Fig. 1: GSDMD can be modified by ubiquitination.
From: E3 ubiquitin ligase SYVN1 is a key positive regulator for GSDMD-mediated pyroptosis
A Lysates from HEK293T cells co-transfected with HA-Ub along with Flag-human GSDMD (hGSDMD) or not were subjected to immunoprecipitation with Flag antibody followed by immunoblotting using an anti-HA antibody. B Immunoprecipitation and immunoblotting of HEK293T cells co-transfected with HA-Ub and Flag-porcine GSDMD (pGSDMD) using anti-Flag and anti-HA antibodies, respectively. C Immunoprecipitation and immunoblotting of HEK293T cells co-transfected with HA-Ub and Flag-mouse GSDMD (mGSDMD) using anti-Flag and anti-HA antibodies, respectively. D, F THP-1 cells were primed with LPS (500 ng/ml) for 4 h, and then treated with or without Nigericin (10 μM). Cell lysates were immunoprecipitated with anti-GSDMD antibody, followed by immunoblotting with anti-GSDMD and anti-Ub antibodies, respectively. Secretion of IL-1β was analyzed using ELISA. LDH release was analyzed using an LDH release assay. E, G THP-1 cells were incubated with Pam3CSK4 (500 ng/ml) for 4 h, and then transfected with or without LPS (2 μg/ml) for 6 h. Cell lysates were immunoprecipitated with anti-GSDMD antibody, followed by immunoblotting with anti-GSDMD and anti-Ub antibodies. IL-1β secretion in THP-1 cells was analyzed using ELISA, whereas LDH release was analyzed using LDH release assay. All results shown are representative of at least three independent experiments.
