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Sample records for akt signal transduction

  1. PI3 kinase directly phosphorylates Akt1/2 at Ser473/474 in the insulin signal transduction pathway

    PubMed Central

    Tsuchiya, A; Kanno, T; Nishizaki, T

    2014-01-01

    Insulin stimulated translocation of the glucose transporter GLUT4 from the cytosol to the plasma membrane in a concentration (1 nM–1 μM)-dependent manner and increased glucose uptake in 3T3-L1 adipocytes. Insulin-induced GLUT4 translocation to the cell surface was prevented by the phosphoinositide 3 kinase (PI3K) inhibitor wortmannin, the 3-phosphoinositide-dependent protein kinase 1 (PDK1) inhibitor BX912 or the Akt1/2 inhibitor MK2206, and by knocking-down PI3K, PDK1 or Akt1/2. Insulin increased phosphorylation of Akt1/2 at Thr308/309 and Ser473/474, to activate Akt1/2, in the adipocytes. Insulin-induced phosphorylation of Akt1/2 was suppressed by wortmannin and knocking-down PI3K, while no significant inhibition of the phosphorylation was obtained with BX912 or knocking-down PDK1. In the cell-free Akt assay, PI3K phosphorylated Akt1 both at Thr308 and Ser473 and Akt2 at Ser474 alone. In contrast, PDK1 phosphorylates Akt1 at Thr308 and Akt2 at Thr309. The results of this study indicate that PI3K activates Akt1, independently of PDK1, and Akt2 by cooperating with PDK1 in the insulin signal transduction pathway linked to GLUT4 translocation. PMID:24169049

  2. Methylmercury, an environmental electrophile capable of activation and disruption of the Akt/CREB/Bcl-2 signal transduction pathway in SH-SY5Y cells

    PubMed Central

    Unoki, Takamitsu; Abiko, Yumi; Toyama, Takashi; Uehara, Takashi; Tsuboi, Koji; Nishida, Motohiro; Kaji, Toshiyuki; Kumagai, Yoshito

    2016-01-01

    Methylmercury (MeHg) modifies cellular proteins via their thiol groups in a process referred to as “S-mercuration”, potentially resulting in modulation of the cellular signal transduction pathway. We examined whether low-dose MeHg could affect Akt signaling involved in cell survival. Exposure of human neuroblastoma SH-SY5Y cells of up to 2 μM MeHg phosphorylated Akt and its downstream signal molecule CREB, presumably due to inactivation of PTEN through S-mercuration. As a result, the anti-apoptotic protein Bcl-2 was up-regulated by MeHg. The activation of Akt/CREB/Bcl-2 signaling mediated by MeHg was, at least in part, linked to cellular defence because either pretreatment with wortmannin to block PI3K/Akt signaling or knockdown of Bcl-2 enhanced MeHg-mediated cytotoxicity. In contrast, increasing concentrations of MeHg disrupted Akt/CREB/Bcl-2 signaling. This phenomenon was attributed to S-mercuration of CREB through Cys286 rather than Akt. These results suggest that although MeHg is an apoptosis-inducing toxicant, this environmental electrophile is able to activate the cell survival signal transduction pathway at lower concentrations prior to apoptotic cell death. PMID:27357941

  3. Methylmercury, an environmental electrophile capable of activation and disruption of the Akt/CREB/Bcl-2 signal transduction pathway in SH-SY5Y cells.

    PubMed

    Unoki, Takamitsu; Abiko, Yumi; Toyama, Takashi; Uehara, Takashi; Tsuboi, Koji; Nishida, Motohiro; Kaji, Toshiyuki; Kumagai, Yoshito

    2016-01-01

    Methylmercury (MeHg) modifies cellular proteins via their thiol groups in a process referred to as "S-mercuration", potentially resulting in modulation of the cellular signal transduction pathway. We examined whether low-dose MeHg could affect Akt signaling involved in cell survival. Exposure of human neuroblastoma SH-SY5Y cells of up to 2 μM MeHg phosphorylated Akt and its downstream signal molecule CREB, presumably due to inactivation of PTEN through S-mercuration. As a result, the anti-apoptotic protein Bcl-2 was up-regulated by MeHg. The activation of Akt/CREB/Bcl-2 signaling mediated by MeHg was, at least in part, linked to cellular defence because either pretreatment with wortmannin to block PI3K/Akt signaling or knockdown of Bcl-2 enhanced MeHg-mediated cytotoxicity. In contrast, increasing concentrations of MeHg disrupted Akt/CREB/Bcl-2 signaling. This phenomenon was attributed to S-mercuration of CREB through Cys286 rather than Akt. These results suggest that although MeHg is an apoptosis-inducing toxicant, this environmental electrophile is able to activate the cell survival signal transduction pathway at lower concentrations prior to apoptotic cell death. PMID:27357941

  4. Stimulatory Effects of Coumestrol on Embryonic and Fetal Development Through AKT and ERK1/2 MAPK Signal Transduction.

    PubMed

    Lim, Whasun; Song, Gwonhwa

    2016-12-01

    Successful establishment of pregnancy is required for fetal-maternal interactions regulating implantation, embryonic development and placentation. A uterine environment with insufficient growth factors and nutrients increases the incidence of intrauterine growth restriction (IUGR) leading to an impaired uterine environment. In the present study, we demonstrated the effects of the phytoestrogen coumestrol on conceptus development in the pig that is regarded as an excellent biomedical animal model for research on IUGR. Results of this study indicated that coumestrol induced migration of porcine trophectoderm (pTr) cells in a concentration-dependent manner. In response to coumestrol, the phosphorylation of AKT, P70S6K, S6, ERK1/2 MAPK, and P90RSK proteins were activated in pTr cells and ERK1/2 MAPK and P90RSK phosphorylation was prolonged for a longer period than for the other proteins. To identify the signal transduction pathway induced by coumestrol, pharmacological inhibitors U0126 (an ERK1/2 inhibitor) and LY294002 (a PI3K inhibitor) were used to pretreat pTr cells. The results showed that coumestrol-induced phosphorylation of ERK1/2 MAPK and P90RSK was blocked by U0126. In addition, the increased phosphorylation in response to coumestrol was completely inhibited following pre-treatment incubation of pTr cells in the presence of LY294002 and U0126. Furthermore, these two inhibitors suppressed the ability of coumestrol to induce migration of pTr cells. Collectively, these findings suggest that coumestrol affects embryonic development through activation of the PI3K/AKT and ERK1/2 MAPK cell signal transduction pathways and improvement in the uterine environment through coumestrol supplementation may provide beneficial effects of enhancing embryonic and fetal survival and development. J. Cell. Physiol. 231: 2733-2740, 2016. © 2016 Wiley Periodicals, Inc. PMID:26991852

  5. Bis(hinokitiolato)zinc complex ([Zn(hkt)2]) activates Akt/protein kinase B independent of insulin signal transduction.

    PubMed

    Naito, Yuki; Yoshikawa, Yutaka; Masuda, Kazufumi; Yasui, Hiroyuki

    2016-07-01

    Since many Zn complexes have been developed to enhance the insulin-like activity and increase the exposure and residence of Zn in the animal body, these complexes are recognized as one of the new candidates with action mechanism different from existing anti-diabetic drugs. However, the molecular mechanism by which Zn complexes exert an anti-DM effect is unknown. Therefore, we evaluated the activity of Zn complexes, especially related to the phosphorylation of insulin signaling pathway components. We focused on the insulin-like effects of the bis(hinokitiolato)zinc complex, [Zn(hkt)2], using 3T3-L1 adipocytes. [Zn(hkt)2] was taken up by cells and induced Akt phosphorylation in a time-dependent manner. Additionally, it showed inhibitory activity against PTP1B and PTEN, which are major negative regulators of insulin signaling. It did not promote the phosphorylation of IR (insulin receptor)-β or IRS (insulin receptor substrate)-1 by itself, but in combination with insulin, it enhanced the phosphorylation of IRβ. We conclude that [Zn(hkt)2] has effects on the proteins of insulin signaling pathway without insulin receptor mediation, and [Zn(hkt)2] promotes insulin function and shows the anti-DM effects. Thus, [Zn(hkt)2] may be the basis for improved DM treatments. PMID:27251140

  6. Activation of the PI3K/Akt signal transduction pathway and increased levels of insulin receptor in protein repair-deficient mice.

    PubMed

    Farrar, Christine; Houser, Carolyn R; Clarke, Steven

    2005-02-01

    Protein L-isoaspartate (D-aspartate) O-methyltransferase is an enzyme that catalyses the repair of isoaspartyl damage in proteins. Mice lacking this enzyme (Pcmt1-/- mice) have a progressive increase in brain size compared with wild-type mice (Pcmt1+/+ mice), a phenotype that can be associated with alterations in the PI3K/Akt signal transduction pathway. Here we show that components of this pathway, including Akt, GSK3beta and PDK-1, are more highly phosphorylated in the brains of Pcmt1-/- mice, particularly in cells of the hippocampus, in comparison with Pcmt1+/+ mice. Examination of upstream elements of this pathway in the hippocampus revealed that Pcmt1-/- mice have increased activation of insulin-like growth factor-I (IGF-I) receptor and/or insulin receptor. Western blot analysis revealed an approximate 200% increase in insulin receptor protein levels and an approximate 50% increase in IGF-I receptor protein levels in the hippocampus of Pcmt1-/- mice. Higher levels of the insulin receptor protein were also found in other regions of the adult brain and in whole tissue extracts of brain, liver, heart and testes of both juvenile and adult Pcmt1-/- mice. There were no significant differences in plasma insulin levels for adult Pcmt1-/- mice during glucose tolerance tests. However, they did show higher peak levels of blood glucose, suggesting a mild impairment in glucose tolerance. We propose that Pcmt1-/- mice have altered regulation of the insulin pathway, possibly as a compensatory response to altered glucose uptake or metabolism or as an adaptive response to a general accumulation of isoaspartyl protein damage in the brain and other tissues. PMID:15659208

  7. Oxidant Stress and Signal Transduction in the Nervous System with the PI 3-K, Akt, and mTOR Cascade

    PubMed Central

    Maiese, Kenneth; Chong, Zhao Zhong; Wang, Shaohui; Shang, Yan Chen

    2012-01-01

    Oxidative stress impacts multiple systems of the body and can lead to some of the most devastating consequences in the nervous system especially during aging. Both acute and chronic neurodegenerative disorders such as diabetes mellitus, cerebral ischemia, trauma, Alzheimer’s disease, Parkinson’s disease, Huntington’s disease, and tuberous sclerosis through programmed cell death pathways of apoptosis and autophagy can be the result of oxidant stress. Novel therapeutic avenues that focus upon the phosphoinositide 3-kinase (PI 3-K), Akt (protein kinase B), and the mammalian target of rapamycin (mTOR) cascade and related pathways offer exciting prospects to address the onset and potential reversal of neurodegenerative disorders. Effective clinical translation of these pathways into robust therapeutic strategies requires intimate knowledge of the complexity of these pathways and the ability of this cascade to influence biological outcome that can vary among disorders of the nervous system. PMID:23203037

  8. The role of the PI3K-Akt signal transduction pathway in Autographa californica multiple nucleopolyhedrovirus infection of Spodoptera frugiperda cells

    SciTech Connect

    Xiao Wei; Yang Yi; Weng Qingbei; Lin Tiehao; Yuan Meijin; Yang Kai; Pang Yi

    2009-08-15

    Many viruses activate the phosphatidylinositol 3-kinase (PI3K)-Akt signaling pathway, thereby modulating diverse downstream signaling pathways associated with antiapoptosis, proliferation, cell cycling, protein synthesis and glucose metabolism, in order to augment their replication. To date, the role of the PI3K-Akt pathway in Baculovirus replication has not been defined. In the present study, we demonstrate that infection of Sf9 cells with Autographa californica multiple nucleopolyhedrovirus (AcMNPV) elevated cellular Akt phosphorylation at 1 h post-infection. The maximum Akt phosphorylation occurred at 6 h post-infection and remained unchanged until 18 h post-infection. The PI3K-specific inhibitor, LY294002, suppressed Akt phosphorylation in a dose-dependent manner, suggesting that AcMNPV-induced Akt phosphorylation is PI3K-dependent. The inhibition of PI3K-Akt activation by LY294002 significantly reduced the viral yield, including a reduction in budded viruses and occlusion bodies. The virus production was reduced only when the inhibitor was added within 24 h of infection, implying that activation of PI3K occurred early in infection. Correspondingly, both viral DNA replication and late (VP39) and very late (POLH) viral protein expression were impaired by LY294002 treatment; LY294002 had no effect on immediate-early (IE1) and early-late (GP64) protein expression. These results demonstrate that the PI3K-Akt pathway is required for efficient Baculovirus replication.

