Fig. 1.

Human AT₁ and AT₂ receptors interact with Mas receptor (MasR) in transfected HEK-293T cells. Immunocytochemistry assays were performed in HEK-293T cells expressing Mas-YFP and AT₁-Rluc receptors (1 μg of cDNA each) (A–C), or Mas-Rluc and AT₂R-YFP (1 μg of cDNA each) (D–F). Images were taken using a Zeiss 880 confocal microscope. Receptors fused to YFP were detected by yellow fluorescence (green), and receptors fused to Rluc were detected by a mouse anti-Rluc antibody and a secondary Cy3 anti-mouse antibody (red). Colocalization is shown in yellow. Cell nuclei were stained with Hoechst (blue). Scale bar: 2 μm. G, H BRET assays were performed in HEK-293T cells transfected with a constant amount of cDNA for Mas-Rluc (0.75 μg) or σ₁R-Rluc (0.75 μg) (as negative control) and increasing amounts of cDNA for AT₁R-YFP (0.5 to 2.5 μg) or AT₂R-YFP (0.5 to 3 μg). Values are the mean ± SEM of 8 independent experiments performed in duplicates. I, J Schematic representation of BRET assays: energy transfer and fluorescence emission at 530 nM only occurs if the BRET donor (Rluc) and the BRET acceptor (YFP) are close