YopK of Yersinia pseudotuberculosis controls translocation of Yop effectors across the eukaryotic cell membrane.
Introduction of anti-host factors into eukaryotic cells by extracellular bacteria is a strategy evolved by several Gram-negative pathogens. In these pathogens, the transport of virulence proteins across the bacterial membranes is governed by closely related type III secretion systems. For pathogenic Yersinia, the protein transport across the eukaryotic cell membrane occurs by a polarized mechanism requiring two secreted proteins, YopB and YopD. YopB was recently shown to induce the formation of a pore in the eukaryotic cell membrane, and through this pore, translocation of Yop effectors is believed to occur (Håkansson et al., 1996b). We have previously shown that YopK of Yersinia pseudotuberculosis is required for the development of a systemic infection in mice. Here, we have analysed the role of YopK in the virulence process in more detail. A yopK-mutant strain was found to induce a more rapid YopE-mediated cytotoxic response in HeLa cells as well as in MDCK-1 cells compared to the wild-type strain. We found that this was the result of a cell-contact-dependent increase in translocation of YopE into HeLa cells. In contrast, overexpression of YopK resulted in impaired translocation. In addition, we found that YopK also influenced the YopB-dependent lytic effect on sheep erythrocytes as well as on HeLa cells. A yopK-mutant strain showed a higher lytic activity and the induced pore was larger compared to the corresponding wild-type strain, whereas a strain overexpressing YopK reduced the lytic activity and the apparent pore size was smaller. The secreted YopK protein was found not to be translocated but, similar to YopB, localized to cell-associated bacteria during infection of HeLa cells. Based on these results, we propose a model where YopK controls the translocation of Yop effectors into eukaryotic cells.
Holmström
A
A
Department of Microbiology, National Defence Research Establishment, Umeå, Sweden. aholmstrom@ume.foa.se
Petterson
J
J
Rosqvist
R
R
Håkansson
S
S
Tafazoli
F
F
Fällman
M
M
Magnusson
K E
KE
Wolf-Watz
H
H
Forsberg
A
A
D000818:Animals; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D004285:Dogs; D005057:Eukaryotic Cells / Q000378:metabolism; D015870:Gene Expression; D006367:HeLa Cells; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D017027:Protein Tyrosine Phosphatases / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D015011:Yersinia pseudotuberculosis / Q000235:genetics / Q000378:metabolism*
D001425:Bacterial Outer Membrane Proteins; D011993:Recombinant Fusion Proteins; C078909:YopB protein, Yersinia; C063239:yopE protein, Yersinia; D017027:Protein Tyrosine Phosphatases; C067571:yopH protein, Yersinia