Fig. 5: Identification of deacetylation sites in XBP1s.

a HEK293T cells were transfected with the indicated plasmids, and total cell lysates were immunoprecipitated with anti-XBP1s antibodies and immunoblotted with anti-Ac-K and anti-Sirt6 antibodies. b Primary hepatocytes isolated from WT and KO mice were treated with Tm (2 μg/ml) and subjected to a co-immunoprecipitation assay for XBP1s deacetylation. c HEK293T cells were transfected with XBP1s with p300 or XBP1s with p300 and Sirt6. The acetylation of XBP1s was determined using LC-MS/MS analysis after in-gel digestion. MS/MS spectra of acetylated XBP1s peptides containing Lys257 (left) and Lys297 (right). d HEK293T cells were transfected with wild type (WT) or mutant XBP1s (K257R and K297R) and then treated with CHX (20 μg/ml) for the indicated times. Relative protein levels of XBP1s were compared (n = 3). e C57BL/6 mice were injected intravenously with 1 × 10⁹ pfu of AdLacZ, AdXBP1s-WT, or AdXBP1s-2KR, and hepatic TG levels were analyzed (n = 4–5). The values are the mean ± SEM. *p < 0.05 and **p < 0.01 versus WT or AdLacZ; ^##p < 0.01 versus AdXBP1s-WT