Related Experiment Videos
Characterization of the DNA-binding and dominant negative activity of v-erbA homodimers
1Division of Endocrinology and Metabolism, University of Mississippi Medical Center, Jackson 39216, USA.
Abstract:
The oncoprotein v-erbA is a mutated form of thyroid hormone receptor alpha1 that is virtually incapable of binding T3. V-erbA is a dominant repressor of transcription induced by thyroid hormone receptors and retinoic acid receptors; however, the genetic targets of v-erbA that lead to oncogenesis are not known. Although v-erbA can bind as monomers and dimers to DNA containing the consensus sequence AGGTCA arranged as direct, inverted, or everted repeats, it is not known which sequence represents the optimal v-erbA-binding site. Determination of the DNA recognition properties of v-erbA would allow a better understanding of the repressor activity of this oncoprotein. The current studies, by using a random DNA selection strategy, have determined that the imperfect everted repeat 5'-TGACC(T/C)NT(A/G)AGGTCAC is the optimal v-erbA homodimer-binding site, where N represents any di- or trinucleotide. Functional studies show that everted repeats containing this sequence are substantially more potent v-erbA response elements than direct or inverted repeats, even though many classic T3 response elements are direct repeats. Thus, v-erbA represses only a subset of T3 response elements. In a similar fashion, v-erbA was found to repress a subset of vitamin D response elements. Of general interest, the data indicate that the two molecules of a transcription factor homodimer do not necessarily have identical DNA-binding specificities.
Insights
The oncoprotein v-erbA, a mutated thyroid hormone receptor, binds DNA optimally as a homodimer to imperfect everted repeats. This finding clarifies v-erbA
Area of Science:
- Molecular Biology
- Oncology
- Endocrinology
Background:
- The oncoprotein v-erbA is a mutated thyroid hormone receptor alpha1.
- v-erbA acts as a dominant repressor of transcription but its genetic targets are unknown.
- Understanding v-erbA's DNA binding is crucial for deciphering its oncogenic mechanisms.
Purpose of the Study:
- To determine the optimal DNA-binding site for the v-erbA oncoprotein.
- To elucidate the DNA recognition properties of v-erbA homodimers.
- To understand the specificity of v-erbA's transcriptional repression.
Main Methods:
- Random DNA selection strategy to identify binding sites.
- Analysis of DNA sequences bound by v-erbA monomers and dimers.
- Functional assays to compare the potency of different v-erbA response elements.
Main Results:
- The optimal v-erbA homodimer-binding site is an imperfect everted repeat: 5'-TGACC(T/C)NT(A/G)AGGTCAC.
- Everted repeats with this sequence are more potent v-erbA response elements than direct or inverted repeats.
- v-erbA represses a specific subset of thyroid hormone and vitamin D response elements.
Conclusions:
- v-erbA exhibits distinct DNA-binding preferences compared to wild-type receptors.
- The identified binding site explains the selective repression of specific gene subsets by v-erbA.
- Transcription factor homodimers may not possess identical DNA-binding specificities for each monomer unit.