The Study On The Evolution Of MACPF Gene Superfamily In Fish

MACPF superfamily members are widely present in biological kingdoms and they play roles in a variety of processes including immune defense,neurodevelopment,cell migration and growth.The members of vertebrate MACPF gene family mainly include complement genes containing MACPF domain(C6,C7,C8α,C8β and C9),perforin,MPEG1(Macrophage-Expressed Gene 1),ASTNs(Astrotactins),and BRINPs(Bone morphogenetic protein/retinoic acid inducible neural-specific proteins).Among them,ASTNs and BRINPs genes are involved in neurodevelopment,while the rest of the members are associated with the immune defense of the host.Existing studies on the evolution of MACPF superfamily members have mainly focused on a single specie or a particular taxon.Moreover,the systematic study on the evolution of MACPF gene family members in fish have not been reported.Therefore,we carried out systematically identification,phylogenetically analysis,synteny and selective pressure analysis of the member of MACPF superfamily in evolutionally representative fish species to reveal the universality and specificity of these genes in fish.In this study,24 fish species at the evolutionary nodes of vertebrates were selected,and 5 tetrapods were selected as reference species.A total of 158 complement genes with MACPF motif(C6,C7,C8α,C8β and C9),187 perforin genes,52 MPEG1 genes,65 ASTNs(ASTN1,ASTN2)and 142 BRINPs(BRINP1,BRINP2 and BRINP3)genes were identified.The results revealed that different amounts of MACPF superfamily members in various fishes.Then,the phylogenetic analysis and structural domain and characteristic motif prediction on fish MACPF superfamily members were performed.It was found that this superfamily in fish has special characteristics compared with those of tetrapod.The phylogenetic results showed that there was only one C7 gene in tetrapod,but there were two C7 genes in teleost fish,which had undergone the third whole genome duplication unique for fish.In this study,based to the phylogenetic results,we speculated that two C7 genes(tentatively named as C7a and C7b)were generated by the common ancestor of bony fishes through whole genome duplication,and they were clustered into one branch on the phylogenetic tree and had their own evolutionary process.2.Multiple copies(316)of perforin genes were always present in cartilaginous and bony fishes,and seven evolutionary branches appeared in the phylogenetic tree.It indicated that the evolutionary history of perforin in fishes is much more complex than in tetrapod with multiple ancestral genes and different evolutionary process.3.There was only one BRINP3 gene in tetrapod,while three BRINP3 genes(tentatively named BRINP3a.1,BRINP3a.2 and BRINP3b in this thesis)were commonly found in spoked finfish.The three BRINP3 genes were clustered in a phylogenetic tree and each of them had their own evolutionary process.Next,synteny analysis was performed for each member of the MACPF superfamily,and the results were concluded as follows:the loci of C6 and C7b were closely linked in fish,which was similar to the positions of the C6 and C7 in tetrapod,and the genes on both sides were conserved.The intra-species synteny analysis and phylogenetic analysis showed that the duplicated genes appearing in fish C6 were proximal duplication genes and fragmental duplication genes.The gene loci of C7a were more conserved in bony fishes and the presence of the same neighboring gene as C7b.C8α and C8β were conserved in most fish and tetrapod with closely contiguous loci and their co-linear regions,except for the Salmonids and Elopomorpha whose C8α and C8β were located on two chromosomes.Synteny analysis showed that C9 was more conserved in fish and tetrapod.The perforin gene had six loci in fish,which were hypothesized to be originated from transposon duplication.ASTN1,BRINP2,ASTN2 and BRINP1 had more conserved syntenic regions in both fish and tetrapod.The fish BRINP3a.1 and BRINP3a.2 loci were closely adjacent and conserved in synteny analysis,and BRINP3b was not on the same chromosome as BRINP3a.1 and BRINP3a.2.Combined with intra-species synteny analysis and phylogenetic analysis,the duplicated genes of ASTNs and BRINPs occurring in fish were dispersed duplicated genes,fragmented duplicated genes and whole genome duplicated genes.Finally,selection pressure analysis was performed on MACPF superfamily members in fish,and the positive selection sites of amino acid were calculated and labeled on the protein 3D structures.The number of positive selection amino acid sites was found to be different in each MACPF superfamily members.There were no positive selection sites for both ASTN1 and BRINPs genes,suggesting that the MACPF superfamily members underwent different adaptive changes.Among them,ASTN1 and BRINPs genes are associated with neurodevelopment and thus have been strongly functionally constrained during fish evolution.In summary,the information of fish MACPF superfamily members was systematically disclosed in the present thesis,their evolution process and the evolutionary relationship between their different isoforms were also deeply analyzed,providing the foundation for further exploration of the functions of fish MACPF superfamily members.