Elucidating altered lipid pathways in daptomycin-resistant pathogens

项目摘要 膜脂的改变，特别是总磷脂酰甘油（PG）的减少，是常见的 达托霉素耐药表型在许多种革兰氏阳性菌中存在。这些修改， 膜脂质含量和组成可以通过脂质生物合成中的直接基因突变发生， 调节膜表达的细胞包膜应激反应系统中的途径或间接突变 和细胞壁生物合成基因。我们先前已经描述了达托霉素中改变的膜脂质- 耐甲氧西林金黄色葡萄球菌（MRSA）、耐万古霉素肠球菌 粪肠杆菌（VRE）和棒状杆菌纹状体，并发现：i）达托霉素耐药性也显着影响 非PG脂质，和ii）仅在具有特定脂肪酸组成的膜脂质中表现出变化 MRSA和VRE。脂质的生物合成是一个有前途的目标，为发展新的治疗方法， 由于达托霉素杀菌作用的脂质依赖性机制和 真核生物和原核生物脂质合成途径的差异。然而，膜脂质体 革兰氏阳性菌的脂质种类和比例以及脂肪酸的种类和比例不同， 这些脂质的组成。这种多样性对寻找共同的脂质途径提出了挑战， 开发破坏达托霉素耐药性，但脂质合成和代谢的中心途径， 可能在不同的达托霉素耐药物种中保守。本项目的长期目标 确定脂质生物合成和代谢中的共同和差异途径， 不同革兰氏阳性菌对达托霉素的耐药性及小分子鉴定 这些途径的调节剂可以逆转达托霉素耐药性。在目标1中，我们将阐明常见和 脂质代谢和生物合成中的差异途径在不同的细菌集合中被修饰， 具有达托霉素抗性的病原体。我们还将研究脂肪酸依赖性的性质，脂质变化， 耐达托霉素MRSA和VRE。在目标2中，我们将用小分子调节达托霉素耐药性 靶向脂质生物合成和代谢，并评估细胞外游离脂肪酸对 达托霉素耐药性我们预计，在不同细菌物种中保守的途径， 达托霉素抗性将是小分子调节的通用靶标，并且这些小分子 将通过影响有利于达托霉素耐药性的脂质和脂肪酸来改善达托霉素敏感性。 该项目将提供新的分子改变的基本理解，有助于 达托霉素耐药性，并确定新的小分子干预措施，可以适用于有效的 用于治疗多种达托霉素抗性病原体感染的治疗剂。

Project Summary Alteration of membrane lipids, particularly the reduction of total phosphatidylglycerols (PGs), is a common daptomycin resistance phenotype across many species of Gram-positive bacteria. These modifications to membrane lipid content and composition can occur through direct genetic mutations in lipid biosynthetic pathways or indirect mutations in cell envelope stress response systems that regulate expression of membrane and cell wall biosynthesis genes. We have previously characterized the altered membrane lipids in daptomycin- resistant strains of methicillin-resistant Staphylococcus aureus (MRSA), vancomycin-resistant Enterococcus faecalis (VRE), and Corynebacterium striatum and found that: i) daptomycin resistance also significantly affected non-PG lipids, and ii) the changes manifested only in membrane lipids with specific fatty acid compositions in MRSA and VRE. Lipid biosynthesis is a promising target for the development of novel therapies for the treatment of daptomycin resistance due to the lipid-dependent mechanism of daptomycin’s bactericidal action and the differences between eukaryotic and prokaryotic lipid biosynthetic pathways. However, the membrane lipidomes of Gram-positive bacteria are diverse in the variety and ratios of lipid classes present and the fatty acid compositions of those lipids. This diversity presents a challenge to finding common lipid pathways that can be exploited to disrupt daptomycin resistance, but the central pathways of lipid synthesis and metabolism are likely to be conserved across diverse daptomycin-resistant species. The long-term goals of this project are to identify the common and differential pathways in lipid biosynthesis and metabolism that contribute to daptomycin resistance among different species of Gram-positive bacteria and to identify small molecule modulators of these pathways that can reverse daptomycin resistance. In Aim 1, we will elucidate common and differential pathways in lipid metabolism and biosynthesis that are modified in a diverse collection of bacterial pathogens with daptomycin resistance. We will also examine the fatty acid-dependent nature of lipid changes in daptomycin-resistant MRSA and VRE. In Aim 2, we will modulate daptomycin resistance with small molecules targeting lipid biosynthesis and metabolism and evaluate the effects of extracellular free fatty acids on daptomycin resistance. We expect that the pathways that are conserved among diverse bacteria species with daptomycin resistance will be universal targets for modulation with small molecules, and these small molecules will improve daptomycin susceptibility by affecting the lipids and fatty acids that favor daptomycin resistance. This project will provide new fundamental understanding of the molecular alterations that contribute to daptomycin resistance and identify novel small molecule interventions that can be adapted into effective therapeutics for treating infections from multiple species of daptomycin-resistant pathogens.