| Products/Services Used | Details | Operation |
|---|---|---|
| Custom Vector Construction | Signaling. Pulldown GST protein protein interaction assay The coding region of the LCK-SH3 domain (183 bp) was cloned into the pcDNA3.1(+)-N-GST vector (GenScript). The resulting vector expresses a fusion protein of LCK-SH3 with GST at the N terminus. Similarly, the coding region of the LCK-LR domain (60 bp), including the coding region of the LCK-LR domain (60 bp), including the WT sequence or mutants K228R and K228Q, was cloned into the pcDNA3.1(+)-N-eGFP vector (GenScript). The resultant vectors express fusion proteins of LCK-LR whether WT or mutants with GFP at the N terminus. Each vector was transfected into 293T cells 3000 Transfection Reagent (Thermo Fisher Scientific). The 293T cell lysates expressing LCK-SH3-GST fusion proteins were applied to glutathione resin (GenScript). The glutathione resin, bound with SH3-GST, was washed five times before the introduction of 293T cell lysates expressing either Empty or LCK-LR (WT, | Get A Quote |
T cell receptor (TCR) signaling is precisely tuned to prevent self-reactivity while allowing protective immunity. Here we found that acetylation modulated TCR signaling. The loss of SIRT2 deacetylase activity in T cells led to amplified calcium mobilization and phosphorylation of key proximal TCR molecules in naive T cells and reversed dampened TCR signaling in anergic T cells. During thymic selection, SIRT2 deficiency lowered the TCR signaling threshold and resulted in a broader TCR repertoire diversity. Mechanistically, we identified acetyl-lysine K228 on the linker region of LCK as a substrate specific for SIRT2 that governed LCK conformation and activity. SIRT2 inhibition in exhausted mouse and human tumor-... More
