Conditional expression of RPA190, the gene encoding the largest subunit of yeast RNA polymerase I: effects of decreased rRNA synthesis on ribosomal protein synthesis.

The synthesis of ribosomal proteins (r proteins) under the conditions of greatly reduced RNA synthesis were studied by using a strain of the yeast Saccharomyces cerevisiae in which the production of the largest subunit (RPA190) of RNA polymerase I was controlled by the galactose promoter. Although growth on galactose medium was normal, the strain was unable to sustain growth when shifted to glucose medium. This growth defect was shown to be due to a preferential decrease in RNA synthesis caused by deprivation of RNA polymerase I. Under these conditions, the accumulation of r proteins decreased to match the rRNA synthesis rate. When proteins were pulse-labeled for short periods, no or only a weak decrease was observed in the differential synthesis rate of several r proteins (L5, L39, L29 and/or L28, L27 and/or S21) relative to those of control cells synthesizing RPA190 from the normal promoter. Degradation of these r proteins synthesized in excess was observed during subsequent chase periods. Analysis of the amounts of mRNAs for L3 and L29 and their locations in polysomes also suggested that the synthesis of these proteins relative to other cellular proteins were comparable to those observed in control cells. However, Northern analysis of several r-protein mRNAs revealed that the unspliced precursor mRNA for r-protein L32 accumulated when rRNA synthesis rates were decreased. This result supports the feedback regulation model in which excess L32 protein inhibits the splicing of its own precursor mRNA, as proposed by previous workers (M. D. Dabeva, M. A. Post-Beittenmiller, and J. R. Warner, Proc. Natl. Acad. Sci. USA 83:5854-5857, 1986).

Download Full-text

Tor Pathway Regulates Rrn3p-dependent Recruitment of Yeast RNA Polymerase I to the Promoter but Does Not Participate in Alteration of the Number of Active Genes

Molecular Biology of the Cell ◽

10.1091/mbc.e03-08-0594 ◽

2004 ◽

Vol 15 (2) ◽

pp. 946-956 ◽

Cited By ~ 117

Author(s):

Jonathan A. Claypool ◽

Sarah L. French ◽

Katsuki Johzuka ◽

Kristilyn Eliason ◽

Loan Vu ◽

...

Keyword(s):

Rna Polymerase ◽

Stationary Phase ◽

Rna Polymerase I ◽

Rrna Synthesis ◽

Synthesis Rate ◽

Transcription Rate ◽

Signaling System ◽

Tor Signaling ◽

Polymerase I ◽

Active Genes

Yeast cells entering into stationary phase decrease rRNA synthesis rate by decreasing both the number of active genes and the transcription rate of individual active genes. Using chromatin immunoprecipitation assays, we found that the association of RNA polymerase I with the promoter and the coding region of rDNA is decreased in stationary phase, but association of transcription factor UAF with the promoter is unchanged. Similar changes were also observed when growing cells were treated with rapamycin, which is known to inhibit the Tor signaling system. Rapamycin treatment also caused a decrease in the amount of Rrn3p-polymerase I complex, similar to stationary phase. Because recruitment of Pol I to the rDNA promoter is Rrn3p-dependent as shown in this work, these data suggest that the decrease in the transcription rate of individual active genes in stationary phase is achieved by the Tor signaling system acting at the Rrn3p-dependent polymerase recruitment step. Miller chromatin spreads of cells treated with rapamycin and cells in post-log phase confirm this conclusion and demonstrate that the Tor system does not participate in alteration of the number of active genes observed for cells entering into stationary phase.

Download Full-text

Cytoplasmic to nuclear translocation of RNA polymerase I is required for lipopolysaccharide-induced nucleolar RNA synthesis and subsequent DNA synthesis in murine B lymphocytes

Journal of Cell Science ◽

10.1242/jcs.92.1.101 ◽

1989 ◽

Vol 92 (1) ◽

pp. 101-109

Author(s):

M.S. Halleck ◽

R.A. Schlegel ◽

K.M. Rose

Keyword(s):

