Aphidicolin triggers a block to replication origin firing in Xenopus egg extracts. Export

@article{citeulike:1182014, abstract = {DNA replication origins are located at random with respect to DNA sequence in Xenopus early embryos and on DNA replicated in Xenopus egg extracts. We have recently shown that origins fire throughout the S phase in Xenopus egg extracts. To study the temporal regulation of origin firing, we have analyzed origin activation in sperm nuclei treated with the DNA polymerase inhibitor aphidicolin. Sperm chromatin was incubated in Xenopus egg extracts in the presence of aphidicolin and transferred to a fresh extract, and digoxigenin-dUTP and biotin-dUTP were added at various times after aphidicolin release to selectively label early and late replicating DNA. Molecular combing analysis of single DNA fibers showed that only a fraction of potential origins were able to initiate in the presence of aphidicolin. After release from aphidicolin, the remaining origins fired asynchronously throughout the S phase. Therefore, initiation during the S phase depends on the normal progression of replication forks assembled at earlier activated origins. Caffeine, an inhibitor of the checkpoint kinases ATR and ATM, did not relieve the aphidicolin-induced block to origin firing. We conclude that a caffeine-insensitive intra-S phase checkpoint regulates origin activation when DNA synthesis is inhibited in Xenopus egg extracts.}, address = {Laboratoire de G\'{e}n\'{e}tique Mol\'{e}culaire, Ecole Normale Sup\'{e}rieure, 46 rue d'Ulm, 75230 Paris Cedex 05, France.}, author = {Marheineke, K. and Hyrien, O.}, citeulike-article-id = {1182014}, citeulike-linkout-0 = {http://dx.doi.org/10.1074/jbc.M100271200}, citeulike-linkout-1 = {http://view.ncbi.nlm.nih.gov/pubmed/11279043}, citeulike-linkout-2 = {http://www.hubmed.org/display.cgi?uids=11279043}, day = {18}, doi = {10.1074/jbc.M100271200}, issn = {0021-9258}, journal = {J Biol Chem}, keywords = {aphidicolin, origins\_of\_replication, xenopus\_laevis}, month = {May}, number = {20}, pages = {17092--17100}, posted-at = {2007-03-23 17:32:12}, priority = {2}, title = {Aphidicolin triggers a block to replication origin firing in Xenopus egg extracts.}, url = {http://dx.doi.org/10.1074/jbc.M100271200}, volume = {276}, year = {2001} }

TY - JOUR ID - citeulike:1182014 L3 - citeulike-article-id:1182014 N2 - DNA replication origins are located at random with respect to DNA sequence in Xenopus early embryos and on DNA replicated in Xenopus egg extracts. We have recently shown that origins fire throughout the S phase in Xenopus egg extracts. To study the temporal regulation of origin firing, we have analyzed origin activation in sperm nuclei treated with the DNA polymerase inhibitor aphidicolin. Sperm chromatin was incubated in Xenopus egg extracts in the presence of aphidicolin and transferred to a fresh extract, and digoxigenin-dUTP and biotin-dUTP were added at various times after aphidicolin release to selectively label early and late replicating DNA. Molecular combing analysis of single DNA fibers showed that only a fraction of potential origins were able to initiate in the presence of aphidicolin. After release from aphidicolin, the remaining origins fired asynchronously throughout the S phase. Therefore, initiation during the S phase depends on the normal progression of replication forks assembled at earlier activated origins. Caffeine, an inhibitor of the checkpoint kinases ATR and ATM, did not relieve the aphidicolin-induced block to origin firing. We conclude that a caffeine-insensitive intra-S phase checkpoint regulates origin activation when DNA synthesis is inhibited in Xenopus egg extracts. IS - 20 JF - J Biol Chem SN - 0021-9258 AD - Laboratoire de Génétique Moléculaire, Ecole Normale Supérieure, 46 rue d'Ulm, 75230 Paris Cedex 05, France. EP - 17100 TI - Aphidicolin triggers a block to replication origin firing in Xenopus egg extracts. VL - 276 SP - 17092 KW - aphidicolin KW - origins_of_replication KW - xenopus_laevis AU - Marheineke, K AU - Hyrien, O PY - 2001/05/18/ UR - http://dx.doi.org/10.1074/jbc.M100271200 ER -