, Tetsuo

    2009-01-01

    The dwg and dwg (Bayer) are allelic mutations of the mouse that are characterized by dwarfism, cataracts, and coat color change in homozygotes. The Ggt1 gene encodes gamma-glutamyltransferase 1 (GGT1), an extracellular membrane-bound enzyme that is critical for glutathione homeostasis. Both the dwg locus and Ggt1 gene are localized on mouse chromosome 10, and the phenotypes of GGT1-deficient mice with targeted disruption of the Ggt1 gene show remarkable similarities with those of dwg/dwg and dwg (Bayer)/dwg (Bayer) mice. This evidence led us to hypothesize that the Ggt1 gene is responsible for dwg and dwg (Bayer) mutations. In this study we characterized dwg mutations by investigating their association with the Ggt1 gene. Histological analysis revealed reduced numbers of proliferative and hypertrophic chondrocytes in the growth plate of dwg/dwg mice, which are characteristic abnormalities observed in GGT1-deficient mice. To identify the causative mutations of dwg mutations, we analyzed the Ggt1 gene in dwg/dwg and dwg (Bayer)/dwg (Bayer) mice. In dwg/dwg mice, 13 nucleotides on exon 7 of the Ggt1 gene were deleted, resulting in the generation of aberrant transcripts due to disrupted pre-mRNA splicing. Furthermore, dwg (Bayer)/dwg (Bayer) mice had a 46.7-kb deletion containing complete coding sequences of Ggt1 and AI646023 genes and the first exon of the Ggt5 gene, which is closely related to the Ggt1 gene as a member of the GGT gene family. These results indicate that both dwg and dwg (Bayer) have defective mutations of the Ggt1 gene. Thus, we concluded that mutations in the Ggt1 gene are responsible for the phenotypes of dwg/dwg and dwg (Bayer)/dwg (Bayer) mice.

  16. Plasminogen Activator Inhibitor-1 (PAI-1) gene 4G/5G alleles frequency distribution in the Lebanese population.

    PubMed

    Shammaa, Dina M R; Sabbagh, Amira S; Taher, Ali T; Zaatari, Ghazi S; Mahfouz, Rami A R

    2008-09-01

    Plasminogen activator inhibitor-1 (PAI-1) is an inhibitor of fibrinolysis. Increased plasma PAI-1 levels play an essential role in the pathogenesis of cardiovascular risk and other diseases associated with thrombosis. The 4G/5G polymorphism of the PAI-1 promoter region has been extensively studied in different populations. We studied 160 healthy unrelated Lebanese individuals using a reverse hybridization PCR assay to detect the 5G/5G, 4G/5G and, 4G/4G genotypes of the PAI-1 gene and the frequencies of the 4G and 5G alleles. We found that 4G/5G genotype was the most prevalent (45.6%) followed by 5G/5G (36.9%) and 4G/4G (17.5%). The frequencies of the 4G and 5G alleles were calculated to be 0.403 and 0.597, respectively. Compared to other ethnic communities, the Lebanese population was found to harbour a relatively high prevalence of the rare 4G allele. This, in turn, may predispose this population to develop cardiovascular diseases and other thrombotic clinical conditions. This study aids to enhance our understanding of the genetic features of the Lebanese population.

  17. WRKY Transcription Factors Phosphorylated by MAPK Regulate a Plant Immune NADPH Oxidase in Nicotiana benthamiana[OPEN

    PubMed Central

    Adachi, Hiroaki; Nakano, Takaaki; Miyagawa, Noriko; Ishihama, Nobuaki; Yoshioka, Miki; Katou, Yuri; Yaeno, Takashi

    2015-01-01

    Pathogen attack sequentially confers pattern-triggered immunity (PTI) and effector-triggered immunity (ETI) after sensing of pathogen patterns and effectors by plant immune receptors, respectively. Reactive oxygen species (ROS) play pivotal roles in PTI and ETI as signaling molecules. Nicotiana benthamiana RBOHB, an NADPH oxidase, is responsible for both the transient PTI ROS burst and the robust ETI ROS burst. Here, we show that RBOHB transactivation mediated by MAPK contributes to R3a/AVR3a-triggered ETI (AVR3a-ETI) ROS burst. RBOHB is markedly induced during the ETI and INF1-triggered PTI (INF1-PTI), but not flg22-tiggered PTI (flg22-PTI). We found that the RBOHB promoter contains a functional W-box in the R3a/AVR3a and INF1 signal-responsive cis-element. Ectopic expression of four phospho-mimicking mutants of WRKY transcription factors, which are MAPK substrates, induced RBOHB, and yeast one-hybrid analysis indicated that these mutants bind to the cis-element. Chromatin immunoprecipitation assays indicated direct binding of the WRKY to the cis-element in plants. Silencing of multiple WRKY genes compromised the upregulation of RBOHB, resulting in impairment of AVR3a-ETI and INF1-PTI ROS bursts, but not the flg22-PTI ROS burst. These results suggest that the MAPK-WRKY pathway is required for AVR3a-ETI and INF1-PTI ROS bursts by activation of RBOHB. PMID:26373453

  18. ABO3, a WRKY transcription factor, mediates plant responses to abscisic acid and drought tolerance in Arabidopsis

    PubMed Central

    Ren, Xiaozhi; Chen, Zhizhong; Liu, Yue; Zhang, Hairong; Zhang, Min; Liu, Qian; Hong, Xuhui; Zhu, Jian-Kang; Gong, Zhizhong

    2011-01-01

    SUMMARY The biological functions of WRKY transcription factors in plants have been widely studied, but their roles in abiotic stress are still not well understood. We isolated an ABA overly sensitive mutant, abo3, which is disrupted by a T-DNA insertion in At1g66600 encoding a WRKY transcription factor AtWRKY63. The mutant was hypersensitive to ABA in both seedling establishment and seedling growth. However, stomatal closure was less sensitive to ABA, and the abo3 mutant was less drought tolerant than the wild type. Northern blot analysis indicated that the expression of the ABA-responsive transcription factor ABF2/AREB1 was markedly lower in the abo3 mutant than in the wild type. The abo3 mutation also reduced the expression of stress-inducible genes RD29A and COR47, especially early during ABA treatment. ABO3 is able to bind the W-box in the promoter of ABF2 in vitro. These results uncover an important role for a WRKY transcription factor in plant responses to ABA and drought stress. PMID:20487379

  19. ABO3, a WRKY transcription factor, mediates plant responses to abscisic acid and drought tolerance in Arabidopsis.

    PubMed

    Ren, Xiaozhi; Chen, Zhizhong; Liu, Yue; Zhang, Hairong; Zhang, Min; Liu, Qian; Hong, Xuhui; Zhu, Jian-Kang; Gong, Zhizhong

    2010-08-01

    The biological functions of WRKY transcription factors in plants have been widely studied, but their roles in abiotic stress are still not well understood. We isolated an ABA overly sensitive mutant, abo3, which is disrupted by a T-DNA insertion in At1g66600 encoding a WRKY transcription factor AtWRKY63. The mutant was hypersensitive to ABA in both seedling establishment and seedling growth. However, stomatal closure was less sensitive to ABA, and the abo3 mutant was less drought tolerant than the wild type. Northern blot analysis indicated that the expression of the ABA-responsive transcription factor ABF2/AREB1 was markedly lower in the abo3 mutant than in the wild type. The abo3 mutation also reduced the expression of stress-inducible genes RD29A and COR47, especially early during ABA treatment. ABO3 is able to bind the W-box in the promoter of ABF2in vitro. These results uncover an important role for a WRKY transcription factor in plant responses to ABA and drought stress.

  20. Gene polymorphisms of novel immunotolerant molecule BTLA: distribution of alleles, genotypes and haplotypes in Polish Caucasian population.

    PubMed

    Partyka, Anna; Woszczyk, Dariusz; Strzała, Tomasz; Szczepańska, Anna; Tomkiewicz, Anna; Frydecka, Irena; Karabon, Lidia

    2015-02-01

    B and T lymphocyte attenuator (BTLA) is one of the members of immunoglobulin superfamily which, like CTLA-4 and PD-1, is involved in down regulation of immune response. Despite the important role of BTLA in maintaining immune homeostasis, relatively little studies were devoted to the relationship of polymorphisms in the gene encoding BTLA with susceptibility to autoimmune disease and cancer. Moreover, all published works were done in Asian populations. BTLA gene is located on chromosome 3 in q13.2 and consists of five exons. The aim of this study was to investigate the alleles, genotypes and haplotypes frequency of selected BTLA gene polymorphisms in Caucasian population originating from Poland. For this study, the single-nucleotide polymorphisms (SNPs) were chosen on the basis of literature data. Additionally, the tag dSNP under linkage equilibrium r (2) > 0.8 and available at the National Center for Biotechnology Information (NCBI) for Caucasian population of rare alleles at a frequency greater than 5 % have been chosen using the NCBI database. The ten BTLA SNPs investigated were: rs1844089, rs2705535, rs9288952, rs9288953, rs1982809, rs2633580, rs2705511, rs2705565, rs76844316, rs16859633. For all SNPs selected on the basis of literature data the significantly different distributions of genotypes between Asian and Caucasian populations were observed.

  1. Interactions between WHITE Genes Carried by a Large Transposing Element and the ZESTE1 Allele in DROSOPHILA MELANOGASTER

    PubMed Central

    Gubb, D.; Roote, J.; McGill, S.; Shelton, M.; Ashburner, M.

    1986-01-01

    TE146, a large transposing element of Drosophila melanogaster, carries two copies of the white and roughest genes in tandem. In consequence, z1 w 11E4; TE146(Z)/+ flies have a zeste (lemon-yellow) eye color. However, one in 103 TE146 chromosomes mutates to a red-eyed form. The majority of these "spontaneous red" (SR) derivatives of TE146 have only one copy of the white gene and are, cytologically, two- to three-banded elements, rather than six-banded as their progenitor. The SR forms of TE146 are also unstable and give zeste-colored forms with a frequency of about one in 104. One such "spontaneous zeste" (SZ) derivative carries duplicated white genes as an inverted, rather than a tandem, repeat. The genetic instability of this inverted repeat form of TE146 is different from that of the original tandem repeat form. In particular, the inverted repeat form frequently produces derivatives with internal rearrangements of the TE and gives a much lower frequency of SR forms. In addition, two novel features of the interaction between w+ alleles in a zeste background have been found. First, copies of w + can become insensitive to suppression by zeste even when paired. Second, an inversion breakpoint may disrupt the pairing between two adjacent w+ alleles, necessary for their suppression by zeste, without physically separating them. PMID:17246318

  2. The frequency of different CGG-repeat alleles in the FMR-1 gene in the general population and special populations

    SciTech Connect

    Holden, J.J.A. |; Chalifoux, M.; Wing, M.

    1994-09-01

    The fragile X (FRAXA) syndrome is the most common inherited form of developmental disability and was the first genetic disorder in which the mechanism of mutation is triplet repeat expansion. The normal fragile X mental retardation-1 gene has 6-52 copies of the CGG-repeat; affected males have extensive amplification, coupled with methylation and gene inactivation; and carriers have between about 55 and 200 copies. There is some overlap in the 45-55 repeat range, with some alleles showing stable and othres unstable transmission. There have been several estimates of the incidence of the FRAXA syndrome, based on testing of special populations using chromosome analysis and the range is 1/750-1/2000. Because of the high burden associated with this syndrome, and in the face of discussions about population screening, it is important to know the actual incidence of mutations in this gene, as well as the distribution of unstable repeats above 45 copes. We have initiated a general population screening to examine 50,000 newborn samples using PCR, and have developed a rapid, inexpensive and reliable method for amplifying the CGG-repeat from Guthrie spots. In the first 1600 samples examined, we found 15 alleles with greater than 45 CGG-repeats, with the highest being 61 repeats.

  3. A Grapevine TTG2-Like WRKY Transcription Factor Is Involved in Regulating Vacuolar Transport and Flavonoid Biosynthesis

    PubMed Central

    Amato, Alessandra; Cavallini, Erika; Zenoni, Sara; Finezzo, Laura; Begheldo, Maura; Ruperti, Benedetto; Tornielli, Giovanni Battista

    2017-01-01

    A small set of TTG2-like homolog proteins from different species belonging to the WRKY family of transcription factors were shown to share a similar mechanism of action and to control partially conserved biochemical/developmental processes in their native species. In particular, by activating P-ATPases residing on the tonoplast, PH3 from Petunia hybrida promotes vacuolar acidification in petal epidermal cells whereas TTG2 from Arabidopsis thaliana enables the accumulation of proanthocyanidins in the seed coat. In this work we functionally characterized VvWRKY26 identified as the closest grapevine homolog of PhPH3 and AtTTG2. When constitutively expressed in petunia ph3 mutant, VvWRKY26 can fulfill the PH3 function in the regulation of vacuolar pH and restores the wild type pigmentation phenotype. By a global correlation analysis of gene expression and by transient over-expression in Vitis vinifera, we showed transcriptomic relationships of VvWRKY26 with many genes related to vacuolar acidification and transport in grapevine. Moreover, our results indicate an involvement in flavonoid pathway possibly restricted to the control of proanthocyanidin biosynthesis that is consistent with its expression pattern in grape berry tissues. Overall, the results show that, in addition to regulative mechanisms and biological roles shared with TTG2-like orthologs, VvWRKY26 can play roles in fleshy fruit development that have not been previously reported in studies from dry fruit species. This study paves the way toward the comprehension of the regulatory network controlling vacuolar acidification and flavonoid accumulation mechanisms that contribute to the final berry quality traits in grapevine. PMID:28105033

  4. L-Methylfolate supplementation in a child with autism and methyltetrahydrofolate reductase, enzyme gene C677TT allele.

    PubMed

    Siscoe, Kim S; Lohr, W David

    2017-03-07

    Errors in folate metabolism may play a role in the pathology of autism spectrum disorders because of increased vulnerability to oxidative stress. We report a case where L-methylfolate supplementation improved symptoms of aggression and disruptive behavior in a child with autism who tested positive for the C677TT allele of the methyltetrahydrofolate reductase enzyme gene. To our knowledge, this is the first report of L-methylfolate administration in this situation. Further controlled studies of L-methylfolate in this population are warranted.

  5. A novel type 2 diabetes risk allele increases the promoter activity of the muscle-specific small ankyrin 1 gene

    PubMed Central

    Yan, Rengna; Lai, Shanshan; Yang, Yang; Shi, Hongfei; Cai, Zhenming; Sorrentino, Vincenzo; Du, Hong; Chen, Huimei

    2016-01-01

    Genome-wide association studies have identified Ankyrin-1 (ANK1) as a common type 2 diabetes (T2D) susceptibility locus. However, the underlying causal variants and functional mechanisms remain unknown. We screened for 8 tag single nucleotide polymorphisms (SNPs) in ANK1 between 2 case-control studies. Genotype analysis revealed significant associations of 3 SNPs, rs508419 (first identified here), rs515071, and rs516946 with T2D (P < 0.001). These SNPs were in linkage disequilibrium (r2 > 0.80); subsequent analysis indicated that the CCC haplotype associated with increased T2D susceptibility (OR 1.447, P < 0.001). Further mapping showed that rs508419 resides in the muscle-specific ANK1 gene promoter. Allele-specific mRNA and protein level measurements confirmed association of the C allele with increased small ANK1 (sAnk1) expression in human skeletal muscle (P = 0.018 and P < 0.001, respectively). Luciferase assays showed increased rs508419-C allele transcriptional activity in murine skeletal muscle C2C12 myoblasts, and electrophoretic mobility-shift assays demonstrated altered rs508419 DNA-protein complex formation. Glucose uptake was decreased with excess sAnk1 expression upon insulin stimulation. Thus, the ANK1 rs508419-C T2D-risk allele alters DNA-protein complex binding leading to increased promoter activity and sAnk1 expression; thus, increased sAnk1 expression in skeletal muscle might contribute to T2D susceptibility. PMID:27121283

  6. Detection of new HLA-DPB1 alleles generated by interallelic gene conversion using PCR amplification of DPB1 second exon sequences from sperm

    SciTech Connect

    Erlich, H.; Zangenberg, G.; Bugawan, T.

    1994-09-01

    The rate at which allelic diversity at the HLA class I and class II loci evolves has been the subject of considerable controversy as have the mechanisms which generate new alleles. The patchwork pattern of polymorphism, particularly within the second exon of the HLA-DPB1 locus where the polymorphic sequence motifs are localized to 6 discrete regions, is consistent with the hypothesis that much of the allelic sequence variation may have been generated by segmental exchange (gene conversion). To measure the rate of new DPB1 variant generation, we have developed a strategy in which DPB1 second exon sequences are amplified from pools of FACS-sorted sperm (n=50) from a heterozygous sperm donor. Pools of sperm from these heterozygous individuals are amplified with an allele-specific primer for one allele and analyzed with sequence-specific oligonucleotide probes (SSOP) complementary to the other allele. This screening procedure, which is capable of detecting a single variant molecule in a pool of parental alleles, allows the identification of new variants that have been generated by recombination and/or gene conversion between the two parental alleles. To control for potential PCR artifacts, the same screening procedure was carried out with mixtures of sperm from DPB1 *0301/*0301 and DPB1 *0401/ 0401 individuals. Pools containing putative new variants DPB1 alleles were analyzed further by cloning into M13 and sequencing the M13 clones. Our current estimate is that about 1/10,000 sperm from these heterozygous individuals represents a new DPB1 allele generated by micro-gene conversion within the second exon.

  7. Limited gene misregulation is exacerbated by allele-specific upregulation in lethal hybrids between Drosophila melanogaster and Drosophila simulans.

    PubMed

    Wei, Kevin H-C; Clark, Andrew G; Barbash, Daniel A

    2014-07-01

    Misregulation of gene expression is often observed in interspecific hybrids and is generally attributed to regulatory incompatibilities caused by divergence between the two genomes. However, it has been challenging to distinguish effects of regulatory divergence from secondary effects including developmental and physiological defects common to hybrids. Here, we use RNA-Seq to profile gene expression in F1 hybrid male larvae from crosses of Drosophila melanogaster to its sibling species D. simulans. We analyze lethal and viable hybrid males, the latter produced using a mutation in the X-linked D. melanogaster Hybrid male rescue (Hmr) gene and compare them with their parental species and to public data sets of gene expression across development. We find that Hmr has drastically different effects on the parental and hybrid genomes, demonstrating that hybrid incompatibility genes can exhibit novel properties in the hybrid genetic background. Additionally, we find that D. melanogaster alleles are preferentially affected between lethal and viable hybrids. We further determine that many of the differences between the hybrids result from developmental delay in the Hmr(+) hybrids. Finally, we find surprisingly modest expression differences in hybrids when compared with the parents, with only 9% and 4% of genes deviating from additivity or expressed outside of the parental range, respectively. Most of these differences can be attributed to developmental delay and differences in tissue types. Overall, our study suggests that hybrid gene misexpression is prone to overestimation and that even between species separated by approximately 2.5 Ma, regulatory incompatibilities are not widespread in hybrids.

  8. [Prognostic implications of GP3a glucoprotein gene PLA1/PLA2 allele in prostatic cancer: pilot results of the study].

    PubMed

    Loran, O B; Itkes, A V; Seregin, A A; Miandina, G I

    2005-01-01

    We studied the role of integrins, primarily, the role of allele distribution of GP3a gene in development of prostatic cancer (PC) and assessment of its prognostic significance. From November 2003 to May 2004 we examined 32 patients with PC: 11 patients with local PC T1-2N0M0; 14 patients with locally advanced cancer T3N0M0 and 7 patients with invasive and/or metastatic cancer T3-4N10-1 or T3-4N0-1M1. The blood from all the patients we studied with PCR for alleles of GP3a gene, PSA. Seventeen patients were found to have alleles PLA1A1, 14(44%)--alleles PLA1A2, 1(3%)--alleles PLA2A2. Alleles PLA1A2 occurred significantly more often than in the population (p < 0.005). The group analysis has found that 8 patients with local PC had alleles PLA1A1, 3 patients--alleles PLA1A2 (27%). We discovered alleles PLA2A2, PLA1A1 and PLA1A2 in 1(7%), 5(36%) and 8(57%) patients with locally advanced PC, respectively. Among patients with metastatic and/or invasive prostatic cancer, there were 4 (57%) and 3 (43%) cases of alleles PLA1A1 and PLA1A2, respectively. Our study demonstrated influence of carriage of PLA2 allele on occurrence of PC risk (5-fold higher) and its invasive forms (10-fold higher and more). Probability to develop local invasion among patients with prostatic cancer--carriers allele PLA1A2 is 6 times higher than among carriers of alleles PLA1A1. A PC course in carriers of alleles PLA1A2 may be characterized by faster development of local invasion and metastasizing vs carriers of alleles PLA1A1. These findings can be used in design of nomograms for prognostication of invasion of clinically small tumors in verification of significance on greater number of the patients.

  9. The murine Cd48 gene: allelic polymorphism in the IgV-like region.

    PubMed

    Cabrero, J G; Freeman, G J; Reiser, H

    1998-12-01

    The murine CD48 molecule is a member of the immunoglobulin superfamily which regulates the activation of T lymphocytes. prior cloning experiments using mRNA from two different mouse strains had yielded discrepant sequences within the IgV-like domain of murine CD48. To resolve this issue, we have directly sequenced genomic DNA of 10 laboratory strains and two inbred strains of wild origin. The results of our analysis reveal an allelic polymorphism within the IgV-like domain of murine CD48.

  10. Allelic diversity of the MHC class II DRB genes in brown bears (Ursus arctos) and a comparison of DRB sequences within the family Ursidae.

    PubMed

    Goda, N; Mano, T; Kosintsev, P; Vorobiev, A; Masuda, R

    2010-11-01

    The allelic diversity of the DRB locus in major histocompatibility complex (MHC) genes was analyzed in the brown bear (Ursus arctos) from the Hokkaido Island of Japan, Siberia, and Kodiak of Alaska. Nineteen alleles of the DRB exon 2 were identified from a total of 38 individuals of U. arctos and were highly polymorphic. Comparisons of non-synonymous and synonymous substitutions in the antigen-binding sites of deduced amino acid sequences indicated evidence for balancing selection on the bear DRB locus. The phylogenetic analysis of the DRB alleles among three genera (Ursus, Tremarctos, and Ailuropoda) in the family Ursidae revealed that DRB allelic lineages were not separated according to species. This strongly shows trans-species persistence of DRB alleles within the Ursidae.

  11. Genetics of unstable alleles of the X chromosome genes isolated from natural populations of Drosophila melanogaster during the outburst of mutation yellow in 1982 to 1991 in Uman`

    SciTech Connect

    Zakharov, I.K.; Skibitskii, E.E.

    1995-08-01

    In 1982, a local increase of frequency of mutation yellow-2, which lasted for a decade, occurred in a population of Drosophila melanogaster from Uman` (Ukraine). Genetic properties (phenotypic difference, mutability, and pecularities of complementation) of alleles yellow-2, isolated from the population during the mutation outburst, and of their revertants, were studied. Allelic diversity, which reflected molecular differences in allele structure, was shown to appear. In addition to mutation yellow, isolated in 1990 from the Uman` population, mutational properties of other sex-linked genes (dusky, miniature, rudimentary, singed, and vermilion) isolated from natural populations in 1986 to 1990, were analyzed. Based on these data, the conclusion was drawn that the presence of unstable alleles in populations is not a sufficient condition for mutation outbursts. Comparative analysis of properties of yellow alleles obtained in different periods of the outburst continues. 17 refs., 4 tabs.

