The Akt1-eNOS Axis Illustrates the Specificity of Kinase-Substrate Relationships in Vivo

Schleicher, Michael; Yu, Jun; Murata, Takahisa; Derakhshan, Berhad; Atochin, Dmitriy N.; Li, Qian; Kashiwagi, Satoshi; Lorenzo, Annarita Di; Harrison, Kathryn; Huang, Paul L.; Sessa, William C.

doi:10.1126/scisignal.2000343

Cited by 86 publications

(103 citation statements)

References 46 publications

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“…Phosphorylation of eNOS at Serine 1177 has been shown to be primarily mediated by activation of AKT (16,17). In this study we observed BMP-mediated AKT phosphorylation, which is indicative of increased AKT activity.…”

Section: Discussionsupporting

confidence: 60%

Bone Morphogenetic Protein Receptor II Is a Novel Mediator of Endothelial Nitric-oxide Synthase Activation

Gangopahyay

Oran

Bauer

et al. 2011

Journal of Biological Chemistry

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Activation of bone morphogenetic protein (BMP) receptor II (BMPRII) promotes pulmonary artery endothelial cell (PAEC) survival, proliferation, and migration. Mutations to BMPRII are associated with the development of pulmonary arterial hypertension (PAH). Endothelial dysfunction, including decreased endothelial nitric-oxide synthase (eNOS) activity and loss of bioactive nitric oxide (NO), plays a prominent role in the development of PAH. We hypothesized that stimulation of BMPRII promotes normal PAEC function by activating eNOS. We report that BMPRII ligands, BMP2 and BMP4, (i) stimulate eNOS phosphorylation at a critical regulatory site, (ii) increase eNOS activity, and (iii) result in canonical changes in eNOS proteinprotein interactions. The stimulation of eNOS activity by BMPRII ligands was largely dependent on protein kinase A (PKA) activation, as demonstrated using the PKA inhibitors H89 and myristoylated PKI(6 -22) amide. PAEC migration stimulated by BMP2 and BMP4 was inhibited by the NOS inhibitor L-nitroarginine methyl ester, providing functional evidence of eNOS activation. Furthermore, BMP2 and BMP4 failed to stimulate eNOS phosphorylation when BMPRII was knocked down by siRNA. Most important to the pathophysiology of the disease, BMP2 and BMP4 failed to stimulate eNOS phosphorylation in PAECs isolated from patients with mutations in the BMPR2 gene. These data demonstrate a new action of BMPs/ BMPRII in the pulmonary endothelium and provide novel mechanistic insight into the pathogenesis of PAH. Pulmonary arterial hypertension (PAH)2 is a devastating disease, characterized by increasing pulmonary arterial pressures, leading to right heart failure and eventually, death (1, 2). Recent studies demonstrate an important role for BMPRII in the pathogenesis of the disease when mutations to the BMPR2 gene were identified as a major underlying cause of heritable or familial PAH (FPAH) (3-5). Approximately 144 different mutations in 210 patients with FPAH have now been identified (6 -8). Additionally, it is estimated that 20% of patients with idiopathic PAH (IPAH) harbor mutations to the BMPR2 gene. These mutations may be a previously undetected inherited mutation or arise from sporadic mutation of the gene.BMPRII is widely expressed in normal tissues. In the lung vasculature, BMPRII is most highly expressed on endothelial cells (9) and at lower levels in smooth muscle cells and fibroblasts. Expression of BMPRII is markedly reduced in the pulmonary vasculature of patients with FPAH and IPAH, regardless of whether they harbor mutations in the BMPR2 gene (9).Pulmonary artery smooth muscle cells (PASMCs) from patients with mutations in BMPRII have a reduced capacity to activate Smad1/5. This is coupled with a reduced ability of BMPs to inhibit proliferation of PASMCs (10). In contrast, the BMPRII ligands BMP2 and BMP4 promote proliferation, migration, and survival in PAECs (11, 12). The opposite effects of BMPs on PAECs and PASMCs provide a convincing model for pulmonary vascular damage in PAH, where a loss of BMP...

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Section: Discussionsupporting

confidence: 60%

Bone Morphogenetic Protein Receptor II Is a Novel Mediator of Endothelial Nitric-oxide Synthase Activation

Gangopahyay

Oran

Bauer

et al. 2011

Journal of Biological Chemistry

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“…Immunostaining demonstrated that the increase in HIF-1α expression occurred in a perivascular distribution in anemic WT but not nNOS −/− mice (Fig. S4), suggesting that NOS-derived NO may play a role in increasing HIF-1α protein levels in the microvasculature (9,27). Collectively, these data suggest that priming of HIF-1α by nNOSderived NO is essential for adaptive responses to severe anemia, possibly modifying S-nitrosothiol signaling (28).…”

Section: Nnos Contributes To Adaptive Cardiovascular Responses Duringmentioning

confidence: 74%

Priming of hypoxia-inducible factor by neuronal nitric oxide synthase is essential for adaptive responses to severe anemia

