Abstract
Relatively small changes in G-protein-coupled receptor kinase (GRK) 3 expression (∼2-fold) profoundly affect α2-adrenergic receptor (AR) function and preferentially regulate neuronal α2A- and α2B-AR signaling. In the present study, we provide evidence that epinephrine (EPI)-induced up-regulation of GRK3 protein expression in two neuronal cell lines, BE(2)-C cells (endogenously express α2A- and β2-AR) and BN17 cells [endogenously express α2B (NG108) and transfected to express β2-AR] is due in part to increased GRK3 gene expression. In both cell lines, the increase in GRK3 transcription occurred via an extracellular signal-regulated kinase (ERK) 1/2-dependent mechanism because the increase in GRK3 mRNA is eliminated in the presence of the mitogen-activated protein kinase/ERK kinase 1/2 inhibitor, U0126 [1,4-diamino-2,3-dicyano-1,4-bis (2-amino phenylthiobutadiene)]. EPI-induced GRK3 mRNA up-regulation also is prevented in the presence of propranolol or phentolamine. Moreover, GRK3 mRNA did not increase in response to EPI treatment in NG108 cells (endogenously express α2B-AR with no β2-AR). Both these results suggest that simultaneous activation of α2- and β2-AR by EPI is required for the ERK1/2-dependent increase in GRK3 mRNA. The EPI-induced increase in GRK3 mRNA was unaffected in the presence of the protein kinase C inhibitor, chelerythrine chloride. Finally, EPI treatment resulted in increased nuclear translocation and accumulation of the transcription factors, Sp-1 and Ap-2, in BE(2)-C cells. Taken together, our results demonstrate the involvement of the ERK1/2 pathway in selective up-regulation of GRK3 mRNA expression, possibly via activation of Sp-1 and Ap-2 transcription factors in neuronal cells.
Footnotes
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This work was supported by the American Heart Association, Texas Affiliate, Inc. (Grant 0555032Y to D.C.E.) and by the National Institutes of Health (Grant DA01738 to K.M.S.).
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Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
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doi:10.1124/jpet.106.116921.
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ABBREVIATIONS: GRK, G-protein-coupled receptor kinase; GPCR, G-protein-coupled receptor; AR, adrenoceptor; CRF, corticotropin-releasing factor; NE, norepinephrine; ERK, extracellular signal-regulated kinase; EPI, epinephrine; DMEM, Dulbecco's modified Eagle's medium; MEK, mitogen-activated protein kinase/ERK kinase; U0126, 1,4-diamino-2,3-dicyano-1,4-bis (2-amino phenylthiobutadiene); RT, reverse transcriptase; PCR, polymerase chain reaction; PAGE, polyacrylamide gel electrophoresis; NG108-15 (NG108), neuroblastoma/glioma hybrid cell; BE(2)-C, human neuroblastoma cell(s); DAPI, 4′,6-diamidino-2-phenylindole; CC, chelerythrine chloride; PBS, phosphate-buffered saline; PBSS, PBS containing 1.2% sucrose; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; PKC, protein kinase C; BPD, bipolar disorder.
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↵1 Current affiliation: Department of Pharmaceutical Sciences, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma.
- Received November 10, 2006.
- Accepted January 23, 2007.
- The American Society for Pharmacology and Experimental Therapeutics
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