Faculty Opinions recommendation of c-Myc binds to human ribosomal DNA and stimulates transcription of rRNA genes by RNA polymerase I.

ABSTRACT PpLSU3, a mobile group I intron in the rRNA genes of Physarum polycephalum, also can home into yeast chromosomal ribosomal DNA (rDNA) (D. E. Muscarella and V. M. Vogt, Mol. Cell. Biol. 13:1023–1033, 1993). By integrating PpLSU3 into the rDNA copies of a yeast strain temperature sensitive for RNA polymerase I, we have shown that the I-PpoI homing endonuclease encoded by PpLSU3 is expressed from an RNA polymerase I transcript. We have also developed a method to integrate mutant forms of PpLSU3 as well as theTetrahymena intron TtLSU1 into rDNA, by expressing I-PpoI in trans. Analysis of I-PpoI expression levels in these mutants, along with subcellular fractionation of intron RNA, strongly suggests that the full-length excised intron RNA, but not RNAs that are further cleaved, serves as or gives rise to the mRNA.

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Gene RRN4 in Saccharomyces cerevisiae encodes the A12.2 subunit of RNA polymerase I and is essential only at high temperatures.

Molecular and Cellular Biology ◽

10.1128/mcb.13.1.114 ◽

1993 ◽

Vol 13 (1) ◽

pp. 114-122 ◽

Cited By ~ 59

Author(s):

Y Nogi ◽

R Yano ◽

J Dodd ◽

C Carles ◽

M Nomura

Keyword(s):

Saccharomyces Cerevisiae ◽

Amino Acid ◽

Amino Acid Sequence ◽

Rna Polymerase ◽

Rna Polymerase I ◽

Rrna Genes ◽

Temperature Sensitive ◽

Polymerase I ◽

Temperature Sensitive Phenotype ◽

Null Mutants

We have previously isolated mutants of Saccharomyces cerevisiae that are primarily defective in transcription of 35S rRNA genes by RNA polymerase I and have identified genes (RRN1 to RRN9) involved in this process. We have now cloned the RRN4 gene by complementation of the temperature-sensitive phenotype of the rrn4-1 mutant and have determined its complete nucleotide sequence. The following results demonstrate that the RRN4 gene encodes the A12.2 subunit of RNA polymerase I. First, RRN4 protein expressed in Escherichia coli reacted with a specific antiserum against A12.2. Second, amino acid sequences of three tryptic peptides obtained from A12.2 were determined, and these sequences are found in the deduced amino acid sequence of the RRN4 protein. The amino acid sequence of the RRN4 protein (A12.2) is similar to that of the RPB9 (B12.6) subunit of yeast RNA polymerase II; the similarity includes the presence of two putative zinc-binding domains. Thus, A12.2 is a homolog of B12.6. We propose to rename the RRN4 gene RPA12. Deletion of RPA12 produces cells that are heat but not cold sensitive for growth. We have found that in such null mutants growing at permissive temperatures, the cellular concentration of A190, the largest subunit of RNA polymerase I, is lower than in the wild type. In addition, the temperature-sensitive phenotype of the rpa12 null mutants can be partially suppressed by RPA190 (the gene for A190) on multicopy plasmids. These results suggest that A12.2 plays a role in the assembly of A190 into a stable polymerase I structure.

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Role of TATA Binding Protein (TBP) in Yeast Ribosomal DNA Transcription by RNA Polymerase I: Defects in the Dual Functions of Transcription Factor UAF Cannot Be Suppressed by TBP

Molecular and Cellular Biology ◽

10.1128/mcb.21.7.2292-2297.2001 ◽

2001 ◽

Vol 21 (7) ◽

pp. 2292-2297 ◽

Cited By ~ 14

Author(s):

Imran Siddiqi ◽

John Keener ◽

Loan Vu ◽

Masayasu Nomura

Keyword(s):

Rna Polymerase ◽

Ribosomal Dna ◽

Binding Protein ◽

Tata Binding Protein ◽

Rna Polymerase I ◽

Pol Ii ◽

Rrna Transcription ◽

Pol I ◽

Mutant Strains ◽

Polymerase I

ABSTRACT Initiation of ribosomal DNA (rDNA) transcription by RNA polymerase I (Pol I) in the yeast Saccharomyces cerevisiae involves upstream activation factor (UAF), core factor, the TATA binding protein (TBP), and Rrn3p in addition to Pol I. We found previously that yeast strains carrying deletions in the UAF component RRN9switch completely to the use of Pol II for rRNA transcription, with no residual Pol I transcription. These polymerase-switched strains initially grow very slowly, but subsequent expansion in the number of rDNA repeats on chromosome XII leads to better growth. Recently, it was reported that TBP overexpression could bypass the requirement of UAF for Pol I transcription in vivo, producing nearly wild-type levels of growth in UAF mutant strains (P. Aprikian, B. Moorefield, and R. H. Reeder, Mol. Cell. Biol. 20:5269–5275, 2000). Here, we demonstrate that deletions in the UAF component RRN5,RRN9, or RRN10 lead to Pol II transcription of rDNA. TBP overexpression does not suppress UAF mutation, and these strains continue to use Pol II for rRNA transcription. We do not find evidence for even low levels of Pol I transcription in UAF mutant strains carrying overexpressed TBP. In diploid strains lacking both copies of the UAF componentRRN9, Pol II transcription of rDNA is more strongly repressed than in haploid strains but TBP overexpression still fails to activate Pol I. These results emphasize that UAF plays an essential role in activation of Pol I transcription and silencing of Pol II transcription of rDNA and that TBP functions to recruit the Pol I machinery in a manner completely dependent on UAF.

