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XPC branch‐point sequence mutations disrupt U2 snRNP binding, resulting in abnormal pre‐mRNA splicing in xeroderma pigmentosum patients
Author(s) -
Khan Sikandar G.,
Yamanegi Koji,
Zheng ZhiMing,
Boyle Jennifer,
Imoto Kyoko,
Oh KyuSeon,
Baker Carl C.,
Gozukara Engin,
Metin Ahmet,
Kraemer Kenneth H.
Publication year - 2010
Publication title -
human mutation
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.981
H-Index - 162
eISSN - 1098-1004
pISSN - 1059-7794
DOI - 10.1002/humu.21166
Subject(s) - xeroderma pigmentosum , biology , rna splicing , nucleotide excision repair , microbiology and biotechnology , point mutation , dna repair , intron , messenger rna , mutation , dna damage , gene , dna , genetics , rna
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 (XP101TMA) or no skin cancer (XP72TMA), 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 XPC protein but XP72TMA had 29% of normal levels. A small amount of XPC protein was detected at sites of localized ultraviolet (UV)‐damaged DNA in XP72TMA cells which then recruited other nucleotide excision repair (NER) proteins. In contrast, XP101TMA cells had no detectable recruitment of XPC or other NER proteins. Post‐UV survival and photoproduct assays revealed greater reduction in DNA repair in XP101TMA cells than in XP72TMA. Thus mutations in XPC BPS resulted in disruption of U2 snRNP‐BPS 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 30:1–9, 2009. Published 2009 Wiley‐Liss, Inc.