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为了揭示水稻白叶枯病菌(Xanthomonas oryzae pv.oryzae,简称Xoo)环二鸟苷酸(c-di-GMP)信号相关蛋白PXO_03945的生物学功能,本研究通过基因同源重组和无标记交换,对PXO_03945基因进行了缺失突变,对野生型、突变体和互补菌株进行表型测定,分析该基因缺失突变对病菌胞内c-di-GMP水平、致病性、运动性、胞外多糖产生和生物膜形成的影响。结果表明,PXO_03945蛋白具有参与c-di-GMP降解的磷酸二酯酶(PDE)的HD-GYP结构域和功能未知的DUF3391结构域。全长基因缺失可导致体内c-di-GMP浓度明显上升。与野生型相比,ΔPXO_03945对水稻品种日本晴的致病性显著下降,而游动性增强,胞外多糖产生和生物膜形成增加。基因互补可以使之恢复。因此,HD-GYP结构域蛋白PXO_03945可能通过降解胞内c-di-GMP,正向调控了致病性,负向调控了运动性、EPS产生和生物膜形成能力。
In order to reveal the biological function of PXO_03945, a Xanthomonas oryzae pv. Oryzae (Xoo) cyclic diGMP signal-related protein PXO_03945 was cloned by homologous recombination and markerless exchange. The deletion of PXO_03945 gene was carried out. The phenotypes of wild-type, mutant and complementary strains were determined. The deletion of this gene was analyzed for the intracellular c-di-GMP level, pathogenicity, motility, extracellular polysaccharide production and The effect of biofilm formation. The results show that the PXO_03945 protein has the HD-GYP domain of phosphodiesterase (PDE) involved in c-di-GMP degradation and the DUF3391 domain of unknown function. The deletion of the full-length gene results in a significant increase in the concentration of c-di-GMP in the body. Compared with the wild type, the pathogenicity of ΔPXO_03945 on the rice variety Nipponbare decreased significantly, while the mobility increased, the production of extracellular polysaccharides and biofilm formation increased. Gene complementation can make it recover. Therefore, HDO-GYP domain protein PXO_03945 may positively regulate pathogenicity by degrading intracellular c-di-GMP and negatively regulate motility, EPS production and biofilm formation.