Phenotypic characterization of a virulence-associated protein, VagH, of Yersinia pseudotuberculosis reveals a tight link between VagH and the type III secretion system.
Recently, a number of attenuated mutants of Yersinia pseudotuberculosis have been identified using a bioinformatics approach. One of the target genes identified in that study was vagH, which the authors now characterized further. VagH shows homology to HemK of Escherichia coli, possessing methyltransferase activity similar to that of HemK, and targeting release factors 1 and 2. Microarray studies comparing the wild-type and the vagH mutant revealed that the mRNA levels of only a few genes were altered in the mutant. By proteome analysis, expression of the virulence determinant YopD was found to be increased, indicating a possible connection between VagH and the virulence plasmid-encoded type III secretion system (T3SS). Further analysis showed that Yop expression and secretion were repressed in a vagH mutant. This phenotype could be suppressed by trans-complementation with the wild-type vagH gene or by deletion of the negative regulator yopD. Also, in a similar manner to a T3SS-negative mutant, the avirulent vagH mutant was rapidly cleared from Peyer's patches and could not reach the spleen after oral infection of mice. In a manner analogous to that of T3SS mutants, the vagH mutant could not block phagocytosis by macrophages. However, a vagH mutant showed no defects in the T3SS-independent ability to proliferate intracellularly and replicated to levels similar to those of the wild-type in macrophages. In conclusion, the vagH mutant exhibits a virulence phenotype similar to that of a T3SS-negative mutant, indicating a tight link between VagH and type III secretion in Y. pseudotuberculosis.
Garbom
Sara
S
Department of Molecular Biology, Umeå University, S-90187 Umeå, Sweden.
Olofsson
Martina
M
Department of Molecular Biology, Umeå University, S-90187 Umeå, Sweden.
Björnfot
Ann-Catrin
AC
Department of Molecular Biology, Umeå University, S-90187 Umeå, Sweden.
Srivastava
Manoj Kumar
MK
Department of Forest Genetics and Plant Physiology, Umeå Plant Science Centre, S-90183 Umeå, Sweden.
Robinson
Victoria L
VL
Biomedical Sciences, Dstl Porton Down, Salisbury, Wiltshire SP4 0JQ, UK.
Oyston
Petra C F
PCF
Biomedical Sciences, Dstl Porton Down, Salisbury, Wiltshire SP4 0JQ, UK.
Titball
Richard W
RW
Biomedical Sciences, Dstl Porton Down, Salisbury, Wiltshire SP4 0JQ, UK.
Wolf-Watz
Hans
H
Department of Molecular Biology, Umeå University, S-90187 Umeå, Sweden.
D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D015169:Colony Count, Microbial; D004195:Disease Models, Animal; D029968:Escherichia coli Proteins / Q000235:genetics; D005260:Female; D017353:Gene Deletion; D020869:Gene Expression Profiling; D008264:Macrophages / Q000382:microbiology; D008780:Methyltransferases / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D051379:Mice; D008810:Mice, Inbred C57BL; D020411:Oligonucleotide Array Sequence Analysis; D010454:Peptide Termination Factors / Q000378:metabolism; D011496:Protein Methyltransferases / Q000235:genetics; D021381:Protein Transport; D020543:Proteome / Q000032:analysis; D012329:RNA, Bacterial / Q000032:analysis / Q000235:genetics; D012333:RNA, Messenger / Q000032:analysis / Q000235:genetics; D037521:Virulence Factors / Q000235:genetics / Q000378:metabolism*; D015011:Yersinia pseudotuberculosis / Q000378:metabolism*; D015012:Yersinia pseudotuberculosis Infections / Q000382:microbiology
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D010454:Peptide Termination Factors; D020543:Proteome; D012329:RNA, Bacterial; D012333:RNA, Messenger; D037521:Virulence Factors; C041460:peptide chain termination release factor 2; D008780:Methyltransferases; D011496:Protein Methyltransferases; C092404:prmC protein, E coli