水稻白叶枯病菌3-羟基脂酰ACP脱水酶基因XoofabZ的功能鉴定

Functional identification of 3-hydroxyacyl-ACP dehydratase gene XoofabZ in Xanthomonas oryzae pv. oryzae

摘要:

【背景】由水稻白叶枯病菌(Xanthomonas oryzae pv. oryzae, Xoo)引起的白叶枯病是水稻生产中最严重的细菌性病害。脂肪酸合成是细菌最重要的基础代谢之一，对细胞生长、膜磷脂和生物活性物质合成尤为重要，但对于该菌脂肪酸合成相关基因的功能知之甚少。【目的】鉴定Xoo PXO99A基因组中PXO_01118 (XoofabZ)基因是否编码3-羟基脂酰ACP脱水酶并参与脂肪酸的从头合成，为防治水稻白叶枯病害提供新思路。【方法】通过异体遗传互补和体外酶学活性检测分析XooFabZ的蛋白活性；进一步利用同源重组原理进行XoofabZ基因的遗传学分析。通过剪叶法检测替换突变株对寄主水稻的致病力。【结果】利用异体遗传互补和体外酶学活性检测手段相结合证明了XooFabZ能在脂肪酸合成的起始反应和脱水反应中发挥3-羟基脂酰ACP脱水酶活性。本研究无法直接获得XoofabZ基因敲除突变株，证明XoofabZ是必需基因。大肠埃希氏菌(Escherichia coli) EcfabZ基因替换突变株对寄主水稻的致病性显著降低。【结论】XoofabZ是水稻白叶枯病菌必需基因，编码3-羟基脂酰ACP脱水酶，参与Xoo脂肪酸从头合成反应和致病力的形成。

Abstract:

[Background] Bacterial leaf blight caused by Xanthomonas oryzae pv. oryzae (Xoo) is the most serious bacterial disease in rice production. Fatty acid synthesis is one of the most important basic metabolisms in bacteria, essential for cell growth, and the synthesis of membrane phospholipid and bioactive substances. However, little is known about the functions of fatty acid synthesis-related genes in Xoo. [Objective] This study aims to investigate whether the gene PXO_01118 (XoofabZ) in the genome of Xoo strain PXO99A encodes 3-hydroxyacyl-ACP dehydratase and participates in de novo fatty acids synthesis, providing new insights into the prevention and control of bacterial leaf blight in rice. [Methods] The protein activity of XooFabZ was analyzed through allogeneic genetic complementation and in vitro enzyme activity detection. Further, genetic analysis of XoofabZ was conducted using homologous recombination principles. The pathogenicity of the replacement mutant strains to the host rice (Oryza sativa L. ssp. Japonica cultivar Nipponbare) was detected by leaf clipping. [Results] Combining allogeneic genetic complementation and in vitro enzyme activity detection to demonstrate that XooFabZ exhibits 3-hydroxyacyl-ACP dehydratase activity in the initiation and dehydration reactions of fatty acid synthesis. It was not possible to directly obtain XoofabZ gene knockout mutants, indicating that XoofabZ is an essential gene in Xoo. The pathogenicity of Escherichia coli EcfabZ gene replacement mutant strains on the host rice was significantly reduced. [Conclusion] XoofabZ is an essential gene in Xoo, encoding 3-hydroxyacyl-ACP dehydratase, participating in de novo fatty acids synthesis reactions, and the formation of pathogenicity in Xoo.