FIGURE 2 from A Druggable FOXA1-Glucocorticoid Receptor Transcriptional Axis Drives Tumor Growth in a Subset of Non-Small Cell Lung Cancer

Identification of a FOXA1–GR interaction through endogenous purification of FOXA1-associated proteins using RIME. A, Schematic overview of the RIME assay. B, Graphic representation of GR protein sequence coverage of unique peptides identified by LC/MS-MS in H441, HCC-44, and H3255 FOXA1 RIME samples. Yellow regions of string represent alignment of peptide sequences to the Mascot database sequences for GR protein. Full list found in Supplementary Table S2. C, Graphic representation of FOXA1 and GR protein sequence coverage as in B) in H441 GR RIME samples. D, Co-IP of FOXA1 and GR in FOXA1-dependent NSCLC cell lines. GR was immunoprecipitated in full serum-containing media and GR, FOXA1, and GAPDH were immunoblotted in the whole-cell lysate (input), IgG IP, and GR IP fractions. E, Volcano plot displaying significantly upregulated (red) or downregulated (green) genes in H2009 doxycycline-inducible shFOXA1 cells compared with shRenilla control cells following 72 hours of 1 μg/mL doxycycline treatment. F, Gene ontology pathway analysis of significant differentially regulated genes between H2009 shFOXA1 cells and H2009 shRenilla control cells. G, qPCR analysis of fold change in FOXA1, SGK1, and DUSP1 mRNA levels in H441 shFOXA1 cells normalized to H441 shRenilla cells treated with 1 μg/mL doxycycline for 48 hours. Values correspond to the mean of three replicates ± SEM. P value calculated by Student two-tailed t test (P < 0.01).