Elucidating altered lipid pathways in daptomycin-resistant pathogens

项目摘要 细胞膜脂质的改变，特别是总磷脂酰甘油(PGs)的减少是常见的 多种革兰氏阳性菌对达托霉素的耐药表型。这些修改 膜脂含量和组成可通过脂类生物合成中的直接基因突变发生 调节细胞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。我们之前已经确定了达托霉素中膜脂变化的特征- 耐甲氧西林金黄色葡萄球菌(MRSA)、耐万古霉素肠球菌 粪霉(VRE)和纹状棒杆菌的研究发现：i)达托霉素耐药性也显著影响 非PG类脂类，以及II)仅表现在具有特定脂肪酸组成的膜脂中的变化 MRSA和VRE。脂质生物合成是开发新的治疗方法的一个很有前途的靶点。 达托霉素杀菌作用的脂质依赖机制和达托霉素耐药性的研究 真核生物和原核生物脂肪生物合成途径的差异。然而，膜脂体 的革兰氏阳性细菌在存在的脂肪类别和脂肪酸的种类和比例上不同 这些脂类的成分。这种多样性给寻找共同的脂质途径带来了挑战，这些途径可能是 被用来破坏达托霉素的耐药性，但脂质合成和代谢的中心途径是 可能在不同的达托霉素耐药物种中保守。这个项目的长期目标是 是确定脂类生物合成和代谢中共同和不同的途径，这些途径有助于 不同种类革兰氏阳性菌对达托霉素的耐药性及小分子鉴定 这些通路的调节器可以逆转达托霉素的耐药性。在目标1中，我们将阐明共同和 脂类代谢和生物合成的不同途径在不同的细菌集合中被修饰 耐达托霉素的病原菌。我们还将研究脂肪变化的脂肪酸依赖性质。 耐达托霉素的耐甲氧西林金黄色葡萄球菌和VRE。在目标2中，我们将用小分子调节达托霉素的耐药性。 靶向脂质生物合成和代谢并评价细胞外游离脂肪酸对血管内皮细胞生长的影响 对达托霉素耐药。我们预计，在不同细菌物种之间保守的途径 达托霉素耐药性将成为小分子调控的通用靶点，而这些小分子 将通过影响有利于达托霉素耐药的脂质和脂肪酸来改善达托霉素的敏感性。 这个项目将提供对分子变化的新的基本理解，这些分子变化有助于 达托霉素耐药并确定可改编为有效的新型小分子干预措施 治疗多种达托霉素耐药病原体感染的治疗方法。

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.