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PMID: 8381119 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

Mitotic repression of transcription in vitro.

The Journal of cell biology ·Vol. 120 ·No. 3 ·1993-02-00 ·Pages 613-24

Hartl P, Gottesfeld J, Forbes DJ

Abstract

A normal consequence of mitosis in eukaryotes is the repression of transcription. Using Xenopus egg extracts shifted to a mitotic state by the addition of purified cyclin, we have for the first time been able to reproduce a mitotic repression of transcription in vitro. Active RNA polymerase III transcription is observed in interphase extracts, but strongly repressed in extracts converted to mitosis. With the topoisomerase II inhibitor VM-26, we demonstrate that this mitotic repression of RNA polymerase III transcription does not require normal chromatin condensation. Similarly; in vitro mitotic repression of transcription does not require the presence of nucleosome structure or involve a general repressive chromatin-binding protein, as inhibition of chromatin formation with saturating amounts of non-specific DNA has no effect on repression. Instead, the mitotic repression of transcription appears to be due to phosphorylation of a component of the transcription machinery by a mitotic protein kinase, either cdc2 kinase and/or a kinase activated by it. Mitotic repression of RNA polymerase III transcription is observed both in complete mitotic cytosol and when a kinase-enriched mitotic fraction is added to a highly simplified 5S RNA transcription reaction. We present evidence that, upon depletion of cdc2 kinase, a secondary protein kinase activity remains and can mediate this in vitro mitotic repression of transcription.

MeSH Terms
Animals CDC2 Protein Kinase/isolation & purification,metabolism Cell-Free System Cyclins/pharmacology Female Interphase/physiology Mitosis/drug effects,physiology Models, Biological Oocytes/cytology,physiology Protamine Kinase/metabolism Protein Kinases/isolation & purification,metabolism RNA Polymerase III/metabolism Templates, Genetic Teniposide/pharmacology Topoisomerase II Inhibitors Transcription Factors/isolation & purification,metabolism Transcription, Genetic/drug effects Xenopus laevis
Chemicals
Cyclins Topoisomerase II Inhibitors Transcription Factors Teniposide Protein Kinases Protamine Kinase CDC2 Protein Kinase RNA Polymerase III
Authors & Affiliations
3 authors, click to expand affiliations / ORCID
Hartl P
Department of Biology, University of California, San Diego, La Jolla 92093-0322.
Gottesfeld J
Forbes D J
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Article Info
Journal
The Journal of cell biology
Abbr.
J Cell Biol
ISSN
0021-9525
DOI
10.1083/jcb.120.3.613
Published
1993-02-00
Pages
613-24
Language
English
Region
United States
NLM ID
0375356
PMCID
PMC2119533
Subset
IM
Grants
NIGMS NIH HHS · R01 GM033279 · United States
NIGMS NIH HHS · GM26453 · United States
NIGMS NIH HHS · GM33279 · United States
External Links
PubMed source DOI: 10.1083/jcb.120.3.613 PMC Full Text View journal details
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