  12. Allele distributions at hybrid incompatibility loci facilitate the potential for gene flow between cultivated and weedy rice in the US.

    PubMed

    Craig, Stephanie M; Reagon, Michael; Resnick, Lauren E; Caicedo, Ana L

    2014-01-01

    The accumulation of independent mutations over time in two populations often leads to reproductive isolation. Reproductive isolation between diverging populations may be reinforced by barriers that occur either pre- or postzygotically. Hybrid sterility is the most common form of postzygotic isolation in plants. Four postzygotic sterility loci, comprising three hybrid sterility systems (Sa, s5, DPL), have been recently identified in Oryza sativa. These loci explain, in part, the limited hybridization that occurs between the domesticated cultivated rice varieties, O. sativa spp. japonica and O. sativa spp. indica. In the United States, cultivated fields of japonica rice are often invaded by conspecific weeds that have been shown to be of indica origin. Crop-weed hybrids have been identified in crop fields, but at low frequencies. Here we examined the possible role of these hybrid incompatibility loci in the interaction between cultivated and weedy rice. We identified a novel allele at Sa that seemingly prevents loss of fertility in hybrids. Additionally, we found wide-compatibility type alleles at strikingly high frequencies at the Sa and s5 loci in weed groups, and a general lack of incompatible alleles between crops and weeds at the DPL loci. Our results suggest that weedy individuals, particularly those of the SH and BRH groups, should be able to freely hybridize with the local japonica crop, and that prezygotic factors, such as differences in flowering time, have been more important in limiting weed-crop gene flow in the past. As the selective landscape for weedy rice changes due to increased use of herbicide resistant strains of cultivated rice, the genetic barriers that hinder indica-japonica hybridization cannot be counted on to limit the flow of favorable crop genes into weeds.

  13. Imprinted chromosomal domains revealed by allele-specific replication timing of the GABRB3 and GABRA5 genes

    SciTech Connect

    LaSalle, J.; Flint, A.; Lalande, M.

    1994-09-01

    The GABRB3 and GABRA5 genes are organized as a cluster in chromosome 15q11-q13. The genes are separated by around 100 kb and arranged in opposite transcriptional orientations. The GABA{sub A} receptor cluster lies near the Angelman and Prader-Willi loci and displays asynchronous DNA replication, suggesting that this region is subject to parental imprinting. In order to further study the association between DNA replication and imprinting, allele-specific replication was assayed by fluorescence in situ hybridization with {lambda}-phage probes from the GABRB3/A5 region and a D15Z1 satellite probe to identify the parental origin of each chromosome. The replication kinetics of each allele was determined by using a flow sorter to fractionate mitogen-stimulated lymphocytes on the basis of cell cycle progression prior to FISH analysis. These kinetic studies reveal a 50-150 kb chromosomal domain extending from the middle of the GABRB3/A5 intergenic region into the GABRA5 5{prime}-UTR which displays maternal replication in early S with paternal replication delayed until the end of S. In contrast, genomic regions on either side of this maternal early replication domain exhibit the opposite pattern with paternal before maternal replication and both alleles replicating in the latter half of S. These results indicate that the GABRB3/A5 region is divided into domains in which replication timing is determined by parental origin. In addition to a loss of asynchronous replication, organization into replication timing domains is also lost in lymphocytes from maternal and paternal uniparental disomy 15 patients suggesting that a chromosome contribution from both parents is required for the establishment of the imprinted replication domains.

  14. Allele mining in the pepper gene pool provided new complementation effects between pvr2-eIF4E and pvr6-eIF(iso)4E alleles for resistance to pepper veinal mottle virus.

    PubMed

    Rubio, Manuel; Nicolaï, Maryse; Caranta, Carole; Palloix, Alain

    2009-11-01

    Molecular cloning of recessive resistance genes to potyviruses in a large range of host species identified the eukaryotic translation initiation factor 4E (eIF4E) as an essential determinant in the outcome of potyvirus infection. Resistance results from a few amino acid changes in the eIF4E protein encoded by the recessive resistance allele that disrupt the direct interaction with the potyviral protein VPg. In plants, several loci encode two protein subfamilies, eIF4E and eIF(iso)4E. While most eIF4E-mediated resistance to potyviruses depends on mutations in a single eIF4E protein, simultaneous mutations in eIF4E (corresponding to the pvr2 locus) and eIF(iso)4E (corresponding to the pvr6 locus) are required to prevent pepper veinal mottle virus (PVMV) infection in pepper. We used this model to look for additional alleles at the pvr2-eIF4E locus that result in resistance when combined with the pvr6-eIF(iso)4E resistant allele. Among the 12 pvr2-eIF4E resistance alleles sequenced in the pepper gene pool, three were shown to have a complementary effect with pvr6-eIF(iso)4E for resistance. Two amino acid changes were exclusively shared by these three alleles and were systematically associated with a second amino acid change, suggesting that these substitutions are associated with resistance expression. The availability of new resistant allele combinations increases the possibility for the durable deployment of resistance against this pepper virus which is prevalent in Africa.

  15. Bi-allelic alterations in DNA repair genes underpin homologous recombination DNA repair defects in breast cancer.

    PubMed

    Mutter, Robert W; Riaz, Nadeem; Ng, Charlotte K Y; Delsite, Rob; Piscuoglio, Salvatore; Edelweiss, Marcia; Martelotto, Luciano G; Sakr, Rita A; King, Tari A; Giri, Dilip D; Drobnjak, Maria; Brogi, Edi; Bindra, Ranjit; Bernheim, Giana; Lim, Raymond S; Blecua, Pedro; Desrichard, Alexis; Higginson, Dan; Towers, Russell; Jiang, Ruomu; Lee, William; Weigelt, Britta; Reis-Filho, Jorge S; Powell, Simon N

    2017-03-15

    Homologous recombination (HR) DNA repair deficient (HRD) breast cancers have been shown to be sensitive to DNA repair targeted therapies. Burgeoning evidence suggests that sporadic breast cancers, lacking germline BRCA1/BRCA2 mutations, may also be HRD. We developed a functional ex vivo RAD51-based test to identify HRD primary breast cancers. An integrated approach examining methylation, gene expression and whole-exome sequencing was employed to ascertain the etiology of HRD. Functional HRD breast cancers displayed genomic features of lack of competent HR, including large-scale state transitions and specific mutational signatures. Somatic and/or germline genetic alterations resulting in bi-allelic loss-of-function of HR genes underpinned functional HRD in 89% of cases, and were observed in only one of the 15 HR-proficient samples tested. These findings indicate the importance of a comprehensive genetic assessment of bi-allelic alterations in the HR pathway to deliver a precision medicine-based approach to select patients for therapies targeting tumor-specific DNA repair defects.

  16. An unstable targeted allele of the mouse Mitf gene with a high somatic and germline reversion rate.

    PubMed

    Bismuth, Keren; Skuntz, Susan; Hallsson, Jón H; Pak, Evgenia; Dutra, Amalia S; Steingrímsson, Eiríkur; Arnheiter, Heinz

    2008-01-01

    The mouse Mitf gene encodes a transcription factor that is regulated by serine phosphorylation and is critical for the development of melanin-containing pigment cells. To test the role of phosphorylation at a particular serine, S73 in exon 2 of Mitf, we used a standard targeting strategy in mouse embryonic stem cells to change the corresponding codon into one encoding an alanine. By chance, we generated an allele in which 85,222 bp of wild-type Mitf sequence are duplicated and inserted into an otherwise correctly targeted Mitf gene. Depending on the presence or absence of a neomycin resistance cassette, this genomic rearrangement leads to animals with a white coat with or without pigmented spots or a gray coat with obligatory white and black spots. Several independent, genetically stable germline revertants that lacked the duplicated wild-type sequence but retained the targeted codon were then derived. These animals were normally pigmented, indicating that the serine-to-alanine mutation is not deleterious to melanocyte development. The fact that mosaic coat reversions occur in all mice lacking the neo-cassette and that approximately 1% of these transmit a reverted allele to their offspring places this mutation among those with the highest spontaneous reversion rates in mammals.

  17. A recombined allele of the lipase gene CEL and its pseudogene CELP confers susceptibility to chronic pancreatitis

    PubMed Central

    Fjeld, Karianne; Johansson, Bente B.; Kirsten, Holger; Ruffert, Claudia; Masson, Emmanuelle; Steine, Solrun J.; Bugert, Peter; Cnop, Miriam; Grützmann, Robert; Mayerle, Julia; Mössner, Joachim; Ringdal, Monika; Schulz, Hans-Ulrich; Sendler, Matthias; Simon, Peter; Sztromwasser, Paweł; Torsvik, Janniche; Scholz, Markus; Tjora, Erling; Férec, Claude; Witt, Heiko; Lerch, Markus M.; Njølstad, Pål R.; Johansson, Stefan; Molven, Anders

    2015-01-01

    Carboxyl-ester lipase is a digestive pancreatic enzyme encoded by the highly polymorphic CEL gene1. Mutations in CEL cause maturity-onset diabetes of the young (MODY) with pancreatic exocrine dysfunction2. Here we identified a hybrid allele (CEL-HYB), originating from a crossover between CEL and its neighboring pseudogene CELP. In a discovery cohort of familial chronic pancreatitis cases, the carrier frequency of CEL-HYB was 14.1% (10/71) compared with 1.0% (5/478) in controls (odds ratio [OR] = 15.5, 95% confidence interval [CI] = 5.1-46.9, P = 1.3 × 10−6). Three replication studies in non-alcoholic chronic pancreatitis cohorts identified CEL-HYB in a total of 3.7% (42/1,122) cases and 0.7% (30/4,152) controls (OR = 5.2, 95% CI = 3.2-8.5, P = 1.2 × 10−11; formal meta-analysis). The allele was also enriched in alcoholic chronic pancreatitis. Expression of CEL-HYB in cellular models revealed reduced lipolytic activity, impaired secretion, prominent intracellular accumulation and induced autophagy. The hybrid variant of CEL is the first chronic pancreatitis gene identified outside the protease/antiprotease system of pancreatic acinar cells. PMID:25774637

  18. Identification of a novel pseudodeficiency allele in the GLB1 gene in a carrier of GM1 gangliosidosis.

    PubMed

    Gort, L; Santamaria, R; Grinberg, D; Vilageliu, L; Chabás, A

    2007-08-01

    The term 'pseudodeficiency' is used in lysosomal storage diseases to denote the situation in which individuals show greatly reduced enzyme activity but remain clinically healthy. Pseudodeficiencies have been reported for several lysosomal hydrolases. GM1 gangliosidosis is a rare autosomal recessive lysosomal storage disorder caused by beta-galactosidase hydrolase deficiency as a result of mutations in the GLB1 gene. Until now, two variants altering the beta-galactosidase activity have been described, p.Arg521Cys and p.Ser532Gly. Here we report the new variant p.Arg595Trp in the GLB1 gene, which markedly reduces beta-galactosidase activity when expressed in COS-1 cells. The variant was identified in the healthy father of a girl with GM1 gangliosidosis. He was a heterozygous compound with p.Arg595Trp in trans with one of the disease-causing mutations identified in his daughter; in leukocytes and plasma he showed lower beta-galactosidase activity than that observed in GM1 gangliosidosis carriers. As this family originated from the Basque Country in the north of Spain, we decided to analyse individuals of Basque and non-Basque origin, finding the p.Arg595Trp allele in 3.2% of Basque and in 0.8% of non-Basque alleles. The detection of the presence of alterations resulting in pseudodeficient activity in leukocytes and plasma is important for the correct diagnosis of GM1 gangliosidosis.

  19. Positive Selection of Deleterious Alleles through Interaction with a Sex-Ratio Suppressor Gene in African Buffalo: A Plausible New Mechanism for a High Frequency Anomaly

    PubMed Central

    van Hooft, Pim; Greyling, Ben J.; Getz, Wayne M.; van Helden, Paul D.; Zwaan, Bas J.; Bastos, Armanda D. S.

    2014-01-01

    Although generally rare, deleterious alleles can become common through genetic drift, hitchhiking or reductions in selective constraints. Here we present a possible new mechanism that explains the attainment of high frequencies of deleterious alleles in the African buffalo (Syncerus caffer) population of Kruger National Park, through positive selection of these alleles that is ultimately driven by a sex-ratio suppressor. We have previously shown that one in four Kruger buffalo has a Y-chromosome profile that, despite being associated with low body condition, appears to impart a relative reproductive advantage, and which is stably maintained through a sex-ratio suppressor. Apparently, this sex-ratio suppressor prevents fertility reduction that generally accompanies sex-ratio distortion. We hypothesize that this body-condition-associated reproductive advantage increases the fitness of alleles that negatively affect male body condition, causing genome-wide positive selection of these alleles. To investigate this we genotyped 459 buffalo using 17 autosomal microsatellites. By correlating heterozygosity with body condition (heterozygosity-fitness correlations), we found that most microsatellites were associated with one of two gene types: one with elevated frequencies of deleterious alleles that have a negative effect on body condition, irrespective of sex; the other with elevated frequencies of sexually antagonistic alleles that are negative for male body condition but positive for female body condition. Positive selection and a direct association with a Y-chromosomal sex-ratio suppressor are indicated, respectively, by allele clines and by relatively high numbers of homozygous deleterious alleles among sex-ratio suppressor carriers. This study, which employs novel statistical techniques to analyse heterozygosity-fitness correlations, is the first to demonstrate the abundance of sexually-antagonistic genes in a natural mammal population. It also has important

  20. Positive selection of deleterious alleles through interaction with a sex-ratio suppressor gene in African Buffalo: a plausible new mechanism for a high frequency anomaly.

    PubMed

    van Hooft, Pim; Greyling, Ben J; Getz, Wayne M; van Helden, Paul D; Zwaan, Bas J; Bastos, Armanda D S

    2014-01-01

    Although generally rare, deleterious alleles can become common through genetic drift, hitchhiking or reductions in selective constraints. Here we present a possible new mechanism that explains the attainment of high frequencies of deleterious alleles in the African buffalo (Syncerus caffer) population of Kruger National Park, through positive selection of these alleles that is ultimately driven by a sex-ratio suppressor. We have previously shown that one in four Kruger buffalo has a Y-chromosome profile that, despite being associated with low body condition, appears to impart a relative reproductive advantage, and which is stably maintained through a sex-ratio suppressor. Apparently, this sex-ratio suppressor prevents fertility reduction that generally accompanies sex-ratio distortion. We hypothesize that this body-condition-associated reproductive advantage increases the fitness of alleles that negatively affect male body condition, causing genome-wide positive selection of these alleles. To investigate this we genotyped 459 buffalo using 17 autosomal microsatellites. By correlating heterozygosity with body condition (heterozygosity-fitness correlations), we found that most microsatellites were associated with one of two gene types: one with elevated frequencies of deleterious alleles that have a negative effect on body condition, irrespective of sex; the other with elevated frequencies of sexually antagonistic alleles that are negative for male body condition but positive for female body condition. Positive selection and a direct association with a Y-chromosomal sex-ratio suppressor are indicated, respectively, by allele clines and by relatively high numbers of homozygous deleterious alleles among sex-ratio suppressor carriers. This study, which employs novel statistical techniques to analyse heterozygosity-fitness correlations, is the first to demonstrate the abundance of sexually-antagonistic genes in a natural mammal population. It also has important

  1. Novel Hypomorphic Alleles of the Mouse Tyrosinase Gene Induced by CRISPR-Cas9 Nucleases Cause Non-Albino Pigmentation Phenotypes

    PubMed Central

    Boitet, Evan R.; Turner, Ashley N.; Johnson, Larry W.; Kennedy, Daniel; Downs, Ethan R.; Hymel, Katherine M.; Gross, Alecia K.; Kesterson, Robert A.

    2016-01-01

    Tyrosinase is a key enzyme in melanin biosynthesis. Mutations in the gene encoding tyrosinase (Tyr) cause oculocutaneous albinism (OCA1) in humans. Alleles of the Tyr gene have been useful in studying pigment biology and coat color formation. Over 100 different Tyr alleles have been reported in mice, of which ≈24% are spontaneous mutations, ≈60% are radiation-induced, and the remaining alleles were obtained by chemical mutagenesis and gene targeting. Therefore, most mutations were random and could not be predicted a priori. Using the CRISPR-Cas9 system, we targeted two distinct regions of exon 1 to induce pigmentation changes and used an in vivo visual phenotype along with heteroduplex mobility assays (HMA) as readouts of CRISPR-Cas9 activity. Most of the mutant alleles result in complete loss of tyrosinase activity leading to an albino phenotype. In this study, we describe two novel in-frame deletion alleles of Tyr, dhoosara (Sanskrit for gray) and chandana (Sanskrit for sandalwood). These alleles are hypomorphic and show lighter pigmentation phenotypes of the body and eyes. This study demonstrates the utility of CRISPR-Cas9 system in generating domain-specific in-frame deletions and helps gain further insights into structure-function of Tyr gene. PMID:27224051

  2. Analysis of nucleotide diversity among alleles of the major bacterial blight resistance gene Xa27 in cultivars of rice (Oryza sativa) and its wild relatives.

    PubMed

    Bimolata, Waikhom; Kumar, Anirudh; Sundaram, Raman Meenakshi; Laha, Gouri Shankar; Qureshi, Insaf Ahmed; Reddy, Gajjala Ashok; Ghazi, Irfan Ahmad

    2013-08-01

    Xa27 is one of the important R-genes, effective against bacterial blight disease of rice caused by Xanthomonas oryzae pv. oryzae (Xoo). Using natural population of Oryza, we analyzed the sequence variation in the functionally important domains of Xa27 across the Oryza species. DNA sequences of Xa27 alleles from 27 rice accessions revealed higher nucleotide diversity among the reported R-genes of rice. Sequence polymorphism analysis revealed synonymous and non-synonymous mutations in addition to a number of InDels in non-coding regions of the gene. High sequence variation was observed in the promoter region including the 5'UTR with 'π' value 0.00916 and 'θ w ' = 0.01785. Comparative analysis of the identified Xa27 alleles with that of IRBB27 and IR24 indicated the operation of both positive selection (Ka/Ks > 1) and neutral selection (Ka/Ks ≈ 0). The genetic distances of alleles of the gene from Oryza nivara were nearer to IRBB27 as compared to IR24. We also found the presence of conserved and null UPT (upregulated by transcriptional activator) box in the isolated alleles. Considerable amino acid polymorphism was localized in the trans-membrane domain for which the functional significance is yet to be elucidated. However, the absence of functional UPT box in all the alleles except IRBB27 suggests the maintenance of single resistant allele throughout the natural population.

  3. Production of early flowering transgenic barley expressing the early flowering allele of Cryptochrome2 gene.

    PubMed

    El-Assal, Salah El-Din; Abd-Alla, Samir M; El-Tarras, Adel A; El-Awady, Mohamed A

    2011-01-01

    This work was carried out in order to develop early flowering barley lines. These lines will be useful to producers by enabling multiple crops within a single season and increasing production. Transgenic barley plants containing the natural early flowering time AtCRY2 allele from the Cape Verde Island (Cvi) ecotype of Arabidopsis have been generated using biolistic transformation. Immature embryo derived calli of two commercially important barley cultivars (El-Dwaser and El-Taif), were transformed using a pCAMBIA-2300 plasmid harboring a genomic fragment containing the AtCRY2-Cvi allele. Transformation was performed utilizing 600 immature embryos for each cultivar. Stable transformation was confirmed in T 0 and T 1 plants by using genomic PCR, RT-PCR and western blot analysis with AtCRY2 specific primers and antibodies, respectively. The transformation efficiency was 5.6% and 3.4% for El-Dwaser and El-Taif cultivars, respectively. Seeds from several T 1 lines were germinated on kanamycin plates and the lines that contained a single locus were selected for further evaluation. The transformed barley plants showed the specific AtCRY2-Cvi flowering phenotype, i.e. early flowering and day length insensitivity, compared to the non transgenic plants. The time to flowering in transgenic T 1 plants was assessed and two lines exhibited flowering more than 25 days earlier than the parental cultivars under short day conditions.

  4. WRKY42 Modulates Phosphate Homeostasis through Regulating Phosphate Translocation and Acquisition in Arabidopsis1[OPEN

    PubMed Central

    Su, Tong; Xu, Qian; Zhang, Fei-Cui; Chen, Yun; Li, Li-Qin; Wu, Wei-Hua; Chen, Yi-Fang

    2015-01-01

    The Arabidopsis (Arabidopsis thaliana) WRKY transcription factor family has more than 70 members, and some of them have been reported to play important roles in plant response to biotic and abiotic stresses. This study shows that WRKY42 regulated phosphate homeostasis in Arabidopsis. The WRKY42-overexpressing lines, similar to the phosphate1 (pho1) mutant, were more sensitive to low-inorganic phosphate (Pi) stress and had lower shoot Pi content compared with wild-type plants. The PHO1 expression was repressed in WRKY42-overexpressing lines and enhanced in the wrky42 wrky6 double mutant. The WRKY42 protein bound to the PHO1 promoter under Pi-sufficient condition, and this binding was abrogated during Pi starvation. These data indicate that WRKY42 modulated Pi translocation by regulating PHO1 expression. Furthermore, overexpression of WRKY42 increased root Pi content and Pi uptake, whereas the wrky42 mutant had lower root Pi content and Pi uptake rate compared with wild-type plants. Under Pi-sufficient condition, WRKY42 positively regulated PHOSPHATE TRANSPORTER1;1 (PHT1;1) expression by binding to the PHT1;1 promoter, and this binding was abolished by low-Pi stress. During Pi starvation, the WRKY42 protein was degraded through the 26S proteasome pathway. Our results showed that AtWRKY42 modulated Pi homeostasis by regulating the expression of PHO1 and PHT1;1 to adapt to environmental changes in Pi availability. PMID:25733771

  5. Different Alleles of a Gene Encoding Leucoanthocyanidin Reductase (PaLAR3) Influence Resistance against the Fungus Heterobasidion parviporum in Picea abies.

    PubMed

    Nemesio-Gorriz, Miguel; Hammerbacher, Almuth; Ihrmark, Katarina; Källman, Thomas; Olson, Åke; Lascoux, Martin; Stenlid, Jan; Gershenzon, Jonathan; Elfstrand, Malin

    2016-08-01

    Despite the fact that fungal diseases are a growing menace for conifers in modern silviculture, only a very limited number of molecular markers for pathogen resistance have been validated in conifer species. A previous genetic study indicated that the resistance of Norway spruce (Picea abies) to Heterobasidion annosum s.l., a pathogenic basidiomycete species complex, is linked to a quantitative trait loci that associates with differences in fungal growth in sapwood (FGS) that includes a gene, PaLAR3, which encodes a leucoanthocyanidin reductase. In this study, gene sequences showed the presence of two PaLAR3 allelic lineages in P. abies. Higher resistance was associated with the novel allele, which was found in low frequency in the four P. abies populations that we studied. Norway spruce plants carrying at least one copy of the novel allele showed a significant reduction in FGS after inoculation with Heterobasidion parviporum compared to their half-siblings carrying no copies, indicating dominance of this allele. The amount of (+) catechin, the enzymatic product of PaLAR3, was significantly higher in bark of trees homozygous for the novel allele. Although we observed that the in vitro activities of the enzymes encoded by the two alleles were similar, we could show that allele-specific transcript levels were significantly higher for the novel allele, indicating that regulation of gene expression is responsible for the observed effects in resistance, possibly caused by differences in cis-acting elements that we observe in the promoter region of the two alleles.

