MHC Ⅱ Antigen Presentation Pathway In Orange-spotted Grouper And Expression Patterns Of Its Immune-related Molecules Involved In The C.irritans Infection

C.irritans is a parasite pathogen that causes severe damage to marine fish culture in Southeast Asia.Previous studies have shown this parasite can cause efficient skin mucosal and systematic humoral immune response.Where and how the MHC antigen presentation occur in the C.irritans infected fish is still unknown.In this study,the tissue distribution of all these key molecules in MHC antigen presentation pathway was tested by Real-time PCR.To verify how they attended the grouper’s antigen presentation against C.irritans infection,then their expression patterns were detected in the mucosal tissues and systematic immune organs.The main results of this paper’ study were as follows:The full-length cDNA of grouper cysteine protease CTSS(GeneBank no.KU179320)was fistly cloned in this study.Alignment analysis showed the grouper CTSS was 56%-79.1% identity to other vertebrate CTSS.The phylogenetic tree analysis illustrated grouper CTSS had a closer relationship with other fish CTSS.The tissue expression analysis showed that CTSB and CTSL were widely expressed in the 13 tested tissues,the higher expression of CTSB was in spleen,head kidney and gills,while the expression level of CTSL was higher in muscle and skin.CTSS was relatively restricted expressed in the tested tissues,and higher expression was found in head kidney and spleen.In gill and head kidney,the highest expression of GILT was detected.MHC ⅡA was mainly higher expressed in thymus,skin,midgut and gill;Followed by gills,spleen,kidney and thymus,the highest expression level of MHC ⅡB was in skin.MHC Ⅰ α was higher expessed in PBLs,gills,head kidney and spleen,with lowest experssion in the stomach and brain.β2m was widely expressed in the tested tissues,with higher exppesion found in PBLs,skin and gill.The genes expression patterns in C.irritans infected grouper’s skin,gill,headkidney and spleen were tested at different time points post infection by Real-time PCR.(1)The most significnat up-regulations of MHC ⅡA were seen in head kidney,while MHC ⅡB mianly showed a trend of up-regulations in skin and spleen.Besides,CTSB,CTSL CTSS and GILT were significantly up-regulated in skin,while significant up-regulations of CTSL were also seen in spleen.(2)In C.irritans challenged groupers,significant up-regulations of MHC Ⅰ α and β2m were seen in skin and spleen.According to these results,C.irritans infection can cause active MHC Ⅱ antigen presentation occurrence in the local mucosal tissues and systematic immune organs.Besides the systematic immune organs,the significant and earlier mucosal antigen presentation indicates the specific immune response may also occur in the local mucosal tissues.As C.irritans are extracellular parasites,the expressions of MHC Ⅰ pathway immue-related genes were also found to be regulated.These results indicate that in the activation of specific cellelar immune response to parasites,grouper MHC Ⅰ may play an important role.