Heat shock in Lactococcus: Regulation and mutants in stress respons

Bacteria respond to stress by induction of specific proteins in order to be able to grow or at least to increase survival. During a temperature upshift, Lactococcus lactis is subjected to a classical heat shock with increased synthesis of the two major chaperone complexes DnaK-GrpE-DnaJ and GroES-GroEL, aswell as ATP-dependent proteases. The main task of the chaperones is to recognise nonnative polypeptides and prevent their aggregation by assisting the correct folding into the native state. The expression of these chaperones are controlled by the negative regulator HrcA wich binds to inverted repeats (CIRCE-elements) located in front of the chaperone genes.
The purpose of the project is by genetical and physiological analysis of a selected set of general stress genes to achieve basic knowledge of the physiological roles of the corresponding proteins during both growth and stress conditions in Lactococcus lactis. Temperature, salt, acid, and survival in stationary phase are the general stress parameters used in the analysis - conditions which are all of industrial importance. The project also includes analysis of the regulatory circuits involved in the stress responses. We aim to isolate a range of mutants with different phenotypes, which are important as tools for the physiological and genetical analyses of stress in Lactococcus. These include 2D-gel analysis, reporter gene fusions to individual stress genes, mRNA analysis by slot blotting, in vitro protein/DNA interactions, and several other molecular biology techniques. Some of the mutants may be used for construction of improved starter cultures.