大豆斑疹病菌胞外纤维素酶编码基因的鉴定及其调控分析  

Identification and regulation analysis of genes encoding extracellular cellulase in soybean pathogen Xanthomonas axonopodis pv. glycines

作　　者：苏如意 王宏杰 戴潇雨 高思涵 王卿怡 冯超颖 张萍华[1] 郭威[1] SU Ru-yi;WANG Hong-jie;DAI Xiao-yu;GAO Si-han;WANG Qing-yi;FENG Chao-ying;ZHANG Ping-hua;GUO Wei(College of Chemistry and Life Sciences,Zhejiang Normal University,Jinhua 321004,China)

机构地区：[1]浙江师范大学化学与生命科学学院,金华321004

出　　处：《植物病理学报》2020年第3期292-300,共9页Acta Phytopathologica Sinica

基　　金：国家级大学生创新创业训练计划(201910345031);浙江省自然科学基金(LY19C130002,LY18C140004)。

摘　　要：植物病原黄单胞菌的胞外纤维素酶基因具有差异性和多样性。大豆斑疹病菌(Xanthomonas axonopodis pv.glycines,Xag)具有胞外纤维素酶活性,但其编码基因和调控特性尚不明确。为了鉴定Xag的胞外纤维素酶编码基因,本研究从NEAU001菌株基因组内筛选出6个候选的纤维素酶编码基因。通过异源表达实验发现,engXCA和egl2基因分别编码的蛋白EngXCA和Egl2具有水解羧甲基纤维素的能力。酶活性测定实验表明,engXCA和egl2基因决定Xag的胞外纤维素酶活性。另外,对已知的10个毒性调控基因突变株进行胞外纤维素酶活性测定,揭示Xag胞外纤维素酶活性受DSF(diffusible signal factor)信号通路和全局性调控子Clp(Crp-like protein)正调控。荧光定量PCR结果进一步证实,RpfF和Clp参与正调控engXCA和egl2基因的mRNA水平。这些结果暗示,DSF信号通路可能通过Clp调控engXCA和egl2基因的表达,实现对Xag胞外纤维素酶活性的调控。The genes encoding extracellular cellulase are different and diverse in plant pathogenic Xanthomonas.Extracellular cellulase contributes to the virulence of Xanthomonas axonopodis pv.glycines(Xag),but its encoding genes and the underlying molecular regulatory mechanism are unclear.In order to identify the genes encoding extracellular cellulase in Xag,6 candidate genes were screened from the whole genome of strain NEAU001.Ectopic expression assay showed that EngXCA and Egl2 hydrolyzed carboxymethylcellulose,and the enzyme activity assay showed that engXCA and egl2 played the major role in the extracellular cellulase activity in Xag.In addition,the enzyme activity assay of 10 mutants in which the core virulence regulatory gene was deleted revealed that the extracellular cellulase activity of Xag was positively regulated by diffuse signal factor(DSF)signaling pathway and the global regulator Clp(Crp-like protein).qRT-PCR experiments further confirmed that both RpfF and Clp positively regulated engXCA and egl2 at the mRNA levels.Taken together,these results suggest that the DSF signaling pathway may regulate the expression of engXCA and egl2 through Clp,resulting in the regulation of the extracellular cellulase activity in Xag.

关 键 词：大豆斑疹病菌 胞外纤维素酶 异源表达 扩散性信号分子 全局性调控子Clp 

分 类 号：S432.42[农业科学—植物病理学]