title: Damage-specific DNA binding protein 1 (DDB1) is involved in ubiquitin-mediated proteolysis of p27Kip1 in response to UV irradiation.
abstract: Damage-specific DNA binding protein 1 (DDB1) is a conserved protein component of the damaged DNA binding protein complex (DDB) that recognizes UV-induced DNA lesions and initiates the nucleotide excision repair process. DDB1 is also part of an E3 ubiquitin-ligase complex that targets a variety of substrates for proteolysis including the cyclin-dependent kinase inhibitor p27(Kip1). The mechanism regulating the trafficking of DDB1 and its relationship with UV irradiation is not known, although cell cycle progression is implicated in the molecular machinery driving DDB1 into the nucleus. We evaluated the involvement of DDB1 in ubiquitination of the cdk inhibitor p27(Kip1) in response to UV irradiation. First, we observed that low and high doses of UV irradiation exert different effects on p27(Kip1) protein levels. Indeed, low but not high UV doses induced p27(Kip1) protein proteolysis in several human cell lines and UV-dependent degradation is dominant over other genotoxic agents such as cisplatin. We also demonstrate that p27(Kip1) reduction is not due to transcriptional regulation and that the proteasome inhibitor MG132 affects p27(Kip1) degradation. We observed that at low UV doses the decrease in p27(Kip1) nuclear protein related with DDB1 translocation into the nucleus; conversely, high doses of UV-induced p27(Kip1) accumulation and unchanged level of DDB1. The knockdown of DDB1 or Skp2 prevents UV-induced degradation of p27(Kip1) suggesting that DDB1 is essential to regulation of p27(kip1) turnover after a mild DNA damage. Our findings support the concept that DDB1 contributes to the activation of DNA repair mechanisms and could be a key factor in regulating the cell cycle in response to UV-induced DNA damage. Although the temporal order with which DDB1 contributes to ubiquitination of p27(Kip1) or initiates the nucleotide excision repair process remains to be established, our results represent a major step towards clarifying these issues.
authors.affiliation: Dipartimento di Biochimica e Biotecnologie Mediche, Università di Napoli Federico II, Via Sergio Pansini 5, 80131 Napoli, Italy.
authors.forename: Barbara
authors.initials: B
authors.lastname: Iovine
authors.forename: Maria Luigia
authors.initials: ML
authors.lastname: Iannella
authors.forename: Maria Assunta
authors.initials: MA
authors.lastname: Bevilacqua
mesh_terms: D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D002843:Chromatin / Q000378:metabolism; D050760:Cyclin-Dependent Kinase Inhibitor p27 / Q000378:metabolism*; D004249:DNA Damage; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D015536:Down-Regulation; D006801:Humans; D046911:Macromolecular Substances / Q000378:metabolism; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D011485:Protein Binding; D021381:Protein Transport / Q000528:radiation effects; D044786:S-Phase Kinase-Associated Proteins / Q000378:metabolism; D014158:Transcription, Genetic / Q000528:radiation effects; D025801:Ubiquitin / Q000378:metabolism*; D044767:Ubiquitin-Protein Ligases / Q000378:metabolism; D014466:Ultraviolet Rays / Q000009:adverse effects*; D000097642:GADD45 Proteins
chemical_list: D018797:Cell Cycle Proteins; D002843:Chromatin; D050760:Cyclin-Dependent Kinase Inhibitor p27; D004268:DNA-Binding Proteins; D046911:Macromolecular Substances; D009687:Nuclear Proteins; D046988:Proteasome Endopeptidase Complex; D044786:S-Phase Kinase-Associated Proteins; D025801:Ubiquitin; D044767:Ubiquitin-Protein Ligases; C436291:CDKN1B protein, human; C091435:DDB1 protein, human; C117669:DDB2 protein, human; C439070:GADD45A protein, human; D000097642:GADD45 Proteins