A novel ribonucleoprotein complex enriched in nucleolar proteins was purified from yeast extracts and constituents were identified by mass spectrometry. When isolated from rapidly growing cells, the assembly contained ribonucleic acid (RNA) polymerase (pol) I, and some of its transcription factors like TATA-binding protein (TBP), Rrn3p, Rrn5p, Rrn7p, and Reb1p along with rRNA processing factors, like Nop1p, Cbf5p, Nhp2p, and Rrp5p. The small nucleolar RNAs (snoRNAs) U3, U14, and MRP were also found to be associated with the complex, which supports accurate transcription, termination, and pseudouridylation of rRNA. Formation of the complex did not depend on pol I, and the complex could efficiently recruit exogenous pol I into active ribosomal DNA (rDNA) transcription units. Visualization of the complex by electron microscopy and immunogold labeling revealed a characteristic cluster-forming network of nonuniform size containing nucleolar proteins like Nop1p and Fpr3p and attached pol I. Our results support the idea that a functional nucleolar subdomain formed independently of the state of rDNA transcription may serve as a scaffold for coordinated rRNA synthesis and processing.
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1 May 2000
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May 01 2000
Association of Yeast RNA Polymerase I with a Nucleolar Substructure Active in Rrna Synthesis and Processing
Stephan Fath,
Stephan Fath
aBiochemie-Zentrum Heidelberg, D-69120 Heidelberg, Germany
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Philipp Milkereit,
Philipp Milkereit
aBiochemie-Zentrum Heidelberg, D-69120 Heidelberg, Germany
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Alexandre V. Podtelejnikov,
Alexandre V. Podtelejnikov
bProtein Interaction Laboratory, Odense University, DK 5230 Odense Middle, Denmark
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Nicolas Bischler,
Nicolas Bischler
cInstitut de Génétique et de Biologie Moléculaire et Cellulaire, F-67404 Illkirch Cedex, France
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Patrick Schultz,
Patrick Schultz
cInstitut de Génétique et de Biologie Moléculaire et Cellulaire, F-67404 Illkirch Cedex, France
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Mirko Bier,
Mirko Bier
aBiochemie-Zentrum Heidelberg, D-69120 Heidelberg, Germany
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Matthias Mann,
Matthias Mann
bProtein Interaction Laboratory, Odense University, DK 5230 Odense Middle, Denmark
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Herbert Tschochner
Herbert Tschochner
aBiochemie-Zentrum Heidelberg, D-69120 Heidelberg, Germany
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Stephan Fath
aBiochemie-Zentrum Heidelberg, D-69120 Heidelberg, Germany
Philipp Milkereit
aBiochemie-Zentrum Heidelberg, D-69120 Heidelberg, Germany
Alexandre V. Podtelejnikov
bProtein Interaction Laboratory, Odense University, DK 5230 Odense Middle, Denmark
Nicolas Bischler
cInstitut de Génétique et de Biologie Moléculaire et Cellulaire, F-67404 Illkirch Cedex, France
Patrick Schultz
cInstitut de Génétique et de Biologie Moléculaire et Cellulaire, F-67404 Illkirch Cedex, France
Mirko Bier
aBiochemie-Zentrum Heidelberg, D-69120 Heidelberg, Germany
Matthias Mann
bProtein Interaction Laboratory, Odense University, DK 5230 Odense Middle, Denmark
Herbert Tschochner
aBiochemie-Zentrum Heidelberg, D-69120 Heidelberg, Germany
Abbreviations used in this paper: CF, core factor; HA, hemagglutinin; MALDI, matrix assisted laser desorption ionization; pol, RNA polymerase; rDNA, ribosomal DNA; snoRNA, small nucleolar RNA; TBP, TATA-binding protein; UAF, upstream activating factor; WCE, whole cell extract.
Received:
August 05 1999
Revision Requested:
March 27 2000
Accepted:
March 27 2000
Online ISSN: 1540-8140
Print ISSN: 0021-9525
© 2000 The Rockefeller University Press
2000
The Rockefeller University Press
J Cell Biol (2000) 149 (3): 575–590.
Article history
Received:
August 05 1999
Revision Requested:
March 27 2000
Accepted:
March 27 2000
Citation
Stephan Fath, Philipp Milkereit, Alexandre V. Podtelejnikov, Nicolas Bischler, Patrick Schultz, Mirko Bier, Matthias Mann, Herbert Tschochner; Association of Yeast RNA Polymerase I with a Nucleolar Substructure Active in Rrna Synthesis and Processing. J Cell Biol 1 May 2000; 149 (3): 575–590. doi: https://doi.org/10.1083/jcb.149.3.575
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