scholarly journals Ndfip1 regulates nuclear Pten import in vivo to promote neuronal survival following cerebral ischemia

2012 ◽  
Vol 196 (1) ◽  
pp. 29-36 ◽  
Author(s):  
Jason Howitt ◽  
Jenny Lackovic ◽  
Ley-Hian Low ◽  
Adam Naguib ◽  
Alison Macintyre ◽  
...  
Keyword(s):  
Cerebral Ischemia ◽  
Tumor Suppression ◽  
Negative Regulator ◽  
Nuclear Entry ◽  
Interacting Protein ◽  
Akt Activation ◽  
Phosphorylated Akt ◽  
Phosphatase And Tensin Homologue

PTEN (phosphatase and tensin homologue deleted on chromosome TEN) is the major negative regulator of phosphatidylinositol 3-kinase signaling and has cell-specific functions including tumor suppression. Nuclear localization of PTEN is vital for tumor suppression; however, outside of cancer, the molecular and physiological events driving PTEN nuclear entry are unknown. In this paper, we demonstrate that cytoplasmic Pten was translocated into the nuclei of neurons after cerebral ischemia in mice. Critically, this transport event was dependent on a surge in the Nedd4 family–interacting protein 1 (Ndfip1), as neurons in Ndfip1-deficient mice failed to import Pten. Ndfip1 binds to Pten, resulting in enhanced ubiquitination by Nedd4 E3 ubiquitin ligases. In vitro, Ndfip1 overexpression increased the rate of Pten nuclear import detected by photobleaching experiments, whereas Ndfip1−/− fibroblasts showed negligible transport rates. In vivo, Ndfip1 mutant mice suffered larger infarct sizes associated with suppressed phosphorylated Akt activation. Our findings provide the first physiological example of when and why transient shuttling of nuclear Pten occurs and how this process is critical for neuron survival.

scholarly journals A role for thioredoxin-interacting protein (Txnip) in cellular creatine homeostasis

2013 ◽  
Vol 305 (2) ◽  
pp. E263-E270 ◽  
Author(s):  
Sevasti Zervou ◽  
Tanmoy Ray ◽  
Natasha Sahgal ◽  
Liam Sebag-Montefiore ◽  
Rebecca Cross ◽  
...  
Keyword(s):  
Feedback Inhibition ◽  
Global Gene Expression ◽  
Negative Regulator ◽  
Specific Response ◽  
Mrna Levels ◽  
Interacting Protein ◽  
Thioredoxin Interacting Protein ◽  
Creatine Uptake

Creatine is important for energy metabolism, yet excitable cells such as cardiomyocytes do not synthesize creatine and rely on uptake via a specific membrane creatine transporter (CrT; SLC6A8). This process is tightly controlled with downregulation of CrT upon continued exposure to high creatine via mechanisms that are poorly understood. Our aim was to identify candidate endogenous CrT inhibitors. In 3T3 cells overexpressing the CrT, creatine uptake plateaued at 3 h in response to 5 mM creatine but peaked 33% higher ( P < 0.01) in the presence of cycloheximide, suggesting CrT regulation depends on new protein synthesis. Global gene expression analysis identified thioredoxin-interacting protein (Txnip) as the only significantly upregulated gene (by 46%) under these conditions ( P = 0.036), subsequently verified independently at mRNA and protein levels. There was no change in Txnip expression with exposure to 5 mM taurine, confirming a specific response to creatine rather than osmotic stress. Small-interfering RNA against Txnip prevented Txnip upregulation in response to high creatine, maintained normal levels of creatine uptake, and prevented downregulation of CrT mRNA. These findings were relevant to the in vivo heart since creatine-deficient mice showed 39.71% lower levels of Txnip mRNA, whereas mice overexpressing the CrT had 57.6% higher Txnip mRNA levels and 28.7% higher protein expression compared with wild types (mean myocardial creatine concentration 124 and 74 nmol/mg protein, respectively). In conclusion, we have identified Txnip as a novel negative regulator of creatine levels in vitro and in vivo, responsible for mediating substrate feedback inhibition and a potential target for modulating creatine homeostasis.

