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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.
Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology). (Wolf-Watz)
Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology). (Wolf-Watz)
Department of Forest Genetics and Plant Physiology, Umeå Plant Science Centre, S-90183 Umeå, Sweden.
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2007 (English)In: Microbiology, ISSN 1350-0872, E-ISSN 1465-2080, Vol. 153, no Pt 5, 1464-73 p.Article in journal (Refereed) Published
Abstract [en]

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.

Place, publisher, year, edition, pages
Society for General Microbiology , 2007. Vol. 153, no Pt 5, 1464-73 p.
URN: urn:nbn:se:umu:diva-16675DOI: 10.1099/mic.0.2006/000323-0PubMedID: 17464060OAI: diva2:156348
Available from: 2007-10-08 Created: 2007-10-08 Last updated: 2011-04-19Bibliographically approved
In thesis
1. Meticulous control of the T3SS of Yersinia is essential for full virulence
Open this publication in new window or tab >>Meticulous control of the T3SS of Yersinia is essential for full virulence
2011 (English)Doctoral thesis, comprehensive summary (Other academic)
Alternative title[sv]
Minutiös kontroll av Yersinias T3SS är essentiellt för fullständig virulens
Abstract [en]

The type III secretion system (T3SS) of pathogenic Yersinia pseudotuberculosis is involved in virulence. The syringe-like secretion system spans both bacterial membranes and is responsible for the ability of Yersinia to transfer toxic proteins (Yop proteins) into the eukaryotic target cell. The T3SS is believed to have evolved from the flagellum and regulation of the T3SS is a complex event that involves a series of regulatory proteins, whereby two are YscP and YscU. In a regulatory model, called the substrate specificity switch, both proteins act together to ensure proper T3SS structure and function by regulating a stop in YscF needle protein export with a shift to Yop effector secretion. YscU undergoes autoproteolysis at a conserved motif consisting of amino acids Asparagine-Proline-Threonine-Histidine (NPTH). Processing generates a C-terminal 10 kDa peptide, YscUCC. Processing is crucial for proper T3SS regulation and function both in vitro and in vivo. Full-length YscU does not support Yop secretion and after cleavage, YscUCC remains attached to the rest of YscU and acts as a negative block on T3S. Relief of this negative block is suggested to occur through displacement of YscUCC from the rest of YscU. Thorough control of many different cellular processes is brought by the heat shock proteins (HSPs) DnaK and DnaJ. Due to their multiple regulatory functions, mutations in the hsp-genes lead to pleiotropic effects. DnaK and DnaJ are essential for proper flagellum driven motion of bacteria, but more so; they ensure proper Yersinia T3SS function in vivo. Furthermore, DnaJ interacts with YscU and may be directly involved in T3SS regulation. Virulence of Yersinia is regulated on many levels. A previously identified virulence associated protein, VagH, is now characterized as an S-adenosyl-methionine dependent methyltransferase. The targets of the methylation activity of VagH are release factors 1 and 2 (RF1 and RF2), that are important for translation termination. The enzymatic activity of VagH is important for Yop secretion and a vagH mutant up-regulates a T3SS negative regulatory protein, YopD. Furthermore, a vagH mutant is avirulent in a mouse infection model, but is not affected in macrophage intracellular survival. The importance of VagH in vivo makes it a possible target for novel antimicrobial therapy.

Place, publisher, year, edition, pages
Umeå: Institutionen för molekylärbiologi, Teknisk-naturvetenskaplig fakultet, Umeå universitet, 2011. 76 p.
Yersinia, type III secretion, YscU, substrate specificity switch, heat shock proteins, VagH, methyltransferase, virulence
National Category
Biochemistry and Molecular Biology
Research subject
Molecular Biology
urn:nbn:se:umu:diva-42577 (URN)978-91-7459-168-2 (ISBN)
Public defence
2011-05-13, Naturvetarhuset, N320, Umeå universitet, Umeå, 10:00 (English)
Available from: 2011-04-19 Created: 2011-04-11 Last updated: 2011-04-19Bibliographically approved

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