Anomalous diffusion of fluorescent probes inside living cell nuclei investigated by spatially-resolved fluorescence correlation spectroscopy

Wachsmuth, Malte; Waldeck, Waldemar; Langowski, Jörg

doi:10.1006/jmbi.2000.3692

Cited by 429 publications

(480 citation statements)

References 45 publications

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“…We found that the concentration of GFP inversely correlates with chromatin density, and the diffusive behavior in all chromatin environments adequately fitted to a 2-diffusive component model (Hinde et al 2010). From the diffusion coefficients derived, we found that GFP diffusion is neither impeded nor dependent upon chromatin density, which is in agreement with reported results (Wachsmuth et al 2000;Dross et al 2009). Thus, if there is a dependence of GFP diffusion on chromatin density, it must manifest itself on a length scale outside the resolution of the FCS technique employed (about 0.3 μm).…”

Section: Molecular Flow Within the Nucleussupporting

confidence: 90%

“…Given the absence of membranes separating intranuclear substructures, it has been postulated that other structural features of the nucleus (e.g., the chromatin itself) must impart divisions which control molecular flows and segregate different activities. A key emerging contributor to genome function is the architectural organisation of the cell nucleus and, given that chromatin fills up to 12% of the cell nucleus, it must be considered a major obstacle to molecular diffusion (Wachsmuth et al 2000;Tini et al 2002).…”

Section: Molecular Flow Within the Nucleusmentioning

confidence: 99%

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Measuring the flow of molecules in cells

Hinde

Cardarelli

2011

Biophys Rev

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No methods proposed thus far have the capability to measure molecular flow in live cells at the single molecule level. Here, we review the potentiality of a newly established method based on the spatial correlation of fluorescence fluctuations at a pair of points in the sample (pair correlation method). The pair correlation function (pCF) offers a unique tool to probe the directionality of intracellular traffic, by measuring the accessibility of the cellular landscape and its role in determining the diffusive routes adopted by molecules. The sensitivity of the pCF method toward detection of barriers means that different structural elements of the cell can be tested in terms of penetrability and mechanisms of regulation imparted on molecular flow. This has been recently demonstrated in a series of studies looking at molecular transport inside live cells. Here, we will review the theory behind detection of barriers to molecular flow, the rules to interpret pCF data, and relevant applications to intracellular transport.

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Section: Molecular Flow Within the Nucleussupporting

confidence: 90%

Section: Molecular Flow Within the Nucleusmentioning

confidence: 99%

Measuring the flow of molecules in cells

Hinde

Cardarelli

2011

Biophys Rev

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“…Thus, it can be treated as a representative, interesting and challenging problem. 15,16 Experimental studies concerning the motion of proteins and lipids in cells were performed using uorescence correlation spectroscopy, [17][18][19][20] pulsed eld gradient NMR 21 and single particle tracking (SPT). 6,7,[22][23][24][25][26][27] In many cases an anomalous diffusion was detected, i.e.…”

Section: Introductionmentioning

confidence: 99%

Simulation of diffusion in a crowded environment

Polanowski

Sikorski

2014

Soft Matter

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We performed extensive and systematic simulation studies of two-dimensional fluid motion in a complex crowded environment. In contrast to other studies we focused on cooperative phenomena that occurred if the motion of particles takes place in a dense crowded system, which can be considered as a crude model of a cellular membrane. Our main goal was to answer the following question: how do the fluid molecules move in an environment with a complex structure, taking into account the fact that motions of fluid molecules are highly correlated. The dynamic lattice liquid (DLL) model, which can work at the highest fluid density, was employed. Within the frame of the DLL model we considered cooperative motion of fluid particles in an environment that contained static obstacles. The dynamic properties of the system as a function of the concentration of obstacles were studied. The subdiffusive motion of particles was found in the crowded system. The influence of hydrodynamics on the motion was investigated via analysis of the displacement in closed cooperative loops. The simulation and the analysis emphasize the influence of the movement correlation between moving particles and obstacles.

