Replication initiation complex formation in the absence of nuclear function in Xenopus.

Liliana Krasinska (1) , Daniel Fisher (1)

1 IGMM - Institut de Génétique Moléculaire de Montpellier

In this article, we study how intercalation-induced changes in chromatin and DNA topology affect chromosomal DNA replication using Xenopus egg extracts. Unexpectedly, intercalation by ethidium or doxorubicin prevents formation of a functional nucleus: although nucleosome formation occurs, DNA decondensation is arrested, membranous vesicles accumulate around DNA but do not fuse to form a nuclear membrane, active transport is abolished and lamins are found on chromatin, but do not assemble into a lamina. DNA replication is inhibited at the stage of initiation complex activation, as shown by molecular combing of DNA and by the absence of checkpoint activation. Replication of single-stranded DNA is not prevented. Surprisingly, in spite of the absence of nuclear function, DNA-replication proteins of pre-replication and initiation complexes are loaded onto chromatin. This is a general phenomenon as initiation complexes could also be seen without ethidium in membrane-depleted extracts which do not form nuclei. These results suggest that DNA or chromatin topology is required for generation of a functional nucleus, and activation, but not formation, of initiation complexes.

Domaines: Génomique, Transcriptomique et Protéomique [q-bio.GN]

Liliana Krasinska, Daniel Fisher. Replication initiation complex formation in the absence of nuclear function in Xenopus.. Nucleic Acids Research, 2009, 37 (7), pp.2238-2248. ⟨10.1093/nar/gkp081⟩. ⟨hal-00368674⟩