Molecular cloning of a carp Jak3 from activated leukocytes

Janus kinases (Jaks) are involved in a signalling pathway leading to tyrosine phosphorylation, which is the key step in transducing cytokine signals from the external environment to the nucleus. We report here the molecular cloning of carp Jak3. A partial cDNA of the Jak homologue was initially identified from a cDNA pool obtained by subtracting the cDNAs from Con A-induced carp head kidney leukocytes with those from uninduced carp head kidney leukocytes. The entire coding sequence was assembled by sequencing both strands of cDNA clones amplified by using anchored PCR reaction. The complete cDNA was cloned by specific primers designed against the partial cDNAs and then sequenced to assure accuracy of the complete sequence. Sequence analysis reveals an ORF encoding a protein of 1026 amino acids, with the unique presence of two carboxy-terminal kinase and pseudokinase domains. The cDNA clone shows ≈52–50%, 50–48.2%, 42–39% and 35–39% amino acid identity with the known Jak3s, Jak2s, Jak1s and Tyk2s, respectively. The sequence of the amino acid residues within the N-terminal JH domains of the carp Jak homologue shows higher similarities with the regions of human Jak3 than with human Jak2. The carp Jak is expressed as a single 4.5 Kb transcript at low levels in the head kidney and the spleen, with the RNA level increasing when leukocytes are activated with con A. The trace transcript was also detected by RT-PCR in the carp liver, muscle and skin, but not in the egg sample. Sequence analysis and pattern of expression suggest that this cDNA represents the carp Jak3 (accession number in GenBank: AF148993).