Effects of human Werner helicase on intrachromosomal homologous recombination mediated DNA deletions in mice.
Werner syndrome (WS) is a rare genetic disorder characterized by accelerated aging and aging-related diseases including cancer. WS is caused by autosomal recessive mutations in the WRN gene, which is involved in genome maintenance although precise functions of WRN are not well understood. To further investigate the role of WRN, we used transgenic mice over-expressing a human helicase mutant WRN gene (hMW). We determined homologous recombination (HR) events leading to 70 kb deletions in the p(un) locus visualized as pigmented cells in the retinal pigment epithelium. hMW mice had an increased spontaneous frequency of DNA deletions compared to control mice, consistent with WRN involvement in HR suppression. In addition, 4-nitroquinoline 1-oxide (4-NQO), which can cause both oxidative stress and DNA adduct formation, significantly increased the frequency of DNA deletions in both control and hMW mice. In order to assess how oxidative stress may modulate this phenotype, we treated mice with the glutathione (GSH) synthesis inhibitor, buthionine sulfoximine (BSO). The frequency of DNA deletions increased significantly in control mice, but not in hMW littermates. To elucidate the cause of this discrepancy, we determined total GSH levels as a measure of anti-oxidative defense. BSO significantly decreased GSH levels in both hMW mice and control mice, while 4-NQO increased GSH levels in all mice. These findings suggest that the reduction of GSH by BSO or compensatory increase of GSH by 4-NQO had little impact on hMW mice in which HR repair is compromised. Therefore, oxidative stress impacts HR repair in hMW mice less than control mice and effects of the mutated gene may be exacerbated by direct DNA damage from 4-NQO. This mouse model of WS in conjunction with different DNA damaging agents may provide insight into mechanisms of genomic instability, DNA repair, and carcinogenesis.
Yamamoto
Mitsuko L
ML
Department of Pathology, Environmental Health and Radiation Oncology, David Geffen School of Medicine and School of Public Health, UCLA, 650 Charles E. Young Drive South, Los Angeles, CA 90024, USA.
Reliene
Ramune
R
Oshima
Junko
J
Schiestl
Robert H
RH
D015112:4-Nitroquinoline-1-oxide / Q000494:pharmacology; D000818:Animals; D019328:Buthionine Sulfoximine / Q000494:pharmacology; D004247:DNA / Q000235:genetics; D004249:DNA Damage; D004260:DNA Repair; D004195:Disease Models, Animal; D005090:Exodeoxyribonucleases / Q000235:genetics* / Q000378:metabolism*; D005260:Female; D042822:Genomic Instability; D005978:Glutathione / Q000378:metabolism; D006801:Humans; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D018384:Oxidative Stress; D010857:Pigment Epithelium of Eye / Q000378:metabolism; D011247:Pregnancy; D053484:RecQ Helicases / Q000235:genetics* / Q000378:metabolism*; D011995:Recombination, Genetic; D017384:Sequence Deletion; D014898:Werner Syndrome / Q000235:genetics / Q000378:metabolism; D000071657:Werner Syndrome Helicase
D019328:Buthionine Sulfoximine; D015112:4-Nitroquinoline-1-oxide; D004247:DNA; D005090:Exodeoxyribonucleases; D053484:RecQ Helicases; C098785:WRN protein, human; D000071657:Werner Syndrome Helicase; D005978:Glutathione