Cloning and Characterization of ATRAP, a Novel Protein That Interacts with the Angiotensin II Type 1 Receptor

Angiotensin II stimulates cellular hypertrophy in cultured vascular smooth muscle and renal proximal tubular cells. This effect is believed to be one of earliest morphological changes of heart and renal failure. However, the precise molecular mechanism involved in angiotensin II-induced hypertrophy is poorly understood. In the present study we report the isolation of a novel angiotensin II type 1 receptor-associated protein. Angiotensin II (Ang II) 1 elicits a wide range of physiological responses in a variety of cell types. This octapeptide hormone plays an important role in the regulation of cardiovascular and renal functions via diverse mechanisms (1), which include arteriolar vasoconstriction, stimulation of aldosterone production, and electrolyte homeostasis (2). Although its vasoactive effects were initially considered to be a unique feature for maintaining blood pressure, recent studies have demonstrated that Ang II exerts many other important actions on the cardiovascular and renal systems. For example, Ang II stimulates the growth of diverse cell types, such as vascular smooth muscle cells (VSMC), cardiac myocytes, and proximal tubular cells (3-6), and it increases the expression of enzymes that produce mediators of inflammation (7,8). These observations suggest that Ang II may play an important role in various cardiovascular and renal diseases associated with abnormal cell growth (cellular hypertrophy or cell proliferation) and inflammation, such as hypertension, congestive heart failure, atherosclerosis, postangioplastic restenosis, and renal failure. Clinical trials and animal studies demonstrated that angiotensin-converting enzyme inhibitors and Ang II receptor blockers prevented vascular, left ventricular, and renal proximal tubular cellular hypertrophy (9 -13), supporting the importance of Ang II in the pathogenesis of cardiovascular and renal diseases. In vitro, Ang II has also been shown to stimulate the growth of VSMC and renal proximal tubular cells (hypertrophic effect) as well as cardiac myocytes and fibroblasts (hyperplastic action) (14 -17).

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confidence: 99%

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confidence: 99%

A Novel Angiotensin II Type 1 Receptor-associated Protein Induces Cellular Hypertrophy in Rat Vascular Smooth Muscle and Renal Proximal Tubular Cells

Guo

Tardif

Ghelima

et al. 2004

“…It appears that the diversity of Ang II actions is regulated at the level of AT1 receptor, where different adaptor proteins may preferentially recruit second messenger pathways in different types of cells to execute the distinct functions of Ang II in the target cells. Therefore understanding the AT1 receptor-associated proteins is a key step to understanding the molecular basis of Ang II signaling.A novel protein, AT1 receptor-associated protein (ATRAP), has been identified in our laboratory and shown to be an intracellular partner of the AT1 receptor, interacting physically with the receptor both in vitro and in vivo (14,15). Functionally, ATRAP is capable of reducing the generation of inositol-1,4,5-triphosphate in an agonist-dependent manner and of enhancing AT1 internalization (14 -16).…”

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confidence: 99%

Identification of Calcium-modulating Cyclophilin Ligand (CAML) as Transducer of Angiotensin II-mediated Nuclear Factor of Activated T Cells (NFAT) Activation

Guo

López-Ilasaca

Dzau

2005

Self Cite

The octapeptide pressor hormone angiotensin II (Ang II), 1 a key player in the renin-angiotensin system, mediates vasoconstriction and regulates salt and fluid homeostasis, contributing to the etiology of hypertension. In addition, Ang II is involved in the pathophysiology of atherosclerosis and cardiac hypertrophy and failure (1-3). Two distinct subtypes of Ang II receptors, type 1 (AT1) and type 2 (AT2), have been characterized (4 -7). Binding of Ang II to the G-protein-coupled receptor AT1, which is thought to mediate most of the biological responses of Ang II, leads to rapid activation of heterotrimeric G proteins (G␣ q/11 , G␣ 12/13 , and G␤␥) that subsequently activate phospholipase C␤ and phospholipase C␥ to generate inositol-1,4,5-triphosphate and diacylglycerol, which in turn cause intracellular calcium mobilization and activation of protein kinase C, respectively (3,8,9). Ang II has also been shown to stimulate phospholipase A2, phospholipase D, tyrosine kinases, Janus kinase (JAK), and mitogenactivated kinases (8, 10 -13). It appears that the diversity of Ang II actions is regulated at the level of AT1 receptor, where different adaptor proteins may preferentially recruit second messenger pathways in different types of cells to execute the distinct functions of Ang II in the target cells. Therefore understanding the AT1 receptor-associated proteins is a key step to understanding the molecular basis of Ang II signaling.A novel protein, AT1 receptor-associated protein (ATRAP), has been identified in our laboratory and shown to be an intracellular partner of the AT1 receptor, interacting physically with the receptor both in vitro and in vivo (14,15). Functionally, ATRAP is capable of reducing the generation of inositol-1,4,5-triphosphate in an agonist-dependent manner and of enhancing AT1 internalization (14 -16). In this study, we analyzed the function of ATRAP in cell signaling via identification of ATRAP-interacting partners. We identified calcium-modulating cyclophilin ligand (CAML) as an ATRAP partner and present evidence that this interaction contributes to the ATRAP-mediated AT1 receptor signaling pathway. MATERIALS AND METHODSReagents and Plasmids-ATRAP full-length cDNA or deletions were cloned into vector pGBKT7 as described (14). Mouse CAML was cloned from a mouse heart cDNA library (Clontech) by standard PCR with a forward primer containing an NdeI site: 5Ј-cgg gaa tcc cat atg gag ccg gtg cct gcg gcc ac-3Ј and a reverse primer containing a BamH1 site: 5Ј-gcg cgg atc ctc agg gta ctt cag gac ccc agt aat c-3Ј. The full-length CAML cDNA was cloned into pGADT7 vector at NdeI and BamH1 cloning sites. Fulllength CAML (aa 1-296), CAML N terminus (aa 1-189), and CAML C terminus (aa 189 -296) were cloned into pGFP-N2 (BioSignal Packard, Montreal, Canada) expression vector with BglII and KpnI. PCR primers were: CAML1-189, 5Ј-aaa gag atc ttt aat acg act c-3Ј and 5Ј-gcg cgg tac caa ata ttc gaa aag atg caa ac-3Ј; CAML189 -296, 5Ј-gga aga tct acc atg ggg ttt cga ttg gtg ggg tgc-3Ј and 5Ј-gcg cgg tac cgg gta c...