Rna Polymerase ◽

Dna Synthesis ◽

B Lymphocytes ◽

Ribosomal Rna ◽

Rna Synthesis ◽

Nuclear Translocation ◽

Rna Polymerase I ◽

Nuclear Accumulation ◽

Rrna Synthesis ◽

Polymerase I

The synthesis of ribosomal RNA (rRNA) in murine B lymphocytes is markedly elevated in response to mitogens such as lipopolysaccharide (LPS). First, to investigate the mechanism involved, antibodies directed against RNA polymerase I, the enzyme responsible for transcription of ribosomal genes, were introduced into the cytoplasm of lymphocytes via red cell-mediated microinjection and the ability of cells to synthesize RNA was examined. Simultaneous immunofluorescence/autoradiography revealed that 7% or less of the cells injected with specific antibodies prior to stimulation were actively synthesizing rRNA 15 or 40 h following LPS addition. In contrast 19% and 27% of cells injected with control IgG were active at these times. Non-ribosomal RNA synthesis was unaffected by the presence of anti-RNA polymerase I antibodies. Since antibodies injected into the cytoplasm were limited to that compartment, these data suggest that rRNA synthesis induced by LPS requires translocation of cytoplasmic RNA polymerase I into the nucleus. Second, to test whether synthesis of rRNA is required for entry into S phase, the effect of anti-RNA polymerase I antibodies on DNA synthesis in response to LPS was evaluated. Only 7% of cells containing anti-RNA polymerase I antibodies had initiated DNA synthesis 40 h after LPS addition whereas 25% of cells containing control IgG were actively synthesizing DNA at that time. These results suggest that nuclear accumulation of RNA polymerase I and increased rRNA synthesis are required for LPS-induced DNA synthesis in B lymphocytes.

Download Full-text

Faculty Opinions recommendation of In exponentially growing Saccharomyces cerevisiae cells, rRNA synthesis is determined by the summed RNA polymerase I loading rate rather than by the number of active genes.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1012188.184671 ◽

2003 ◽

Author(s):

Jonathan R Warner

Keyword(s):

Saccharomyces Cerevisiae ◽

Rna Polymerase ◽

Loading Rate ◽

Rna Polymerase I ◽

Rrna Synthesis ◽

Polymerase I ◽

Active Genes

Download Full-text

Selective inhibition by zinc of RNA synthesis initiation in the RNA polymerase I reaction

FEBS Letters ◽

10.1016/0014-5793(79)80241-0 ◽

1979 ◽

Vol 99 (1) ◽

pp. 29-32 ◽

Cited By ~ 3

Author(s):

Yoshikuni Nagamine ◽

Den'ichi Mizuno ◽

Shunji Natori

Keyword(s):

Rna Polymerase ◽

Rna Synthesis ◽

Selective Inhibition ◽

Rna Polymerase I ◽

Polymerase I

Download Full-text

A specialized form of RNA polymerase I, essential for initiation and growth-dependent regulation of rRNA synthesis, is disrupted during transcription

The EMBO Journal ◽

10.1093/emboj/17.13.3692 ◽

1998 ◽

Vol 17 (13) ◽

pp. 3692-3703 ◽

Cited By ~ 89

Author(s):

P. Milkereit

Keyword(s):

Rna Polymerase ◽

Rna Polymerase I ◽

Rrna Synthesis ◽

Specialized Form ◽

Polymerase I

Download Full-text

Nucleolar protein upstream binding factor is sequestered into adenovirus DNA replication centres during infection without affecting RNA polymerase I location or ablating rRNA synthesis

Journal of Cell Science ◽

10.1242/jcs.02982 ◽

2006 ◽

Vol 119 (12) ◽

pp. 2621-2631 ◽

Cited By ~ 31

Author(s):

F. J. Lawrence

Keyword(s):

Dna Replication ◽

Rna Polymerase ◽

Rna Polymerase I ◽

Nucleolar Protein ◽

Rrna Synthesis ◽

Binding Factor ◽

Upstream Binding Factor ◽

Polymerase I

Download Full-text

The RNA polymerase I transcription machinery

Biochemical Society Symposium ◽

10.1042/bss0730203 ◽

2006 ◽

Vol 73 ◽

pp. 203-216 ◽

Cited By ~ 82

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.

Download Full-text

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.

Download Full-text