Characterization and expression analysis of a gC1qR gene from Macrobrachium nipponense under ammonia-N stress

Abstract

Ammonia-N is the most common environmental stress for prawn culture, and gC1qR is a multifunctional and multiligand binding protein with a significant role in the innate immunity in crustaceans. In the present study, a gC1qR (MngC1qR) cDNA was identified from the freshwater prawn, Macrobrachium nipponense (M. nipponense). The full-length cDNA of MngC1qR was 1164 bp, with an open reading frame (ORF) of 774 bp encoding 257 amino acids. Multiple alignment analysis showed that gC1qR in M. nipponense and other crustaceans shared high similarity. The expression level of MngC1qR and the change of antioxidant enzyme (super oxide dismutase (SOD) and catalase (CAT)), phenoloxidase (PO) and lysozyme (LSZ) were investigated in M. nipponense after ammonia-N exposure. The SOD and CAT activities in hepatopancreas were significantly increased at 12 h and 24 h, respectively, and decreased significantly at 48 h. The activities of PO and LSZ in serum was significantly decreased from 12 h to 96 h. Quantitative real-time PCR (qRT-PCR) demonstrated that MngC1qR was expressed in all examined tissues, with the high expression level in the hepatopancreas and gills. The expression of MngC1qR was significantly up-regulated at 12 h in the tissues of hepatopancrea and gills under ammonia-N stress. Western blot analysis further indicated that the MngC1qR protein was distributed in all tissues, with the highest level in hepatopancreas, followed by gills and heart, which was consistent with the qRT-PCR results. RNA interference analysis showed that the knockdown of MngC1qR could significantly decrease the transcription level of MngC1qR and led to the high mortality. Furthermore, the expression level of MngC1qR was found to be induced significantly under ammonia-N stress. Findings of the present study indicate that high concentration of ammonia-N has significantly effects on the M. nipponense and MngC1qR may be involved in the immune defense against ammonia-N stress.