  6. Inactive allele-specific methylation and chromatin structure of the imprinted gene U2af1-rs1 on mouse chromosome 11

    SciTech Connect

    Shibata, Hideo; Yoshino, Kiyoshi; Kamiya, Mamoru

    1996-07-01

    The imprinted U2Af1-rs1 gene that maps to mouse chromosome 11 is predominately expressed from the paternal allele. We examined the methylation of genomic sequences in and around the U2af1-rs1 locus to establish the extent of sequence modifications that accompanied the silencing of the maternal allele. The analysis of HapII or HhaI sites showed that the silent maternal allele was hypermethylated in a block of CpG sequences that covered more than 10 kb. By comparison, the expressed paternal allele was unmethylated from a CpG island upstream of the transcribed region through 2 kb. An analysis of DNaseI hypersensitivity of a putative promoter of U2af1-rs1 showed an open chromatin conformation only on the unmethylated, expressed paternal allele. These results suggest that allele-specific hypermethylation covering the gene and its upstream CpG island plays a role in maternal allele repression of U2af1-rs1, which is reflected in altered chromatin conformation of DNaseI hypersensitive sites. 9 refs., 2 figs.

  7. In-Frame and Unmarked Gene Deletions in Burkholderia cenocepacia via an Allelic Exchange System Compatible with Gateway Technology.

    PubMed

    Fazli, Mustafa; Harrison, Joe J; Gambino, Michela; Givskov, Michael; Tolker-Nielsen, Tim

    2015-06-01

    Burkholderia cenocepacia is an emerging opportunistic pathogen causing life-threatening infections in immunocompromised individuals and in patients with cystic fibrosis, which are often difficult, if not impossible, to treat. Understanding the genetic basis of virulence in this emerging pathogen is important for the development of novel treatment regimes. Generation of deletion mutations in genes predicted to encode virulence determinants is fundamental to investigating the mechanisms of pathogenesis. However, there is a lack of appropriate selectable and counterselectable markers for use in B. cenocepacia, making its genetic manipulation problematic. Here we describe a Gateway-compatible allelic exchange system based on the counterselectable pheS gene and the I-SceI homing endonuclease. This system provides efficiency in cloning homology regions of target genes and allows the generation of precise and unmarked gene deletions in B. cenocepacia. As a proof of concept, we demonstrate its utility by deleting the Bcam1349 gene, encoding a cyclic di-GMP (c-di-GMP)-responsive regulator protein important for biofilm formation.

  8. In-Frame and Unmarked Gene Deletions in Burkholderia cenocepacia via an Allelic Exchange System Compatible with Gateway Technology

    PubMed Central

    Fazli, Mustafa; Harrison, Joe J.; Gambino, Michela; Givskov, Michael

    2015-01-01

    Burkholderia cenocepacia is an emerging opportunistic pathogen causing life-threatening infections in immunocompromised individuals and in patients with cystic fibrosis, which are often difficult, if not impossible, to treat. Understanding the genetic basis of virulence in this emerging pathogen is important for the development of novel treatment regimes. Generation of deletion mutations in genes predicted to encode virulence determinants is fundamental to investigating the mechanisms of pathogenesis. However, there is a lack of appropriate selectable and counterselectable markers for use in B. cenocepacia, making its genetic manipulation problematic. Here we describe a Gateway-compatible allelic exchange system based on the counterselectable pheS gene and the I-SceI homing endonuclease. This system provides efficiency in cloning homology regions of target genes and allows the generation of precise and unmarked gene deletions in B. cenocepacia. As a proof of concept, we demonstrate its utility by deleting the Bcam1349 gene, encoding a cyclic di-GMP (c-di-GMP)-responsive regulator protein important for biofilm formation. PMID:25795676

  9. Severe hypertriglyceridemia in a patient heterozygous for a lipoprotein lipase gene allele with two novel missense variants.

    PubMed

    Kassner, Ursula; Salewsky, Bastian; Wühle-Demuth, Marion; Szijarto, Istvan Andras; Grenkowitz, Thomas; Binner, Priska; März, Winfried; Steinhagen-Thiessen, Elisabeth; Demuth, Ilja

    2015-09-01

    Rare monogenic hyperchylomicronemia is caused by loss-of-function mutations in genes involved in the catabolism of triglyceride-rich lipoproteins, including the lipoprotein lipase gene, LPL. Clinical hallmarks of this condition are eruptive xanthomas, recurrent pancreatitis and abdominal pain. Patients with LPL deficiency and severe or recurrent pancreatitis are eligible for the first gene therapy treatment approved by the European Union. Therefore the precise molecular diagnosis of familial hyperchylomicronemia may affect treatment decisions. We present a 57-year-old male patient with excessive hypertriglyceridemia despite intensive lipid-lowering therapy. Abdominal sonography showed signs of chronic pancreatitis. Direct DNA sequencing and cloning revealed two novel missense variants, c.1302A>T and c.1306G>A, in exon 8 of the LPL gene coexisting on the same allele. The variants result in the amino-acid exchanges p.(Lys434Asn) and p.(Gly436Arg). They are located in the carboxy-terminal domain of lipoprotein lipase that interacts with the glycosylphosphatidylinositol-anchored HDL-binding protein (GPIHBP1) and are likely of functional relevance. No further relevant mutations were found by direct sequencing of the genes for APOA5, APOC2, LMF1 and GPIHBP1. We conclude that heterozygosity for damaging mutations of LPL may be sufficient to produce severe hypertriglyceridemia and that chylomicronemia may be transmitted in a dominant manner, at least in some families.

  10. Allele and genotype frequencies of polymorphisms in cytokine genes in ethnic Russian individuals from Moscow, Russia.

    PubMed

    Shadrina, Alexandra; Voronina, Elena; Zolotukhin, Igor; Filipenko, Maxim

    2017-02-01

    Two hundred and twenty eight ethnic Russian individuals from Moscow, Russia, were genotyped at 14 single nucleotide polymorphisms CCL2 A-2578G; VEGFA C-2578A, G-634C, and C+936T; TNF G+419A and G-308A; IL1A G-889A; IL1RN T+1018C; IL6G-174C and G-572C; IFNG T+874A; IL1B C-511T; IL10 A+1082G; TGFB1 C-509T. Genotypes were determined using real-time polymerase chain reaction with TaqMan probes and polymerase chain reaction followed by melting analysis of dual-labeled probe. Genotype distribution was in accordance with Hardy-Weinberg equilibrium for all studied polymorphisms. Genotype data are available in the Allele Frequencies Net Database under identifier AFND 3367 and the population name "Russia Moscow Cytokine".

  11. Pepper CabZIP63 acts as a positive regulator during Ralstonia solanacearum or high temperature-high humidity challenge in a positive feedback loop with CaWRKY40.

    PubMed

    Shen, Lei; Liu, Zhiqin; Yang, Sheng; Yang, Tong; Liang, Jiaqi; Wen, Jiayu; Liu, Yanyan; Li, Jiazhi; Shi, Lanping; Tang, Qian; Shi, Wei; Hu, Jiong; Liu, Cailing; Zhang, Yangwen; Lin, Wei; Wang, Rongzhang; Yu, Huanxin; Mou, Shaoliang; Hussain, Ansar; Cheng, Wei; Cai, Hanyang; He, Li; Guan, Deyi; Wu, Yang; He, Shuilin

    2016-04-01

    CaWRKY40 is known to act as a positive regulator in the response of pepper (Capsicum annuum) to Ralstonia solanacearum inoculation (RSI) or high temperature-high humidity (HTHH), but the underlying mechanism remains elusive. Herein, we report that CabZIP63, a pepper bZIP family member, participates in this process by regulating the expression of CaWRKY40. CabZIP63 was found to localize in the nuclei, be up-regulated by RSI or HTHH, bind to promoters of both CabZIP63(pCabZIP63) and CaWRKY40(pCaWRKY40), and activate pCabZIP63- and pCaWRKY40-driven β-glucuronidase expression in a C- or G-box-dependent manner. Silencing of CabZIP63 by virus-induced gene silencing (VIGS) in pepper plants significantly attenuated their resistance to RSI and tolerance to HTHH, accompanied by down-regulation of immunity- or thermotolerance-associated CaPR1, CaNPR1, CaDEF1, and CaHSP24. Hypersensitive response-mediated cell death and expression of the tested immunity- and thermotolerance-associated marker genes were induced by transient overexpression (TOE) of CabZIP63, but decreased by that of CabZIP63-SRDX. Additionally, binding of CabZIP63 to pCaWRKY40 was up-regulated by RSI or HTHH, and the transcript level of CaWRKY40 and binding of CaWRKY40 to the promoters of CaPR1, CaNPR1, CaDEF1 and CaHSP24 were up-regulated by TOE of CabZIP63. On the other hand, CabZIP63 was also up-regulated transcriptionally by TOE of CaWRKY40. The data suggest collectively that CabZIP63 directly or indirectly regulates the expression of CaWRKY40 at both the transcriptional and post-transcriptional level, forming a positive feedback loop with CaWRKY40 during pepper's response to RSI or HTHH. Altogether, our data will help to elucidate the underlying mechanism of crosstalk between pepper's response to RSI and HTHH.

  12. Pepper CabZIP63 acts as a positive regulator during Ralstonia solanacearum or high temperature–high humidity challenge in a positive feedback loop with CaWRKY40

    PubMed Central

    Shen, Lei; Liu, Zhiqin; Yang, Sheng; Yang, Tong; Liang, Jiaqi; Wen, Jiayu; Liu, Yanyan; Li, Jiazhi; Shi, Lanping; Tang, Qian; Shi, Wei; Hu, Jiong; Liu, Cailing; Zhang, Yangwen; Lin, Wei; Wang, Rongzhang; Yu, Huanxin; Mou, Shaoliang; Hussain, Ansar; Cheng, Wei; Cai, Hanyang; He, Li; Guan, Deyi; Wu, Yang; He, Shuilin

    2016-01-01

    CaWRKY40 is known to act as a positive regulator in the response of pepper (Capsicum annuum) to Ralstonia solanacearum inoculation (RSI) or high temperature–high humidity (HTHH), but the underlying mechanism remains elusive. Herein, we report that CabZIP63, a pepper bZIP family member, participates in this process by regulating the expression of CaWRKY40. CabZIP63 was found to localize in the nuclei, be up-regulated by RSI or HTHH, bind to promoters of both CabZIP63 (pCabZIP63) and CaWRKY40 (pCaWRKY40), and activate pCabZIP63- and pCaWRKY40-driven β-glucuronidase expression in a C- or G-box-dependent manner. Silencing of CabZIP63 by virus-induced gene silencing (VIGS) in pepper plants significantly attenuated their resistance to RSI and tolerance to HTHH, accompanied by down-regulation of immunity- or thermotolerance-associated CaPR1, CaNPR1, CaDEF1, and CaHSP24. Hypersensitive response-mediated cell death and expression of the tested immunity- and thermotolerance-associated marker genes were induced by transient overexpression (TOE) of CabZIP63, but decreased by that of CabZIP63-SRDX. Additionally, binding of CabZIP63 to pCaWRKY40 was up-regulated by RSI or HTHH, and the transcript level of CaWRKY40 and binding of CaWRKY40 to the promoters of CaPR1, CaNPR1, CaDEF1 and CaHSP24 were up-regulated by TOE of CabZIP63. On the other hand, CabZIP63 was also up-regulated transcriptionally by TOE of CaWRKY40. The data suggest collectively that CabZIP63 directly or indirectly regulates the expression of CaWRKY40 at both the transcriptional and post-transcriptional level, forming a positive feedback loop with CaWRKY40 during pepper’s response to RSI or HTHH. Altogether, our data will help to elucidate the underlying mechanism of crosstalk between pepper’s response to RSI and HTHH. PMID:26936828

  13. Canine DLA-79 gene: an improved typing method, identification of new alleles and its role in graft rejection and graft-versus-host disease.

    PubMed

    Venkataraman, G M; Geraghty, D; Fox, J; Graves, S S; Zellmer, E; Storer, B E; Torok-Storb, B J; Storb, R

    2013-04-01

    Developing a preclinical canine model that predicts outcomes for hematopoietic cell transplantation in humans requires a model that mimics the degree of matching between human donor and recipient major histocompatibility complex (MHC) genes. The polymorphic class I and class II genes in mammals are typically located in a single chromosome as part of the MHC complex. However, a divergent class I gene in dogs, designated dog leukocyte antigen-79 (DLA-79), is located on chromosome 18 while other MHC genes are on chromosome 12. This gene is not taken into account while DLA matching for transplantation. Though divergent, this gene shares significant similarity in sequence and exon-intron architecture with other class I genes, and is transcribed. Little is known about the polymorphisms of DLA-79 and their potential role in transplantation. This study was aimed at exploring the reason for high rate of rejection seen in DLA-matched dogs given reduced intensity conditioning, in particular, the possibility that DLA-79 allele mismatches may be the cause. We found that about 82% of 407 dogs typed were homozygous for a single, reference allele. Owing to the high prevalence of a single allele, 87 of the 108 dogs (∼80%) transplanted were matched for DLA-79 with their donor. In conclusion, we have developed an efficient method to type alleles of a divergent MHC gene in dogs and identified two new alleles. We did not find any statistical correlation between DLA-79 allele disparity and graft rejection or graft-versus-host disease, among our transplant dogs.

  14. Discovery, evaluation and distribution of haplotypes and new alleles of the Photoperiod-A1 gene in wheat.

    PubMed

    Muterko, Alexandr; Kalendar, Ruslan; Cockram, James; Balashova, Irina

    2015-05-01

    Photoperiod response in wheat is determined to a large extent by the homoeologous series of Photoperiod 1 (Ppd1) genes. In this study, Ppd-A1 genomic sequences from the 5' UTR and promoter region were analysed in 104 accessions of six tetraploid wheat species (Triticum dicoccoides, T. dicoccum, T. turgidum, T. polonicum, T. carthlicum, T. durum) and 102 accessions of six hexaploid wheat species (T. aestivum, T. compactum, T. sphaerococcum, T. spelta, T. macha, T. vavilovii). This data was supplemented with in silico analysis of publicly available sequences from 46 to 193 accessions of diploid and tetraploid wheat, respectively. Analysis of a region of the Ppd-A1 promoter identified thirteen haplotypes, which were divided in two haplogroups. Distribution of the Ppd-A1 haplogroups and haplotypes in wheat species, and their geographical distributions were analysed. Polymerase chain reaction combined with a heteroduplex mobility assay was subsequently used to efficiently discriminate between Ppd-A1 alleles, allowing identification of the Ppd-A1b haplotypes and haplogroups. The causes of anomalous migration of Ppd-A1 heteroduplexes in gels were found to be the localization of mismatches relative to the center of fragment, the cumulative effect of neighbouring polymorphic sites, and the location of mismatches within A/T-tracts. Analysis of the Ppd-A1 5' UTR in hexaploid wheat revealed a novel mutation within the "photoperiod critical" region in a subset of T. compactum accessions. This putative photoperiod insensitive allele (designated Ppd-A1a.4) includes a 684 bp deletion which spans region in common with deletions previously identified in other photoperiod insensitive Ppd1 alleles.

  15. Genotype and allele frequencies of drug-metabolizing enzymes and drug transporter genes affecting immunosuppressants in the Spanish white population.

    PubMed

    Bosó, Virginia; Herrero, María J; Buso, Enrique; Galán, Juan; Almenar, Luis; Sánchez-Lázaro, Ignacio; Sánchez-Plumed, Jaime; Bea, Sergio; Prieto, Martín; García, María; Pastor, Amparo; Sole, Amparo; Poveda, José Luis; Aliño, Salvador F

    2014-04-01

    Interpatient variability in drug response can be widely explained by genetically determined differences in metabolizing enzymes, drug transporters, and drug targets, leading to different pharmacokinetic and/or pharmacodynamic behaviors of drugs. Genetic variations affect or do not affect drug responses depending on their influence on protein activity and the relevance of such proteins in the pathway of the drug. Also, the frequency of such genetic variations differs among populations, so the clinical relevance of a specific variation is not the same in all of them. In this study, a panel of 33 single nucleotide polymorphisms in 14 different genes (ABCB1, ABCC2, ABCG2, CYP2B6, CYP2C19, CYP2C9, CYP3A4, CYP3A5, MTHFR, NOD2/CARD15, SLCO1A2, SLCO1B1, TPMT, and UGT1A9), encoding for the most relevant metabolizing enzymes and drug transporters relating to immunosuppressant agents, was analyzed to determine the genotype profile and allele frequencies in comparison with HapMap data. A total of 570 Spanish white recipients and donors of solid organ transplants were included. In 24 single nucleotide polymorphisms, statistically significant differences in allele frequency were observed. The largest differences (>100%) occurred in ABCB1 rs2229109, ABCG2 rs2231137, CYP3A5 rs776746, NOD2/CARD15 rs2066844, TPMT rs1800462, and UGT1A9 rs72551330. In conclusion, differences were recorded between the Spanish and other white populations in terms of allele frequency and genotypic distribution. Such differences may have implications in relation to dose requirements and drug-induced toxicity. These data are important for further research to help explain interindividual pharmacokinetic and pharmacodynamic variability in response to drug therapy.

  16. Protective Effect of R Allele of PON1 Gene on the Coronary Artery Disease in the Presence of Specific Genetic Background

    PubMed Central

    Balcerzyk, Anna; Zak, Iwona; Krauze, Jolanta

    2008-01-01

    Background: Genetic susceptibility to CAD may be determined by polymorphic variants of genes encoding isoforms involved in the processes important in the pathogenesis of atherosclerosis, including lipids disorders. Participation of single polymorphic variants is relatively small, however its significance may increase in the presence of specific genetic or environmental background. Aim: The aim of the study was an evaluation a possible association between single polymorphic variants of PON1, APOE, ABCA1 and PPARA genes and CAD and looking for specific multigene genotype patterns which differentiate study groups. Materials and methods: We studied 358 subjects:178 patients with angiographically confirmed CAD and 180 blood donors without history of CAD. Polymorphisms were genotyped using PCR-RFLP method. Results: We observed statistically significant differences in the frequencies of R allele and R allele carriers of PON1 gene between CAD and controls. The distribution of genotypes and alleles of other analyzed genes did not differentiate the study groups, however the presence of specific genotypes (APOE– ɛ3ɛ3, ɛ3ɛ2, ABCA1 – AG, PPARA – GG) increased the protective effect of R allele. Conclusion: The present study revealed an independent protective association between carrier-state of PON1 R allele and CAD. This protective effect was especially strong in the presence of specific genotype arrangements of other analyzed genes. PMID:18219093

  17. The homeologous Zea mays gigantea genes: characterization of expression and novel mutant alleles

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The two homeologous Zea mays gigantea (gi) genes, gi1 and gi2, arose from the last genome duplication event in the maize lineage. Homologs of these genes in other species are required for correct circadian rhythms and proper regulation of growth and development. Here we characterized the expression ...

  18. WRKY Transcription Factors: Molecular Regulation and Stress Responses in Plants

    PubMed Central

    Phukan, Ujjal J.; Jeena, Gajendra S.; Shukla, Rakesh K.

    2016-01-01

    Plants in their natural habitat have to face multiple stresses simultaneously. Evolutionary adaptation of developmental, physiological, and biochemical parameters give advantage over a single window of stress but not multiple. On the other hand transcription factors like WRKY can regulate diverse responses through a complicated network of genes. So molecular orchestration of WRKYs in plant may provide the most anticipated outcome of simultaneous multiple responses. Activation or repression through W-box and W-box like sequences is regulated at transcriptional, translational, and domain level. Because of the tight regulation involved in specific recognition and binding of WRKYs to downstream promoters, they have become promising candidate for crop improvement. Epigenetic, retrograde and proteasome mediated regulation enable WRKYs to attain the dynamic cellular homeostatic reprograming. Overexpression of several WRKYs face the paradox of having several beneficial affects but with some unwanted traits. These overexpression-associated undesirable phenotypes need to be identified and removed for proper growth, development and yeild. Taken together, we have highlighted the diverse regulation and multiple stress response of WRKYs in plants along with the future prospects in this field of research. PMID:27375634

  19. Alleles of the yeast Pms1 mismatch-repair gene that differentially affect recombination- and replication-related processes.

    PubMed Central

    Welz-Voegele, Caroline; Stone, Jana E; Tran, Phuoc T; Kearney, Hutton M; Liskay, R Michael; Petes, Thomas D; Jinks-Robertson, Sue

    2002-01-01

    Mismatch-repair (MMR) systems promote eukaryotic genome stability by removing errors introduced during DNA replication and by inhibiting recombination between nonidentical sequences (spellchecker and antirecombination activities, respectively). Following a common mismatch-recognition step effected by MutS-homologous Msh proteins, homologs of the bacterial MutL ATPase (predominantly the Mlh1p-Pms1p heterodimer in yeast) couple mismatch recognition to the appropriate downstream processing steps. To examine whether the processing steps in the spellchecker and antirecombination pathways might differ, we mutagenized the yeast PMS1 gene and screened for mitotic separation-of-function alleles. Two alleles affecting only the antirecombination function of Pms1p were identified, one of which changed an amino acid within the highly conserved ATPase domain. To more specifically address the role of ATP binding/hydrolysis in MMR-related processes, we examined mutations known to compromise the ATPase activity of Pms1p or Mlh1p with respect to the mitotic spellchecker and antirecombination activities and with respect to the repair of mismatches present in meiotic recombination intermediates. The results of these analyses confirm a differential requirement for the Pms1p ATPase activity in replication vs. recombination processes, while demonstrating that the Mlh1p ATPase activity is important for all examined MMR-related functions. PMID:12454061

  20. A recombined allele of the lipase gene CEL and its pseudogene CELP confers susceptibility to chronic pancreatitis.

    PubMed

    Fjeld, Karianne; Weiss, Frank Ulrich; Lasher, Denise; Rosendahl, Jonas; Chen, Jian-Min; Johansson, Bente B; Kirsten, Holger; Ruffert, Claudia; Masson, Emmanuelle; Steine, Solrun J; Bugert, Peter; Cnop, Miriam; Grützmann, Robert; Mayerle, Julia; Mössner, Joachim; Ringdal, Monika; Schulz, Hans-Ulrich; Sendler, Matthias; Simon, Peter; Sztromwasser, Paweł; Torsvik, Janniche; Scholz, Markus; Tjora, Erling; Férec, Claude; Witt, Heiko; Lerch, Markus M; Njølstad, Pål R; Johansson, Stefan; Molven, Anders

    2015-05-01

    Carboxyl ester lipase is a digestive pancreatic enzyme encoded by the CEL gene. Mutations in CEL cause maturity-onset diabetes of the young as well as pancreatic exocrine dysfunction. Here we describe a hybrid allele (CEL-HYB) originating from a crossover between CEL and its neighboring pseudogene, CELP. In a discovery series of familial chronic pancreatitis cases, we observed CEL-HYB in 14.1% (10/71) of cases compared to 1.0% (5/478) of controls (odds ratio (OR) = 15.5; 95% confidence interval (CI) = 5.1-46.9; P = 1.3 × 10(-6) by two-tailed Fisher's exact test). In three replication studies of nonalcoholic chronic pancreatitis, we identified CEL-HYB in a total of 3.7% (42/1,122) cases and 0.7% (30/4,152) controls (OR = 5.2; 95% CI = 3.2-8.5; P = 1.2 × 10(-11); formal meta-analysis). The allele was also enriched in alcoholic chronic pancreatitis. Expression of CEL-HYB in cellular models showed reduced lipolytic activity, impaired secretion, prominent intracellular accumulation and induced autophagy. These findings implicate a new pathway distinct from the protease-antiprotease system of pancreatic acinar cells in chronic pancreatitis.