  9. Functional significance of the signal transduction pathways Akt and Erk in ovarian follicles: in vitro and in vivo studies in cattle and sheep

    PubMed Central

    Ryan, Kate E; Glister, Claire; Lonergan, Pat; Martin, Finian; Knight, Phil G; Evans, Alexander CO

    2008-01-01

    Background The intracellular signalling mechanisms that regulate ovarian follicle development are unclear; however, we have recently shown differences in the Akt and Erk signalling pathways in dominant compared to subordinate follicles. The aim of this study was to investigate the effects of inhibiting Akt and Erk phosphorylation on IGF- and gonadotropin- stimulated granulosa and theca cell function in vitro, and on follicle development in vivo. Methods Bovine granulosa and theca cells were cultured for six days and stimulated with FSH and/or IGF, or LH in combination with PD98059 (Erk inhibitor) and/or LY294002 (Akt inhibitor) and their effect on cell number and hormone secretion (estradiol, activin-A, inhibin-A, follistatin, progesterone and androstenedione) determined. In addition, ovarian follicles were treated in vivo with PD98059 and/or LY294002 in ewes on Day 3 of the cycle and follicles were recovered 48 hours later. Results We have shown that gonadotropin- and IGF-stimulated hormone production by granulosa and theca cells is reduced by treatment with PD98059 and LY294002 in vitro. Furthermore, treatment with PD98059 and LY294002 reduced follicle growth and oestradiol production in vivo. Conclusion These results demonstrate an important functional role for the Akt and Erk signalling pathways in follicle function, growth and development. PMID:19014654

  10. Reliable Signal Transduction

    NASA Astrophysics Data System (ADS)

    Wollman, Roy

    Stochasticity inherent to biochemical reactions (intrinsic noise) and variability in cellular states (extrinsic noise) degrade information transmitted through signaling networks. We analyzed the ability of temporal signal modulation - that is dynamics - to reduce noise-induced information loss. In the extracellular signal-regulated kinase (ERK), calcium (Ca(2 +)) , and nuclear factor kappa-B (NF- κB) pathways, response dynamics resulted in significantly greater information transmission capacities compared to nondynamic responses. Theoretical analysis demonstrated that signaling dynamics has a key role in overcoming extrinsic noise. Experimental measurements of information transmission in the ERK network under varying signal-to-noise levels confirmed our predictions and showed that signaling dynamics mitigate, and can potentially eliminate, extrinsic noise-induced information loss. By curbing the information-degrading effects of cell-to-cell variability, dynamic responses substantially increase the accuracy of biochemical signaling networks.

  11. Meeting Report: Teaching Signal Transduction

    ERIC Educational Resources Information Center

    Kramer, IJsbrand; Thomas, Geraint

    2006-01-01

    In July, 2005, the European Institute of Chemistry and Biology at the campus of the University of Bordeaux, France, hosted a focused week of seminars, workshops, and discussions around the theme of "teaching signal transduction." The purpose of the summer school was to offer both junior and senior university instructors a chance to reflect on the…

  12. Signal Transduction to the Permeability Transition Pore

    PubMed Central

    Rasola, Andrea; Sciacovelli, Marco; Pantic, Boris; Bernardi, Paolo

    2010-01-01

    The permeability transition pore (PTP) is an inner mitochondrial membrane channel that has been thoroughly characterized functionally, yet remains an elusive molecular entity. The best characterized PTP-regulatory component, cyclophilin (CyP) D, is a matrix protein that favors pore opening. CyP inhibitors, CyPD null animals, and in situ PTP readouts have established the role of PTP as an effector mechanism of cell death, and the growing definition of PTP signaling mechanisms. This review briefly covers the functional features of the PTP and the role played by its dysregulation in disease pathogenesis. Recent progress on PTP modulation by kinase/phosphatase signal transduction is discussed, with specific emphasis on hexokinase and on the Akt-ERK-GSK3 axis, which might modulate the PTP through CyPD phosphorylation. PMID:20153328

  13. Phosphorylation in halobacterial signal transduction.

    PubMed Central

    Rudolph, J; Tolliday, N; Schmitt, C; Schuster, S C; Oesterhelt, D

    1995-01-01

    Regulated phosphorylation of proteins has been shown to be a hallmark of signal transduction mechanisms in both Eubacteria and Eukarya. Here we demonstrate that phosphorylation and dephosphorylation are also the underlying mechanism of chemo- and phototactic signal transduction in Archaea, the third branch of the living world. Cloning and sequencing of the region upstream of the cheA gene, known to be required for chemo- and phototaxis in Halobacterium salinarium, has identified cheY and cheB analogs which appear to form part of an operon which also includes cheA and the following open reading frame of 585 nucleotides. The CheY and CheB proteins have 31.3 and 37.5% sequence identity compared with the known signal transduction proteins CheY and CheB from Escherichia coli, respectively. The biochemical activities of both CheA and CheY were investigated following their expression in E.coli, isolation and renaturation. Wild-type CheA could be phosphorylated in a time-dependent manner in the presence of [gamma-32P]ATP and Mg2+, whereas the mutant CheA(H44Q) remained unlabeled. Phosphorylated CheA was dephosphorylated rapidly by the addition of wild-type CheY. The mutant CheY(D53A) had no effect on phosphorylated CheA. The mechanism of chemo- and phototactic signal transduction in the Archaeon H.salinarium, therefore, is similar to the two-component signaling system known from chemotaxis in the eubacterium E.coli. Images PMID:7556066

  14. Protein Regulation in Signal Transduction.

    PubMed

    Lee, Michael J; Yaffe, Michael B

    2016-01-01

    SUMMARYCells must respond to a diverse, complex, and ever-changing mix of signals, using a fairly limited set of parts. Changes in protein level, protein localization, protein activity, and protein-protein interactions are critical aspects of signal transduction, allowing cells to respond highly specifically to a nearly limitless set of cues and also to vary the sensitivity, duration, and dynamics of the response. Signal-dependent changes in levels of gene expression and protein synthesis play an important role in regulation of protein levels, whereas posttranslational modifications of proteins regulate their degradation, localization, and functional interactions. Protein ubiquitylation, for example, can direct proteins to the proteasome for degradation or provide a signal that regulates their interactions and/or location within the cell. Similarly, protein phosphorylation by specific kinases is a key mechanism for augmenting protein activity and relaying signals to other proteins that possess domains that recognize the phosphorylated residues. PMID:27252361

  15. Meeting Report: Teaching Signal Transduction

    PubMed Central

    Kramer, IJsbrand; Thomas, Geraint

    2006-01-01

    In July, 2005, the European Institute of Chemistry and Biology at the campus of the University of Bordeaux, France, hosted a focused week of seminars, workshops, and discussions around the theme of “teaching signal transduction.” The purpose of the summer school was to offer both junior and senior university instructors a chance to reflect on the development and delivery of their teaching activities in this area. This was achieved by combining open seminars with restricted access workshops and discussion events. The results suggest ways in which systems biology, information and communication technology, Web-based investigations, and high standard illustrations might be more effectively and efficiently incorporated into modern cell biology courses. PMID:17012185

  16. 14-3-3ζ up-regulates hypoxia-inducible factor-1α in hepatocellular carcinoma via activation of PI3K/Akt/NF-кB signal transduction pathway

    PubMed Central

    Tang, Yufu; Lv, Pengfei; Sun, Zhongyi; Han, Lei; Luo, Bichao; Zhou, Wenping

    2015-01-01

    14-3-3ζ protein, a member of 14-3-3 family, plays important roles in multiple cellular processes. Our previous study showed that 14-3-3ζ could bind to regulate the expression of hypoxia-inducible factor-1α (HIF-1α), which is induced by hypoxia and a crucial factor for induction of tumor metastasis. Moreover, we also have confirmed the response of 14-3-3ζ to hypoxia in our unpublished data as well. Thus, in the present study, we attempted to reveal that whether the regulation effect of 14-3-3ζ on HIF-1α functioned in a similar pattern as hypoxia. Stable regulation of 14-3-3ζ in human HCC cell line SMMC-772 and HCC-LM3 was achieved. The regulation of 14-3-3ζ on HIF-1α mRNA transcription was evaluated by luciferase activity assay and quantitative real-time PCR (qPCR). The effect of 14-3-3ζ on the production of HIF-1α and pathways determining HIF-1α’s response to hypoxia was assessed using western blotting assay. Our results showed that regulation of 14-3-3ζ expression influenced the activity of HIF-1α, phosphatidyl inositol 3-kinase (PI3K), Akt, extracellular signal-regulated kinase 1/2 (ERK1/2), and nuclear factor kappa B (NF-кB). Blocking of these pathways using indicated inhibitors revealed that 14-3-3ζ enhanced the production of HIF-1α via the activation of PI3K/Akt/NF-кB pathway, which was identical to hypoxia induced HIF-1α expression. For the first time, our study described the key role of 14-3-3ζ in the HIF-1α production in HCC cells. And the molecule exerted its function on HIF-1α both by directly binding to it and via PI3K/Akt/NF-кB signal transduction pathway. PMID:26884855

  17. Gravitational Effects on Signal Transduction

    NASA Technical Reports Server (NTRS)

    Sytkowski, Arthur J.

    1999-01-01

    An understanding of the mechanisms by which individual cells perceive gravity and how these cells transduce and respond to gravitational stimuli is critical for the development of long-term manned space flight experiments. We now propose to use a well-characterized model erythroid cell system and to investigate gravitational perturbations of its erythropoietin (Epo) signaling pathway and gene regulation. Cells will be grown at 1-G and in simulated microgravity in the NASA Rotating Wall Vessel bioreactor (RWV). Cell growth and differentiation, the Epo-receptor, the protein kinase C pathway to the c-myc gene, and the protein phosphatase pathway to the c-myb gene will be studied and evaluated as reporters of gravitational stimuli. The results of these experiments will have impact on the problems of 1) gravitational sensing by individual cells, and 2) the anemia of space flight. This ground-based study also will serve as a Space Station Development Study in gravitational effects on intracellular signal transduction.

  18. SENTRA, a database of signal transduction proteins.

    SciTech Connect

    D'Souza, M.; Romine, M. F.; Maltsev, N.; Mathematics and Computer Science; PNNL

    2000-01-01

    SENTRA, available via URL http://wit.mcs.anl.gov/WIT2/Sentra/, is a database of proteins associated with microbial signal transduction. The database currently includes the classical two-component signal transduction pathway proteins and methyl-accepting chemotaxis proteins, but will be expanded to also include other classes of signal transduction systems that are modulated by phosphorylation or methylation reactions. Although the majority of database entries are from prokaryotic systems, eukaroytic proteins with bacterial-like signal transduction domains are also included. Currently SENTRA contains signal transduction proteins in 34 complete and almost completely sequenced prokaryotic genomes, as well as sequences from 243 organisms available in public databases (SWISS-PROT and EMBL). The analysis was carried out within the framework of the WIT2 system, which is designed and implemented to support genetic sequence analysis and comparative analysis of sequenced genomes.

  19. Sentra, a database of signal transduction proteins.

    SciTech Connect

    Maltsev, N.; Marland, E.; Yu, G. X.; Bhatnagar, S.; Lusk, R.; Mathematics and Computer Science

    2002-01-01

    Sentra (http://www-wit.mcs.anl.gov/sentra) is a database of signal transduction proteins with the emphasis on microbial signal transduction. The database was updated to include classes of signal transduction systems modulated by either phosphorylation or methylation reactions such as PAS proteins and serine/threonine kinases, as well as the classical two-component histidine kinases and methyl-accepting chemotaxis proteins. Currently, Sentra contains signal transduction proteins from 43 completely sequenced prokaryotic genomes as well as sequences from SWISS-PROT and TrEMBL. Signal transduction proteins are annotated with information describing conserved domains, paralogous and orthologous sequences, and conserved chromosomal gene clusters. The newly developed user interface supports flexible search capabilities and extensive visualization of the data.

  20. Meeting report: Signal transduction meets systems biology

    PubMed Central

    2012-01-01

    In the 21st century, systems-wide analyses of biological processes are getting more and more realistic. Especially for the in depth analysis of signal transduction pathways and networks, various approaches of systems biology are now successfully used. The EU FP7 large integrated project SYBILLA (Systems Biology of T-cell Activation in Health and Disease) coordinates such an endeavor. By using a combination of experimental data sets and computational modelling, the consortium strives for gaining a detailed and mechanistic understanding of signal transduction processes that govern T-cell activation. In order to foster the interaction between systems biologists and experimentally working groups, SYBILLA co-organized the 15th meeting “Signal Transduction: Receptors, Mediators and Genes” together with the Signal Transduction Society (STS). Thus, the annual STS conference, held from November 7 to 9, 2011 in Weimar, Germany, provided an interdisciplinary forum for research on signal transduction with a major focus on systems biology addressing signalling events in T-cells. Here we report on a selection of ongoing projects of SYBILLA and how they were discussed at this interdisciplinary conference. PMID:22546078

  1. Fluorescence polarization assays in signal transduction discovery.

    PubMed

    Sportsman, J Richard; Daijo, Janet; Gaudet, Elizabeth A

    2003-05-01

    Fluorescence polarization (FP) has become widely employed for high throughput screening used in pharmaceutical drug discovery. Assays of important signal transduction targets are now adapted to FP. In this review we examine assays for cyclic adenosine monophosphate, phosphodiesterases, and protein kinases and phosphatases using FP competitive immunoassays and a direct enzymatic method called IMAP. PMID:12678698

  2. Signal transduction mechanisms in plants: an overview

    NASA Technical Reports Server (NTRS)

    Clark, G. B.; Thompson, G. Jr; Roux, S. J.

