title: Poly(ADP-ribose)-mediated interplay of XPA and PARP1 leads to reciprocal regulation of protein function.
abstract: Poly(ADP-ribose) (PAR) is a complex and reversible post-translational modification that controls protein function and localization through covalent modification of, or noncovalent binding to target proteins. Previously, we and others characterized the noncovalent, high-affinity binding of the key nucleotide excision repair (NER) protein XPA to PAR. In the present study, we address the functional relevance of this interaction. First, we confirm that pharmacological inhibition of cellular poly(ADP-ribosyl)ation (PARylation) impairs NER efficacy. Second, we demonstrate that the XPA-PAR interaction is mediated by specific basic amino acids within a highly conserved PAR-binding motif, which overlaps the DNA damage-binding protein 2 (DDB2) and transcription factor II H (TFIIH) interaction domains of XPA. Third, biochemical studies reveal a mutual regulation of PARP1 and XPA functions showing that, on the one hand, the XPA-PAR interaction lowers the DNA binding affinity of XPA, whereas, on the other hand, XPA itself strongly stimulates PARP1 enzymatic activity. Fourth, microirradiation experiments in U2OS cells demonstrate that PARP inhibition alters the recruitment properties of XPA-green fluorescent protein to sites of laser-induced DNA damage. In conclusion, our results reveal that XPA and PARP1 regulate each other in a reciprocal and PAR-dependent manner, potentially acting as a fine-tuning mechanism for the spatio-temporal regulation of the two factors during NER.
authors: Molecular Toxicology Group, Department of Biology, University of Konstanz, Germany; Konstanz Research School Chemical Biology, University of Konstanz, Germany.
authors: Jan M F
authors: JM
authors: Fischer
authors: Oliver
authors: O
authors: Popp
authors: Daniel
authors: D
authors: Gebhard
authors: Sebastian
authors: S
authors: Veith
authors: Arthur
authors: A
authors: Fischbach
authors: Sascha
authors: S
authors: Beneke
authors: Alfred
authors: A
authors: Leitenstorfer
authors: Jörg
authors: J
authors: Bergemann
authors: Martin
authors: M
authors: Scheffner
authors: Elisa
authors: E
authors: Ferrando-May
authors: Aswin
authors: A
authors: Mangerich
authors: Alexander
authors: A
authors: Bürkle
mesh_terms: D000595:Amino Acid Sequence; D016384:Consensus Sequence; D004260:DNA Repair; D006367:HeLa Cells; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D000071137:Poly (ADP-Ribose) Polymerase-1; D011064:Poly Adenosine Diphosphate Ribose / Q000378:metabolism*; D011065:Poly(ADP-ribose) Polymerases / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D011499:Protein Processing, Post-Translational; D061987:Sf9 Cells; D051760:Xeroderma Pigmentosum Group A Protein / Q000737:chemistry / Q000378:metabolism*
chemical_list: C497314:XPA protein, human; D051760:Xeroderma Pigmentosum Group A Protein; D011064:Poly Adenosine Diphosphate Ribose; C499320:PARP1 protein, human; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases
keywords: ARTD1; host cell reactivation assay; laser microirradiation; nucleotide excision repair; xeroderma pigmentosum