Fig 2: NLRP3 blockade prevents PKD auto-phosphorylation.(A) Immunoblot analysis of whole cell lysates from THP-1 cells treated with nigericin for the indicated times ± MCC950 (1 µM), AFN700 (3 µM) or CGP084892 (20 µM), monitoring total PKD3 as well as auto-phosphorylated PKD and gasdermin D cleavage. (B) Immunoblot analysis of whole cell lysates from THP-1-WT or NLRP3-KO cells treated with nigericin for the indicated times, auto-phosphorylated PKD, as well as full and N-terminally cleaved gasdermin D.

Fig 3: PKD3 expression is positively correlated with expression of FASN and mSREBP1 in prostate cancer tissues. Tissue microarray slides containing 80 prostate cancer samples, 8 adjacent normal prostate tissue and 8 prostate tissue were subjected to IHC analysis for PKD3, FASN and mSREBP1. A. Representative images depicted at 400x magnification for PKD3 and FASN antibody staining of different Gleason scores are shown (left panels). Spearman rank test were used to study the correlations between PKD3 and FASN expression. (r=0.580, p<0.001) (right panels). B. Representative images depicted at 400x magnification for PKD3 and mSREBP1 antibody staining of different Gleason scores are shown (left panels), Spearman rank test were used to study the correlations between PKD3 and mSREBP1 expression. (r=0.381, p<0.001) (right panels).

Fig 4: PKD3 overexpression enhances IFN-?-induced PD-L1 expression in DOK cells. (A and B) The levels of PKD3 and PD-L1 protein were examined by western blot analysis and immunofluorescence. The PKD3 overexpression plasmid was transfected into DOK cells using Lipofectamine 2000. (C and D) Cells transfected with the PKD3 overexpression plasmid and cells transfected with control shRNA were cultured in the presence or absence of IFN-? (20 ng/ml) for 24 h. The expression levels of PKD3 and PD-L1 were determined by western blot analysis. In addition, the cell surface expression of PD-L1 was analyzed by flow cytometry. Data are presented as the means ± SD (n=3). ***P<0.001. PKD3, protein kinase D3; PD-L1, programmed death ligand-1; IFN-?, interferon-?.

Fig 5: IFN-? induces the activation of related factors regulating PD-L1 expression in OSCC cells. (A) Cancer cells were stimulated with IFN-? (20 ng/ml) for the indicated periods of time. The activation of related factors regulating PD-L1 expression was analyzed by western blot analysis. (B) The line graphs show densitometric analysis of changes in the abundance of phosphorylated substrates and total proteins normalized to GAPDH to account for loading variability. (C) PD-L1 mRNA expression was assayed by RT-qPCR. The cells were treated as described in (A). Data are presented as the means ± SD (n=3). IFN-?, interferon-?; PKD3, protein kinase D3; PD-L1, programmed death ligand-1.