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Binding site analysis of c-Myb: screening of potential binding sites by using the mutation matrix derived from
1Tsukuba Life Science Center, Institute of Physical and Chemical Research (RIKEN), Tsukuba, Ibaraki 305, Japan.
Abstract:
The c-Myb oncoprotein is known to bind to multiple sites in the promoters of target genes. We have developed a protocol to screen the binding site of c-Myb by using the systematic binding data derived form measurements of binding affinity for oligonucleotide containing a known Myb-binding site and its complete single mutants. We first applied the method to predict the binding affinity for the known binding sites and compared with available experimental data. The predicated binding sites agree with many putative binding sites of known target promoters. However, there are some binding sites not predicated by the analysis. These sequences deviate from the consensus sequence derived from the binding analyses. In the light of the structure of Myb-DNA complex, these results indicate that different DNA-binding modes may be used by c-Myb to recognize different classes of binding sites. We also screened the sequence database for potential Myb-binding sites, and found sequences of several promoters that have not been identified experimentally but could be the target for c-Myb.
Insights
Researchers developed a new protocol to screen c-Myb oncoprotein binding sites. This method predicts binding affinity, revealing potential new target genes for c-Myb and its varied DNA-binding modes.
Area of Science:
- Molecular Biology
- Genetics
- Biochemistry
Background:
- The c-Myb oncoprotein is a transcription factor known to interact with numerous target gene promoters.
- Understanding c-Myb's specific DNA binding is crucial for deciphering gene regulation.
Purpose of the Study:
- To develop and validate a protocol for systematically screening and predicting c-Myb oncoprotein binding sites.
- To investigate the DNA-binding modes of c-Myb in relation to its binding site sequences.
Main Methods:
- Developed a protocol utilizing systematic binding data from affinity measurements of oligonucleotides with known Myb-binding sites and single mutants.
- Applied the protocol to predict binding affinities for known sites and compared predictions with experimental data.
- Screened sequence databases for potential c-Myb binding sites.
Main Results:
- The developed protocol successfully predicted binding affinities for many known c-Myb binding sites.
- Identified discrepancies where predicted binding sites deviated from the consensus sequence, suggesting varied DNA-binding modes.
- Discovered potential novel c-Myb binding sites in promoter sequences not previously identified experimentally.
Conclusions:
- The findings suggest that c-Myb may employ different DNA-binding modes to recognize diverse classes of binding sites.
- The protocol offers a valuable tool for identifying novel c-Myb target genes and understanding its regulatory mechanisms.