Nonhomologous end-joining promotes resistance to DNA damage in the absence of an ADP-ribosyltransferase that signals DNA single strand breaks

Abstract

Summary ADP-ribosylation of proteins at DNA lesions by ADP-ribosyltransferases (ARTs) is an early response to DNA damage. The best defined role of ADP-ribosylation in the DNA damage response is in repair of single strand breaks (SSBs). Recently, we initiated a study of how ADP-ribosylation regulates DNA repair in Dictyostelium and found that two ARTs (Adprt1b and Adprt2) are required for tolerance of cells to SSBs, and a third ART (Adprt1a) promotes nonhomologous end-joining (NHEJ). Here we report that disruption of adprt2 results in accumulation of DNA damage throughout the cell cycle following exposure to agents that induce base damage and DNA SSBs. Although ADP-ribosylation is evident in adprt2 -cells exposed to methylmethanesulfonate (MMS), disruption of adprt1a and adprt2 in combination abolishes this response and further sensitises cells to this agent, indicating that in the absence of Adprt2, Adprt1a signals MMS-induced DNA lesions to promote resistance of cells to DNA damage. As a consequence of defective signalling of SSBs by Adprt2, Adprt1a is required to assemble NHEJ factors in chromatin, and disruption of the NHEJ pathway in combination with adprt2 increases sensitivity of cells to MMS. Taken together, these data indicate overlapping functions of different ARTs in signalling DNA damage, and illustrate a critical requirement for NHEJ in maintaining cell viability in the absence of an effective SSB response.

The role of ADP-ribosylation in regulating DNA interstrand crosslink repair

“…This is based on our observations that (1) substantial nuclear ADP-ribosylation is observed in response to the ICL-inducing agent cisplatin, (2) enrichment of APL in chromatin in response to cisplatin is dependent on its macrodomain and the ART Adprt2, and (3) the adprt2 − strain is sensitive to cisplatin. Our data in Dictyostelium (Couto et al, 2013, 2011; Pears et al, 2012), in addition to those of others in vertebrates (Gibson and Kraus, 2012), implicate ARTs in repair of SSBs and DSBs. Therefore, it is possible that ARTs are detecting these or similar DNA architectures following processing of cisplatin-induced DNA damage, rather than the ICL directly.…”

“…Similar to in humans, two Dictyostelium ARTs (Adprt1b and Adprt2) are required for tolerance of cells to DNA SSBs, whereas a third ART (Adprt1a) is required to promote NHEJ of DNA DSBs (Couto et al, 2013, 2011). We therefore considered whether any of these ARTs are similarly required for the cellular response to cisplatin.…”

“…Our previous work indicates that loss of Adprt2 results in increased DNA DSBs following exposure of cells to DNA alkylating agents and that this is subsequently signalled by Adprt1a to promote NHEJ and cell survival in response to these genotoxins (Couto et al, 2013). Given the redundancy between Adprt1a and Adprt2 in signalling cisplatin-induced DNA damage, we considered whether similar mechanisms are being employed in response to this variety of DNA damage.…”

“…However, although Adprt2 is required for tolerance to DNA SSBs, the enrichment of APL in chromatin, an event that is dependent on Adprt2, does not occur in response to canonical base damage induced by MMS or 4-NQO. Similarly, no gross enrichment of APL is observed in chromatin following DNA DSBs, and the Adprt2-null strain is not sensitive to agents that induce this variety of DNA damage (Couto et al, 2013). Therefore, we believe Adprt2-mediated ADP-ribosylation induced by cisplatin is not induced by these DNA damage types directly, or if so, it is in the context of these DNA structures being produced as a consequence of DNA ICL processing.…”

The Role of PARPs in DNA Strand Break Repair

“…This suggests that Adprt1a may be a functional homologue of PARP3. In contrast, D. discoideum Adprt1b and Adprt2 have overlapping functions with PARP1 and PARP2, and primarily appear to function in SSBR [169,170].…”