    2001-01-01

    This article provides an overview on recent advances in some of the basic signalling mechanisms that participate in a wide variety of stimulus-response pathways. The mechanisms include calcium-based signalling, G-protein-mediated-signalling and signalling involving inositol phospholipids, with discussion on the role of protein kinases and phosphatases interspersed. As a further defining feature, the article highlights recent exciting findings on three extracellular components that have not been given coverage in previous reviews of signal transduction in plants, extracellular calmodulin, extracellular ATP, and integrin-like receptors, all of which affect plant growth and development.

  3. The ethylene signal transduction pathway in Arabidopsis

    NASA Technical Reports Server (NTRS)

    Kieber, J. J.; Evans, M. L. (Principal Investigator)

    1997-01-01

    The gaseous hormone ethylene is an important regulator of plant growth and development. Using a simple response of etiolated seedlings to ethylene as a genetic screen, genes involved in ethylene signal transduction have been identified in Arabidopsis. Analysis of two of these genes that have been cloned reveals that ethylene signalling involves a combination of a protein (ETR1) with similarity to bacterial histidine kinases and a protein (CTR1) with similarity to Raf-1, a protein kinase involved in multiple signalling cascades in eukaryotic cells. Several lines of investigation provide compelling evidence that ETR1 encodes an ethylene receptor. For the first time there is a glimpse of the molecular circuitry underlying the signal transduction pathway for a plant hormone.

  4. Gravitational Effects on Signal Transduction

    NASA Technical Reports Server (NTRS)

    Sytkowski, Arthur J.

    1999-01-01

    The purpose of this study was to investigate in ground-based experiments, the effect of microgravity on in vitro erythroid differentiation triggered by the hematopoietic growth factor erythropoietin (Epo) and to begin to determine whether this is associated with the anemia of space flight. We chose to use a model cell culture system with which we have had a long and successful experience. These cells, designated Rauscher murine erythroleukemia, grow independently in suspension culture. We first compared the growth rate of Rauscher cells under conditions of simulated microgravity with that of cells grown at 1XG in standard tissue culture flasks. Therefore, since there were fewer cells in the RWV at each specified time, glucose consumption per cell was increased in simulated microgravity. We next began to study the effect of simulated microgravity on erythropoietin induced differentiation of these cells. In another experiment, we allow the cells to grown in flasks or in the RWV for 24 hours prior to the addition of Epo. We initiated studies of c-myb, a proto-oncogene the down-regulation of which is necessary for erythroid differentiation. These preliminary results suggest that simulated microgravity interferes with the signal to c-myb. This may be part of the mechanism that blocks differentiation. A flight experiment is planned within the next 18- 24 months.

  5. Studying Cellular Signal Transduction with OMIC Technologies.

    PubMed

    Landry, Benjamin D; Clarke, David C; Lee, Michael J

    2015-10-23

    In the gulf between genotype and phenotype exists proteins and, in particular, protein signal transduction systems. These systems use a relatively limited parts list to respond to a much longer list of extracellular, environmental, and/or mechanical cues with rapidity and specificity. Most signaling networks function in a highly non-linear and often contextual manner. Furthermore, these processes occur dynamically across space and time. Because of these complexities, systems and "OMIC" approaches are essential for the study of signal transduction. One challenge in using OMIC-scale approaches to study signaling is that the "signal" can take different forms in different situations. Signals are encoded in diverse ways such as protein-protein interactions, enzyme activities, localizations, or post-translational modifications to proteins. Furthermore, in some cases, signals may be encoded only in the dynamics, duration, or rates of change of these features. Accordingly, systems-level analyses of signaling may need to integrate multiple experimental and/or computational approaches. As the field has progressed, the non-triviality of integrating experimental and computational analyses has become apparent. Successful use of OMIC methods to study signaling will require the "right" experiments and the "right" modeling approaches, and it is critical to consider both in the design phase of the project. In this review, we discuss common OMIC and modeling approaches for studying signaling, emphasizing the philosophical and practical considerations for effectively merging these two types of approaches to maximize the probability of obtaining reliable and novel insights into signaling biology. PMID:26244521

  6. Advances in Targeting Signal Transduction Pathways

    PubMed Central

    McCubrey, James A.; Steelman, Linda S.; Chappell, William H.; Sun, Lin; Davis, Nicole M.; Abrams, Stephen L.; Franklin, Richard A.; Cocco, Lucio; Evangelisti, Camilla; Chiarini, Francesca; Martelli, Alberto M.; Libra, Massimo; Candido, Saverio; Ligresti, Giovanni; Malaponte, Grazia; Mazzarino, Maria C.; Fagone, Paolo; Donia, Marco; Nicoletti, Ferdinando; Polesel, Jerry; Talamini, Renato; Bäsecke, Jörg; Mijatovic, Sanja; Maksimovic-Ivanic, Danijela; Milella, Michele; Tafuri, Agostino; Dulińska-Litewka, Joanna; Laidler, Piotr; D'Assoro, Antonio B.; Drobot, Lyudmyla; Umezawa, Kazuo; Montalto, Giuseppe; Cervello, Melchiorre; Demidenko, Zoya N.

    2012-01-01

    Over the past few years, significant advances have occurred in both our understanding of the complexity of signal transduction pathways as well as the isolation of specific inhibitors which target key components in those pathways. Furthermore critical information is being accrued regarding how genetic mutations can affect the sensitivity of various types of patients to targeted therapy. Finally, genetic mechanisms responsible for the development of resistance after targeted therapy are being discovered which may allow the creation of alternative therapies to overcome resistance. This review will discuss some of the highlights over the past few years on the roles of key signaling pathways in various diseases, the targeting of signal transduction pathways and the genetic mechanisms governing sensitivity and resistance to targeted therapies. PMID:23455493

  7. Targeting Signaling Transduction Pathways in Bladder Cancer.

    PubMed

    Abbosh, Phillip H; McConkey, David J; Plimack, Elizabeth R

    2015-12-01

    Systemic therapy for urothelial carcinoma (UC) of the bladder has largely revolved around cytotoxic chemotherapy regimens. However, several recent clinical trials have explored the roles of targeted therapies which specifically inhibit signal transduction pathways. Simultaneously, a rationale for such therapies has come to the forefront of management of this disease because an overabundance of signaling pathways are genetically deranged as a result of point mutation or copy number alteration (CNA) as identified by several recent next generation sequencing (NGS) studies. Importantly, these derangements are found in all stages of disease, and therefore targeted therapies hold promise as a next step in the evolution of the medical management of both localized and metastatic UCC. We review the rationale for and progress in studying inhibition of signal transduction as a means of treatment of UCC. PMID:26472299

  8. Signal transduction in T lymphocytes in microgravity

    NASA Technical Reports Server (NTRS)

    Cogoli, A.

    1997-01-01

    More than 120 experiments conducted in space in the last 15 years have shown that dramatic changes are occurring in several types of single cells during their exposure to microgravity. One focus of today's research on cells in space is on signal transduction, especially those steps involving the cytoskeleton and cell-cell interactions. Signal transduction is often altered in microgravity as well as in hypergravity. This leads to changes in cell proliferation, genetic expression and differentiation. Interesting examples are leukocytes, HeLa cells, epidermoid cells and osteoblastic cells. Signalling pathways were studied in T lymphocytes in microgravity by several investigators after the discovery that mitogenic activation in vitro is virtually nil at 0g. T cells are a good model to study signal transduction because three extracellular signals (mitogen, IL-1 and IL-2) are required for full activation, and two classical pathways (via proteins G and PKC) are activated within the cell. In addition, low molecular weight GTP-binding proteins (Ras and Rap) are interacting with the cytoskeleton. The data at 0g support the notion that the expression of IL-2 receptor is inhibited at 0g, while mitogen binding and the transmission of IL-1 by accessory cells occur normally. In addition, alterations of the cytoskeleton suggest that the interaction with Rap proteins is disturbed. Data obtained with phorbol esters indicate that the function of PKC is changed in microgravity. Similar conclusions are drawn from the results with epidermoid cells A431.

  9. Akt signaling dynamics in individual cells

    PubMed Central

    Gross, Sean M.; Rotwein, Peter

    2015-01-01

    ABSTRACT The protein kinase Akt (for which there are three isoforms) is a key intracellular mediator of many biological processes, yet knowledge of Akt signaling dynamics is limited. Here, we have constructed a fluorescent reporter molecule in a lentiviral delivery system to assess Akt kinase activity at the single cell level. The reporter, a fusion between a modified FoxO1 transcription factor and clover, a green fluorescent protein, rapidly translocates from the nucleus to the cytoplasm in response to Akt stimulation. Because of its long half-life and the intensity of clover fluorescence, the sensor provides a robust readout that can be tracked for days under a range of biological conditions. Using this reporter, we find that stimulation of Akt activity by IGF-I is encoded into stable and reproducible analog responses at the population level, but that single cell signaling outcomes are variable. This reporter, which provides a simple and dynamic measure of Akt activity, should be compatible with many cell types and experimental platforms, and thus opens the door to new insights into how Akt regulates its biological responses. PMID:26040286

  10. Automated modelling of signal transduction networks

    PubMed Central

    2002-01-01

    Background Intracellular signal transduction is achieved by networks of proteins and small molecules that transmit information from the cell surface to the nucleus, where they ultimately effect transcriptional changes. Understanding the mechanisms cells use to accomplish this important process requires a detailed molecular description of the networks involved. Results We have developed a computational approach for generating static models of signal transduction networks which utilizes protein-interaction maps generated from large-scale two-hybrid screens and expression profiles from DNA microarrays. Networks are determined entirely by integrating protein-protein interaction data with microarray expression data, without prior knowledge of any pathway intermediates. In effect, this is equivalent to extracting subnetworks of the protein interaction dataset whose members have the most correlated expression profiles. Conclusion We show that our technique accurately reconstructs MAP Kinase signaling networks in Saccharomyces cerevisiae. This approach should enhance our ability to model signaling networks and to discover new components of known networks. More generally, it provides a method for synthesizing molecular data, either individual transcript abundance measurements or pairwise protein interactions, into higher level structures, such as pathways and networks. PMID:12413400

  11. Calcium and signal transduction in plants

    NASA Technical Reports Server (NTRS)

    Poovaiah, B. W.; Reddy, A. S.

    1993-01-01

    Environmental and hormonal signals control diverse physiological processes in plants. The mechanisms by which plant cells perceive and transduce these signals are poorly understood. Understanding biochemical and molecular events involved in signal transduction pathways has become one of the most active areas of plant research. Research during the last 15 years has established that Ca2+ acts as a messenger in transducing external signals. The evidence in support of Ca2+ as a messenger is unequivocal and fulfills all the requirements of a messenger. The role of Ca2+ becomes even more important because it is the only messenger known so far in plants. Since our last review on the Ca2+ messenger system in 1987, there has been tremendous progress in elucidating various aspects of Ca(2+) -signaling pathways in plants. These include demonstration of signal-induced changes in cytosolic Ca2+, calmodulin and calmodulin-like proteins, identification of different Ca2+ channels, characterization of Ca(2+) -dependent protein kinases (CDPKs) both at the biochemical and molecular levels, evidence for the presence of calmodulin-dependent protein kinases, and increased evidence in support of the role of inositol phospholipids in the Ca(2+) -signaling system. Despite the progress in Ca2+ research in plants, it is still in its infancy and much more needs to be done to understand the precise mechanisms by which Ca2+ regulates a wide variety of physiological processes. The purpose of this review is to summarize some of these recent developments in Ca2+ research as it relates to signal transduction in plants.

  12. Activity Dependent Signal Transduction in Skeletal Muscle

    NASA Technical Reports Server (NTRS)

    Hamilton, Susan L.

    1999-01-01

    The overall goals of this project are: 1) to define the initial signal transduction events whereby the removal of gravitational load from antigravity muscles, such as the soleus, triggers muscle atrophy, and 2) to develop countermeasures to prevent this from happening. Our rationale for this approach is that, if countermeasures can be developed to regulate these early events, we could avoid having to deal with the multiple cascades of events that occur downstream from the initial event. One of our major findings is that hind limb suspension causes an early and sustained increase in intracellular Ca(2+) concentration ([Ca (2+)](sub i)). In most cells the consequences of changes in ([Ca (2+)](sub i))depend on the amplitude, frequency and duration of the Ca(2+) signal and on other factors in the intracellular environment. We propose that muscle remodeling in microgravity represents a change in the balance among several CA(2+) regulated signal transduction pathways, in particular those involving the transcription factors NFAT and NFkB and the pro-apoptotic protein BAD. Other Ca(2+) sensitive pathways involving PKC, ras, rac, and CaM kinase II may also contribute to muscle remodeling.

