Abstract

The ADP-ribosyl hydrolases PARG and ARH3 counteract PARP enzymatic activity by removing ADP-ribosylation. PARG and ARH3 activities have a synthetic lethal effect; however, the specific molecular mechanisms underlying this response remain unknown.

Here, we show that the PARG and ARH3 synthetic lethality is enhanced further in the presence of DNA alkylating agents, suggesting that the inability to revert ADP-ribosylation primarily affects the repair of alkylated DNA bases. ARH3 knockout cells, treated with PARG inhibitor and alkylating genotoxins, accumulated single-stranded DNA and DNA damage, resulting in G2/M cell cycle arrest and apoptosis.

Furthermore, we reveal a reduction in PARP1/PARP2 levels in ARH3-deficient cells treated with PARG inhibitor due to excessive ADP-ribosylation, which may contribute to alkylating agents’ vulnerability.

Collectively, these results uncover the potential of targeting ADP-ribosyl hydrolases in combination with alkylating agents for cancer therapy and provide insights into the mechanisms underlying the synthetic lethal effect.

Introduction

Poly(ADP-ribose) polymerases (PARPs) transfer single or multiple units of ADP-ribose from NAD to target proteins and nucleic acids, a process called ADP-ribosylation.

PARP1 and PARP2 (PARP1/2) play essential roles in maintaining genome stability. Mechanistically, DNA breaks activate PARP1/2, whose poly(ADP-ribosyl)ation (PARylation) represents one of the earliest events in response to single-strand breaks (SSBs), double-strand breaks (DSBs), and disturbed replication forks.

By ADP-ribosylating protein substrates, PARP1/2 trigger the recruitment and regulation of DNA repair enzymes and chromatin remodellers, thereby facilitating DNA repair. Accordingly, PARP inhibitors (PARPis) represent an established treatment strategy for ovarian, breast, pancreatic, prostate, and small-cell lung cancers harboring defects in DNA repair pathways.

After DNA damage, large amounts of serine-linked ADP-ribose (Ser-ADPr) and chains of ADP-ribose are formed. To synthesize Ser-ADPr, the accessory factor HPF1 forms a complex with PARP1 or PARP2 to catalyze the initial serine mono-ADP-ribose adduct, which can be further extended by PARP1/2 alone into poly(ADP-ribose) (PAR) chains.

Without HPF1, activated PARP1/2 can also attach ADP-ribose to other amino acids in proteins, such as glutamate, or target nucleic acids

This is one of the scientific articles published by one or more synbrAIn collaborators and data scientists.

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