基于转录组解析淡玫红鹅膏环肽基因家族特征及其系统发育

Characteristics and phylogeny of the cyclic peptide gene family in Amanita pallidorosea: based on transcriptome analysis

摘要:

[背景] 淡玫红鹅膏（Amanita pallidorosea）是鹅膏属檐托鹅膏组的一种剧毒鹅膏菌，其子实体内含有丰富的鹅膏环肽毒素，但其编码毒环肽和相关肽的基因家族“MSDIN”尚待深入研究。[目的] 探究淡玫红鹅膏中编码毒环肽及相关肽的基因家族成员的多样性、保守性及其系统发育。[方法] 采用Illumina HiSeq 2000平台对淡玫红鹅膏转录组进行测序，使用TBLASTn软件对MSDIN基因家族进行检索，并设计特异性引物进行PCR验证，通过生物信息学比对分析MSDIN基因家族成员的种类和序列构成，通过重建分子系统发育树了解其演化历程。[结果] 通过对MSDIN基因家族的查找，从转录组数据获得了60条环肽编码基因，经PCR验证其可编码32条环肽，包括α-鹅膏毒肽、β-鹅膏毒肽和羧基二羟鬼笔毒肽。本研究报道了8条新环肽。分子系统发育树分析显示，鹅膏环肽分为鹅膏毒肽、鬼笔毒肽和未知功能环肽3大支。系统发育结合保守序列推测出7条潜在的新毒肽。[结论] 淡玫红鹅膏具有丰富的鹅膏环肽资源，利用转录组测序能够系统挖掘鹅膏环肽新资源，为其整体结构解析奠定基础。

Abstract:

[Background] Amanita pallidorosea is a lethal Amanita species in the sect. Phalloideae, containing abundant cyclic peptide toxins. However, its gene family (MSDIN) encoding the toxic peptides and related peptides needs further systematic study. [Objective] This paper aims to explore the diversity, conservation, and phylogeny of genes encoding toxic peptides and related peptides in A. pallidorosea. [Methods] Illumina HiSeq 2000 platform was used for the transcriptome sequencing of A. pallidorosea and TBLASTn for searching MSDIN genes. Specific primers were designed for PCR verification. The types and sequence composition of MSDIN genes were analyzed with the bioinformatics method, and the phylogenetic tree was reconstructed to clarify the evolution. [Results] A total of 60 genes encoding cyclic peptides were obtained from the transcriptome data, which encoded 32 cyclic peptides as verified by PCR, including α-amanitin, β-amanitin, and phallacidin. Among them, 8 new cyclic peptide sequences were reported for the first time in this study. Molecular phylogenetic analysis showed that all cyclic peptides were classified into three clades: amatoxins, phallotoxins, and cyclic peptides of unknown function. Seven potential and new toxic peptides were predicted according to phylogenetic analysis and conserved sequences. [Conclusion] A. pallidorosea boasts abundant cyclic peptides. Transcriptome sequencing can help discover new resources of cyclic peptides in A. pallidorosea, which lays the foundation for the overall structure analysis.