Related Experiment Videos
Protein complexes bearing myc-like antigenicity recognize two distinct DNA sequences
Y Negishi1, S M Iguchi-Ariga, H Ariga
1Faculty of Pharmaceutical Sciences, Hokkaido University, Sapporo, Japan.
Abstract:
The products of the myc proto-oncogene family have been suggested to carry functions important for cell proliferation, differentiation and neoplasia, but the molecular mechanisms of such biological effects have not yet been clarified. We have previously reported that the c-myc protein or protein(s) complexed with the c-myc protein bind to a specific sequence in the region upstream of the c-myc gene, where there exist an origin of cellular DNA replication (ori) and also a transcriptional enhancer. It was recently reported that the c-myc protein forms complexes with a novel helix-loop-helix leucine zipper protein (Max), and that the Myc-Max complex specifically recognizes a DNA sequence different from the sequence we have defined in the c-myc gene. In this report we examine the nuclear extract prepared from human Raji cells for binding to the two different sequences which have been reported as specific binding sequences for c-myc protein or c-myc protein complexes. The binding to one sequence was inhibited in the presence of excess amount of the other sequence, suggesting that the same protein(s) may be necessary for binding to either of the two sequences. Since binding to both sequences was cancelled by pretreatment of the Raji extract with an anti-c-myc protein antibody, it is suggested that proteins carrying myc-like antigenicity are necessary for efficient binding of the nucleoprotein complexes to the two distinct DNA sequences.
Insights
The c-myc protein binds to DNA sequences crucial for cell growth and cancer. This study suggests myc-like proteins are essential for binding to distinct DNA sites, clarifying oncogene mechanisms.
Area of Science:
- Molecular Biology
- Oncogenesis
- Cellular Regulation
Background:
- The myc proto-oncogene family plays roles in cell proliferation, differentiation, and neoplasia.
- The precise molecular mechanisms underlying these effects remain unclear.
- Previous research identified c-myc binding to a specific upstream sequence of its own gene, containing a DNA replication origin and enhancer.
Purpose of the Study:
- To investigate the binding of c-myc protein or its complexes to two distinct DNA sequences.
- To clarify the molecular mechanisms of c-myc's biological functions.
- To determine if the same proteins are involved in binding to both identified DNA sequences.
Main Methods:
- Nuclear extracts from human Raji cells were used.
- Electrophoretic mobility shift assays (EMSA) were employed to study DNA-protein interactions.
- Competition assays with excess unlabeled DNA sequences were performed.
- Antibody inhibition assays using an anti-c-myc antibody were conducted.
Main Results:
- Binding to one DNA sequence was inhibited by the presence of the other, indicating shared protein requirements.
- Pretreatment of nuclear extracts with an anti-c-myc antibody abolished binding to both sequences.
- These findings suggest that proteins with myc-like antigenicity are necessary for binding to both distinct DNA sequences.
Conclusions:
- The study provides evidence that myc-like proteins are essential for the binding of nucleoprotein complexes to specific DNA sequences.
- This suggests a conserved mechanism for c-myc protein interaction with DNA, potentially involving shared protein partners.
- Clarifying these binding mechanisms is crucial for understanding the role of myc in cell proliferation, differentiation, and cancer development.