RUNX Poly(ADP-Ribosyl)ation and BLM Interaction Facilitate the Fanconi Anemia Pathway of DNA Repair

Abstract

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.

RUNX transcription factors: orchestrators of development

“…CBFβ acts as a non-DNA-binding regulatory element, which allosterically increases the DNA-binding affinity and stability of the complex by interacting with RUNX through the RHD (Bravo et al, 2001;Huang et al, 2001;Tang et al, 2000;Yan et al, 2004). Of note, CBFβindependent functions of RUNX have also been suggested in the literature (Bresciani et al, 2014), including a non-transcriptional role for RUNX1 and RUNX3 in the Fanconi anemia DNA-repair pathway (Tay et al, 2018;Wang et al, 2014b). Usually regarded as weak transcription factors by themselves, RUNX proteins might have the potential to act as 'pioneer' transcription factors, which are able to engage condensed chromatin to facilitate its opening, and promote the recruitment of other transcriptional regulators (Zaret and Carroll, 2011).…”

“…Likewise, premature senescence was induced in murine embryonic fibroblasts by individually overexpressing one of the three RUNX genes (Kilbey et al, 2007). Studies in Runx1/Runx3 double knockout (DKO) mice have exposed functional redundancy between these genes in the Fanconi anemia DNA-repair pathway, independent of their transcriptional role (Tay et al, 2018;Wang et al, 2014b). Finally, Morita and colleagues have shown compensatory mechanisms between the three RUNX factors in the context of leukemia (Morita et al, 2017).…”

“…These mice die within 25 weeks of induction of the DKO phenotype, as a result of either bone marrow failure (BMF) or myeloproliferative disorders. Such seemingly contradictory phenotypes are reminiscent of Fanconi anemia, an inherited BMF syndrome caused by defective DNA repair, in which RUNX1/3 play crucial roles (Tay et al, 2018;Wang et al, 2014b). The reduction of all mature hematopoietic lineage compartments observed in DKO mice appears to be responsible for the BMF phenotype, which occurs in spite of the HSPC expansion (Wang et al, 2014b).…”

Targeting dePARylation for cancer therapy

“…PAR chains due to the negatively charged phosphates of ADPR bring a lot of anionic charges to the damaged chromatin and the negative charges alter the chemical and biological properties of the acceptor proteins. The acceptor protein of PARylation includes histones (H1, H2A, and H2B) [41,42], DNA protein kinases [43,44], p53 [45], Ku complex [3,46], DNA glycosylase 8-oxoguanine glycosylase 1 (OGG1) [47], PCNA [48], RUNX [49], etc. Since DNA is negatively charged; charge repulsion between PAR and DNA modulates the chromatin structure at the damaged loci.…”

PrimPol primase mediates replication traverse of DNA interstrand crosslinks

“…A quantitative proteomic analysis (enrichment ratio and statistical significance of proteins identified in WT vs KO cells) confirmed the efficient IP of PrimPol and the co-precipitation of known interacting factors RPA and SSBP1 (Figure 1B). Amongst the new potential PrimPol-interacting proteins were HERC2, involved in the resolution of G4 quadruplex structures (Wu et al, 2018) and several proteins linked to the recognition and repair of ICL lesions: MHF1 (part of the FANCM complex; Ciccia et al, 2007;Yan et al, 2010;Wang et al, 2013); RUNX1 (Tay et al, 2018); and BLM, RMI1 and RMI2 (three components of the BTR complex; Manthei and Keck, 2013). Both MHF1 and BTR participate in the ICL traverse reaction Ling et al, 2016).…”