Overview of the intranuclear steps of expression of a BR1/BR2 gene. Processes taking place at the gene locus, in the interchromatin space and at the NPC are listed. BR gene chromatin in black, RNA polymerase II elongation complex in purple, pre-mRNPs in light blue, mRNPs in dark blue and ribosomes in orange. The NPC and nuclear membrane are in grey

Cotranscriptional processes at the 3′ end of the BR1/BR2 gene. a Schematic representation. Colour code as in Fig. 2. Excision of intron 4 (i4, black box) is initiated rapidly. About 0.6 kb downstream the poly(A) signal (grey box), transcription termination, 3′ end cleavage and polyadenylation are coordinated. The pre-mRNP is retained at the gene during the initial polyadenylation, and splicing is stimulated at this stage. Approximately 3 kb of DNA, as extended chromatin, is present downstream the site of release of the pre-mRNP, before being folded into compact chromatin (black curved line). Released mRNP in dark blue. b EM image of the 3′ end of the gene (adopted from Ericsson et al. 1989). The arrows indicate the chromatin fibre, extending from the final pre-mRNP into the compact chromatin. The scale bar represents 50 nm

The behaviour of BR1/BR2 mRNPs at the NPC. Schematic representations and EM images (adopted from Mehlin et al. 1992) of the corresponding steps. a, a′ The mRNP (in dark blue) binds to the tip of the basket (in grey). b, b′ It then enters into the basket and docks at the entrance of the central channel. c, c′ Subsequently, the mRNP changes conformation as it is translocated through the central channel with the 5′ end first. d, d′ At the cytoplasmic side, initiation of translation can occur rapidly at the exit of the central channel. Orange dots in (d) and arrows in (d′) indicate ribosomes. The scale bar represents 100 nm

Cotranscriptional synthesis, assembly and processing of BR1/BR2 and BR3 pre-mRNPs. a The proximal (0–7 kb) and middle (7–35 kb) regions of the BR1/BR2 gene is shown schematically. The bent arrow indicates the transcription start site. The RNA polymerase II elongation complex in purple and the pre-mRNPs in light blue. The proximal region contains three introns (i1–i3, black boxes) and exons 1–3, followed by the long (approximately 35 kb) exon 4. The continuous recruitment of components to the pre-mRNPs (listed components have been demonstrated) and the continuous assembly of the pre-mRNPs are indicated by red arrows. The processing events of the pre-mRNPs are indicated (red lines). Assembly of the exon sequences results in a 7-nm pre-mRNP fibre that is further folded into a 19-nm fibre, followed by organisation into higher order structures. Interaction between the pre-mRNP and chromatin may involve pre-mRNP-bound actin. b Schematic representation of part of the BR3 gene. The BR3 gene contains multiple short introns and exons. Three nascent transcript and splicing (NTS) complexes are shown. Each complex consists of a pre-mRNP (light blue), including splicing factors and an RNA polymerase II elongation complex (purple). b′ EM 3D reconstruction of a corresponding BR3 gene segment with three NTS complexes (adopted from Wetterberg et al. 2001). The repeated assembly and structural dynamics of the spliceosome along the multi-intron gene dominate the structure of the pre-mRNPs. The scale bar represents 10 nm. c, d EM images of nascent BR1/BR2 pre-mRNPs in the middle region of the gene (adopted from Skoglund et al. 1983). The 19-nm fibre and the higher order granular structure are seen in several pre-mRNPs. The granular structure (arrows) gets bigger and denser as more RNA protein is added (compare c and d). The scale bar represents 50 nm

The BR1/BR2 mRNPs in the interchromatin space. a EM image of BR mRNPs in situ. The BR mRNPs (50 nm in diameter) in the interchromatin are indicated by long arrows. For comparison, short arrows indicate ribosomal subunits. N nucleus, C cytoplasm. The scale bar represents 100 nm. b The 3D structure of the BR1/BR2 mRNP (50 nm in diameter) (adopted from Mehlin et al. 1995, © The Rockefeller University Press. The Journal of Cell Biology, 1995, 129:1205–1216, Fig. 7). The numbers (1–6) on the mRNP indicate multiple contact points with the inner ring of the NPC during export. The numbers outside the mRNP (1–4) indicate described domains of the mRNP. The scale bar represents 10 nm. c The released BR1/BR2 mRNPs move randomly inside the interchromatin space and become part of a population of mRNPs, from which individual mRNPs (stochastically) bind to the basket of the NPCs. Colour code as in Fig. 1. c′ EM image of a section through a polytene nucleus (adopted from Singh et al. 1999). Newly synthesised (red dots) and old (blue dots) BR1/BR2 mRNPs are randomly distributed. The highest concentration of mRNPs is at the gene locus (BR). PC polytene chromosomes, Nu nucleolus, N nucleus, C cytoplasm. The scale bar represents 5 μm. d Within the interchromatin space, the random movement of the mRNP is occasionally interrupted by transient interactions (light grey, thin lines) with interchromatin structures (dark grey, thick line). d′ EM 3D reconstruction of a BR1/BR2 mRNP interacting with an interchromatin fibre structure (adopted from Miralles et al. 2000, © The Rockefeller University Press. The Journal of Cell Biology, 2000, 148:271–282, Fig. 2d, part IV). Numbers 1–4 refer to the described domains of the mRNP. The scale bar represents 20 nm