TGF-ß promotes genomic instability after loss of RUNX3

Studies of genomic instability have historically focused on intrinsic mechanisms rather than extrinsic mechanisms based in the tumor microenvironment (TME). TGF-ß is the most abundantly secreted cytokine in the TME where it imparts various aggressive characteristics including invasive migration, drug resistance and epithelial-to-mesenchymal transition (EMT). Here we show that TGF-ß also promotes genomic instability in the form of DNA double strand breaks (DSB) in cancer cells which lack the tumor suppressor gene RUNX3. Loss of RUNX3 resulted in transcriptional downregulation of the redox regulator heme oxygenase-1 (HO-1 or HMOX1). Consequently, elevated oxidative DNA damage disrupted genomic integrity and triggered cellular senescence, which was accompanied by tumor-promoting inflammatory cytokine expression and acquisition of the senescence-associated secretory phenotype (SASP). Recapitulating the above findings, tumors harbouring a TGF-ß gene expression signature and RUNX3 loss exhibited higher levels of genomic instability. In summary, RUNX3 creates an effective barrier against further TGF-ß-dependent tumor progression by preventing genomic instability. These data suggest a novel cooperation between cancer cell-extrinsic TGF-ß signaling and cancer cell-intrinsic RUNX3 inactivation as aggravating factors for genomic instability. Overall design: A549 cells were seeded into 6-cm dishes at the cell density of 0.2X106 cells/dish and transfected with control siRNA or RUNX3 siRNA. 3 days later, cells were trypsinized and subjected to a second round of siRNA. 24 hours later, cells were exposed to either vehicle control or TGF-ß for 48 hours followed by RNA sequencing. C1 and C2 samples are biological replicates of control SiRNA transfection; C-1TGF and C-2TGF are biological replicates following TGF-ß for 48 hours. R1 and R2 samples are biological replicates following RUNX3 SiRNA; R-1TGF and R-2TGF are biological replicates following TGF-ß for 48 hours of cells transfected with RUNX3 siRNA.