CRISPR-Cas和毒素-抗毒素系统VapBC协同调控结核分枝杆菌抗胁迫的分子机制

适应性免疫系统CRISPR-Cas和毒素-抗毒素(TA)系统是调控细菌生长和存活的两种重要调控系统，尽管关于它们单独调控细菌生存的报道较多，但两个系统协同调控细菌抗胁迫的机制还鲜有报道。申请人前期发现结核分枝杆菌的CRISPR-Cas系统中Cas2与TA系统VapBC22中毒素VapC22蛋白存在直接的物理相互作用，且初步证实Cas2能够抑制VapC22的毒性。本项目拟在分别研究CRISPR-Cas和TA系统调控结核分枝杆菌在不同时期抗饥饿、药物、氧化压力等胁迫生长的基础上，阐明Cas2和毒素VapC22的相互作用对两个调控系统的影响，并系统地揭示Cas2与VapC22互作在结核分枝杆菌协同抗胁迫中的调控作用。研究成果有望揭示细菌CRISPR-Cas和TA系统之间的内在联系，丰富CRISPR-Cas和TA系统的调控功能，发掘结核分枝杆菌胁迫应答新机制，为控制结核感染提供了新的线索和思路。

The CRISPR-Cas, an adaptive immune system, and Toxin-antitoxin systems play important roles on regulating bacterial growth and survival. The separate impacts of these two systems on bacterial growth were showed in many studies. However, the synergy effect of the two systems on regulation of bacterial resistance in stress environment is unknown. In the preliminary study, we found that the Cas2 protein of the CRISPR-Cas system of Mycobacterium tuberculosis (Mtb) directly interacts with the toxin VapC22 of the VapBC22 toxin-antitoxin system in vitro, and further we confirmed that Cas2 inhibits the cytotoxicity of VapC22 and restores the growth of Mtb. On this basis of studying the regulation of the CRISPR-Cas and VapBC22 on the growth of Mtb under the stress of starvation, drugs and oxidative pressure in different periods, we propose to systematically demonstrate the interaction between VapC22 toxin and Cas2 using multiple techniques, as well as the regulatory mode and synergistic effect of Cas2 and VapBC22 on regulation of the survival of Mtb in stress environment. In addition, we will elucidate the synergistic effect of CRISPR-Cas and toxin-antitoxin systems on regulation of the stress-inducible survival of Mtb. In summary, we aim to reveal the intrinsic connection between the CRISPR-Cas and toxin-antitoxin systems and the results may provide new insight into the survival of Mtb in stress environment. Our study will provide new clues and ideas for the ultimate control of TB infection.