Kinetics of poly(ADP-ribosyl)ation, but not PARP1 itself, determines the cell fate in response to DNA damage in vitro and in vivo

Abstract

One of the fastest cellular responses to genotoxic stress is the formation of poly(ADP-ribose) polymers (PAR) by poly(ADP-ribose)polymerase 1 (PARP1, or ARTD1). PARP1 and its enzymatic product PAR regulate diverse biological processes, such as DNA repair, chromatin remodeling, transcription and cell death. However, the inter-dependent function of the PARP1 protein and its enzymatic activity clouds the mechanism underlying the biological response. We generated a PARP1 knock-in mouse model carrying a point mutation in the catalytic domain of PARP1 (D993A), which impairs the kinetics of the PARP1 activity and the PAR chain complexity in vitro and in vivo, designated as hypo-PARylation. PARP1D993A/D993A mice and cells are viable and show no obvious abnormalities. Despite a mild defect in base excision repair (BER), this hypo-PARylation compromises the DNA damage response during DNA replication, leading to cell death or senescence. Strikingly, PARP1D993A/D993A mice are hypersensitive to alkylation in vivo, phenocopying the phenotype of PARP1 knockout mice. Our study thus unravels a novel regulatory mechanism, which could not be revealed by classical loss-of-function studies, on how PAR homeostasis, but not the PARP1 protein, protects cells and organisms from acute DNA damage.

PARP1 inhibition alleviates injury in ARH3-deficient mice and human cells

“…The PARP1 inhibitor veliparib, partially attenuated oxidative stress-induced cell death. More than 90% of PAR is generated by PARP1 activation (43). While the PAR accumulation in cytoplasm is responsible for cell death under oxidative stress in ARH3-deficient mice and in the patient, we speculate that PARP1 inhibition may have therapeutic potential.…”

The Enigmatic Function of PARP1: From PARylation Activity to PAR Readers

“…Homozygous PARP1-D993A mutant mice showed no obvious phenotypic alterations, but were hypersensitive to alkylating agents and died six days after intraperitoneal injection of the MNU (N-methyl-N-nitroso urea); the wild-type control did not show any sign of disease. Despite a significant delay of PARylation after DNA damage, the homozygous PARP1-D993A mutant cells showed only a mild impairment of BER [64]. The conclusion drawn from this is that although neither one of PARP1 protein itself and its complete activity is necessary for the development, the kinetics of PARylation and the complexity of PAR are important for the final outcome of biological response to genotoxic stress in vivo [64].…”

“…Despite a significant delay of PARylation after DNA damage, the homozygous PARP1-D993A mutant cells showed only a mild impairment of BER [64]. The conclusion drawn from this is that although neither one of PARP1 protein itself and its complete activity is necessary for the development, the kinetics of PARylation and the complexity of PAR are important for the final outcome of biological response to genotoxic stress in vivo [64].…”

Coordination of DNA Base Excision Repair by Protein-Protein Interactions

“…Complexes of PAR-PARP1 with damaged DNA have been detected by atomic force microscopy [72]. Recently, kinetics of poly(ADP-ribosyl)ation and PAR homeostasis (but not the PARP1 protein) have been proposed to play a primary role in protection of cells from acute DNA damage [73]. Hence, the formation of BER complexes on the damaged DNA can be regulated via either poly(ADP-ribosyl)ation of proteins or their interactions with PAR polymer synthesized by PARP1 and PARP2.…”