Molecular Cloning And A Preliminary Study On The Function Of MAVS From Black Carp

MAVS(mitochondria antiviral signaling protein) plays an important role in the host cellular innate immune response against microbial pathogens. In this study, MAVS has been cloned and characterized from black carp(Mylopharyngodon piceus). The full-length cDNA of black carp MAVS(bcMAVS) consists of 2352 nucleotides and the predicted bcMAVS protein contains 579 amino acids with a calculated molecular mass of 62.4 kDa and a theoretical isoelectric point of 4.83. Structural analysis showed that bcMAVS is composed of functional domains including an N-terminal CARD(caspase activation and recruitment domain), a central proline-rich domain, a putative TRAF2-binding motif and a C-terminal TM domain, which is similar to mammalian MAVS. Sequence alignment revealed that the CARD domain of bcMAVS and other known MAVS from different fish and mammals share higher similarity(53.3%-97.6%),which suggests CARD domain of MAVS is conservative in the species from fish to human.bcMAVS is constitutively transcribed in all the selected tissues including gill, kidney,heart, intestine, liver, muscle, skin and spleen; bcMAVS mRNA level in intestine, liver, muscle increased but decreased in spleen right after GCRV or SVCV infection. Multiple bands of bcMAVS were detected in western blot when it was expressed in tissue culture, which is similar to mammalian MAVS. Immunofluorescence assay determined that bcMAVS is a mitochondria protein and luciferase reporter assay demonstrated that bcMAVS could induce zebrafish IFN and EPC IFN expression in tissue culture.The next antiviral assay of bc MAVS in EPC cell lines suggest that overexpression of bc MAVS protect EPC from SVCV infection,including the decrease of virus plaque,the delay of time and the lower virus titer in comparison with the control group.Data generated in this manuscript has built a solid foundation for further elucidating the function mechanism of bcMAVS in the innate immune system of black carp.