Involvement of the heat shock proteins DnaK/DnaJ in Yersinia T3S
(English)Manuscript (preprint) (Other academic)
Yersinia pseudotuberculosis uses a type III secretion system (T3SS) to secrete and deliver effectors called Yops into target cells. These processes are highly regulated and the pathogen senses cell contact and respond accordingly by inducing Yop-effector expression. A key component of the T3SS is the YscF needle present on the surface of the pathogen. It has been suggested that the bacterium can switch from needle export to Yop expression and secretion and that this substrate switch is important for proper regulation during infection. YscU is an essential protein regulating the substrate switch and autoproteolysis of YscU is essential for accurate T3SS regulation. To study regulation of Yop translocation in more detail, we generated mutants defective for expression of the heat shock proteins (HSPs) DnaJ and DnaK, since earlier studies had indicated a role of these proteins in regulation of effector translocation in Salmonella. The dnaJ mutant and the double dnaK/J mutant showed significant defects in Yop translocation, but surprisingly both mutants were able to secrete Yops in vitro much like the wild type. However, both mutants showed a changed export pattern of the YscF needle with a pronounced increased export of the YscF needle protein after incubation in calcium containing media. This phenotype was linked to defects in YscU autoproteolysis and in this respect the hsp-mutants were identical to earlier identified autoprocessing defective mutants in YscU (Single amino acid exchange mutants N263A and P264A). The hsp-mutants and the processing mutants accumulated full-length YscU, which surprisingly was associated with the outer membrane, while the processed form of YscU was found in the inner membrane fraction. The dnaJ and dnaK/J mutants were strongly affected in YscU autoproteolysis, which indicates a possible direct role for DnaJ in this process. Indeed a specific interaction between YscU and DnaJ could be found suggesting a direct role of the HSPs in regulation of the substrate switch in the T3SS.
Biochemistry and Molecular Biology
Research subject Molecular Biology
IdentifiersURN: urn:nbn:se:umu:diva-42567OAI: oai:DiVA.org:umu-42567DiVA: diva2:409757