The Importance of Tyrosine Phosphorylation in Angiotensin II Signaling

doi:10.1016/S1050-1738(96)00053-9

Trends in Cardiovascular Medicine

Volume 6, Issue 6, August 1996, Pages 179-187

https://doi.org/10.1016/S1050-1738(96)00053-9 Get rights and content

Abstract

Angiotensin II plays a critical role in the regulation of vascular resistance and intravascular volume. Virtually all of the physiologic effects of angiotensin II are mediated by the AT₁ receptor, a seven-transmembrane spanning receptor. Although G proteins play an important role in the signaling of this class of receptors, it has become increasingly clear that tyrosine phosphorylation is also intimately involved in AT₁ receptor signaling. In response to angiotensin II, both smooth muscle and glomerular mesangial cells tyrosine phosphorylate the γ isoform of phospholipase C. This is critical to downstream signaling events, including the intracellular generation of 1,4,5-inositol triphosphate. The soluble cytoplasmic kinase Src appears to be activated by angiotensin II and to play an important role in the phosphorylation of phospholipase C. Angiotensin II, acting through the AT₁ receptor, causes Jak kinase phosphorylation and activation. This, in turn, leads to STAT phosphorylation and translocation to the nucleus. Finally, we present data that indicate that angiotensin II activates Ras and leads to Ras—Raf-1 complex formation. Activation of this pathway also appears to require active Src. These studies provide compelling evidence that tyrosine phosphorylation plays an important role in the signaling of angiotensin II. The exact biochemical mechanism by which a seven-transmembrane receptor stimulates intracellular kinases to be elucidated. (Trends Cardiovasc Med 1996;6:179-187).

Section snippets

• Protein Phosphorylation and Dephosphorylation

The existence of regulatory kinases was suggested by Fischer and Krebs (1955)and Rall et al. (1957)when they observed both an active phosphorylated and an inactive dephosphorylated form of the enzyme glycogen phosphorylase. Ten years later, the first protein kinase isolated was the cAMP-dependent protein kinase, PKA (Walsh et al. 1968). An enormous number of other protein kinases and phosphoprotein phosphatases have now been characterized; it is predicted that there are over a thousand such

• Angiotensin II Stimulated Activation of Phospholipase C

Our group has investigated the role of angiotension II-induced tyrosine phosphorylation in three separate signaling pathways. One pathway is the activation of PLC, leading to the intracellular generation of 1,4,5-IP₃ and diacylglycerol. This signaling pathway is activated almost immediately upon ligand binding and serves as the stimulus for many downstream signaling events, including the very rapid release of intracellular calcium stores (Berridge 1987). To understand this basic signaling

• Angiotensin II and the Jak-STAT Pathway

In 1992, Dr. Arnold Katz published an article entitled “Is Angiotensin II a Growth Factor Masquerading as a Vasopressor?” (Katz et al. 1992). This article underlined a host of observations owing that angiotensin II acts as a smooth muscle growth factor both in vivo and in vitro. For instance, in RASM cells, Chiu et al. (1991)showed that adding angiotensin II (10⁻₇M) to quiescent cells increased protein synthesis 45% and DNA synthesis 56% after 24 h (Chiu et al. 1991). Both responses were

• Angiotensin II and the Ras Pathway

The best understood signaling pathway associated with cell growth is activated when ligands such as PDGF bind to their cell surface receptors. These receptors contain intracellular kinase domains, which become active and lead to a cascade of protein phosphorylation events (Pimental 1994, Van Der Geer et al. 1994). Central to this cascade is the membrane-associated protein Ras. Ras is similar to heterotrimeric G proteins in that it becomes activated when bound GDP is exchanged with GTP (Bollag

• Conclusion

Until very recently, the importance of protein phosphorylation in signaling cascades initiated by seven-transmembrane receptors had not been fully appreciated. Our data strongly suggest that within minutes of ligand binding to the AT₁ receptor, protein tyrosine phosphorylation plays a major role in PLC-γ1, Jak-STAT, and Ras signaling pathway activation. At present, the exact biochemical role of heterotrimeric G-protein components in this activation is not fully understood. The exact

Acknowledgements

This work was supported by National Institute of Health grants DK39777, DK44280, and DK45215, as well as Grants-In-Aid from the American Heart Association and the Georgia affiliate of the American Heart Association. M.B.M. is an American Heart Association Minority Developmental Award Scientist.

