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  • The EMBO Journal (2003) 22, 6148 - 6160
  • doi:10.1093/emboj/cdg576

Thymine-rich single-stranded DNA activates Mcm4/6/7 helicase on Y-fork and bubble-like substrates

Zhiying You1, Yukio Ishimi2, Takeshi Mizuno3,4, Kaoru Sugasawa3,4, Fumio Hanaoka3,4,5 and Hisao Masai1

  1. Department of Cell Biology, Tokyo Metropolitan Institute of Medical Science, 18-22 Honkomagome 3-chome, Bunkyo-ku, Tokyo 113-8613, Japan
  2. Biomolecular Science and Technology Department, Mitsubishi Kagaku Institute of Life Sciences, 11 Minamiooya, Machida, Tokyo 194-8511, Japan
  3. Cellular Physiology Laboratory, RIKEN (The Institute of Physical and Chemical Research), Hirosawa 2-1, Wako-shi, Saitama 351-0198, Japan
  4. CREST, Japan Science and Technology Corporation, Hirosawa 2-1, Wako-shi, Saitama 351-0198, Japan
  5. Graduate School of Frontier Biosciences, Osaka University, 1-3 Yamada-oka, Suita, Osaka 565-0871, Japan

Correspondence to:

Hisao Masai, E-mail: hmasai@rinshoken.or.jp

Received 30 July 2003; Accepted 25 September 2003; Revised 9 September 2003

The presence of multiple clusters of runs of asymmetric adenine or thymine is a feature commonly found in eukaryotic replication origins. Here we report that the helicase and ATPase activities of the mammalian Mcm4/6/7 complex are activated specifically by thymine stretches. The Mcm helicase is specifically activated by a synthetic bubble structure which mimics an activated replication origin, as well as by a Y-fork structure, provided that a single-stranded DNA region of sufficient length is present in the unwound segment or 3' tail, respectively, and that it carries clusters of thymines. Sequences derived from the human lamin B2 origin can serve as a potent activator for the Mcm helicase, and substitution of its thymine clusters with guanine leads to loss of this activation. At the fork, Mcm displays marked processivity, expected for a replicative helicase. These findings lead us to propose that selective activation by stretches of thymine sequences of a fraction of Mcm helicases loaded onto chromatin may be the determinant for selection of initiation sites on mammalian genomes.

  • Keywords:

    • initiation of DNA replication,
    • Mcm helicase,
    • replication bubble or fork,
    • replication origin,
    • thymine stretch