MOLECULAR INSIGHTS INTO LYMPHOCYTE CYTOSOLIC PROTEIN 2 (LCP2); EXPRESSION ANALYSIS OF Sebastes schlegelii AFTER ACTIVATING T-CELL RECEPTORS FROM IMMUNE STIMULATION

D.S. Liyanage, G.I. Godahewa and Jehee Lee
 
Department of Marine Life Sciences & Fish Vaccine Research Center
Jeju National University
Jeju Self-Governing Province 63243, Republic of Korea.
Email: dileepasripal@gmail.com
 

Lymphocyte cytosolic protein 2 (LCP2), also known as SLP-76, is a cytosolic adapter protein that nucleates signaling complexes generated by immunoreceptors. It is mostly studied in T lymphocytes and associated with many proteins. These molecules recruited by LCP2 launch the intracellular propagation of molecular signals leading to downstream responses including calcium mobilization, activation of MAP kinases, integrin activation, cytoskeletal reorganization and ultimately full-blown T cell responses. According to in-silico analysis performed, LCP2 protein belongs to the SAM, Atrophin-1 and SH2 super families; possesses inducibly phosphorylated tyrosines in N-terminus, a central proline-rich domain and C-terminal SH2 domain. LCP2 translated sequence consisted of 507 amino acids. Pairwise alignment results showed that highest sequence similarity to Larimichthys crocea with 78.3 % identity and 84.0 % similarity and N-linked glycosylation sites can be found at 48NLSD51 and 286NRTN289. The LCP2 protein showed signal peptides at 1-20 residues.

Quantitative real time PCR indicated, the highest expression was in blood followed by spleen, gill among fourteen different tissues checked from healthy rockfish. As these tissues actively participated in immune responses, intensive expression of LCP2 can be observed. Then blood and spleen tissues were subjected to the immune challenge with Lipopolysaccharides (LPS), Poly IC and Streptococcus iniae to observe variation in the transcriptional pattern of LCP2. According to the results, LCP2 showed a significant response in spleen towards all the stimuli at 12 h to 48 h post infection. Following the T cell receptor engagement to the immune stimulation in the spleen, growth factor receptor-bound protein 2 (Grb2)-related adapter protein 2 (GRAP2) binds to the LCP2 while making a complex with the phospholipase C (PLC) and it binds to linker of activation T cells (LAT1), stabilizing and activating the T cells downstream responses.  The LCP2 actively participating in the T-cell related immune responses and it can be identified as an immunologically important gene in Korean rockfish.