scholarly journals The Adapter Protein ZIP Binds Grb14 and Regulates Its Inhibitory Action on Insulin Signaling by Recruiting Protein Kinase Cζ

2002 ◽  
Vol 22 (20) ◽  
pp. 6959-6970 ◽  
Author(s):  
Bertrand Cariou ◽  
Dominique Perdereau ◽  
Katia Cailliau ◽  
Edith Browaeys-Poly ◽  
Véronique Béréziat ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Insulin Receptor ◽  
Insulin Signaling ◽  
Inhibitory Action ◽  
Catalytic Domain ◽  
Inhibitory Effect ◽  
Negative Regulator ◽  
Interacting Protein

ABSTRACT Grb14 is a member of the Grb7 family of adapters and acts as a negative regulator of insulin-mediated signaling. Here we found that the protein kinase Cζ (PKCζ) interacting protein, ZIP, interacted with Grb14. Coimmunoprecipitation experiments demonstrated that ZIP bound to both Grb14 and PKCζ, thereby acting as a link in the assembly of a PKCζ-ZIP-Grb14 heterotrimeric complex. Mapping studies indicated that ZIP interacted through its ZZ zinc finger domain with the phosphorylated insulin receptor interacting region (PIR) of Grb14. PKCζ phosphorylated Grb14 under in vitro conditions and in CHO-IR cells as demonstrated by in vivo labeling experiments. Furthermore, Grb14 phosphorylation was increased under insulin stimulation, suggesting that the PKCζ-ZIP-Grb14 complex is involved in insulin signaling. The PIR of Grb14, which also interacts with the catalytic domain of the insulin receptor (IR) and inhibits its activity, was preferentially phosphorylated by PKCζ. Interestingly, the phosphorylation of Grb14 by PKCζ increased its inhibitory effect on IR tyrosine kinase activity in vitro. The role of ZIP and Grb14 in insulin signaling was further investigated in vivo in Xenopus laevis oocytes. In this model, ZIP potentiated the inhibitory action of Grb14 on insulin-induced oocyte maturation. Importantly, this effect required the recruitment of PKCζ and the phosphorylation of Grb14, providing in vivo evidences for a regulation of Grb14-inhibitory action by ZIP and PKCζ. Together, these results suggest that Grb14, ZIP, and PKCζ participate in a new feedback pathway of insulin signaling.

BI 836845, a fully human IGF ligand neutralizing antibody, to improve the efficacy of rapamycin by blocking rapamycin-induced AKT activation.

2012 ◽  
Vol 30 (15_suppl) ◽  
pp. 3092-3092 ◽  
Author(s):  
Paul J. Adam ◽  
Katrin Friedbichler ◽  
Marco H. Hofmann ◽  
Thomas Bogenrieder ◽  
Eric Borges ◽  
...  
Keyword(s):  
Cell Lines ◽  
Cancer Cell ◽  
Feedback Loop ◽  
Cancer Cell Lines ◽  
Phase I Clinical Trials ◽  
Intrinsic Resistance ◽  
Akt Activation ◽  
Phosphorylated Akt

