Molecular cloning and characterization of lymphocyte cell kinase from humphead snapper (Lutjanus sanguineus)

Abstract

Lymphocyte cell kinase (LCK) belongs to the Src family of tyrosine kinases, which involves in the proliferation control of lymphocytes. In this study, we cloned the LCK gene of humphead snapper (Lutjanus sanguineus) (designed as LsLCK). Sequence analysis showed that the full-length cDNA of LsLCK was 2279 bp, contained a 1506-bp open reading frame (ORF), encoding a polypeptide of 501 amino acids. The deduced amino acid possessed the typical structural features of known LCK proteins, including four Src homology (SH) domains arranged as the SH1 domain followed by a regulatory C-terminal tail (COOH-domain), SH2 and SH3 adapter domains and SH4 domain which required for membrane attachment and CD4/CD8 binding. Fluorescent quantitative real-time PCR analysis indicated that LsLCK transcripts were expressed mainly in thymus, spleen and head kidney in healthy fish. Moreover, the mRNA expressions in these tissues were significantly up-regulated after challenge with Vibrio harveyi. The results of immunohistochemistry showed that LsLCK protein localized distinctly in cytoplasm of cell in thymus, spleen and head kidney. Taken together, these findings indicated that LsLCK may play an important role in the immune response of humphead snapper against bacterial infection.

Honokiol inhibitsVibrio harveyihemolysin virulence by reducing its haemolytic activity

“…All samples were analysed in triplicate. The relative expression levels of the genes were detected by using Mon Amp ChemoHS qPCR Mix (Monad) in the StepOnePlus Real‐Time PCR Instrument (Applied Biosystems), analysed by the 2 −ΔΔCt method (Huang et al, ), and the independent sample t test was used to determine significant differences between the groups; p < .05 was significant, and p < .01 was extremely significant.…”