Processing of targeted psoralen cross-links in Xenopus oocytes

David Segal, A. Fawad Faruqi, Peter M. Glazer, Dana Carroll

Research output: Contribution to journalArticle

7 Scopus citations

Abstract

Psoralen cross-links have been shown to be both mutagenic and recombinagenic in bacterial, yeast, and mammalian cells. Double-strand breaks (DSBs) have been implicated as intermediates in the removal of psoralen cross-links. Recent work has suggested that site-specific mutagenesis and recombination might be achieved through the use of targeted psoralen adducts. The fate of plasmids containing psoralen adducts was evaluated in Xenopus oocytes, an experimental system that has well-characterized recombination capabilities and advantages in the analysis of intermediates in DNA metabolism. Psoralen adducts were delivered to a specific site by a triplex- forming oligonucleotide. These lesions are dearly recognized and processed in oocytes, since mutagenesis was observed at the target site. The spectrum of induced mutations was compared with that found in similar studies in mammalian cells. Plasmids carrying multiple random adducts were preferentially degraded, perhaps due to the introduction of DSBs. However, when DNAs carrying site-specific adducts were examined, no plasmid loss was observed and removal of cross-links was found to be very slow. Sensitive assays for DSB-dependent homologous recombination were performed with substrates with one or two cross-link sites. No adduct-stimulated recombination was observed with a single lesion, and only very low levels were observed with paired lesions, even when a large proportion of the cross- links was removed by the oocytes. We conclude that DSBs or other recombinagenic structures are not efficiently formed at psoralen adducts in Xenopus oocytes. While psoralen is not a promising reagent for stimulating site-specific recombination, it is effective in inducing targeted mutations.

Original languageEnglish (US)
Pages (from-to)6645-6652
Number of pages8
JournalMolecular and Cellular Biology
Volume17
Issue number11
StatePublished - Nov 1997
Externally publishedYes

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics
  • Cell Biology

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  • Cite this

    Segal, D., Fawad Faruqi, A., Glazer, P. M., & Carroll, D. (1997). Processing of targeted psoralen cross-links in Xenopus oocytes. Molecular and Cellular Biology, 17(11), 6645-6652.