p53 localizes to intranucleolar regions distinct from the ribosome production compartments.

The tumor suppressor p53 has been implicated in the regulation of ribosome biogenesis based on its inhibitory effect on RNA polymerase I (pol I)-dependent transcription. Consistent with this, p53 has been described in nucleoli, albeit under specific experimental conditions. Since data on the intranucleolar localization of p53 are controversial, we have analyzed in detail its subnucleolar distribution. Our results show that p53 does not localize to one of the well-known structural components of the nucleolus involved in ribosome biogenesis, but rather occupies distinct intranucleolar regions that constitute nucleolar cavities. When cells were treated with the proteasome inhibitor MG132, the size and frequency of p53-containing nucleolar cavities increased, and the protein partially colocalized with inactivated proteasomes. Importantly, p53 did not colocalize with pol I at the transcription sites in fibrillar centers (FCs) as has previously been reported. The observed intranucleolar distribution and accumulation of p53 raises the question of how the protein influences rDNA transcription in vivo.

Intranucleolar sites of ribosome biogenesis defined by the localization of early binding ribosomal proteins.

Considerable efforts are being undertaken to elucidate the processes of ribosome biogenesis. Although various preribosomal RNP complexes have been isolated and molecularly characterized, the order of ribosomal protein (r-protein) addition to the emerging ribosome subunits is largely unknown. Furthermore, the correlation between the ribosome assembly pathway and the structural organization of the dedicated ribosome factory, the nucleolus, is not well established. We have analyzed the nucleolar localization of several early binding r-proteins in human cells, applying various methods, including live-cell imaging and electron microscopy. We have located all examined r-proteins (S4, S6, S7, S9, S14, and L4) in the granular component (GC), which is the nucleolar region where later pre-ribosomal RNA (rRNA) processing steps take place. These results imply that early binding r-proteins do not assemble with nascent pre-rRNA transcripts in the dense fibrillar component (DFC), as is generally believed, and provide a link between r-protein assembly and the emergence of distinct granules at the DFC-GC interface.

The chromatin remodelling complex WSTF-SNF2h interacts with nuclear myosin 1 and has a role in RNA polymerase I transcription.

Nuclear actin and myosin 1 (NM1) are key regulators of gene transcription. Here, we show by biochemical fractionation of nuclear extracts, protein-protein interaction studies and chromatin immunoprecipitation assays that NM1 is part of a multiprotein complex that contains WICH, a chromatin remodelling complex containing WSTF (Williams syndrome transcription factor) and SNF2h. NM1, WSTF and SNF2h were found to be associated with RNA polymerase I (Pol I) and ribosomal RNA genes (rDNA). RNA interference-mediated knockdown of NM1 and WSTF reduced pre-rRNA synthesis in vivo, and antibodies to WSTF inhibited Pol I transcription on pre-assembled chromatin templates but not on naked DNA. The results indicate that NM1 cooperates with WICH to facilitate transcription on chromatin.