  1. On the use of allelic transmission rates for assessing gene-by-environment interaction in case-parent trios.

    PubMed

    Shin, Ji-Hyung; McNeney, Brad; Graham, Jinko

    2010-09-01

    Allelic transmission rates from parents to cases are frequently stratified by an environmental risk factor E and compared, with heterogeneity interpreted as gene-environment interaction or GxE. Though generally invalid, such analyses continue to appear. We revisit why heterogeneity is not equivalent to GxE in a range of settings not considered previously. The objective is a fuller understanding of the bias in transmission rates and what is driving it. Extending previously published findings, we derive parental mating-type probabilities in cases and use them to obtain transmission rates, which we then compare to GxE. Through simulation, we investigate the practical implications of the bias for a transmission-based test of GxE. We find that general population characteristics distort the picture of GxE obtained from transmission rates: the stratum-specific mating-type probabilities under G - E dependence and the allele frequency under independence. Furthermore, the transmission-based test has inflated error rates relative to a likelihood-based test. Our investigation provides further insight into how and why transmission-based tests and descriptive summaries can mislead about GxE. For exploring GxE, we suggest graphical displays of the transmission rates within parental mating types, as they are robust to population stratification and the penetrance model.

  2. EcoTILLING-Based Association Mapping Efficiently Delineates Functionally Relevant Natural Allelic Variants of Candidate Genes Governing Agronomic Traits in Chickpea

    PubMed Central

    Bajaj, Deepak; Srivastava, Rishi; Nath, Manoj; Tripathi, Shailesh; Bharadwaj, Chellapilla; Upadhyaya, Hari D.; Tyagi, Akhilesh K.; Parida, Swarup K.

    2016-01-01

    The large-scale mining and high-throughput genotyping of novel gene-based allelic variants in natural mapping population are essential for association mapping to identify functionally relevant molecular tags governing useful agronomic traits in chickpea. The present study employs an alternative time-saving, non-laborious and economical pool-based EcoTILLING approach coupled with agarose gel detection assay to discover 1133 novel SNP allelic variants from diverse coding and regulatory sequence components of 1133 transcription factor (TF) genes by genotyping in 192 diverse desi and kabuli chickpea accessions constituting a seed weight association panel. Integrating these SNP genotyping data with seed weight field phenotypic information of 192 structured association panel identified eight SNP alleles in the eight TF genes regulating seed weight of chickpea. The associated individual and combination of all SNPs explained 10–15 and 31% phenotypic variation for seed weight, respectively. The EcoTILLING-based large-scale allele mining and genotyping strategy implemented for association mapping is found much effective for a diploid genome crop species like chickpea with narrow genetic base and low genetic polymorphism. This optimized approach thus can be deployed for various genomics-assisted breeding applications with optimal expense of resources in domesticated chickpea. The seed weight-associated natural allelic variants and candidate TF genes delineated have potential to accelerate marker-assisted genetic improvement of chickpea. PMID:27148286

  3. [The differences of the effects of Vrd1 and Ppd-D1 gene alleles on winterhardiness, frost resistance, and yield in winter wheat].

    PubMed

    Mokanu, N V; Faĭt, V I

    2008-01-01

    The influence of allelic differences of Vrd1 and Ppd-D1 genes on winterhardiness, frost resistance, yield and its components was studied in recombinant-inbred F5 lines of Odesskaya 16/Bezostaya 1. From 9 to 15% differences in the resistance of recombinant-inbred lines were determined by alternative alleles of Vrd1 gene and 10-16% of Ppd-D1 gene. Interaction of vrd1 and Ppd-D1a alleles led to the higher winterhardiness and frost resistance of tillered plants during the winter. At the same time the significant increase of the period to heading, plant height and the tendency of yield reduction were revealed for vrd1 vrd1 Ppd-D1a Ppd-D1a lines when compared to the lines of Vrd1 Vrd1 Ppd-D1a Ppd-D1a genotype.

  4. OsWRKY62 is a negative regulator of basal and Xa21-mediated defense against Xanthomonas oryzae pv. oryzae in rice

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The rice Xa21 gene, which confers resistance to the bacterial pathogen Xanthomonas oryzae pv. oryzae (Xoo), encodes a receptor-like kinase. Few components involved in transducing the Xa21-mediated defense response have yet been identified. It is reported that XA21 binds to a WRKY transcription fac...

  5. [Double mutant alleles in the EXT1 gene not previously reported in a teenager with hereditary multiple exostoses].

    PubMed

    Cammarata-Scalisi, Francisco; Cozar, Mónica; Grinberg, Daniel; Balcells, Susana; Asteggiano, Carla G; Martínez-Domenech, Gustavo; Bracho, Ana; Sánchez, Yanira; Stock, Frances; Delgado-Luengo, Wilmer; Zara-Chirinos, Carmen; Chacín, José Antonio

    2015-04-01

    Hereditary forms of multiple exostoses, now called EXT1/EXT2-CDG within Congenital Disorders of Glycosylation, are the most common benign bone tumors in humans and clinical description consists of the formation of several cartilage-capped bone tumors, usually benign and localized in the juxta-epiphyseal region of long bones, although wide body dissemination in severe cases is not uncommon. Onset of the disease is variable ranging from 2-3 years up to 13-15 years with an estimated incidence ranging from 1/18,000 to 1/50,000 cases in European countries. We present a double mutant alleles in the EXT1 gene not previously reported in a teenager and her family with hereditary multiple exostoses.

  6. Divergence and gene flow among Darwin's finches: a genome-wide view of adaptive radiation driven by interspecies allele sharing

    PubMed Central

    Palmer, Daniela H.; Kronforst, Marcus R.

    2015-01-01

    A recent analysis of the genomes of Darwin's finches revealed extensive interspecies allele sharing throughout the history of the radiation and identified a key locus responsible for morphological evolution in this group. The radiation of Darwin's finches on the Galápagos archipelago has long been regarded as an iconic study system for field ecology and evolutionary biology. Coupled with an extensive history of field work, these latest findings affirm the increasing acceptance of introgressive hybridization, or gene flow between species, as a significant contributor to adaptive evolution. Here we review and discuss these findings in relation to both classical work on Darwin's finches and contemporary work showing similar evolutionary signatures in other biological systems. The continued unification of genomic data with field biology promises to further elucidate the molecular basis of adaptation in Darwin's finches and well beyond. PMID:26200327

  7. No allelic association between Parkinson`s disease and dopamine D2, D3, and D4 receptor gene polymorphisms

    SciTech Connect

    Nanko, S.; Hattori, M.; Dai, X.Y.

    1994-12-15

    Parkinson`s disease is thought to be caused by a combination of unknown environmental, genetic, and degenerative factors. Evidence from necropsy brain samples and pharmacokinetics suggests involvement of dopamine receptors in the pathogenesis or pathophysiology of Parkinson`s disease. Genetic association studies between Parkinson`s disease and dopamine D2, D3 and D4 receptor gene polymorphisms were conducted. The polymorphism was examined in 71 patients with Parkinson`s disease and 90 controls. There were no significant differences between two groups in allele frequencies at the D2, D3, and D4 dopamine receptor loci. Our findings do not support the hypothesis that susceptibility to Parkinson`s disease is associated with the dopamine receptor polymorphisms examined. 35 refs., 2 tabs.

  8. Genetic Adaptation to Climate in White Spruce Involves Small to Moderate Allele Frequency Shifts in Functionally Diverse Genes.

    PubMed

    Hornoy, Benjamin; Pavy, Nathalie; Gérardi, Sébastien; Beaulieu, Jean; Bousquet, Jean

    2015-11-11

    Understanding the genetic basis of adaptation to climate is of paramount importance for preserving and managing genetic diversity in plants in a context of climate change. Yet, this objective has been addressed mainly in short-lived model species. Thus, expanding knowledge to nonmodel species with contrasting life histories, such as forest trees, appears necessary. To uncover the genetic basis of adaptation to climate in the widely distributed boreal conifer white spruce (Picea glauca), an environmental association study was conducted using 11,085 single nucleotide polymorphisms representing 7,819 genes, that is, approximately a quarter of the transcriptome.Linear and quadratic regressions controlling for isolation-by-distance, and the Random Forest algorithm, identified several dozen genes putatively under selection, among which 43 showed strongest signals along temperature and precipitation gradients. Most of them were related to temperature. Small to moderate shifts in allele frequencies were observed. Genes involved encompassed a wide variety of functions and processes, some of them being likely important for plant survival under biotic and abiotic environmental stresses according to expression data. Literature mining and sequence comparison also highlighted conserved sequences and functions with angiosperm homologs.Our results are consistent with theoretical predictions that local adaptation involves genes with small frequency shifts when selection is recent and gene flow among populations is high. Accordingly, genetic adaptation to climate in P. glauca appears to be complex, involving many independent and interacting gene functions, biochemical pathways, and processes. From an applied perspective, these results shall lead to specific functional/association studies in conifers and to the development of markers useful for the conservation of genetic resources.

  9. Genetic Adaptation to Climate in White Spruce Involves Small to Moderate Allele Frequency Shifts in Functionally Diverse Genes

    PubMed Central

    Hornoy, Benjamin; Pavy, Nathalie; Gérardi, Sébastien; Beaulieu, Jean; Bousquet, Jean

    2015-01-01

    Understanding the genetic basis of adaptation to climate is of paramount importance for preserving and managing genetic diversity in plants in a context of climate change. Yet, this objective has been addressed mainly in short-lived model species. Thus, expanding knowledge to nonmodel species with contrasting life histories, such as forest trees, appears necessary. To uncover the genetic basis of adaptation to climate in the widely distributed boreal conifer white spruce (Picea glauca), an environmental association study was conducted using 11,085 single nucleotide polymorphisms representing 7,819 genes, that is, approximately a quarter of the transcriptome. Linear and quadratic regressions controlling for isolation-by-distance, and the Random Forest algorithm, identified several dozen genes putatively under selection, among which 43 showed strongest signals along temperature and precipitation gradients. Most of them were related to temperature. Small to moderate shifts in allele frequencies were observed. Genes involved encompassed a wide variety of functions and processes, some of them being likely important for plant survival under biotic and abiotic environmental stresses according to expression data. Literature mining and sequence comparison also highlighted conserved sequences and functions with angiosperm homologs. Our results are consistent with theoretical predictions that local adaptation involves genes with small frequency shifts when selection is recent and gene flow among populations is high. Accordingly, genetic adaptation to climate in P. glauca appears to be complex, involving many independent and interacting gene functions, biochemical pathways, and processes. From an applied perspective, these results shall lead to specific functional/association studies in conifers and to the development of markers useful for the conservation of genetic resources. PMID:26560341

  10. Lack of association between TaqI A1 Allele of dopamine D2 receptor gene and alcohol-use disorders in Atayal natives of Taiwan

    SciTech Connect

    Chia-Hsiang Chen; Shih-Hsiang Chien; Hai-Gwo Hwu

    1996-09-20

    Association studies between the A1 allele of the dopamine D2 receptor (DRD2) gene TaqI A polymorphism and alcoholism remain controversial. A recent study from Japan demonstrated that the A1 allele is associated with severe alcoholism in the Japanese population. We were interested in knowing if this association also exists in the Atayals of Taiwan, who were found to have a higher prevalence of alcohol-use disorders than the Han Chinese in Taiwan. Genotype and allele frequencies were determined in alcohol-abusing, alcohol-dependent, and nonalcoholic control Atayal natives in Taiwan. A1 allele frequencies in alcohol-dependent, alcohol-abusing, and normal control Atayals were 0.39, 0.42, and 0.39, respectively. No difference in A1 allele frequency was found among these three groups. Our data do not support the hypothesis that the A1 allele of the TaqI A polymorphism of the DRD2 gene increases susceptibility to alcohol-use disorders in the Atayals of Taiwan. 18 refs., 1 tab.

  11. Allelic variants of IL1R1 gene associate with severe hand osteoarthritis

    PubMed Central

    2010-01-01

    Background In search for genes predisposing to osteoarthritis (OA), several genome wide scans have provided evidence for linkage on 2q. In this study we targeted a 470 kb region on 2q11.2 presenting the locus with most evidence for linkage to severe OA of distal interphalangeal joints (DIP) in our genome wide scan families. Methods We genotyped 32 single nucleotide polymorphisms (SNPs) in this 470 kb region comprising six genes belonging to the interleukin 1 superfamily and monitored for association with individual SNPs and SNP haplotypes among severe familial hand OA cases (material extended from our previous linkage study; n = 134), unrelated end-stage bilateral primary knee OA cases (n = 113), and population based controls (n = 436). Results Four SNPs in the IL1R1 gene, mapping to a 125 kb LD block, provided evidence for association with hand OA in family-based and case-control analysis, the strongest association being with SNP rs2287047 (p-value = 0.0009). Conclusions This study demonstrates an association between severe hand OA and IL1R1 gene. This gene represents a highly relevant biological candidate since it encodes protein that is a known modulator of inflammatory processes associated with joint destruction and resides within a locus providing consistent evidence for linkage to hand OA. As the observed association did not fully explain the linkage obtained in the previous study, it is plausible that also other variants in this genome region predispose to hand OA. PMID:20353565

  12. Screens for Extragenic Mutations That Fail to Complement Act1 Alleles Identify Genes That Are Important for Actin Function in Saccharomyces Cerevisiae

    PubMed Central

    Welch, M. D.; Vinh, DBN.; Okamura, H. H.; Drubin, D. G.

    1993-01-01

    Null mutations in SAC6 and ABP1, genes that encode actin-binding proteins, failed to complement the temperature-sensitive phenotype caused by a mutation in the ACT1 gene. To identify novel genes whose protein products interact with actin, mutations that fail to complement act1-1 or act1-4, two temperature-sensitive alleles of ACT1, were isolated. A total of 14 extragenic noncomplementing mutations and 12 new alleles of ACT1 were identified in two independent screens. The 14 extragenic noncomplementing mutations represent alleles of at least four different genes, ANC1, ANC2, ANC3 and ANC4 (Actin NonComplementing). Mutations in the ANC1 gene were shown to cause osmosensitivity and defects in actin organization; phenotypes that are similar to those caused by act1 mutations. We conclude that the ANC1 gene product plays an important role in actin cytoskeletal function. The 12 new alleles of ACT1 will be useful for further elucidation of the functions of actin in yeast. PMID:8243992

  13. A two allele DNA polymorphism of the human phenylethanolamine N-methyltransferase (hPNMT) gene identified by HGIA I

    SciTech Connect

    Hoehe, M.R.; Berrettini, W.H. ); Baetge, E.E. )

    1989-01-25

    An 8 kb DNA fragment (Eco RI) of the human phenylethanolamine N-methyltransferase gene (hPNMT), selected from a 14 kb Eco RI fragment isolated from a lambda Charon 3A human lymphocyte genomic library and subcloned into pUC18, was used as a probe. This human genomic fragment contained the first 1,923 bp of 5{prime} flanking DNA, the hPNMT structural gene spanning 2,070 bp in total (composed of three exons (225, 208, 524 bp) and two introns (1,000 and 113 bp)), and 3.8 kb of 3{prime} flanking DNA (1). Hybridization of human genomic DNA digested with HgiA I identifies a two allele polymorphism with bands at 3.0 kb (A) and 2.5 kb (B). The hPNMT gene has been assigned to chromosome 17. Co-dominant segregation in two families with two generations was observed. The number of meioses scorred was 18.

  14. Allele-specific marker development and selection efficiencies for both flavonoid 3'-hydroxylase and flavonoid 3',5'-hydroxylase genes in soybean subgenus soja.

    PubMed

    Guo, Yong; Qiu, Li-Juan

    2013-06-01

    Color is one of the phenotypic markers mostly used to study soybean (Glycine max L. Merr.) genetic, molecular and biochemical processes. Two P450-dependent mono-oxygenases, flavonoid 3'-hydroxylase (F3'H; EC1.14.3.21) and flavonoid 3',5'-hydroxylase (F3'5'H, EC1.14.13.88), both catalyzing the hydroxylation of the B-ring in flavonoids, play an important role in coloration. Previous studies showed that the T locus was a gene encoding F3'H and the W1 locus co-segregated with a gene encoding F3'5'H in soybean. These two genetic loci have identified to control seed coat, flower and pubescence colors. However, the allelic distributions of both F3'H and F3'5'H genes in soybean were unknown. In this study, three novel alleles were identified (two of four alleles for GmF3'H and one of three alleles for GmF3'5'H). A set of gene-tagged markers was developed and verified based on the sequence diversity of all seven alleles. Furthermore, the markers were used to analyze soybean accessions including 170 cultivated soybeans (G. max) from a mini core collection and 102 wild soybeans (G. soja). For both F3'H and F3'5'H, the marker selection efficiencies for pubescence color and flower color were determined. The results showed that one GmF3'H allele explained 92.2 % of the variation in tawny and two gmf3'h alleles explained 63.8 % of the variation in gray pubescence colors. In addition, two GmF3'5'H alleles and one gmF3'5'h allele explained 94.0 % of the variation in purple and 75.3 % in white flowers, respectively. By the combination of the two loci, seed coat color was determined. In total, 90.9 % of accessions possessing both the gmf3'h-b and gmf3'5'h alleles had yellow seed coats. Therefore, seed coat colors are controlled by more than two loci.

  15. Allelic and genotypic frequencies in polymorphic Booroola fecundity gene and their association with multiple birth and postnatal growth in Chhotanagpuri sheep

    PubMed Central

    Oraon, Thanesh; Singh, D. K.; Ghosh, Mayukh; Kullu, S. S.; Kumar, Rajesh; Singh, L. B.

    2016-01-01

    Aim: Chhotanagpuri breed of sheep reared for mutton in Jharkhand, India, having problem of low litter size and body weight. The response of genetic improvement for traits with low heritability through traditional selection method is time-consuming. Therefore, marker-assisted selection based on a polymorphism study of suitable candidate gene can response quickly. Thus, this study was aimed at identification of different allelic and genotypic frequencies of Booroola fecundity (FecB) gene and its association with multiple birth and postnatal growth in Chhotanagpuri sheep. Materials and Methods: DNA isolation and gene-specific amplification of FecB gene was performed from blood samples of from 92 Chhotanagpuri lambs maintained under similar feeding and management conditions. Custom nucleotide sequencing and single-strand conformational polymorphism analysis were performed to identify different genotypes with respect to the target gene. Statistical analysis was performed for determination of allelic and genotypic frequencies of FecB gene polymorphisms and its association with multiple birth and postnatal growth of lambs from birth to 52 weeks age. Results: “AA,” “AB,” and “BB” genotypes were found at locus-1 as it is polymorphic for FecB gene while locus-2 was found to be monomorphic for FecB gene. Higher frequency of “A” allele at locus-1 was found in single born lambs, whereas “B” allele was predominant among multiple born lambs. The lambs having “BB” genotype weighed significantly (p≤0.01) heavier than those of “AB” and “AA” genotype at 52 weeks of age. Conclusion: “BB” genotype has emerged as favored genotype for multiple births and better growth indicator. Therefore, homozygous lambs for “B” allele should be selected and utilized in breeding program for better growth rate. PMID:27956784

  16. Single-gene speciation with pleiotropy: effects of allele dominance, population size, and delayed inheritance.

    PubMed

    Yamamichi, Masato; Sasaki, Akira

    2013-07-01

    Single-gene speciation is considered to be unlikely, but an excellent example is found in land snails, in which a gene for left-right reversal has given rise to new species multiple times. This reversal might be facilitated by their small population sizes and maternal effect (i.e., "delayed inheritance," in which an individual's phenotype is determined by the genotype of its mother). Recent evidence suggests that a pleiotropic effect of the speciation gene on antipredator survival may also promote speciation. Here we theoretically demonstrate that, without a pleiotropic effect, in small populations the fixation probability of a recessive mutant is higher than a dominant mutant, but they are identical for large populations and sufficiently weak selection. With a pleiotropic effect that increases mutant viability, a dominant mutant has a higher fixation probability if the strength of viability selection is sufficiently greater than that of reproductive incompatibility, whereas a recessive mutant has a higher fixation probability otherwise. Delayed inheritance increases the fixation probability of a mutant if viability selection is sufficiently weaker than reproductive incompatibility. Our results clarify the conflicting effects of viability selection and positive frequency-dependent selection due to reproductive incompatibility and provide a new perspective to single-gene speciation theory.

  17. Evidence of extensive non-allelic gene conversion among LTR elements in the human genome

    PubMed Central

    Trombetta, Beniamino; Fantini, Gloria; D’Atanasio, Eugenia; Sellitto, Daniele; Cruciani, Fulvio

    2016-01-01

    Long Terminal Repeats (LTRs) are nearly identical DNA sequences found at either end of Human Endogenous Retroviruses (HERVs). The high sequence similarity that exists among different LTRs suggests they could be substrate of ectopic gene conversion events. To understand the extent to which gene conversion occurs and to gain new insights into the evolutionary history of these elements in humans, we performed an intra-species phylogenetic study of 52 LTRs on different unrelated Y chromosomes. From this analysis, we obtained direct evidence that demonstrates the occurrence of ectopic gene conversion in several LTRs, with donor sequences located on both sex chromosomes and autosomes. We also found that some of these elements are characterized by an extremely high density of polymorphisms, showing one of the highest nucleotide diversities in the human genome, as well as a complex patchwork of sequences derived from different LTRs. Finally, we highlighted the limits of current short-read NGS studies in the analysis of genetic diversity of the LTRs in the human genome. In conclusion, our comparative re-sequencing analysis revealed that ectopic gene conversion is a common event in the evolution of LTR elements, suggesting complex genetic links among LTRs from different chromosomes. PMID:27346230

  18. Comprehensively Evaluating cis-Regulatory Variation in the Human Prostate Transcriptome by Using Gene-Level Allele-Specific Expression

    PubMed Central

    Larson, Nicholas B.; McDonnell, Shannon; French, Amy J.; Fogarty, Zach; Cheville, John; Middha, Sumit; Riska, Shaun; Baheti, Saurabh; Nair, Asha A.; Wang, Liang; Schaid, Daniel J.; Thibodeau, Stephen N.

    2015-01-01

    The identification of cis-acting regulatory variation in primary tissues has the potential to elucidate the genetic basis of complex traits and further our understanding of transcriptomic diversity across cell types. Expression quantitative trait locus (eQTL) association analysis using RNA sequencing (RNA-seq) data can improve upon the detection of cis-acting regulatory variation by leveraging allele-specific expression (ASE) patterns in association analysis. Here, we present a comprehensive evaluation of cis-acting eQTLs by analyzing RNA-seq gene-expression data and genome-wide high-density genotypes from 471 samples of normal primary prostate tissue. Using statistical models that integrate ASE information, we identified extensive cis-eQTLs across the prostate transcriptome and found that approximately 70% of expressed genes corresponded to a significant eQTL at a gene-level false-discovery rate of 0.05. Overall, cis-eQTLs were heavily concentrated near the transcription start and stop sites of affected genes, and effects were negatively correlated with distance. We identified multiple instances of cis-acting co-regulation by using phased genotype data and discovered 233 SNPs as the most strongly associated eQTLs for more than one gene. We also noted significant enrichment (25/50, p = 2E−5) of previously reported prostate cancer risk SNPs in prostate eQTLs. Our results illustrate the benefit of assessing ASE data in cis-eQTL analyses by showing better reproducibility of prior eQTL findings than of eQTL mapping based on total expression alone. Altogether, our analysis provides extensive functional context of thousands of SNPs in prostate tissue, and these results will be of critical value in guiding studies examining disease of the human prostate. PMID:25983244

  19. Comprehensively evaluating cis-regulatory variation in the human prostate transcriptome by using gene-level allele-specific expression.

