Functional analysis of Shigella VirG domains essential for interaction with vinculin and actin-based motility

被引:77
作者
Suzuki, T
Saga, S
Sasakawa, C
机构
[1] UNIV TOKYO,INST MED SCI,DEPT BACTERIOL,MINATO KU,TOKYO 108,JAPAN
[2] AICHI HUMAN SERV CTR,INST DEV RES,DEPT MORPHOL,KASUGAI,AICHI 48003,JAPAN
关键词
D O I
10.1074/jbc.271.36.21878
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The VirG (IcsA) protein of Shigella is required for recruitment of host actin filament (F-actin) by intracellularly motile bacteria. An N-terminal 80-kDa VirG portion (alpha-domain) is exposed on the bacterial surface, while the following C-terminal 37-kDa portion (beta-core) is embedded in the outer membrane. Here, we report that the surface exposed alpha-domain of VirG possesses two distinct functional domains; one is the N-terminal two-thirds portion of the alpha-domain which is required for eliciting F-actin assembly on the bacteria in infected cells, and the other one is the rest of the C-terminal portion of the VirG alpha-domain, which is essential for the asymmetric distribution of VirG on the bacterial surface, Furthermore, we found that vinculin, an actin-binding cytoskeletal protein, accumulates on the surface of bacteria expressing VirG in infected cells, and that the distribution of vinculin coincided with the distribution of VirG and assembled F-actin. The vinculin accumulation depended on the expression of the alpha-domain VirG portion required for F-actin assembly, but the recruitment of vinculin on Shigella appeared prior to the appearance of F-actin in the infected cells. Analysis of proteins interacting with VirG using Xenopus laevis eggs extracts revealed that vinculin was a protein that bound to the alpha-domain portion. This was further confirmed using purified chicken gizzard vinculin, in that the 95-kDa vinculin head part, but not the 30-kDa tail part, directly bound to the alpha-domain portion. These results suggest a possible role for vinculin in recruitment of F-actin to the VirG moiety exposed on Shigella in infected mammalian cells.
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页码:21878 / 21885
页数:8
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