Related Experiment Videos
DNA binding specificity of the CCAAT-binding factor CBF/NF-Y
1Department of Molecular Genetics, The University of Texas M. D. Anderson Cancer Center, Houston, Texas 77030, USA.
The Journal of Biological Chemistry
|October 23, 1997
Summary
CCAAT-binding factor (CBF) interacts with DNA, specifically the CCAAT motif and flanking sequences. This interaction is crucial for regulating transcription in higher eukaryotic genes.
Area of Science:
- Molecular Biology
- Genetics
- Biochemistry
Background:
- CCAAT-binding factor (CBF) is a heterotrimeric protein complex.
- CBF is known to bind DNA sequences containing CCAAT motifs.
Purpose of the Study:
- To analyze the interactions of recombinant CBF with DNA.
- To determine the specific DNA sequences required for CBF binding.
- To investigate the role of flanking sequences in CBF-DNA affinity.
Main Methods:
- Hydroxyl radical footprinting assays.
- Methylation interference assays.
- Polymerase chain reaction (PCR)-mediated random binding site selection.
Main Results:
- CBF binding protects three DNA regions, with sequence-specific contacts localized to the CCAAT motif.
- Methylation interference confirmed sequence-specific binding within the CCAAT motif.
- CBF binding affinity is influenced by nucleotide sequences flanking the CCAAT motif.
- Selected CBF binding sites closely resemble CCAAT motifs in higher eukaryotic promoters.
Conclusions:
- CBF binds DNA in a sequence-specific manner, primarily through the CCAAT motif and its flanking regions.
- The identified binding sites suggest CBF's role in regulating transcription of numerous eukaryotic genes.