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Replication focus-forming activity 1 and the Werner syndrome gene product

Nature Genetics volume 19, pages 375–378 (1998)Cite this article

Abstract

The initiation of DNA replication involves a minimum of four factors: a specific DNA sequence (origin), an initiator protein which binds to the origin, a helicase that unwinds the origin and a protein that binds single-stranded DNA that stabilizes the unwound origin1. In eukaryotic cells, the origin recognition complex2 (ORC) is the initiator protein and replication protein A (RPA; ref. 3) is the single-stranded DNA-binding protein. However, the helicase has not been identified and the nature of origins remains elusive, except in the case of Saccharomyces cerevisiae. A unique feature of eukaryotic DNA replication is that it occurs at a few-hundred discrete foci4. It has thus been proposed that a real origin must contain a specific DNA sequence and must be attached to replication foci. Using Xenopus laevis egg extracts, we have identified and purified a 170-kD protein, focus-forming activity 1 (FFA-1), which is required for the formation of replication foci5. Here we report that FFA-1 has DNA-helicase activity. Moreover, it is a homologue of the human Werner syndrome gene product WRN, a protein associated with premature ageing in humans6.

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Figure 1: ATP-dependent formation of RPA foci.
Figure 2: DNA-dependent ATPase activity of FFA-1.
Figure 3: DNA helicase activity of FFA-1.
Figure 4: Co-purification of ATPase, helicase, FFA-1 and a 170-kD protein.
Figure 5: Distribution, immunodepletion and immunofluorescence staining of p170.
Figure 6: Sequence alignment of FFA-1 and the human Werner syndrome protein (hWRN) performed by the Macvector program (Clustal).

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Acknowledgements

This research was supported by a grant from the National Institute of Health to J.N. and a grant from the V Foundation to H.Y.

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Authors and Affiliations

  1. Fox Chase Cancer Center, W450, 7701 Burholme Avenue, Philadelphia, 19111, Pennsylvania, USA

    Hong Yan & Chin-Yi Chen

  2. Cold Spring Harbor Laboratory, Cold Spring Harbor, 11724, New York, USA

    Ryuji Kobayashi

  3. University of California at San Diego, Dept of Biology, La Jolla, 92093-0347, California, USA

    John Newport

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  1. Hong Yan
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  4. John Newport
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Correspondence to Hong Yan.

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Yan, H., Chen, CY., Kobayashi, R. et al. Replication focus-forming activity 1 and the Werner syndrome gene product . Nat Genet 19, 375–378 (1998). https://doi.org/10.1038/1263

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