“…Over the past few years, many groups have identified novel intracellular proteins that directly interact with C-terminal tails of GPCRs and function as scaffolds to regulate receptor trafficking or signaling (16,17). ATRAP is one such example of a novel protein that selectively interacts with the AT1 receptor C terminus and down-regulates its activity (18,19). Regarding the AT2 receptor, recent studies have documented a direct interaction of its C-terminal tail with ErbB3, a member of the EGF receptor family (20,21), and with the transcription factor promyelocytic zinc finger containing protein (PLZF) abundantly expressed in the heart (22).…”

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confidence: 99%

Trans-inactivation of Receptor Tyrosine Kinases by Novel Angiotensin II AT2 Receptor-interacting Protein, ATIP

Nouet

Amzallag

et al. 2004

Self Cite

171

189

Negative regulation of mitogenic pathways is a fundamental process that remains poorly characterized. The angiotensin II AT2 receptor is a rare example of a 7-transmembrane domain receptor that negatively cross-talks with receptor tyrosine kinases to inhibit cell growth. In the present study, we report the molecular cloning of a novel protein, ATIP1 (AT2-interacting protein), which interacts with the C-terminal tail of the AT2 receptor, but not with those of other receptors such as angiotensin AT1, bradykinin BK2, and adrenergic ␤ 2 receptor. ATIP1 defines a family of at least four members that possess the same domain of interaction with the AT2 receptor, contain a large coiled-coil region, and are able to dimerize. Ectopic expression of ATIP1 in eukaryotic cells leads to inhibition of insulin, basic fibroblast growth factor, and epidermal growth factor-induced ERK2 activation and DNA synthesis, and attenuates insulin receptor autophosphorylation, in the same way as the AT2 receptor. The inhibitory effect of ATIP1 requires expression, but not ligand activation, of the AT2 receptor and is further increased in the presence of Ang II, indicating that ATIP1 cooperates with AT2 to transinactivate receptor tyrosine kinases. Our findings therefore identify ATIP1 as a novel early component of growth inhibitory signaling cascade.The potent vasoactive peptide angiotensin II (Ang II) 1 is also an important regulator of cellular proliferation and hypertrophy. This peptide binds to two main subtypes of receptors (AT1 and AT2) that both belong to the superfamily of G proteincoupled receptors (GPCR) but display opposite biological and physiological effects. The AT1 receptor mediates most of the known cardiovascular and central actions of Ang II. This subtype has mitogenic and trophic effects in many tissues and cell types and transduces multiple intracellular signaling cascades typically associated with GPCR activation. In contrast, AT2 behaves like a "natural antagonist" of the AT1 subtype on most physiological functions, and induces anti-proliferative and proapoptotic effects in vitro and in vivo (for reviews, see Refs. 1-5).The AT2 receptor activates unconventional signaling pathways that in most cases do not involve coupling to classical regulatory G proteins. A growing body of evidence indicates that anti-growth effects of the AT2 receptor are associated with activation of tyrosine phosphatases and inhibition of protein kinases, which ultimately lead to inhibition of extracellular regulated kinase (ERK2). In many cell types, AT2 is functionally coupled to the Src homology 2 domain-containing tyrosine phosphatase SHP-1 (6 -8). This phosphatase has been shown to play a central role in the AT2 signaling cascades leading to inhibition of AT1-induced PYK2 and Jun kinase (9), AT1-transactivated EGF receptor tyrosine kinase (10), and insulin-induced phosphatidylinositol 3-kinase and Akt activation (11).AT2 negatively cross-talks with receptor tyrosine kinases (RTK) such as bFGF, EGF, and insulin receptors (10, 12, 13) by targeti...