PDF(Pubmed)
Abstract：
Gram-negative bacteria have a unique cell surface that can be modified to maintain bacterial fitness in diverse environments. A well-defined example is the modification of the lipid A component of lipopolysaccharide (LPS), which promotes resistance to polymyxin antibiotics and antimicrobial peptides. In many organisms, such modifications include the addition of the amine-containing constituents 4-amino-4-deoxy-l-arabinose (l-Ara4N) and phosphoethanolamine (pEtN). Addition of pEtN is catalyzed by EptA, which uses phosphatidylethanolamine (PE) as its substrate donor, resulting in production of diacylglycerol (DAG). DAG is then quickly recycled into glycerophospholipid (GPL) synthesis by the DAG kinase A (DgkA) to produce phosphatidic acid, the major GPL precursor. Previously, we hypothesized that loss of DgkA recycling would be detrimental to the cell when LPS is heavily modified. Instead, we found that DAG accumulation inhibits EptA activity, preventing further degradation of PE, the predominant GPL of the cell. However, DAG inhibition of pEtN addition results in complete loss of polymyxin resistance. Here, we selected for suppressors to find a mechanism of resistance independent of DAG recycling or pEtN modification. Disrupting the gene encoding the adenylate cyclase, cyaA, fully restored antibiotic resistance without restoring DAG recycling or pEtN modification. Supporting this, disruptions of genes that reduce CyaA-derived cAMP formation (e.g., ptsI) or disruption of the cAMP receptor protein, Crp, also restored resistance. We found that loss of the cAMP-CRP regulatory complex was necessary for suppression and that resistance arises from a substantial increase in l-Ara4N-modified LPS, bypassing the need for pEtN modification. IMPORTANCE Gram-negative bacteria can alter the structure of their LPS to promote resistance to cationic antimicrobial peptides, including polymyxin antibiotics. Polymyxins are considered last-resort antibiotics for treatment against multidrug-resistant Gram-negative organisms. Here, we explore how changes in general metabolism and carbon catabolite repression pathways can alter LPS structure and influence polymyxin resistance.
摘要：
革兰氏阴性细菌具有独特的细胞表面，可以对其进行修饰以在不同的环境中保持细菌适应性。一个明确的例子是脂多糖（LPS）的脂质A成分的修饰，促进对多粘菌素抗生素和抗菌肽的抗性。在许多生物体中，这种修饰包括加入含胺成分4-氨基-4-脱氧-1-阿拉伯糖(1-Ara4N)和磷酸乙醇胺(pEtN)。pEtN的添加由EptA催化，它使用磷脂酰乙醇胺（PE）作为其底物供体，产生二酰基甘油(DAG)。然后通过DAG激酶A（DgkA）将DAG快速再循环到甘油磷脂（GPL）合成中，以产生磷脂酸，主要的GPL前体。以前,我们假设当LPS被大量修饰时，DgkA再循环的损失将对细胞有害。相反，我们发现DAG积累抑制了EptA活性，防止PE进一步降解，细胞的主要GPL。然而,pEtN添加的DAG抑制导致多粘菌素抗性的完全丧失。这里,我们选择抑制剂以找到一种与DAG回收或pEtN修饰无关的耐药机制。破坏编码腺苷酸环化酶的基因，cyaA,完全恢复抗生素耐药性，而不恢复DAG回收或pEtN修饰。支持这一点,破坏减少CyaA衍生的cAMP形成的基因(例如，ptsI）或cAMP受体蛋白的破坏，Crp,也恢复了抵抗力。我们发现cAMP-CRP调节复合物的丢失对于抑制是必要的，并且抗性是由l-Ara4N修饰的LPS的大量增加引起的。绕过pEtN修改的需要。重要性革兰氏阴性菌可以改变其LPS的结构，以促进对阳离子抗菌肽的抗性，包括多粘菌素抗生素.多粘菌素被认为是治疗耐多药革兰氏阴性菌的最后手段抗生素。这里,我们探讨了一般代谢和碳分解代谢物抑制途径的变化如何改变LPS结构并影响多粘菌素抗性。
