Cell cycle-dependent mobility of Cdc45 in living cells determined by fluorescence correlation spectroscopy.
Broderick, Ronan Ramadurai, Sivaramakrishnan Togashi, Denisio M. Ryder, Alan G. Langowski, Jorg
Broderick, Ronan
Ramadurai, Sivaramakrishnan
Togashi, Denisio M.
Ryder, Alan G.
Langowski, Jorg
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2012
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R. Broderick, S. Ramadurai, K. Toth, D. Togashi, A. G. Ryder, J. Langwoski, and H.P. Nasheuer. (2012) 'Cell cycle-dependent mobility of Cdc45 in living cells determined by fluorescence correlation spectroscopy'. Plos One, 7 (4):e35537-e35537.
Abstract
Eukaryotic DNA replication is a dynamic process requiring the co-operation of specific replication proteins. We measured the mobility of eGFP-Cdc45 by Fluorescence Correlation Spectroscopy (FCS) in vivo in asynchronous cells and in cells synchronized at the G1/S transition and during S phase. Our data show that eGFP-Cdc45 mobility is faster in G1/S transition compared to S phase suggesting that Cdc45 is part of larger protein complex formed in S phase. Furthermore, the size of complexes containing Cdc45 was estimated in asynchronous, G1/S and S phase-synchronized cells using gel filtration chromatography; these findings complemented the in vivo FCS data. Analysis of the mobility of eGFP-Cdc45 and the size ofcomplexes containing Cdc45 and eGFP-Cdc45 after UVC-mediated DNA damage revealed no significant changes in diffusion rates and complex sizes using FCS and gel filtration chromatography analyses. This suggests that after UV-damage, Cdc45 is still present in a large multi-protein complex and that its mobility within living cells is consistently similar following UVC-mediated DNA damage.
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Public Library of Science
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http://dx.doi.org/10.1371/journal.pone.0035537
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Attribution-NonCommercial-NoDerivs 3.0 Ireland