title: Random mutagenesis identifies a C-terminal region of YopD important for Yersinia type III secretion function.
abstract: A common virulence mechanism among bacterial pathogens is the use of specialized secretion systems that deliver virulence proteins through a translocation channel inserted in the host cell membrane. During Yersinia infection, the host recognizes the type III secretion system mounting a pro-inflammatory response. However, soon after they are translocated, the effectors efficiently counteract that response. In this study we sought to identify YopD residues responsible for type III secretion system function. Through random mutagenesis, we identified eight Y. pseudotuberculosis yopD mutants with single amino acid changes affecting various type III secretion functions. Three severely defective mutants had substitutions in residues encompassing a 35 amino acid region (residues 168-203) located between the transmembrane domain and the C-terminal putative coiled-coil region of YopD. These mutations did not affect regulation of the low calcium response or YopB-YopD interaction but markedly inhibited MAPK and NFκB. [corrected] activation. When some of these mutations were introduced into the native yopD gene, defects in effector translocation and pore formation were also observed. We conclude that this newly identified region is important for YopD translocon function. The role of this domain in vivo remains elusive, as amino acid substitutions in that region did not significantly affect virulence of Y. pseudotuberculosis in orogastrically-infected mice.
authors: Clinical Laboratory Science, School of Health, Technology and Management, Stony Brook University, Stony Brook, New York, United States of America; Department of Molecular Genetics and Microbiology, Center for Infectious Diseases, School of Medicine, Stony Brook University, Stony Brook, New York, United States of America.
authors: Rebecca
authors: R
authors: Solomon
authors: Clinical Laboratory Science, School of Health, Technology and Management, Stony Brook University, Stony Brook, New York, United States of America; Department of Molecular Genetics and Microbiology, Center for Infectious Diseases, School of Medicine, Stony Brook University, Stony Brook, New York, United States of America.
authors: Weibing
authors: W
authors: Zhang
authors: Clinical Laboratory Science, School of Health, Technology and Management, Stony Brook University, Stony Brook, New York, United States of America.
authors: Grace
authors: G
authors: McCrann
authors: Department of Molecular Genetics and Microbiology, Center for Infectious Diseases, School of Medicine, Stony Brook University, Stony Brook, New York, United States of America.
authors: James B
authors: JB
authors: Bliska
authors: Clinical Laboratory Science, School of Health, Technology and Management, Stony Brook University, Stony Brook, New York, United States of America; Department of Molecular Genetics and Microbiology, Center for Infectious Diseases, School of Medicine, Stony Brook University, Stony Brook, New York, United States of America.
authors: Gloria I
authors: GI
authors: Viboud
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism* / Q000633:toxicity; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D005260:Female; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D008810:Mice, Inbred C57BL; D020929:Mitogen-Activated Protein Kinase Kinases / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism; D017434:Protein Structure, Tertiary; D014774:Virulence / Q000502:physiology; D015011:Yersinia pseudotuberculosis / Q000378:metabolism*
chemical_list: D001425:Bacterial Outer Membrane Proteins; D016328:NF-kappa B; C078910:YopD protein, Yersinia; C063239:yopE protein, Yersinia; D020929:Mitogen-Activated Protein Kinase Kinases; D002118:Calcium