Tsui

Marsden²,

Mazer³

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

103

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Cells sense and respond to changes in oxygen concentration through gene regulatory processes that are fundamental to survival. Surprisingly, little is known about how anemia affects hypoxia signaling. Because nitric oxide synthases (NOSs) figure prominently in the cellular responses to acute hypoxia, we defined the effects of NOS deficiency in acute anemia. In contrast to endothelial NOS or inducible NOS deficiency, neuronal NOS (nNOS) −/− mice demonstrated increased mortality during anemia. Unlike wild-type (WT) animals, anemia did not increase cardiac output (CO) or reduce systemic vascular resistance (SVR) in nNOS −/− mice. At the cellular level, anemia increased expression of HIF-1α protein and HIF-responsive mRNA levels (EPO, VEGF, GLUT1, PDK1) in the brain of WT, but not nNOS −/− mice, despite comparable reductions in tissue PO 2 . Paradoxically, nNOS −/− mice survived longer during hypoxia, retained the ability to regulate CO and SVR, and increased brain HIF-α protein levels and HIF-responsive mRNA transcripts. Real-time imaging of transgenic animals expressing a reporter HIF-α(ODD)-luciferase chimeric protein confirmed that nNOS was essential for anemia-mediated increases in HIF-α protein stability in vivo. S -nitrosylation effects the functional interaction between HIF and pVHL. We found that anemia led to nNOS-dependent S -nitrosylation of pVHL in vivo and, of interest, led to decreased expression of GSNO reductase. These findings identify nNOS effects on the HIF/pVHL signaling pathway as critically important in the physiological responses to anemia in vivo and provide essential mechanistic insight into the differences between anemia and hypoxia.

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“…Consistent with the observed local increases in NO production in brain vasculature after rapamycin or ACh treatment, both drugs induced phosphorylation of endothelial NO synthase (eNOS) at Ser1176, which is critical for its activation. 54 Ser1176 phosphorylation of eNOS after rapamycin and after ACh treatment proceeded with a comparable time course and showed maximal Ser1176-eNOS phosphorylation 1 minute after administration ( Figures 4G and H). To determine whether rapamycin-induced NO release and vasodilation were dependent on NO synthase (NOS) activity, we pretreated experimental animals with a NOS inhibitor (L-NAME, which becomes a functional inhibitor, L-NAA, after hydrolysis by esterases) immediately before the administration of rapamycin or ACh.…”

Section: Increased Vascular Density Without Changes In Glucosementioning

confidence: 91%

“…66,68 Restoration of vascular density in AD mice required NO synthase activity, suggesting that mTOR has a critical role in the inhibition of NO release in brain vascular endothelium during the progression of AD-like disease in mice ( Figure 5). Of note, rapamycin did not affect vascular density or CBF in WT mice, suggesting that restoration of vascular integrity by chronic mTOR inhibition involves a pathway activated by chronic injury 54 such as that associated with the progression of neurodegeneration in AD mice. Because mTOR controls key metabolic functions in most cell types, and pharmacologically inhibiting mTOR extends lifespan and healthspan, [18][19][20] retarding multiple, but not all, aspects of aging in mice, it is possible that mTOR may be involved in several different, specific processes of complex disease mechanisms that enable neurodegeneration.…”

Section: Discussionmentioning

confidence: 99%

“…Pretreatment with L-NAME abrogated both NO release at brain endothelium and vasodilation induced by rapamycin administration ( Figure 4E) as well as by ACh (not shown) as previously demonstrated. 54 The observed localization and time course of NO release ( Figure 4D), which rule out an involvement of the inducible isoform of NOS (iNOS), and the phosphorylation of eNOS at Ser1176 (Figures 4G and H) suggest that NO release and vasodilation by acute rapamycin treatment may depend on eNOS activation.…”

Section: Increased Vascular Density Without Changes In Glucosementioning

confidence: 99%

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Chronic Rapamycin Restores Brain Vascular Integrity and Function Through NO Synthase Activation and Improves Memory in Symptomatic Mice Modeling Alzheimer’s Disease

Lin¹,

Zheng

Halloran

et al. 2013

J Cereb Blood Flow Metab

177

201

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Vascular pathology is a major feature of Alzheimer's disease (AD) and other dementias. We recently showed that chronic administration of the target-of-rapamycin (TOR) inhibitor rapamycin, which extends lifespan and delays aging, halts the progression of AD-like disease in transgenic human (h)APP mice modeling AD when administered before disease onset. Here we demonstrate that chronic reduction of TOR activity by rapamycin treatment started after disease onset restored cerebral blood flow (CBF) and brain vascular density, reduced cerebral amyloid angiopathy and microhemorrhages, decreased amyloid burden, and improved cognitive function in symptomatic hAPP (AD) mice. Like acetylcholine (ACh), a potent vasodilator, acute rapamycin treatment induced the phosphorylation of endothelial nitric oxide (NO) synthase (eNOS) and NO release in brain endothelium. Administration of the NOS inhibitor L-NG-Nitroarginine methyl ester reversed vasodilation as well as the protective effects of rapamycin on CBF and vasculature integrity, indicating that rapamycin preserves vascular density and CBF in AD mouse brains through NOS activation. Taken together, our data suggest that chronic reduction of TOR activity by rapamycin blocked the progression of AD-like cognitive and histopathological deficits by preserving brain vascular integrity and function. Drugs that inhibit the TOR pathway may have promise as a therapy for AD and possibly for vascular dementias.

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The Akt1-eNOS Axis Illustrates the Specificity of Kinase-Substrate Relationships in Vivo

Cited by 86 publications

References 46 publications

Bone Morphogenetic Protein Receptor II Is a Novel Mediator of Endothelial Nitric-oxide Synthase Activation

Bone Morphogenetic Protein Receptor II Is a Novel Mediator of Endothelial Nitric-oxide Synthase Activation

Priming of hypoxia-inducible factor by neuronal nitric oxide synthase is essential for adaptive responses to severe anemia

Chronic Rapamycin Restores Brain Vascular Integrity and Function Through NO Synthase Activation and Improves Memory in Symptomatic Mice Modeling Alzheimer’s Disease

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