蓝细菌碳氮代谢协同的转录调控机制

Cyanobacteria, as one of the most ancient microbes in the world, have played and are playing very important role in maintaining the proper ratio of O2 and CO2 in the atmosphere. The fine regulation of metabolism homeostasis of two major substance, nitrogen and carbon, make these bacteria a long history and robust living capacity. The homeostasis is mainly achieved by regulating the CO2 concentration mechanism and nitrogen assimilation via four global regulators: CcmR, CmpR, NtcA and NtcB. Interestingly, CcmR, CmpR and NtcB belong to a so-called LysR family of transcriptional factors, despite they possess a rather different effector binding domain. Our systematic investigation on NtcA and preliminary results of CcmR showed that the CO2 concentration mechanism and nitrogen assimilation crosstalk to each other; and moreover, are perfectly coordinated by a couple transcriptional factors which sense the intracellular concentration of metabolites such as 2-OG and 2-PG. In this project, we will apply the two model organisms of cyanobacteria, Synechocystis sp. PCC 6803 and Anabaena sp. PCC 7120 to systematically explore the 3D structure, molecular/biophysical function and mechanism of these three LysR family transcriptional factors. The findings on the effector molecules will enable us to build a direct link between the two key metabolism pathways and expression regulation of corresponding genes. These findings will not only help illustrating the molecular mechanism of cyanobacterial carbon/nitrogen metabolism homeostasis, and also guide the genetic engineering of economic plants.

蓝细菌的悠久历史和顽强生命力得益于利用碳浓缩机制和氮同化两大途径来实现碳氮代谢平衡的精密调控。这两大蓝细菌特有的代谢途径的转录调控主要由包括CcmR、CmpR、NtcA和NtcB等转录因子来完成。而且NtcB、cmpR和ccmR都是属于LysR家族的转录因子，只是它们感应不同的效应子。我们的前期结果表明，通过感应代谢中间物2-磷酸乙醇酸和α-酮戊二酸，蓝细菌完美实现了碳氮代谢的偶联和协同调控。我们将以集胞藻Synechocystis sp. PCC 6803和丝状蓝细菌Anabaena sp. PCC 7120为研究对象，深入研究NtcB、CcmR和CmpR的结构和功能，以提炼LysR家族转录因子的共同特征和作用机理。效应分子和靶基因表达谱的研究将建立关键代谢途径与基因表达调控之间的直接联系，不仅有助于阐明蓝细菌碳氮代谢平衡转录调控的分子机制，而且可以指导经济植物的抗逆和增产的品种改良。

蓝细菌的碳氮平衡由多个转录因子的协同调控完成，包括NdhR、CmpR、NtcA和NtcB等。本项目顺利完成了计划书的任务目标，取得的主要研究成果如下：1) 通过结构生物学、生化和遗传学等手段，阐明了NdhR通过结合不同的代谢小分子(2-PG和2-OG)，快速响应环境中的碳氮比例变化，协同调控碳氮代谢的分子机制 (PNAS 2018)。2) 基于NtcA和NtcB同时结合RNA聚合酶的转录激活复合物NtcB-NtcA-TAC三维结构分析，结合生化实验阐明了双转录因子顺次协同激活碳代谢转录的调控机理(论文整理中)。3) 阐明分子伴侣协助光合作用固碳酶RuBisCO组装和成熟的分子机理，发现RuBisCO组装和成熟过程的多层次精细动态调控网络，加深了对于RuBisCO组装和功能的认识，为RuBisCO的工程改造并应用于光合固碳效率提升奠定了理论基础 (Nature Plants 2020)。