The HspR protein functions as a negative regulator of chaperone and protease gene expression in a diversity of bacteria. Here we have identified, cloned and deleted the Deinococcus radiodurans HspR homologue, DR0934. Delta hspR mutants exhibit moderate growth defects when shifted to mild heat shock temperatures, but are severely impaired for survival at 48 degrees C. Using quantitative reverse transcription polymerase chain reaction and global transcriptional analysis, we have identified 14 genes that are derepressed in the absence of stress in the delta hspR background, 11 of which encode predicted chaperones and proteases, including dnaKJgrpE, ftsH, lonB, hsp20 and clpB. Promoter mapping indicated that the transcription of these genes initiates from a promoter bearing a sigma70-type consensus, and that putative HspR binding sites (HAIR) were present in the 5'-untranslated regions. Electrophoretic mobility shift assays indicated that HspR binds to these promoters at the HAIR site in vitro. These results strongly suggest that DR0934 encodes the HspR-like global negative regulator of D. radiodurans that directly represses chaperone and protease gene expression by binding to the HAIR site in close proximity to promoter regions.