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Interaction of the H4 autonomously replicating sequence core consensus sequence and its 3'-flanking domain
1Department of Microbiology, School of Medicine, University of Virginia, Charlottesville 22908.
Molecular and Cellular Biology
|December 1, 1989
Summary
Yeast autonomously replicating sequence (ARS) elements require a specific orientation between the core consensus sequence and its 3'-flanking DNA for function. Additional near-consensus sequences in the flanking region are not essential for H4 ARS activity.
Area of Science:
- Molecular Biology
- Yeast Genetics
Background:
- Autonomously replicating sequence (ARS) elements in yeast are crucial for DNA replication initiation.
- ARS elements consist of a conserved core consensus sequence and a variable 3'-flanking region.
Purpose of the Study:
- To investigate the functional relationship between the H4 ARS core consensus sequence and its 3'-flanking domain.
- To determine the minimal sequence requirements and orientation dependency for H4 ARS function.
Main Methods:
- Deletional analysis to define minimal functional ARS fragments.
- Oligonucleotide mutagenesis to invert the core consensus sequence.
- Site-directed mutagenesis to disrupt near-consensus sequences in the 3'-flanking region.
Main Results:
- A minimum of 66 bp of 3'-flanking DNA is necessary for full H4 ARS function.
- Inversion of the core consensus sequence abolished ARS function, highlighting the importance of orientation.
- Disruption of multiple near-consensus sequences (8/11 bp match or better) did not impair ARS function.
Conclusions:
- The relative orientation of the core consensus and 3'-flanking domain is critical for H4 ARS function.
- Specific near-consensus sequences within the 3'-flanking region are not required for H4 ARS activity.