Figure 6
From: Exchange factors directly activated by cAMP mediate melanocortin 4 receptor-induced gene expression
Significant role for EPACs in MC4R-promoted c-fos and TRH mRNA induction.
qRT-PCR experiments were performed with cDNAs derived from serum-starved mHypoA-2/10-CRE cells pretreated with KT-5720 (5 μM), ESI-09 (20 μM) or the carrier DMSO (0.1 or 0.2 respectively) for 30 min and then stimulated with 1 μM of α-MSH for 20 min. In (A) specific primer pairs against TRH and in (B) against c-fos were used. 5 independent cDNAs for each condition were analysed twice in triplicates, data normalized to β-actin and to control cells and expressed as the mean ± S.E.M. Crossing points (Cp) were determined by the software supplied with the LightCycler® 480 and data analysed by the ΔΔCP method (0.5(target gene–actin)MSH-(target gene–actin)basal). Asterisks indicate a significant difference between ESI-09 or KT-5720 and DMSO-treated cells using the two-sample Student’s t-test. Hashed signs indicate a significant difference to 1.0 using the one-sample Student’s t-test.
