Redundancy between nucleases required for homologous recombination promotes PARP inhibitor resistance in the eukaryotic model organism Dictyostelium

Abstract

ADP-ribosyltransferases promote repair of DNA single strand breaks and disruption of this pathway by Poly(ADP-ribose) polymerase (PARP) inhibitors (PARPi) is toxic to cells with defects in homologous recombination (HR). Here, we show that this relationship is conserved in the simple eukaryote Dictyostelium and exploit this organism to define mechanisms that drive resistance of the HR-deficient cells to PARPi. Dictyostelium cells disrupted in exonuclease I, a critical factor for HR, are sensitive to PARPi. Deletion of exo1 prevents the accumulation of Rad51 in chromatin induced by PARPi, resulting in DNA damage being channelled through repair by non-homologous end-joining (NHEJ). Inactivation of NHEJ supresses the sensitivity of exo1− cells to PARPi, indicating this pathway drives synthetic lethality and that in its absence alternative repair mechanisms promote cell survival. This resistance is independent of alternate-NHEJ and is instead achieved by re-activation of HR. Moreover, inhibitors of Mre11 restore sensitivity of dnapkcs−exo1− cells to PARPi, indicating redundancy between nucleases that initiate HR can drive PARPi resistance. These data inform on mechanism of PARPi resistance in HR-deficient cells and present Dictyostelium as a convenient genetic model to characterize these pathways.

Dictyostelium discoideum as a Model to Assess Genome Stability Through DNA Repair

“…Treatment of D. discoideum with the PARP inhibitor NU1025 leads to chromatin recruitment of the HR protein Rad51, indicating that, similar to humans, DNA damage induced by PARP inhibitors is repaired by HR. Consistent with this model, strains lacking the nuclease Exo1 that are defective in HR, show synthetic lethality with PARP inhibitors (Kolb et al, 2017). The conservation of this phenomenon with humans makes D. discoideum an attractive system to identify mutations which lead to resistance of HR-deficient strains to PARP inhibitors and therefore potentially cause resistance of tumors to PARP inhibitor treatment.…”

“…For example, this approach demonstrated that resistance to PARP inhibitors observed by disrupting NHEJ in an exo1 − background (dnapkcs − exo1 − cells) is mediated by promoting end-resection to reactivate HR. Small molecule inhibitors of Mre11, a second nuclease involved in strand resection during HR, restores sensitivity of dnapkcs − exo1 − cells to PARP inhibitors (Kolb et al, 2017). This highlights redundancy between the nucleases Exo1 and Mre11 in HR and suggests combination approaches to avoid resistance in cells deficient in one nuclease.…”

Linking DNA repair and cell cycle progression through serine ADP-ribosylation of histones

“…The h3b − , adprt1a − adprt2 − , ku80 − and e xo1 − strains have been previously described 35 , 37 , 38 , 42 , 45 . The Flag-Rad51 expression plasmid has been previously described 42 .…”

Dictyostelium discoideum as a pharmacological model system to study the mechanisms of medicinal drugs and natural products

“…The role of cisplatin on gene expression in the model has been analysed (Van Driessche et al, 2007), in addition to its regulation of mitogen activated protein kinase phosphatases (Moncho-Amor et al, 2011) and in DNA repair pathways (Gunn et al, 2016). The cellular functions of other anti-cancer treatments, such as bestatin and Poly (ADPribose) polymerase inhibitors have also been analysed in D. discoideum (Kolb et al, 2017, Poloz et al, 2012. Finally, Dictyostelium strains have also been shown to provide a source of novel compounds with anti-proliferative properties that could thus be used as cancer treatments (Honma et al, 2018, Kubohara and.…”