RXLR类效应分子PITG_21645.2的基因序列分析及功能验证

Sequence analysis and functional verification of a RXLR effector PITG_21645. 2

马铃薯晚疫病黑龙江菌株效应分子PITG_21645.2在本氏烟(Nicotiana benthamiana)上可以抑制由INF1引起的过敏性细胞坏死(Hypersensitive response,HR)。为验证其为致病疫霉致病的重要效应分子,克隆了10个致病疫霉菌株以及3个同属卵菌菌株中的PITG_21645.2同源基因,它们在氨基酸序列上相似度为93%~100%。与INF1共表达的结果显示,仅有PITG_21645.2^(MP903)丧失了抑制HR的功能。其中PITG_21645.2^(MP903)的第31位氨基酸存在特异性,编码丝氨酸,其余同源基因在该位点均编码天冬酰胺。PITG_21645.2^(MP903)回复突变体(S31N)能够恢复该基因抑制INF1引起HR的功能,说明第31位氨基酸是影响抑制功能的关键位点。另外,PITG_21645.2还能抑制BAX、大豆疫霉PsojNIP和效应分子Avh238在本氏烟上激发的HR,而PITG_21645.2^(MP903)都丧失抑制功能。本氏烟上表达PITG_21645.2能够促进致病疫霉在寄主上的定殖,从而提示PITG_21645.2在致病疫霉的侵染中发挥致病性功能。

PITG_21645.2,an effector of Phytophthora infestans isolates Heilongjiang（ HLJ）,could inhibit the hypersensitive response induced by INF1 in Nicotiana benthamiana. To identify that it is one of important effectors,the homologous genes of PITG_21645.2 were cloned from 10 isolates of P. infestans and three other oomycetes species including Phytophthora capsici,Phytophthora sojae and Phytophthora parasitica. They shared 93%to 100% identity with each other for the amino acid sequence. After co-infiltration with INF1,only PITG_21645.2^MP903,which contains the Serine differentiated to Asparagine of those homologues at the position of 31,couldn’t suppressed the HR triggered by INF1. And the site directed mutagenesis of the 31 th amino acid of PITG_21645.2^MP903 from Serine to Asparagine could restored the inhibition function. These implied that the 31 thposition is one of the key sites to suppress the INF1-triggered HR. Meanwhile,we identified that PITG_21645.2 HLJ but not PITG_21645.2^MP903 could also suppress the HR triggered by BAX,or other two secreted peptides PsojNIP and Avh238 of P. sojae,in N. benthamiana. Moreover,the transient expression of PITG_21645.2 HLJ could also enhance the susceptibility to P. infestans in N. benthamiana. It indicates that PITG_21645.2 plays important roles during the infection process for P. infestans.