Mechanisms Study Of Differential Expression Of Antimicrobial Peptides In Dark-spotted Frog(Rana Nigromaculate) By Bacterial Induction

Plenty of antimicrobial peptides(AMPs)from Amphibian provide a rich template for developing antimicrobial drug.In addition,antimicrobial peptides are important effector molecules of the natural immune system of organisms.They have attracted much attention due to their wide antibacterial spectrum,strong specificity,and high antibacterial efficiency.Many antimicrobial peptides have been used as new antibacterial drugs for clinical development.Studies have shown that the mechanism of action of antimicrobial peptides is different from that of traditional antibiotics.It mainly acts on the cell membrane of microorganisms and is not easy to cause microbial resistance.Therefore,it is extremely necessary to study it.However,the mechanisms of AMPs diveristy was unknown.Black-spotted frog has the characteristics of wide survival range and moderate types of antimicrobial peptides.It is an ideal research object of antimicrobial peptide immune pathways in amphibians.Firstly,dark-spotted frog(Rana nigromaculata)was induced by various bacteria,then the transcriptome sequencing were performed.Our study will reveal the generation mechanism of AMPs of R.nigromaculata,and provide understanding for Amphibian immune mechanism and theoretical basis for the utilization of AMPs.The main findings are as follows:(1)Transcriptome data of the back skin of the R.nigromaculataAfter being stimulated by different pathogenic microorganisms,the skin of the black spotted frog produced different antimicrobial peptide expression profiles.High-throughput sequencing was performed on the back skin of the black-spotted frog stimulated by different pathogens,and the transcriptome data of different groups were obtained by processing the original data,splicing and assembling the transcripts.(2)The expression profile of the back skin of R.nigromaculataThe results of GO enrichment show that the differential genes of each group of R.nigromaculata are mainly enriched in the signal pathways of defense response,extracellular region,peptidase inhibitor activity,peptidase regulator activity,enzyme inhibitor activity,endopeptidase inhibitor activity,endopeptidase regulator activity,enzyme regulator activity.The results of KEGG enrichment showed that the differential genes of each group of black-spotted frogs were mainly enriched in B cell receptor signaling pathway,NF-kappa B signaling pathway,PI3K-Akt signaling pathway and Phospholipase D signaling pathway,the Calcium signaling pathway,Natural killer cell mediated cytotoxicity.(3)Prediction of antimicrobial peptide-related genes produced by R.nigromaculataWe predict that the Cluster-115471.109485 gene may be related to the immune response of R.nigromaculata to Gram-negative bacteria,and the Cluster-115471.47742 gene may be related to the immune response of R.nigromaculata to Gram-positive bacteria.Conclusion:The signal pathway of the black-spotted frog that is stimulated by pathogens to produce antimicrobial peptides is similar to that of mammals and Drosophila.The NF-?B signaling pathway,Toll-like receptor signaling pathway,and chemokine signaling pathway are all involved,and we predict Cluster-115471.109485 and Cluster-115471.47742 genes are key genes in the pathway of antimicrobial peptide production.