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Multiple silencer elements are involved in regulating the chicken vimentin gene
1Department of Biochemistry and Molecular Biophysics, Medical College of Virginia/Virginia Commonwealth University, Richmond 23298.
Molecular and Cellular Biology
|February 1, 1994
Summary
Researchers identified new silencer elements (SE1, SE2) regulating chicken vimentin gene expression. Multiple silencer elements enhance gene down-regulation, involving a 95 kDa protein binding to purine-rich DNA sites.
Area of Science:
- Molecular Biology
- Gene Regulation
Background:
- Vimentin is an intermediate filament protein with tissue- and development-specific expression.
- A previously identified silencer element (SE3) down-regulates the chicken vimentin gene.
Purpose of the Study:
- To identify and characterize additional silencer elements involved in chicken vimentin gene regulation.
- To investigate the mechanism of negative regulation by these silencer elements.
Main Methods:
- Transfection analysis to assess silencer element activity.
- Gel mobility shift assays to detect protein binding.
- Southwestern (DNA-protein) blot analysis to determine protein molecular mass.
- DNase I footprinting to identify protein binding sites.
Main Results:
- Two new silencer elements (SE1 and SE2) were identified, in addition to SE3.
- All three silencer elements are functionally active, with optimal silencing requiring at least two elements.
- A 95 kDa protein binds to all identified silencer elements and SE3 subregions.
- DNase I footprinting revealed that the protein binds to purine-rich DNA sequences.
Conclusions:
- Multiple silencer elements contribute to the negative regulation of the chicken vimentin gene.
- A specific 95 kDa protein mediates this regulation by binding to purine-rich sites.
- These findings may have implications for understanding gene regulation in other contexts.