3092 Background: Analogs of rapamycin (rapalogs) targeting mammalian target of rapamycin complex 1 (mTORC1) have shown clinical activity in several cancers. Nonetheless, preclinical and clinical data suggest that there may be intrinsic resistance to rapalogs through a feedback loop which activates upstream signaling when mTORC1 is blocked. BI 836845 is a fully human antibody, currently in phase I clinical trials, which potently neutralizes both IGF-1 and IGF-2. We tested whether BI 836845 is able to improve the efficacy of rapamycin by inhibiting upstream signaling in preclinical models. Methods: Cancer cell lines were profiled in vitro and in vivo for sensitivity to BI 836845 and rapamycin, alone or in combination. Mitogenic signaling was examined by measuring levels of phosphorylated AKT (pAKT) using Western blot analysis. IGF bioactivity was determined using a cellular IGF-1R phosphorylation ELISA. Results: The combination of BI 836845 and rapamycin was more effective than either agent alone at inhibiting the proliferation of Ewing’s sarcoma cells cultured in vitro as well as in a nude mouse xenograft model in vivo. Analysis of cell signaling upstream of mTOR demonstrated that treatment with rapamycin alone resulted in elevated pAKT, indicating feedback loop activation. BI 836845 treatment alone or in combination with rapamycin inhibited AKT phosphorylation, demonstrating that the rapamycin-induced increase in pAKT was due to elevated IGF bioactivity. Consistent with this we demonstrated that rapamycin increased IGF bioactivity in mice and that this could be inhibited by BI 836845. We extended these studies to include other cancer cell lines and profiled the correlation between improved efficacy of the combination with BI 836845 inhibition of rapamycin-induced feedback. A correlation has been observed for cancer cells derived from several indications. Conclusions: Rapamycin treatment increases AKT activation via elevated IGF ligand bioactivity. This effect can be inhibited by BI 836845, thus explaining the improved pre-clinical efficacy seen when both agents are combined. These data provide a rationale for the clinical combination of rapalogs and BI 836845.

Rapamycin Analogs Reduce mTORC2 Signaling and Inhibit AKT Activation in AML.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 156-156 ◽  
Author(s):  
Zhihong Zeng ◽  
Dos D. Sarbassov ◽  
Francis J. Giles ◽  
Ismael Samudio ◽  
Karen W.L. Yee ◽  
...  
Keyword(s):  
Cyclin D1 ◽  
Peripheral Blood ◽  
Therapeutic Potential ◽  
Absolute Lymphocyte Count ◽  
Mtor Inhibition ◽  
Akt Activation ◽  
Glut 1 ◽  
Phosphorylated Akt

Abstract The mTOR complex 2 (mTORC2) containing mTOR and rictor is thought to be rapamycin-insensitive, and was recently shown to regulate the pro-survival kinase AKT by phosphorylation on Ser473 (Sarbassov Science 2005;307 and Mol Cell 2006;22). We investigated the molecular effects of mTOR inhibition by rapamycin analog CCI-779 in AML cells. Unexpectedly, CCI-779 not only inhibited the mTOR complex 1 (mTORC1) containing mTOR and raptor with decreased phosphorylation of p70S6K, 4EPB1 and reduction in Glut-1 mRNA, but also blocked AKT activation via inhibition of mTORC2 formation. This resulted in suppression of phosphorylation of the direct AKT substrate FKHR and decreased transcription of D-Cyclins in AML cell line and 5 of 8 primary AML samples in vitro. Similar observations were made in samples from patients with hematological malignancies who were treated with the rapamycin analogs temsirolimus or everolimus: the levels of Ser473 phosphorylated AKT decreased in 3/5 patient samples at 1 or 24 hour(s) of temsirolimus treatment, and in 6/8 patient samples treated with everolimus. In the 9 samples in which AKT was inhibited, ≥2-fold decrease in Cyclin D1 mRNA was observed in 5, Cyclin D2 in 3, both, Cyclin D1 and D2 in 1 sample, and Glut-1 in 4 patient samples. In 7 of the 9 patients in whom AKT was inhibited, a >50% decrease in peripheral blood absolute blast count (3 AML, 1 ALL) or absolute lymphocyte count (1 CLL) for >1 week duration was documented, and two patients with RAEB-1 had improvements in platelet counts, one fulfilling the criteria for clinical response (hematological improvement). No change in peripheral blood counts or progression of leukemia was seen in 6 patients. Of these, decrease in pAKT was observed in 2, no change in 3 and increase in 1 (Fisher exact two-tailed p= 0.021). Altogether, our study provides first evidence that rapamycin analogs inhibit AKT signaling in primary AML cells both in vitro and in vivo, and support the therapeutic potential of mTOR inhibition strategies in leukemias.

scholarly journals Increased mechanical strain imposed on murine lungs during ventilation in vivo depresses airway responsiveness and activation of protein kinase Akt

