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. Author manuscript; available in PMC: 2007 Mar 21.
Published in final edited form as: J Biol Chem. 2006 Aug 31;281(44):33467–33476. doi: 10.1074/jbc.M603436200

FIGURE 5. PDGF stimulates Ser-133 phosphorylation of CREB and binding to the NOR1 promoter through the ERK-MAPK signaling pathway.

FIGURE 5

A, serum-deprived SMC were stimulated with PDGF (25 ng/ml) and analyzed for Ser-133-phosphorylated CREB by Western blotting. Cohybridization for total CREB was employed as control. B, serum-deprived SMC were preincubated with vehicle or PD98059 (10 μmol/liter) for 30 min and stimulated with PDGF (25 ng/ml) for 15 min. Whole cell lysates were analyzed for Ser-133-phosphorylated CREB and phosphorylated p44/42 MAPK. Cohybridization for total CREB and total p44/42 MAPK was employed as control. C, quiescent SMC were stimulated with PDGF (25 ng/ml) for the indicated time points. Following chromatin immunoprecipitation (IP) using a Ser-133-phospho-CREB antibody (4 μg) or control IgG, PCR analysis was performed using primer pairs that cover the CRE sites between −79 and −46 of the NOR1 promoter. Total extract (Input) was used as positive PCR control. D, serum-deprived SMC were preincubated with vehicle or PD98059 (10 μmol/liter) for 30 min prior to stimulation with PDGF (25 ng/ml) for 30 min. Chromatin immunoprecipitation of Ser-133-phosphorylated CREB to the endogenous NOR1 promoter was performed as described in C. All autoradiograms shown are representative of three independently performed experiments. DMSO indicates Me2SO.