Type III secretion decreases bacterial and host survival following phagocytosis of Yersinia pseudotuberculosis by macrophages.
Yersinia pseudotuberculosis uses a plasmid (pYV)-encoded type III secretion system (T3SS) to translocate a set of effectors called Yops into infected host cells. YopJ functions to induce apoptosis, and YopT, YopE, and YopH act to antagonize phagocytosis in macrophages. Because Yops do not completely block phagocytosis and Y. pseudotuberculosis can replicate in macrophages, it is important to determine if the T3SS modulates host responses to intracellular bacteria. Isogenic pYV-cured, pYV(+) wild-type, and yop mutant Y. pseudotuberculosis strains were allowed to infect bone marrow-derived murine macrophages at a low multiplicity of infection under conditions in which the survival of extracellular bacteria was prevented. Phagocytosis, the intracellular survival of the bacteria, and the apoptosis of the infected macrophages were analyzed. Forty percent of cell-associated wild-type bacteria were intracellular after a 20-min infection, allowing the study of the macrophage response to internalized pYV(+) Y. pseudotuberculosis. Interestingly, macrophages restricted survival of pYV(+) but not pYV-cured or DeltayopB Y. pseudotuberculosis within phagosomes: only a small fraction of the pYV(+) bacteria internalized replicated by 24 h. In addition, approximately 20% of macrophages infected with wild-type pYV(+) Y. pseudotuberculosis died of apoptosis after 20 h. Analysis of yop mutants expressing catalytically inactive effectors revealed that YopJ was important for apoptosis, while a role for YopE, YopH, and YopT in modulating macrophage responses to intracellular bacteria could not be identified. Apoptosis was reduced in Toll-like receptor 4-deficient macrophages, indicating that cell death required signaling through this receptor. Treatment of macrophages harboring intracellular pYV(+) Y. pseudotuberculosis with chloramphenicol reduced apoptosis, indicating that the de novo bacterial protein synthesis was necessary for cell death. Our finding that the presence of a functional T3SS impacts the survival of both bacterium and host following phagocytosis of Y. pseudotuberculosis suggests new roles for the T3SS in Yersinia pathogenesis.
Zhang
Yue
Y
Department of Molecular Genetics and Microbiology, Center for Infectious Diseases, SUNY Stony Brook, Stony Brook, New York 11794-5222, USA. yzhang@ms.cc.sunysb.edu
Murtha
James
J
Roberts
Margaret A
MA
Siegel
Richard M
RM
Bliska
James B
JB
D000818:Animals; D017209:Apoptosis; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics / Q000502:physiology; D001426:Bacterial Proteins / Q000235:genetics / Q000502:physiology; D003546:Cysteine Endopeptidases / Q000235:genetics / Q000502:physiology; D017353:Gene Deletion; D008264:Macrophages / Q000276:immunology* / Q000382:microbiology* / Q000648:ultrastructure; D051379:Mice; D050296:Microbial Viability; D046529:Microscopy, Electron, Transmission; D010587:Phagocytosis; D010588:Phagosomes / Q000382:microbiology; D010957:Plasmids; D021381:Protein Transport; D017027:Protein Tyrosine Phosphatases / Q000235:genetics / Q000502:physiology; D051197:Toll-Like Receptor 4 / Q000172:deficiency / Q000276:immunology; D037521:Virulence Factors / Q000235:genetics; D015011:Yersinia pseudotuberculosis / Q000276:immunology* / Q000378:metabolism* / Q000648:ultrastructure
D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D051197:Toll-Like Receptor 4; D037521:Virulence Factors; C109227:YopP protein, Yersinia; C116117:YopT protein, Yersinia; C063239:yopE protein, Yersinia; D017027:Protein Tyrosine Phosphatases; C067571:yopH protein, Yersinia; D003546:Cysteine Endopeptidases