  13. Targeting prostate cancer based on signal transduction and cell cycle pathways

    PubMed Central

    Lee, John T.; Lehmann, Brian D.; Terrian, David M.; Chappell, William H.; Stivala, Franca; Libra, Massimo; Martelli, Alberto M.; Steelman, Linda S.

    2008-01-01

    Prostate cancer remains a leading cause of death in men despite increased capacity to diagnose at earlier stages. After prostate cancer has become hormone independent, which often occurs after hormonal ablation therapies, it is difficult to effectively treat. Prostate cancer may arise from mutations and dysregulation of various genes involved in regulation signal transduction (e.g., PTEN, Akt, etc.,) and the cell cycle (e.g., p53, p21Cip1, p27Kip1, Rb, etc.,). This review focuses on the aberrant interactions of signal transduction and cell cycle genes products and how they can contribute to prostate cancer and alter therapeutic effectiveness. PMID:18594202

  14. Striatal Signal Transduction and Drug Addiction

    PubMed Central

    Philibin, Scott D.; Hernandez, Adan; Self, David W.; Bibb, James A.

    2011-01-01

    Drug addiction is a severe neuropsychiatric disorder characterized by loss of control over motivated behavior. The need for effective treatments mandates a greater understanding of the causes and identification of new therapeutic targets for drug development. Drugs of abuse subjugate normal reward-related behavior to uncontrollable drug-seeking and -taking. Contributions of brain reward circuitry are being mapped with increasing precision. The role of synaptic plasticity in addiction and underlying molecular mechanisms contributing to the formation of the addicted state are being delineated. Thus we may now consider the role of striatal signal transduction in addiction from a more integrative neurobiological perspective. Drugs of abuse alter dopaminergic and glutamatergic neurotransmission in medium spiny neurons of the striatum. Dopamine receptors important for reward serve as principle targets of drugs abuse, which interact with glutamate receptor signaling critical for reward learning. Complex networks of intracellular signal transduction mechanisms underlying these receptors are strongly stimulated by addictive drugs. Through these mechanisms, repeated drug exposure alters functional and structural neuroplasticity, resulting in transition to the addicted biological state and behavioral outcomes that typify addiction. Ca2+ and cAMP represent key second messengers that initiate signaling cascades, which regulate synaptic strength and neuronal excitability. Protein phosphorylation and dephosphorylation are fundamental mechanisms underlying synaptic plasticity that are dysregulated by drugs of abuse. Increased understanding of the regulatory mechanisms by which protein kinases and phosphatases exert their effects during normal reward learning and the addiction process may lead to novel targets and pharmacotherapeutics with increased efficacy in promoting abstinence and decreased side effects, such as interference with natural reward, for drug addiction. PMID

  15. Signal transduction by the growth hormone receptor

    SciTech Connect

    Waters, M.J.; Rowlinson, S.W.; Clarkson, R.W.

    1994-12-31

    It has been proposed that dimerization of identical receptor subunits by growth hormone (GH) is the mechanism of signal transduction across the cell membrane. We present here data with analogs of porcine GH (pGH), with GH receptors (GHR) mutated in the dimerization domain and with monoclonal antibodies to the GHR which indicate that dimerization is necessary but not sufficient for transduction. We also report nuclear uptake of GH both in vivo and in vitro, along with nuclear localization of the receptor and GH-binding protein (GHBP). This suggests that GH acts directly at the nucleus, and one possible target for this action is a rapid increase in transcription of C/EBP delta seen in 3T3-F442A cells in response to GH. This tyrosine kinase-dependent event may be an archetype for induction of other immediate early gene transcription factors which then interact to determine the programming of the subsequent transcriptional response to GH. 29 refs., 1 fig., 1 tab.

  16. The Membrane and Lipids as Integral Participants in Signal Transduction: Lipid Signal Transduction for the Non-Lipid Biochemist

    ERIC Educational Resources Information Center

    Eyster, Kathleen M.

    2007-01-01

    Reviews of signal transduction have often focused on the cascades of protein kinases and protein phosphatases and their cytoplasmic substrates that become activated in response to extracellular signals. Lipids, lipid kinases, and lipid phosphatases have not received the same amount of attention as proteins in studies of signal transduction.…

  17. Targeting the Raft-Associated Akt Signaling in Hepatocellular Carcinoma

    PubMed Central

    Liu, Yuan; Lv, Ji-Yun; Shi, Jian-Fei; Yang, Mei; Liu, Shu-Hong; Li, Zhi-Wei; Wang, Hong-Bo; Zhang, Shao-Geng; Liu, Zhen-Wen; Ding, Jin-Biao; Xu, Dong-Ping; Zhao, Jing-Min

    2014-01-01

    Caveolin-1 and flotillin-1 are considered as markers of lipid rafts which can be regarded as sorting platforms for targeted transport of transmembrane proteins and are involved in fundamental cellular events such as signal transduction, cell adhesion, lipid/protein sorting, and human cancer. We addressed caveolin-1 and flotillin-1 expression in 90 human hepatocellular carcinoma (HCC) and adjacent noncancerous tissues (ANT) samples by SDS-PAGE and immunoblotting with specific antibodies. Significant caveolin-1 and flotillin-1 overexpression was found in HCC tissues compared to ANT and was confirmed by immunohistochemistry. Raft-associated Akt signaling pathway components involved in the regulation of cell survival were altered by western blotting in HCC microdomain-enriched subcellular fractions purified from paired HCC and ANT samples. Our results demonstrated that the activity of raft-associated but not total membrane Akt determines its cellular functions. Lipid rafts differ in different types of tissues, which allows for the possibility of tissue-type-specific targeting for cell survival. PMID:25243186

  18. Signal transduction by guanine nucleotide binding proteins.

    PubMed

    Spiegel, A M

    1987-01-01

    High affinity binding of guanine nucleotides and the ability to hydrolyze bound GTP to GDP are characteristics of an extended family of intracellular proteins. Subsets of this family include cytosolic initiation and elongation factors involved in protein synthesis, and cytoskeletal proteins such as tubulin (Hughes, S.M. (1983) FEBS Lett. 164, 1-8). A distinct subset of guanine nucleotide binding proteins is membrane-associated; members of this subset include the ras gene products (Ellis, R.W. et al. (1981) Nature 292, 506-511) and the heterotrimeric G-proteins (also termed N-proteins) (Gilman, A.G. (1984) Cell 36, 577-579). Substantial evidence indicates that G-proteins act as signal transducers by coupling receptors (R) to effectors (E). A similar function has been suggested but not proven for the ras gene products. Known G-proteins include Gs and Gi, the G-proteins associated with stimulation and inhibition, respectively, of adenylate cyclase; transducin (TD), the G-protein coupling rhodopsin to cGMP phosphodiesterase in rod photoreceptors (Bitensky, M.W. et al. (1981) Curr. Top. Membr. Transp. 15, 237-271; Stryer, L. (1986) Annu. Rev. Neurosci. 9, 87-119), and Go, a G-protein of unknown function that is highly abundant in brain (Sternweis, P.C. and Robishaw, J.D. (1984) J. Biol. Chem. 259, 13806-13813; Neer, E.J. et al. (1984) J. Biol. Chem. 259, 14222-14229). G-proteins also participate in other signal transduction pathways, notably that involving phosphoinositide breakdown. In this review, I highlight recent progress in our understanding of the structure, function, and diversity of G-proteins. PMID:2435586

  19. EDITORIAL: Special section on signal transduction Special section on signal transduction

    NASA Astrophysics Data System (ADS)

    Shvartsman, Stanislav

    2012-08-01

    This special section of Physical Biology focuses on multiple aspects of signal transduction, broadly defined as the study of the mechanisms by which cells communicate with their environment. Mechanisms of cell communication involve detection of incoming signals, which can be chemical, mechanical or electromagnetic, relaying these signals to intracellular processes, such as cytoskeletal networks or gene expression systems, and, ultimately, converting these signals to responses such as cell differentiation or death. Given the multiscale nature of signal transduction systems, they must be studied at multiple levels, from the identities and structures of molecules comprising signal detection and interpretation networks, to the systems-level properties of these networks. The 11 papers in this special section illustrate some of the most exciting aspects of signal transduction research. The first two papers, by Marie-Anne Félix [1] and by Efrat Oron and Natalia Ivanova [2], focus on cell-cell interactions in developing tissues, using vulval patterning in worm and cell fate specification in mammalian embryos as prime examples of emergent cell behaviors. Next come two papers from the groups of Julio Saez-Rodriguez [3] and Kevin Janes [4]. These papers discuss how the causal relationships between multiple components of signaling systems can be inferred using multivariable statistical analysis of empirical data. An authoritative review by Zarnitsyna and Zhu [5] presents a detailed discussion of the sequence of signaling events involved in T-cell triggering. Once the structure and components of the signaling systems are determined, they can be modeled using approaches that have been successful in other physical sciences. As two examples of such approaches, reviews by Rubinstein [6] and Kholodenko [7], present reaction-diffusion models of cell polarization and thermodynamics-based models of gene regulation. An important class of models takes the form of enzymatic networks

  20. COMP-angiopoietin 1 increases proliferation, differentiation, and migration of stem-like cells through Tie-2-mediated activation of p38 MAPK and PI3K/Akt signal transduction pathways

    SciTech Connect

    Kook, Sung-Ho; Lim, Shin-Saeng; Cho, Eui-Sic; Lee, Young-Hoon; Han, Seong-Kyu; Lee, Kyung-Yeol; Kwon, Jungkee; Hwang, Jae-Won; Bae, Cheol-Hyeon; Seo, Young-Kwon; Lee, Jeong-Chae

    2014-12-12

    Highlights: • COMP-Ang1 induces Tie-2 activation in BMMSCs, but not in primary osteoblasts. • Tie-2 knockdown inhibits COMP-Ang1-stimulated proliferation and osteoblastogenesis. • Tie-2 knockdown prevents COMP-Ang1-induced activation of PI3K/Akt and p38 MAPK. • COMP-Ang1 induces migration of cells via activation of PI3K/Akt and CXCR4 pathways. • COMP-Ang1 stimulates in vivo migration of PDLSCs into a calvarial defect site of rats. - Abstract: Recombinant COMP-Ang1, a chimera of angiopoietin-1 (Ang1) and a short coiled-coil domain of cartilage oligomeric matrix protein (COMP), is under consideration as a therapeutic agent capable of inducing the homing of cells with increased angiogenesis. However, the potentials of COMP-Ang1 to stimulate migration of mesenchymal stem cells (MSCs) and the associated mechanisms are not completely understood. We examined the potential of COMP-Ang1 on bone marrow (BM)-MSCs, human periodontal ligament stem cells (PDLSCs), and calvarial osteoblasts. COMP-Ang1 augmented Tie-2 induction at protein and mRNA levels and increased proliferation and expression of runt-related transcription factor 2 (Runx2), osterix, and CXCR4 in BMMSCs, but not in osteoblasts. The COMP-Ang1-mediated increases were inhibited by Tie-2 knockdown and by treating inhibitors of phosphoinositide 3-kinase (PI3K), LY294002, or p38 mitogen-activated protein kinase (MAPK), SB203580. Phosphorylation of p38 MAPK and Akt was prevented by siRNA-mediated silencing of Tie-2. COMP-Ang1 also induced in vitro migration of BMMSCs and PDLSCs. The induced migration was suppressed by Tie-2 knockdown and by CXCR4-specific peptide antagonist or LY294002, but not by SB203580. Furthermore, COMP-Ang1 stimulated the migration of PDLSCs into calvarial defect site of rats. Collectively, our results demonstrate that COMP-Ang1-stimulated proliferation, differentiation, and migration of progenitor cells may involve the Tie-2-mediated activation of p38 MAPK and PI3K/Akt pathways.

  1. Angiotensin II Signaling in Human Preadipose Cells: Participation of ERK1,2-Dependent Modulation of Akt

    PubMed Central

    Dünner, Natalia; Quezada, Carolina; Berndt, F. Andrés; Cánovas, José; Rojas, Cecilia V.

