Microenvironment and effect of energy depletion in the nucleus analyzed by mobility of multiple oligomeric EGFPs |
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Authors: | Pack Changi Saito Kenta Tamura Mamoru Kinjo Masataka |
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Institution: | Laboratory of Supramolecular Biophysics, Research Institute for Electronic Science, Hokkaido University, Sapporo 060-0812, Japan. |
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Abstract: | Four different tandem EGFPs were constructed to elucidate the nuclear microenvironment by quantifying its diffusional properties in both aqueous solution and the nuclei of living cells. Diffusion of tandem EGFP was dependent on the length of the protein as a rod-like molecule or molecular ruler in solution. On the other hand, we found two kinds of mobility, fast diffusional mobility and much slower diffusional mobility depending on cellular compartments in living cells. Diffusion in the cytoplasm and the nucleoplasm was mainly measured as fast diffusional mobility. In contrast, diffusion in the nucleolus was complex and mainly much slower diffusional mobility, although both the fast and the slow diffusional mobilities were dependent on the protein length. Interestingly, we found that diffusion in the nucleolus was clearly changed by energy depletion, even though the diffusion in the cytoplasm and the nucleoplasm was not changed. Our results suggest that the nucleolar microenvironment is sensitive to energy depletion and very different from the nucleoplasm. |
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Keywords: | cps count per second Cyt cytoplasm 2-DG 2-deoxyglucose DT diffusion time EGFPn tandemly linked oligomeric EGFP FAF fluorescence autocorrelation function MR molecular ruler FCS fluorescence correlation spectroscopy FRAP fluorescence recovery after photobleach mRFP monomeric red fluorescent protein NP nucleoplasm NL nucleolus SPT single particle tracking TSAT Trichostatin A |
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