Antimicrobial peptides(AMPs) are a group of gene-encoded small peptides that play pivotal roles in the host immune system of multicellular organisms.Cathelicidins are an important family of AMPs that exclusively exist...Antimicrobial peptides(AMPs) are a group of gene-encoded small peptides that play pivotal roles in the host immune system of multicellular organisms.Cathelicidins are an important family of AMPs that exclusively exist in vertebrates. Many cathelicidins have been identified from mammals, birds, reptiles and fish. To date, however, cathelicidins from amphibians are poorly understood. In the present study, two novel cathelicidins(OL-CATH1 and 2) were identified and studied from the odorous frog Odorrana livida.Firstly, the cDNAs encoding the OL-CATHs(780 and735 bp in length, respectively) were successfully cloned from a lung cDNA library constructed for the frog. Multi-sequence alignment was carried out to analyze differences between the precursors of the OL-CATHs and other representative cathelicidins.Mature peptide sequences of OL-CATH1 and 2 were predicted(33 amino acid residues) and their secondary structures were determined(OL-CATH1 showed a random-coil conformation and OL-CATH2 demonstrated α-helical conformation). Furthermore,OL-CATH1 and 2 were chemically synthesized and their in vitro functions were determined. Antimicrobial and bacterial killing kinetic analyses indicated that OL-CATH2 demonstrated relatively moderate and rapid antimicrobial potency and exhibited strong anti-inflammatory activity. At very low concentrations(10 μg/mL), OL-CATH2 significantly inhibited the lipopolysaccharide(LPS)-induced transcription and production of pro-inflammatory cytokines TNF-α, IL-1βand IL-6 in mouse peritoneal macrophages. In contrast, OL-CATH1 did not exhibit any detectableantimicrobial or anti-inflammatory activities. Overall,identification of these OL-CATHs from O. livida enriches our understanding of the functions of cathelicidins in the amphibian immune system. The potent antimicrobial and anti-inflammatory activities of OL-CATH2 highlight its potential as a novel candidate in anti-infective drug development.展开更多
Antimicrobial peptides（AMPs）,a class of gene-encoded peptides,are the first line of immune system to defense microbial invasions in multicellular organisms.Cathelicidins are an important family of AMPs that have bee...Antimicrobial peptides（AMPs）,a class of gene-encoded peptides,are the first line of immune system to defense microbial invasions in multicellular organisms.Cathelicidins are an important family of AMPs that have been identified exclusively in vertebrates.However,up to now,cathelicidins from amphibians are poorly understood.In the present study,we reported the identification and characterization of two novel cathelicidins（FM-CATH1 and FMCATH2） from the frog Fejervarya multistriata.The c DNA sequences encoding FM-CATHs were successfully cloned from the constructed lung c DNA library of F.multistriata.Both of the c DNA sequences encoding FM-CATHs are 447 bp in length,and the deduced mature peptides of FM-CATHs are composed of 34 residues.Structural analysis indicated that FM-CATH1 and FM-CATH2 mainly assume amphipathic alpha-helical conformations.Antimicrobial and bacterial killing kinetic analysis indicated that both FM-CATH1 and FM-CATH2 possess potent,broad-spectrum and rapid antimicrobial potency.And cytoplasmic membrane permeabilization analysis indicated that FM-CATH1 and FMCATH2 kill bacteria by inducing the permeabilization of bacterial membrane.Besides direct antimicrobial activities,FM-CATHs also exhibited significant inhibitory effect on the formation of bacterial biofilms at low concentrations below 1×MIC.Furthermore,FM-CATH1 and FM-CATH2 exhibited potent anti-inflammatory activities by inhibiting LPS-induced transcription and production of pro-inflammatory cytokines TNF-α,IL-1β,and IL-6 in mouse peritoneal macrophages.Meanwhile,FM-CATHs showed relatively low cytotoxic activity against mammalian normal and tumor cell lines,and low hemolytic activity against human erythrocytes.In summary,the identification of FM-CATHs provides novel clues for our understanding of the roles of cathelicidins in amphibian immune systems.The potent antimicrobial,biofilm inhibitory,anti-inflammatory activities,and low cytotoxicity of FM-CATHs imply their great potential in novel antibiotics development.展开更多