    2013-01-01

    The renin-angiotensin system expressed in adipose tissue has been implicated in the modulation of adipocyte formation, glucose metabolism, triglyceride accumulation, lipolysis, and the onset of the adverse metabolic consequences of obesity. As we investigated angiotensin II signal transduction mechanisms in human preadipose cells, an interplay of extracellular-signal-regulated kinases 1 and 2 (ERK1,2) and Akt/PKB became evident. Angiotensin II caused attenuation of phosphorylated Akt (p-Akt), at serine 473; the p-Akt/Akt ratio decreased to 0.5±0.2-fold the control value without angiotensin II (p<0.001). Here we report that the reduction of phosphorylated Akt associates with ERK1,2 activities. In the absence of angiotensin II, inhibition of ERK1,2 activation with U0126 or PD98059 resulted in a 2.1±0.5 (p<0.001) and 1.4±0.2-fold (p<0.05) increase in the p-Akt/Akt ratio, respectively. In addition, partial knockdown of ERK1 protein expression by the short hairpin RNA technique also raised phosphorylated Akt in these cells (the p-Akt/Akt ratio was 1.5±0.1-fold the corresponding control; p<0.05). Furthermore, inhibition of ERK1,2 activation with U0126 prevented the reduction of p-Akt/Akt by angiotensin II. An analogous effect was found on the phosphorylation status of Akt downstream effectors, the forkhead box (Fox) proteins O1 and O4. Altogether, these results indicate that angiotensin II signaling in human preadipose cells involves an ERK1,2-dependent attenuation of Akt activity, whose impact on the biological functions under its regulation is not fully understood. PMID:24098385

  2. Engineering key components in a synthetic eukaryotic signal transduction pathway

    PubMed Central

    Antunes, Mauricio S; Morey, Kevin J; Tewari-Singh, Neera; Bowen, Tessa A; Smith, J Jeff; Webb, Colleen T; Hellinga, Homme W; Medford, June I

    2009-01-01

    Signal transduction underlies how living organisms detect and respond to stimuli. A goal of synthetic biology is to rewire natural signal transduction systems. Bacteria, yeast, and plants sense environmental aspects through conserved histidine kinase (HK) signal transduction systems. HK protein components are typically comprised of multiple, relatively modular, and conserved domains. Phosphate transfer between these components may exhibit considerable cross talk between the otherwise apparently linear pathways, thereby establishing networks that integrate multiple signals. We show that sequence conservation and cross talk can extend across kingdoms and can be exploited to produce a synthetic plant signal transduction system. In response to HK cross talk, heterologously expressed bacterial response regulators, PhoB and OmpR, translocate to the nucleus on HK activation. Using this discovery, combined with modification of PhoB (PhoB-VP64), we produced a key component of a eukaryotic synthetic signal transduction pathway. In response to exogenous cytokinin, PhoB-VP64 translocates to the nucleus, binds a synthetic PlantPho promoter, and activates gene expression. These results show that conserved-signaling components can be used across kingdoms and adapted to produce synthetic eukaryotic signal transduction pathways. PMID:19455134

  3. Effect of bisphenol A on SOCS-3 and insulin signaling transduction in 3T3-L1 adipocytes.

    PubMed

    Dai, Yue-E; Chen, Wei; Qi, Humin; Liu, Qian-Qi

    2016-07-01

    The aim of the present study was to investigate whether environmental endocrine disrupting chemical, bisphenol A (BPA), affects secretion of suppressor of cytokine signaling 3 (SOCS-3) and insulin signaling transduction in 3T3-L1 adipocytes. 3T3-L1 adipocytes were treated for 0, 2, 6, 12 and 24 h with BPA at 80 µM in serum‑deprived medium. Reverse transcription-quantitative polymerase chain reaction and western blotting were performed to detect the mRNA expression levels of SOCS‑3 and protein expression levels of SOCS‑3, insulin receptor substrate 1 (IRS‑1), phosphorylated (p)‑IRS‑1, Akt and p‑Akt. The levels of p‑IRS‑1, Akt and p‑Akt in cultures treated for 6 h with BPA were also analyzed by immunofluorescence. The SOCS‑3 mRNA and protein expression levels were decreased in the 6, 12 and 24 h groups. The levels of p‑IRS‑1 and p‑Akt protein were markedly downregulated, while the level of IRS‑1 and Akt protein remained unaltered among these groups, which was consistent with the results observed using immunofluorescence. BPA may inhibit insulin signal transduction and result in the occurrence of insulin resistance via promoting the expression of SOCS-3. PMID:27176707

  4. Modelling protein functional domains in signal transduction using Maude

    NASA Technical Reports Server (NTRS)

    Sriram, M. G.

    2003-01-01

    Modelling of protein-protein interactions in signal transduction is receiving increased attention in computational biology. This paper describes recent research in the application of Maude, a symbolic language founded on rewriting logic, to the modelling of functional domains within signalling proteins. Protein functional domains (PFDs) are a critical focus of modern signal transduction research. In general, Maude models can simulate biological signalling networks and produce specific testable hypotheses at various levels of abstraction. Developing symbolic models of signalling proteins containing functional domains is important because of the potential to generate analyses of complex signalling networks based on structure-function relationships.

  5. Signal Transduction in Histidine Kinases: Insights from New Structures

    PubMed Central

    Bhate, Manasi P.; Molnar, Kathleen S.; Goulian, Mark; DeGrado, William F.

    2015-01-01

    Histidine kinases (HKs) are major players in bacterial signaling. There has been an explosion of new HK crystal structures in the last five years. We globally analyze the structures of HKs to yield insights into the mechanisms by which signals are transmitted to and across protein structures in this family. We interpret known enzymological data in the context of new structural data to show how asymmetry across the dimer interface is a key feature of signal transduction in HKs, and discuss how different HK domains undergo asymmetric-to-symmetric transitions during signal transduction and catalysis. A thermodynamic framework for signaling that encompasses these various properties is presented and the consequences of weak thermodynamic coupling are discussed. The synthesis of observations from enzymology, structural biology, protein engineering and thermodynamics paves the way for a deeper molecular understanding of histidine kinase signal transduction. PMID:25982528

  6. Mechanism and evolution of cytosolic Hedgehog signal transduction

    PubMed Central

    Wilson, Christopher W.; Chuang, Pao-Tien

    2010-01-01

    Hedgehog (Hh) signaling is required for embryonic patterning and postnatal physiology in invertebrates and vertebrates. With the revelation that the primary cilium is crucial for mammalian Hh signaling, the prevailing view that Hh signal transduction mechanisms are conserved across species has been challenged. However, more recent progress on elucidating the function of core Hh pathway cytosolic regulators in Drosophila, zebrafish and mice has confirmed that the essential logic of Hh transduction is similar between species. Here, we review Hh signaling events at the membrane and in the cytosol, and focus on parallel and divergent functions of cytosolic Hh regulators in Drosophila and mammals. PMID:20530542

  7. Akt signaling in platelets and thrombosis

    PubMed Central

    Woulfe, Donna S

    2010-01-01

    Akt is a Ser–Thr kinase with pleiotropic effects on cell survival, growth and metabolism. Recent evidence from gene-deletion studies in mice, and analysis of human platelets treated with Akt inhibitors, suggest that Akt regulates platelet activation, with potential consequences for thrombosis. Akt activation is regulated by the level of phosphoinositide 3-phosphates, and proteins that regulate concentrations of this lipid also regulate Akt activation and platelet function. Although the effectors through which Akt contributes to platelet activation are not definitively known, several candidates are discussed, including endothelial nitric oxide synthase, glycogen synthase kinase 3β, phosphodiesterase 3A and the integrin β3 tail. Selective inhibitors of Akt isoforms or of proteins that contribute to its activation, such as individual PI3K isoforms, may make attractive targets for antithrombotic therapy. This review summarizes the current literature describing Akt activity and its regulation in platelets, including speculation regarding the future of Akt or its regulatory pathways as targets for the development of antithrombotic therapies. PMID:20352060

  8. Antitumor Activity of Tenacissoside H on Esophageal Cancer through Arresting Cell Cycle and Regulating PI3K/Akt-NF-κB Transduction Cascade

    PubMed Central

    Jia, Yong-sen; Hu, Xue-qin; Gabriella, Hegyi; Qin, Li-juan; Meggyeshazi, Nora

    2015-01-01

    Objective. The purpose of the study was to elucidate the molecular mechanism of tenacissoside H (TDH) inhibiting esophageal carcinoma infiltration and proliferation. Methods. In vitro, EC9706 cells were treated with TDH. Cells proliferation and cell cycle were assayed. PI3K and NF-κB mRNAs expression were determined by real time PCR. In vivo, model of nude mice with tumor was established. Mice were treated with TDH. Inhibition ratio of tumor volume was calculated. PCNA expression was examined. Protein expression in PI3K/Akt-NF-κB signaling pathway was determined. Results. In vitro, TDH significantly inhibited cells proliferation in a time-and-dose-dependent manner. TDH arrested the cell cycle in S phase and significantly inhibited PI3K and NF-κB mRNA expression, compared with blank controlled group (P < 0.05). In vivo, TDH strongly inhibits tumor growth and volume. PCNA expression was significantly decreased after treatment of TDH. TDH downregulated proteins expression in PI3K/Akt-NF-κB transduction cascade (P < 0.05). Conclusion. TDH inhibited esophageal carcinoma infiltration and proliferation both in vitro and in vivo. The anticancer activity has relation to arresting the cell cycle at the S phase, inhibited the PCNA expression of transplanted tumors in nude mice, and regulated the protein expression in the PI3K/Akt-NF-κB transduction cascade. PMID:26495015

  9. MOLECULAR MECHANISMS OF RECEPTOR KINASE ACTION IN BRASSINOSTEROID SIGNAL TRANSDUCTION

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Brassinosteroids (BRs) regulate multiple aspects of plant growth and development and require an active BRASSINOSTEROID INSENSITIVE 1 (BRI1) and BRI1-ASSOCIATED RECEPTOR KINASE 1 (BAK1) for hormone perception and signal transduction. To examine early events in BR signaling, we used co-immunoprecipita...

  10. Developing a synthetic signal transduction system in plants.

    PubMed

    Morey, Kevin J; Antunes, Mauricio S; Albrecht, Kirk D; Bowen, Tessa A; Troupe, Jared F; Havens, Keira L; Medford, June I

    2011-01-01

    One area of focus in the emerging field of plant synthetic biology is the manipulation of systems involved in sensing and response to environmental signals. Sensing and responding to signals, including ligands, typically involves biological signal transduction. Plants use a wide variety of signaling systems to sense and respond to their environment. One of these systems, a histidine kinase (HK) based signaling system, lends itself to manipulation using the tools of synthetic biology. Both plants and bacteria use HKs to relay signals, which in bacteria can involve as few as two proteins (two-component systems or TCS). HK proteins are evolutionarily conserved between plants and bacteria and plant HK components have been shown to be functional in bacteria. We found that this conservation also applies to bacterial HK components which can function in plants. This conservation of function led us to hypothesize that synthetic HK signaling components can be designed and rapidly tested in bacteria. These novel HK signaling components form the foundation for a synthetic signaling system in plants, but typically require modifications such as codon optimization and proper targeting to allow optimal function. We describe the process and methodology of producing a synthetic signal transduction system in plants. We discovered that the bacterial response regulator (RR) PhoB shows HK-dependent nuclear translocation in planta. Using this discovery, we engineered a partial synthetic pathway in which a synthetic promoter (PlantPho) is activated using a plant-adapted PhoB (PhoB-VP64) and the endogenous HK-based cytokinin signaling pathway. Building on this work, we adapted an input or sensing system based on bacterial chemotactic binding proteins and HKs, resulting in a complete eukaryotic signal transduction system. Input to our eukaryotic signal transduction system is provided by a periplasmic binding protein (PBP), ribose-binding protein (RBP). RBP interacts with the membrane

  11. Signal transduction pathways involved in mechanotransduction in bone cells

    SciTech Connect

    Liedert, Astrid . E-mail: astrid.liedert@uni-ulm.de; Kaspar, Daniela; Blakytny, Robert; Claes, Lutz; Ignatius, Anita

    2006-10-13

    Several in vivo and in vitro studies with different loading regimens showed that mechanical stimuli have an influence on proliferation and differentiation of bone cells. Prerequisite for this influence is the transduction of mechanical signals into the cell, a phenomenon that is termed mechanotransduction, which is essential for the maintenance of skeletal homeostasis in adults. Mechanoreceptors, such as the integrins, cadherins, and stretch-activated Ca{sup 2+} channels, together with various signal transduction pathways, are involved in the mechanotransduction process that ultimately regulates gene expression in the nucleus. Mechanotransduction itself is considered to be regulated by hormones, the extracellular matrix of the osteoblastic cells and the mode of the mechanical stimulus.

