Cloning and characterization of ATRAP, a novel protein that interacts with the angiotensin II type 1 receptor

The carboxyl-terminal cytoplasmic domain of the angiotensin II type 1 (AT(1)) receptor has recently been shown to interact with several classes of cytoplasmic proteins that regulate different aspects of AT(1) receptor physiology. Employing yeast two-hybrid screening of a mouse kidney cDNA library with the carboxyl-terminal cytoplasmic domain of the murine AT(1a) receptor as a bait, we have isolated a novel protein with a predicted molecular mass of 18 kDa, which we have named ATRAP (for AT(1) receptor-associated protein). ATRAP interacts specifically with the carboxyl-terminal domain of the AT(1a) receptor but not with those of ansotensin II type 2 (AT(2)), m(3) muscarinic acetylcholine, bradykinin B-2, endothelin B, and beta(2)-adrenergic receptors, The mRNA of ATRAP was abundantly expressed in kidney, heart, and testis but was poorly expressed in lung, liver, spleen, and brain. The ATRAP-AT(1a) receptor association was confirmed by affinity chromatography, by specific co-immunoprecipitation of the two proteins, and by fluorescence microscopy, showing co-localization of these proteins in intact cells. Overexpression of ATRAP in COS-7 cells caused a marked inhibition of AT(1a) receptor-mediated activation of phospholipase C without affecting m(3) receptor-mediated activation. In conclusion, we have isolated a novel protein that interacts specifically with the carboxyl-terminal cytoplasmic domain of the AT(1a) receptor and affects AT(1a) receptor signaling.