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. Author manuscript; available in PMC: 2021 Mar 14.
Published in final edited form as: Nat Cell Biol. 2020 Sep 14;22(10):1264–1275. doi: 10.1038/s41556-020-0575-z

Fig. 4: nPD-L1 as co-activator together with p-Y705-Stat3 transcriptionally activates GSDMC expression in response to hypoxia.

Fig. 4:

(a–d) GSDMC mRNA (a and c) and protein (b and d) levels in the indicated MDA-MB-231 stable transfectants were determined by RT-qPCR and immunoblotting, respectively. Data are shown as mean ± SD of n = 3 biologically independent experiments. Blots are representative of three independent experiments. N, normoxia; H, hypoxia. (e and f), GSDMC promoter luciferase reporter plasmids were transfected into the indicated MDA-MB-231 stable transfectants and assayed for luciferase reporter activities. Data are shown as mean ± SD of n = 3 biologically independent experiments. (g) GSDMC promoter luciferase reporter plasmids were co-transfected with PD-L1 and Stat3 in HEK293T cells and assayed for luciferase activities. Data are shown as mean ± SD of n = 3 biologically independent experiments. Pro-WT, wild-type GSDMC promoter; Pro-Mut, GSDMC promoter with predicted p-Y705-Stat3 binding site mutation (red). (h) Sequential ChIP-PCR analysis of interactions between PD-L1, p-Y705-Stat3, and GSDMC promoter. VEGF promoter which interacts with HIF1α and p-Y705-Stat3 under hypoxia as a positive control. The experiment was repeated three times with similar results. P, MDA-MB-231 parental cells; KO, MDA-MB-231-PD-L1-knockout cells. P values of all statistical analysis were determined by two-sided Student’s t-test. Statistical source data and unprocessed blots are provided in Source Data Fig. 4.