A mechanism for relief of replication blocks by activation of unused origins and age-dependent change in the caffeine susceptibility in xeroderma pigmentosum variant

https://doi.org/10.1016/0921-8777(91)90043-OGet rights and content

Abstract

The first XP16KO-I (age 42 without cancer) and the second XP16KO-II (age 52 after carcinogenesis) skin fibroblasts of a xeroderma pigmentosum variant (XPV) were studied. First, caffeine had no effect on XP16KO-I cells, but it potentiated UV killing and inhibited the recovery of DNA synthesis and the elongation of nascent DNA after UV irradiation in XP16KO-II cells, indicating an age-dependent change from caffeine resistance to sensitivity. These results confirmed a previous similar change in an unrelated XPV subject and the existence of 2 XPV subgroups, caffeine-sensitive and -resistant. Further, processing of post-excision breaks was also slightly more defective in XP16KO-II than in XP16KO-I cells. Second, nascent DNA pulse-labeled at 1.5 h after 5 J/m2 indicated an initial complete blockage of elongation by each dimer in XP16KO-II cells and less complete blocks in XP16KO-I and normal cells. The nascent DNA in 5 J/m2-irradiated XP16KO-II cells was 4 μm (8 × 106 DA) long for the peak fraction, with a possible range up to 20 μm of average replicon size, indicating activation of at least 4 μm-spaced new origins around 1.5 h post irradiation. Post-labeling chase without caffeine allowed nearly normal elongation to the present maximum molecular weight of 2.60–2.80 × 108 in 3–4 h in irradiated XP16KO-I and -II cells. Thus, the activation of unused origins at shorter spaces is a main mechanism for relief and the reduction of almost all unreplicated regions or blocks initially formed by 2 dimers in trans on both leading strands between converging forks. Post-labeling addition of 1 mM caffeine increased perpetuated blocks to a frequency of about 10% of the initial number of dimers in 4 h in XP16KO-II cells, but not in XP16KO-I and normal cells.

References (34)

  • W.K. Kaufmann et al.

    Mechanisms of inhibition of DNA replication by ultraviolet light in normal human and xeroderma pigmentosum fibroblasts

    J. Mol. Biol.

    (1981)
  • R.B. Painter

    Effect of caffeine on DNA synthesis in irradiated and unirradiated mammalian cells

    J. Mol. Biol.

    (1980)
  • R.B. Painter

    Inhibition and recovery of DNA synthesis in human cells after exposure to ultraviolet light

    Mutation Res.

    (1985)
  • S. Squires et al.

    Initial rates of DNA incision in UV-irradiated human cells: differences between normal, xeroderma pigmentosum and tumour cells

    Mutation Res.

    (1982)
  • S. Tornaletti et al.

    Studies on DNA binding of caffeine and derivatives: Evidence of intercalation by DNA-unwinding experiments

    Biochim. Biophys. Acta

    (1989)
  • C.A. Berger et al.

    Pyrimidine dimers block simian virus 40 replication forks

    Mol. Cell. Biol.

    (1986)
  • P.G. Burk et al.

    Ultraviolet-stimulated thymidine incorporation in xeroderma pigmentosum lymphocytes

    J. Lab. Clin. Med.

    (1971)
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