稻瘟病菌丝氨酸/苏氨酸磷酸酶MoPpz1的功能研究分析

稻瘟病菌（Magnaporthe oryzae）是一种在水稻不同生育期均能引起病害的真菌。在真核生物中,蛋白激酶CK2高度保守,包含2个调节亚基和2个催化亚基。稻瘟病菌中MoCKb1和MoCKb2分别编码MoCK2的2个调节亚基,MoCKa编码稻瘟病菌MoCK2的催化亚基。运用反向遗传学策略和同源重组原理,对MoCKa-GFP的pull-down实验结果中获得的一个丝氨酸/苏氨酸磷酸酶MoPpz1进行功能分析。经过生物信息学研究分析发现Ppz1在不同的真菌中具有不同的功能,同时与其他模式真菌的Ppz1蛋白序列具有较高同源性。在获得基因敲除突变体后,通过表型分析发现,与野生型Ku80相比,ΔMoppz1突变体菌落的营养生长和产孢量等方面均无明显差异,但分生孢子的萌发和附着胞的形成滞后于野生型菌株,且对水稻的致病能力减弱。激光共聚焦显微分析表明,MoPpz1定位在菌丝、分生孢子和附着胞的细胞质中。综上所述,蛋白磷酸酶MoPpz1可能参与稻瘟病菌分生孢子萌发、附着胞形成以及对水稻致病性的调控。

Functional Analysis of Serine/Threonine Phosphatase MoPpz1 in Magnaporthe oryzae

Magnaporthe oryzae is a fungal pathogen causing blast disease in different stages of rice. In eukaryotes, the highly conserved protein kinase CK2 contains two regulatory subunits and two catalytic subunits. MoCK2, CK2 homologus in M. oryzae, is encoding by three genes, MoCKb1 and MoCKb2 encode two different regulatory subunits, and MoCKa encodes its catalytic subunit. MoPpz1, one of serine/threonine phosphatase obtained by pull-down assay of MoCKa-GFP in M. oryzae, was researched at its function by reverse genetic approache and homologous recombination principle in our study. It was found that Ppz1 had different functions in various fungi by bioinformatics analysis, and showed that Ppz1 and it’s homolog proteins in other model fungi had highly identity in the protein sequences. ΔMoppz1 mutant obtained by gene knock-out and genotyping screen, while the mutant was not affected in vegetative growth and conidial production, the mutant showed development delay of conidium from germination to appressorium and reduced pathogenicity to rice after phenotypic assay comparing with wildtype strain Ku80. Confocal laser scanning microscopy analysis showed that MoPpz1 was localized in the cytoplasm of hyphae, conidium and appressorium. In conclusion, the protein phosphatase MoPpz1 may be involved in the regulation of conidial germination, appressorial formation in M. oryzae and pathogenicity to rice.