Enhanced DNA accessibility and increased DNA damage induced by the absence of poly(ADP-ribose) hydrolysis
Yiran Zhou1, Xiaoxing Feng, David W Koh
1Department of Pharmaceutical Sciences, College of Pharmacy, Washington State University, Pullman, Washington 99164, USA.
Abstract:
Poly(ADP-ribose) (PAR) is a therapeutic target primarily identified through inhibiting its synthesis by PAR polymerase-1 (PARP-1). However, inhibiting its hydrolysis by PAR glycohydrolase (PARG) has therapeutic potential in cancer. Unknown is the effect of elevated PAR levels on cellular processes and if this effect can enhance the therapeutic value of PARG. Here, we demonstrate in PARG null embryonic trophoblast stem (TS) cells that the absence of PAR hydrolysis led to PAR-modified histones H1, H2A, and H2B. To determine if this led to the differential vulnerability of DNA to modification, TS cells were treated with DNA-modifying agents. The results demonstrate increased DNA laddering by micrococcal nuclease and an increased amount of DNA intercalation by acridine orange in PARG null-TS cells. This increased access to PARG null-TS cell DNA was further verified by the detection of increased DNA damage following treatment with UV radiation and a minimal dose of the DNA-alkylating agent N-methyl-N'-nitro-N-nitrosoguanidine. Further, this DNA damage was predominantly unrepaired 12 h after treatment in PARG null-TS cells. Finally, TS cells were treated with DNA-modifying chemotherapeutic agents. The results demonstrate up to 4-fold increases in cell death in PARG null-TS cells after treatment with epirubicin or sub-IC(50) doses of cisplatin and cyclophosphamide. Taken together, we provide compelling evidence that increased DNA access induced by the absence of PARG enhances the efficacy of DNA-modifying agents. Thus, this study demonstrates that greater DNA accessibility, increased DNA damage, and increased cell death all contribute to the PARG null cell phenotype in response to genotoxic stress.
Insights
Inhibiting PAR glycohydrolase (PARG) increases DNA accessibility and damage in cells lacking PARG. This enhances cancer chemotherapy efficacy by increasing cell death from DNA-modifying agents.
Area of Science:
- Cellular Biology
- Biochemistry
- Genetics
Background:
- Poly(ADP-ribose) (PAR) is a therapeutic target, with PAR polymerase-1 (PARP-1) inhibition being a common strategy.
- Inhibiting PAR hydrolysis by PAR glycohydrolase (PARG) presents a novel therapeutic avenue, particularly in cancer treatment.
- The cellular effects of elevated PAR levels and their potential to enhance PARG-based therapies remain largely unexplored.
Purpose of the Study:
- To investigate the impact of absent PAR hydrolysis on cellular processes in embryonic trophoblast stem (TS) cells.
- To determine if elevated PAR levels enhance the therapeutic efficacy of DNA-modifying agents.
- To elucidate the role of PARG in DNA accessibility and repair.
Main Methods:
- Generated PARG null embryonic trophoblast stem (TS) cells to study the absence of PAR hydrolysis.
- Treated TS cells with DNA-modifying agents, UV radiation, and chemotherapeutic drugs.
- Assessed DNA accessibility, DNA damage, DNA repair, and cell death.
- Quantified PAR-modified histones (H1, H2A, H2B).
Main Results:
- PARG null TS cells exhibited PAR-modified histones H1, H2A, and H2B.
- Absence of PAR hydrolysis led to increased DNA laddering and intercalation, indicating greater DNA accessibility.
- PARG null TS cells showed increased DNA damage following UV radiation and N-methyl-N'-nitro-N-nitrosoguanidine treatment, with predominantly unrepaired damage.
- PARG null TS cells demonstrated up to 4-fold increases in cell death when treated with epirubicin, cisplatin, and cyclophosphamide.
Conclusions:
- The absence of PARG leads to increased DNA accessibility and susceptibility to damage.
- Elevated PAR levels, due to inhibited hydrolysis, enhance the efficacy of DNA-modifying chemotherapeutic agents.
- Targeting PARG could be a viable strategy to improve cancer therapy outcomes by increasing genotoxic stress sensitivity.
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