Characterization of ADP ribosylation factor 1 gene from Exopalaemon carinicauda and its immune response to pathogens challenge and ammonia-N stress

Abstract

ADP ribosylation factors (Arf), as highly conserved small guanosine triphosphate (GTP)-binding proteins, participates in intracellular trafficking and organelle structure. In this study, a full-length cDNA of Arf1 (designated EcArf1) was cloned from Exopalaemon carinicauda by using rapid amplification of cDNA ends (RACE) approaches. The full-length cDNA of EcArf1 was 1428bp, which contains an open reading frame (ORF) of 549bp, encoding a 182 amino-acid polypeptide with the predicted molecular weight of 20.69kDa and estimated isoelectric point was 7.24. Sequence analysis revealed that the conserved Arf protein family signatures were identified in EcArf1. The deduced amino acid sequence of EcArf1 shared high identity (95%-98%) with that of other species and clustered together with Arf1 of other shrimp in the NJ phylogenetic tree, indicating that EcArf1 should be a member of the Arf1 family. Quantitative real-time RT-qPCR analysis indicated that EcArf1 was expressed in hemocytes, hepatopancreas, gills, muscle, ovary, intestine, stomach and heart, and the most abundant level was in hemocytes and gills, which were also the two main target tissues of pathogen infection and environmental stress. After Vibrio parahaemolyticus challenge, EcArf1 transcripts level significantly increased in hemocytes and hepatopancreas at 3h and 6h, respectively. The expression of EcArf1 in hemocytes and hepatopancreas significantly up-regulated at 12h and 6h respectively, and down-regulated at 72h and 48h, respectively. EcArf1 expression in hepatopancreas and gills both significantly increased at 6h and decreased at 24h under ammonia-N stress. The results suggested that EcArf1 might be involved in immune responses to pathogens (V.parahaemolyticus and WSSV) challenge and ammonia-N stress in E.carinicauda.