基金supported by grants from the Jiangsu Students' Innovation and Entrepreneurship Training Program(2017suda098)the National Natural Science Foundation of China(31772455)+2 种基金Natural Science Foundation of Jiangsu Province(BK20160336 and BK20171214)Natural Science Foundation of College in Jiangsu Province(16KJB350004)Suzhou Science and Technology Development Project(SYN201504 and SNG2017045)
文摘Antimicrobial peptides(AMPs) are a group of gene-encoded small peptides that play pivotal roles in the host immune system of multicellular organisms.Cathelicidins are an important family of AMPs that exclusively exist in vertebrates. Many cathelicidins have been identified from mammals, birds, reptiles and fish. To date, however, cathelicidins from amphibians are poorly understood. In the present study, two novel cathelicidins(OL-CATH1 and 2) were identified and studied from the odorous frog Odorrana livida.Firstly, the cDNAs encoding the OL-CATHs(780 and735 bp in length, respectively) were successfully cloned from a lung cDNA library constructed for the frog. Multi-sequence alignment was carried out to analyze differences between the precursors of the OL-CATHs and other representative cathelicidins.Mature peptide sequences of OL-CATH1 and 2 were predicted(33 amino acid residues) and their secondary structures were determined(OL-CATH1 showed a random-coil conformation and OL-CATH2 demonstrated α-helical conformation). Furthermore,OL-CATH1 and 2 were chemically synthesized and their in vitro functions were determined. Antimicrobial and bacterial killing kinetic analyses indicated that OL-CATH2 demonstrated relatively moderate and rapid antimicrobial potency and exhibited strong anti-inflammatory activity. At very low concentrations(10 μg/mL), OL-CATH2 significantly inhibited the lipopolysaccharide(LPS)-induced transcription and production of pro-inflammatory cytokines TNF-α, IL-1βand IL-6 in mouse peritoneal macrophages. In contrast, OL-CATH1 did not exhibit any detectableantimicrobial or anti-inflammatory activities. Overall,identification of these OL-CATHs from O. livida enriches our understanding of the functions of cathelicidins in the amphibian immune system. The potent antimicrobial and anti-inflammatory activities of OL-CATH2 highlight its potential as a novel candidate in anti-infective drug development.
基金supported by the grants BK20160336 from the Natural Science Foundation of Jiangsu Province to Yan CHEN16KJB350004 from the Natural Science Foundation of College in Jiangsu Province to Yipeng WANG+1 种基金SYN201407 and SYN201504 from the Suzhou Science and Technology Development Project to Yipeng WANG2015NY06 from the Haimen Science and Technology Development Project to Yipeng WANG
文摘Antimicrobial peptides（AMPs）,a class of gene-encoded peptides,are the first line of immune system to defense microbial invasions in multicellular organisms.Cathelicidins are an important family of AMPs that have been identified exclusively in vertebrates.However,up to now,cathelicidins from amphibians are poorly understood.In the present study,we reported the identification and characterization of two novel cathelicidins（FM-CATH1 and FMCATH2） from the frog Fejervarya multistriata.The c DNA sequences encoding FM-CATHs were successfully cloned from the constructed lung c DNA library of F.multistriata.Both of the c DNA sequences encoding FM-CATHs are 447 bp in length,and the deduced mature peptides of FM-CATHs are composed of 34 residues.Structural analysis indicated that FM-CATH1 and FM-CATH2 mainly assume amphipathic alpha-helical conformations.Antimicrobial and bacterial killing kinetic analysis indicated that both FM-CATH1 and FM-CATH2 possess potent,broad-spectrum and rapid antimicrobial potency.And cytoplasmic membrane permeabilization analysis indicated that FM-CATH1 and FMCATH2 kill bacteria by inducing the permeabilization of bacterial membrane.Besides direct antimicrobial activities,FM-CATHs also exhibited significant inhibitory effect on the formation of bacterial biofilms at low concentrations below 1×MIC.Furthermore,FM-CATH1 and FM-CATH2 exhibited potent anti-inflammatory activities by inhibiting LPS-induced transcription and production of pro-inflammatory cytokines TNF-α,IL-1β,and IL-6 in mouse peritoneal macrophages.Meanwhile,FM-CATHs showed relatively low cytotoxic activity against mammalian normal and tumor cell lines,and low hemolytic activity against human erythrocytes.In summary,the identification of FM-CATHs provides novel clues for our understanding of the roles of cathelicidins in amphibian immune systems.The potent antimicrobial,biofilm inhibitory,anti-inflammatory activities,and low cytotoxicity of FM-CATHs imply their great potential in novel antibiotics development.
