Abstract 293: c-Myc Inactivation by PP2A is Necessary for Dopaminergic Activity in the Proximal Tubule

Abstract

We have previously shown that c-Myc regulates the expression of G protein-coupled receptor kinase 4 (GRK4). The inhibition of c-Myc can revert a dopamine-1 receptor (D 1 R) uncoupled renal proximal tubule cell (uRPTC) to normal (nRPTC). The c-Myc inhibitor, 10074-G5 (30 μM, 3 hr), restored coupling of D 1 R mediated adenylyl cyclase stimulation in uRPTCs, causing a significant increase in cAMP levels (16.21% + 0.97; p<0.01; n=47) in response to the dopamine D 1 R/D 5 R agonist, fenoldopam (FEN), than when compared to cells pre-treated with DMSO vehicle (VEH). We have also shown that FEN leads to PP2A activation. Therefore we tested the hypothesis that an inhibitory role of PP2A on c-Myc may play a role in regulating dopaminergic function. PMA (100 nM) was added to either nRPTC or uRPTC for 3hrs. In nRPTC, the total c-Myc from immunoprecipitation increased by 138.7% + 4.1%; p<0.01; n=3 with respect to VEH, while in uRPTC the total c-Myc increased by 214.5% + 11.3%; p<0.01; n=3. The PP2A/Myc association decreased by 46.0% + 1.16%; p<0.05; n=3 with respect VEH in nRPTC and decreased in uRPTC by 46.9% + 8.71%; p<0.05; n=3. These data suggest that c-Myc is activated only when PP2A binding is low. Addition of FEN (3 hrs) decreased the total c-Myc in nRPTC by 22.6% + 9.1%; p=0.06; n=3 but not in uRPTC. In nRPTC, the PP2A/Myc association increased by 56.6% + 15.1%; p <0.05; n=3 when compared to VEH, but showed no response in uRPTC. Addition of natriuretic peptide angiotensin-III (Ang III, 10 nM) decreased the expression of c-Myc by 29.4% + 16.8%; p<0.05; n=3, but showed no response in uRPTC. The PP2A/Myc binding increased by 109.3% + 30.1%; p<0.05; n=3 in nRPTC, with no increase in uRPTC. Here we show for the first time in nRPTC that both FEN and Ang III decrease total c-Myc while increasing PP2A/Myc co-immunoprecipitation, leading to inactivation of c-Myc. In uRPTC, no significant change in the total c-Myc or the association of PP2A/Myc was detected in response to FEN or Ang III. This suggests that both a lack of expressional control of c-Myc, in addition to a lack of inhibition of c-Myc by PP2A in uRPTC, may have a role in the etiology of D 1 R uncoupling and human renal proximal tubule-related hypertension. Therefore, c-Myc inhibitors may provide a new approach for treating hypertension.