Fig. 2

RAMP1 overexpression increases Gαi3, PKA, CREB and YAP expression. RAMP1 OE and VECs were treated with DOX (5 μg/ml) for 48 h when the following experiments were conducted. A Western blot analysis of Gαi3, PKA, CREB, pCREB (S133), YAP and pYAP (S127). Total protein was stained as an endogenous control. B Quantification of the protein band intensities in A. The data were normalized to LNF. C qPCR analysis of the YAP mRNA levels. The data were normalized to the amount of β-actin mRNA. D Western blot analysis of cytoplasmic and nuclear fractions of CREB and YAP. Total protein was stained as an endogenous control. E Quantification of the protein band intensities in D. The data were normalized to LNF. C-CREB means cytoplasmic CREB, N-CREB means nuclear CREB, C-YAP means cytoplasmic YAP, and N-YAP means nuclear YAP. F Confocal immunofluorescence of CREB (red), Actin (green) and DAPI (blue). Scale bar = 20 μm. G Confocal immunofluorescence of YAP (red), Actin (green) and DAPI (blue). Scale bar = 20 μm. The experiments were performed in triplicate. The data are presented as means ± SD, and significant differences were evaluated using unpaired t test. *P < 0.05, **P < 0.01 and ***P < 0.005