MOLECULAR CHARACTERISTICS AND ANTIVIRAL EFFECTS OF ATG10 IN LARGE YELLOW CROAKER (LARIMICHTHYS CROCEA)

Large yellow croaker (Larimichthys crocea) is an important economic marine fish species in China. In recent years, the frequent outbreaks of infectious diseases caused by bacteria or viruses have led to great economic losses to the large yellow croaker aquaculture. Thus, the understanding of immune mechanisms of large yellow croaker will contribute to prevent and control these diseases. Autophagy is an essential and widely conserved cellular degradation process that mediates the elimination of defective proteins and organelles. Autophagy is regulated by various autophagy-related genes (ATG), of which the ATG10 gene has been demonstrated to play a vital role in the immune response. To study the function of fish ATG10 in immune response, the ATG10 gene (LcATG10) was cloned from large yellow croaker (Larimichthys crocea). The open reading frame (ORF) of LcATG10 cDNA is 747 nucleotides long, encoding a protein of 248 amino acids with an Autophagy_act_C domain. Multiple amino acid sequence alignment showed that LcATG10 shared a high homology with ATG10 molecules in other known bony fishes. Phylogenetic analysis revealed that LcATG10 and other fish ATG10 molecules were clustered into a major clade, separated from the clade formed by the amphibian, avian, and mammalian ATG10s. Real-time qPCR results showed that the LcATG10 were widely expressed in tested tissues and immune-related cells of large yellow croaker. Poly (I:C) induced the mRNA levels of LcATG10 in the head kidney and spleen tissues with 5.9- and 9.4- fold, respectively, and it also increased LcATG10 in primary head kidney granulocytes and head kidney cell line (LYCK) cells of large yellow croaker. LcATG10-overexpression in epithelioma papulosum cyprinid (EPC) cells decreased the cytopathic effects (CPEs) caused by spring viremia of carp virus (SVCV) infection and reduced the viral titer of SVCV in the culture supernatant to 10^3.55 TCID₅₀/mL. LcATG10-overexpression significantly down-regulated the expression levels of three viral genes (SVCV-G, SVCV-M, and SVCV-P) to 0.022-, 0.015- and 0.022-folds of those in control group, respectively. These results indicate that LcATG10 played an important role in the antiviral immune response of large yellow croaker, which provides a basis for in-depth study of the molecular mechanism of autophagy in the antiviral immunity of large yellow croaker.