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Heterogeneous nuclear ribonucleoprotein K is a DNA-binding transactivator
1Laboratory of Pathology, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892.
The Journal of Biological Chemistry
|March 3, 1995
Summary
Heterogeneous nuclear ribonucleoprotein K (hnRNP K) is a DNA-binding protein that activates gene expression. It specifically binds single-stranded DNA, with at least two CT repeats required for stable complex formation.
Area of Science:
- Molecular Biology
- Gene Regulation
- Protein-DNA Interactions
Background:
- Heterogeneous nuclear ribonucleoprotein K (hnRNP K) previously shown to bind the CT element in the human c-myc gene.
- hnRNP K activates CT reporter-driven gene expression in vivo.
Purpose of the Study:
- To characterize the DNA and protein requirements for hnRNP K interaction with the CT element.
- To elucidate the binding preferences and domain requirements of hnRNP K for DNA binding and transcriptional activation.
Main Methods:
- Electrophoretic mobility shift assay (EMSA) using specific oligoribonucleotide and deoxyribonucleotide probes.
- Competition assays with RNA and DNA competitors.
- Deletion analysis of hnRNP K.
Main Results:
- hnRNP K preferentially binds single-stranded DNA over RNA or double-stranded DNA.
- Specific binding requires at least two CT repeats separated by at least three nucleotides, indicating stereochemical constraints.
- hnRNP K possesses multiple non-overlapping DNA-binding domains, each containing at least one K homology motif.
- The N-terminal 35 residues of hnRNP K are essential for transactivation of the CT element.
Conclusions:
- hnRNP K functions as a DNA-binding protein.
- hnRNP K acts as a transcriptional activator, with specific DNA-binding and N-terminal domains mediating its function.