Journal of Molecular Biology
Volume 186, Issue 1, 5 November 1985, Pages 129-136
Journal home page for Journal of Molecular Biology

Origin of adenovirus DNA replication: Role of the nuclear factor I binding site in vivo

https://doi.org/10.1016/0022-2836(85)90263-3Get rights and content

Abstract

An assay is described that detects in vivo a single round of initiation and DNA synthesis directed by a linear molecule containing an exposed single copy of an adenovirus (Ad) origin of replication. This and a previously described assay, which measures multiple rounds of DNA replication, were used to identify DNA sequences within the Ad2 and Ad4 origins of replication that are important for ori function. Linear DNA molecules containing sequences from the Ad2 or Ad4 genome termini were cotransfected with homologous and heterologous helper virus, and net amounts of DNA synthesis were compared. Linear molecules containing the Ad4 inverted terminal repeats were replicated 20-fold better in the presence of the homologous helper, whereas both Ad2 and Ad4 inverted terminal repeats were utilized efficiently by Ad4. DNA sequence analysis of the Ad2 ori and the corresponding region in Ad4 indicated that, although there are only ten variant base-pairs, eight are located within the Ad2 DNA sequence recognized by the cellular protein nuclear factor I. This protein is required to achieve the maximal rate of Ad2 DNA replication in vitro, and these differences therefore identify DNA sequences that are crucial to Ad2 ori function. The Ad4 ITR does not contain a functional nuclear factor I binding site, and deletion analysis has demonstrated that this region of the Ad4 genome is not required for ori function. In contrast to Ad2, the DNA sequences required for the initiation of Ad4 DNA replication were shown to reside entirely within the terminal 18 base-pairs of the Ad4 inverted terminal repeat.

References (45)

  • P. Aleström et al.

    Gene

    (1982)
  • J.R. Arrand et al.

    J. Mol. Biol

    (1979)
  • E.A. Carusi

    Virology

    (1977)
  • S.V. Desiderio et al.

    J. Mol. Biol

    (1981)
  • G. Geier et al.

    J. Biol. Chem

    (1979)
  • D. Hanahan

    J. Mol. Biol

    (1983)
  • R.T. Hay et al.

    Cell

    (1982)
  • R.T. Hay et al.

    J. Mol. Biol

    (1984)
  • R.L. Lechner et al.

    Cell

    (1977)
  • D.R. Rawlins et al.

    Cell

    (1984)
  • D.M.K. Rekosh et al.

    Cell

    (1977)
  • A.J. Robinson et al.

    Virology

    (1973)
  • F. Sanger et al.

    J. Mol. Biol

    (1980)
  • U. Siebenlist et al.

    Cell

    (1984)
  • E.M. Southern

    J. Mol. Biol

    (1975)
  • B.W. Stillman et al.

    Cell

    (1981)
  • B.W. Stillman et al.

    Cell

    (1982)
  • M. Temple et al.

    Virology

    (1981)
  • O. Tokunaga et al.

    Gene

    (1982)
  • J. Vieira et al.

    Gene

    (1982)
  • E. Wimmer

    J. Mol. Biol

    (1972)
  • M.D. Biggin et al.
  • Cited by (0)

    Present address: Department of Biochemistry and Microbiology, St Andrews University, Fife KY16 9AL, Scotland.

    View full text