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An SNP in the Human GRK3 Gene Promoter‐Luciferase Construct Eliminates Hormone‐Induced Activation of GRK3 Transcription
Author(s) -
Eikenburg Douglas,
Salim Samina,
Taneja Manish
Publication year - 2009
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.23.1_supplement.944.2
Subject(s) - luciferase , promoter , transcription factor , reporter gene , single nucleotide polymorphism , snp , microbiology and biotechnology , biology , chemistry , transfection , gene expression , gene , genotype , genetics
Epinephrine (EPI) and corticotrophin releasing factor (CRF) selectively increase GRK3 expression by an ERK1/2‐dependent activation of the transcription factors Sp‐1 and Ap‐2 in two neuronal cell lines, BE(2)‐C and CATH.a cells. Moreover, a single nucleotide polymorphism (SNP; ‐382G/A) in the promoter region of the human GRK3 gene reportedly decreases GRK3 expression, and increases the risk for bipolar disorder (BPD). In this study we generated plasmids to express a WT human GRK3 promoter (1.3kb, ‐1597 to ‐297)‐luciferase reporter construct containing the predicted Sp‐1 and Ap‐2 binding sites and a second construct containing the SNP. BE(2)‐C cells and CATH.a cells were transfected to express each construct and the effects of EPI and CRF on luciferase reporter activity, respectively, were studied. EPI/CRF treatment (1‐8h) increased WT GRK3 promoter driven luciferase activity; this was prevented by the MEK1/2 inhibitor, U1026. EPI/CRF treatment (1‐8h) failed to stimulate SNP‐GRK3 promoter driven luciferase activity. These results demonstrate that our WT GRK3 promoter construct accurately models regulation of endogenous GRK3 gene expression and that the SNP in the GRK3 promoter eliminates the ability of the stress hormones EPI/CRF to stimulate ERK‐dependent GRK3 expression. This mechanism may contribute to the reduced GRK3 levels observed in cells from BPD patients with the SNP. (Supported by NARSAD)