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Initiation of DNA replication at the human beta-globin 3' enhancer
Alla Buzina1, Mirit I Aladjem, John L Kolman
1Developmental Biology Program, Hospital for Sick Children Toronto, Ontario, Canada.
Nucleic Acids Research
|August 9, 2005
Summary
Researchers identified a new DNA replication origin in the human beta-globin gene. This finding reveals that a mammalian enhancer cooperates with nearby sequences to form an efficient replication module.
Area of Science:
- Genetics
- Molecular Biology
- Epigenetics
Background:
- The human beta-globin gene's DNA replication origin involves an initiation region (IR) and flanking elements.
- While upstream and IR core sequences contain replicator modules, the downstream auxiliary element's function was unclear.
Purpose of the Study:
- To identify functional sequences within the downstream auxiliary element of the beta-globin gene's replication origin.
- To characterize the role of the 3' enhancer in DNA replication initiation and replicator module formation.
Main Methods:
- Benzoylated-naphthoylated DEAE (BND) cellulose purification and nascent strand abundance assays were employed.
- FRT/EGFP-FLP integration system was used to insert 3' enhancer fragments into an ectopic site in murine erythroleukemia (MEL) cells.
Main Results:
- Replication initiation was localized to the beta-globin 3' enhancer on human chromosome 11 in Hu11 MEL cells.
- The 1.6 kb downstream auxiliary element functions as a third replicator module (bGRep-E) in erythroid cells.
- The minimal 3' enhancer requires cooperation with adjacent sequences for efficient replication initiation.
Conclusions:
- The beta-globin replicator possesses multiple initiation sites within three distinct, closely located replicator modules.
- Mammalian enhancers can collaborate with adjacent sequences to establish functional replicator modules, expanding our understanding of DNA replication origins.