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Visualization of replication initiation and elongation in Drosophila

Chorion gene amplification in the ovaries of Drosophila melanogaster is a powerful system for the study of metazoan DNA replication in vivo. Using a combination of high-resolution confocal and deconvolution microscopy and quantitative realtime PCR, we found that initiation and elongation occur durin...

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Autores principales: Claycomb, Julie M., MacAlpine, David M., Evans, James G., Bell, Stephen P., Orr-Weaver, Terry L.
Formato: Texto
Lenguaje:English
Publicado: The Rockefeller University Press 2002
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2173051/
https://www.ncbi.nlm.nih.gov/pubmed/12403810
http://dx.doi.org/10.1083/jcb.200207046
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author Claycomb, Julie M.
MacAlpine, David M.
Evans, James G.
Bell, Stephen P.
Orr-Weaver, Terry L.
author_facet Claycomb, Julie M.
MacAlpine, David M.
Evans, James G.
Bell, Stephen P.
Orr-Weaver, Terry L.
author_sort Claycomb, Julie M.
collection PubMed
description Chorion gene amplification in the ovaries of Drosophila melanogaster is a powerful system for the study of metazoan DNA replication in vivo. Using a combination of high-resolution confocal and deconvolution microscopy and quantitative realtime PCR, we found that initiation and elongation occur during separate developmental stages, thus permitting analysis of these two phases of replication in vivo. Bromodeoxyuridine, origin recognition complex, and the elongation factors minichromosome maintenance proteins (MCM)2–7 and proliferating cell nuclear antigen were precisely localized, and the DNA copy number along the third chromosome chorion amplicon was quantified during multiple developmental stages. These studies revealed that initiation takes place during stages 10B and 11 of egg chamber development, whereas only elongation of existing replication forks occurs during egg chamber stages 12 and 13. The ability to distinguish initiation from elongation makes this an outstanding model to decipher the roles of various replication factors during metazoan DNA replication. We utilized this system to demonstrate that the pre–replication complex component, double-parked protein/cell division cycle 10–dependent transcript 1, is not only necessary for proper MCM2–7 localization, but, unexpectedly, is present during elongation.
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spelling pubmed-21730512008-05-01 Visualization of replication initiation and elongation in Drosophila Claycomb, Julie M. MacAlpine, David M. Evans, James G. Bell, Stephen P. Orr-Weaver, Terry L. J Cell Biol Article Chorion gene amplification in the ovaries of Drosophila melanogaster is a powerful system for the study of metazoan DNA replication in vivo. Using a combination of high-resolution confocal and deconvolution microscopy and quantitative realtime PCR, we found that initiation and elongation occur during separate developmental stages, thus permitting analysis of these two phases of replication in vivo. Bromodeoxyuridine, origin recognition complex, and the elongation factors minichromosome maintenance proteins (MCM)2–7 and proliferating cell nuclear antigen were precisely localized, and the DNA copy number along the third chromosome chorion amplicon was quantified during multiple developmental stages. These studies revealed that initiation takes place during stages 10B and 11 of egg chamber development, whereas only elongation of existing replication forks occurs during egg chamber stages 12 and 13. The ability to distinguish initiation from elongation makes this an outstanding model to decipher the roles of various replication factors during metazoan DNA replication. We utilized this system to demonstrate that the pre–replication complex component, double-parked protein/cell division cycle 10–dependent transcript 1, is not only necessary for proper MCM2–7 localization, but, unexpectedly, is present during elongation. The Rockefeller University Press 2002-10-28 /pmc/articles/PMC2173051/ /pubmed/12403810 http://dx.doi.org/10.1083/jcb.200207046 Text en Copyright © 2002, The Rockefeller University Press This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 4.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/4.0/).
spellingShingle Article
Claycomb, Julie M.
MacAlpine, David M.
Evans, James G.
Bell, Stephen P.
Orr-Weaver, Terry L.
Visualization of replication initiation and elongation in Drosophila
title Visualization of replication initiation and elongation in Drosophila
title_full Visualization of replication initiation and elongation in Drosophila
title_fullStr Visualization of replication initiation and elongation in Drosophila
title_full_unstemmed Visualization of replication initiation and elongation in Drosophila
title_short Visualization of replication initiation and elongation in Drosophila
title_sort visualization of replication initiation and elongation in drosophila
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2173051/
https://www.ncbi.nlm.nih.gov/pubmed/12403810
http://dx.doi.org/10.1083/jcb.200207046
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