    PubMed

    Larson, Nicholas B; McDonnell, Shannon; French, Amy J; Fogarty, Zach; Cheville, John; Middha, Sumit; Riska, Shaun; Baheti, Saurabh; Nair, Asha A; Wang, Liang; Schaid, Daniel J; Thibodeau, Stephen N

    2015-06-04

    The identification of cis-acting regulatory variation in primary tissues has the potential to elucidate the genetic basis of complex traits and further our understanding of transcriptomic diversity across cell types. Expression quantitative trait locus (eQTL) association analysis using RNA sequencing (RNA-seq) data can improve upon the detection of cis-acting regulatory variation by leveraging allele-specific expression (ASE) patterns in association analysis. Here, we present a comprehensive evaluation of cis-acting eQTLs by analyzing RNA-seq gene-expression data and genome-wide high-density genotypes from 471 samples of normal primary prostate tissue. Using statistical models that integrate ASE information, we identified extensive cis-eQTLs across the prostate transcriptome and found that approximately 70% of expressed genes corresponded to a significant eQTL at a gene-level false-discovery rate of 0.05. Overall, cis-eQTLs were heavily concentrated near the transcription start and stop sites of affected genes, and effects were negatively correlated with distance. We identified multiple instances of cis-acting co-regulation by using phased genotype data and discovered 233 SNPs as the most strongly associated eQTLs for more than one gene. We also noted significant enrichment (25/50, p = 2E-5) of previously reported prostate cancer risk SNPs in prostate eQTLs. Our results illustrate the benefit of assessing ASE data in cis-eQTL analyses by showing better reproducibility of prior eQTL findings than of eQTL mapping based on total expression alone. Altogether, our analysis provides extensive functional context of thousands of SNPs in prostate tissue, and these results will be of critical value in guiding studies examining disease of the human prostate.

  20. Variants of the mannose-binding lectin gene in the Benin population: heterozygosity for the p.G57E allele may confer a selective advantage.

    PubMed

    Dossou-Yovo, Omer Placide; Lapoumeroulie, Claudine; Hauchecorne, Michelle; Zaccaria, Isabelle; Ducrocq, Rolande; Krishnamoorthy, Rajagopal; Rahimy, Mohamed Chérif; Elion, Jacques

    2007-12-01

    Human mannose-binding lectin (MBL) plays an important role in innate immunity. MBL deficiency is associated with mutations in the promoter region and in exon 1 of the MBL2 gene. Such deficiency has been correlated with elevated incidence of infections in infancy and in immunocompromised adults. We determined the distribution profile of the MBL2 gene variants in the general population of Benin (West Africa) and in a vulnerable subset of children with sickle cell disease (SCD) (SS homozygotes). Five hundred forty-two healthy individuals (274 newborns, 268 adults) and 128 patients with SCD (35 newborns, 93 children) were screened for the common variant alleles in the MBL2 secretor haplotype region (exon 1 and promoter). The p.G57E variant allele was the most frequent allele compared to p.G54D (27.5% vs. 1.6%, respectively). The p.R52C allele was not found in this population. There was no difference in allele or genotype frequencies between healthy newborns and newborns with SCD. Alleles associated with MBL deficiency were more frequent in adults than in newborns (69.8% vs. 57.3%, respectively; p = 0.002). This enrichment was exclusively due to an elevated proportion of heterozygotes for the p.G57E allele (47.0% vs. 35.3%, respectively; p = 0.004), supporting a potential selective advantage of this genotype. Our results, compared to those reported in other African countries, support the implication of the MBL2 gene in various major infections in Africa, such as meningitis and tuberculosis in HIV-positive patients.

  1. Variants of the mannose-binding lectin gene in the Benin population: heterozygosity for the p.G57E allele may confer a selective advantage. 2007.

    PubMed

    Dossou-Yovo, Omer Placide; Lapoumeroulie, Claudine; Hauchecorne, Michelle; Zaccaria, Isabelle; Ducrocq, Rolande; Krishnamoorthy, Rajagopal; Rahimy, Mohamed Chérif; Elion, Jacques

    2009-12-01

    Human mannose- binding lectin (MBL) plays an important role in innate immunity. MBL deficiency is associated with mutations in the promoter region and in exon 1 of the MBL2 gene. Such deficiency has been correlated with elevated incidence of infections in infancy and in immunocompromised adults. We determined the distribution profile of the MBL2 gene variants in the general population of Benin (West Africa) and in a vulnerable subset of children with sickle cell disease (SCD) (SS homozygotes). Five hundred forty-two healthy individuals (274 newborns, 268 adults) and 128 patients with SCD (35 newborns, 93 children) were screened for the common variant alleles in the MBL2 secretor haplotype region (exon 1 and promoter). The p.G57E variant allele was the most frequent allele compared to p.G54D (27.5% vs. 1.6%, respectively). The p.R52C allele was not found in this population. There was no difference in allele or genotype frequencies between healthy newborns and newborns with SCD. Alleles associated with MBL deficiency were more frequent in adults than in newborns (69.8% vs. 57.3%, respectively; p=0.002). This enrichment was exclusively due to an elevated proportion of heterozygotes for the p.G57E allele (47.0% vs. 35.3%,respectively; p=0.004), supporting a potential selective advantage of this genotype. Our results, compared to those reported in other African countries, support the implication of the MBL2 gene in various major infections in Africa, such as meningitis and tuberculosis in HIV- positive patients.

  2. Characterization of a protein kinase gene in allelic association with the spinal muscular atrophy locus

    SciTech Connect

    Wang, C.H.; Carter, T.A.; Kleyn, P.W.

    1994-09-01

    A protein kinase gene has been identified from a 400 Kb minimal genetic region which defines the spinal muscular atrophy (SMA) locus. A highly polymorphic microsatellite marker (D5S1414) isolated from a yeast artificial chromosome (YAC) clone within this interval detects linkage disequilibrium with the SMA locus in 32 Polish families (Yule`s coefficient: 0.92) and maps to an intron of the protein kinase gene. Exon amplification was used to isolate coding sequences from a YAC-derived phage subclone containing D5S1414. Five exons were identified and a GenBank search using the BLAST program showed complete homology of these exons with a protein kinase gene. The gene is expressed in all tissues checked to far. Full-length cDNAs have been identified from both normal and SMA brain libraries and by reverse-transcriptase (RT) PCR from RNA of various tissues. The cDNA sequences will be reported. The genomic sequences flanking each exon were determined by direct sequencing of the homologous phage. The marker D5S1414 was located within the intronic sequence between exons 6 and 7. To screen for disease mutations, PCR was performed across each exon including the flanking splice sites in normal controls and SMA samples shown to be homozygous across the region by haplotyping. Comparative sequence analysis of the products together with the RT-PCR from normal and SMA brain RNA has identified several candidate polymorphisms. To date, the most interesting lead is an intronic polymorphism possibly affecting exon splicing in a homozygous SMA patient. An updated mutation search will be reported.

  3. Analysis of delta-globin gene alleles in the Sicilian population: identification of five new mutations.

    PubMed

    Giambona, Antonino; Passarello, Cristina; Ruggeri, Gaetano; Renda, Disma; Teresi, Pietro; Anzà, Maurizio; Maggio, Aurelio

    2006-12-01

    Although delta-globin gene (HBD MIM#142000) mutations have no clinical implications, co-inheritance of beta- and delta-thalassemia may lead to misdiagnosis. Among 7,153 samples studied for beta-thalassemia, 205 samples with lower than expected HbA2 levels were selected for our analysis and 183 samples (2.5%) were positive for delta-globin gene mutations. Twelve different mutations were detected, and among these five have not been not previously described (HbA2-Catania HBD c.8A-->T, HbA2-Corleone HBD c.41C-->A, HbA2-Ventimiglia HBD c.212C-->G, HbA2-Montechiaro HBD c.260C-->A, and HbA2-Bagheria HBD c.422C-->T). This study suggests that delta-globin gene defects are very common in Sicily. Thus, these mutations need to be considered during beta-thalassemia screening to avoid false negative results in the detection of at-risk couples.

  4. Detection of Favorable QTL Alleles and Candidate Genes for Lint Percentage by GWAS in Chinese Upland Cotton.

    PubMed

    Su, Junji; Fan, Shuli; Li, Libei; Wei, Hengling; Wang, Caixiang; Wang, Hantao; Song, Meizhen; Zhang, Chi; Gu, Lijiao; Zhao, Shuqi; Mao, Guangzhi; Wang, Chengshe; Pang, Chaoyou; Yu, Shuxun

    2016-01-01

    Improving cotton yield is a major breeding goal for Chinese upland cotton. Lint percentage is an important yield component and a critical economic index for cotton cultivars, and raising the lint percentage has a close relationship to improving cotton lint yield. To investigate the genetic architecture of lint percentage, a diversity panel consisting of 355 upland cotton accessions was grown, and the lint percentage was measured in four different environments. Genotyping was performed with specific-locus amplified fragment sequencing (SLAF-seq). Twelve single-nucleotide polymorphisms (SNPs) associated with lint percentage were detected via a genome-wide association study (GWAS), in which five SNP loci distributed on chromosomes At3 (A02) and At4 (A08) and contained two major-effect QTLs, which were detected in the best linear unbiased predictions (BLUPs) and in more than three environments simultaneously. Furthermore, favorable haplotypes (FHs) of two major-effect QTLs and 47 putative candidate genes in the two linkage disequilibrium (LD) blocks of these associated loci were identified. The expression levels of these putative candidate genes were estimated using RNA-seq data from ten upland cotton tissues. We found that Gh_A02G1268 was very highly expressed during the early fiber development stage, whereas the gene was poorly expressed in the seed. These results implied that Gh_A02G1268 may determine the lint percentage by regulating seed and fiber development. The favorable QTL alleles and candidate genes for lint percentage identified in this study will have high potential for improving lint yield in future Chinese cotton breeding programs.

  5. Detection of Favorable QTL Alleles and Candidate Genes for Lint Percentage by GWAS in Chinese Upland Cotton

    PubMed Central

    Su, Junji; Fan, Shuli; Li, Libei; Wei, Hengling; Wang, Caixiang; Wang, Hantao; Song, Meizhen; Zhang, Chi; Gu, Lijiao; Zhao, Shuqi; Mao, Guangzhi; Wang, Chengshe; Pang, Chaoyou; Yu, Shuxun

    2016-01-01

    Improving cotton yield is a major breeding goal for Chinese upland cotton. Lint percentage is an important yield component and a critical economic index for cotton cultivars, and raising the lint percentage has a close relationship to improving cotton lint yield. To investigate the genetic architecture of lint percentage, a diversity panel consisting of 355 upland cotton accessions was grown, and the lint percentage was measured in four different environments. Genotyping was performed with specific-locus amplified fragment sequencing (SLAF-seq). Twelve single-nucleotide polymorphisms (SNPs) associated with lint percentage were detected via a genome-wide association study (GWAS), in which five SNP loci distributed on chromosomes At3 (A02) and At4 (A08) and contained two major-effect QTLs, which were detected in the best linear unbiased predictions (BLUPs) and in more than three environments simultaneously. Furthermore, favorable haplotypes (FHs) of two major-effect QTLs and 47 putative candidate genes in the two linkage disequilibrium (LD) blocks of these associated loci were identified. The expression levels of these putative candidate genes were estimated using RNA-seq data from ten upland cotton tissues. We found that Gh_A02G1268 was very highly expressed during the early fiber development stage, whereas the gene was poorly expressed in the seed. These results implied that Gh_A02G1268 may determine the lint percentage by regulating seed and fiber development. The favorable QTL alleles and candidate genes for lint percentage identified in this study will have high potential for improving lint yield in future Chinese cotton breeding programs. PMID:27818672

  6. Endochondral ossification pathway genes and postmenopausal osteoporosis: Association and specific allele related serum bone sialoprotein levels in Han Chinese.

    PubMed

    Zhang, Yunzhi; Liu, Haiyan; Zhang, Chen; Zhang, Tianxiao; Zhang, Bo; Li, Lu; Chen, Gang; Fu, Dongke; Wang, KunZheng

    2015-11-16

    Osteoporosis is a systemic skeletal disorder characterized by reduced bone mineral density (BMD) and disrupted bone architecture, predisposing the patient to increased fracture risk. Evidence from early genetic epidemiological studies has indicated a major role for genetics in the development of osteoporosis and the variation in BMD. In this study, we focused on two key genes in the endochondral ossification pathway, IBSP and PTHLH. Over 9,000 postmenopausal Han Chinese women were recruited, and 54 SNPs were genotyped. Two significant SNPs within IBSP, rs1054627 and rs17013181, were associated with BMD and postmenopausal osteoporosis by the two-stage strategy, and rs17013181 was also significantly associated with serum IBSP levels. Moreover, one haplotype (rs12425376-rs10843047-rs42294) covering the 5' end of PTHLH was associated with postmenopausal osteoporosis. Our results provide evidence for the association of these two key endochondral ossification pathway genes with BMD and osteoporosis in postmenopausal Han Chinese women. Combined with previous findings, we provide evidence that a particular SNP in IBSP has an allele-specific effect on mRNA levels, which would, in turn, reflect serum IBSP levels.

  7. Induction of antimicrobial activities in heterologous streptomycetes using alleles of the Streptomyces coelicolor gene absA1.

    PubMed

    McKenzie, Nancy L; Thaker, Maulik; Koteva, Kalinka; Hughes, Donald W; Wright, Gerard D; Nodwell, Justin R

    2010-04-01

    The bacterial genus Streptomyces is endowed with a remarkable secondary metabolism that generates an enormous number of bioactive small molecules. Many of these genetically encoded small molecules are used as antibiotics, anticancer agents and as other clinically relevant therapeutics. The rise of resistant pathogens has led to calls for renewed efforts to identify antimicrobial activities, including expanded screening of streptomycetes. Indeed, it is known that most strains encode >20 secondary metabolites and that many, perhaps most of these, have not been considered for their possible therapeutic use. One roadblock is that many strains do not express their secondary metabolic gene clusters efficiently under laboratory conditions. As one approach to this problem, we have used alleles of a pleiotropic regulator of secondary metabolism from Streptomyces coelicolor to activate secondary biosynthetic gene clusters in heterologous streptomycetes. In one case, we demonstrate the activation of pulvomycin production in S. flavopersicus, a metabolite not previously attributed to this species. We find that the absA1-engineered strains produced sufficient material for purification and characterization. As a result, we identified new, broad-spectrum antimicrobial activities for pulvomycin, including a potent antimicrobial activity against highly antibiotic-resistant Gram-negative and Gram-positive pathogens.

  8. Wound Induced Tanscriptional Regulation of Benzylisoquinoline Pathway and Characterization of Wound Inducible PsWRKY Transcription Factor from Papaver somniferum

    PubMed Central

    Singh, Seema; Phukan, Ujjal J.; Gupta, M. M.; Shanker, Karuna; Shukla, Rakesh Kumar

    2013-01-01

    Wounding is required to be made in the walls of the green seed pod of Opium poppy prior exudation of latex. To withstand this kind of trauma plants regulate expression of some metabolites through an induced transcript level. 167 unique wound-inducible ESTs were identified by a repetitive round of cDNA subtraction after 5 hours of wounding in Papaver somniferum seedlings. Further repetitive reverse northern analysis of these ESTs revealed 80 transcripts showing more than two fold induction, validated through semi-quantitative RT-PCR & real time expression analysis. One of the major classified categories among identified ESTs belonged to benzylisoquinoline transcripts. Tissue specific metabolite analysis of benzylisoquinoline alkaloids (BIAs) in response to wounding revealed increased accumulation of narcotine and papaverine. Promoter analysis of seven transcripts of BIAs pathway showed the presence of W-box cis-element with the consensus sequence of TGAC, which is the proposed binding site for WRKY type transcription factors. One of the Wound inducible ‘WRKY’ EST isolated from our subtracted library was made full-length and named as ‘PsWRKY’. Bacterially expressed PsWRKY interacted with the W-box element having consensus sequence TTGACT/C present in the promoter region of BIAs biosynthetic pathway genes. PsWRKY further activated the TYDC promoter in yeast and transiently in tobacco BY2 cells. Preferential expression of PsWRKY in straw and capsule and its interaction with consensus W-box element present in BIAs pathway gene transcripts suggest its possible involvement in the wound induced regulation of BIAs pathway. PMID:23382823

  9. Identification and Characterization of Multiple Intermediate Alleles of the Key Genes Regulating Brassinosteroid Biosynthesis Pathways

    PubMed Central

    Du, Junbo; Zhao, Baolin; Sun, Xin; Sun, Mengyuan; Zhang, Dongzhi; Zhang, Shasha; Yang, Wenyu

    2017-01-01

    Most of the early identified brassinosteroid signaling and biosynthetic mutants are null mutants, exhibiting extremely dwarfed phenotypes and male sterility. These null mutants are usually unable to be directly transformed via a routinely used Agrobacterium-mediated gene transformation system and therefore are less useful for genetic characterization of the brassinosteroid (BR)-related pathways. Identification of intermediate signaling mutants such as bri1–5 and bri1–9 has contributed drastically to the elucidation of BR signaling pathway using both genetic and biochemical approaches. However, intermediate mutants of key genes regulating BR biosynthesis have seldom been reported. Here we report identification of several intermediate BR biosynthesis mutants mainly resulted from leaky transcriptions due to the insertions of T-DNAs in the introns. These mutants are semi-dwarfed and fertile and capable to be transformed. These intermediate mutants could be useful tools for future discovery and analyses of novel components regulating BR biosynthesis and catabolism via genetic modifier screen. PMID:28138331

  10. Distribution of Porphyromonas gingivalis biotypes defined by alleles of the kgp (Lys-gingipain) gene.

    PubMed

    Nadkarni, Mangala A; Nguyen, Ky-Anh; Chapple, Cheryl C; DeCarlo, Arthur A; Jacques, Nicholas A; Hunter, Neil

    2004-08-01

    Paired subgingival plaque samples representing the most-diseased and least-diseased sites were collected from 34 adult patients with diagnosed chronic periodontitis. The percentage of Porphyromonas gingivalis relative to the total anaerobic and gram-negative bacterial load at each site was determined by real-time PCR. Based on variations in the noncatalytic C terminus of the Lys-gingipain (Kgp), it was reasoned that DNA sequence variation in the 3'-coding region of the kgp gene might determine functional biotypes. Perusal of the available sequence information in GenBank indicated three such forms of the kgp gene corresponding to P. gingivalis strains HG66, 381, and W83. Analysis of patient samples revealed the presence of a fourth genotype (W83v) that showed duplication of a sequence recognized by the W83 reverse primer. The four biotypes, HG66, 381, W83, and W83v, were present in the study group in the ratio 8:11:6:5, respectively. Each subject was colonized by one predominant biotype, and only three patients were colonized by a trace amount of a second biotype.

  11. A genetic polymorphism in coumarin 7-hydroxylation: Sequence of the human CYP2A genes and identification of variant CYP2A6 alleles

    SciTech Connect

    Fernandez-Salguero, P.; Hoffman, S.M.G.; Mohrenweiser, H.

    1995-09-01

    A group of human cytochrome P450 genes encompassing the CYP2A, CYP2B, and CYP2F subfamilies were cloned and assembled into a 350-kb contig localized on the long arm of chromosome 19. Three complete CYP2A genes - CYP2A6, CYP2A7, and CYP2A13 - plus two pseudogenes truncated after exon 5 were identified and sequenced. A variant CYP2A6 allele that differed from the corresponding CYP2A6 and CYP2A7 cDNAs previously sequenced was found and was designated CYP2A6{nu}2. Sequence differences in the CY-P2A6{nu}2 gene are restricted to regions encompassing exons 3, 6, and 8, which bear sequence relatedness with the corresponding exons of the CYP2A7 gene, located downstream and centromeric of CYP2A6{nu}2, suggesting recent gene-conversion events. The sequencing of all the CYP2A genes allowed the design of a PCR diagnostic test for the normal CYP2A6 allele, the CYP2A6{nu}2 allele, and a variant - designated CYP2A6{nu}1 - that encodes an enzyme with a single inactivating amino acid change. These variant alleles were found in individuals who were deficient in their ability to metabolize the CYP2A6 probe drug coumarin. The allelic frequencies of CYP2A6{nu}1 and CYP2A6{nu}2 differed significantly between Caucasian, Asian, and African-American populations. These studies establish the existence of a new cytochrome P450 genetic polymorphism. 30 refs., 4 figs., 2 tabs.

  12. Radiosensitivity of Human Fibroblasts is Associated With Amino Acid Substitution Variants in Susceptible Genes And Correlates With The Number of Risk Alleles

    SciTech Connect

    Alsbeih, Ghazi . E-mail: galsbeih@kfshrc.edu.sa; El-Sebaie, Medhat; Al-Harbi, Najla; Al-Buhairi, Muneera; Al-Hadyan, Khaled; Al-Rajhi, Nasser

    2007-05-01

    Purpose: Genetic predictive markers of radiosensitivity are being sought for stratifying radiotherapy for cancer patients and risk assessment of radiation exposure. We hypothesized that single nucleotide polymorphisms in susceptible genes are associated with, and the number of risk alleles has incremental effect on, individual radiosensitivity. Methods and Materials: Six amino acid substitution variants (ATM 1853 Asp/Asn G>A, p53 72 Arg/Pro G>C, p21 31 Ser/Arg C>A, XRCC1 399 Arg/Gln G>A, XRCC3 241 Thr/Met C>T, and TGF{beta}1 10 Leu/Pro T>C) were genotyped by direct sequencing in 54 fibroblast strains of different radiosensitivity. Results: The clonogenic survival fraction at 2 Gy range was 0.15-0.50 (mean, 0.34, standard deviation, 0.08). The mean survival fraction at 2 Gy divided the cell strains into radiosensitive (26 cases) and normal (28 controls). A significant association was observed between the survival fraction at 2 Gy and ATM 1853 Asn, XRCC3 241 Met, and TGF{beta}1 10 Leu alleles (p = 0.05, p = 0.02, and p = 0.02, respectively). The p53 72 Arg allele showed a borderline association (p = 0.07). The number of risk alleles increased with increasing radiosensitivity, and the group comparison showed a statistically significant difference between the radiosensitive and control groups (p {<=}0.001). Conclusion: The results of our study have shown that single nucleotide polymorphisms in susceptible genes influence cellular radiation response and that the number of risk alleles has a combined effect on radiosensitivity. Individuals with multiple risk alleles could be more susceptible to radiation effects than those with fewer risk alleles. These results may have implications in predicting normal tissue reactions to radiotherapy and risk assessment of radiation exposure.

  13. The distribution of repressive histone modifications on silenced FMR1 alleles provides clues to the mechanism of gene silencing in fragile X syndrome.

