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PMID: 8413256 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

Initiation and termination of DNA replication in human rRNA genes.

Molecular and cellular biology ·Vol. 13 ·No. 10 ·1993-10-00 ·Pages 6600-13

Little RD, Platt TH, Schildkraut CL

Abstract

We have used the multicopy human rRNA genes as a model system to study replication initiation and termination in mammalian chromosomes. Enrichment for replicating molecules was achieved by isolating S-phase enriched populations of cells by centrifugal elutriation, purification of DNA associated with the nuclear matrix, and a chromatographic procedure that enriches for molecules containing single-stranded regions, a characteristic of replication forks. Two-dimensional agarose gel electrophoresis techniques were used to demonstrate that replication appears to initiate at multiple sites throughout most of the 31-kb nontranscribed spacer (NTS) of human ribosomal DNA but not within the 13-kb transcription unit or adjacent regulatory elements. Although initiation events were detected throughout the majority of the NTS, some regions may initiate more frequently than others. Termination of replication, the convergence of opposing replication forks, was found throughout the ribosomal DNA repeat units, and, in some repeats, specifically at the junction of the 3' end of the transcription unit and the NTS. This site-specific termination of replication is the result of pausing of replication forks near the sites of transcription termination. The naturally occurring multicopy rRNA gene family offers a unique system to study mammalian DNA replication without the use of chemical synchronization agents.

MeSH Terms
Animals CHO Cells Cricetinae DNA Replication Electrophoresis, Gel, Two-Dimensional Humans RNA, Ribosomal/genetics Restriction Mapping
Chemicals
RNA, Ribosomal
Authors & Affiliations
3 authors, click to expand affiliations / ORCID
Little R D
Department of Cell Biology, Albert Einstein College of Medicine, Bronx, New York 10461.
Platt T H
Schildkraut C L
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Article Info
Journal
Molecular and cellular biology
Abbr.
Mol Cell Biol
ISSN
0270-7306
DOI
10.1128/mcb.13.10.6600-6613.1993
Published
1993-10-00
Pages
6600-13
Language
English
Region
United States
NLM ID
8109087
PMCID
PMC364718
Subset
IM
Grants
NCI NIH HHS · NCI P30-CA13330 · United States
NIGMS NIH HHS · R01 GM45751 · United States
External Links
PubMed source DOI: 10.1128/mcb.13.10.6600-6613.1993 PMC Full Text View journal details
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