Identification and characterization of multiple lck genes in ginbuna crucian carp Carassius auratus langsdorfii

Lck is a member of the Src-related protein tyrosine kinase family. In mammals, it is predominantly expressed in T cells, and plays crucial role in T-cell maturation and activation. In this report, the cloning, characterization, and expression analysis of lck genes from the S3n strain of ginbuna crucian carp Carassius auratus langsdorfii is described. Three distinct lck homologs, designated as gblck-1, gblck-2, and gblck-3 were isolated from ginbuna thymus. The deduced amino acid sequences of gblck-1 and gblck-2 cDNAs exhibit conserved lck essential motif and overall structures, but gblck-3 lacks an essential region related to regulation of Src kinase activity. Reverse transcriptase polymerase chain reaction (RT-PCR) analysis demonstrated that gblck-1 was expressed in peripheral blood leukocytes (PBLs), lymphoid organs, and mucosal tissues, as has been reported for T-cell marker genes, i.e. CD3, TCR, and lck in other fish species. In contrast, the expression of gblck-2 and gblck-3 were observed only in PBLs and lymphoid organs. In the sorted lymphocytes from kidney leukocytes using antiginbuna IgM monoclonal antibody, only sIg(-) lymphocytes expressed lck genes as well as the TCR beta gene. Collectively, these results suggest that lck is a useful marker for the identification of T cells.