Abstract
The gene responsible for Werner's syndrome (WRN) contains a region homologous to the Escherichia coli RecQ type DNA helicase and was thought to code for a DNA helicase belonging to this helicase family. However, no evidence has been shown before to substantiate this prediction. Here, we show data that the product of the WRN gene is indeed a DNA helicase. The gene product, a polypeptide with a relative molecular mass of 170 kDa, expressed in the insect Spodoptera frugiperda (Sf21) cell and purified by affinity column chromatography contained both the ATPase and DNA unwinding activities characteristic of DNA helicase. Expressions in Sf21, as well as in HeLa cells, showed that the WRN DNA helicase is exclusively transported to the nucleoplasm, which is consistent with its function in DNA metabolism. Our studies on strand displacement suggest that WRN helicase can unwind not only a duplex DNA, but also an RNA-DNA heteroduplex, while the latter reaction seems less efficient. Enzymological features learned from the purified WRN helicase are discussed with respect to the biological function, which remains to be clarified.
Full Text
The Full Text of this article is available as a PDF (253.9 KB).
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Ellis N. A., Groden J., Ye T. Z., Straughen J., Lennon D. J., Ciocci S., Proytcheva M., German J. The Bloom's syndrome gene product is homologous to RecQ helicases. Cell. 1995 Nov 17;83(4):655–666. doi: 10.1016/0092-8674(95)90105-1. [DOI] [PubMed] [Google Scholar]
- Ellis N. A. Mutation-causing mutations. Nature. 1996 May 9;381(6578):110–111. doi: 10.1038/381110a0. [DOI] [PubMed] [Google Scholar]
- Epstein C. J., Martin G. M., Schultz A. L., Motulsky A. G. Werner's syndrome a review of its symptomatology, natural history, pathologic features, genetics and relationship to the natural aging process. Medicine (Baltimore) 1966 May;45(3):177–221. doi: 10.1097/00005792-196605000-00001. [DOI] [PubMed] [Google Scholar]
- Goto M., Horiuchi Y., Tanimoto K., Ishii T., Nakashima H. Werner's syndrome: analysis of 15 cases with a review of the Japanese literature. J Am Geriatr Soc. 1978 Aug;26(8):341–347. doi: 10.1111/j.1532-5415.1978.tb03681.x. [DOI] [PubMed] [Google Scholar]
- Goto M., Imamura O., Kuromitsu J., Matsumoto T., Yamabe Y., Tokutake Y., Suzuki N., Mason B., Drayna D., Sugawara M. Analysis of helicase gene mutations in Japanese Werner's syndrome patients. Hum Genet. 1997 Feb;99(2):191–193. doi: 10.1007/s004390050336. [DOI] [PubMed] [Google Scholar]
- Goto M., Miller R. W., Ishikawa Y., Sugano H. Excess of rare cancers in Werner syndrome (adult progeria). Cancer Epidemiol Biomarkers Prev. 1996 Apr;5(4):239–246. [PubMed] [Google Scholar]
- Goto M., Tanimoto K., Horiuchi Y., Sasazuki T. Family analysis of Werner's syndrome: a survey of 42 Japanese families with a review of the literature. Clin Genet. 1981 Jan;19(1):8–15. doi: 10.1111/j.1399-0004.1981.tb00660.x. [DOI] [PubMed] [Google Scholar]
- Irino N., Nakayama K., Nakayama H. The recQ gene of Escherichia coli K12: primary structure and evidence for SOS regulation. Mol Gen Genet. 1986 Nov;205(2):298–304. doi: 10.1007/BF00430442. [DOI] [PubMed] [Google Scholar]
- Lohman T. M. Helicase-catalyzed DNA unwinding. J Biol Chem. 1993 Feb 5;268(4):2269–2272. [PubMed] [Google Scholar]
- Lombard D. B., Guarente L. Cloning the gene for Werner syndrome: a disease with many symptoms of premature aging. Trends Genet. 1996 Aug;12(8):283–286. doi: 10.1016/0168-9525(96)30065-6. [DOI] [PubMed] [Google Scholar]
- Ma L., Siemssen E. D., Noteborn H. M., van der Eb A. J. The xeroderma pigmentosum group B protein ERCC3 produced in the baculovirus system exhibits DNA helicase activity. Nucleic Acids Res. 1994 Oct 11;22(20):4095–4102. doi: 10.1093/nar/22.20.4095. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Matsumoto T., Imamura O., Yamabe Y., Kuromitsu J., Tokutake Y., Shimamoto A., Suzuki N., Satoh M., Kitao S., Ichikawa K. Mutation and haplotype analyses of the Werner's syndrome gene based on its genomic structure: genetic epidemiology in the Japanese population. Hum Genet. 1997 Jul;100(1):123–130. doi: 10.1007/s004390050477. [DOI] [PubMed] [Google Scholar]
- Mushegian A. R., Bassett D. E., Jr, Boguski M. S., Bork P., Koonin E. V. Positionally cloned human disease genes: patterns of evolutionary conservation and functional motifs. Proc Natl Acad Sci U S A. 1997 May 27;94(11):5831–5836. doi: 10.1073/pnas.94.11.5831. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Salk D., Au K., Hoehn H., Martin G. M. Cytogenetic aspects of Werner syndrome. Adv Exp Med Biol. 1985;190:541–546. doi: 10.1007/978-1-4684-7853-2_27. [DOI] [PubMed] [Google Scholar]
- Seki M., Enomoto T., Yanagisawa J., Hanaoka F., Ui M. Further characterization of DNA helicase activity of mouse DNA-dependent adenosinetriphosphatase B (DNA helicase B). Biochemistry. 1988 Mar 8;27(5):1766–1771. doi: 10.1021/bi00405a057. [DOI] [PubMed] [Google Scholar]
- Seki M., Yanagisawa J., Kohda T., Sonoyama T., Ui M., Enomoto T. Purification of two DNA-dependent adenosinetriphosphatases having DNA helicase activity from HeLa cells and comparison of the properties of the two enzymes. J Biochem. 1994 Mar;115(3):523–531. doi: 10.1093/oxfordjournals.jbchem.a124369. [DOI] [PubMed] [Google Scholar]
- Yu C. E., Oshima J., Fu Y. H., Wijsman E. M., Hisama F., Alisch R., Matthews S., Nakura J., Miki T., Ouais S. Positional cloning of the Werner's syndrome gene. Science. 1996 Apr 12;272(5259):258–262. doi: 10.1126/science.272.5259.258. [DOI] [PubMed] [Google Scholar]
- Yu C. E., Oshima J., Wijsman E. M., Nakura J., Miki T., Piussan C., Matthews S., Fu Y. H., Mulligan J., Martin G. M. Mutations in the consensus helicase domains of the Werner syndrome gene. Werner's Syndrome Collaborative Group. Am J Hum Genet. 1997 Feb;60(2):330–341. [PMC free article] [PubMed] [Google Scholar]