草菇谷胱甘肽S-转移酶编码基因(vv-gto1)的序列分析及其差异表达

Sequence characterization and differential expression of a glutathione S-transferase gene vv-gto1 from Volvariella volvacea

摘要:

摘要:【目的】通过对草菇组学数据的分析，获得一个谷胱甘肽S-转移酶(GSTs)编码基因(vv-gto1)，对其基因结构、序列特征及表达情况进行分析，探索GSTs 在食用菌生长发育过程中的作用机制，并为其他食用菌GSTs编码基因的发现提供依据。【方法】应用ZOOM软件将基因组与转录组的测序读段(reads)与基因组拼接序列进行定位确定基因全长及其序列准确性，并对基因结构进行可视化分析。采用MEGA 5.1进行多序列比对及进化树分析，对草菇GTO1进行功能归类及同其他真菌的亲缘关系分析。结合表达谱、蛋白质组数据分析该基因在草菇不同生长时期的差异表达，并应用荧光定量PCR进行验证。【结果】vv-gto1全长2083bp，含有11个外显子、10个内含子，编码356个氨基酸;5'端非翻译区长度为305bp、包含1个内含子区域，3'端非翻译区为86bp;RNA加工过程中有两个内含子存在保留现象，由内含子保留所形成的转录本均无法翻译形成正确的保守功能域。vv-gto1全长序列都获得超过50个reads的准确定位，证明该区段的基因组测序及组装结果正确可靠。进化树结果显示，草菇GTO1属于Omega类第I小类谷胱甘肽S-转移酶，与黄孢原毛平革菌的GTO1、GTO2亲缘关系最近。数字基因表达谱、荧光定量PCR及蛋白质组的分析结果表明，vv-gto1在异核体菌丝H1521的表达量最高。【结论】本研究首次在草菇中获得一个Omega类谷胱甘肽S-转移酶编码基因，该基因在异核体菌丝生物学功能的行使过程中可能起到重要作用，同时也暗示草菇异核体菌丝具备较强的抗逆性。另外，vv-gto1可以通过形成不同的可变剪接体来调节基因的转录和翻译，最终影响蛋白质的功能行使。

Abstract:

Abstract:［Objective］ Based on the analysis of omics data of Volvariella volvacea，a gene encoding glutathione Stransferase (GSTs) named vv-gto1 was obtained．To reveal the role of GSTs in the growth and development in edible fungi，the structure，the sequence characters and the expression profile of a GST gene vv-gto1 of Volvariella volvacea were analyzed．［Methods］ZOOM software was used to map sequencing read ( reads) from genome and transcriptome against the splicing sequence of genome，to confirm the complete length and the accuracy of the gene sequence，and to visualize gene structure．The MEGA 5.1 was used to do the multiple sequence alignment and phylogenetic tree analysis． Ｒeal time fluorescent quantitative PCR was used to determine the expression levels of vv-gto1 at different growth periods of Volvariella volvacea． ［Ｒesults］The full sequence of vv-gto1 covered 2083bp，containing 11 exons and 10 introns，and encoded a protein with 356 amino acids．5'UTＲ was 305bp which contains one intron region，and 3'UTＲ was 86bp． Two intron retentions could be recognized during RNA processing，and the transcripts formed by the intron retention could not translate the correct conservative functional domains．The full-length of vv-gto1 had more than 50 accurate positioning genome sequencing reads，suggesting that genome sequencing and assembly results are accurate and reliable． The phylogenetic tree showed that GTO1 of Volvariella volvacea belonged to the subclass I of the Omega class of glutathione S-transferase superfamily，and had the closest relationship with GTO1 and GTO2 in Phanerochaete chrysosporium． The analysis of digital gene expression profiling，fluorescence quantitative PCＲ and proteomics showed that vv-gto1 had the highest expression level in the heterokaryotic hyphae．［Conclusions］This is the first time to obtain a gene encoding glutathione S-transferase from Volvariella volvacea which belongs to Omega class．Our study showed that the gene may play an important role during the special biological functions of heterokaryotic hyphae． This study also suggested that Volvariella volvacea heterokaryotic hyphae in H1521 had stronger resistance ability than other samples． In addition，vv-gto1 could form different alternative splicesome to regulate gene transcription and translation，and ultimately affect the function of the protein．