Identification of two carcinin isoforms (MnCarc1 and MnCarc2) and their function in the antimicrobial immunity of Macrobrachium nipponense.

Carcinin, a member of the crustin family, plays important roles in crustacean innate immunity. In this study, we identified two carcinin isoforms (MnCarc1 and MnCarc2) produced by alternative splicing from Macrobrachium nipponense. The full length of MnCarc1 and MnCarc2 cDNA are 1554 and 1495 bp with 687 and 609 bp open reading frame-encoding proteins that contain 228 and 202 amino acids, respectively. The genome of carcinin has nine exons and eight introns. MnCarc1 transcript contains all nine exons, whereas MnCarc2 only contains eight exons and lacks exon 4. MnCarc1 and MnCarc2 proteins contain a signal peptide, cysteine-rich regions, and a whey acidic protein domain. The phylogenetic tree shows that MnCarc1 and MnCarc2 are not grouped with other crustins and carcinins. MnCarc1 and MnCarc2 form a subgroup. MnCarc1 and MnCarc2 are widely distributed in various tissues. The expression of MnCarc1 and MnCarc2 were evidently upregulated at multiple time points in hemocytes and the intestine of M. nipponense after white spot syndrome virus, Vibrio parahaemolyticus, and Staphylococcus aureus challenges. Further studies showed that knockdown of MnDorsal or MnStat transcription factor could remarkably inhibit the upregulated expression of MnCarc1 and MnCarc2 caused by viral or bacterial challenges. In addition, recombinant MnCarc1 and MnCarc2 proteins could bind to various bacteria and polysaccharides and inhibit the growth of S. aureus and V. parahaemolyticus in vitro. This study indicated that carcinins from M. nipponense were involved in prawns innate immunity.