The mammalian cell nucleus contains numerous sub-compartments, which have been implicated in essential processes such as transcription and splicing1,2. The mechanisms by which nuclear compartments are formed and maintained are unclear. More fundamentally, it is not known how proteins move within the cell nucleus. We have measured the kinetic properties of proteins in the nucleus of living cells using photobleaching techniques. Here we show that proteins involved in diverse nuclear processes move rapidly throughout the entire nucleus. Protein movement is independent of energy, which indicates that proteins may use a passive mechanism of movement. Proteins rapidly associate and dissociate with nuclear compartments. Using kinetic modelling, we determined residence times and steady-state fluxes of molecules in two main nuclear compartments. These data show that many nuclear proteins roam the cell nucleus in vivo and that nuclear compartments are the reflection of the steady-state association/dissociation of its ‘residents’ with the nucleoplasmic space. Our observations have conceptual implications for understanding nuclear architecture and how nuclear processes are organized in vivo.
Lamond,A. I. & Earnshaw,W. C. Structure and function in the nucleus. Science 280, 547– 553 (1998).
Misteli,T. & Spector,D. L. The cellular organization of gene expression. Curr. Opin. Cell Biol. 10, 322 –331 (1998).
Matera,A. G. Nuclear bodies: multifaceted subdomains of the interchromatin space. Trends Cell Biol. 9, 302–309 (1999).
Bustin,M. Regulation of DNA-dependent activities by the functional motifs of the high-mobility-group chromosomal proteins. Mol. Cell. Biol. 19, 5237–5246 (1999).
Hock,R., Wilde,F., Scheer,U. & Bustin,M. Dynamic relocation of chromosomal protein HMG-17 in the nucleus is dependent on transcriptional activity. EMBO J. 17, 6992– 7001 (1998).
Ge,H. & Manley,J. L. A protein factor, ASF, controls cell specific alternative splicing of SV40 early pre-mRNA in vitro. Cell 62, 24–34 (1990).
Krainer,A. R., Conway,G. C. & Kozak, D. The essential pre-mRNA splicing factor SF2 influences 5′ splice site selection by activating proximal sites. Cell 62, 35–42 ( 1990).
Cáceres,J. F., Misteli,T., Screaton,G., Spector,D. L. & Krainer, A. R. Role of the modular domains of SR-proteins in subnuclear localization and alternative splicing specificity. J. Cell Biol. 138, 225–238 ( 1997).
Scheer,U. & Hock,R. Structure and function of the nucleolus. Curr. Opin. Cell Biol. 11, 385– 390 (1999).
Olson,M. O. J., Dundre,M. & Szebeni,A. The nucleolus: An old factory with unexpected capabilities. Trends. Cell Biol. (in the press).
Ochs,R. L., Lischwe,M. A., Spohn,W. H. & Busch,H. Fibrillarin: a new protein of the nucleolus identified by autoimmune sera. Biol. Cell 54, 123–133 ( 1985).
Misteli,T., Cáceres,J. F. & Spector, D. L. The dynamics of a pre-mRNA splicing factor in living cells. Nature 387, 523– 527 (1997).
White,J. & Stelzer,E. Photobleaching GFP reveals protein dynamics inside live cells. Trends Cell Biol. 9, 61–65 (1999).
Seksek,O., Biwersi,J. & Verkman, A. S. Translational diffusion of macromolecule-sized solutes in cytoplasm and nucleus. J. Cell Biol. 138, 131–142 (1997).
Yokoe,H. & Meyer,T. Spatial dynamics of GFP-tagged proteins investigated by local fluorescence enhancement. Nature Biotechnol. 14, 1252–1256 ( 1996).
Houtsmuller,A. B. et al. Action of DNA repair endonuclease ERCC1/XPF in living cells. Science 284, 958–961 (1999).
Kanda,T., Sullivan,K. F. & Wahl, G. M. Histone–GFP fusion protein enables sensitive analysis of chromosome dynamics in living mammalian cells. Curr. Biol. 26, 377–385 ( 1998).
Foster,D. M. et al. in Proceedings of the Simulation in Health Science Conference 87–90 (Society for Computer Simulation, San Diego, 1994).
Bell,B. M., Burke,J. V. & Schumitzky, A. A relative weighting method for estimating parameters and variances in multiple data sets. Computational Statistics and Data Analysis 22, 119–135 (1996).
Hanamura,A., Cáceres,J. F., Mayeda, A., Franza,B. A. & Krainer,A. R. Regulated tissue-specific expression of antagonistic pre-mRNA splicing factors. RNA 4, 430–444 (1998).
Politz,J. C., Browne,E. S., Wolf,D. E. & Pederson,T. Intranuclear diffusion and hybridization state of oligonucleotides measured by fluorescence correlation spectroscopy in living cells. Proc. Natl Acad. Sci. USA 95, 6043–6048 (1998).
Daneholt,B. Pre-mRNP particles: From gene to nuclear pore. Curr. Biol. 11, R412–R415 (1999).
Politz,J. C., Tuft,R. A., Pederson,T. & Singer,R. H. Movement of nuclear poly(A) RNA throughout the interchromatin space in living cells. Curr. Biol. 9, 285–291 ( 1999).
Singh,O. P., Bjorkroth,B., Masich,S., Wieslander,L. & Daneholt, B. The intranuclear movement of Balbiani ring premessenger ribonucleoprotein particles. Exp. Cell Res. 251, 135–146 (1999).
Sleeman,J. E., Platani,M., Kreiv,J. P., Lamond,A. I. Dynamic interactions between splicing snRNPs, coiled bodies and nucleoli revealed using snRNP protein fusions to the green fluorescent protein. Exp. Cell Res. 243, 290–304 (1998).
Boudonck,K., Dolan,L. & Shaw,P. J. The movement of coiled bodies visualized in living plant cells by the green fluorescent protein. Mol. Biol. Cell 10, 2297–2307 (1999).
Jolly,C., Usson,Y. & Morimoto,R. I. Rapid and reversible relocalization of heat shock factor 1 within seconds to nuclear stress granules. Proc. Natl Acad. Sci. USA 96, 6769–6774 ( 1999).
Misteli,T. & Spector,D. L. Serine/threonine phosphatase 1 modulates the subnuclear distribution of pre-mRNA splicing factors. Mol. Biol. Cell 7, 1559–1572 (1996).
Dingwall,C., Black,S. J., Kearsey,S. E., Cox,L. S. & Laskey,R. A. Nucleoplasmin cDNA sequence reveals polyglutamic acid tracts and a cluster of sequences homologous to putative nuclear localization signals. EMBO J. 6, 69–74 (1987).
Axelrod,D., Koppel,D. E., Schlessinger, J., Elson,E. & Webb,W. W. Mobility measurement by analysis of fluorescence photobleaching recovery kinetics. Biophys. J. 16, 1055–1069 (1976).
We thank M. Dundr, G. Wahl, R. Hock and M. Bustin for providing GFP-fibrillarin, GFP-H2B and GFP-HMG-17 clones, respectively, and M. Bustin for anti HMG-17 antibody. We thank M. Bustin for critical reading of the manuscript.
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Phair, R., Misteli, T. High mobility of proteins in the mammalian cell nucleus. Nature 404, 604–609 (2000). https://doi.org/10.1038/35007077
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