Calmodulin Gene of Blunt Snout Bream (Megalobrama amblycephala): Molecular Characterization and Differential Expression after Aeromonas hydrophila and Cadmium Challenges

Abstract

Calmodulin (Calm), a crucial Ca2+ sensor, plays an important role in calcium-dependent signal transduction cascades. However, the expression and the relevance of Calm in stress and immune response have not been characterized in Megalobrama amblycephala. In this study, we identified the full-length cDNA of Calm (termed MaCalm) in blunt snout bream M. amblycephala, and analyzed MaCalm expression patterns in response to cadmium and Aeromonas hydrophila challenges. MaCalm was 1603 bp long, including a 5′-terminal untranslated region (UTR) of 97 bp, a 3′-terminal UTR of 1056 bp and an open reading frame (ORF) of 450 bp encoding a polypeptide of 149 amino acids with a calculated molecular weight (MW) of 16.84 kDa and an isoelectric point (pI) of 4.09. Usually, MaCalm contains four conservative EF hand motifs. The phylogenetic tree analysis indicated that the nucleotide sequence of MaCalm specifically clustered with Ctenopharyngodon idella with high identity (98.33%). Tissue distribution analysis demonstrated that the ubiquitous expression of MaCalm mRNA was found in all tested tissues, with the highest expression in the brain and the lowest expression in muscle. MaCalm showed significant upregulation at 14 d and 28 d post exposure to varying concentrations of cadmium in the liver; HSP70 transcripts in the liver significantly upregulated at 14 d post exposure to different concentrations of cadmium. Moreover, in response to the A. hydrophila challenge in vivo, MaCalm transcripts in the liver first increased and then decreased, but MaCalm transcripts in the kidney declined gradually with prolonged infection. After the A. hydrophila challenge, the expression level of HSP70 was significantly downregulated at 24 h in the liver and its expression level was notably downregulated at 12 h and at 24 h in the kidney. Collectively, our results suggest that MaCalm possesses vital roles in stress and immune response in M. amblycephala.

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