Role Of Transcriptional Regulation Protein CarD Of Deinococcus Radiodurans In Response To Heat Shock

Sublethal doses of heat stress induce a cellular response, the heat-shock response, which protects cells and organisms from severe damage, promotes the recovery of normal cellular and physiological activities. In order to face heat stress, the cells of E.coli transiently increase the level of their heat-shock proteins.Inducing the heat-shock regulon by a and a32 provides thermotolerance for E.coli.The heat-shock response of Bacillus subtilis is controlled by dnakandgroESLregulon, aB-dependent stress proteins and other heat-inducible proteases/chaperones by initiating and inhibiting related genes. Furthermore, there is another mechanism, stringent reponse, which regulates in a globle level, this mechanism is not aimed at a particular gene but directly effects on ribosomal proteins and RNA polymerase. As a necessary protein in mycobacterium, CarD regulated the transcription of rRNA by interacting withRNA polymerase. CarD had the same function as DksA in E.coli, it regulated the transcription of rRNA by stringent reponse.Whether there is the similar regulatory mechanism in Deinococcus radiodurans is still unclear.There is the carD deletion mutant in lab. Deinococcus radiodurans and △ card mutant were used to study the regulation mechanism of carD gene on the heat-shock response. The main results are as follows:1. Under normal growth condition, the lack of CarD had no effect onthe growth rate of Deinococcus radiodurans.2. The viability of the mutant strain △ carD was significantlylower than the wild type Rl after heat shock（48℃）for 4 h. There were the most distinct phenotypes after heat shock for 5h.3. The result of real-time PCR proved thatCarDmight have a role in the regulation of heat-shock related genes by inducing the expression of heat shock proteins after heat shock for 1 h.4. Biolog analysis showed thatthe using of carbon source like undine, inosine and methyl pyruvate increased in themutant strain △ carD compared to wild-type D. radioduransRl, while the utilization rate of glutamine-glutamic acid, acetic acid, turanose, sucrose, D-ribose and α-D-glucosesignificantly decreased when strains lacked carD.5. The cell division ofthe mutant strain △carD was asymmetric, most appearred dyad form. After heat shock for 4 h,the cells of △carDwere mostly broken, the rest turn wrinkled and soft, the materialsof the cytoplasm also became loose.6. The protein was about 19.9 KDa, protein expression with chitin tag shows that minority of CarD was in the supernatant, while most of it was in precipitation, it may be related to the nature ofthe protein itself.7. CarD might affect the transcription of gene DR₁314, dnaJl, DR₀326 and DR₀972, but it maybe not a directly-related effect.In the summary, CarD presumably regulated the heat shock response by stringent reponseina global level withoutaiming at a particular gene in Deinococcus radiodurans.