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HUMAN MUTATION, vol.31, no.2, pp.167-175, 2010 (SCI-Expanded)
Mutations in two branch-point sequences (BPS) in intron 3 of the XPC DNA repair gene affect pre-mRNA splicing in association with xeroderma pigmentosum (XP) with many skin cancers (X-P101TMA) or no skin cancer (XT72TMA), respectively. To investigate the mechanism of these abnormalities we now report that transfection of minigenes with these mutations revealed abnormal XPC pre-mRNA splicing that mimicked pre-mRNA splicing in the patients' cells. DNA oligonucleotide-directed RNase H digestion demonstrated that mutations in these BPS disrupt U2 snRNP-BPS interaction. XP101TMA cells had no detectable XTC protein but XT72TMA had 29% of normal levels. A small amount of XTC protein was detected at sites of localized ultraviolet (UV),damaged DNA in XT72TMA cells which then recruited other nucleotide excision repair (NER) proteins. In contrast, XP101TMA cells had no detectable recruitment of XTC or other NER proteins. Post-UV survival and photoproduct assays revealed greater reduction in DNA repair in XP101TMA cells than in XT72TMA. Thus mutations in XPC BPS resulted in disruption of U2 snRNPBPS interaction leading to abnormal pre-mRNA splicing and reduced XPC protein. At the cellular level these changes were associated with features of reduced DNA repair including diminished NER protein recruitment reduced post-UV survival and impaired photoproduct removal. Hum Mutat 31:167-175, 2010. Published 2009 Wiley-Liss, Inc.