| In vivo quantitative studies of dynamic intracellular processes using fluorescence correlation spectroscopy. | |
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MedLine Citation:
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PMID: 16603490 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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It has been a significant challenge to quantitatively study the dynamic intracellular processes in live cells. These studies are essential for a thorough understanding of the underlying mechanisms regulating the signaling pathways and the transitions between cell cycle stages. Our studies of Cdc20, an important mitotic checkpoint protein, throughout the cell cycle demonstrate that fluorescence correlation spectroscopy is a powerful tool for in vivo quantitative studies of dynamic intracellular processes. In this study, Cdc20 is found to be present primarily in a large complex (>1 Mda) during interphase with a diffusion constant of 1.8+/-0.1 microm2/s and a concentration of 76+/-24 nM, consistent with its association with the APC/C. During mitosis, however, a proportion of Cdc20 dissociates from APC/C at a rate of 12 pM/s into a soluble pool with a diffusion constant of 19.5+/-5.0 microm2/s, whose size is most consistent with free Cdc20. This free pool accumulates to 50% of total Cdc20 (approximately 40 nM) during chronic activation of the mitotic checkpoint but disappears during mitotic exit at a rate of 31 pM/s. The observed changes in the biochemical assembly states of Cdc20 closely correlate to the known temporal pattern of the activity of APC/CCdc20 in mitosis. Photon counting histograms reveal that both complexes contain only a single molecule of Cdc20. The underlying mechanisms of the activities of APC/CCdc20 throughout the cell cycle are discussed in light of our experimental observations. |
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Authors:
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Zifu Wang; Jagesh V Shah; Michael W Berns; Don W Cleveland |
Publication Detail:
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Type: Journal Article; Research Support, N.I.H., Extramural; Research Support, U.S. Gov't, Non-P.H.S. Date: 2006-04-07 |
Journal Detail:
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Title: Biophysical journal Volume: 91 ISSN: 0006-3495 ISO Abbreviation: Biophys. J. Publication Date: 2006 Jul |
Date Detail:
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Created Date: 2006-06-15 Completed Date: 2006-08-16 Revised Date: 2009-11-18 |
Medline Journal Info:
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Nlm Unique ID: 0370626 Medline TA: Biophys J Country: United States |
Other Details:
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Languages: eng Pagination: 343-51 Citation Subset: IM |
Affiliation:
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Developmental Biology Center, Beckman Laser Institute, and Department of Biomedical Engineering, University of California, Irvine, and Ludwig Institute for Cancer Research, La Jolla, California, USA. zifuw@uci.edu |
Export Citation:
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| MeSH Terms | |
Descriptor/Qualifier:
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Cell Cycle
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physiology* Cell Cycle Proteins / metabolism* Cell Line Intracellular Fluid / metabolism Kinetics Microscopy, Fluorescence, Multiphoton / methods* Potoroidae Spectrometry, Fluorescence / methods* |
| Grant Support | |
ID/Acronym/Agency:
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GM29513/GM/NIGMS NIH HHS; GM66051/GM/NIGMS NIH HHS; RR-14892/RR/NCRR NIH HHS |
| Chemical | |
Reg. No./Substance:
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0/Cell Cycle Proteins; 156288-95-8/CDC20 protein, human |
| Comments/Corrections | |
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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