条形柄锈菌小麦专化型葡萄糖-6-磷酸脱氢酶基因PsG6PDH1的表达特征及功能分析  

Expression and functional characterizations of a glucose-6-phosphatase dehydrogenase gene PsG6PDH1 from Puccinia striiformis f. sp. tritici

作　　者：王秋玲[1] 李丹[1] 怀宝玉 郑佩晶 康振生[2] 刘杰[1] 

机构地区：[1]旱区作物逆境生物学国家重点实验室西北农林科技大学生命科学学院,陕西杨凌712100 [2]旱区作物逆境生物学国家重点实验室西北农林科技大学植物保护学院,陕西杨凌712100

出　　处：《菌物学报》2016年第10期1199-1207,共9页Mycosystema

基　　金：国家自然科学基金(31000078);中央高校基本科研业务费(QN2011027)~~

摘　　要：葡萄糖-6-磷酸脱氢酶是磷酸戊糖途径中的关键限速酶。基于已测序的条形柄锈菌小麦专化型基因组序列,利用RT-PCR方法克隆了该病菌葡萄糖-6-磷酸脱氢酶PsG6PDH1的全长cDNA序列(1 497bp),编码498个氨基酸的蛋白。编码蛋白含有葡萄糖-6-磷酸脱氢酶的保守功能域。系统进化分析发现,PsG6PDH1与禾柄锈菌小麦专化型的G6PDH聚为一簇。qRT-PCR分析表明,PsG6PDH1在病菌侵染早期的表达明显上调,其中侵染24h时表达量最高,为对照夏孢子的30倍。将PsG6PDH1导入酿酒酵母G6PDH缺失突变体中成功表达,并表现出较强的葡萄糖-6-磷酸脱氢酶活性,明显酵母增强了菌株对过氧化氢的耐受性。由此推测,PsG6PDH1可能参与了条形柄锈菌小麦专化型在侵染寄主时抵御寄主的氧化胁迫反应。研究结果为进一步研究该病菌基础代谢及侵染机理奠定一定的理论基础。The PsG6PDH1 gene, encoding glucose-6-phosphate dehydrogenase, which is a key enzyme in the pentose phosphate path, was cloned using reverse-transcriptional-PCR（RT-PCR） based on Puccinia striiformis f. sp. tritici（Pst） genome sequence. The c DNA sequence of PsG6PDH1 consists of 1 497 nucleotides, encoding a polypeptide of 498 amino acid containing conserved G6 PDH domains. The phylogenetic analysis revealed that PsG6PDH1 was clustered with the G6 PDH sequence from Puccinia graminis f. sp. tritici. q RT-PCR analysis showed that the expression of PsG6PDH1 was obviously up-regulated during the early stages of Pst infection and then reached the maximum at 24 h post inoculation（hpi）, which is 30-fold as high as that of the control. The function of PsG6PDH1 was identified via mutant complementation. PsG6PDH1 was expressed in Saccharomyces cerevisiae mutant lacking G6 PDH gene. The results show that PsG6PDH1 was successfully obtained and exhibited higher enzymatic activity of glucose-6-phosphate dehydrogenase. In addition, introducing PsG6PDH1 into the S. cerevisiae mutant can enhance its tolerance against hydrogen peroxide, indicating that PsG6PDH1 may play a role in resisting active oxygen stress in the host defence response. This work elucidates the function of PsG6PDH1 in Pst and provides a theoretical basis for revealing the molecular mechanism of Pst metabolism and infection.

关 键 词：条形柄锈菌小麦专化型 葡萄糖-6-磷酸脱氢酶 实时定量PCR 过氧化氢 

分 类 号：S435.121.4[农业科学—农业昆虫与害虫防治]