拜式梭菌Sigma-54转录调控网络的比较基因组学重构与实验鉴定

Clostridium beijerinckii is an important industrial microorganisms with the capability of producing a wealth of bulk chemicals and fuels. However, the complex regulatory network in this organism remains largely unexplored. A gene encoding Sigma-54 was identified in the Clostridium beijerinckii genome. Our preliminary studies have showed that the inactivation of Sigma-54 resulted in serious impact on the solvent production in Clostridium beijerincki, while the internal mechanism of this Sigma-54-dependent control is still unclear. In this study, we will combine bioinformatic skills with experimental technology for the analysis of sigma-54-controlled transcriptional regulation. Based on the identification of EBPs proteins that interact with Sigma-54, we will genomic reconstruct the regulons of Sigma-54 and EBPs in clostridia species. The key targets regulated by the representative master EBPs and Sigma-54 of will be further analyzed to elucidate how they affect solvent synthesis efficiency and ratio, as well as the way for EBPs to receive the signal, and the physiological function of the EBPs domains and their relationships, thus revealing the regulatory mechanism of Sigma-54 controlled solvent-producing in Clostridium beijerinckii. This study will greatly improve our understanding of the metabolic regulation mechanism in industrial clostridium species, which is important for bioengineering applications and achieving the promotion of the solvent-producing ability in clostridia.

拜式梭菌可发酵生成多种大宗化学品及燃料，是一种重要的工业微生物。然而，目前对其复杂的调控网络仍然了解甚少。在拜式梭菌中预测到转录因子Sigma-54。前期工作表明，Sigma-54的缺失严重影响了拜式梭菌的溶剂生成，但是Sigma-54具体的调控靶点和调控机制尚待研究。本项目拟结合生物信息学和实验技术研究Sigma-54的转录调控网络，识别与Sigma-54相互作用的增强子结合蛋白EBPs，在比较基因组学重构Sigma-54 和各个EBPs的调节子（regulons）的基础上，深入解析代表性关键EBPs和Sigma-54的调控靶点及其影响溶剂合成效率和产物比例的作用机制，阐明关键EBPs的信号感受机制、不同结构域的功能以及它们之间的相互关系，从而揭示Sigma-54对于拜式梭菌溶剂生成的调控机制，提高对于拜式梭菌等重要工业梭菌的代谢调控机制的认识，为提高梭菌的溶剂合成能力提供理论基础。

拜式梭菌是一种重要的工业菌，可发酵生成多种大宗化学品及燃料。然而，目前对其复杂的调控网络仍然了解甚少。我们的前期工作发现，Sigma-54的缺失严重影响了拜式梭菌的溶剂生成，但是Sigma-54如何调控，具体的调控靶点和调控机制均有待研究。本项目结合了生物信息学和实验技术研究梭菌Sigma-54的转录调控网络和调控功能。我们利用比较基因组学技术重构了拜氏梭菌在内的57株梭菌的Sigma-54调节单元，发现了上百个新的调控靶点，预测了超20个新的Sigma54依赖的增强子结合蛋白EBPs。研究发现在拜氏梭菌中Sigma54主要调控中心碳代谢，尤其是发酵途径的重要基因。我们鉴定了若干参与乙醇、丁醇、2,3-丁二醇等重要溶剂生成的关键靶基因，同时解析一个关键EBP-AdhR与Sigma-54共同调控溶剂生成的调控作用，并揭示了AdhR作为一种新型细胞氧化还原感受器，通过调控溶剂生成的关键基因的表达来平衡胞内氧化还原势的机制。以上研究结果不仅提高了对于拜式梭菌等重要工业梭菌的代谢调控机制的认识，也为提高梭菌的溶剂合成能力提供了理论基础。