ANGIOTENSIN-(1-7) INDUCES MAS RECEPTOR INTERNALIZATION

M. GIRONACCI; H. ADAMO; G. CORRADI; R. SANTOS; P. ORTIZ; O. CARRETERO

Milan

Congreso; 21sth European Meeting on Hypertension and cardiovascular Prevention; 2011

Objective: Angiotensin (Ang) (1-7) is the endogenous ligand for the G protein-coupled receptorMas, a receptor (R) coupled to cardiac, renal and cerebral protective responses. Physiologicalevidence suggests that Mas R undergoes agonist-dependent desensitization, but underlyingmolecular mechanism regulating R activity is unknown. We investigated the hypothesis that Mas Rdesensitizes and internalizes upon stimulation with Ang-(1-7).Design and Method: We generated a chimera between the Mas R and the fluorescent protein YFP(Mas-YFP R). Mas-YFP transfected HEK 293T cells were incubated with Ang-(1-7) and the relativecellular distribution of Mas-YFP R was observed by confocal microscopy.Results: In resting cells, Mas-YFP R was mostly localized on the cell membrane. Ang-(1-7) induceda redistribution in fluorescence after 5 min stimulation changing the localization of Mas-YFP R tointracellular vesicles of various sizes. Following the time course of [125I]Ang-(1-7) endocytosiswe observed that up to 65.1+8.7 % of Mas-YFP R underwent endocytosis after 20 min from theplasma membrane which was blocked by the Mas receptor antagonist. Mas-YFP R colocalizes withRab5, the early endosome antigen 1 and AP-50, indicating that Mas-YFP R is internalized throughclathrin-coated pits and targeted to early endosomes after Ang-(1-7) stimulation. Mas-YFP R alsocolocalized with caveolin-1 suggesting that at some point of R trafficking Mas-YFP R traversescaveolin-1 positive compartments.Conclusion: Mas R undergoes endocytosis upon stimulation with Ang-(1-7) and this event mayexplain the desensitization of Mas R responsiveness. In this way, Mas R activity and density may betightly controlled by the cell.