The Journal of Neuroscience, July 8, 2009, 29(27):8790-8797; doi:10.1523/JNEUROSCI.1289-09.2009
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Cellular/Molecular
The gad2 Promoter Is a Transcriptional Target of Estrogen Receptor (ER) 
and ERβ: A Unifying Hypothesis to Explain Diverse Effects of Estradiol
Edward D. Hudgens,
Lan Ji,
Clifford D. Carpenter, and
Sandra L. Petersen
Department of Biology, University of Massachusetts Amherst, Amherst, Massachusetts 01003
Correspondence should be addressed to Dr. Sandra L. Petersen, Department of Biology, 611 North Pleasant Street, University of Massachusetts Amherst, Amherst, MA 01002. Email: sandyp{at}bio.umass.edu
Estradiol (E2) regulates a wide range of neural functions, many of which require activation of estrogen receptor 
(ER) and/or ERβ, ligand-gated transcriptional regulators. Surprisingly, very few neural gene targets of ERs have been identified, and these cannot easily explain the myriad effects of E2. GABA regulates most of the same neural functions as E2, and GABAergic neurons throughout the brain contain ER. Therefore, we examined whether E2 directly regulates expression of glutamic acid decarboxylase 2 (gad2), the enzyme primarily responsible for GABA synthesis for synaptic release. Using dual luciferase assays, we found that E2, but not other gonadal steroids, stimulated the activity of a 2691 bp rat gad2 promoter reporter construct. Activation required either ER or ERβ, and ERβ did not repress ER-mediated transactivation. Site-directed mutagenesis studies identified three estrogen response elements (EREs) with cell-specific functions. An ERE at –711 upstream of the gad2 translational start site was essential for transactivation in both MCF-7 breast cancer cells and SN56.B5.G4 neural cells, but an ERE at –546 enhanced transcription only in neural cells. A third ERE at –1958 was inactive in neural cells but exerted potent transcriptional repression in E2-treated MCF-7 cells. Chromatin immunoprecipitation assays in mouse GABAergic N42 cells confirmed that E2 induced ER binding to a DNA fragment containing sequences corresponding to the –546 and –711 EREs of the rat promoter. Based on these data, we propose that direct transcriptional regulation of gad2 may explain, at least in part, the ability of E2 to impact such a diverse array of neural functions.
Received March 16, 2009;
revised May 9, 2009;
accepted May 29, 2009.
Correspondence should be addressed to Dr. Sandra L. Petersen, Department of Biology, 611 North Pleasant Street, University of Massachusetts Amherst, Amherst, MA 01002. Email: sandyp{at}bio.umass.edu
