Abstract
Exposure to ultraviolet radiation (UVR) promotes the formation of UVR-induced, DNA helix distorting photolesions such as (6-4) pyrimidine-pyrimidone photoproducts and cyclobutane pyrimidine dimers. Effective repair of such lesions by the nucleotide excision repair (NER) pathway is required to prevent DNA mutations and chromosome aberrations. Poly(ADP-ribose) polymerase-1 (PARP-1) is a zinc finger protein with well documented involvement in base excision repair. PARP-1 is activated in response to DNA damage and catalyzes the formation of poly(ADP-ribose) subunits that assist in the assembly of DNA repair proteins at sites of damage. In this study, we present evidence for PARP-1 contributions to NER, extending the knowledge of PARP-1 function in DNA repair beyond the established role in base excision repair. Silencing the PARP-1 protein or inhibiting PARP activity leads to retention of UVR-induced photolesions. PARP activation following UVR exposure promotes association between PARP-1 and XPA, a central protein in NER. Administration of PARP inhibitors confirms that poly(ADP-ribose) facilitates PARP-1 association with XPA in whole cell extracts, in isolated chromatin complexes, and in vitro. Furthermore, inhibition of PARP activity decreases UVR-stimulated XPA chromatin association, illustrating that these relationships occur in a meaningful context for NER. These results provide a mechanistic link for PARP activity in the repair of UVR-induced photoproducts.
Highlights
Little is known regarding the role of Poly(ADP-ribose) polymerase-1 (PARP-1) in ultraviolet radiation (UVR)-induced photolesion repair
A 60% reduction in PARP-1 protein levels was detected by Western blotting (Fig. 1, A and B), which corresponded to a 60% reduction in basal PAR levels (Fig. 1C, normalized to the untreated (NT)) and a 70% reduction in PARP activation following a single dose of 3 kJ/m2 solar-simulated UVR (Fig. 1C, 1h)
To expand on the above findings and examine the role of PARP activity during the repair process, the PARP inhibitor DPQ [27, 28] was used. This inhibitor significantly decreased UVR-stimulated PARP activity as measured by PAR production (Fig. 2A) and resulted in retention of 6-4 PPs (Fig. 2B) and cyclobutane pyrimide dimers (CPDs) (Fig. 2C). These findings demonstrate that reduction of PARP activity, either by PARP-1 silencing or chemical inhibition, promotes retention of UVR-induced photolesions
Summary
Little is known regarding the role of PARP-1 in UVR-induced photolesion repair. Results: PARP inhibitors decrease PARP-1-XPA associations and reduce chromatin binding of XPA. Inhibition of PARP activity decreases UVR-stimulated XPA chromatin association, illustrating that these relationships occur in a meaningful context for NER These results provide a mechanistic link for PARP activity in the repair of UVR-induced photoproducts. Solar UVR exposure forms DNA photoproducts such as cyclobutane pyrimide dimers (CPDs) and [] pyrimidine-pyrimidone photoproducts (6-4 PPs), which are helix distorting lesions repaired predominantly by nucleotide excision repair (NER) [2, 3] If such lesions are retained, they may lead to mutations, chromosome aberrations, and cellular malfunctions including cell death, senescence, and cancer [4, 5]. These results confirm a role for PARP-1 in NER but suggest a mechanistic link for PARP activity in the repair of UVR-induced photoproducts
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