Related Experiment Videos
Target-sequence recognition by separate-type Cys2/His2 zinc finger proteins in plants
1Laboratory of Developmental Biology, National Institute of Agrobiological Resources, 2-1-2 Kannondai, Tsukuba, Ibaraki 305, Japan.
The Journal of Biological Chemistry
|September 20, 1996
Summary
Plant EPF transcription factors bind DNA using two zinc fingers that recognize spaced AGT sequences. DNA binding specificity depends on spacing and intervening sequences, with different EPFs showing varied sensitivity.
Area of Science:
- Molecular Biology
- Plant Science
- Genetics
Background:
- EPF (Epithelial Proliferation Factor) family proteins are plant transcription factors featuring Cys2/His2-type zinc finger motifs.
- Unlike typical cluster-type zinc finger proteins binding contiguous triplets, EPFs possess longer spacers between zinc fingers (HC-link).
Purpose of the Study:
- To investigate the DNA binding mechanism of EPF transcription factors.
- To elucidate how EPFs recognize specific target DNA sequences, focusing on the role of zinc fingers and spacing.
Main Methods:
- Direct evidence generation for zinc finger-DNA interactions.
- Analysis of DNA binding affinities and specificity using varying spacer lengths and sequences.
- Site-directed mutagenesis to assess the contribution of individual zinc fingers to DNA binding.
Main Results:
- EPF zinc fingers directly contact two tandem AGT core sequences separated by approximately 13 base pairs.
- DNA binding affinity is sensitive to spacer length and significantly influenced by the intervening DNA sequence (GC-rich enhances specificity).
- EPF1 exhibits lower spacing sensitivity compared to EPF2-5, indicating differential recognition mechanisms within the family.
- Mutagenesis studies confirm synergistic contribution of both zinc fingers for high-affinity DNA binding.
Conclusions:
- EPFs employ a unique DNA recognition strategy involving both core sequence and spacing, distinguishing them from other Cys2/His2 zinc finger proteins.
- Differential sensitivity to spacing allows various EPF family members to target distinct genes.
- Synergistic action of the two zinc fingers is crucial for robust DNA binding.