DeBord Kristin L, Galanopoulos Nicholas S, Schneewind Olaf
Committee on Microbiology, University of Chicago, Chicago, Illinois 60637, USA.
J Bacteriol. 2003 Jun;185(12):3499-507. doi: 10.1128/JB.185.12.3499-3507.2003.
Pathogenic Yersinia species use a virulence-plasmid encoded type III secretion pathway to escape the innate immune response and to establish infections in lymphoid tissues. At least 22 secretion machinery components are required for type III transport of 14 different Yop proteins, and 10 regulatory factors are responsible for activating this pathway in response to environmental signals. Although the genes for these products are located on the 70-kb virulence plasmid of Yersinia, this extrachromosomal element does not appear to harbor genes that provide for the sensing of environmental signals, such as calcium-, glutamate-, or serum-sensing proteins. To identify such genes, we screened transposon insertion mutants of Y. enterocolitica W22703 for defects in type III secretion and identified ttsA, a chromosomal gene encoding a polytopic membrane protein. ttsA mutant yersiniae synthesize reduced amounts of Yops and display a defect in low-calcium-induced type III secretion of Yop proteins. ttsA mutants are also severely impaired in bacterial motility, a phenotype which is likely due to the reduced expression of flagellar genes. All of these defects were restored by complementation with plasmid-encoded wild-type ttsA. LcrG is a repressor of the Yersinia type III pathway that is activated by an environmental calcium signal. Mutation of the lcrG gene in a ttsA mutant strain restored the type III secretion of Yop proteins, although the double mutant strain secreted Yops in the presence and absence of calcium, similar to the case for mutants that are defective in lcrG gene function alone. To examine the role of ttsA in the establishment of infection, we measured the bacterial dose required to produce an acute lethal disease following intraperitoneal infection of mice. The ttsA insertion caused a greater-than-3-log-unit reduction in virulence compared to that of the parental strain.
致病性耶尔森氏菌属利用一种毒力质粒编码的III型分泌途径来逃避先天免疫反应，并在淋巴组织中建立感染。III型转运14种不同的Yop蛋白至少需要22种分泌机制成分，10种调节因子负责响应环境信号激活该途径。尽管这些产物的基因位于耶尔森氏菌的70 kb毒力质粒上，但这个染色体外元件似乎不含有提供环境信号感知功能的基因，如钙、谷氨酸或血清感知蛋白。为了鉴定此类基因，我们筛选了小肠结肠炎耶尔森氏菌W22703的转座子插入突变体，以寻找III型分泌缺陷，从而鉴定出ttsA，这是一个编码多跨膜蛋白的染色体基因。ttsA突变的耶尔森氏菌合成的Yop蛋白量减少，并且在低钙诱导的Yop蛋白III型分泌中表现出缺陷。ttsA突变体在细菌运动性方面也严重受损，这种表型可能是由于鞭毛基因表达降低所致。通过用质粒编码的野生型ttsA进行互补，所有这些缺陷都得到了恢复。LcrG是耶尔森氏菌III型途径的一种阻遏物，可被环境钙信号激活。ttsA突变菌株中lcrG基因的突变恢复了Yop蛋白的III型分泌，尽管双突变菌株在有钙和无钙的情况下都分泌Yop蛋白，这与仅lcrG基因功能有缺陷的突变体情况类似。为了研究ttsA在感染建立过程中的作用，我们测量了小鼠腹腔感染后产生急性致死性疾病所需的细菌剂量。与亲本菌株相比，ttsA插入导致毒力降低超过3个对数单位。
