Structural biology: Pivotal findings for a transcription machine.

Crystal structures of the complete RNA Polymerase I complex are now revealed. These structures link opening and closing of the DNA binding cleft of this enzyme to control of transcription.

Topoisomerase IIα promotes activation of RNA polymerase I transcription by facilitating pre-initiation complex formation.

Type II DNA topoisomerases catalyse DNA double-strand cleavage, passage and re-ligation to effect topological changes. There is considerable interest in elucidating topoisomerase II roles, particularly as these proteins are targets for anti-cancer drugs. Here we uncover a role for topoisomerase IIα in RNA polymerase I-directed ribosomal RNA gene transcription, which drives cell growth and proliferation and is upregulated in cancer cells.

Basic mechanisms in RNA polymerase I transcription of the ribosomal RNA genes.

RNA Polymerase (Pol) I produces ribosomal (r)RNA, an essential component of the cellular protein synthetic machinery that drives cell growth, underlying many fundamental cellular processes. Extensive research into the mechanisms governing transcription by Pol I has revealed an intricate set of control mechanisms impinging upon rRNA production. Pol I-specific transcription factors guide Pol I to the rDNA promoter and contribute to multiple rounds of transcription initiation, promoter escape, elongation and termination.

TAF1B is a TFIIB-like component of the basal transcription machinery for RNA polymerase I.

Transcription by eukaryotic RNA polymerases (Pols) II and III and archaeal Pol requires structurally related general transcription factors TFIIB, Brf1, and TFB, respectively, which are essential for polymerase recruitment and initiation events. A TFIIB-like protein was not evident in the Pol I basal transcription machinery. We report that TAF1B, a subunit of human Pol I basal transcription factor SL1, is structurally related to TFIIB/TFIIB-like proteins, through predicted amino-terminal zinc ribbon and cyclin-like fold domains.

RNA polymerase I-specific subunits promote polymerase clustering to enhance the rRNA gene transcription cycle.

RNA polymerase I (Pol I) produces large ribosomal RNAs (rRNAs). In this study, we show that the Rpa49 and Rpa34 Pol I subunits, which do not have counterparts in Pol II and Pol III complexes, are functionally conserved using heterospecific complementation of the human and Schizosaccharomyces pombe orthologues in Saccharomyces cerevisiae. Deletion of RPA49 leads to the disappearance of nucleolar structure, but nucleolar assembly can be restored by decreasing ribosomal gene copy number from 190 to 25.

FACT facilitates chromatin transcription by RNA polymerases I and III.

Efficient transcription elongation from a chromatin template requires RNA polymerases (Pols) to negotiate nucleosomes. Our biochemical analyses demonstrate that RNA Pol I can transcribe through nucleosome templates and that this requires structural rearrangement of the nucleosomal core particle. The subunits of the histone chaperone FACT (facilitates chromatin transcription), SSRP1 and Spt16, co-purify and co-immunoprecipitate with mammalian Pol I complexes.

Structure and function of ribosomal RNA gene chromatin.

Transcription of the major ribosomal RNAs by Pol I (RNA polymerase I) is a key determinant of ribosome biogenesis, driving cell growth and proliferation in eukaryotes. Hundreds of copies of rRNA genes are present in each cell, and there is evidence that the cellular control of Pol I transcription involves adjustments to the number of rRNA genes actively engaged in transcription, as well as to the rate of transcription from each active gene. Chromatin structure is inextricably linked to rRNA gene activity, and the present review highlights recent advances in this area.

A novel TBP-associated factor of SL1 functions in RNA polymerase I transcription.

In mammalian RNA polymerase I transcription, SL1, an assembly of TBP and associated factors (TAFs), is essential for preinitiation complex formation at ribosomal RNA gene promoters in vitro. We provide evidence for a novel component of SL1, TAF(I)41 (MGC5306), which functions in Pol I transcription. TAF(I)41 resides at the rDNA promoter in the nucleolus and co-purifies and co-immunoprecipitates with SL1.

