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Supplementary Table 1 shows the differential expression analysis of RNA sequencing data. A list of genes that are differentially expressed upon TGF-beta treatment as well as upon RUNX3-Knockdown are shown.

This file contains Supplementary Figures 1-8. Fig.S1 shows Characterization of TGF-β-dependent EMT. Fig.S2 shows an analysis of endogenous DNA damage accumulation in the presence of TGF-β. Fig.S3 shows the Upregulation of NOX4 and downregulation of NRF-dependent genes upon TGF-β exposure.Fig.S4 displays that RNA-sequencing analysis unveils spontaneous EMT upon RUNX3 knockdown. Fig.S5 shows the regulation of HMOX1 expression by RUNX3. Figure.S6 shows how BACH1-KD in RUNX3-depleted cells rescues the accumulation of DNA damage. Fig.S7 displays RUNX1 depletion results in DNA damage accumulation and senescence in the presence of TGF-β. Fig.S8 shows HMOX1 depletion results in heightened ROS production and senescence in the presence of TGF-β.

Supplementary Methods, Legends for Supplementary Figures,Legend for Table 1 and References

Adaptive evolution to cellular stress is a process implicated in a wide range of biological and clinical phenomena. Two major routes of adaptation have been identified: non‐genetic changes, which allow expression of different phenotypes in novel environments, and genetic variation achieved by selection of fitter phenotypes. While these processes are broadly accepted, their temporal and epistatic features in the context of cellular evolution and emerging drug resistance are contentious. In this manuscript, we generated hypomorphic alleles of the essential nuclear pore complex (NPC) gene NUP58. By dissecting early and long‐term mechanisms of adaptation in independent clones, we observed that early physiological adaptation correlated with transcriptome rewiring and upregulation of genes known to interact with the NPC; long‐term adaptation and fitness recovery instead occurred via focal amplification of NUP58 and restoration of mutant protein expression. These data support the concept that early phenotypic plasticity allows later acquisition of genetic adaptations to a specific impairment. We propose this approach as a genetic model to mimic targeted drug therapy in human cells and to dissect mechanisms of adaptation., Generation of hypomorphic alleles of NUP58 in human cells using CRISPR‐Cas9 provides a proof‐of‐principle demonstration that early non‐genetic changes allow later acquisition of genetic adaptations to a specific impairment.

Summary: The Fanconi anemia (FA) pathway is a pivotal genome maintenance network that orchestrates the repair of DNA interstrand crosslinks (ICLs). The tumor suppressors RUNX1 and RUNX3 were shown to regulate the FA pathway independent of their canonical transcription activities, by controlling the DNA damage-dependent chromatin association of FANCD2. Here, in further biochemical characterization, we demonstrate that RUNX3 is modified by PARP-dependent poly(ADP-ribosyl)ation (PARylation), which in turn allows RUNX binding to DNA repair structures lacking transcription-related RUNX consensus motifs. SILAC-based mass spectrometric analysis revealed significant association of RUNX3 with core DNA repair complexes, including PARP1, even in unstressed cells. After DNA damage, the increased interaction between RUNX3 and BLM facilitates efficient FANCD2 chromatin localization. RUNX-Walker motif mutations from breast cancers are impaired for DNA damage-inducible PARylation, unveiling a potential mechanism for FA pathway inactivation in cancers. Our results reinforce the emerging paradigm that RUNX proteins are tumor suppressors with genome gatekeeper function. : Tay et al. demonstrate that the tumor suppressor genes RUNX1 and RUNX3 have an important regulatory role in the genome maintenance pathway controlled by FANCD2. DNA damage induces PARP-dependent PARylation of RUNX proteins, leading to their interaction with BLM to control the loading of FANCD2 on DNA damage sites. Keywords: RUNX1, RUNX3, Fanconi anemia, interstrand crosslink repair, PARP1, poly(ADP-ribosyl)ation, BLM, SILAC, DNA repair, FANCD2