The CtsR regulator of stress response is active as a dimer and specifically degraded in vivo at 37 degrees C

CtsR ((c) under bar lass (t) under bar hree (s) under bar tress gene (r) un der bar epressor) negatively regulates the expression of class III heat sho ck genes (clpP, clpE and the clpC operon) by binding to a directly repeated heptanucleotide operator sequence (A/GGTCAAA NAN A/GGTCAAA). CtsR-dependen t genes are expressed at a low level at 37 degreesC and are strongly induce d under heat shock conditions. We performed a structure/function analysis o f the CtsR protein, which is highly conserved among low G+C Gram-positive b acteria. Random chemical mutagenesis, in vitro cross-linking, in vivo co-ex pression of wild-type and mutant forms of CtsR and the construction of chim eric proteins with the DNA-binding domain of the lambda CI repressor allowe d us to identify three different functional domains within CtsR: a helix-tu rn-helix DNA-binding domain, a dimerization domain and a putative heat-sens ing domain. We provide evidence suggesting that CtsR is active as a dimer. Transcriptional analysis of a clpP'-bgaB fusion and/or Western blotting exp eriments using antibodies directed against the CtsR protein indicate that C lpP and ClpX are involved in CtsR degradation at 37 degreesC. This in turn leads to a low steady-state level of CtsR within the cell, as CtsR negative ly autoregulates its own synthesis. This is the first example of degradatio n of a repressor of stress response genes by the Clp ATP-dependent protease .