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Phenotypic characterization of a virulence-associated protein, VagH, of Yersinia pseudotuberculosis reveals a tight link between VagH and the type III secretion system

Martina Olofsson^1,

Manoj Kumar Srivastava^2,

¹ Department of Molecular Biology, Umeå University, S-90187 Umeå, Sweden
² Department of Forest Genetics and Plant Physiology, Umeå Plant Science Centre, S-90183 Umeå, Sweden
³ Biomedical Sciences, Dstl Porton Down, Salisbury, Wiltshire SP4 0JQ, UK

Correspondence
Hans Wolf-Watz
Hans.wolf-watz{at}molbiol.umu.se

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.

Present address: Division of Investigative Science, Department of Metabolic Medicine, Imperial College London, Hammersmith Hospital, Du Cane Road, London W12 0NN, UK.

Present address: Department of Biochemistry, J. C. Bose Institute of Life Sciences, Bundelkhand University, Jhansi (UP) 284 128, India.

The array data discussed in this publication have been deposited in µG@Sbase (accession no. E-BUGS-48; http://bugs.sgul.ac.uk/E-BUGS-48) and also ArrayExpress (accession no. E-BUGS-48). The array design is available in µG@Sbase (accession no. A-BUGS-11; http://bugs.sgul.ac.uk/A-BUGS-11) and also ArrayExpress (accession no. A-BUGS-11).