Ru Binglu, Hao Xinchang, Li Wenhao, Peng Qin, Miao Jianqiang, Liu Xili
State Key Laboratory of Crop Stress Biology for Arid Areas, College of Plant Protection, Northwest A&F University, 3 Taicheng Road, Yangling, Xianyang 712100, China.
Department of Plant Pathology, College of Plant Protection, China Agricultural University, 2 Yuanmingyuanxi Road, Beijing 100193, China.
J Fungi (Basel). 2023 Jun 28;9(7):709. doi: 10.3390/jof9070709.
Proteins containing both FYVE and serine/threonine kinase catalytic (STKc) domains are exclusive to protists. However, the biological function of these proteins in oomycetes has rarely been reported. In the genome database, we identified five proteins containing FYVE and STKc domains, which we named PsZFPK1, PsZFPK2, PsZFPK3, PsZFPK4, and PsZFPK5. In this study, we characterized the biological function of PsZFPK1 using a CRISPR/Cas9-mediated gene replacement system. Compared with the wild-type strain, P6497, the -knockout mutants exhibited significantly reduced growth on a nutrient-rich V8 medium, while a more pronounced defect was observed on a nutrient-poor Plich medium. The -knockout mutants also showed a significant increase in sporangium production. Furthermore, PsZFPK1 was found to be essential for oospore production and complete virulence but dispensable for the stress response in . The N-terminal region, FYVE and STKc domains, and T602 phosphorylation site were found to be vital for the function of PsZFPK1. Conversely, these domains were not required for the localization of PsZFPK1 protein in the cytoplasm. Our results demonstrate that PsZFPK1 plays a critical role in vegetative growth, sporangium formation, oospore production, and virulence in .
同时含有FYVE和丝氨酸/苏氨酸激酶催化（STKc）结构域的蛋白质仅存在于原生生物中。然而，这些蛋白质在卵菌中的生物学功能鲜有报道。在基因组数据库中，我们鉴定出了五种含有FYVE和STKc结构域的蛋白质，将其命名为PsZFPK1、PsZFPK2、PsZFPK3、PsZFPK4和PsZFPK5。在本研究中，我们使用CRISPR/Cas9介导的基因替换系统对PsZFPK1的生物学功能进行了表征。与野生型菌株P6497相比，基因敲除突变体在营养丰富的V8培养基上生长显著降低，而在营养贫瘠的Plich培养基上观察到更明显的缺陷。基因敲除突变体的孢子囊产量也显著增加。此外，发现PsZFPK1对卵孢子的产生和完全致病性至关重要，但对其应激反应并非必需。发现N端区域、FYVE和STKc结构域以及T602磷酸化位点对PsZFPK1的功能至关重要。相反，这些结构域对于PsZFPK1蛋白在细胞质中的定位并非必需。我们的结果表明，PsZFPK1在营养生长、孢子囊形成、卵孢子产生以及致病力方面发挥着关键作用。