    PubMed

    Kumari, Daman; Usdin, Karen

    2010-12-01

    Fragile X syndrome (FXS) is the most common heritable cause of intellectual disability and the most common known cause of autism. Most cases of FXS result from the expansion of a CGG·CCG repeat in the 5' UTR of the FMR1 gene that leads to gene silencing. It has previously been shown that silenced alleles are associated with histone H3 dimethylated at lysine 9 (H3K9Me2) and H3 trimethylated at lysine 27 (H3K27Me3), modified histones typical of developmentally repressed genes. We show here that these alleles are also associated with elevated levels of histone H3 trimethylated at lysine 9 (H3K9Me3) and histone H4 trimethylated at lysine 20 (H4K20Me3). All four of these modified histones are present on exon 1 of silenced alleles at levels comparable to that seen on pericentric heterochromatin. The two groups of histone modifications show a different distribution on fragile X alleles: H3K9Me2 and H3K27Me3 have a broad distribution, whereas H3K9Me3 and H4K20Me3 have a more focal distribution with the highest level of these marks being present in the vicinity of the repeat. This suggests that the trigger for gene silencing may be local to the repeat itself and perhaps involves a mechanism similar to that involved in the formation of pericentric heterochromatin.

  14. The Dopamine Receptor D4 7-Repeat Allele and Prenatal Smoking in ADHD-Affected Children and Their Unaffected Siblings: No Gene-Environment Interaction

    ERIC Educational Resources Information Center

    Altink, Marieke E.; Arias-Vasquez, Alejandro; Franke, Barbara; Slaats-Willemse, Dorine I. E.; Buschgens, Cathelijne J. M.; Rommelse, Nanda N. J.; Fliers, Ellen A.; Anney, Richard; Brookes, Keeley-Joanne; Chen, Wai; Gill, Michael; Mulligan, Aisling; Sonuga-Barke, Edmund; Thompson, Margaret; Sergeant, Joseph A.; Faraone, Stephen V.; Asherson, Philip; Buitelaar, Jan K.

    2008-01-01

    Background: The dopamine receptor D4 ("DRD4") 7-repeat allele and maternal smoking during pregnancy are both considered as risk factors in the aetiology of attention deficit hyperactivity disorder (ADHD), but few studies have been conducted on their interactive effects in causing ADHD. The purpose of this study is to examine the gene by…

  15. Genealogical analyses of rabies virus strains from Brazil based on N gene alleles.

    PubMed Central

    Heinemann, M. B.; Fernandes-Matioli, F. M. C.; Cortez, A.; Soares, R. M.; Sakamoto, S. M.; Bernardi, F.; Ito, F. H.; Madeira, A. M. B. N.; Richtzenhain, L. J.

    2002-01-01

    Thirty rabies virus isolates from cows and vampire bats from different regions of São Paulo State, Southeastern Brazil and three rabies vaccines were studied genetically. The analysis was based on direct sequencing of PCR-amplified products of 600 nucleotides coding for the amino terminus of nucleoprotein gene. The sequences were checked to verify their genealogical and evolutionary relationships and possible implication for health programmes. Statistical data indicated that there were no significant genetic differences between samples isolated from distinct hosts, from different geographical regions and between samples collected in the last two decades. According to the HKA test, the variability observed in the sequences is probably due to genetic drift. Since changes in genetic material may produce modifications in the protein responsible for immunogenicity of virus, which may eventually cause vaccine failure in herds, we suggest that continuous efforts in monitoring genetic diversity in rabies virus field strains, in relation to vaccine strains, must be conducted. PMID:12113496

  16. Association between allelic variation due to short tandem repeats in tRNA gene of Entamoeba histolytica and clinical phenotypes of amoebiasis.

    PubMed

    Jaiswal, Virendra; Ghoshal, Ujjala; Mittal, Balraj; Dhole, Tapan N; Ghoshal, Uday C

    2014-05-01

    Genotypes of Entamoeba histolytica (E. histolytica) may contribute clinical phenotypes of amoebiasis such as amoebic liver abscess (ALA), dysentery and asymptomatic cyst passers state. Hence, we evaluated allelic variation due to short tandem repeats (STRs) in tRNA gene of E. histolytica and clinical phenotypes of amoebiasis. Asymptomatic cyst passers (n=24), patients with dysentery (n=56) and ALA (n=107) were included. Extracted DNA from stool (dysentery, asymptomatic cyst passers) and liver aspirate was amplified using 6 E. histolytica specific tRNA-linked STRs (D-A, A-L, N-K2, R-R, S-Q, and S(TGA)-D) primers. PCR products were subjected to sequencing. Association between allelic variation and clinical phenotypes was analyzed. A total of 9 allelic variations were found in D-A, 8 in A-L, 4 in N-K2, 5 in R-R, 10 in S(TAG)-D and 7 in S-Q loci. A significant association was found between allelic variants and clinical phenotypes of amoebiasis. This study reveals that allelic variation due to short tandem repeats (STRs) in tRNA gene of E. histolytica is associated different clinical outcome of amoebiasis.

  17. Mucopolysaccharidosis VI (Maroteaux-Lamy Syndrome): Six unique arylsulfatase B gene alleles causing variable disease phenotypes

    SciTech Connect

    Isbrandt, D.; Arlt, G.; Figura, K. von; Peters, C.; Brooks, D.A.; Hopwood, J.J.

    1994-03-01

    Mucopolysaccharidosis type VI, or Maroteaux-Lamy syndrome, is a lysosomal storage disorder caused by a deficiency of the enzyme arylsulfatase B (ASB), also known as N-acetylgalactosamine-4-sulfatase. Multiple clinical phenotypes of this autosomal recessively inherited disease have been described. Recent isolation and characterization of the human ASB gene facilitated the analysis of molecular defects underlying the different phenotypes. Conditions for PCR amplification of the entire open reading frame from genomic DNA and for subsequent direct automated DNA sequencing of the resulting DNA fragments were established. Besides two polymorphisms described elsewhere that cause methionine-for-valine substitutions in the arylsulfatase B gene, six new mutations in six patients were detected: four point mutations resulting in amino acid substitutions, a 1-bp deletion, and a 1-bp insertion. The point mutations were two G-to-A and two T-to-C transitions. The G-to-A transitions cause an arginine-for-glycine substitution at residue 144 in a homoallelic patient with a severe disease phenotype and a tyrosine-for-cysteine substitution at residue 521 in a potentially heteroallelic patient with the severe form of the disease. The T-to-C transitions cause an arginine-for-cysteine substitution at amino acid residue 192 in a homoallelic patient with mild symptoms and a proline-for-leucine substitution at amino acid 321 in a homoallelic patient with the intermediate form. The insertion between nucleotides T1284 and G1285 resulted in a loss of the 100 C-terminal amino acids of the wild-type protein and in the deletion of nucleotide C1577 in a 39-amino-acid C-terminal extension of the ASB polypeptide. Both mutations were detected in homoallelic patients with the severe form of the disease. Expression of mutant cDNAs encoding the four amino acid substitutions and the deletion resulted in reduction of both ASB protein levels and arylsulfatase enzyme activity. 25 refs., 4 figs.

  18. Population genetic evidence for rapid changes in intraspecific diversity and allelic cycling of a specialist defense gene in Zea.

    PubMed

    Tiffin, Peter; Hacker, Robert; Gaut, Brandon S

    2004-09-01

    Two patterns of plant defense gene evolution are emerging from molecular population genetic surveys. One is that specialist defenses experience stronger selection than generalist defenses. The second is that specialist defenses are more likely to be subject to balancing selection, i.e., evolve in a manner consistent with balanced-polymorphism or trench-warfare models of host-parasite coevolution. Because most of the data of specialist defenses come from Arabidopsis thaliana, we examined the genetic diversity and evolutionary history of three defense genes in two outcrossing species, the autotetraploid Zea perennis and its most closely related extant relative the diploid Z. diploperennis. Intraspecific diversity at two generalist defenses, the protease inhibitors wip1 and mpi, were consistent with a neutral model. Like previously studied genes in these taxa, wip1 and mpi harbored similar levels of diversity in Z. diploperennis and Z. perennis. In contrast, the specialist defense hm2 showed strong although distinctly different departures from a neutral model in the two species. Z. diploperennis appears to have experienced a strong and recent selective sweep. Using a rejection-sampling coalescent method, we estimate the strength of selection on Z. diploperennis hm2 to be approximately 3.0%, which is approximately equal to the strength of selection on tb1 during maize domestication. Z. perennis hm2 harbors three highly diverged alleles, two of which are found at high frequency. The distinctly different patterns of diversity may be due to differences in the phase of host-parasite coevolutionary cycles, although higher hm2 diversity in Z. perennis may also reflect reduced efficacy of selection in the autotetraploid relative to its diploid relative.

  19. Characterization of three active transposable elements recently inserted in three independent DFR-A alleles and one high-copy DNA transposon isolated from the Pink allele of the ANS gene in onion (Allium cepa L.).

    PubMed

    Kim, Sunggil; Park, Jee Young; Yang, Tae-Jin

    2015-06-01

    Intact retrotransposon and DNA transposons inserted in a single gene were characterized in onions (Allium cepa) and their transcription and copy numbers were estimated in this study. While analyzing diverse onion germplasm, large insertions in the DFR-A gene encoding dihydroflavonol 4-reductase (DFR) involved in the anthocyanin biosynthesis pathway were found in two accessions. A 5,070-bp long terminal repeat (LTR) retrotransposon inserted in the active DFR-A (R4) allele was identified from one of the large insertions and designated AcCOPIA1. An intact ORF encoded typical domains of copia-like LTR retrotransposons. However, AcCOPIA1 contained atypical 'TG' and 'TA' dinucleotides at the ends of the LTRs. A 4,615-bp DNA transposon was identified in the other large insertion. This DNA transposon, designated AcCACTA1, contained an ORF coding for a transposase showing homology with the CACTA superfamily transposable elements (TEs). Another 5,073-bp DNA transposon was identified from the DFR-A (TRN) allele. This DNA transposon, designated AchAT1, belonged to the hAT superfamily with short 4-bp terminal inverted repeats (TIRs). Finally, a 6,258-bp non-autonomous DNA transposon, designated AcPINK, was identified in the ANS-p allele encoding anthocyanidin synthase, the next downstream enzyme to DFR in the anthocyanin biosynthesis pathway. AcPINK also possessed very short 3-bp TIRs. Active transcription of AcCOPIA1, AcCACTA1, and AchAT1 was observed through RNA-Seq analysis and RT-PCR. The copy numbers of AcPINK estimated by mapping the genomic DNA reads produced by NextSeq 500 were predominantly high compared with the other TEs. A series of evidence indicated that these TEs might have transposed in these onion genes very recently, providing a stepping stone for elucidation of enormously large-sized onion genome structure.

  20. Both gene amplification and allelic loss occur at 14q13.3 in lung cancer

    PubMed Central

    Harris, Thomas; Pan, Qiulu; Sironi, Juan; Lutz, Dionne; Tian, Jianmin; Sapkar, Jana; Perez-Soler, Roman; Keller, Steven; Locker, Joseph

    2010-01-01

    Purpose Because loss of Nkx2-8 increases lung cancer in the mouse, we studied suppressive mechanisms in human lung cancer. Experimental Design NKX2-8 is located within 14q13.3, adjacent to its close relative TTF1/NKX2-1. We first analyzed loss of heterozygosity (LOH) of 14q13.3 in 45 matched human lung cancer and control specimens. DNA from tumors with LOH was then analyzed with high-density SNP arrays. For correlation with this genetic analysis, we quantified expression of Nkx2-8 and TTF1 mRNA in tumors. Finally, suppressive function of Nkx2-8 was assessed via colony formation assays in 5 lung cancer cell lines. Results 13/45 (29%) tumors had LOH. In 6 tumors, most adenocarcinomas, LOH was caused by gene amplification. The 0.8 Mb common region of amplification included MBIP, SFTA, TTF1, NKX2-8, and PAX9. In 4 squamous or adenosquamous cancers, LOH was caused by deletion. In 3 other tumors, LOH resulted from whole chromosome mechanisms (14−, 14+, or aneuploidy). The 1.2 Mb common region of deletion included MBIP, SFTA, TTF1, NKX2-8, PAX9, SLC25A21, and MIPOL1. Most tumors had low expression of Nkx2-8. Nevertheless, sequencing did not show NKX2-8 mutations that could explain the low expression. TTF1 overexpression, in contrast, was common and usually independent of Nkx2-8 expression. Finally, stable transfection of Nkx2-8 selectively inhibited growth of H522 lung cancer cells. Conclusions 14q13.3, which contains NKX2-8, is subject to both amplification and deletion in lung cancer. Most tumors have low expression of NKX2-8, and its expression can inhibit growth of some lung cancer cells. PMID:21148747

  1. Simultaneous genotyping of single-nucleotide polymorphisms in alcoholism-related genes using duplex and triplex allele-specific PCR with two-step thermal cycles.

    PubMed

    Shirasu, Naoto; Kuroki, Masahide

    2014-01-01

    We developed a time- and cost-effective multiplex allele-specific polymerase chain reaction (AS-PCR) method based on the two-step PCR thermal cycles for genotyping single-nucleotide polymorphisms in three alcoholism-related genes: alcohol dehydrogenase 1B, aldehyde dehydrogenase 2 and μ-opioid receptor. Applying MightyAmp(®) DNA polymerase with optimized AS-primers and PCR conditions enabled us to achieve effective and selective amplification of the target alleles from alkaline lysates of a human hair root, and simultaneously to determine the genotypes within less than 1.5 h using minimal lab equipment.

  2. Identification of Expanded Alleles of the "FMR1" Gene in the CHildhood Autism Risks from Genes and Environment (CHARGE) Study

    ERIC Educational Resources Information Center

    Tassone, Flora; Choudhary, Nimrah S.; Tassone, Federica; Durbin-Johnson, Blythe; Hansen, Robin; Hertz-Picciotto, Irva; Pessah, Isaac

    2013-01-01

    Fragile X syndrome (FXS) is a neuro-developmental disorder characterized by intellectual disabilities and autism spectrum disorders (ASD). Expansion of a CGG trinucleotide repeat (greater than 200 repeats) in the 5'UTR of the fragile X mental retardation gene, is the single most prevalent cause of cognitive disabilities. Several screening studies…

  3. [Cloning and analyzing of rice blast resistance gene Pi-ta+ allele from Jinghong erect type of common wild rice (Oryza rufipogon Griff) in Yunnan].

    PubMed

    Geng, Xian-Sheng; Yang, Ming-Zhi; Huang, Xing-Qi; Cheng, Zai-Quan; Fu, Jian; Sun, Tao; Li, Jun

    2008-01-01

    A 4,672 bp DNA sequence including the whole coding region and partial non-coding region of rice blast resistance gene Pi-ta+ has been cloned from Jinghong erect type of common wild rice (Oryza rufipogon Griff) in Yunnan by polymerase chain reaction method. The coding region shares 99.86% and 98.78% identity with the corresponding regions of the reported cultivated rice Yashiro-mochi and Yuanjiang type of common wild rice respectively. There are 4 nucleotides difference in the coding region and 6 in intron of the cloned Pi-ta+ gene,compared with Pi-ta from Yashiro-mochi. Pi-ta+ gene in Jinghong erect type of common wild rice has been proved to be a rare existing Pi-ta+ allele, because there was a alanine rather than a serine at the position 918 within the predicted amino acid sequence of PITA. Pi-ta+ allele can cause disease resistance response to rice blast pathogens in plant cells. Differences in DNA sequence, deduced amino acid sequence and antibacterial spectrum may make the Pi-ta+ allele new resistant characteristics. Finding and cloning of Pi-ta+ allele from Jinghong erect type of common wild rice in Yunnan provides a basement for further utilization of the wild rice resources.

  4. Short communication: duplication in the 5'-flanking region of the beta-lactoglobulin gene is linked to the BLG A allele.

    PubMed

    Braunschweig, M H

    2007-12-01

    beta-Lactoglobulin (beta-LG) is the major whey protein in the milk of cows and other ruminants. It is well established that the predominant genetic variants beta-LG A and B are differentially expressed. Extensive investigation of the genetic variation in the promoter region of the BLG gene revealed the existence of specific haplotypes associated with the A and B variants. However, the genetic basis for the differentially expressed BLG A and B alleles is still elusive. In this study additional genetic variation further upstream in the 5'-flanking region of the BLG gene was identified, including 6 single nucleotide substitutions, a single nucleotide deletion, and a 7-bp duplication. Comparison of DNA sequences showed that the investigated 5'-flanking region is highly conserved between ruminants, and the duplication g.-1885_-1879dupCTCTCGC and the substitution g.-1888A>G are only found in the BLG A and D alleles in cattle. The cytosine at position g.-1957 and the thymines at positions g.-2008 and g.-2049 are only found in BLG B alleles of cattle. It is suggested that the described genetic variability contributes to the differential allelic expression of the BLG gene.

  5. Authentication of official Da-huang by sequencing and multiplex allele-specific PCR of a short maturase K gene.

    PubMed

    Xu, Guojie; Wang, Xueyong; Liu, Chunsheng; Li, Weidong; Wei, Shengli; Liu, Ying; Cheng, Xiaoli; Liu, Juan

    2013-02-01

    Rhubarb (official Da-huang) is an important medicinal herb in Asia. Many adulterants of official Da-huang have been discovered in Chinese markets in recent years, which has resulted in adverse effects in medicinal treatment. Here, novel molecular markers based on a short maturase K (matK) gene were developed for authenticating official Da-huang. This study showed that all the species from official Da-huang were clustered together in one clade in the polygenetic trees based on short matK. Two highly conserved single nucleotide polymorphisms of short matK were mined in the species from official Da-huang. Based on these polymophisms, four improved specific primers of official Da-huang were successfully developed that generated reproducible specific bands. These results suggest that the short matK sequence can be considered as a favorable candidate for distinguishing official Da-huang from its adulterants. The established multiplex allele-specific PCR was determined to be simple and accurate and may serve as a preferable tool for authentication of official Da-huang. In addition, we suggest that short-sized specific bands be developed to authenticate materials used in traditional Chinese medicine.

  6. Allele-specific expression of mutated in colorectal cancer (MCC) gene and alternative susceptibility to colorectal cancer in schizophrenia

    PubMed Central

    Wang, Yang; Cao, Yanfei; Huang, Xiaoye; Yu, Tao; Wei, Zhiyun; McGrath, John; Xu, Fei; Bi, Yan; Li, Xingwang; Yang, Fengping; Li, Weidong; Zou, Xia; Peng, Zhihai; Xiao, Yanzeng; Zhang, Yan; He, Lin; He, Guang

    2016-01-01

    Evidence has indicated that the incidence of colorectal cancer (CRC) among schizophrenia is lower than normal. To explore this potential protective effect, we employed an innovative strategy combining association study with allele-specific expression (ASE) analysis in MCC gene. We first genotyped four polymorphisms within MCC in 312 CRC patients, 270 schizophrenia patients and 270 controls. Using the MassArray technique, we performed ASE measurements in a second sample series consisting of 50 sporadic CRC patients, 50 schizophrenia patients and 52 controls. Rs2227947 showed significant differences between schizophrenia cases and controls, and haplotype analysis reported some significant discrepancies among these three subject groups. ASE values of rs2227948 and rs2227947 presented consistently differences between CRC (or schizophrenia) patients and controls. Of the three groups, highest frequencies of ASE in MCC were concordantly found in CRC group, whereas lowest frequencies of ASE were observed in schizophrenia group. Similar trends were confirmed in both haplotype frequencies and ASE frequencies (i.e. CRC > control > schizophrenia). We provide a first indication that MCC might confer alterative genetic susceptibility to CRC in individuals with schizophrenia promising to shed more light on the relationship between schizophrenia and cancer progression. PMID:27226254

  7. Dynamic subnuclear relocalization of WRKY40, a potential new mechanism of ABA-dependent transcription factor regulation.

    PubMed

    Geilen, Katja; Böhmer, Maik

    2015-01-01

    The phytohormone ABA plays a major role during plant development, e.g. seed maturation and seed germination, and during adaptation to abiotic stresses like stomatal aperture regulation. The three closely related WRKY transcription factors WRKY18, WRKY40 and WRKY60 function in ABA signal transduction. We recently demonstrated that WRKY18 and WRKY40 but not WRKY60 localize to nuclear bodies in A. thaliana mesophyll protoplasts. WRKY40, a negative regulator of ABA-dependent inhibition of seed germination, relocalizes from PNBs to the nucleoplasm in the presence of ABA in a dynamic and phosphorylation-dependent manner. We propose that subnuclear relocalization of WRKY40 might constitute a new regulatory mechanism of ABA-dependent modulation of transcription factor activity.

  8. Structures of and allelic diversity and relationships among the major outer membrane protein (ompA) genes of the four chlamydial species.

    PubMed Central

    Kaltenboeck, B; Kousoulas, K G; Storz, J

    1993-01-01

    DNA sequences coding for 81% of the ompA gene from 24 chlamydial strains, representing all chlamydial species, were determined from DNA amplified by polymerase chain reactions. Chlamydial strains of serovars and strains with similar chromosomal restriction fragment length polymorphism had identical ompA DNA sequences. The ompA sequences were segregated into 23 different ompA alleles and aligned with each other, and phylogenetic relationships among them were inferred by neighbor-joining and maximum parsimony analyses. The neighbor-joining method produced a single phylogram which was rooted at the branch between two major clusters. One cluster included all Chlamydia trachomatis ompA alleles (trachoma group). The second cluster was composed of three major groups of ompA alleles: psittacosis group (alleles MN, 6BC, A22/M, B577, LW508, FEPN, and GPIC), pneumonia group (Chlamydia pneumoniae AR388 with the allele KOALA), and polyarthritis group (ruminant and porcine chlamydial alleles LW613, 66P130, L71, and 1710S with propensity for polyarthritis). These groups were distinguished through specific DNA sequence signatures. Maximum parsimony analysis yielded two equally most parsimonious phylograms with topologies similar to the ompA tree of neighbor joining. Two phylograms constructed from chlamydial genomic DNA distances had topologies identical to that of the ompA phylogram with respect to branching of the chlamydial species. Human serovars of C. trachomatis with essentially identical genomes represented a single taxonomic unit, while they were divergent in the ompA tree. Consistent with the ompA phylogeny, the porcine isolate S45, previously considered to be Chlamydia psittaci, was identified as C. trachomatis through biochemical characteristics. These data demonstrate that chlamydial ompA allelic relationships, except for human serovars of C. trachomatis, are cognate with chromosomal phylogenies. Images PMID:8419295

  9. The Variability of Sesquiterpenes Emitted from Two Zea mays Cultivars Is Controlled by Allelic Variation of Two Terpene Synthase Genes Encoding Stereoselective Multiple Product Enzymes

    PubMed Central

    Köllner, Tobias G.; Schnee, Christiane; Gershenzon, Jonathan; Degenhardt, Jörg

    2004-01-01

    The mature leaves and husks of Zea mays release a complex blend of terpene volatiles after anthesis consisting predominantly of bisabolane-, sesquithujane-, and bergamotane-type sesquiterpenes. The varieties B73 and Delprim release the same volatile constituents but in significantly different proportions. To study the molecular genetic and biochemical mechanisms controlling terpene diversity and distribution in these varieties, we isolated the closely related terpene synthase genes terpene synthase4 (tps4) and tps5 from both varieties. The encoded enzymes, TPS4 and TPS5, each formed the same complex mixture of sesquiterpenes from the precursor farnesyl diphosphate but with different proportions of products. These mixtures correspond to the sesquiterpene blends observed in the varieties B73 and Delprim, respectively. The differences in the stereoselectivity of TPS4 and TPS5 are determined by four amino acid substitutions with the most important being a Gly instead of an Ala residue at position 409 at the catalytic site of the enzyme. Although both varieties contain tps4 and tps5 alleles, their differences in terpene composition result from the fact that B73 has only a single functional allele of tps4 and no functional alleles of tps5, whereas Delprim has only a functional allele of tps5 and no functional alleles of tps4. Lack of functionality was shown to be attributable to frame-shift mutations or amino acid substitutions that greatly reduce the activity of their encoded proteins. Therefore, the diversity of sesquiterpenes in these two maize cultivars is strongly influenced by single nucleotide changes in the alleles of two terpene synthase genes. PMID:15075399

  10. A mutant allele of the SUP45 (SAL4) gene of Saccharomyces cerevisiae shows temperature-dependent allosuppressor and omnipotent suppressor phenotypes.

