Fig. 3
From: In vivo screening identifies GATAD2B as a metastasis driver in KRAS-driven lung cancer
GATAD2B drives KRAS-dependent HBEC tumor growth and metastasis. a Immunoblot analysis of total and activated KRAS (via RAF1 pulldown assay) extracted from HBEC cells expressing GFP or KRASG12D in the presence or absence of Dox. b Differentially expressed mRNAs in HBECs engineered with Dox-inducible alleles encoding wild-type (WT) or mutationally activated (G12D) KRAS (n = 3 each) in the presence or absence of Dox. c Transwell invasion assay of HBEC-iKRASG12D cells stably expressing GFP or GATAD2B in the presence or absence of Dox (**p < 0.01, 2 way- ANOVA; n = 3 each). Representative images of transwell chambers shown in bottom panel. d Tumor growth curves illustrating the failure of HBEC-iKRASG12D tumor formation in the absence of KRASG12D induction [Off Dox; GFP (Orange), GATAD2B (Black)], rapid tumor formation in GATAD2B (Red) vs. GFP control (Blue) in the presence of Dox, and the ability to toggle KRAS expression from On to Off (On > Off) to demonstrate requirement of KRAS for tumor maintenance in GATAD2B (Purple) vs. GFP control (Green) tumors. e Confirmation of human GATAD2B overexpression (left, 200 μm scale bar) and enhanced KI67 staining (right, 100 μm scale bar) via immunohistochemicial analysis of HBEC-iKRASG12D SQ tumors expressing GATAD2B vs. GFP from d. f Number of barcode sequencing reads from metastatic tissues (lymph node, lung) isolated from mice harboring HBEC-iKRASG12D-GATAD2B SQ tumors (On Dox) from d. NGS barcode sequencing was performed to quantitate presence of GATAD2B-specific barcode. Note that no metastases were observed in mice harboring iKRASG12D-GATAD2B SQ tumors, thus barcode sequencing was not performed. g Anchorage-independent colony formation assays for KRAS mutant (H23, A549, CALU-1) and KRAS wild-type (H1437, H1568) NSCLC cells (N = 3). shNT = Non-targeting control
