MOLECULAR CHARACTERIZATION AND EXPRESSION ANALYSIS OF SECRETORY IMMUNOGLOBULIN M (IGM) HEAVY CHAIN GENE IN ROHU Labeo rohita

The major humoral component of adaptive immune system is Immunoglobulins (Igs) of which Immunoglobulin M (IgM) is the major isotype among teleost Igs and is involved in systemic and mucosal immunity. IgM heavy chain molecules are present in two different forms namely, secretory and membrane IgM. The information on structural and functional properties of immunoglobulin genes in rohu and other Indian Major Carps is lacking. Knowledge on the adaptive immune system including structural and genetic organization of immunoglobulins is of potential importance to devise effective prophylactic measures to manage diseases in cultured fish. The present study was aimed to explore IgM heavy chain gene and its expression profile in healthy rohu and fishes challenged with bacteria.

Full-length IgM heavy chain cDNA of rohu was sequenced using RACE PCR. The IgM gene consists of 1994 bp encoding a polypeptide of 576 amino acid residues including a leader peptide, variable (VH) and constant (CH1-CH2-CH3-CH4) domains confirming the secretory form of IgM. The sequence carries conserved residues such as cysteine, tryptophan and amino acid motifs. The predicted 3D model confirmed various domains of rohu IgM heavy chain (Fig. 1). Phylogenetic tree analysis revealed that rohu IgM shared the same cluster with that of other cyprinid fishes.  

Tissue distribution analysis was carried out using quantitative real-time PCR which showed the predominant level of IgM heavy chain gene expression in kidney followed by spleen and intestine (Fig. 2). Further, IgM heavy chain gene expression in rohu kidney was found to be upregulated and reached maximum at 7 days post-challenge with Aeromonas hydrophila (Fig. 3). This is the first report of full-length secretory IgM heavy chain gene of rohu. Besides, IgM heavy chain gene was highly expressed in major lymphoid tissues and bacterial challenge upregulated its expression thus confirming its role in adaptive humoral immune response.