Initiation of conjugative transfer of IncC conjugative plasmids and associated genomic islands circulating in Gammaproteobacteria

启动 IncC 接合质粒和在 Gammaproteobacteria 中循环的相关基因组岛的接合转移

• 批准号: RGPIN-2021-02814
• 负责人: Burrus, Vincent
• 金额: $ 3.64万
• 依托单位: Université de Sherbrooke
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2022
• 资助国家: 加拿大
• 起止时间: 2022-01-01 至 2023-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=744998
• 关键词: Initiation; conjugative; transfer; IncC; plasmids

Knowledge of the molecular mechanisms that drive the dissemination of antibiotic resistance genes is essential to fight the emergence of multidrug-resistant bacteria. Conjugative plasmids of the incompatibility group C (IncC) are key contributors to the multidrug resistance of many pathogenic species of Enterobacteriaceae and Vibrionaceae. Reports associating IncC plasmids with extended-spectrum ß-lactamases and carbapenemases are unsettlingly rising worldwide in enteric pathogens recovered from the environment, food products, food-producing animals, humans and their pets. IncC plasmids also drive the propagation of multidrug resistance-conferring genomic islands such as the Salmonella genomic island 1 (SGI1) that acts as a parasite of IncC plasmids. IncC plasmids code for a conjugative transfer machinery, also known as a type IV secretion system, that secretes genetic material (DNA) from a donor into a recipient cell. Currently, the molecular mechanisms mediating this intercellular DNA transfer are not well understood. Translocation of DNA is initiated at the origin of transfer (oriT) by the relaxase TraI that belongs to a family of HD phosphodiesterases and requires a MobI auxiliary protein. Conjugative transfer initiated by an HD phosphodiesterase is unusual and suggests a unique and novel mechanism that needs to be revealed. SGI1 interferes with the initiation of transfer of IncC plasmids likely by inhibiting the interaction between TraI and MobI. Furthermore, data mining of bacterial genome databases suggests that IncC plasmids can drive the dissemination of a completely new family of genomic islands distantly related to SGI1 from and to a broad range of bacterial species. The main goal of this proposal is to characterize the mechanism of conjugative transfer initiation of IncC plasmids, and the pathways used by SGI1 and related genomic islands to interfere with this process. More specifically, our objectives are to identify all the proteins involved in DNA processing at oriT and the secretion of DNA into the recipient cells as well as the protein translocation signals that allow recognition and secretion of these proteins through the type IV secretion system encoded by IncC plasmids. We will also determine the mechanisms by which SGI1 interferes with these functions to ensure its own propagation and the destabilization of the IncC plasmids. Finally, we will investigate the role of IncC plasmids in the mobility of putative SGI1-like genomic islands that are widespread in the genome of marine-dwelling microorganisms. This study will yield fundamental insights about the basic biology IncC plasmids and fill important knowledge gaps in their role in promoting transfer of unrelated genomic islands. On the long term, these studies will provide strong foundations for the discovery of new potential molecular targets that could be used for the development of tools aimed at tackling the progression of multidrug resistance.

了解驱动抗生素抗性基因传播的分子机制对于对抗多重耐药细菌的出现至关重要。不相容性 C 群 (IncC) 的接合质粒是肠杆菌科和弧菌科许多致病菌多重耐药性的关键因素。在世界范围内从环境、食品、食用动物、人类及其宠物中回收的肠道病原体中，将 IncC 质粒与超广谱 β-内酰胺酶和碳青霉烯酶相关联的报告令人不安地不断增加。 IncC 质粒还驱动赋予多药耐药性的基因组岛的传播，例如充当 IncC 质粒寄生虫的沙门氏菌基因组岛 1 (SGI1)。 IncC 质粒编码接合转移机制，也称为 IV 型分泌系统，将遗传物质 (DNA) 从供体细胞分泌到受体细胞中。目前，介导这种细胞间 DNA 转移的分子机制尚不清楚。 DNA 易位由松弛酶 TraI 在转移起点 (oriT) 启动，TraI 属于 HD 磷酸二酯酶家族，需要 MobI 辅助蛋白。 HD 磷酸二酯酶引发的接合转移是不寻常的，这表明需要揭示一种独特且新颖的机制。 SGI1 可能通过抑制 TraI 和 MobI 之间的相互作用来干扰 IncC 质粒转移的启动。此外，细菌基因组数据库的数据挖掘表明，IncC 质粒可以驱动与 SGI1 远缘相关的全新基因组岛家族在广泛的细菌物种之间传播。该提案的主要目标是表征 IncC 质粒接合转移起始的机制，以及 SGI1 和相关基因组岛干扰该过程的途径。更具体地说，我们的目标是识别 oriT 的 DNA 加工和 DNA 分泌到受体细胞中所涉及的所有蛋白质，以及允许通过 IncC 质粒编码的 IV 型分泌系统识别和分泌这些蛋白质的蛋白质易位信号。我们还将确定 SGI1 干扰这些功能的机制，以确保其自身增殖和 IncC 质粒的不稳定。最后，我们将研究 IncC 质粒在假定的 SGI1 样基因组岛的移动性中的作用，这些岛广泛存在于海洋微生物的基因组中。这项研究将产生关于基础生物学 IncC 质粒的基本见解，并填补其在促进无关基因组岛转移中的作用的重要知识空白。从长远来看，这些研究将为发现新的潜在分子靶点提供坚实的基础，这些靶点可用于开发旨在应对多药耐药性进展的工具。