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| Antitumor drug adozelesin differentially affects active and silent origins of DNA replication in yeast checkpoint kinase mutants. | ||
| Y Wang, T A Beerman, D Kowalski | ||
| Cancer Res (2001), 61(9):3787-94 | ||
| The antitumor drug adozelesin is a potent cytotoxic DNA-damaging agent. Here we determined how adozelesin affects chromosomal DNA replication at a molecular level in a yeast model system and examined the influence of checkpoint kinase genes, the human homologues of which are mutated in cancer. Analysis of replication intermediates using two-dimensional gel electrophoresis showed that adozelesin inhibited the activity of a replication origin and stalled replication fork progression through chromosomal DNA at the origin. RAD53 and MEC1 protein kinase genes, homologues of human CHK2 and ATM, respectively, regulate an intra-S-phase DNA damage checkpoint and, when mutated, permit unchecked replication of damaged DNA in S-phase. Mutations in these genes did not abrogate adozelesin-induced inhibition of origin activity and fork progression at the replication origin. However, novel replication intermediates indicative of DNA breaks were detected only in the rad53 mutant, suggesting a role for the wild-type gene in maintaining chromosome integrity in the presence of the drug. In contrast to the inhibition of the active replication origin by adozelesin, normally silent origins present in the same chromosome were activated by adozelesin in rad53 and mec1 mutant cells. Thus, an antitumor drug that damages DNA can induce an abnormal replication pattern in a chromosome by activating silent origins, depending upon defects in yeast checkpoint kinase genes, the homologues of which are mutated in cancer. Implications of an abnormal replication pattern for the epigenetic regulation of gene expression are discussed. | ||
OriDB annotation of this paper: |
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Origin site(s) analysed by ARS assay: |
None curated. | |
Origin site(s) analysed by 2D gel analysis: |
ARS303, ARS320, ARS305. | |
Predicted ACS element: |
None curated. | |
Experimentally confirmed ACS element: |
None curated. | |
Activation ('firing') time determined: |
None curated. | |
Activation ('firing') efficiency determined: |
None curated. | |
Analysed by microarray: |
None curated. | |
Other analysis: |
None curated. | |
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