Fig. 5: Selective autophagic degradation of PROCR via p62 is inhibited under irradiation.
From: PROCR diminishes the efficacy of radiation by impairing T-cell-mediated antitumour immunity
A Flow cytometry analysis showing PROCR expression of EpCAM+ cells from fresh NPC tissues, treated with or without irradiation (n = 5 independent experiments). B Flow cytometry analysis showing PROCR expression of HONE1 cells after treatment of radiation for the indicated time intervals. C RT-qPCR detection of PROCR mRNA expression of HONE1 cells after treatment of radiation for the indicated time intervals. D Immunoblots (left) and corresponding greyscale analysis (right) of PROCR in HONE1 cells, treated with CHX for the indicated times, with or without exposure to irradiation. E PROCR protein level in HONE1 cells treated with DMSO, MG132, CQ and EBSS, with or without radiation. F Immunofluorescence showing localization of PROCR (green) relative to LC3B (red) positive autophagosomes in HONE1 cells, with or without radiation. Graph shows the percentage co-localization (n = 15 fields). Scale, 10μm. The boxplots indicate the median (centre), 25th, and 75th percentiles (box boundaries), and minimum and maximum (the whiskers). G Coimmunoprecipitation (Co-IP) and immunoblot analysis of HONE1 cells transfected with MYC-tagged PROCR together with FLAG-tagged TOLLIP, SQSTM1, NDP52, NBR1, TAX1BP1, BNIP3L or OPTN. H Cell lysates of HONE1 cells were subjected to immunoprecipitation with IgG or anti-p62 antibody. Immunoblot analysis with indicated antibodies were shown. I Flow cytometry analysis showing PROCR expression of parental (sgGFP) HONE1 cells, p62-knockout (sgSQSTM1) HONE1 cells, and p62-knockout HONE1 cells subjected to radiation treatment (sgSQSTM1 + IR). J Flow cytometry analysis showing PROCR expression of parental (sgGFP) HONE1 cells, p62-knockout (sgSQSTM1) cells, p62-rescue (sgSQSTM1 + SQSTM1-OE) cells and p62-rescue cells subjected to radiation treatment (sgSQSTM1 + SQSTM1-OE + IR). K RT-qPCR detection of IL-6 mRNA expression of parental (sgGFP) and p62-knockout (sgSQSTM1) HONE1 cells. n = 3 independent experiments (B–K). One of three experiments is shown (D, E, G, H). Data are shown as mean ± SD (B–D, I, J, K). P values were calculated using paired t test (A), two-tailed unpaired t test (F, K), one-way ANOVA (B, C, I, J), or two-way ANOVA (D). Source data are provided as a Source Data file.