2013 ◽  
Vol 114 (11) ◽  
pp. 1506-1510 ◽  
Author(s):  
Z. Xue ◽  
W. Zhang ◽  
L. P. Desai ◽  
H. Gao ◽  
S. J. Gunst ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Mechanical Strain ◽  
Airway Responsiveness ◽  
Negative Regulator ◽  
High Peep ◽  
Mechanical Loads ◽  
Akt Activation ◽  
Isolated Lung

Continuous positive airway pressure (CPAP) administered to tracheostomized rabbits and ferrets for 4 days or 2 wk suppresses bronchial reactivity in vivo and suppresses airway reactivity in lobes and tracheal segments isolated from these animals. In vitro studies of canine tracheal smooth muscle tissues indicate that mechanical loading suppresses the activation of the growth regulatory kinase, Akt, and that Akt is a negative regulator of smooth muscle differentiation. The transduction of mechanical signals in the tracheal tissues in vitro is mediated by integrin-associated adhesion complexes. To determine whether airway responsiveness and Akt activation are modulated by mechanical loads applied for short time periods to the airways of living animals in vivo, mice were mechanically ventilated for 2 h with high (5 cmH2O) or low (0–1 cmH2O) positive end-expiratory pressure (PEEP) and then ventilated at low PEEP for 30 min. Ventilation of mice with PEEP in vivo for 2 h depressed airway responsiveness to methacholine measured in vivo subsequent to the PEEP treatment. Airway narrowing in vitro in intraparenchymal airways in isolated lung slices and contractile responses of isolated tracheal segments in vitro were suppressed for at least 6 h subsequent to the in vivo exposure to PEEP. Tracheal segments isolated from high PEEP-treated mice exhibited significantly lower levels of Akt activation than tracheae from low PEEP-treated mice. The results indicate that mechanical loads imposed in vivo result in physiological and biochemical changes in the airway tissues after a relatively short 2-h period of in vivo loading.

scholarly journals Targeting of C-Terminal Binding Protein (CtBP) by ARF Results in p53-Independent Apoptosis

2006 ◽  
Vol 26 (6) ◽  
pp. 2360-2372 ◽  
Author(s):  
Seema Paliwal ◽  
Sandhya Pande ◽  
Ramesh C. Kovi ◽  
Norman E. Sharpless ◽  
Nabeel Bardeesy ◽  
...  
Keyword(s):  
Tumor Suppression ◽  
Negative Regulator ◽  
Interacting Proteins ◽  
Yeast Two Hybrid ◽  
Induced Apoptosis ◽  
Interfering Rna ◽  
Two Hybrid ◽  
Potent Tumor

ABSTRACT ARF encodes a potent tumor suppressor that antagonizes MDM2, a negative regulator of p53. ARF also suppresses the proliferation of cells lacking p53, and loss of ARF in p53-null mice, compared with ARF or p53 singly null mice, results in a broadened tumor spectrum and decreased tumor latency. To investigate the mechanism of p53-independent tumor suppression by ARF, potential interacting proteins were identified by yeast two-hybrid screen. The antiapoptotic transcriptional corepressor C-terminal binding protein 2 (CtBP2) was identified, and ARF interactions with both CtBP1 and CtBP2 were confirmed in vitro and in vivo. Interaction with ARF resulted in proteasome-dependent CtBP degradation. Both ARF-induced CtBP degradation and CtBP small interfering RNA led to p53-independent apoptosis in colon cancer cells. ARF induction of apoptosis was dependent on its ability to interact with CtBP, and reversal of ARF-induced CtBP depletion by CtBP overexpression abrogated ARF-induced apoptosis. CtBP proteins represent putative targets for p53-independent tumor suppression by ARF.

scholarly journals Anti-Cancer Properties of Theaflavins

Molecules ◽  
2021 ◽  
Vol 26 (4) ◽  
pp. 987
Author(s):  
Eric J. O’Neill ◽  
Deborah Termini ◽  
Alexandria Albano ◽  
Evangelia Tsiani
Keyword(s):  
Signaling Pathways ◽  
Cancer Progression ◽  
Treatment Strategies ◽  
B Cell Lymphoma ◽  
Black Tea ◽  
Phosphorylated Akt ◽  
Phenolic Components ◽  
Anti Cancer