    PubMed

    Stansfield, I; Akhmaloka; Tuite, M F

    1995-04-01

    Using a plasmid-based termination-read-through assay, the sal4-2 conditional-lethal (temperature-sensitive) allele of the SUP45 (SAL4) gene was shown to enhance the efficiency of the weak ochre suppressor tRNA SUQ5 some 10-fold at 30 degrees C. Additionally, this allele increased the suppressor efficiency of SRM2-2, a weak tRNA(Gln) ochre suppressor, indicating that the allosuppressor phenotype is not SUQ5-specific. A sup+ sal4-2 strain also showed a temperature-dependent omnipotent suppressor phenotype, enhancing readthrough of all three termination codons. Combining the sal4-2 allele with an efficient tRNA nonsense suppressor (SUP4) increased the temperature-sensitivity of that strain, indicating that enhanced nonsense suppressor levels contribute to the conditional-lethality conferred by the sal4-2 allele. However, UGA suppression levels in a sup+ sal4-2 strain following a shift to the non-permissive temperature reached a maximum significantly below that exhibited by a non-temperature sensitive SUP4 suppressor strain. Enhanced nonsense suppression may not therefore be the primary cause of the conditional-lethality of this allele. These data indicate a role for Sup45p in translation termination, and possibly in an additional, as yet unidentified, cellular process.

  11. About the origin of European spelt ( Triticum spelta L.): allelic differentiation of the HMW Glutenin B1-1 and A1-2 subunit genes.

    PubMed

    Blatter, R H E; Jacomet, S; Schlumbaum, A

    2004-01-01

    To investigate the origin of European spelt ( Triticum spelta L., genome AABBDD) and its relation to bread wheat ( Triticum aestivum L., AABBDD), we analysed an approximately 1-kb sequence, including a part of the promoter and the coding region, of the high-molecular-weight (HMW) glutenin B1-1 and A1-2 subunit genes in 58 accessions of hexa- and tetraploid wheat from different geographical regions. Six Glu-B1-1 and five Glu-A1-2 alleles were identified based on 21 and 19 informative sites, respectively, which suggests a polyphyletic origin of the A- and B-genomes of hexaploid wheat. In both genes, a group of alleles clustered in a distinct, so-called beta subclade. High frequencies of alleles from the Glu-B1-1 and Glu-A1-2 beta subclades differentiated European spelt from Asian spelt and bread wheat. This indicates different origins of European and Asian spelt, and that European spelt does not derive from the hulled progenitors of bread wheat. The conjoint differentiation of alleles of the A- and B-genome in European spelt suggests the introgression of a tetraploid wheat into free-threshing hexaploid wheat as the origin of European spelt.

  12. Frequencies of 32 base pair deletion of the (Delta 32) allele of the CCR5 HIV-1 co-receptor gene in Caucasians: a comparative analysis.

    PubMed

    Lucotte, Gérard

    2002-05-01

    The CCR5 gene encodes for the co-receptor for the major macrophage-tropics strains of human immunodeficiency virus (HIV-1), and a mutant allele of this gene (Delta 32) provide to homozygotes a strong resistance against infection by HIV. The frequency of the Delta 32 allele was investigated in 40 populations of 8842 non-infected subjects coming from Europe, the Middle-East and North Africa. A clear north-south decreasing gradient was evident for Delta 32 frequencies, with a significant correlation coefficient (r=0.83). The main frequency value of Delta 32 for Sweden, Norway, Denmark, Finland and Iceland (0.134) is significantly (chi(2)=63.818, P<0.001) highest than the Delta 32 mean value, indicating that probably the Vikings might have been instrumental in disseminating the Delta 32 allele during the eighth to the tenth centuries during historical times. Possibly variola virus has discriminated the Delta 32 carriers in Europe since the eighth century AD, explaining the high frequency of the Delta 32 allele in Europe today.

  13. A lin-45 raf enhancer screen identifies eor-1, eor-2 and unusual alleles of Ras pathway genes in Caenorhabditis elegans.

    PubMed

    Rocheleau, Christian E; Howard, Robyn M; Goldman, Alissa P; Volk, Mandy L; Girard, Laura J; Sundaram, Meera V

    2002-05-01

    In Caenorhabditis elegans, the Ras/Raf/MEK/ERK signal transduction pathway controls multiple processes including excretory system development, P12 fate specification, and vulval cell fate specification. To identify positive regulators of Ras signaling, we conducted a genetic screen for mutations that enhance the excretory system and egg-laying defects of hypomorphic lin-45 raf mutants. This screen identified unusual alleles of several known Ras pathway genes, including a mutation removing the second SH3 domain of the sem-5/Grb2 adaptor, a temperature-sensitive mutation in the helical hairpin of let-341/Sos, a gain-of-function mutation affecting a potential phosphorylation site of the lin-1 Ets domain transcription factor, a dominant-negative allele of ksr-1, and hypomorphic alleles of sur-6/PP2A-B, sur-2/Mediator, and lin-25. In addition, this screen identified multiple alleles of two newly identified genes, eor-1 and eor-2, that play a relatively weak role in vulval fate specification but positively regulate Ras signaling during excretory system development and P12 fate specification. The spectrum of identified mutations argues strongly for the specificity of the enhancer screen and for a close involvement of eor-1 and eor-2 in Ras signaling.

  14. Functional screening of willow alleles in Arabidopsis combined with QTL mapping in willow (Salix) identifies SxMAX4 as a coppicing response gene.

    PubMed

    Salmon, Jemma; Ward, Sally P; Hanley, Steven J; Leyser, Ottoline; Karp, Angela

    2014-05-01

    Willows (Salix spp.) are important biomass crops due to their ability to grow rapidly with low fertilizer inputs and ease of cultivation in short-rotation coppice cycles. They are relatively undomesticated and highly diverse, but functional testing to identify useful allelic variation is time-consuming in trees and transformation is not yet possible in willow. Arabidopsis is heralded as a model plant from which knowledge can be transferred to advance the improvement of less tractable species. Here, knowledge and methodologies from Arabidopsis were successfully used to identify a gene influencing stem number in coppiced willows, a complex trait of key biological and industrial relevance. The strigolactone-related More AXillary growth (MAX) genes were considered candidates due to their role in shoot branching. We previously demonstrated that willow and Arabidopsis show similar response to strigolactone and that transformation rescue of Arabidopsis max mutants with willow genes could be used to detect allelic differences. Here, this approach was used to screen 45 SxMAX1, SxMAX2, SxMAX3 and SxMAX4 alleles cloned from 15 parents of 11 mapping populations varying in shoot-branching traits. Single-nucleotide polymorphism (SNP) frequencies were locus dependent, ranging from 29.2 to 74.3 polymorphic sites per kb. SxMAX alleles were 98%-99% conserved at the amino acid level, but different protein products varying in their ability to rescue Arabidopsis max mutants were identified. One poor rescuing allele, SxMAX4D, segregated in a willow mapping population where its presence was associated with increased shoot resprouting after coppicing and colocated with a QTL for this trait.

  15. A nonsense nucleotide substitution in the oculocutaneous albinism II gene underlies the original pink-eyed dilution allele (Oca2(p)) in mice.

    PubMed

    Shoji, Haruka; Kiniwa, Yukiko; Okuyama, Ryuhei; Yang, Mu; Higuchi, Keiichi; Mori, Masayuki

    2015-01-01

    The original pink-eyed dilution (p) on chromosome 7 is a very old spontaneous mutation in mice. The oculocutaneous albinism II (Oca2) gene has previously been identified as the p gene. Oca2 transcripts have been shown to be absent in the skin of SJL/J mice with the original p mutant allele (Oca2(p)); however, the molecular genetic lesion underlying the original Oca2(p) allele has never been reported. The NCT mouse (commonly known as Nakano cataract mouse) has a pink-eyed dilution phenotype, which prompted us to undertake a molecular genetic analysis of the Oca2 gene of this strain. Our genetic linkage analysis suggests that the locus for the pink-eyed dilution phenotype of NCT is tightly linked to the Oca2 locus. PCR cloning and nucleotide sequence analysis indicates that the NCT mouse has a nonsense nucleotide substitution at exon 7 of the Oca2 gene. Examination of three mouse strains (NZW/NSlc, SJL/J, and 129X1/SvJJmsSlc) with the original Oca2(p) allele revealed the presence of a nonsense nucleotide substitution identical to that in the NCT strain. RT-PCR analysis revealed that the Oca2 transcripts were absent in the skin of NCT mice, suggesting intervention of the nonsense-mediated mRNA decay pathway. Collectively, the data in this study indicate that the nonsense nucleotide substitution in the Oca2 gene underlies the Oca2(p) allele. Our data also indicate that the NCT mouse can be used not only as a cataract model, but also as a model for human type II oculocutaneous albinism.

  16. Linkage disequilibrium between the M470V variant and the IVS8 polyT alleles of the CFTR gene in CBAVD.

    PubMed Central

    de Meeus, A; Guittard, C; Desgeorges, M; Carles, S; Demaille, J; Claustres, M

    1998-01-01

    Congenital bilateral absence of the vas deferens (CBAVD) is a cause of male sterility mostly resulting from mutations in the cystic fibrosis transmembrane regulator (CFTR) gene. The most common defect is the 5T variant at the branch/acceptor site of intron 8, which induces high levels of exon 9 skipping leading to non-functional protein. However, this 5T variant has incomplete penetrance and variable expressivity, suggesting that some other regulatory factors may modulate the splicing of exon 9. To identify such factors, we report here the genetic analysis of a polymorphic locus, M470V, located in exon 10 of the CFTR gene in 60 patients with CBAVD, compared to a normal control population. The statistical analysis showed strong linkage disequilibrium between the 5T allele and the V allele of the M470V polymorphism in the CBAVD population, but not in the normal population. The V allele in a gene carrying 5T could, however, contribute to lowering the level of normal transcripts, as already suggested by in vitro transcriptional studies. These genetic findings, together with previous studies, suggest involvement of the M470V variant in the modulation of the splicing of exon 9 of the CFTR gene. PMID:9678705

  17. Risk alleles of genes with monoallelic expression are enriched in gain-of-function variants and depleted in loss-of-function variants for neurodevelopmental disorders.

    PubMed

    Savova, V; Vinogradova, S; Pruss, D; Gimelbrant, A A; Weiss, L A

    2017-03-07

    Over 3000 human genes can be expressed from a single allele in one cell, and from the other allele-or both-in neighboring cells. Little is known about the consequences of this epigenetic phenomenon, monoallelic expression (MAE). We hypothesized that MAE increases expression variability, with a potential impact on human disease. Here, we use a chromatin signature to infer MAE for genes in lymphoblastoid cell lines and human fetal brain tissue. We confirm that across clones MAE status correlates with expression level, and that in human tissue data sets, MAE genes show increased expression variability. We then compare mono- and biallelic genes at three distinct scales. In the human population, we observe that genes with polymorphisms influencing expression variance are more likely to be MAE (P<1.1 × 10(-6)). At the trans-species level, we find gene expression differences and directional selection between humans and chimpanzees more common among MAE genes (P<0.05). Extending to human disease, we show that MAE genes are under-represented in neurodevelopmental copy number variants (CNVs) (P<2.2 × 10(-10)), suggesting that pathogenic variants acting via expression level are less likely to involve MAE genes. Using neuropsychiatric single-nucleotide polymorphism (SNP) and single-nucleotide variant (SNV) data, we see that genes with pathogenic expression-altering or loss-of-function variants are less likely MAE (P<7.5 × 10(-11)) and genes with only missense or gain-of-function variants are more likely MAE (P<1.4 × 10(-6)). Together, our results suggest that MAE genes tolerate a greater range of expression level than biallelic expression (BAE) genes, and this information may be useful in prediction of pathogenicity.Molecular Psychiatry advance online publication, 7 March 2017; doi:10.1038/mp.2017.13.

  18. Starch phosphorylation in potato tubers is influenced by allelic variation in the genes encoding glucan water dikinase, starch branching enzymes I and II, and starch synthase III.

    PubMed

    Carpenter, Margaret A; Joyce, Nigel I; Genet, Russell A; Cooper, Rebecca D; Murray, Sarah R; Noble, Alasdair D; Butler, Ruth C; Timmerman-Vaughan, Gail M

    2015-01-01

    Starch phosphorylation is an important aspect of plant metabolism due to its role in starch degradation. Moreover, the degree of phosphorylation of starch determines its physicochemical properties and is therefore relevant for industrial uses of starch. Currently, starch is chemically phosphorylated to increase viscosity and paste stability. Potato cultivars with elevated starch phosphorylation would make this process unnecessary, thereby bestowing economic and environmental benefits. Starch phosphorylation is a complex trait which has been previously shown by antisense gene repression to be influenced by a number of genes including those involved in starch synthesis and degradation. We have used an association mapping approach to discover genetic markers associated with the degree of starch phosphorylation. A diverse collection of 193 potato lines was grown in replicated field trials, and the levels of starch phosphorylation at the C6 and C3 positions of the glucosyl residues were determined by mass spectrometry of hydrolyzed starch from tubers. In addition, the potato lines were genotyped by amplicon sequencing and microsatellite analysis, focusing on candidate genes known to be involved in starch synthesis. As potato is an autotetraploid, genotyping included determination of allele dosage. Significant associations (p < 0.001) were found with SNPs in the glucan water dikinase (GWD), starch branching enzyme I (SBEI) and the starch synthase III (SSIII) genes, and with a SSR allele in the SBEII gene. SNPs in the GWD gene were associated with C6 phosphorylation, whereas polymorphisms in the SBEI and SBEII genes were associated with both C6 and C3 phosphorylation and the SNP in the SSIII gene was associated with C3 phosphorylation. These allelic variants have potential as genetic markers for starch phosphorylation in potato.

  19. A commonly carried allele of the obesity-related FTO gene is associated with reduced brain volume in the healthy elderly.

    PubMed

    Ho, April J; Stein, Jason L; Hua, Xue; Lee, Suh; Hibar, Derrek P; Leow, Alex D; Dinov, Ivo D; Toga, Arthur W; Saykin, Andrew J; Shen, Li; Foroud, Tatiana; Pankratz, Nathan; Huentelman, Matthew J; Craig, David W; Gerber, Jill D; Allen, April N; Corneveaux, Jason J; Stephan, Dietrich A; DeCarli, Charles S; DeChairo, Bryan M; Potkin, Steven G; Jack, Clifford R; Weiner, Michael W; Raji, Cyrus A; Lopez, Oscar L; Becker, James T; Carmichael, Owen T; Thompson, Paul M

    2010-05-04

    A recently identified variant within the fat mass and obesity-associated (FTO) gene is carried by 46% of Western Europeans and is associated with an approximately 1.2 kg higher weight, on average, in adults and an approximately 1 cm greater waist circumference. With >1 billion overweight and 300 million obese persons worldwide, it is crucial to understand the implications of carrying this very common allele for the health of our aging population. FTO is highly expressed in the brain and elevated body mass index (BMI) is associated with brain atrophy, but it is unknown how the obesity-associated risk allele affects human brain structure. We therefore generated 3D maps of regional brain volume differences in 206 healthy elderly subjects scanned with MRI and genotyped as part of the Alzheimer's Disease Neuroimaging Initiative. We found a pattern of systematic brain volume deficits in carriers of the obesity-associated risk allele versus noncarriers. Relative to structure volumes in the mean template, FTO risk allele carriers versus noncarriers had an average brain volume difference of approximately 8% in the frontal lobes and 12% in the occipital lobes-these regions also showed significant volume deficits in subjects with higher BMI. These brain differences were not attributable to differences in cholesterol levels, hypertension, or the volume of white matter hyperintensities; which were not detectably higher in FTO risk allele carriers versus noncarriers. These brain maps reveal that a commonly carried susceptibility allele for obesity is associated with structural brain atrophy, with implications for the health of the elderly.

  20. ThWRKY4 from Tamarix hispida Can Form Homodimers and Heterodimers and Is Involved in Abiotic Stress Responses.

    PubMed

    Wang, Liuqiang; Zheng, Lei; Zhang, Chunrui; Wang, Yucheng; Lu, Mengzhu; Gao, Caiqiu

    2015-11-13

    WRKY proteins are a large family of transcription factors that are involved in diverse developmental processes and abiotic stress responses in plants. However, our knowledge of the regulatory mechanisms of WRKYs participation in protein-protein interactions is still fragmentary, and such protein-protein interactions are fundamental in understanding biological networks and the functions of proteins. In this study, we report that a WRKY protein from Tamarix hispida, ThWRKY4, can form both homodimers and heterodimers with ThWRKY2 and ThWRKY3. In addition, ThWRKY2 and ThWRKY3 can both bind to W-box motif with binding affinities similar to that of ThWRKY4. Further, the expression patterns of ThWRKY2 and ThWRKY3 are similar to that of ThWRKY4 when plants are exposed to abscisic acid (ABA). Subcellular localization shows that these three ThWRKY proteins are nuclear proteins. Taken together, these results demonstrate that ThWRKY4 is a dimeric protein that can form functional homodimers or heterodimers that are involved in abiotic stress responses.

  1. Involvement of NtERF3 in the cell death signalling pathway mediated by SIPK/WIPK and WRKY1 in tobacco plants.

    PubMed

    Ogata, T; Okada, H; Kawaide, H; Takahashi, H; Seo, S; Mitsuhara, I; Matsushita, Y

    2015-09-01

    We previously reported that one of the ethylene response factors (ERFs), NtERF3, and other members of the subgroup VIII-a ERFs of the AP2/ERF family exhibit cell death-inducing ability in tobacco leaves. In this study, we focused on the involvement of NtERF3 in a cell death signalling pathway in tobacco plants, particularly downstream of NtSIPK/NtWIPK and NtWRKY1, which are mitogen-activated protein kinases and a phosphorylation substrate of NtSIPK, respectively. An ERF-associated amphiphilic repression (EAR) motif-deficient NtERF3b mutant (NtERF3bΔEAR) that lacked cell death-inducing ability suppressed the induction of cell death caused by NtERF3a. The transient co-expression of NtERF3bΔEAR suppressed the hypersensitive reaction (HR)-like cell death induced by NtSIPK and NtWRKY1. The induction of cell death by NtSIPK and NtWRKY1 was also inhibited in transgenic plants expressing NtERF3bΔEAR. Analysis of gene expression, ethylene production and cell death symptoms in salicylic acid-deficient tobacco plants suggested the existence of some feedback regulation in the HR cell death signalling pathway mediated by SIPK/WIPK and WRKY1. Overall, these results suggest that NtERF3 functions downstream of NtSIPK/NtWIPK and NtWRKY1 in a cell death signalling pathway, with some feedback regulation.

  2. IMPre: An Accurate and Efficient Software for Prediction of T- and B-Cell Receptor Germline Genes and Alleles from Rearranged Repertoire Data

    PubMed Central

    Zhang, Wei; Wang, I-Ming; Wang, Changxi; Lin, Liya; Chai, Xianghua; Wu, Jinghua; Bett, Andrew J.; Dhanasekaran, Govindarajan; Casimiro, Danilo R.; Liu, Xiao

    2016-01-01

    Large-scale study of the properties of T-cell receptor (TCR) and B-cell receptor (BCR) repertoires through next-generation sequencing is providing excellent insights into the understanding of adaptive immune responses. Variable(Diversity)Joining [V(D)J] germline genes and alleles must be characterized in detail to facilitate repertoire analyses. However, most species do not have well-characterized TCR/BCR germline genes because of their high homology. Also, more germline alleles are required for humans and other species, which limits the capacity for studying immune repertoires. Herein, we developed “Immune Germline Prediction” (IMPre), a tool for predicting germline V/J genes and alleles using deep-sequencing data derived from TCR/BCR repertoires. We developed a new algorithm, “Seed_Clust,” for clustering, produced a multiway tree for assembly and optimized the sequence according to the characteristics of rearrangement. We trained IMPre on human samples of T-cell receptor beta (TRB) and immunoglobulin heavy chain and then tested it on additional human samples. Accuracy of 97.7, 100, 92.9, and 100% was obtained for TRBV, TRBJ, IGHV, and IGHJ, respectively. Analyses of subsampling performance for these samples showed IMPre to be robust using different data quantities. Subsequently, IMPre was tested on samples from rhesus monkeys and human long sequences: the highly accurate results demonstrated IMPre to be stable with animal and multiple data types. With rapid accumulation of high-throughput sequence data for TCR and BCR repertoires, IMPre can be applied broadly for obtaining novel genes and a large number of novel alleles. IMPre is available at https://github.com/zhangwei2015/IMPre. PMID:27867380

  3. Overexpression of AaWRKY1 Leads to an Enhanced Content of Artemisinin in Artemisia annua

    PubMed Central

    Jiang, Weimin; Fu, Xueqing; Pan, Qifang; Tang, Yueli; Shen, Qian; Lv, Zongyou; Yan, Tingxiang; Shi, Pu; Li, Ling; Zhang, Lida; Wang, Guofeng; Sun, Xiaofen; Tang, Kexuan

    2016-01-01

    Artemisinin is an effective component of drugs against malaria. The regulation of artemisinin biosynthesis is at the forefront of artemisinin research. Previous studies showed that AaWRKY1 can regulate the expression of ADS, which is the first key enzyme in artemisinin biosynthetic pathway. In this study, AaWRKY1 was cloned, and it activated ADSpro and CYPpro in tobacco using dual-LUC assay. To further study the function of AaWRKY1, pCAMBIA2300-AaWRKY1 construct under 35S promoter was generated. Transgenic plants containing AaWRKY1 were obtained, and four independent lines with high expression of AaWRKY1 were analyzed. The expression of ADS and CYP, the key enzymes in artemisinin biosynthetic pathway, was dramatically increased in AaWRKY1-overexpressing A. annua plants. Furthermore, the artemisinin yield increased significantly in AaWRKY1-overexpressing A. annua plants. These results showed that AaWRKY1 increased the content of artemisinin by regulating the expression of both ADS and CYP. It provides a new insight into the mechanism of regulation on artemisinin biosynthesis via transcription factors in the future. PMID:27064403

  4. Allelic variation of polyphenol oxidase (PPO) genes located on chromosomes 2A and 2D and development of functional markers for the PPO genes in common wheat.

