Related Experiment Videos
Negative modulator of the rat D2 dopamine receptor gene
T Minowa1, M T Minowa, M M Mouradian
1Genetic Pharmacology Unit, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, Maryland 20892.
The Journal of Biological Chemistry
|April 15, 1994
Summary
Researchers identified two DNA sequences that regulate the D2 dopamine receptor gene. One sequence (D2Neg-A) acts independently, while the other (D2Neg-B) interacts with Sp1 and a novel protein to control gene transcription.
Area of Science:
- Molecular Biology
- Neuroscience
- Gene Regulation
Background:
- The rat D2 dopamine receptor gene promoter is TATA-less, featuring an initiator-like sequence and Sp1 binding sites.
- A previously identified negative modulator region (-217 to -76) influences D2 gene transcription.
Purpose of the Study:
- To delineate functional DNA sequences within the negative modulator of the rat D2 dopamine receptor gene.
- To identify the trans-acting factors interacting with these regulatory elements.
Main Methods:
- Deletion analysis of the D2 gene promoter fused to a chloramphenicol acetyltransferase reporter gene in NB41A3 cells.
- DNase I footprinting, gel mobility shift assays, and UV cross-linking experiments.
- Analysis of nuclear extracts from rat striatum and D2-expressing cells.
Main Results:
- Two cis-acting negative regulatory elements were identified: D2Neg-A (-160 to -135) and D2Neg-B (-116 to -76).
- D2Neg-A reduced transcription independently, while D2Neg-B decreased transcription by 45% through interaction with Sp1 and a novel 130-kDa nuclear factor.
- This novel factor, recognizing Sp1 binding sites, is present in both NB41A3 cells and rat striatum.
Conclusions:
- The D2 dopamine receptor gene's transcription is precisely controlled by distinct negative regulatory elements.
- Sp1 and a novel 130-kDa protein are key interacting factors with the D2Neg-B element, contributing to gene regulation in neuronal cells.