Cancer is a disease characterized by aberrant proliferative and apoptotic signaling pathways, leading to uncontrolled proliferation of cancer cells combined with enhanced survival and evasion of cell death. Current treatment strategies are sometimes ineffective in eradicating more aggressive, metastatic forms of cancer, indicating the need to develop novel therapeutics targeting signaling pathways which are essential for cancer progression. Historically, plant-derived compounds have been utilized in the production of pharmaceuticals and chemotherapeutic compounds for the treatment of cancer, including paclitaxel and docetaxel. Theaflavins, phenolic components present in black tea, have demonstrated anti-cancer potential in cell cultures in vitro and in animal studies in vivo. Theaflavins have been shown to inhibit proliferation, survival, and migration of many cancer cellswhile promoting apoptosis. Treatment with theaflavins has been associated with increased levels of cleaved poly (ADP-ribose) polymerase (PARP) and cleaved caspases-3, -7, -8, and -9, all markers of apoptosis, and increased expression of the proapoptotic marker Bcl-2-associated X protein (Bax) and concomitant reduction in the antiapoptotic marker B-cell lymphoma 2 (Bcl-2). Additionally, theaflavin treatment reduced phosphorylated Akt, phosphorylated mechanistic target of rapamycin (mTOR), phosphatidylinositol 3-kinase (PI3K), and c-Myc levels with increased expression of the tumour suppressor p53. This review summarizes the current in vitro and in vivo evidence available investigating the anti-cancer effects of theaflavins across various cancer cell lines and animal models.

scholarly journals LINGO-1 shRNA protects the brain against ischemia/reperfusion injury by inhibiting the activation of NF-κB and JAK2/STAT3

Human Cell ◽  
2021 ◽  
Author(s):  
Jiaying Zhu ◽  
Zhu Zhu ◽  
Yipin Ren ◽  
Yukang Dong ◽  
Yaqi Li ◽  
...  
Keyword(s):  
Cerebral Ischemia ◽  
Nuclear Translocation ◽  
Ischemia Reperfusion Injury ◽  
Ischemia Reperfusion ◽  
Glucose Deprivation ◽  
Artery Occlusion ◽  
Mice Model ◽  
Down Regulation

AbstractLINGO-1 may be involved in the pathogenesis of cerebral ischemia. However, its biological function and underlying molecular mechanism in cerebral ischemia remain to be further defined. In our study, middle cerebral artery occlusion/reperfusion (MACO/R) mice model and HT22 cell oxygen–glucose deprivation/reperfusion (OGD/R) were established to simulate the pathological process of cerebral ischemia in vivo and in vitro and to detect the relevant mechanism. We found that LINGO-1 mRNA and protein were upregulated in mice and cell models. Down-regulation LINGO-1 improved the neurological symptoms and reduced pathological changes and the infarct size of the mice after MACO/R. In addition, LINGO-1 interference alleviated apoptosis and promoted cell proliferation in HT22 of OGD/R. Moreover, down-regulation of LINGO-1 proved to inhibit nuclear translocation of p-NF-κB and reduce the expression level of p-JAK2 and p-STAT3. In conclusion, our data suggest that shLINGO-1 attenuated ischemic injury by negatively regulating NF-KB and JAK2/STAT3 pathways, highlighting a novel therapeutic target for ischemic stroke.

scholarly journals Hyperglycaemia up-regulates placental growth factor (PlGF) expression and secretion in endothelial cells via suppression of PI3 kinase-Akt signalling and activation of FOXO1

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Samir Sissaoui ◽  
Stuart Egginton ◽  
Ling Ting ◽  
Asif Ahmed ◽  
Peter W. Hewett
Keyword(s):  
Endothelial Cells ◽  
Growth Factor ◽  
Endothelial Dysfunction ◽  
Akt Activation ◽  
Forkhead Box ◽  
Pi3k Activity ◽  
Binding Sequence

