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Saccharomyces cerevisiae Cdc6 stimulates Abf1 DNA binding activity
1Division of Hematology/Oncology, Childrens Hospital Los Angeles, University of Southern California School of Medicine 90027, USA.
The Journal of Biological Chemistry
|January 27, 1998
Summary
Yeast Cdc6 protein stimulates Abf1 DNA binding, crucial for DNA replication and transcription. This interaction, mediated by Cdc6 binding to DNA, suggests Cdc6 acts as an architectural factor in replication complex assembly.
Area of Science:
- Molecular Biology
- Yeast Genetics
- DNA Replication
Background:
- ARS binding factor 1 (Abf1) is essential for DNA replication and transcription in Saccharomyces cerevisiae.
- Cdc6 protein's role in regulating Abf1 DNA binding is not fully understood.
Purpose of the Study:
- To investigate how yeast Cdc6 protein influences Abf1 DNA binding activities.
- To elucidate the mechanism by which Cdc6 modulates Abf1's function.
Main Methods:
- Utilized a cdc6-1 mutant strain to assess Abf1 binding activity.
- Performed in vitro binding assays with recombinant Abf1 and GST-Cdc6 fusion protein.
- Conducted deletion analysis of Cdc6 and competitive binding assays with distamycin A.
Main Results:
- Abf1 DNA binding activity was significantly reduced (approx. 20-fold) in the cdc6-1 mutant.
- Recombinant GST-Cdc6 protein strongly stimulated Abf1 binding to ARS1 DNA in vitro.
- Cdc6's N-terminus contains the stimulating region, and Cdc6 competes with distamycin A for DNA binding, suggesting a DNA-mediated interaction.
Conclusions:
- Yeast Cdc6 protein stimulates Abf1 DNA binding activity, likely through interaction with DNA rather than direct physical contact with Abf1.
- Cdc6 may function as an architectural factor, facilitating the assembly of functional DNA replication initiation complexes.