  12. Neuropilins are positive regulators of Hedgehog signal transduction.

    PubMed

    Hillman, R Tyler; Feng, Brian Y; Ni, Jun; Woo, Wei-Meng; Milenkovic, Ljiljana; Hayden Gephart, Melanie G; Teruel, Mary N; Oro, Anthony E; Chen, James K; Scott, Matthew P

    2011-11-15

    The Hedgehog (Hh) pathway is essential for vertebrate embryogenesis, and excessive Hh target gene activation can cause cancer in humans. Here we show that Neuropilin 1 (Nrp1) and Nrp2, transmembrane proteins with roles in axon guidance and vascular endothelial growth factor (VEGF) signaling, are important positive regulators of Hh signal transduction. Nrps are expressed at times and locations of active Hh signal transduction during mouse development. Using cell lines lacking key Hh pathway components, we show that Nrps mediate Hh transduction between activated Smoothened (Smo) protein and the negative regulator Suppressor of Fused (SuFu). Nrp1 transcription is induced by Hh signaling, and Nrp1 overexpression increases maximal Hh target gene activation, indicating the existence of a positive feedback circuit. The regulation of Hh signal transduction by Nrps is conserved between mammals and bony fish, as we show that morpholinos targeting the Nrp zebrafish ortholog nrp1a produce a specific and highly penetrant Hh pathway loss-of-function phenotype. These findings enhance our knowledge of Hh pathway regulation and provide evidence for a conserved nexus between Nrps and this important developmental signaling system. PMID:22051878

  13. Signal transduction of stress via ceramide.

    PubMed Central

    Mathias, S; Peña, L A; Kolesnick, R N

    1998-01-01

    The sphingomyelin (SM) pathway is a ubiquitous, evolutionarily conserved signalling system analogous to conventional systems such as the cAMP and phosphoinositide pathways. Ceramide, which serves as second messenger in this pathway, is generated from SM by the action of a neutral or acidic SMase, or by de novo synthesis co-ordinated through the enzyme ceramide synthase. A number of direct targets for ceramide action have now been identified, including ceramide-activated protein kinase, ceramide-activated protein phosphatase and protein kinase Czeta, which couple the SM pathway to well defined intracellular signalling cascades. The SM pathway induces differentiation, proliferation or growth arrest, depending on the cell type. Very often, however, the outcome of signalling through this pathway is apoptosis. Mammalian systems respond to diverse stresses with ceramide generation, and recent studies show that yeast manifest a form of this response. Thus ceramide signalling is an older stress response system than the caspase/apoptotic death pathway, and hence these two pathways must have become linked later in evolution. Signalling of the stress response through ceramide appears to play a role in the development of human diseases, including ischaemia/reperfusion injury, insulin resistance and diabetes, atherogenesis, septic shock and ovarian failure. Further, ceramide signalling mediates the therapeutic effects of chemotherapy and radiation in some cells. An understanding of the mechanisms by which ceramide regulates physiological and pathological events in specific cells may provide new targets for pharmacological intervention. PMID:9794783

  14. MAPK Cascades in Guard Cell Signal Transduction

    PubMed Central

    Lee, Yuree; Kim, Yun Ju; Kim, Myung-Hee; Kwak, June M.

    2016-01-01

    Guard cells form stomata on the epidermis and continuously respond to endogenous and environmental stimuli to fine-tune the gas exchange and transpirational water loss, processes which involve mitogen-activated protein kinase (MAPK) cascades. MAPKs form three-tiered kinase cascades with MAPK kinases and MAPK kinase kinases, by which signals are transduced to the target proteins. MAPK cascade genes are highly conserved in all eukaryotes, and they play crucial roles in myriad developmental and physiological processes. MAPK cascades function during biotic and abiotic stress responses by linking extracellular signals received by receptors to cytosolic events and gene expression. In this review, we highlight recent findings and insights into MAPK-mediated guard cell signaling, including the specificity of MAPK cascades and the remaining questions. PMID:26904052

  15. MAPK Cascades in Guard Cell Signal Transduction.

    PubMed

    Lee, Yuree; Kim, Yun Ju; Kim, Myung-Hee; Kwak, June M

    2016-01-01

    Guard cells form stomata on the epidermis and continuously respond to endogenous and environmental stimuli to fine-tune the gas exchange and transpirational water loss, processes which involve mitogen-activated protein kinase (MAPK) cascades. MAPKs form three-tiered kinase cascades with MAPK kinases and MAPK kinase kinases, by which signals are transduced to the target proteins. MAPK cascade genes are highly conserved in all eukaryotes, and they play crucial roles in myriad developmental and physiological processes. MAPK cascades function during biotic and abiotic stress responses by linking extracellular signals received by receptors to cytosolic events and gene expression. In this review, we highlight recent findings and insights into MAPK-mediated guard cell signaling, including the specificity of MAPK cascades and the remaining questions. PMID:26904052

  16. Protein phosphorylation and its role in archaeal signal transduction.

    PubMed

    Esser, Dominik; Hoffmann, Lena; Pham, Trong Khoa; Bräsen, Christopher; Qiu, Wen; Wright, Phillip C; Albers, Sonja-Verena; Siebers, Bettina

    2016-09-01

    Reversible protein phosphorylation is the main mechanism of signal transduction that enables cells to rapidly respond to environmental changes by controlling the functional properties of proteins in response to external stimuli. However, whereas signal transduction is well studied in Eukaryotes and Bacteria, the knowledge in Archaea is still rather scarce. Archaea are special with regard to protein phosphorylation, due to the fact that the two best studied phyla, the Euryarchaeota and Crenarchaeaota, seem to exhibit fundamental differences in regulatory systems. Euryarchaeota (e.g. halophiles, methanogens, thermophiles), like Bacteria and Eukaryotes, rely on bacterial-type two-component signal transduction systems (phosphorylation on His and Asp), as well as on the protein phosphorylation on Ser, Thr and Tyr by Hanks-type protein kinases. Instead, Crenarchaeota (e.g. acidophiles and (hyper)thermophiles) only depend on Hanks-type protein phosphorylation. In this review, the current knowledge of reversible protein phosphorylation in Archaea is presented. It combines results from identified phosphoproteins, biochemical characterization of protein kinases and protein phosphatases as well as target enzymes and first insights into archaeal signal transduction by biochemical, genetic and polyomic studies. PMID:27476079

  17. Beyond microarrays: Finding key transcription factors controlling signal transduction pathways

    PubMed Central

    Kel, Alexdander; Voss, Nico; Jauregui, Ruy; Kel-Margoulis, Olga; Wingender, Edgar

    2006-01-01

    Background Massive gene expression changes in different cellular states measured by microarrays, in fact, reflect just an "echo" of real molecular processes in the cells. Transcription factors constitute a class of the regulatory molecules that typically require posttranscriptional modifications or ligand binding in order to exert their function. Therefore, such important functional changes of transcription factors are not directly visible in the microarray experiments. Results We developed a novel approach to find key transcription factors that may explain concerted expression changes of specific components of the signal transduction network. The approach aims at revealing evidence of positive feedback loops in the signal transduction circuits through activation of pathway-specific transcription factors. We demonstrate that promoters of genes encoding components of many known signal transduction pathways are enriched by binding sites of those transcription factors that are endpoints of the considered pathways. Application of the approach to the microarray gene expression data on TNF-alpha stimulated primary human endothelial cells helped to reveal novel key transcription factors potentially involved in the regulation of the signal transduction pathways of the cells. Conclusion We developed a novel computational approach for revealing key transcription factors by knowledge-based analysis of gene expression data with the help of databases on gene regulatory networks (TRANSFAC® and TRANSPATH®). The corresponding software and databases are available at . PMID:17118134

  18. Protein phosphorylation and its role in archaeal signal transduction

    PubMed Central

    Esser, Dominik; Hoffmann, Lena; Pham, Trong Khoa; Bräsen, Christopher; Qiu, Wen; Wright, Phillip C.; Albers, Sonja-Verena; Siebers, Bettina

    2016-01-01

    Reversible protein phosphorylation is the main mechanism of signal transduction that enables cells to rapidly respond to environmental changes by controlling the functional properties of proteins in response to external stimuli. However, whereas signal transduction is well studied in Eukaryotes and Bacteria, the knowledge in Archaea is still rather scarce. Archaea are special with regard to protein phosphorylation, due to the fact that the two best studied phyla, the Euryarchaeota and Crenarchaeaota, seem to exhibit fundamental differences in regulatory systems. Euryarchaeota (e.g. halophiles, methanogens, thermophiles), like Bacteria and Eukaryotes, rely on bacterial-type two-component signal transduction systems (phosphorylation on His and Asp), as well as on the protein phosphorylation on Ser, Thr and Tyr by Hanks-type protein kinases. Instead, Crenarchaeota (e.g. acidophiles and (hyper)thermophiles) only depend on Hanks-type protein phosphorylation. In this review, the current knowledge of reversible protein phosphorylation in Archaea is presented. It combines results from identified phosphoproteins, biochemical characterization of protein kinases and protein phosphatases as well as target enzymes and first insights into archaeal signal transduction by biochemical, genetic and polyomic studies. PMID:27476079

  19. Signal transduction and chemotaxis in mast cells.

    PubMed

    Draber, Petr; Halova, Ivana; Polakovicova, Iva; Kawakami, Toshiaki

    2016-05-01

    Mast cells play crucial roles in both innate and adaptive arms of the immune system. Along with basophils, mast cells are essential effector cells for allergic inflammation that causes asthma, allergic rhinitis, food allergy and atopic dermatitis. Mast cells are usually increased in inflammatory sites of allergy and, upon activation, release various chemical, lipid, peptide and protein mediators of allergic reactions. Since antigen/immunoglobulin E (IgE)-mediated activation of these cells is a central event to trigger allergic reactions, innumerable studies have been conducted on how these cells are activated through cross-linking of the high-affinity IgE receptor (FcεRI). Development of mature mast cells from their progenitor cells is under the influence of several growth factors, of which the stem cell factor (SCF) seems to be the most important. Therefore, how SCF induces mast cell development and activation via its receptor, KIT, has been studied extensively, including a cross-talk between KIT and FcεRI signaling pathways. Although our understanding of the signaling mechanisms of the FcεRI and KIT pathways is far from complete, pharmaceutical applications of the knowledge about these pathways are underway. This review will focus on recent progresses in FcεRI and KIT signaling and chemotaxis. PMID:25941081

  20. Aged black garlic extract inhibits HT29 colon cancer cell growth via the PI3K/Akt signaling pathway.

    PubMed

    Dong, Menghua; Yang, Guiqing; Liu, Hanchen; Liu, Xiaoxu; Lin, Sixiang; Sun, Dongning; Wang, Yishan

    2014-03-01

    Accumulating evidence indicates that aged black garlic extract (ABGE) may prove beneficial in preventing or inhibiting oncogenesis; however, the underlying mechanisms have not been fully elucidated. The present study aimed to investigate the effects of ABGE on the proliferation and apoptosis of HT29 colon cancer cells. Our results demonstrated that ABGE inhibited HT29 cell growth via the induction of apoptosis and cell cycle arrest. We further investigated the phosphatidylinositol 3-kinase/protein kinase B (PI3K/Akt) signal transduction pathway and the molecular mechanisms underlying the ABGE-induced inhibition of HT29 cell proliferation. We observed that ABGE may regulate the function of the PI3K/Akt pathway through upregulating PTEN and downregulating Akt and p-Akt expression, as well as suppressing its downstream target, 70-kDa ribosomal protein S6 kinase 1, at the mRNA and protein levels. In conclusion, these findings suggest that the PI3K/Akt signal transduction pathway is crucial for the development of colon cancer. ABGE inhibited the growth and induced apoptosis in HT29 cells through the inhibition of the PI3K/Akt pathway, suggesting that ABGE may be effective in the prevention and treatment of colon cancer in humans. PMID:24649105

  1. Parameter estimate of signal transduction pathways

    PubMed Central

    Arisi, Ivan; Cattaneo, Antonino; Rosato, Vittorio

    2006-01-01

    Background The "inverse" problem is related to the determination of unknown causes on the bases of the observation of their effects. This is the opposite of the corresponding "direct" problem, which relates to the prediction of the effects generated by a complete description of some agencies. The solution of an inverse problem entails the construction of a mathematical model and takes the moves from a number of experimental data. In this respect, inverse problems are often ill-conditioned as the amount of experimental conditions available are often insufficient to unambiguously solve the mathematical model. Several approaches to solving inverse problems are possible, both computational and experimental, some of which are mentioned in this article. In this work, we will describe in details the attempt to solve an inverse problem which arose in the study of an intracellular signaling pathway. Results Using the Genetic Algorithm to find the sub-optimal solution to the optimization problem, we have estimated a set of unknown parameters describing a kinetic model of a signaling pathway in the neuronal cell. The model is composed of mass action ordinary differential equations, where the kinetic parameters describe protein-protein interactions, protein synthesis and degradation. The algorithm has been implemented on a parallel platform. Several potential solutions of the problem have been computed, each solution being a set of model parameters. A sub-set of parameters has been selected on the basis on their small coefficient of variation across the ensemble of solutions. Conclusion Despite the lack of sufficiently reliable and homogeneous experimental data, the genetic algorithm approach has allowed to estimate the approximate value of a number of model parameters in a kinetic model of a signaling pathway: these parameters have been assessed to be relevant for the reproduction of the available experimental data. PMID:17118160

  2. Measurement of constitutive MAPK and PI3K/AKT signaling activity in human cancer cell lines

    PubMed Central

    Paraiso, Kim H.T.; Van Der Kooi, Kaisa; Messina, Jane L.; Smalley, Keiran S. M.

