scholarly journals Migration of rat RNA polymerase I into chick erythrocyte nuclei undergoing reactivation in chick-rat heterokaryons.

1983 ◽  
Vol 97 (5) ◽  
pp. 1641-1643 ◽  
Author(s):  
U Scheer ◽  
G Lanfranchi ◽  
K M Rose ◽  
W W Franke ◽  
N R Ringertz
Keyword(s):  
Rna Polymerase ◽  
Immunofluorescence Microscopy ◽  
Sendai Virus ◽  
Specific Protein ◽  
Rna Polymerase I ◽  
Rrna Genes ◽  
Rrna Synthesis ◽  
Time Interval ◽  
Reactivation Process ◽  
Polymerase I

Transcriptionally inactive chick erythrocyte nuclei were reactivated by Sendai virus-induced fusion of erythrocytes with rat L6J1 myoblasts. We used antibodies to trace the appearance of a specific protein engaged in transcription of a defined class of genes, those coding for rRNA, during reactivation. Using immunofluorescence microscopy, we found increasing amounts of rat RNA polymerase I to appear, during a certain period of time after fusion, in the reforming nucleoli of the chick nuclei. Amounts of rat RNA polymerase I sufficient to be detected by immunofluorescence microscopy had accumulated in the newly developed chick nucleoli 72-190 h after fusion was initiated. This time interval coincides with the time when chick rRNA synthesis can first be detected. The results raise the possibility that during these stages of the reactivation process chick rRNA genes are transcribed by heterologous RNA polymerase I molecules of rat origin.

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

1996 ◽  
Vol 133 (2) ◽  
pp. 225-234 ◽  
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.

Three-dimensional organization of active rRNA genes within the nucleolus

2002 ◽  
Vol 115 (16) ◽  
pp. 3297-3307 ◽  
Author(s):  
Thierry Cheutin ◽  
Marie-Françoise O'Donohue ◽  
Adrien Beorchia ◽  
Marc Vandelaer ◽  
Hervé Kaplan ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Electron Tomography ◽  
Three Dimensional ◽  
Spatial Arrangement ◽  
Ultrastructural Level ◽  
Rna Polymerase I ◽  
Rrna Genes ◽  
Rrna Synthesis ◽  
Fibrillar Component ◽  
Polymerase I

In this work, we have localized transcribing rRNA genes at the ultrastructural level and described their three-dimensional organization within the nucleolus by electron tomography. Isolated nucleoli, which exhibit a reduced transcriptional rate, were used to determine the sites of initial BrUTP incorporation (i.e. rRNA synthesis by the transcriptional machinery). Using pulse-chase experiments with BrUTP and an elongation inhibitor,cordycepin, it was possible to precisely localize the initial sites of BrUTP incorporation. Our data show that BrUTP incorporation initially takes place in the fibrillar centers and that elongating rRNAs rapidly enter the surrounding dense fibrillar component. Furthermore, we investigated the spatial arrangement of RNA polymerase I molecules within the whole volume of the fibrillar centers. Electron tomography was performed on thick sections of cells that had been labeled with anti-RNA polymerase I antibodies prior to embedding. Detailed tomographic analyses revealed that RNA polymerase I molecules are mainly localized within discrete clusters. In each of them, RNA polymerase I molecules were grouped as several coils, 60 nm in diameter. Overall, these findings have allowed us to propose a model for the three-dimensional organization of transcribing rDNA genes within the nucleolus.

scholarly journals Compositional reorganization of the nucleolus in budding yeast mitosis

2019 ◽  
Vol 30 (5) ◽  
pp. 591-606 ◽  
Author(s):  
Philipp Girke ◽  
Wolfgang Seufert
Keyword(s):  
Rna Polymerase ◽  
Budding Yeast ◽  
Protein Composition ◽  
Rna Polymerase I ◽  
Rrna Genes ◽  
Rrna Synthesis ◽  
Rdna Transcription ◽  
Unequal Distribution ◽  
Polymerase I ◽  
Nucleolar Segregation

