1A6/DRIM, a Novel t-UTP, Activates RNA Polymerase I Transcription and Promotes Cell Proliferation

BACKGROUND: Ribosome biogenesis is required for protein synthesis and cell proliferation. Ribosome subunits are assembled in the nucleolus following transcription of a 47S ribosome RNA precursor by RNA polymerase I and rRNA processing to produce mature 18S, 28S and 5.8S rRNAs. The 18S rRNA is incorporated into the ribosomal small subunit, whereas the 28S and 5.8S rRNAs are incorporated into the ribosomal large subunit. Pol I transcription and rRNA processing are coordinated processes and this coordination has been demonstrated to be mediated by a subset of U3 proteins known as t-UTPs. Up to date, five t-UTPs have been identified in humans but the mechanism(s) that function in the t-UTP(s) activation of Pol I remain unknown. In this study we have identified 1A6/DRIM, which was identified as UTP20 in our previous study, as a t-UTP. In the present study, we investigated the function and mechanism of 1A6/DRIM in Pol I transcription. METHODOLOGY/PRINCIPAL FINDINGS: Knockdown of 1A6/DRIM by siRNA resulted in a decreased 47S pre-rRNA level as determined by Northern blotting. Ectopic expression of 1A6/DRIM activated and knockdown of 1A6/DRIM inhibited the human rDNA promoter as evaluated with luciferase reporter. Chromatin immunoprecipitation (ChIP) experiments showed that 1A6/DRIM bound UBF and the rDNA promoter. Re-ChIP assay showed that 1A6/DRIM interacts with UBF at the rDNA promoter. Immunoprecipitation confirmed the interaction between 1A6/DRIM and the nucleolar acetyl-transferase hALP. It is of note that knockdown of 1A6/DRIM dramatically inhibited UBF acetylation. A finding of significance was that 1A6/DRIM depletion, as a kind of nucleolar stress, caused an increase in p53 level and inhibited cell proliferation by arresting cells at G1. CONCLUSIONS: We identify 1A6/DRIM as a novel t-UTP. Our results suggest that 1A6/DRIM activates Pol I transcription most likely by associating with both hALP and UBF and thereby affecting the acetylation of UBF

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. In this, UBF is fundamentally different from archetypal activators of transcription. Our data imply that UBF exerts its stimulatory effect on RNA synthesis, after PIC formation, promoter opening and first phosphodiester bond formation and before elongation. We provide evidence to suggest that UBF activates transcription in the transition between initiation and elongation, at promoter escape by Pol I. This novel role for UBF in promoter escape would allow control of rRNA synthesis at active rDNA repeats, independent of and complementary to the promoter-specific targeting of SL1 and Pol I during PIC assembly. We posit that stimulation of promoter escape could be a general mechanism of activator function

NoRC-dependent nucleosome positioning silences rRNA genes

Previous studies have established that the Snf2h-containing chromatin remodeling complex NoRC mediates epigenetic silencing of a subset of rRNA genes (rDNA) by recruiting enzymatic activities that modify histones and methylate DNA. Here we have analyzed nucleosome positions at the murine rDNA promoter and show that active and silent rDNA copies are characterized not only by specific epigenetic marks but also by differently positioned nucleosomes. At active genes the promoter-bound nucleosome covers nucleotides from −157 to −2, whereas at silent genes the nucleosome is positioned 25 nucleotides further downstream. We provide evidence that NoRC is the molecular machine that shifts the promoter-bound nucleosome downstream of the transcription start site into a translational position that is unfavorable for transcription complex formation

DNA methyltransferase inhibition may limit cancer cell growth by disrupting ribosome biogenesis

Mutations in the pattern of CpG methylation imprinting of the human genome have been correlated with a number of diseases including cancer. In particular, aberrant imprinting of tumor suppressor genes by gain of CpG methylation has been observed in many cancers and thus represents an important alternative pathway to gene mutation and tumor progression. Inhibitors of DNA methylation display therapeutic effects in the treatment of certain cancers and it has been assumed that these effects are due to the reversal of mutant gene imprinting. However, significant reactivation of imprinted tumor suppressor genes is rarely observed in vivo following treatment with DNA methylation inhibitors. A recent study revealed an unexpected requirement for CpG methylation in the synthesis and assembly of the ribosome, an essential function for cell growth and proliferation. As such, the data provide an unforeseen explanation of the action of DNA methylation inhibitors in restricting cancer cell growth