Casein kinase 2 associates with initiation-competent RNA polymerase I and has multiple roles in ribosomal DNA transcription.

Mammalian RNA polymerase I (Pol I) complexes contain a number of associated factors, some with undefined regulatory roles in transcription. We demonstrate that casein kinase 2 (CK2) in human cells is associated specifically only with the initiation-competent Pol Ibeta isoform and not with Pol Ialpha. Chromatin immunoprecipitation analysis places CK2 at the ribosomal DNA (rDNA) promoter in vivo.

UBF activates RNA polymerase I transcription by stimulating promoter escape.

Ribosomal RNA gene transcription by RNA polymerase I (Pol I) is the driving force behind ribosome biogenesis, vital to cell growth and proliferation. The key activator of Pol I transcription, UBF, has been proposed to act by facilitating recruitment of Pol I and essential basal factor SL1 to rDNA promoters. However, we found no evidence that UBF could stimulate recruitment or stabilization of the pre-initiation complex (PIC) in reconstituted transcription assays.

RNA polymerase I-specific subunit CAST/hPAF49 has a role in the activation of transcription by upstream binding factor.

Eukaryotic RNA polymerases are large complexes, 12 subunits of which are structurally or functionally homologous across the three polymerase classes. Each class has a set of specific subunits, likely targets of their cognate transcription factors. We have identified and characterized a human RNA polymerase I (Pol I)-specific subunit, previously identified as ASE-1 (antisense of ERCC1) and as CD3epsilon-associated signal transducer (CAST), and here termed CAST or human Pol I-associated factor of 49 kDa (hPAF49), after mouse orthologue PAF49.

The RNA polymerase I transcription machinery.

The rRNAs constitute the catalytic and structural components of the ribosome, the protein synthesis machinery of cells. The level of rRNA synthesis, mediated by Pol I (RNA polymerase I), therefore has a major impact on the life and destiny of a cell. In order to elucidate how cells achieve the stringent control of Pol I transcription, matching the supply of rRNA to demand under different cellular growth conditions, it is essential to understand the components and mechanics of the Pol I transcription machinery.

TBP-TAF complex SL1 directs RNA polymerase I pre-initiation complex formation and stabilizes upstream binding factor at the rDNA promoter.

Knowledge of the role of components of the RNA polymerase I transcription machinery is paramount to understanding regulation of rDNA expression. We describe key findings for the roles of essential transcription factor SL1 and activator upstream binding factor (UBF). We demonstrate that human SL1 can direct accurate Pol I transcription in the absence of UBF and can interact with the rDNA promoter independently and stably, consistent with studies of rodent SL1 but contrary to previous reports of human SL1.

RNA-polymerase-I-directed rDNA transcription, life and works.

In the extensive network of interdependent biochemical processes required for cell growth and division, there is mounting evidence that ribosomal DNA transcription by RNA polymerase I (pol I) not only drives cell growth via its direct role in production of the ribosomal RNA (rRNA) component of the protein-synthesis machinery, but that it is also crucial in determining the fate of the cell. Considerable progress has been made in recent years towards understanding both the function of components of the pol I transcription machinery and how cells accomplish the tight control of pol I transcription, balancing the supply of rRNA with demand under different growth conditions.

Phosphatidylinositol 3-kinase and mTOR signaling pathways regulate RNA polymerase I transcription in response to IGF-1 and nutrients.

Regulation of ribosomal RNA gene transcription by RNA polymerase I (Pol I) is fundamental to ribosome biogenesis and therefore protein translation capacity and cell growth, yet little is known of the key signaling cascades involved. We show here that insulin-like growth factor-1 (IGF-1)-induced Pol I transcription in HEK293 cells is entirely dependent on phosphatidylinositol 3-kinase (PI3K) activity and, additionally, is modulated by the mammalian target of rapamycin (mTOR), which coordinates Pol I transcription with the availability of amino acids. The mitogen-activated protein kinase (MAPK) pathway is weakly stimulated by IGF-1 in these cells and partly contributes to Pol I transcription regulation.