Spatial organization of RNA polymerase II transcription in the nucleus

Abstract

In eukaryotic cells, mRNA synthesis is carried out by large, multifunctional complexes that are also involved in coordinating transcription with other nuclear processes. This survey focuses on the distribution and structural arrangement of these complexes within the nucleus, in relationship with the discrete positioning of particular chromosomal loci. To better understand the link between the spatial organization of the nucleus and the regulation of gene expression, it is necessary to combine information from biochemical studies with results from microscopic observations of preserved nuclear structures. Recent experimental approaches have made this possible. The subnuclear locations of specific chromosome loci, RNA transcripts, RNA polymerases, and transcription and pre-mRNA-processing factors can now be observed with computer-assisted microscopy and specific molecular probes. The results indicate that RNA polymerase II (RNAPII) transcription takes place at discrete sites scattered throughout the nucleoplasm, and that these sites are also the locations of pre-mRNA processing. Transcribing polymerases appear to be grouped into clusters at each transcription site. Cell cycle-dependent zones of transcription and processing factors have been identified, and certain subnuclear domains appear specialized for expression or silencing of particular genes. The arrangement of transcription in the nucleus is dynamic and depends on its transcriptional activity, with the RNAPII itself playing a central role in marshalling the large complexes involved in gene expression.

Figure 1

Schematic representation of the subnuclear localization of pre-mRNA transcription and processing components as observed by immunofluorescence labeling and confocal microscopy analyses. (A) Immunodetection with antibodies to the RNAPII phosphorylated CTD reveals RNAPII localized in the nucleus in thousands of tiny clusters. These foci, which occupy <1% of the nuclear volume, overlap with the sites of pre-mRNA synthesis detected by labeling the nascent transcripts with BrdU. In general, transcription and processing factors colocalize with the RNAPII foci, although particular factors can be observed at other locations that may represent storage pools. In cells that are actively transcribing, RNAPII transcription foci are diffusely distributed throughout the nucleus (bottom). When transcription is inhibited, sites contract to minimal domains (top). (B) Two-dimensional schematic representation of an RNAPII transcription factory (inner dashed circle) and its surrounding region of chromosome loops and transcripts with associated processing factors (outer dashed circle). In this model, genes are recruited to and transcribed by attached polymerases. Most genes are transcribed by a single RNAPII, although particular genes may have several transcribing polymerases.

Figure 2

Possible models for the reorganization of RNAPII transcription factories as a function of the overall transcriptional activity of the cell. Nuclear components are known to undergo dynamic rearrangements as the transcriptional status of either endogenous or newly introduced genes changes. While transcriptionally active RNAPII molecules are clustered in transcription factories (right), the arrangement of inactive RNAPIIs is not known. Inactive RNAPIIs may be grouped (top left) or dissociated from any nuclear structure (bottom left).