Mechanisms of propagation of antibiotic resistance-conferring mobile genetic elements in Enterobacteriaceae and Vibrionaceae

肠杆菌科和弧菌科中赋予抗生素抗性的可移动遗传元件的传播机制

• 批准号: RGPIN-2016-04365
• 负责人: Burrus, Vincent
• 金额: $ 3.21万
• 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: 2018
• 资助国家: 加拿大
• 起止时间: 2018-01-01 至 2019-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=646583
• 关键词: Mechanisms; propagation; antibiotic; resistance; conferring

Knowledge of the genetic determinants and molecular mechanisms that drive the mobility of antibiotic resistance genes is essential to fight the emergence and spread of multidrug resistant bacteria. The integrative and conjugative elements of the SXT/R391 family (SRIs) and the conjugative plasmids of the A/C incompatibility group (ACPs), are major vectors of multidrug resistance genes in pathogenic Enterobacteriaceae and Vibrionaceae. SRIs is responsible for antibiotic resistance of the seventh pandemic lineage of Vibrio cholerae, the etiologic agent of cholera, which remains a major cause of mortality and morbidity on a global scale. Meanwhile, reports associating ACPs with extended-spectrum β-lactamases and carbapenemases are unsettlingly rising worldwide in enteric pathogens recovered from the environment, food-producing animals, food products and humans. ACPs also facilitate the propagation of the multidrug resistance-conferring Salmonella Genomic Island 1 (SGI1). Because ACPs and SRIs derive from a common ancestor, they code for highly similar conjugative transfer machineries, which mediate the translocation of genetic material from donor to recipient cells. Currently, the molecular mechanisms mediating these transfers are not well understood. Translocation of ACP and SRI DNA is initiated at the origin of transfer (oriT) by the relaxase TraI and the auxiliary protein MobI, both encoded by ACPs and SRIs. TraI belongs to an atypical family of relaxases, which are predicted HD phosphodiesterases and seems to require a MobI-like auxiliary protein to function, suggesting a completely unique and novel mechanism of conjugative transfer initiation. We anticipate that TraI and MobI are key factors involved in the ACP-mediated transfer of genomic islands such as SGI1.***The main goal of this proposal is to characterize the mechanism of conjugative transfer initiation of ACPs and SRIs, and the molecular interactions between the conjugative machinery of ACPs and genomic islands that they mobilize. More specifically, we propose to identify all the partner proteins that are involved in DNA processing at oriT. Moreover, we aim at discovering the protein translocation signals that allow recognition and secretion of proteins through the type IV secretion system encoded by ACPs and SRIs. We will also functionally map with single-nucleotide accuracy the oriT to determine what specific sequence features promote assembly of a high-order complex at this locus. Finally, we will investigate the interactions between the ACP-encoded relaxosome and genomic islands that are mobilized by ACPs.***Our results will yield fundamental insights about the basic biology ACPs and SRIs, thereby providing the scientific community with strong foundations and 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移动的遗传决定因素和分子机制对于对抗多重耐药细菌的出现和传播至关重要。 SXT/R391家族（SRIs）的整合和接合元件以及A/C不相容组（ACP）的接合质粒是致病性肠杆菌科和弧菌科多药耐药基因的主要载体。 SRI 导致霍乱弧菌第七种大流行谱系的抗生素耐药性，霍乱弧菌是霍乱的病原体，仍然是全球范围内死亡和发病的主要原因。与此同时，在世界范围内从环境、食用动物、食品和人类中回收的肠道病原体中，将 ACP 与超广谱 β-内酰胺酶和碳青霉烯酶相关的报告令人不安地不断增加。 ACP 还促进具有多重耐药性的沙门氏菌基因组岛 1 (SGI1) 的传播。由于 ACP 和 SRI 源自共同的祖先，因此它们编码高度相似的接合转移机制，介导遗传物质从供体细胞到受体细胞的易位。目前，介导这些转移的分子机制尚不清楚。 ACP 和 SRI DNA 的易位由松弛酶 TraI 和辅助蛋白 MobI 在转移起点 (oriT) 启动，两者均由 ACP 和 SRI 编码。 TraI 属于非典型松弛酶家族，被预测为 HD 磷酸二酯酶，并且似乎需要类似 MobI 的辅助蛋白才能发挥作用，这表明接合转移起始的完全独特且新颖的机制。我们预计 TraI 和 MobI 是参与 ACP 介导的基因组岛（如 SGI1）转移的关键因素。***本提案的主要目标是表征 ACP 和 SRI 接合转移起始机制，以及 ACP 接合机制与其动员的基因组岛之间的分子相互作用。更具体地说，我们建议鉴定 oriT 参与 DNA 加工的所有伙伴蛋白。此外，我们的目标是发现允许通过 ACP 和 SRI 编码的 IV 型分泌系统识别和分泌蛋白质的蛋白质易位信号。我们还将以单核苷酸精度对 oriT 进行功能图谱，以确定哪些特定序列特征促进该位点处高阶复合物的组装。最后，我们将研究 ACP 编码的松弛体和 ACP 动员的基因组岛之间的相互作用。***我们的结果将产生关于基础生物学 ACP 和 SRI 的基本见解，从而为科学界提供坚实的基础和新的潜在分子靶标，可用于开发旨在解决多药耐药性进展的工具。