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Trans-dominant negative mutants of Fos and Jun
L J Ransone1, J Visvader, P Wamsley
1Molecular Biology and Virology Laboratory, Salk Institute, San Diego, CA 92138.
Abstract:
Jun and Fos nuclear oncoproteins form a complex that regulates transcription from promoters containing activator protein AP-1 binding sites. The leucine-zipper and basic-region domains of both Fos and Jun are necessary for formation of the heterodimer that binds to DNA. Reciprocal mutations in the basic region of Fos or Jun can influence the binding of the heterodimer to DNA, implying a symmetrical binding site. DNA-binding mutants of Jun exhibit increased affinity for Fos and are capable of suppressing wild-type Fos-Jun DNA-binding activity. In contrast, mutations in the basic domain of Fos, which prevent binding to DNA in association with Jun, do not significantly diminish the ability of the wild-type heterodimer to bind to DNA. These dominant negative mutants are functional in vivo and can be exploited to study the role of Fos and Jun in normal and transformed cells.
Insights
Jun and Fos oncoproteins form a transcription-regulating complex. Mutations reveal insights into their DNA binding and interactions, aiding study of their roles in cells.
Area of Science:
- Molecular Biology
- Cell Biology
- Genetics
Background:
- Jun and Fos are nuclear oncoproteins that form a complex.
- This complex regulates transcription via activator protein 1 (AP-1) binding sites.
- Specific domains, including the leucine-zipper and basic-region, are crucial for heterodimer formation and DNA binding.
Purpose of the Study:
- To investigate the roles of specific domains in Fos and Jun proteins during DNA binding.
- To explore the implications of reciprocal mutations in the basic regions of Fos and Jun.
- To characterize the function of dominant-negative mutants in vivo for cellular studies.
Main Methods:
- Site-directed mutagenesis of Jun and Fos basic regions.
- Analysis of heterodimer formation and DNA-binding affinity.
- In vivo functional assays using dominant-negative mutants.
Main Results:
- Reciprocal mutations in the basic region of Fos or Jun affect heterodimer DNA binding, suggesting a symmetrical interaction.
- Jun mutants with impaired DNA binding show increased Fos affinity and can suppress wild-type Fos-Jun activity.
- Fos mutants that abolish DNA binding do not significantly impair wild-type heterodimer binding.
Conclusions:
- The study elucidates the functional symmetry and domain requirements for Fos-Jun heterodimer DNA binding.
- Dominant-negative mutants of Fos and Jun are effective tools for studying their roles in cellular processes.
- Findings contribute to understanding gene regulation by AP-1 transcription factors in normal and transformed cells.