Molecular Identification, Functional Characterization And The Evolution Analysis Of The Complement Component C1q And Immunoglobulin Genes In Teleosts

This article focused on the molecular identification,functional characterization, and evolution analysis of complement component C1q genes and immunoglobulin genes in teleost fish.In part one,based on the bioinformatics method such as synteny comparative analysis approach,we successfully identified and cloned the C1qA,B,C chains in Zebrafish(Danio rerio).The gene expression profiling in different tissues were detected.It was found that these genes are globally expressed and the mRNA expressions of Zebrafish C1qA could be detected at all stages during the development. Furthermore,the evolution analysis was performed the elucidate the evolution relationships of C1q family members in teleost fish such as Zebrafish(Danio rerio), Fugu(Fugu rubripe) and Tetraodon(Tetraodon nigroviridis),as well as in other vertebrate species.By phylogenetic analysis,we revealed an evolutionary relationship between C1q and EMu superfamily.Finally,utilizing the soluble recombinant protein which expressed in E.coli,we analyzed the functional characterization of the globular head regions of C1qA(ghA),B(ghB),and C(ghC) chains.The results showed that the ghA,ghB and ghC could bind to heat-aggregated human IgM and IgG and inhibit C1q mediated complement classical pathway.Moreover,it could bind to heat-aggregated and cross linked zebrafish IgM,but it could not bind to cross linked zebrafish IgZ.In part two,we cloned multiple immunoglobulin genes in zebrafish.Two genes, named IgM and IgZ,have been reported previously.And the other two novel immunoglobulin genes,including IgZ-2 and IgMm2,were identified for the first time. According to the structure characterizations,the IgMm2 gene was supposed to be formed by the directly splicing of the transmemebrane region to CH1 of IgM.The IgZ-2 showed high homologies to the newly identified IgZ.It was expressed in the kidney,skin,and intestine and we hypothesis that it may related to mucosal immunity. It was firstly appeared 14 days after fertilization.C1q is the first subcomponent of the classical pathway of complement activation and a major connecting link between classical pathway-driven innate immunity and IgG- or IgM- mediated acquired immunity.It could activate the classical complement pathway via binding of its globular heads to the Fc regions of aggregated IgG or IgM. In the present study,we cloned the C1qA,C1qB,and C1qC chain in zebrafish fish for the first time.Also,we identified several immunoglobulin genes in zebrafish.Thus, we performed an ELISA analysis to find a relationship between C1q and immunoglobulins and studied the classical pathway in teleost fish.In teleost fish,the classical pathway was identified,which showed similar properties as it in the mammals.And the C1q in teleost fish also has the similar functional characterizations as its mammalian counterpart.