杨树海藻糖-6-磷酸合成酶基因家族序列与功能分化的意义

项目名称： 杨树海藻糖-6-磷酸合成酶基因家族序列与功能分化的意义

项目编号： No.30800873

项目类型： 青年科学基金项目

立项/批准年度： 2009

项目学科： 生物科学

项目作者： 杨海灵

作者单位： 北京林业大学

项目金额： 18万元

中文摘要： 基因家族的进化及其功能分化是后基因组时代进化生物学研究的核心问题。我们以海藻糖-6-磷酸合成酶（TPS）基因家族为研究对象，从毛果杨全基因组中鉴定了12个TPS基因，系统进化分析发现这些基因分为两种类型，第一种有两个成员，含有16个内含子，而第二种有10个成员，仅含有2个内含子，这些基因在杨树基因组中主要是通过全基因组加倍进行扩张。蛋白质活性位点分析发现仅第一类TPS蛋白具有结合底物的功能残基。分子进化分析发现第一类TPS基因的进化速率比第二类TPS更慢，而且受到更强的负选择压力。基因表达模式分析发现，在12个TPS基因中，有3个基因在所有组织和所有胁迫条件下都不表达，这3个TPS基因可能变成了假基因，而其余的9个基因在所有的组织和所有的胁迫条件下都表达，说明这些基因是组织构成型表达的基因。从杨树中克隆了PtTPS1，PtTPS5和PtTPS9基因，并在大肠杆菌中表达并纯化了蛋白，酶学活性分析发现仅仅PtTPS1具有催化UDP-G和G-6-P合成海藻糖6磷酸的酶学活性。我们的研究说明了杨树两类TPS基因的序列分化导致了功能的分化，这种功能分化对植物适应性功能的建立可能具有重要作用。

中文关键词： 海藻糖-6-磷酸合成酶；基因家族；分子进化；基因表达；功能分化.

英文摘要： Gene family is a group of related genes of similar sequence and function resulting from multiple duplications and subsequent mutational variation among the different copies. In this study, we identified 12 trehalose-6-phosphate synthase (TPS) genes from Populus genome. The 12 TPS genes were divided into two groups, the first group only cotained two copies with 16 introns, another group had 10 copies with 2 introns. Segmental duplications contributed significantly to the expansion of this gene family. Proteins sequence analysis showed only the first group of TPS proteins had conserved active sites conbinding to substrates UDP-G and G-6-P. PAML analysis showed the evolutional rates of the first TPS genes were slower than the second TPS genes, moreover, the first TPS genes have evolved under stronger purify selection than the second TPS genes. RT-PCR did not detected expression of three Populus TPS genes, indicating these three genes might be pseudogenes. Among 12 Populus TPS genes, nine were expressed in all tissues under all growth conditions and in response to stress treatments. Based on phylogenetic relationship, we cloned three TPS genes (PtTPS1, PtTPS5 and PtTPS9) from Populus genomes. The three TPS proteins were expressed in E.coli and recombinant proteins were purified using Ni affinity chromatography. The only PtTPS1 protein showed enzymatic activity to UDP-G and G-6-P. In conclusion, our findings revealed the role of sequence divergence in functional divergence of gene family.

英文关键词： TPS; Gene family; Molecular evolution; Gene expression; Funtional divergence.