The relative expression of mutated XPB genes results in xeroderma pigmentosum/Cockayne's syndrome or trichothiodystrophy cellular phenotypes

Abstract

The human XPB DNA helicase is a subunit of the DNA repair/basal transcription factor TFIIH, involved in early steps of the nucleotide excision repair pathway. Two distinct clinical phenotypes, xeroderma pigmentosum associated with Cockayne's syndrome (XP/CS) and trichothiodystrophy (TTD), can be due to mutations in the XPB gene. In the present work, we studied cellular DNA repair properties of skin fibro-blasts from two patients mutated in the XPB gene: an XP/CS patient cell (XPCS2BA) with a T296C (F99S) transition and a TTD patient cell (TTD6VI) exhibiting an A355C (T119P) transversion. Both cells are clearly associated with different levels of alterations in their response to UV light. To establish the relationship between the relative expression level of these two alleles and DNA repair properties, we transfected SV40-transformed XPCS2BA (XPCS2BASV) cells with a plasmid (pTTD6VI) carrying the XPB-A355C cDNA and examined DNA repair properties after UV irradiation (cell survival, unscheduled DNA synthesis and kinetics of photoproduct removal) in stable transfectants. We isolated three clones, which express the XPB-A355C gene (Cl-5) or the XPB-T296C gene (Cl-14) or both genes (Cl-19). This con-stitutes a model system allowing us to correlate the relative expression levels of the XPB-A355C (TTD) and XPB-T296C (XP/CS) genes with various DNA repair properties. Overexpression of the XPB-A355C (TTD) gene in an XP/CS cell gives rise to a cellular phenotype of increased repair similar to that of TTD6VI cells, while equal expression of the two mutated genes leads to an intermediate cellular phenotype between XP/CS and TTD.