转录因子SlyA调控腐败希瓦氏菌生物被膜形成的分子机制研究

腐败希瓦氏菌在环境修复和微生物燃料电池等领域具有广泛应用，生物被膜的形成能提高其环境修复效率和产电性能。同时腐败希瓦氏菌是常见的条件致病菌和食品腐败菌，生物被膜会使其难于被清除，严重威胁人类健康和食品安全。然而目前对腐败希瓦氏菌形成生物被膜的研究还处于起步阶段。在前期工作中，我们首次发现核心转录因子SlyA通过调控胞内c-di-GMP的含量显著抑制生物被膜的形成，但SlyA参与的具体的信号转导通路尚不清楚。本研究拟通过ChIP-seq，EMSA技术寻找SlyA调控的下游靶基因；利用5′-RACE和DNase I足迹法分析SlyA对靶基因的调控模式；通过DNA亲和捕获和启动子探测技术寻找调控slyA转录的上游调控因子；同时研究SlyA与c-di-GMP之间的相互调控关系，进而揭示SlyA调控腐败希瓦氏菌形成生物被膜的分子机制，为其生物被膜的充分利用和解决其生物被膜引起的危害奠定理论基础

Shewanella putrefaciens has been widely applied in environmental remediation and microbial fuel cell, and the biofilm formation can improve the efficiency of environmental remediation and power generation. Moreover, Shewanella putrefaciens is a common opportunistic pathogen and food spoilage bacteria, and biofilm make it hard to be cleaned up, which seriously threaten human health and food safety. However, the study of biofilm formation by Shewanella putrefaciens is still in its infancy. In our previous study, we found for the first time that core transcriptional factor SlyA significantly inhibits biofilm formation by regulating intracellular c-di-GMP level. However, the specific signal transduction pathways involved by SlyA remain unclear. This proposal aims to look for the downstream target genes regulated by SlyA through the methods of ChIP-seq and EMSA, to analyze regulatory patterns of SlyA on target genes by DNase I footprinting and 5′-RACE, to look for the upstream regulators that modulate the transcription of SlyA by DNA affinity capture assay and transposon mutagenesis technique, and to investigate the regulatory relationship between SlyA and c-di-GMP. This proposal can promote the studies on molecular mechanism of Shewanella putrefaciens biofilm formation and to establish theoretical basis for controlling biofilm formation.