The nucleolus is a membraneless organelle of the nucleus and the site of rRNA synthesis, maturation, and assembly into preribosomal particles. The nucleolus, organized around arrays of rRNA genes (rDNA), dissolves during prophase of mitosis in metazoans, when rDNA transcription ceases, and reforms in telophase, when rDNA transcription resumes. No such dissolution and reformation cycle exists in budding yeast, and the precise course of nucleolar segregation remains unclear. By quantitative live-cell imaging, we observed that the yeast nucleolus is reorganized in its protein composition during mitosis. Daughter cells received equal shares of preinitiation factors, which bind the RNA polymerase I promoter and the rDNA binding barrier protein Fob1, but only about one-third of RNA polymerase I and the processing factors Nop56 and Nsr1. The distribution bias was diminished in nonpolar chromosome segregation events observable in dyn1 mutants. Unequal distribution, however, was enhanced by defects in RNA polymerase I, suggesting that rDNA transcription supports nucleolar segregation. Indeed, quantification of pre-rRNA levels indicated ongoing rDNA transcription in yeast mitosis. These data, together with photobleaching experiments to measure nucleolar protein dynamics in anaphase, consolidate a model that explains the differential partitioning of nucleolar components in budding yeast mitosis.

scholarly journals Human Nopp140, Which Interacts with RNA Polymerase I: Implications for rRNA Gene Transcription and Nucleolar Structural Organization

1999 ◽  
Vol 19 (12) ◽  
pp. 8536-8546 ◽  
Author(s):  
Hung-Kai Chen ◽  
Chi-Yun Pai ◽  
Jing-Yi Huang ◽  
Ning-Hsing Yeh
Keyword(s):  
Amino Acids ◽  
Rna Polymerase ◽  
Gene Transcription ◽  
Ectopic Expression ◽  
Rna Polymerase I ◽  
Rrna Genes ◽  
Rrna Synthesis ◽  
Rrna Gene ◽  
Amino Terminal ◽  
Polymerase I

ABSTRACT Nopp140 is thought to shuttle between nucleolus and cytoplasm. However, the predominant nucleolar localization of Nopp140 homologues from different species suggests that Nopp140 is also involved in events occurring within the nucleolus. In this study, we demonstrated that the largest subunit of RNA polymerase I, RPA194, was coimmunoprecipitated with the human Nopp140 (hNopp140). Such an interaction is mediated through amino acids 204 to 382 of hNopp140. By double immunofluorescence, hNopp140 was colocalized with RNA polymerase I at the rDNA (rRNA genes) transcription active foci in the nucleolus. These results suggest that Nopp140 can interact with RNA polymerase I in vivo. Transfected cells expressing the amino-terminal half of hNopp140, hNopp140N382 (amino acids 1 to 382), displayed altered nucleoli with crescent-shaped structures. This phenotype is reminiscent of the segregated nucleoli induced by actinomycin D treatment, which is known to inhibit rRNA synthesis. Consistently, the hNopp140N382 protein mislocalized the endogenous RNA polymerase I and shut off cellular rRNA gene transcription as revealed by an in situ run-on assay. These dominant negative effects of the mutant hNopp140N382 suggest that Nopp140 plays an essential role in rDNA transcription. Interestingly, ectopic expression of hNopp140 to a very high level caused the formation of a transcriptionally inactive spherical structure occupying the entire nucleolar area which trapped the RNA polymerase I, fibrillarin, and hNopp140 but excluded the nucleolin. The mislocalizations of these nucleolar proteins after hNopp140 overexpression imply that Nopp140 may also play roles in maintenance of nucleolar integrity.

scholarly journals I-PpoI, the Endonuclease Encoded by the Group I Intron PpLSU3, Is Expressed from an RNA Polymerase I Transcript

1998 ◽  
Vol 18 (10) ◽  
pp. 5809-5817 ◽  
Author(s):  
Jue Lin ◽  
Volker M. Vogt
Keyword(s):  
Rna Polymerase ◽  
Homing Endonuclease ◽  
Group I Intron ◽  
Subcellular Fractionation ◽  
Rna Polymerase I ◽  
Rrna Genes ◽  
Temperature Sensitive ◽  
Group I ◽  
Polymerase I ◽  
Mutant Forms

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.

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

1993 ◽  
Vol 13 (1) ◽  
pp. 114-122 ◽  
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.

scholarly journals The RNA polymerase I transcription machinery

2006 ◽  
Vol 73 ◽  
pp. 203-216 ◽  
Author(s):  
Jackie Russell ◽  
Joost C.B.M. Zomerdijk
Keyword(s):  
Rna Polymerase ◽  
Mammalian Cells ◽  
Growth Conditions ◽  
Rna Polymerase I ◽  
Cellular Growth ◽  
Rrna Synthesis ◽  
Transcription Machinery ◽  
Pol I ◽  
A Cell ◽  
Polymerase I

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. In this review, we discuss: (i) the molecular composition and functions of the Pol I enzyme complex and the two main Pol I transcription factors, SL1 (selectivity factor 1) and UBF (upstream binding factor); (ii) the interplay between these factors during pre-initiation complex formation at the rDNA promoter in mammalian cells; and (iii) the cellular control of the Pol I transcription machinery.

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