    2014-01-01

    The growth and survival of cancer cells is often driven by constitutive activity in the mitogen activated protein kinase (MAPK) and phospho-inositide 3-kinase (PI3K)/AKT signaling pathways. Activity in these signal transduction cascades is known to contribute to the uncontrolled growth and resistance to apoptosis that characterizes tumor progression. There is now a great deal of interest in therapeutically targeting these pathways in cancer using small molecule inhibitors. In this chapter we describe methods to measure constitutive MAPK and AKT activity in melanoma cell lines, with a focus upon Western blotting, phospho-flow cytometry and immunofluorescence staining techniques. PMID:21036250

  3. Signal transduction and Th17 cell differentiation

    PubMed Central

    O’Shea, John J.; Steward-Tharp, Scott M.; Laurence, Arian; Watford, Wendy T.; Wei, Lai; Adamson, Adewole S.; Fan, Samuel

    2009-01-01

    The paradigm of effector T helper cell differentiation into either Th1 or Th2 lineages has been notably shaken by the discovery of a third lineage of cells that selectively produce interleukin (IL)-17. Characterization of this new subset, referred to as Th17, has provided exciting new insights into immunoregulation, host defense and the pathogenesis of autoimmune diseases. Additionally, the discovery of this T cell subset has offered a fresh look at such concepts as lineage commitment and terminal differentiation. The transcriptional regulatory events and epigenetic modifications that control these processes are diverse and complex, and despite the rapid pace at which data continues to accumulate, many questions remain to be answered. Here we review our current understanding of the signaling pathways, molecular interactions and transcriptional events that lead to Th17 differentiation and effector function, as well as the epigenetic modifications that accompany them. PMID:19379825

  4. Signal transduction in mammalian oocytes during fertilization.

    PubMed

    Machaty, Zoltan

    2016-01-01

    Mammalian embryo development begins when the fertilizing sperm triggers a series of elevations in the oocyte's intracellular free Ca(2+) concentration. The elevations are the result of repeated release and re-uptake of Ca(2+) stored in the smooth endoplasmic reticulum. Ca(2+) release is primarily mediated by the phosphoinositide signaling system of the oocyte. The system is stimulated when the sperm causes the hydrolysis of phosphatidylinositol 4,5-bisphosphate (PIP2) into inositol 1,4,5-trisphosphate (IP3) and diacylglycerol (DAG); IP3 then binds its receptor on the surface of the endoplasmic reticulum that induces Ca(2+) release. The manner in which the sperm generates IP3, the Ca(2+) mobilizing second messenger, has been the subject of extensive research for a long time. The sperm factor hypothesis has eventually gained general acceptance, according to which it is a molecule from the sperm that diffuses into the ooplasm and stimulates the phosphoinositide cascade. Much evidence now indicates that the sperm-derived factor is phospholipase C-zeta (PLCζ) that cleaves PIP2 and generates IP3, eventually leading to oocyte activation. A recent addition to the candidate sperm factor list is the post-acrosomal sheath WW domain-binding protein (PAWP), whose role at fertilization is currently under debate. Ca(2+) influx across the plasma membrane is also important as, in the absence of extracellular Ca(2+), the oscillations run down prematurely. In pig oocytes, the influx that sustains the oscillations seems to be regulated by the filling status of the stores, whereas in the mouse other mechanisms might be involved. This work summarizes the current understanding of Ca(2+) signaling in mammalian oocytes. PMID:26453398

  5. Spatial organization and signal transduction at intercellular junctions

    PubMed Central

    Manz, Boryana N.; Groves, Jay T.

    2013-01-01

    The coordinated organization of cell membrane receptors into diverse micrometre-scale spatial patterns is emerging as an important theme of intercellular signalling, as exemplified by immunological synapses. Key characteristics of these patterns are that they transcend direct protein–protein interactions, emerge transiently and modulate signal transduction. Such cooperativity over multiple length scales presents new and intriguing challenges for the study and ultimate understanding of cellular signalling. As a result, new experimental strategies have emerged to manipulate the spatial organization of molecules inside living cells. The resulting spatial mutations yield insights into the interweaving of the spatial, mechanical and chemical aspects of intercellular signalling. PMID:20354536

  6. Signal transduction by the Wnt family of ligands.

    PubMed Central

    Dale, T C

    1998-01-01

    The Wnt genes encode a large family of secreted polypeptides that mediate cell-cell communication in diverse developmental processes. The loss or inappropriate activation of Wnt expression has been shown to alter cell fate, morphogenesis and mitogenesis. Recent progress has identified Wnt receptors and components of an intracellular signalling pathway that mediate Wnt-dependent transcription. This review will highlight this 'core' Wnt signal-transduction pathway, but also aims to reveal the potential diversity of Wnt signalling targets. Particular attention will be paid to the overlap between developmental biology and oncogenesis, since recent progress shows Wnt signalling forms a paradigm for an interdisciplinary approach. PMID:9425102

  7. Epstein-Barr virus LMP1 modulates lipid raft microdomains and the vimentin cytoskeleton for signal transduction and transformation.

    PubMed

    Meckes, David G; Menaker, Nathan F; Raab-Traub, Nancy

    2013-02-01

    The Epstein-Barr virus (EBV) is an important human pathogen that is associated with multiple cancers. The major oncoprotein of the virus, latent membrane protein 1 (LMP1), is essential for EBV B-cell immortalization and is sufficient to transform rodent fibroblasts. This viral transmembrane protein activates multiple cellular signaling pathways by engaging critical effector molecules and thus acts as a ligand-independent growth factor receptor. LMP1 is thought to signal from internal lipid raft containing membranes; however, the mechanisms through which these events occur remain largely unknown. Lipid rafts are microdomains within membranes that are rich in cholesterol and sphingolipids. Lipid rafts act as organization centers for biological processes, including signal transduction, protein trafficking, and pathogen entry and egress. In this study, the recruitment of key signaling components to lipid raft microdomains by LMP1 was analyzed. LMP1 increased the localization of phosphatidylinositol 3-kinase (PI3K) and its activated downstream target, Akt, to lipid rafts. In addition, mass spectrometry analyses identified elevated vimentin in rafts isolated from LMP1 expressing NPC cells. Disruption of lipid rafts through cholesterol depletion inhibited PI3K localization to membranes and decreased both Akt and ERK activation. Reduction of vimentin levels or disruption of its organization also decreased LMP1-mediated Akt and ERK activation and inhibited transformation of rodent fibroblasts. These findings indicate that LMP1 reorganizes membrane and cytoskeleton microdomains to modulate signal transduction. PMID:23152522

  8. Soliton growth-signal transduction in topologically quantized T cells

    NASA Astrophysics Data System (ADS)

    Matsson, Leif

    1993-09-01

    A model for growth-signal transduction of the T cell and its growth factor, interleukin-2, is presented. It is obtained as a generalization of the usual rate equation and is founded on the observation that a definite number of receptor occupations must take place in order to promote transition to the S phase and subsequent DNA replication. The generalized rate equation is identified as the equation of motion of a Lagrangian field theory of Ginzburg-Landau (Goldstone) type. However it is not an ad hoc model but is a microscopic theory of the interaction of interleukin-2 and its receptor. The topological quantum number of the model is related to the observed definite number of receptor occupations required to elicit growth-signal transduction. Individual receptor quanta, up to this limit, are subjected to a type of Bose condensation. This collective excitation constitutes the growth signal in the form of a topological kink soliton which is then launched by the next potential receptor occupation that makes the interaction repulsive. The model provides a possible long-absent explanation of the triggering mechanism for growth-signal transduction by means of the ambivalent interaction, which switches sign after a definite number of receptor occupations. Moreover, it offers an explanation of how Nature screens out fractional signals in the growth-signal-transduction process of T cells. Although the model is derived for assumed point-like cells and certain other restrictions, the obtained dose-response curves are in striking agreement with proliferation data from studies of both the leukemic T cell line MLA-144 from gibbon ape and normal human T cells in, and without, the presence of monoclonal anti-Tac antibodies.

  9. Mapping growth-factor-modulated Akt signaling dynamics.

    PubMed

    Gross, Sean M; Rotwein, Peter

    2016-05-15

    Growth factors alter cellular behavior through shared signaling cascades, raising the question of how specificity is achieved. Here, we have determined how growth factor actions are encoded into Akt signaling dynamics by real-time tracking of a fluorescent sensor. In individual cells, Akt activity was encoded in an analog pattern, with similar latencies (∼2 min) and half-maximal peak response times (range of 5-8 min). Yet, different growth factors promoted dose-dependent and heterogeneous changes in signaling dynamics. Insulin treatment caused sustained Akt activity, whereas EGF or PDGF-AA promoted transient signaling; PDGF-BB produced sustained responses at higher concentrations, but short-term effects at low doses, actions that were independent of the PDGF-α receptor. Transient responses to EGF were caused by negative feedback at the receptor level, as a second treatment yielded minimal responses, whereas parallel exposure to IGF-I caused full Akt activation. Small-molecule inhibitors reduced PDGF-BB signaling to transient responses, but only decreased the magnitude of IGF-I actions. Our observations reveal distinctions among growth factors that use shared components, and allow us to capture the consequences of receptor-specific regulatory mechanisms on Akt signaling. PMID:27044757

  10. Perturbation in T cell signal transduction pathway in microgravity

    NASA Astrophysics Data System (ADS)

    Kulkarni, A.; Yamauchi, K.; Taga, M.; Odle, J.; Sundaresan, A.; Pellis, N.

    T lymphocytes are regulatory and effector components of the immune system. It has been documented that T cell function is down regulated in microgravity of space flight and also in microgravity analogs. Lymphocyte signal transduction and the function of its effector elements are essential for proper functioning of T cells in any environment. We have shown that T cell mediated responses are down regulated in the microgravity analogs; in vivo antiorthostatic suspension mouse model as well as the in vitro culture system of Bioreactor (BIO). One of the postulated mechanisms for this effect is perturbation in signal transduction mechanisms via disruption of cytoskeleton due to the tensile force acting on cell membranes. Using BIO cultured mouse splenocytes we analyzed T cell signaling molecules associated with T cell receptor (TcR) and essential in signal transduction and cellular function. ZAP-70, a protein tyrosine kinase, is unaltered in 1g, however, is decreased 50% in the BIO at 96 hrs. SLP-76 levels drop more than 50% early in Bio samples at 24 and 48 hrs. LAT was unchanged. Once activated, ZAP-70 interacts with and phosphorylates Vav, SLP-76 and LAT proteins resulting in one of the complexs, namely SLP- 76/Vav, which putatively plays a regulatory role in TcR signal transduction pathway, perhaps via the actin cytoskeleton. Thus the decrease in SLP -76 at earlier time point could lead to ineffective recruitment and activation of cytoskeleton. Further studies are underway to delineate the mechanisms of T cell down regulation in microgravity. (Supported by NASA NCC8-168 grant, ADK)

  11. Casein kinase 2 dependent phosphorylation of neprilysin regulates receptor tyrosine kinase signaling to Akt.

    PubMed

    Siepmann, Martin; Kumar, Sathish; Mayer, Günter; Walter, Jochen

    2010-01-01

    Neprilysin (NEP) is a type II membrane metalloproteinase that cleaves physiologically active peptides at the cell surface thus regulating the local concentration of these peptides available for receptor binding and signal transduction. In addition, the cytoplasmic N-terminal domain of NEP interacts with the phosphatase and tensin homologue deleted on chromosome 10 (PTEN) thereby regulating intracellular signaling via Akt. Thus, NEP serves dual functions in extracellular and intracellular signal transduction. Here, we show that NEP undergoes phosphorylation at serine residue 6 within the N-terminal cytoplasmic domain. In vitro and cell culture experiments demonstrate that Ser 6 is efficiently phosphorylated by protein kinase CK2. The phosphorylation of the cytoplasmic domain of NEP inhibits its interaction with PTEN. Interestingly, expression of a pseudophosphorylated NEP variant (Ser6Asp) abrogates the inhibitory effect of NEP on insulin/insulin-like growth factor-1 (IGF-1) stimulated activation of Akt. Thus, our data demonstrate a regulatory role of CK2 in the interaction of NEP with PTEN and insulin/IGF-1 signaling. PMID:20957047

  12. Signal Transduction Cascades Regulating Fungal Development and Virulence

    PubMed Central

    Lengeler, Klaus B.; Davidson, Robert C.; D'souza, Cletus; Harashima, Toshiaki; Shen, Wei-Chiang; Wang, Ping; Pan, Xuewen; Waugh, Michael; Heitman, Joseph

    2000-01-01

    Cellular differentiation, mating, and filamentous growth are regulated in many fungi by environmental and nutritional signals. For example, in response to nitrogen limitation, diploid cells of the yeast Saccharomyces cerevisiae undergo a dimorphic transition to filamentous growth referred to as pseudohyphal differentiation. Yeast filamentous growth is regulated, in part, by two conserved signal transduction cascades: a mitogen-activated protein kinase cascade and a G-protein regulated cyclic AMP signaling pathway. Related signaling cascades play an analogous role in regulating mating and virulence in the plant fungal pathogen Ustilago maydis and the human fungal pathogens Cryptococcus neoformans and Candida albicans. We review here studies on the signaling cascades that regulate development of these and other fungi. This analysis illustrates both how the model yeast S. cerevisiae can serve as a paradigm for signaling in other organisms and also how studies in other fungi provide insights into conserved signaling pathways that operate in many divergent organisms. PMID:11104818

  13. Maxwell's demon in biochemical signal transduction with feedback loop.

    PubMed

    Ito, Sosuke; Sagawa, Takahiro

    2015-01-01

    Signal transduction in living cells is vital to maintain life itself, where information transfer in noisy environment plays a significant role. In a rather different context, the recent intensive research on 'Maxwell's demon'-a feedback controller that utilizes information of individual molecules-have led to a unified theory of information and thermodynamics. Here we combine these two streams of research, and show that the second law of thermodynamics with information reveals the fundamental limit of the robustness of signal transduction against environmental fluctuations. Especially, we find that the degree of robustness is quantitatively characterized by an informational quantity called transfer entropy. Our information-thermodynamic approach is applicable to biological communication inside cells, in which there is no explicit channel coding in contrast to artificial communication. Our result could open up a novel biophysical approach to understand information processing in living systems on the basis of the fundamental information-thermodynamics link. PMID:26099556

  14. Maxwell's demon in biochemical signal transduction with feedback loop

    NASA Astrophysics Data System (ADS)

    Ito, Sosuke; Sagawa, Takahiro

    2015-06-01

    Signal transduction in living cells is vital to maintain life itself, where information transfer in noisy environment plays a significant role. In a rather different context, the recent intensive research on `Maxwell's demon'--a feedback controller that utilizes information of individual molecules--have led to a unified theory of information and thermodynamics. Here we combine these two streams of research, and show that the second law of thermodynamics with information reveals the fundamental limit of the robustness of signal transduction against environmental fluctuations. Especially, we find that the degree of robustness is quantitatively characterized by an informational quantity called transfer entropy. Our information-thermodynamic approach is applicable to biological communication inside cells, in which there is no explicit channel coding in contrast to artificial communication. Our result could open up a novel biophysical approach to understand information processing in living systems on the basis of the fundamental information-thermodynamics link.