AbstractPlacenta growth factor (PlGF) is a pro-inflammatory angiogenic mediator that promotes many pathologies including diabetic complications and atherosclerosis. Widespread endothelial dysfunction precedes the onset of these conditions. As very little is known of the mechanism(s) controlling PlGF expression in pathology we investigated the role of hyperglycaemia in the regulation of PlGF production in endothelial cells. Hyperglycaemia stimulated PlGF secretion in cultured primary endothelial cells, which was suppressed by IGF-1-mediated PI3K/Akt activation. Inhibition of PI3K activity resulted in significant PlGF mRNA up-regulation and protein secretion. Similarly, loss or inhibition of Akt activity significantly increased basal PlGF expression and prevented any further PlGF secretion in hyperglycaemia. Conversely, constitutive Akt activation blocked PlGF secretion irrespective of upstream PI3K activity demonstrating that Akt is a central regulator of PlGF expression. Knock-down of the Forkhead box O-1 (FOXO1) transcription factor, which is negatively regulated by Akt, suppressed both basal and hyperglycaemia-induced PlGF secretion, whilst FOXO1 gain-of-function up-regulated PlGF in vitro and in vivo. FOXO1 association to a FOXO binding sequence identified in the PlGF promoter also increased in hyperglycaemia. This study identifies the PI3K/Akt/FOXO1 signalling axis as a key regulator of PlGF expression and unifying pathway by which PlGF may contribute to common disorders characterised by endothelial dysfunction, providing a target for therapy.

scholarly journals Coordinated Regulation of Insulin Signaling by the Protein Tyrosine Phosphatases PTP1B and TCPTP

2005 ◽  
Vol 25 (2) ◽  
pp. 819-829 ◽  
Author(s):  
Sandra Galic ◽  
Christine Hauser ◽  
Barbara B. Kahn ◽  
Fawaz G. Haj ◽  
Benjamin G. Neel ◽  
...  
Keyword(s):  
Insulin Signaling ◽  
Tyrosine Phosphatase ◽  
Protein Tyrosine Phosphatases ◽  
Negative Regulator ◽  
Mitogen Activated Protein Kinase ◽  
Protein Tyrosine ◽  
Akt Activation ◽  
Protein Levels ◽  
Mitogen Activated Protein

ABSTRACT The protein tyrosine phosphatase PTP1B is a negative regulator of insulin signaling and a therapeutic target for type 2 diabetes. Our previous studies have shown that the closely related tyrosine phosphatase TCPTP might also contribute to the regulation of insulin receptor (IR) signaling in vivo (S. Galic, M. Klingler-Hoffmann, M. T. Fodero-Tavoletti, M. A. Puryer, T. C. Meng, N. K. Tonks, and T. Tiganis, Mol. Cell. Biol. 23:2096-2108, 2003). Here we show that PTP1B and TCPTP function in a coordinated and temporally distinct manner to achieve an overall regulation of IR phosphorylation and signaling. Whereas insulin-induced phosphatidylinositol 3-kinase/Akt signaling was prolonged in both TCPTP−/− and PTP1B−/− immortalized mouse embryo fibroblasts (MEFs), mitogen-activated protein kinase ERK1/2 signaling was elevated only in PTP1B-null MEFs. By using phosphorylation-specific antibodies, we demonstrate that both IR β-subunit Y1162/Y1163 and Y972 phosphorylation are elevated in PTP1B−/− MEFs, whereas Y972 phosphorylation was elevated and Y1162/Y1163 phosphorylation was sustained in TCPTP−/− MEFs, indicating that PTP1B and TCPTP differentially contribute to the regulation of IR phosphorylation and signaling. Consistent with this, suppression of TCPTP protein levels by RNA interference in PTP1B−/− MEFs resulted in no change in ERK1/2 signaling but caused prolonged Akt activation and Y1162/Y1163 phosphorylation. These results demonstrate that PTP1B and TCPTP are not redundant in insulin signaling and that they act to control both common as well as distinct insulin signaling pathways in the same cell.

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