A novel mechanism of rifamycin resistance in Mycobacterium abscessus mediated by a putative helicase

项目总结： 脓肿分枝杆菌(Mab)是一种快速生长的NTM，可引起皮肤和软组织感染。 以及慢性肺损伤患者的肺部感染。MAB脱颖而出，成为世界上 对抗生素具有抗药性的微生物物种，使其感染难以治疗。特别引人注目的是 它对利福平(RIF)的耐药性，利福平是治疗包括结核病在内的许多分枝杆菌感染的一线药物。 RIF通过与细菌RNA聚合酶的b亚基结合来抑制全球RNA的合成；此外，它还 也已知由于抑制DNAA的表达而抑制复制启动。固有的RIF 到目前为止，单抗的抗性归因于ADP-核糖基转移酶(ARR)活性的存在 核糖化RIF导致其失活。然而，我们最近发现了另一种 决定簇MAB_3189c-一种假定的解旋酶，在单抗中具有高水平的RIF抗性。 MAB_3189c的表达是RIF诱导的，可能受RIF相关元件(RAE)的调控 依赖转录因子。在本项目中，我们将确定Mab3189c的调节和功能 在RIF抵抗中。在目标1中，我们将研究Mab3189c是否通过直接或间接方式介导RIF抵抗 保护RNAP免受RIF的作用，从而使全球RNA合成继续进行，或通过 减轻在ORIM对RIF敏感的复制启动，从而能够在RIF存在下生长。在AIM 2我们建议鉴定RAE依赖的MAB_3189c诱导所需的调节蛋白。这个 这些发现将为长期研究提供一个平台，以深入了解Mab3189c依赖的RIF 单抗的耐药性，以及设计治疗单抗感染的新策略。

Project Summary: Mycobacterium abscessus (Mab) is a rapidly growing NTM causing skin and soft tissue infections as well as pulmonary infections in patients with chronic lung damage. Mab stands apart as one of the most antibiotic resistant microbial species, making its infections incredibly difficult to treat. Particularly striking is its resistance to rifampicin (RIF), a frontline drug for many mycobacterial infections including tuberculosis. RIF inhibits global RNA synthesis by binding to the b-subunit of bacterial RNA polymerase; additionally it is also known to inhibit replication initiation due to an inhibition of dnaA expression. The intrinsic RIF resistance in Mab has so far been attributed to the presence of an ADP-ribosyltransferase (Arr) activity that ribosylates RIF leading to its inactivation. However, we have recently identified an additional determinant, MAB_3189c - a putative helicase, that confers high levels of RIF resistance in Mab. MAB_3189c expression is RIF inducible and is likely regulated by a RIF-associated element (RAE) dependent transcription factor. In this project we will determine the regulation and function of Mab3189c in RIF resistance. In Aim 1 we will investigate if Mab3189c mediates RIF resistance by either directly protecting RNAP against the action of RIF thereby enabling global RNA synthesis to continue, or by alleviating RIF-sensitive replication initiation at oriM thereby enabling growth in the presence of RIF. In Aim 2 we propose to identify the regulatory protein required for RAE-dependent induction of MAB_3189c. The findings will provide a platform for a long-term study to gain insights into Mab3189c dependent RIF resistance in Mab, and design of new strategies to treat Mab infections.