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The mechanisms leading to development of restenosis are complex, involving multiple processes that include vascular remodeling and neointimal hyperplasia.4,5 Neointimal hyperplasia after percutaneous coronary interventions occurs in response to growth factors that promote the migration and proliferation of vascular smooth muscle cells at the site of injury.6–13 While stent placement reduces vessel recoil and lessens negative vascular remodeling, neointimal hyperplasia with in-stent restenosis frequently results.57
Arterial restenosis after angioplasty/stenting has hindered coronary artery disease treatment, especially in diabetics. We theorized that gastrin-releasing peptide (GRP) antagonists and growth hormone-releasing hormone (GHRH) antagonists might decrease neointimal hyperplasia and restenosis in diabetic rats after common carotid arterial balloon injury.
Two separate experiments were conducted to test the effects of a GRP antagonist (RC-3095) and a GHRH antagonist (MZ-4-71) on vascular smooth muscle (VSM) growth. In a preliminary in vitro experiment non-injured human aortic vascular smooth muscle (VSM) proliferation was compared between growth media and control. In a second in vivo experiment, intimal and medial area, intima/media ratio (IM) and percent stenosis were compared between injured carotid arteries in twelve Zucker type II obese rats treated with subcutaneously injected RC-3095, MZ-4-71, or control media.
In the in vitro experiment, decreased VSM cell growth was observed in GRP antagonist (p < 0.05) and GHRH antagonist groups (p < 0.05) compared to the control group. In the in vivo experiment, the GRP antagonist group had a decreased IM ratio (1.63 ± 0.41, p < 0.05) and an increased area of stenosis (98.78% ± 1.48 p = NS) compared to control (2.38 ± 1.09) while the GHRH antagonist group had decreased IM ratio (1.33 ± 0.58 SD, p < 0.05) and percent area of stenosis (78.84% ± 24.97, p < 0.05) compared to control (2.38 ± 1.09).
The significant decrease in both IM ratio and percent area of stenosis in the GHRH antagonist group supports the hypothesis that this peptide may reduce neointimal hyperplasia and restenosis.
Angiotensin II stimulates a novel angiotensin II type 1 receptor-associated protein, GLP gene expression in rat kidney proximal tubular cells
2006, Journal of Cardiothoracic-Renal Research
We have recently demonstrated that a novel angiotensin II (Ang II) type 1 receptor-associated protein, GLP induces cellular hypertrophy and Ang II stimulated GLP mRNA expression in a time-, dose-, and AT₁ receptor-dependent manner in rat immortalized renal proximal tubular cells (IRPTCs). The present studies investigated which signaling pathways are involved in Ang II stimulated GLP mRNA expression in IRPTCs. Cellular GLP mRNA expression was determined by reverse transcription polymerase chain reaction. The effect of Ang II was blocked by epidermal growth factor receptor inhibitor, AG1478, PKC inhibitor, GF109203X, MAPK kinase inhibitor, PD98059, antioxidants, taurine and tiron, and NADH/NADPH oxidase inhibitor, DPI, whereas, no effects were observed on Ang II-induced GLP mRNA expression with p38 MAPK inhibitors, SB203580 and PD169316, and Janus kinase 2 inhibitor, AG490. IRPTCs stably expressing GLP gene significantly increased reactive oxygen species (ROS) generation compared to control IRPTCs analyzed by lucigenin assay. Furthermore, NADH/NADPH oxidase inhibitor, DPI, and antioxidants taurine and tiron inhibited GLP-induced total protein content and de novo protein synthesis. These studies demonstrated that the stimulatory action of Ang II on GLP mRNA expression in IRPTCs is mediated by multiple signal mechanisms, transactivation of EGF receptor and subsequent MAPK activation, PKC activation and ROS generation. Furthermore, the cellular hypertrophic effect of GLP gene in IRPTCs is mediated at least in part via ROS generation.
Angiotensin II signaling and HB-EGF shedding via metalloproteinase in glomerular mesangial cells
2001, Kidney International
Angiotensin II signaling and HB-EGF shedding via metalloproteinase in glomerular mesangial cells.
Angiotensin II (Ang II) has been implicated in the development of glomerulosclerosis by stimulating fibronectin (FN) synthesis. The processing and release of heparin binding–endothelin growth factor (HB-EGF) are activated by protein kinase C (PKC) and Ca²⁺ signaling. We studied the roles of HB-EGF and endothelial growth factor (EGF) receptor (EGFR) in Ang II-induced FN expression using mesangial cells.
Mesangial cells were prepared from mouse kidneys by the explant method and cells were used at passages 4 and 5.
Ang II stimulated FN mRNA levels dose-dependently with a maximal increase (3.4-fold) after 12 hours of incubation. This action was completely inhibited by PKC inhibitors and slightly blocked by Ca²⁺ chelating agents. FN mRNA accumulation by Ang II was abolished by tyrosine kinase inhibitors, a specific inhibitor for EGFR (AG1478) and extracellular signal-regulated kinase (ERK) inactivation. Addition of neutralizing anti–HB-EGF antibody, as well as pretreatment with heparin or the metalloproteinase inhibitor batimastat abolished induction of FN expression by Ang II. In mesangial cells stably transfected with a chimeric construct containing HB-EGF and alkaline phosphatase (ALP) genes, ALP activity in incubation medium was rapidly increased by Ang II (1.7-fold at 0.5 min) and reached a 4.1-fold increase at two minutes. Ang II phosphorylated EGFR (maximal at 2 min) and ERK (maximal at 8 min) in a PKC- and metalloproteinase-dependent manner. Ang II stimulated the expression and release of transforming growth factor-β (TGF-β) via EGFR-mediated signaling, and the released TGF-β also contributed to Ang II-mediated FN expression via EGFR transactivation.
