6 Histochem Cell Biol (2008) 129:5–11 Fig. 1 Nuclear architecture and function. The mammalian cell nucle- us contains a large number of non-random and structural features. a Nuclear lamins (green) form a network of architectural elements at the periphery. This nuclear lamina is implicated in mechanical support of the nucleus, organization of chromatin (red/blue) in the nuclear inte- rior, as a platform for signaling events and in the sequestration of nu- clear proteins. b, c Genomes are non-randomly organized within the »10 m, chromatin is estimated to only occupy about 15% of the nuclear volume. A key question in understanding nuclear architecture is how protein and RNAs move within the nucleus. This is a key issue if we are to gain a full appreciation of what the nuclear environment is like. More importantly, the mode of nuclear motion of molecules has physiological implications as it aVects how proteins Wnd their target sequences in the genome, how RNAs are exported from the nucleus upon transcription and how proteins are sequestered within the nucleus for regulatory purposes. Some initial insights into these issues are summarized in broad terms here. The motion of RNA and proteins in the cell nucleus We have long known that proteins and RNAs must be able to move within the cell nucleus. Messenger RNAs traverse the nucleus and are promptly exported from the nucleus after their synthesis and complete processing. Newly syn- thesized proteins on the other hand are imported into the nucleus and rapidly disperse throughout its volume, imply- ing eVective intranuclear traYcking. Furthermore, proteins rapidly relocalize within the nucleus upon experimental or physiological changes in conditions. For example, inhibi- tion of protein synthesis leads to rapid and dramatic relocal- ization of a large number of nucleolar proteins (Andersen et al. 2005), and pre-mRNA splicing factors swiftly respond to transcriptional activation and inhibition (Jiménez-García and Spector 1993; Misteli et al. 1997). How proteins and RNAs move within the nucleus and how they Wnd their targets, however, was unknown for long. It was not clear whether movements are energy dependent, whether they occur by directed transport or what the speed of intranuclear traYcking is. These fundamental questions have recently been answered (Misteli 2001). 123 nucleus. b Each chromosome occupies a distinct chromosome territory (red chromosome 14; green chromosome 17 in human breast epithelial cells). c Pericentromeric heterochromatin regions (white) from multi- ple chromosomes often cluster into chromocenters. Bar 6.3 m. Image courtesy: a Gianluca Pegoraro, b Karen Meaburn, c Manjari Mazum- dar, all National Cancer Institute, NIH The investigation of intranuclear protein and RNA dynamics was made possible by the development of tech- niques to visualize and measure the motion of these mole- cules by time-lapse microscopy. Tracking of proteins by photobleaching methods revealed a surprising degree and speed of intranuclear traYcking. The analysis of tracers such as GFP or dextran which do not undergo speciWc inter- actions with DNA or other proteins indicated that the nuclear environment is permissive for rapid diVusional motion (Görisch et al. 2005; Misteli 2001; Seksek et al. 1997). The diVusion of molecules within the nucleus is only limited by the steric constraints imposed by chromatin and nuclear bodies (Görisch et al. 2005; Misteli 2001; Seksek et al. 1997). The typically measured diVusion coeYcients of 10–100 m2 s¡1 are similar to those observed in the cytoplasm and are only »5 times less than that in solution. As expected for diVusion, this rapid motion is energy-independent and non-directional. RNA motion within the nucleus is similarly rapid and non-directional. Several methodological approaches involving Xuorescently labeled, microinjected, engineered or endogenous RNAs demonstrate that ribosomal RNAs as well as polyA-RNAs move freely in a non-directional man- ner with a diVusion coeYcient of 0.03–0.1 m2 s¡1 within the nucleus (Ritland-Politz et al. 2006; Shav-Tal et al. 2004). A diVusion coeYcient of this magnitude is suYcient to ensure transport of an RNA particle from deep within the nucleus to the cytoplasm within a few minutes, consistent with biochemical observations on kinetics of RNA matura- tion and transport. Thus energy-independent, diVusion- based movement of RNA particles alone can account for the observed kinetics of RNA export, and no active mecha- nisms are required to ensure rapid export. An impressive demonstration of the non-directional motion of mRNA comes from studies in which a nascent RNA is visualized at its site of transcription and its export to the cytoplasm mea-