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Inhibition of poly(ADP-ribose) polymerase activity is insufficient to induce tetraploidy
C M Simbulan-Rosenthal1, D S Rosenthal, R Luo
1Department of Biochemistry and Molecular Biology, Georgetown University School of Medicine, 3900 Reservoir Road NW, Washington, DC 20007, USA.
Abstract:
Poly(ADP-ribose) polymerase (PARP) knockout mice are resistant to murine models of human diseases such as cerebral and myocardial ischemia, traumatic brain injury, diabetes, Parkinsonism, endotoxic shock and arthritis, implicating PARP in the pathogenesis of these diseases. Potent selective PARP inhibitors are therefore being evaluated as novel therapeutic agents in the treatment of these diseases. Inhibition or depletion of PARP, however, increases genomic instability in cells exposed to genotoxic agents. We recently demonstrated the presence of a genomically unstable tetraploid population in PARP(-/-) fibroblasts and its loss after stable transfection with PARP cDNA. To elucidate whether the genomic instability is attributable to PARP deficiency or lack of PARP activity, we investigated the effects of PARP inhibition on development of tetraploidy. Immortalized wild-type and PARP(-/-) fibroblasts were exposed for 3 weeks to 20 microM GPI 6150 (1,11b-dihydro-[2H:]benzopyrano[4,3,2-de]isoquinolin-3-one), a novel small molecule specific competitive inhibitor of PARP (K(i) = 60 nM) and one of the most potent PARP inhibitors to date (IC(50) = 0.15 microM). Although GPI 6150 initially decreased cell growth in wild-type cells, there was no effect on cell growth or viability after 24 h. GPI 6150 inhibited endogenous PARP activity in wild-type cells by approximately 91%, to about the residual levels in PARP(-/-) cells. Flow cytometric analysis of unsynchronized wild-type cells exposed for 3 weeks to GPI 6150 did not induce the development of tetraploidy, suggesting that, aside from its catalytic function, PARP may play other essential roles in the maintenance of genomic stability.
Insights
Poly(ADP-ribose) polymerase (PARP) inhibition does not cause genomic instability in wild-type cells. This suggests PARP has roles beyond its catalytic function in maintaining genomic stability.
Area of Science:
- Molecular Biology
- Genetics
- Biochemistry
Background:
- Poly(ADP-ribose) polymerase (PARP) knockout mice show resistance to various diseases, implicating PARP in disease pathogenesis.
- PARP inhibitors are explored as therapeutics, but PARP inhibition can increase genomic instability.
- Previous studies linked PARP deficiency to genomic instability and tetraploidy in fibroblasts.
Purpose of the Study:
- To determine if genomic instability observed in PARP deficiency is due to lack of PARP activity or its catalytic function.
- To investigate the effect of PARP inhibition on tetraploidy development in wild-type cells.
Main Methods:
- Wild-type and PARP(-/-) fibroblasts were treated with GPI 6150, a potent PARP inhibitor.
- Cells were exposed to the inhibitor for 3 weeks.
- Flow cytometry was used to analyze the development of tetraploidy.
Main Results:
- GPI 6150 effectively inhibited endogenous PARP activity in wild-type cells.
- Long-term exposure to GPI 6150 did not induce tetraploidy in wild-type fibroblasts.
- PARP inhibition did not replicate the genomic instability seen in PARP-deficient cells.
Conclusions:
- Genomic instability in PARP deficiency may not solely result from the loss of catalytic activity.
- PARP might possess additional functions crucial for maintaining genomic stability.
- Further research is needed to understand PARP's non-catalytic roles in genome integrity.