Abstract
Cockayne syndrome (CS) is a human disease characterized by sensitivity to sunlight, severe neurological abnormalities, and accelerated aging. CS has two complementation groups, CS-A and CS-B. The CSB gene encodes the CSB protein with 1493 amino acids. We previously reported that the CSB protein is involved in cellular repair of 8-hydroxyguanine, an abundant lesion in oxidatively damaged DNA and that the putative helicase motif V/VI of the CSB may play a role in this process. The present study investigated the role of the CSB protein in cellular repair of 8-hydroxyadenine (8-OH-Ade), another abundant lesion in oxidatively damaged DNA. Extracts of CS-B-null cells and mutant cells with site-directed mutation in the motif VI of the putative helicase domain incised 8-hydroxyadenine in vitro less efficiently than wild type cells. Furthermore, CS-B-null and motif VI mutant cells accumulated more 8-hydroxyadenine in their genomic DNA than wild type cells after exposure to gamma-radiation at doses of 2 or 5 Gy. These results suggest that the CSB protein contributes to cellular repair of 8-OH-Ade and that the motif VI of the putative helicase domain of CSB is required for this activity.
Highlights
Cockayne syndrome (CS) is a human disease characterized by sensitivity to sunlight, severe neurological abnormalities, and accelerated aging
We investigated the possibility that CSB is involved in cellular repair of 8-OH-Ade, a major lesion in oxidatively damaged DNA. 8-OH-Ade repair was assessed and compared in wild type, CS-B-null, and putative CS-B helicase motif VI mutant cells
The present study shows that CS-B-null and motif VI mutant cells are deficient in incision of 8-OH-Ade
Summary
Vol 277, No 34, Issue of August 23, pp. 30832–30837, 2002 Printed in U.S.A. The Cockayne Syndrome Group B Gene Product Is Involved in Cellular Repair of 8-Hydroxyadenine in DNA*. We previously reported that the CSB protein is involved in cellular repair of 8-hydroxyguanine, an abundant lesion in oxidatively damaged DNA and that the putative helicase motif V/VI of the CSB may play a role in this process. The present study investigated the role of the CSB protein in cellular repair of 8-hydroxyadenine (8-OHAde), another abundant lesion in oxidatively damaged DNA. CS-B cells have a pronounced defect in repair of UV radiationinduced DNA damage in actively transcribed genes [4]. CS-B cells may be deficient in other processes besides TCR of UV radiation-induced DNA damage. We investigated the role of CSB in cellular repair of 8-OH-Ade. CSB mutants were expressed in CS-Bnull cells and in vitro incision of 8-OH-Ade was analyzed using WCEs from these cell lines. Mass spectrometry (LC/MS) with the isotope dilution technique [17]
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