sRNA在红球菌P14降解多环芳烃中的调控功能研究

Microbial degradation is usually considered as the main pathway of bioremediation of the polycyclicaromatic hydrocarbons (PAHs) in the environment. sRNA, as a post-transcriptional regulator which plays an important role in various metabolic pathways and physiological activities in microorganisms, has become a hot scientific topic in the current world, however, the function of sRNAs in the degradation of PAHs is still unknown. Rhodococcus sp. P14 is a bacterium from ocean with a strong ability of degradation of many PAHs. The genome and genes with functions in PAHs degradation were studied previously. In this project, Rhodococcus sp 14 is used as the object. This project intends to predict and identify sRNAs with the potential functions in the degradation of pyrene which is a kind of PAHs compounds through the use of dRNA-seq as well as Northern Blot. Moreover, qPCR and EMSA are used to clarify the functions of sRNAs on their target genes which are related to the pyrene degradation. Finally, this project intends to illustrate the role of sRNAs in Rhodococcus sp. P14 during PAHs degradation. The results will help to reveal the molecular mechanism of the degradation of PAHs.

微生物降解是环境中多环芳烃生物修复的主要途径。微生物中，sRNA作为一种可在各种代谢途径及生理活动中起重要作用的转录后调控因子，近年来成为国际上的研究热点，但对sRNA在多环芳烃降解中的调控功能还未见报道。红球菌P14是一种可高效降解多种多环芳烃的海洋细菌，前期课题组对其基因组及多环芳烃降解相关基因进行了深入研究。本项目以红球菌P14为研究对象，利用dRNA-seq和Northern Blot等技术，预测并鉴定与一种多环芳烃化合物芘降解相关的sRNA，进一步通过qPCR与EMSA等技术，明确sRNA对芘降解相关靶基因的调控作用，从而阐明sRNA在红球菌P14多环芳烃降解中的调控功能。预期成果可为阐明多环芳烃降解机制提供理论基础。

细菌在多环芳烃污染生物修复中具有重要作用。sRNA作为一种转录后调控因子，在细菌各种代谢途径及生理活动中起重要调控功能，但对sRNA在细菌降解多环芳烃中的调控功能还未知。本项目以多环芳烃高效降解菌株红球菌P14为研究对象，利用生物信息学手段，通过分析红球菌P14多环芳烃降解过程的转录组学数据，鉴定了在多环芳烃降解中表达量具有显著变化的12个sRNA，其具有调控多环芳烃降解的潜力。同时，发现在红球菌P14降解多环芳烃中,共有108个相关基因表达量显著提高，通过预测发现其中18个基因为sRNA调控的靶基因,其中双加氧酶BaaA的表达可能受predsRNA57的调控。通过基因克隆异源表达及酶学分析手段，发现BaaA具有羟化蒽及苯并蒽的能力，表明其在多环芳烃降解中具有重要功能，PredsRNA57可通过调控靶基因BaaA的表达，从而参与多环芳烃降解的调控。另外，本项目对红球菌P14基因组进行分析结合多环芳烃降解中间产物检测，在红球菌P14构建了多环芳烃降解途径，结果表明多环芳烃可降解生成琥珀先乙酰辅酶A及乙酰辅酶A，进入三羧酸循环生成二氧化碳及能量。除上述研究成果外，本项目研究发现红球菌P14中NarL调控蛋白具有抑制功能基因单加氧酶CYP108j1的表达能力及受ArsR1调控的双加氧酶PidY在多环芳烃降解中具有重要功能。上述成果为阐明细菌降解多环芳烃机制提供理论基础，也为红球菌P14在环境多环芳烃生物修复中的应用提供新的思路。