Ang II-mediated FN expression was regulated by autocrine effects of HB-EGF and TGF-β, suggesting a novel paradigm for cross-talk between Ang II and growth factor receptor signaling pathways.
Angiotensin II receptors in endothelial cells
1997, General Pharmacology
- 1.
  1. Angiotensin II (Ang II), the main effector of the renin-angiotensin system, exerts its vasoconstrictory and trophic actions on smooth muscle cells via AT₁ receptors. However, Ang II does not act only on smooth muscle cells, as Ang II receptors are also present in endothelial cells.
- 2.
  2. The receptor type on these cells differs depending on the origin of the endothelium and the species. The rat endothelial receptors are mostly of the AT₁ type, but AT₂ receptors have also been found. The pharmacological characteristics of the AT₁ receptors on endothelial cells are similar to those of other cell types.
- 3.
  3. Ang II stimulates phospholipase C and phospholipase A₂ activation via the AT₁ receptor in endothelial cells. Ang II also stimulates the tyrosine phosphorylation of several proteins in these cells.
- 4.
  4. Some studies suggest that the AT₁ receptor mediates the release of vasodilator molecules by endothelial cells and could modulate Ang II effect on smooth muscle cells. Ang II may also inhibit endothelial cell growth via the AT₂ receptor. Finally, endothelial Ang II receptors may be implicated in the regulation of fibrinolysis.
Role of Janus kinase/signal transducer and activator of transcription and mitogen-activated protein kinase cascades in angiotensin II- and platelet-derived growth factor-induced vascular smooth muscle cell proliferation
1997, Journal of Biological Chemistry
In vascular smooth muscle cells, the induction of early growth response genes involves the Janus kinase (JAK)/signal transducer and activators of transcription (STAT) and the Ras/Raf-1/mitogen-activated protein kinase cascades. In the present study, we found that electroporation of antibodies against MEK1 or ERK1 abolished vascular smooth muscle cell proliferation in response to either platelet-derived growth factor or angiotensin II. However, anti-STAT1 or -STAT3 antibody electroporation abolished proliferative responses only to angiotensin II and not to platelet-derived growth factor. AG-490, a specific inhibitor of the JAK2 tyrosine kinase, prevented proliferation of vascular smooth muscle cells, complex formation between JAK2 and Raf-1, the tyrosine phosphorylation of Raf-1, and the activation of ERK1 in response to either angiotensin II or platelet-derived growth factor. However, AG-490 had no effect on angiotensin II- or platelet-derived growth factor-induced Ras/Raf-1 complex formation. Our results indicate that: 1) STAT proteins play an essential role in angiotensin II-induced vascular smooth muscle cell proliferation, 2) JAK2 plays an essential role in the tyrosine phosphorylation of Raf-1, and 3) convergent mitogenic signaling cascades involving the cytosolic kinases JAK2, MEK1, and ERK1 mediate vascular smooth muscle cell proliferation in response to both growth factor and G protein-coupled receptors.
Dependence on the motif YIPP for the physical association of Jak2 kinase with the intracellular carboxyl tail of the angiotensin II AT<inf>1</inf> receptor
1997, Journal of Biological Chemistry
Angiotensin II is the effector molecule of the renin-angiotensin system. Virtually all of its biochemical actions are mediated through a single class of cell-surface receptors called AT₁. These receptors contain the structural features of the seven-transmembrane, G-protein-coupled receptor superfamily. Angiotensin II, acting through the AT₁ receptor, also stimulates the Jak/STAT pathway by inducing ligand-dependent Jak2 tyrosine phosphorylation and activation. Here, we show that a glutathione S-transferase fusion protein containing the carboxyl-terminal 54 amino acids of the rat AT_1A receptor physically binds to Jak2 in an angiotensin II-dependent manner. Deletional analysis, using both in vitro protocols and cell transfection analysis, showed that this association is dependent on the AT_1Areceptor motif YIPP (amino acids 319–322). The wild-type AT_1A receptor can induce Jak2 tyrosine phosphorylation. In contrast, an AT_1A receptor lacking the YIPP motif is unable to induce ligand-dependent phosphorylation of Jak2. Competition experiments with synthetic peptides suggest that each of the YIPP amino acids, including tyrosine 319, is important in Jak2 binding to the AT_1A receptor. The binding of the AT_1A receptor to the intracellular tyrosine kinase Jak2 supports the concept that the seven-transmembrane superfamily of receptors can physically associate with enzymatically active intracellular proteins, creating a signaling complex mechanistically similar to that observed with growth factor and cytokine receptors.

