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High Sensitivity Measurement of Transcription Factor-DNA Binding Affinities by Competitive Titration Using Fluorescence Microscopy
Published on: February 7, 2019
DNA binding properties of TAF1 isoforms with two AT-hooks
Chad E Metcalf1, David A Wassarman
1Department of Biomolecular Chemistry, University of Wisconsin School of Medicine and Public Health, Madison, Wisconsin 53706, USA.
The Journal of Biological Chemistry
|August 9, 2006
Summary
TATA-binding protein-associated factor 1 (TAF1) isoforms directly bind DNA using AT-hook motifs, anchoring to gene promoters. Alternative splicing of TAF1 influences gene transcription in Drosophila.
Area of Science:
- Molecular Biology
- Genetics
- Biochemistry
Background:
- TFIID is crucial for RNA polymerase II transcription.
- TAF1 is a TFIID component, but its promoter localization mechanism is unclear.
- Known TFIID DNA-binding components are limited.
Purpose of the Study:
- Investigate direct DNA binding by TAF1 isoforms.
- Determine the role of AT-hook motifs in TAF1 DNA interaction.
- Elucidate TAF1's contribution to gene promoter targeting.
Main Methods:
- Electrophoretic mobility shift assays (EMSA).
- Alanine-scanning mutagenesis of AT-hook motifs.
- Analysis of TAF1 isoform binding to Drosophila gene promoters.
Main Results:
- Drosophila TAF1 isoforms (TAF1-2, TAF1-4) bind DNA independently of TAF2.
- DNA binding is mediated by cooperative AT-hook motifs.
- TAF1-2 binds AT-rich DNA minor groove, preferring AAT sequences.
- Alternative splicing of AT-hook impacts DNA binding affinity.
Conclusions:
- AT-hooks anchor TAF1 isoforms to AT-rich gene promoters.
- TAF1 isoforms contribute to promoter specificity.
- Alternative splicing of TAF1 isoforms may regulate gene transcription.
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