Investigation of EscA as a chaperone for the Edwardsiella tarda type III secretion system putative translocon component EseC.
Edwardsiella tarda is an important Gram-negative enteric pathogen affecting both animals and humans. It possesses a type III secretion system (T3SS) essential for pathogenesis. EseB, EseC and EseD have been shown to form a translocon complex after secretion, while EscC functions as a T3SS chaperone for EseB and EseD. In this paper we identify EscA, a protein required for accumulation and proper secretion of another translocon component, EseC. The escA gene is located upstream of eseC and the EscA protein has the characteristics of T3SS chaperones. Cell fractionation experiments indicated that EscA is located in the cytoplasm and on the cytoplasmic membrane. Mutation with in-frame deletion of escA greatly decreased the secretion of EseC, while complementation of escA restored the wild-type secretion phenotype. The stabilization and accumulation of EseC in the cytoplasm were also affected in the absence of EscA. Mutation of escA did not affect the transcription of eseC but reduced the accumulation level of EseC as measured by using an EseC-LacZ fusion protein in Ed. tarda. Co-purification and co-immunoprecipitation studies demonstrated a specific interaction between EscA and EseC. Further analysis showed that residues 31-137 of EseC are required for EseC-EscA interaction. Mutation of EseC residues 31-137 reduced the secretion and accumulation of EseC in Ed. tarda. Finally, infection experiments showed that mutations of EscA and residues 31-137 of EseC increased the LD(50) by approximately 10-fold in blue gourami fish. These results indicated that EscA functions as a specific chaperone for EseC and contributes to the virulence of Ed. tarda.
Wang
Bo
B
Graduate University of Chinese Academy of Sciences, Beijing 100049, PR China.|Institute of Oceanology, Chinese Academy of Sciences, Qingdao 266071, PR China.
Mo
Zhao Lan
ZL
Institute of Oceanology, Chinese Academy of Sciences, Qingdao 266071, PR China.
Mao
Yun Xiang
YX
Ocean University of China, Qingdao 266003, PR China.
Zou
Yu Xia
YX
Institute of Oceanology, Chinese Academy of Sciences, Qingdao 266071, PR China.
Xiao
Peng
P
Graduate University of Chinese Academy of Sciences, Beijing 100049, PR China.|Institute of Oceanology, Chinese Academy of Sciences, Qingdao 266071, PR China.
Li
Jie
J
Ocean University of China, Qingdao 266003, PR China.
Yang
Jia Yin
JY
Graduate University of Chinese Academy of Sciences, Beijing 100049, PR China.|Institute of Oceanology, Chinese Academy of Sciences, Qingdao 266071, PR China.
Ye
Xu Hong
XH
Ocean University of China, Qingdao 266003, PR China.
Leung
Ka Yin
KY
Department of Biological Sciences, Faculty of Science, National University of Singapore, 117543 Singapore.
Zhang
Pei Jun
PJ
Institute of Oceanology, Chinese Academy of Sciences, Qingdao 266071, PR China.