东湖假单胞菌HYS中的精氨酸琥珀酰转移酶途径缺失对秀丽隐杆线虫毒性的影响

Arginine succinyltransferase pathway deficiency in Pseudomonas donghuensis HYS affects its toxicity to Caenorhabditis elegans

摘要:

【背景】东湖假单胞菌HYS是本实验室从武汉东湖水域中分离并鉴定的一株高产铁载体细菌，HYS菌株对秀丽隐杆线虫具有较强毒性。前期研究发现HYS中编码精氨酸琥珀酰转移酶基因(argS)的插入突变可导致其对线虫毒性明显减弱。【目的】探究argS基因功能及其如何参与细菌毒性，为后续深入研究HYS菌株的毒性机制提供理论依据。【方法】采用生物信息学比对、遗传分析和生理生化实验确认argS基因的生物学功能及其参与的精氨酸琥珀酰转移酶(arginine succinyltransferase，AST)途径与细菌毒性的关系。【结果】生物信息学比对结果显示argS编码精氨酸琥珀酰转移酶，其蛋白序列与铜绿假单胞菌中精氨酸琥珀酰转移酶β亚基具有高达88%的相似度；缺失argS导致菌株不能利用精氨酸作为唯一碳源进行生长；精氨酸脱羧酶(arginine decarboxylase，ADC)、精氨酸脱氢酶(arginine dehydrogenase，ADH)以及精氨酸脱亚胺酶(arginine deiminase，ADI)途径中关键基因缺失菌株均可正常利用精氨酸作为唯一碳源，且对线虫并无明显毒性减弱现象；添加外源精氨酸导致菌株对线虫的减毒效果更加明显，且菌株产铁载体能力显著下降。【结论】东湖假单胞菌HYS中AST途径可以通过影响菌株铁载体合成来影响其对秀丽隐杆线虫的毒性，本研究为深入了解假单胞菌精氨酸代谢和致病性机制提供了新依据。

Abstract:

[Background] Pseudomonas donghuensis HYS is a high-siderophore-yielding strain isolated and identified by our laboratory from the Wuhan East Lake. HYS has strong toxicity to Caenorhabditis elegans. Previous studies have found that insertions in the argS gene in HYS can significantly reduce its toxicity to C. elegans. [Objective] To explore the function of argS gene and how it affects bacterial toxicity, and to provide a theoretical basis for further study on the mechanism of HYS toxicity. [Methods] The biological function of argS gene and the relationship between the arginine succinyltransferase (AST) pathway and bacterial toxicity were investigated with bioinformatics comparison, genetic analysis, physiological assays, and biochemical experiments. [Results] The argS gene in HYS encodes a protein that shares 88% similarity to the arginine succinyltransferase β subunit in Pseudomonas aeruginosa. The deletion of argS prevented the HYS strain from using arginine as the sole carbon source for growth. Deletion strains of key genes in arginine decarboxylase (ADC) pathway, arginine dehydrogenase (ADH) pathway and arginine deiminase (ADI) pathway could normally use arginine as the sole carbon source and had no significantly attenuated effect on the toxicity to C. elegans. Added arginine to these strains significantly attenuates the toxicity, and decreases their ability in producing siderophores. [Conclusion] AST pathway in Pseudomonas donghuensis HYS can affect the toxicity to C. elegans by affecting the synthesis of the siderophores. This study lays the foundation for further studies into the mechanism of arginine metabolism and pathogenicity in Pseudomonas.