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BRIEF REVIEWSThe Importance of Tyrosine Phosphorylation in Angiotensin II Signaling

Abstract

Section snippets

• Protein Phosphorylation and Dephosphorylation

• Angiotensin II Stimulated Activation of Phospholipase C

• Angiotensin II and the Jak-STAT Pathway

• Angiotensin II and the Ras Pathway

• Conclusion

Acknowledgements

Semin Immunol

Trends Biochem Sci

J Biol Chem

J Biol Chem

J Biol Chem

J Biol Chem

Curr Opin Cell Biol

Biochem Biophys Res Commun

J Biol Chem

Biochim Biophys Acta

J Biol Chem

Angiotensin II increases inositol triphosphate and calcium in vascular smooth muscle cells

Hypertension

Inositol triphosphate and diacylglycerol: two interacting second messengers

Annu Rev Biochem

Angiotensin II stimulates sis-inducing factor-like DNA binding activity

J Biol Chem

Activation of the STAT pathway by angiotensin II in T3CHO/ATA1 cells

J Biol Chem

Regulators and effectors of ras proteins

Annu Rev Cell Biol

1002 protein phosphatases

Annu Rev Cell Biol

Angiotensin II-1 receptors mediated both vasoconstrictor and hypertrophic responses in rat aortic smooth muscle cells

Receptor

BRIEF REVIEWS
The Importance of Tyrosine Phosphorylation in Angiotensin II Signaling

Activation of the STAT pathway by angiotensin II in T3CHO/AT_A1 cells