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“…In fluorescent photobleaching experiments and in the uncaging experiments described here, the measured average mobility of the various nuclear particles is usually at least fivefold slower than that observed in solution. However, when more detailed analyses are carried out, it has been found in many cases that the entire population of molecules is not moving at this reduced rate, as one would expect for movement through a viscous solution, but instead, multiple populations of molecules with different mobilities are present (e.g., Politz et al, 1998;Wachsmuth et al, 2000;Platani et al, 2002;Pederson, 2002). This is consistent with anomalous subdiffusion, where mobility is constrained either by transient binding to and/or collisions with nuclear entities or by corralling within confinement zones, both of which phenomena impede free diffusion and give rise to multiple subpopulations of molecules moving with different mobilities (Feder et al, 1996;Saxton, 2001).…”

mentioning

confidence: 99%

Diffusion-based Transport of Nascent Ribosomes in the Nucleus

Politz

Tuft

Pederson

2003

MBoC

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Although the complex process of ribosome assembly in the nucleolus is beginning to be understood, little is known about how the ribosomal subunits move from the nucleolus to the nuclear membrane for transport to the cytoplasm. We show here that large ribosomal subunits move out from the nucleolus and into the nucleoplasm in all directions, with no evidence of concentrated movement along directed paths. Mobility was slowed compared with that expected in aqueous solution in a manner consistent with anomalous diffusion. Once nucleoplasmic, the subunits moved in the same random manner and also sometimes visited another nucleolus before leaving the nucleus. INTRODUCTIONIn eukaryotes, rRNA transcription and ribosome assembly take place in the nucleolus. Nascent ribosomes then exit the nucleolus and move into the cytoplasm. Multiple ribosomal proteins assemble with rRNA in the nucleolus and may facilitate proper processing of the pre-rRNA primary transcript (Fatica and Tollervey, 2002). Additionally, a significant number of nonribosomal proteins bind to these formative ribosomes in the nucleolus and also in the nucleoplasm after the nascent ribosomal subunits leave the nucleolus Kuersten et al., 2001;Nissan et al., 2002). The binding of particular proteins in a prescribed order is probably necessary for nucleocytoplasmic transport of the processed ribosomal subunits (e.g., Milkereit et al., 2001). The nuclear export of both the large and small ribosomal subunits has been shown to be dependent, at least indirectly, on the Ran-GTPase cycle and the exportin CRM1, and it is likely that CRM1-mediated export of 60S subunits requires the adaptor protein NMD3 (Moy and Silver, 1999;Ho et al., 2000;Gadal et al., 2001;Thomas and Kutay, 2003;Trotta et al., 2003). In situ hybridization experiments in fission yeast have indicated that rRNA may accumulate along short tracks when near the nuclear pores but nonetheless appears to exit from all the pores, not just those near the nucleolus (Léger-Silvestre et al., 1999). In situ hybridization studies in mammalian cells have primarily addressed the distribution of rRNA within the nucleolus (Huang, 2002) rather than the routes of extranucleolar rRNA traffic, because although high signal representing rRNA is routinely detected in both the nucleolus and the cytoplasm, nucleoplasmic signal which might represent ribosomes moving to the nuclear periphery is not easily detectable by in situ hybridization (PuvionDutilleul et al., 1991;Lazdins et al., 1997; J.C.R. Politz, unpublished observations). Therefore, very little is known about the spatial pattern or mechanism of rRNA movement from the nucleolus into the nucleoplasm and then to the nuclear pores for export (Cullen, 2000;Kuersten et al., 2001;Lei and Silver, 2002), although all three classes of eukaryotic RNAs, rRNA, mRNA, and pol III transcripts have been shown to exit from all the nuclear pores (Dworetzky and Feldherr, 1988;Pante et al., 1997;Mattaj and Englmeier, 1998).In previous studies in live cells (Politz et al., 1998, we investigat...

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Anomalous diffusion of fluorescent probes inside living cell nuclei investigated by spatially-resolved fluorescence correlation spectroscopy

Cited by 429 publications

References 45 publications

Measuring the flow of molecules in cells

Measuring the flow of molecules in cells

Simulation of diffusion in a crowded environment

Diffusion-based Transport of Nascent Ribosomes in the Nucleus

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