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Time-lapse, photoactivation, and photobleaching imaging of nucleolar assembly after mitosis

Research output: Chapter in Book/Report/Conference proceedingChapterpeer-review

Danièle Hernandez-Verdun, Emilie Louvet, Eleonora Muro

Original languageEnglish
Title of host publicationImaging Gene Expression
Subtitle of host publicationMethods and Protocols
EditorsYaron Shav-Tal
PublisherHumana Press
Pages337-350
Number of pages14
ISBN (Print)9781627035255
DOIs
PublishedSep 2013

Publication series

NameMethods in Molecular Biology
Volume1042
ISSN (Print)1064-3745

King's Authors

Research Groups

  • King's College London

Abstract

Nucleolus assembly starts in telophase with the benefit of building blocks passing through mitosis and lasts until cytokinesis generating the two independent interphasic cells. Several approaches make it possible to follow the dynamics of fluorescent molecules in live cells. Here, three complementary approaches are described to measure the dynamics of proteins during nucleolar assembly after mitosis: (1) rapid two-color 4-D imaging time-lapse microscopy that demonstrates the relative localization and movement of two proteins, (2) photoactivation that reveals the directionality of migration from the activated area, and (3) fluorescence recovery after photobleaching (FRAP) that measures the renewing of proteins in the bleached area. We demonstrate that the order of recruitment of the processing machineries into nucleoli results from differential sorting of intermediate structures assembled during telophase, the prenucleolar bodies.

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