    PubMed

    He, X Y; He, Z H; Zhang, L P; Sun, D J; Morris, C F; Fuerst, E P; Xia, X C

    2007-06-01

    Polyphenol oxidase (PPO) activity is highly related to the undesirable browning of wheat-based end products, especially Asian noodles. Characterization of PPO genes and the development of their functional markers are of great importance for marker-assisted selection in wheat breeding. In the present study, complete genomic DNA sequences of two PPO genes, one each located on chromosomes 2A and 2D and their allelic variants were characterized by means of in silico cloning and experimental validation. Sequences were aligned at both DNA and protein levels. Two haplotypes on chromosome 2D showed 95.2% sequence identity at the DNA level, indicating much more sequence diversity than those on chromosome 2A with 99.6% sequence identity. Both of the PPO genes on chromosomes 2A and 2D contain an open reading frame (ORF) of 1,731 bp, encoding a PPO precursor peptide of 577 amino acids with a predicted molecular mass of approximately 64 kD. Two complementary dominant STS markers, PPO16 and PPO29, were developed based on the PPO gene haplotypes located on chromosome 2D; they amplify a 713-bp fragment in cultivars with low PPO activity and a 490-bp fragment in those with high PPO activity, respectively. The two markers were mapped on chromosome 2DL using a doubled haploid population derived from the cross Zhongyou 9507/CA9632, and a set of nullisomic-tetrasomic lines and ditelosomic line 2DS of Chinese Spring. QTL analysis indicated that the PPO gene co-segregated with the two STS markers and was closely linked to SSR marker Xwmc41 on chromosome 2DL, explaining from 9.6 to 24.4% of the phenotypic variance for PPO activity across three environments. In order to simultaneously detect PPO loci on chromosomes 2A and 2D, a multiplexed marker combination PPO33/PPO16 was developed and yielded distinguishable DNA patterns in a number of cultivars. The STS marker PPO33 for the PPO gene on chromosome 2A was developed from the same gene sequences as PPO18 that we reported previously, and

  5. Allelic asymmetry of the Lethal hybrid rescue (Lhr) gene expression in the hybrid between Drosophila melanogaster and D. simulans: confirmation by using genetic variations of D. melanogaster.

    PubMed

    Shirata, Mika; Araye, Quenta; Maehara, Kazunori; Enya, Sora; Takano-Shimizu, Toshiyuki; Sawamura, Kyoichi

    2014-02-01

    In the cross between Drosophila melanogaster females and D. simulans males, hybrid males die at the late larval stage, and the sibling females also die at later stages at high temperatures. Removing the D. simulans allele of the Lethal hybrid rescue gene (Lhr (sim) ) improves the hybrid incompatibility phenotypes. However, the loss-of-function mutation of Lhr (sim) (Lhr (sim0) ) does not rescue the hybrid males in crosses with several D. melanogaster strains. We first describe the genetic factor possessed by the D. melanogaster strains. It has been suggested that removing the D. melanogaster allele of Lhr (Lhr (mel) ), that is Lhr (mel0) , does not have the hybrid male rescue effect, contrasting to Lhr (sim0) . Because the expression level of the Lhr gene is known to be Lhr (sim) > Lhr (mel) in the hybrid, Lhr (mel0) may not lead to enough of a reduction in total Lhr expression. Then, there is a possibility that the D. melanogaster factor changes the expression level to Lhr (sim) < Lhr (mel) . But in fact, the expression level was Lhr (sim) > Lhr (mel) in the hybrid irrespectively of the presence of the factor. At last, we showed that Lhr (mel0) slightly improves the viability of hybrid females, which was not realized previously. All of the present results are consistent with the allelic asymmetry model of the Lhr gene expression in the hybrid.

  6. Physical and genetic mapping of the serpin gene cluster at 14q32.1: allelic association and a unique haplotype associated with alpha 1-antitrypsin deficiency.

    PubMed Central

    Byth, B. C.; Billingsley, G. D.; Cox, D. W.

    1994-01-01

    The alpha 1-antitrypsin (PI) gene is part of a cluster of structurally related serine protease inhibitor genes localized at chromosome 14q32.1, a cluster that includes the alpha 1-antichymotrypsin (AACT), protein C inhibitor (PCI), and corticosteroid-binding globulin (CBG) genes and the alpha 1-antitrypsin-like pseudogene (PIL). The order of the genes is refined here by genetic mapping using simple tandem repeat polymorphisms (STRPs) and by physical mapping in YACs. The order of the genes is (centromere)-CBG-PIL-PI-PCI-AACT-(telomere). Analysis of DNA haplotypes comprising STRP and RFLP markers in the serpin genes reveals considerable allelic association throughout the cluster. Furthermore, the common alpha 1-antitrypsin deficiency allele, PI*Z, has a unique DNA haplotype at the CBG, PIL, and PI loci, which extends over 60 kb in 97% of cases and in 44% of cases includes the PCI and AACT loci. This unique haplotype will be of use in examining a number of other diseases, particularly those with an inflammatory component, thought to be associated with alpha 1-antitrypsin deficiency or partial deficiency. Images Figure 1 Figure 3 PMID:7912884

  7. [Allelic state of the molecular marker for the golden nematode (Globodera rostochiensis) resistance gene H1 among Ukrainian and world cultivars of potato (Solanum tuberosum ssp. tuberosum)].

    PubMed

    Karelov, A V; Pilipenko, L A; Kozub, N A; Bondus, R A; Borzykh, A U; Sozinov, I A; Blium, Ia B; Sozinov, A A

    2013-01-01

    The purpose of our investigation was determination of allelic state of the H1 resistance gene against the pathotypes Ro1 and Ro4 of golden potato cyst nematode (Globodera rostochiensis) among Ukrainian and world potato (Solanum tuberosum ssp. tuberosum) cultivars. The allelic condition of the TG689 marker was determined by PCR with DNA samples isolated from tubers of potato and primers, one pair of which flanks the allele-specific region and the other one was used for the control of DNA quality. Among analyzed 77 potato cultivars the allele of marker associated with the H1-type resistance was found in 74% of Ukrainian and 90% foreign ones although some of those cultivars proved to be susceptible to the golden potato nematode in field. The obtained data confirm the presence of H1-resistance against golden nematode pathotypes Ro1 and Ro4 among the Ukrainian potato cultivars and efficiency of the used marker within the accuracy that has been declared by its authors.

  8. Polymorphism of the bovine POU1F1 gene: allele frequencies and effects on milk production in three Iranian native breeds and Holstein cattle of Iran.

    PubMed

    Zakizadeh, S; Reissmann, M; Rahimi, G; Javaremi, A Nejati; Reinecke, P; Mirae-Ashtiani, S R; Shahrbabak, M Moradi

    2007-08-01

    The aim of this study was to estimate the allele frequencies in polymorphic site of exon six of POU1F1 gene in three Iranian native and Holstein cattle. Genomic DNA was extracted from 3 Iranian native cattle breeds, including 97 Mazandarani, 87 Sarabi, 112 Golpaygani and also 110 Holstein cattle. A 451 bp fragment of intron 5 and exon 6 were amplified and digested with HinfI restriction enzyme. Frequencies of allele A were 0.37, 0.27, 0.34 and 0.21 for Mazandarani, Sarabi, Golpaygani and Holstein cattle, respectively. Significant differences in genotype frequencies were found between Mazandarani or Golpaygani and Holstein cattle. No significant differences in genotype frequencies were found between Sarabi and Holstein cattle. Transition A to G in nucleotide 1256 is responsible for HinfI(-) allele. No significant association was observed between POU1F1 polymorphism and milk production. Differences in allelic frequency between native Bos indicus breeds (Mazandarani, Golpaygani) and Holstein at the present study might be due to differences in origin breeds, low number of samples and/or as the effect of natural selection in native breeds.

  9. A dominant truncation allele identifies a gene, STE20, that encodes a putative protein kinase necessary for mating in Saccharomyces cerevisiae.

    PubMed Central

    Ramer, S W; Davis, R W

    1993-01-01

    This work reports the identification, characterization, and nucleotide sequence of STE20, a newly discovered gene involved in the Saccharomyces cerevisiae mating response pathway, to date one of the best understood signal transduction pathways. STE20 encodes a putative serine/threonine-specific protein kinase with a predicted molecular mass of 102 kDa. Its expression pattern is similar to that of several other protein kinases in the mating response pathway. Deletion of the kinase domain of STE20 causes sterility in both haploid mating types. This sterility can be partially suppressed by high-level production of STE12 but is not suppressible by high levels of STE4 or a dominant STE11 truncation allele. A truncation allele of STE20 was isolated that can activate the mating response pathway in the absence of exogenous mating pheromone. This allele causes dominant growth arrest that cannot be suppressed by deletions of STE4, STE5, STE7, STE11, or STE12. The allele is able to suppress the mating defect of a strain in which the STE20 kinase domain has been deleted, but not the mating defects of strains carrying mutations in STE4, STE5, STE7, STE11, or STE12. Images PMID:8421676

  10. Allele-specific germ cell epimutation in the spacer promoter of the 45S ribosomal RNA gene after Cr(III) exposure

    SciTech Connect

    Shiao, Y.-H. . E-mail: shiao@mail.ncifrcf.gov; Crawford, Erik B.; Anderson, Lucy M.; Patel, Pritesh; Ko, Kinarm

    2005-06-15

    Paternal exposure of mice to Cr(III) causes increased tumor risk in offspring; an epigenetic mechanism has been hypothesized. Representational difference analysis of gene methylation in sperm revealed hypomethylation in the 45S ribosomal RNA (rRNA) gene after Cr(III) exposure, compared with controls. The most striking effects were seen in the rRNA spacer promoter, a region in the intergenic region of rRNA gene clusters that can influence transcription. Methylation of the rRNA spacer promoter has not been studied heretofore. Sperm DNAs from Cr(III)-treated and control mice were modified by the bisulfite method followed by PCR amplification of the spacer promoter, including 27 CpG sites. Cloning and dideoxy sequencing identified sequence variants (T or G at base -2214) in the spacer promoter. The T allele had less DNA methylation than the G allele in control mice (17 of 17 clones vs. 42 of 72 clones, P = 0.0004). In spite of diversity of sperm DNA methylation patterns, the DNA clones from Cr(III)-exposed mice had fewer methylated CpG sites, by an average of 19% (P < 0.0001). This difference was limited to the G allele. The pyrosequencing technique was applied to quantify the percentage of methylation directly from amplified PCR products. Strikingly, for nine CpG sites including the spacer promoter core region, hypomethylation was highly significant in the Cr(III)-treated group (paired T test, P < 0.0001). Thus, one allele of the 45S rRNA spacer promoter is hypomethylated in sperm germ cells after Cr(III) exposure. This epimutation may lead to increase of tumor risk in the offspring.

  11. Allelic differences between Europeans and Chinese for CREB1 SNPs and their implications in gene expression regulation, hippocampal structure and function, and bipolar disorder susceptibility.

    PubMed

    Li, M; Luo, X-J; Rietschel, M; Lewis, C M; Mattheisen, M; Müller-Myhsok, B; Jamain, S; Leboyer, M; Landén, M; Thompson, P M; Cichon, S; Nöthen, M M; Schulze, T G; Sullivan, P F; Bergen, S E; Donohoe, G; Morris, D W; Hargreaves, A; Gill, M; Corvin, A; Hultman, C; Toga, A W; Shi, L; Lin, Q; Shi, H; Gan, L; Meyer-Lindenberg, A; Czamara, D; Henry, C; Etain, B; Bis, J C; Ikram, M A; Fornage, M; Debette, S; Launer, L J; Seshadri, S; Erk, S; Walter, H; Heinz, A; Bellivier, F; Stein, J L; Medland, S E; Arias Vasquez, A; Hibar, D P; Franke, B; Martin, N G; Wright, M J; Su, B

    2014-04-01

    Bipolar disorder (BD) is a polygenic disorder that shares substantial genetic risk factors with major depressive disorder (MDD). Genetic analyses have reported numerous BD susceptibility genes, while some variants, such as single-nucleotide polymorphisms (SNPs) in CACNA1C have been successfully replicated, many others have not and subsequently their effects on the intermediate phenotypes cannot be verified. Here, we studied the MDD-related gene CREB1 in a set of independent BD sample groups of European ancestry (a total of 64,888 subjects) and identified multiple SNPs significantly associated with BD (the most significant being SNP rs6785[A], P=6.32 × 10(-5), odds ratio (OR)=1.090). Risk SNPs were then subjected to further analyses in healthy Europeans for intermediate phenotypes of BD, including hippocampal volume, hippocampal function and cognitive performance. Our results showed that the risk SNPs were significantly associated with hippocampal volume and hippocampal function, with the risk alleles showing a decreased hippocampal volume and diminished activation of the left hippocampus, adding further evidence for their involvement in BD susceptibility. We also found the risk SNPs were strongly associated with CREB1 expression in lymphoblastoid cells (P<0.005) and the prefrontal cortex (P<1.0 × 10(-6)). Remarkably, population genetic analysis indicated that CREB1 displayed striking differences in allele frequencies between continental populations, and the risk alleles were completely absent in East Asian populations. We demonstrated that the regional prevalence of the CREB1 risk alleles in Europeans is likely caused by genetic hitchhiking due to natural selection acting on a nearby gene. Our results suggest that differential population histories due to natural selection on regional populations may lead to genetic heterogeneity of susceptibility to complex diseases, such as BD, and explain inconsistencies in detecting the genetic markers of these diseases among

  12. Allele-specific transcriptional activity of the variable number of tandem repeats in 5' region of the DRD4 gene is stimulus specific in human neuronal cells.

    PubMed

    Paredes, U M; Quinn, J P; D'Souza, U M

    2013-03-01

    The dopamine receptor D4 (DRD4) gene includes several variable number of tandem repeat loci that have been suggested to modulate DRD4 gene expression patterns. Previous studies showed differential basal activity of the two most common variants of a tandem repeat (120 bp per repeat unit) located in the 5' region adjacent to the DRD4 promoter in human cell lines. In this communication, we further characterized the ability of this polymorphic repeat to elicit tissue-, allele- and stimuli-specific transcriptional activity in vitro. The short and long variants of the DRD4 5' tandem repeat were cloned into a luciferase reporter gene construct containing the SV40 promoter. The luciferase constructs were cotransfected with expression vectors of two ubiquitously expressed human transcription factors (TFs), CCCTC-binding factor (CTCF) and upstream stimulatory factor 2 (USF2), into human cell lines and primary cultures of neonate rat cortex and luciferase activity measured. Overexpression with these TFs resulted in differential cell- and allele-specific transcriptional activities of the luciferase constructs. The results of our experiments show that variants of this tandem repeat in the 5' promoter of the DRD4 gene will direct differential reporter gene transcriptional activity in a cell-type-specific manner dependent on the signal pathways activated.

  13. Hybrid weakness in a rice interspecific hybrid is nitrogen-dependent, and accompanied by changes in gene expression at both total transcript level and parental allele partitioning

    PubMed Central

    Lin, Xiuyun; Wang, Jie; Yu, Jiamiao; Sun, Yue; Miao, Yiling; Li, Qiuping; Sanguinet, Karen A.; Liu, Bao

    2017-01-01

    Background Hybrid weakness, a phenomenon opposite to heterosis, refers to inferior growth and development in a hybrid relative to its pure-line parents. Little attention has been paid to the phenomenological or mechanistic aspect of hybrid weakness, probably due to its rare occurrence. Methodology/Principal findings Here, using a set of interspecific triploid F1 hybrids between Oryza sativa, ssp. japonica (genome AA) and a tetraploid wild rice species, O. alta (genome, CCDD), we investigated the phenotypic and physiological differences between the F1 hybrids and their parents under normal and nitrogen-limiting conditions. We quantified the expression levels of 21 key genes involved in three important pathways pertinent to the assayed phenotypic and physiological traits by real-time qRT-PCR. Further, we assayed expression partitioning of parental alleles for eight genes in the F1 hybrids relative to the in silico “hybrids” (parental cDNA mixture) under both normal and N-limiting conditions by using locus-specific cDNA pyrosequencing. Conclusions/Significance We report that the F1 hybrids showed weakness in several phenotypic traits at the final seedling-stage compared with their corresponding mid-parent values (MPVs). Nine of the 21 studied genes showed contrasted expression levels between hybrids and parents (MPVs) under normal vs. N-limiting conditions. Interestingly, under N-limiting conditions, the overtly enhanced partitioning of maternal allele expression in the hybrids for eight assayed genes echo their attenuated hybrid weakness in phenotypes, an observation further bolstered by more resemblance of hybrids to the maternal parent under N-limiting conditions compared to normal conditions in a suite of measured physiological traits. Our observations suggest that both overall expression level and differential partitioning of parental alleles of critical genes contribute to condition-specific hybrid weakness. PMID:28248994

  14. Dominance relationships among mutant alleles of regulatory gene araC in the Escherichia coli B/R L-arabinose operon.

    PubMed

    Sheppard, D E

    1986-11-01

    The araBAD operon of Escherichia coli B/r is positively and negatively regulated by the araC+ regulatory protein. Mutations in gene araC can result in a variety of different regulatory phenotypes: araC null mutants (those carrying a null allele exhibiting no repressor or activator activity) are unable to achieve operon induction; araC-constitutive (araCc) mutants are partially constitutive, inducible by D-fucose, and resistant to catabolite repression; araCh mutants are hypersensitive to catabolite repression; and araCi mutants are resistant to catabolite repression. Various mutant alleles of gene araC were cloned into a derivative of plasmid pBR322 by in vivo recombination. Various heterozygous araC allelic combinations were constructed by transformation. Analysis of isomerase (araA) specific activity levels under various growth conditions indicated the following dominance relationships with regard to sensitivity to catabolite repression: araCh greater than araC+ greater than (araCc and araCi) greater than araC. It was concluded that the araCh protein may form a repressor complex that is refractory to removal by cyclic AMP receptor protein-cyclic AMP complex. This was interpreted in terms of the known nucleoprotein interactions between ara regulatory proteins and ara regulatory DNA.

  15. Evidence that the penetrance of mutations at the RP11 locus causing dominant retinitis pigmentosa is influenced by a gene linked to the homologous RP11 allele.

    PubMed Central

    McGee, T L; Devoto, M; Ott, J; Berson, E L; Dryja, T P

    1997-01-01

    A subset of families with autosomal dominant retinitis pigmentosa (RP) display reduced penetrance with some asymptomatic gene carriers showing no retinal abnormalities by ophthalmic examination or by electroretinography. Here we describe a study of three families with reduced-penetrance RP. In all three families the disease gene appears to be linked to chromosome 19q13.4, the region containing the RP11 locus, as defined by previously reported linkage studies based on five other reduced-penetrance families. Meiotic recombinants in one of the newly identified RP11 families and in two of the previously reported families serve to restrict the disease locus to a 6-cM region bounded by markers D19S572 and D19S926. We also compared the disease status of RP11 carriers with the segregation of microsatellite alleles within 19q13.4 from the noncarrier parents in the newly reported and the previously reported families. The results support the hypothesis that wild-type alleles at the RP11 locus or at a closely linked locus inherited from the noncarrier parents are a major factor influencing the penetrance of pathogenic alleles at this locus. PMID:9345108

  16. Determination of cis/trans phase of variations in the MC1R gene with allele-specific PCR and single base extension.

    PubMed

    Mengel-From, Jonas; Børsting, Claus; Sanchez, Juan J; Eiberg, Hans; Morling, Niels

    2008-12-01

    The MC1R gene encodes a protein with key regulatory functions in the melanin synthesis. A multiplex PCR and a multiplex single base extension protocol were established for genotyping six exonic MC1R variations highly penetrant for red hair (R), four exonic MC1R variations weakly penetrant for red hair (r), two frameshift variations highly penetrant for red hair (R) and three variations in the promoter region. We genotyped 600 individuals from Denmark using either CE or MALDI-TOF MS as the detection platform. A total of 62 individuals were genotyped R/R and among the 62 individuals, 57 had red hair and five had blond hair colour. Two different R alleles may be located in cis (RR/-) position or trans (R/R) position, and the phenotype associated with RR/- and R/R may be different. Two allele-specific PCRs were established with primers targeting the -G445A variation in the MC1R promoter and the allele-specific PCR products were used in the multiplex single base extension assay. In all 62 individuals, the MC1R variants were situated in trans position. Another 18 individuals with red hair colour were either genotyped R/- or R/r, suggesting that other genes influence hair colour.

  17. Evidence for convergent nucleotide evolution and high allelic turnover rates at the complementary sex determiner gene of Western and Asian honeybees.

    PubMed

    Hasselmann, Martin; Vekemans, Xavier; Pflugfelder, Jochen; Koeniger, Nikolaus; Koeniger, Gudrun; Tingek, Salim; Beye, Martin

    2008-04-01

    Our understanding of the impact of recombination, mutation, genetic drift, and selection on the evolution of a single gene is still limited. Here we investigate the impact of all these evolutionary forces at the complementary sex determiner (csd) gene that evolves under a balancing mode of selection. Females are heterozygous at the csd gene and males are hemizygous; diploid males are lethal and occur when csd is homozygous. Rare alleles thus have a selective advantage, are seldom lost by the effect of genetic drift, and are maintained over extended periods of time when compared with neutral polymorphisms. Here, we report on the analysis of 17, 19, and 15 csd alleles of Apis cerana, Apis dorsata, and Apis mellifera honeybees, respectively. We observed great heterogeneity of synonymous (piS) and nonsynonymous (piN) polymorphisms across the gene, with a consistent peak in exons 6 and 7. We propose that exons 6 and 7 encode the potential specifying domain (csd-PSD) that has accumulated elevated nucleotide polymorphisms over time by balancing selection. We observed no direct evidence that balancing selection favors the accumulation of nonsynonymous changes at csd-PSD (piN/piS ratios are all <1, ranging from 0.6 to 0.95). We observed an excess of shared nonsynonymous changes, which suggest that strong evolutionary constraints are operating at csd-PSD resulting in the independent accumulation of the same nonsynonymous changes in different alleles across species (convergent evolution). Analysis of csd-PSD genealogy revealed relatively short average coalescence times ( approximately 6 Myr), low average synonymous nucleotide diversity (piS < 0.09), and a lack of trans-specific alleles that substantially contrasts with previously analyzed loci under strong balancing selection. We excluded the possibility of a burst of diversification after population bottlenecking and intragenic recombination as explanatory factors, leaving high turnover rates as the explanation for this

  18. CKM Gene G (Ncoi-) Allele Has a Positive Effect on Maximal Oxygen Uptake in Caucasian Women Practicing Sports Requiring Aerobic and Anaerobic Exercise Metabolism

    PubMed Central

    Gronek, Piotr; Holdys, Joanna; Kryściak, Jakub; Stanisławski, Daniel

    2013-01-01

    The search for genes with a positive influence on physical fitness is a difficult process. Physical fitness is a trait determined by multiple genes, and its genetic basis is then modified by numerous environmental factors. The present study examines the effects of the polymorphism of creatine kinase (CKM) gene on VO2max – a physiological index of aerobic capacity of high heritability. The study sample consisted of 154 men and 85 women, who were students of the University School of Physical Education in Poznań and athletes practicing various sports, including members of the Polish national team. The study revealed a positive effect of a rare G (NcoI−) allele of the CKM gene on maximal oxygen uptake in Caucasian women practicing sports requiring aerobic and anaerobic exercise metabolism. Also a tendency was noted in individuals with NcoI−/− (GG) and NcoI−/+ (GA) genotypes to reach higher VO2max levels. PMID:24511349

  19. CKM Gene G (Ncoi-) Allele Has a Positive Effect on Maximal Oxygen Uptake in Caucasian Women Practicing Sports Requiring Aerobic and Anaerobic Exercise Metabolism.

    PubMed

    Gronek, Piotr; Holdys, Joanna; Kryściak, Jakub; Stanisławski, Daniel

    2013-12-18

    The search for genes with a positive influence on physical fitness is a difficult process. Physical fitness is a trait determined by multiple genes, and its genetic basis is then modified by numerous environmental factors. The present study examines the effects of the polymorphism of creatine kinase (CKM) gene on VO2max - a physiological index of aerobic capacity of high heritability. The study sample consisted of 154 men and 85 women, who were students of the University School of Physical Education in Poznań and athletes practicing various sports, including members of the Polish national team. The study revealed a positive effect of a rare G (NcoI-) allele of the CKM