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Nucleolin Is Required for RNA Polymerase I Transcription In Vivo

Molecular and Cellular Biology ◽

10.1128/mcb.01584-06 ◽

2006 ◽

Vol 27 (3) ◽

pp. 937-948 ◽

Cited By ~ 76

Author(s):

Brenden Rickards ◽

S. J. Flint ◽

Michael D. Cole ◽

Gary LeRoy

Keyword(s):

Rna Polymerase ◽

Rna Polymerase Ii ◽

Rna Polymerase I ◽

Rrna Genes ◽

Accessory Proteins ◽

Naked Dna ◽

Nucleolar Chromatin ◽

Polymerase I

ABSTRACT Eukaryotic genomes are packaged with histones and accessory proteins in the form of chromatin. RNA polymerases and their accessory proteins are sufficient for transcription of naked DNA, but not of chromatin, templates in vitro. In this study, we purified and identified nucleolin as a protein that allows RNA polymerase II to transcribe nucleosomal templates in vitro. As immunofluorescence confirmed that nucleolin localizes primarily to nucleoli with RNA polymerase I, we demonstrated that nucleolin allows RNA polymerase I transcription of chromatin templates in vitro. The results of chromatin immunoprecipitation experiments established that nucleolin is associated with chromatin containing rRNA genes transcribed by RNA polymerase I but not with genes transcribed by RNA polymerase II or III. Knockdown of nucleolin by RNA interference resulted in specific inhibition of RNA polymerase I transcription. We therefore propose that an important function of nucleolin is to permit RNA polymerase I to transcribe nucleolar chromatin.

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Drug-induced dispersal of transcribed rRNA genes and transcriptional products: immunolocalization and silver staining of different nucleolar components in rat cells treated with 5,6-dichloro-beta-D-ribofuranosylbenzimidazole.

The Journal of Cell Biology ◽

10.1083/jcb.99.2.672 ◽

1984 ◽

Vol 99 (2) ◽

pp. 672-679 ◽

Cited By ~ 57

Author(s):

U Scheer ◽

B Hügle ◽

R Hazan ◽

K M Rose

Keyword(s):

Rna Polymerase ◽

Organizer Region ◽

Nucleolar Organizer Region ◽

Ribosomal Subunit ◽

Rna Polymerase I ◽

Rrna Genes ◽

Rrna Gene ◽

Linear Distribution ◽

Nucleolar Material ◽

Polymerase I

Upon incubation of cultured rat cells with the adenosine analogue 5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole (DRB), nucleoli reversibly dissociate into their substructures, disperse throughout the nuclear interior, and form nucleolar "necklaces". We have used this experimental system, which does not inhibit transcription of the rRNA genes, to study by immunocytochemistry the distribution of active rRNA genes and their transcriptional products during nucleolar dispersal and recovery to normal morphology. Antibodies to RNA polymerase I allow detection of template-engaged polymerase, and monoclonal antibodies to a ribosomal protein (S1) of the small ribosomal subunit permit localization of nucleolar preribosomal particles. The results show that, under the action of DRB transcribed rRNA, genes spread throughout the nucleoplasm and finally appear in the form of several rows, each containing several (up to 30) granules positive for RNA polymerase I and argyrophilic proteins. Nucleolar material containing preribosomal particles also appears in granular structures spread over the nucleoplasm but its distribution is distinct from that of rRNA gene-containing granules. We conclude that, although transcriptional units and preribosomal particles are both redistributed in response to DRB, these entities retain their individuality as functionally defined subunits. We further propose that each RNA polymerase-positive granular unit represents a single transcription unit and that each continuous array of granules ("string of nucleolar beads") reflects the linear distribution of rRNA genes along a nucleolar organizer region. Based on the total number of polymerase I-positive granules we estimate that a minimum of 60 rRNA genes are active during interphase of DRB-treated rat cells.

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In vivo evidence that TATA-binding protein/SL1 colocalizes with UBF and RNA polymerase I when rRNA synthesis is either active or inactive.

The Journal of Cell Biology ◽

10.1083/jcb.133.2.225 ◽

1996 ◽

Vol 133 (2) ◽

pp. 225-234 ◽

Cited By ~ 97

Author(s):

P Jordan ◽

M Mannervik ◽

L Tora ◽

M Carmo-Fonseca

Keyword(s):

Rna Polymerase ◽

Actinomycin D ◽

Binding Protein ◽

Tata Binding Protein ◽

Rna Polymerase I ◽

Rrna Genes ◽

Rrna Synthesis ◽

Rrna Transcription ◽

Polymerase I

Here we show that the TATA-binding protein (TBP) is localized in the nucleoplasm and in the nucleolus of mammalian cells, consistent with its known involvement in transcription by RNA polymerase I, II, and III. In the nucleolus of actively growing cells, TBP colocalizes with upstream binding factor (UBF) and RNA polymerase I at the sites of rRNA transcription. During mitosis, when rRNA synthesis is down-regulated, TBP colocalizes with TBP-associated factors for RNA polymerase I (TAF(I)s), UBF, and RNA polymerase I on the chromosomal regions containing the rRNA genes. Treatment of cells with a low concentration of actinomycin D inhibits rRNA synthesis and causes a redistribution of the rRNA genes that become concentrated in clusters at the periphery of the nucleolus. A similar redistribution was observed for the major components of the rRNA transcription machinery (i.e., TBP, TAF(I)s, UBF, and RNA polymerase I), which still colocalized with each other. Furthermore, anti-TBP antibodies are shown to coimmunoprecipitate TBP and TAF(I)63 in extracts prepared from untreated and actinomycin D-treated cells. Collectively, the data indicate that in vivo TBP/promoter selectivity factor, UBF, and RNA polymerase I remain associated with both active and inactive rRNA genes.

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