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PMID: 2160586 Published · ppublish English Journal Article Research Support, U.S. Gov't, Non-P.H.S. Research Support, U.S. Gov't, P.H.S.

Nucleosome assembly of simian virus 40 DNA in a mammalian cell extract.

Molecular and cellular biology ·Vol. 10 ·No. 6 ·1990-06-00 ·Pages 2863-73

Banerjee S, Cantor CR

Abstract

We report here a mammalian cell-free system that can support chromatin assembly. Effective nucleosome assembly in HeLa cell extracts occurred at 125 to 200 mM KCl or potassium glutamate. At this physiological K+ ion concentration, two types of chromatin assembly were observed. The first was interfered with by Mg2+. Other cations such as Mn2+, Ca2+, Fe3+, and spermidine also inhibited this type of nucleosome assembly. The second type of assembly occurred in the presence of Mg2+ and at least equimolar ATP. However, even in the presence of ATP, excess Mg2+ inhibited assembly and promoted catenation of DNA; these effects could be circumvented by excess ATP, GTP, EDTA, or polyglutamic acid. The critical DNA concentration for optimum assembly in both pathways suggested a stoichiometric association of histones with DNA. The spacing of nucleosomes formed by both types of assembly on linear and circular DNA was reasonably regular, but chromatin assembled in the presence of ATP and Mg2+ was more stable.

MeSH Terms
Adenosine Triphosphate/metabolism Anions Cations Cell Transformation, Viral Cell-Free System Chromatin/metabolism DNA, Superhelical/genetics DNA, Viral/genetics Edetic Acid/pharmacology HeLa Cells/metabolism Histones/metabolism Humans Kinetics Magnesium/pharmacology Nucleosomes/drug effects,metabolism Osmolar Concentration Simian virus 40/genetics,metabolism
Chemicals
Anions Cations Chromatin DNA, Superhelical DNA, Viral Histones Nucleosomes Adenosine Triphosphate Edetic Acid Magnesium
Authors & Affiliations
2 authors, click to expand affiliations / ORCID
Banerjee S
Department of Genetics and Development, College of Physicians and Surgeons, Columbia University, New York, New York 10032.
Cantor C R
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Article Info
Journal
Molecular and cellular biology
Abbr.
Mol Cell Biol
ISSN
0270-7306
DOI
10.1128/mcb.10.6.2863-2873.1990
Published
1990-06-00
Pages
2863-73
Language
English
Region
United States
NLM ID
8109087
PMCID
PMC360648
Subset
IM
Grants
NCI NIH HHS · CA 39782 · United States
NIGMS NIH HHS · GM 14825 · United States
External Links
PubMed source DOI: 10.1128/mcb.10.6.2863-2873.1990 PMC Full Text View journal details
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