Targeted inactivation of the hrcA repressor gene in cyanobacteria

To determine if the CIRCE/HrcA system operates in cyanobacteria, we have inactivated the hrcA repressor gene in Synechocystis sp. PCC 6803 by gene targeting. In the hrcA mutant, the groESL1 operon and the groEL2 gene, both of which have the CIRCE operator in their upstream regions, were derepressed at 30°C without affecting expression of other major heat-shock genes. However, expression of these groE genes in the mutant was not fully derepressed. Their transcription increased further upon heat shock, and was initiated from the same sites as those used under normal conditions. This suggests that their expression is regulated by at least two different mechanisms, a negative one controlled by HrcA and an unknown positive one. The heat-induced expression of clpB1 and htpG was greatly repressed by the absence of HrcA. The hrcA mutant which constitutively overexpressed GroEL displayed improved cellular thermotolerance and also reduced photobleaching of phycocyanin under heat stress conditions.