title: Combined Systems Approaches Reveal a Multistage Mode of Action of a Marine Antimicrobial Peptide against Pathogenic Escherichia coli and Its Protective Effect against Bacterial Peritonitis and Endotoxemia.
abstract: A marine arenicin-3 derivative, N4, displayed potent antibacterial activity against Gram-negative bacteria, but its antibacterial mode of action remains elusive. The mechanism of action of N4 against pathogenic Escherichia coli was first researched by combined cytological and transcriptomic techniques in this study. The N4 peptide permeabilized the outer membrane within 1 min, disrupted the plasma membrane after 0.5 h, and localized in the cytoplasm within 5 min. Gel retardation and circular dichroism (CD) spectrum analyses demonstrated that N4 bound specifically to DNA and disrupted the DNA conformation from the B type to the C type. N4 inhibited 21.1% of the DNA and 20.6% of the RNA synthesis within 15 min. Several hallmarks of apoptosis-like cell death were exhibited by N4-induced E. coli, such as cell cycle arrest in the replication (R) and division(D) phases, reactive oxygen species production, depolarization of the plasma membrane potential, and chromatin condensation within 0.5 h. Deformed cell morphology, disappearance of the plasma membrane, leakage of the contents, and ghost cell formation were demonstrated by transmission electron microscopy, and nearly 100% of the bacteria were killed by N4. A total of 428 to 663 differentially expressed genes are involved in the response to N4, which are associated mainly with membrane biogenesis (53.9% to 56.7%) and DNA binding (13.3% to 14.9%). N4-protected mice that were lethally challenged with lipopolysaccharide (LPS) exhibited reduced levels of interleukin-6 (IL-6), IL-1β, and tumor necrosis factor alpha (TNF-α) in serum and protected the lungs from LPS-induced injury. These data facilitate an enhanced understanding of the mechanisms of marine antimicrobial peptides (AMPs) against Gram-negative bacteria and provide guidelines in developing and applying novel multitarget AMPs in the field of unlimited marine resources as therapeutics.
authors: Key Laboratory of Feed Biotechnology, Ministry of Agriculture, Beijing, China.|Gene Engineering Laboratory, Feed Research Institute, Chinese Academy of Agricultural Sciences, Beijing, China.
authors: Xiumin
authors: X
authors: Wang
authors: Key Laboratory of Feed Biotechnology, Ministry of Agriculture, Beijing, China.|Gene Engineering Laboratory, Feed Research Institute, Chinese Academy of Agricultural Sciences, Beijing, China.
authors: Da
authors: D
authors: Teng
authors: Key Laboratory of Feed Biotechnology, Ministry of Agriculture, Beijing, China.|Gene Engineering Laboratory, Feed Research Institute, Chinese Academy of Agricultural Sciences, Beijing, China.
authors: Ruoyu
authors: R
authors: Mao
authors: Key Laboratory of Feed Biotechnology, Ministry of Agriculture, Beijing, China.|Gene Engineering Laboratory, Feed Research Institute, Chinese Academy of Agricultural Sciences, Beijing, China.
authors: Na
authors: N
authors: Yang
authors: Key Laboratory of Feed Biotechnology, Ministry of Agriculture, Beijing, China.|Gene Engineering Laboratory, Feed Research Institute, Chinese Academy of Agricultural Sciences, Beijing, China.
authors: Ya
authors: Y
authors: Hao
authors: Key Laboratory of Feed Biotechnology, Ministry of Agriculture, Beijing, China wangjianhua@caas.cn.|Gene Engineering Laboratory, Feed Research Institute, Chinese Academy of Agricultural Sciences, Beijing, China.
authors: Jianhua
authors: J
authors: Wang
mesh_terms: D000818:Animals; D000900:Anti-Bacterial Agents / Q000627:therapeutic use*; D023181:Antimicrobial Cationic Peptides / Q000378:metabolism; D002462:Cell Membrane / Q000187:drug effects; D002942:Circular Dichroism; D019446:Endotoxemia / Q000188:drug therapy*; D004926:Escherichia coli / Q000187:drug effects / Q000472:pathogenicity*; D005260:Female; D006090:Gram-Negative Bacteria / Q000187:drug effects / Q000472:pathogenicity; D016905:Gram-Negative Bacterial Infections / Q000188:drug therapy; D053583:Interleukin-1beta / Q000378:metabolism; D015850:Interleukin-6 / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology; D051379:Mice; D008826:Microbial Sensitivity Tests; D046529:Microscopy, Electron, Transmission; D010538:Peritonitis / Q000188:drug therapy*; D017382:Reactive Oxygen Species / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
chemical_list: D000900:Anti-Bacterial Agents; D023181:Antimicrobial Cationic Peptides; D053583:Interleukin-1beta; D015850:Interleukin-6; D008070:Lipopolysaccharides; D017382:Reactive Oxygen Species; D014409:Tumor Necrosis Factor-alpha
keywords: Escherichia coli; N4; lipopolysaccharide; marine antimicrobial peptides; mechanism