Research objectives and content
Regulation of bacterial responses to stress often involves control of activity and stability of sigma factors. This project aims to dissect such regulatory mechanisms using E. coli sigma32 and sigmaS. Sigma32 controls heat shock genes encoding chaperones and proteases. Heat shock increases stability and activity sigma32 thereby increasing expression of heat shock genes. The DnaK and DnaJ chaperones mediate this regulation by stress-dependent association with sigma32 thereby repressing its activity and controlling its degradation by the protease FtsH. SigmaS controls general stress genes which protect cells from starvation damage. Starvation prevents sigmaS degradation by the ClpXP protease in a DnaK-dependent process. For both sigma factors genetic evidence suggests that control of their stability and activity requires short segments within the sigma32 and sigmaS polypeptides that are in similar relative position. Using genetic and biochemical tools it will be investigated whether these segments represent regulatory modules. It will then be attempted to identify residues which are essential for protease and ligand (DnaK, DnaJ) recognition. Identification of the recognition principles are relevant to the understanding of other stress regulations involving similar control mechanisms of sigma factors (e.g. SigB in B. subtilis) and eucaryotic transcription factors (e. g. . heat shock transcription factor).
Training content (objective, benefit and expected impact)
The planned project is expected to identify molecular principles governing the regulation of two important stress responses in E. coli. The results may serve as a paradigm for understanding other stress responses in bacteria and the heat shock response in eucaryotes. It may also allow to manipulate the content of chaperones in bacteria which is of interest to biotechnology since it may allow to increase the yield of soluble recombinant protein. This project could give me the opportunity to acquire novel experimental techniques (e. g. protein purification, analysis of protein interactions, crosslinking, in vitro transcription assays) which are essential for my future scientific career.
Links with industry / industrial relevance (22)
Collaborations of the host laboratory with Boehringer (Mannheim) and Hoffmann-La Roche (Basel) exist, aiming at using chaperones to avoid aggregation of recombinant proteins.