A Cell-Based β-Lactamase Reporter Gene Assay for the CREB Signaling Pathway
Menghang Xia*, 1, Vicky Guo2, Ruili Huang1, James Inglese1, Marshall Nirenberg2, Christopher P Austin1
1 NIH Chemical Genomics Center, National Institutes of Health, Bethesda, MD 20892, USA
2 Laboratory of Biochemical Genetics, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD 20892, USA
The Cyclic-AMP Response Element Binding (CREB) proteins comprise a family of transcription factors that stimulate or repress the expression of a wide variety of genes by binding to nucleotide sequences known as cAMP Response Elements. CREB-mediated transcription has been implicated in a wide variety of important physiological processes, including long-term memory, and enhancement of CREB signaling has been suggested as an attractive therapeutic strategy for human memory disorders. To identify small molecule compounds that enhance CREB pathway signaling, we have optimized and validated a cell-based β-lactamase reporter gene CREB pathway assay in 1536-well plate format. The LOPAC library of 1280 compounds was screened in triplicate in this assay on a quantitative high throughput screening (qHTS) platform. A variety of compounds which affect known members of the CREB pathway were identified as active, including twelve known phosphodiesterase (PDE) inhibitors, and forskolin, a known activator of adenylate cyclase, thus validating the assay’s performance. This qHTS platform assay will facilitate identification of novel small molecule CREB signaling enhancers, which will be useful for chemical genetic dissection of the CREB pathway and as starting points for potentially memory-enhancing therapeutics.
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* Address correspondence to this author at the NIH Chemical Genomics Center, National Institutes of Health, 9800 Medical Center Drive, MSC 3370, Bethesda, MD 20892-3370, USA; Tel: 301-217-5718; Fax: 301-217-5736; E-mail: email@example.com