  15. Hypergravity signal transduction and gene expression in cultured mammalian cells

    NASA Technical Reports Server (NTRS)

    Kumei, Y.; Whitson, P. A.

    1994-01-01

    A number of studies have been conducted during space flight and with clinostats and centrifuges, suggesting that gravity effects the proliferation and differentiation of mammalian cells in vitro. However, little is known about the mechanisms by which mammalian cells respond to changes in gravitational stress. This paper summarizes studies designed to clarify the effects of hypergravity on the cultured human HeLa cells and to investigate the mechanism of hypergravity signal transduction in these cells.

  16. IKVAV regulates ERK1/2 and Akt signalling pathways in BMMSC population growth and proliferation

    PubMed Central

    Li, B; Qiu, T; Zhang, P; Wang, X; Yin, Y; Li, S

    2014-01-01

    Objectives The molecular mechanism of bone marrow mesenchymal stem cell (BMMSC) population growth and proliferation, induced by Isoleucyl-lysyl-valyl-alanyl-valine (IKVAV), was explored in this study. Materials and methods IKVAV peptides were synthesized by the solid-phase method. Influence of IKVAV on BMMSC population growth and proliferation were investigated by assays of CCK-8, flow cytometry, real-time PCR and western blotting. Results IKVAV peptide was found to induce proliferation and proliferating cell nuclear antigen (PCNA) synthesis of BMMSC in a dose- and time-dependent manner. Cell cycle analysis showed that the proportion of IKVAV-treated BMMSC in S phase in was higher than controls. Western blot results suggested that mitogen-activated protein kinase/extracellular signal-regulated kinase (MAPK/ERK) and phosphatidylinositol 3-kinase/protein kinase B (PI3K/Akt) signalling pathways were activated by IKVAV by enhancing phosphorylation levels of ERK1/2 and Akt in the BMMSCs. Meanwhile, phosphorylation levels of ERK1/2 and Akt were partially blocked by ERK1/2 inhibitor (PD98059) and Akt inhibitor (wortmannin), respectively. Conclusions Our results demonstrated that IKVAV stimulated BMMSC population growth and proliferation by activating MAPK/ERK1/2 and PI3K/Akt signalling pathways. This study is the first to reveal an enhancement effect of IKVAV peptide on BMMSC at the signal transduction level, and the outcome could provide experimental evidence for application of IKVAV-grafted scaffolds in the field of BMMSC-based tissue engineering. PMID:24617901

  17. Signal transduction and information processing in mammalian taste buds

    PubMed Central

    2013-01-01

    The molecular machinery for chemosensory transduction in taste buds has received considerable attention within the last decade. Consequently, we now know a great deal about sweet, bitter, and umami taste mechanisms and are gaining ground rapidly on salty and sour transduction. Sweet, bitter, and umami tastes are transduced by G-protein-coupled receptors. Salty taste may be transduced by epithelial Na channels similar to those found in renal tissues. Sour transduction appears to be initiated by intracellular acidification acting on acid-sensitive membrane proteins. Once a taste signal is generated in a taste cell, the subsequent steps involve secretion of neurotransmitters, including ATP and serotonin. It is now recognized that the cells responding to sweet, bitter, and umami taste stimuli do not possess synapses and instead secrete the neurotransmitter ATP via a novel mechanism not involving conventional vesicular exocytosis. ATP is believed to excite primary sensory afferent fibers that convey gustatory signals to the brain. In contrast, taste cells that do have synapses release serotonin in response to gustatory stimulation. The postsynaptic targets of serotonin have not yet been identified. Finally, ATP secreted from receptor cells also acts on neighboring taste cells to stimulate their release of serotonin. This suggests that there is important information processing and signal coding taking place in the mammalian taste bud after gustatory stimulation. PMID:17468883

  18. Molecular mechanisms in signal transduction at the membrane

    PubMed Central

    Groves, Jay T; Kuriyan, John

    2013-01-01

    Signal transduction originates at the membrane, where the clustering of signaling proteins is a key step in transmitting a message. Membranes are difficult to study, and their influence on signaling is still only understood at the most rudimentary level. Recent advances in the biophysics of membranes, surveyed in this review, have highlighted a variety of phenomena that are likely to influence signaling activity, such as local composition heterogeneities and long-range mechanical effects. We discuss recent mechanistic insights into three signaling systems—Ras activation, Ephrin signaling and the control of actin nucleation—where the active role of membrane components is now appreciated and for which experimentation on the membrane is required for further understanding. PMID:20495561

  19. Development of a new model system to dissect isoform specific Akt signalling in adipocytes.

    PubMed

    Kajno, Esi; McGraw, Timothy E; Gonzalez, Eva

    2015-06-15

    Protein kinase B (Akt) kinases are critical signal transducers mediating insulin action. Genetic studies revealed that Akt1 and Akt2 signalling differentially contribute to sustain lipid and glucose homoeostasis; however Akt isoform-specific effectors remain elusive due to the lack of a suitable model system to mechanistically interrogate Akt isoform-specific signalling. To overcome those technical limitations we developed a novel model system that provides acute and specific control of signalling by Akt isoforms. We generated mutants of Akt1 and Akt2 resistant to the allosteric Akt inhibitor MK-2206. We then developed adipocyte cell lines, in which endogenous Akt1 or Akt2 has been replaced by their corresponding drug-resistant Akt mutant. Treatment of those cells with MK-2206 allowed for acute and specific control of either Akt1 or Akt2 function. Our data showed that Akt1(W80A) and Akt2(W80A) mutants are resistant to MK-2206, dynamically regulated by insulin and able to signal to Akt downstream effectors. Analyses of insulin action in this cellular system showed that Akt1 and Akt2 are both able to mediate insulin regulation of the transcription factor forkhead box O1 (FoxO1) and the glucose transporter 4 (GLUT4), revealing a redundant role for these Akt kinases in the control of glucose transport into fat cells. In contrast, Akt1 signalling is uniquely required for adipogenesis, by controlling the mitotic clonal expansion (MCE) of pre-adipocytes that precedes white adipose cell differentiation. Our data provide new insights into the role of Akt kinases in glucose transport and adipogenesis and support our model system as a valuable tool for the biochemical characterization of signalling by specific Akt isoforms. PMID:25856301

  20. Deciphering Parameter Sensitivity in the BvgAS Signal Transduction

    PubMed Central

    Mapder, Tarunendu; Talukder, Srijeeta; Chattopadhyay, Sudip; Banik, Suman K.

    2016-01-01

    To understand the switching of different phenotypic phases of Bordetella pertussis, we propose an optimized mathematical framework for signal transduction through BvgAS two-component system. The response of the network output to the sensory input has been demonstrated in steady state. An analysis in terms of local sensitivity amplification characterizes the nature of the molecular switch. The sensitivity analysis of the model parameters within the framework of various correlation coefficients helps to decipher the contribution of the modular structure in signal propagation. Once classified, the model parameters are tuned to generate the behavior of some novel strains using simulated annealing, a stochastic optimization technique. PMID:26812153

  1. Determinants of specificity in two-component signal transduction.

    PubMed

    Podgornaia, Anna I; Laub, Michael T

    2013-04-01

    Maintaining the faithful flow of information through signal transduction pathways is critical to the survival and proliferation of organisms. This problem is particularly challenging as many signaling proteins are part of large, paralogous families that are highly similar at the sequence and structural levels, increasing the risk of unwanted cross-talk. To detect environmental signals and process information, bacteria rely heavily on two-component signaling systems comprised of sensor histidine kinases and their cognate response regulators. Although most species encode dozens of these signaling pathways, there is relatively little cross-talk, indicating that individual pathways are well insulated and highly specific. Here, we review the molecular mechanisms that enforce this specificity. Further, we highlight recent studies that have revealed how these mechanisms evolve to accommodate the introduction of new pathways by gene duplication. PMID:23352354

  2. Repression of AKT signaling by ARQ 092 in cells and tissues from patients with Proteus syndrome

    PubMed Central

    Lindhurst, Marjorie J.; Yourick, Miranda R.; Yu, Yi; Savage, Ronald E.; Ferrari, Dora; Biesecker, Leslie G.

    2015-01-01

    A somatic activating mutation in AKT1, c.49G>A, pGlu17Lys, that results in elevated AKT signaling in mutation-positive cells, is responsible for the mosaic overgrowth condition, Proteus syndrome. ARQ 092 is an allosteric pan-AKT inhibitor under development for treatment in cancer. We tested the efficacy of this drug for suppressing AKT signaling in cells and tissues from patients with Proteus syndrome. ARQ 092 reduced phosphorylation of AKT and downstream targets of AKT in a concentration-dependent manner in as little as two hours. While AKT signaling was suppressed with ARQ 092 treatment, cells retained their ability to respond to growth factor stimulation by increasing pAKT levels proportionally to untreated cells. At concentrations sufficient to decrease AKT signaling, little reduction in cell viability was seen. These results indicate that ARQ 092 can suppress AKT signaling and warrants further development as a therapeutic option for patients with Proteus syndrome. PMID:26657992

  3. Reduced modeling of signal transduction – a modular approach

    PubMed Central

    Koschorreck, Markus; Conzelmann, Holger; Ebert, Sybille; Ederer, Michael; Gilles, Ernst Dieter

    2007-01-01

    Background Combinatorial complexity is a challenging problem in detailed and mechanistic mathematical modeling of signal transduction. This subject has been discussed intensively and a lot of progress has been made within the last few years. A software tool (BioNetGen) was developed which allows an automatic rule-based set-up of mechanistic model equations. In many cases these models can be reduced by an exact domain-oriented lumping technique. However, the resulting models can still consist of a very large number of differential equations. Results We introduce a new reduction technique, which allows building modularized and highly reduced models. Compared to existing approaches further reduction of signal transduction networks is possible. The method also provides a new modularization criterion, which allows to dissect the model into smaller modules that are called layers and can be modeled independently. Hallmarks of the approach are conservation relations within each layer and connection of layers by signal flows instead of mass flows. The reduced model can be formulated directly without previous generation of detailed model equations. It can be understood and interpreted intuitively, as model variables are macroscopic quantities that are converted by rates following simple kinetics. The proposed technique is applicable without using complex mathematical tools and even without detailed knowledge of the mathematical background. However, we provide a detailed mathematical analysis to show performance and limitations of the method. For physiologically relevant parameter domains the transient as well as the stationary errors caused by the reduction are negligible. Conclusion The new layer based reduced modeling method allows building modularized and strongly reduced models of signal transduction networks. Reduced model equations can be directly formulated and are intuitively interpretable. Additionally, the method provides very good approximations especially for

  4. Spatial Regulation and the Rate of Signal Transduction Activation

    PubMed Central

    Batada, Nizar N; Shepp, Larry A; Siegmund, David O; Levitt, Michael

    2006-01-01

    Of the many important signaling events that take place on the surface of a mammalian cell, activation of signal transduction pathways via interactions of cell surface receptors is one of the most important. Evidence suggests that cell surface proteins are not as freely diffusible as implied by the classic fluid mosaic model and that their confinement to membrane domains is regulated. It is unknown whether these dynamic localization mechanisms function to enhance signal transduction activation rate or to minimize cross talk among pathways that share common intermediates. To determine which of these two possibilities is more likely, we derive an explicit equation for the rate at which cell surface membrane proteins interact based on a Brownian motion model in the presence of endocytosis and exocytosis. We find that in the absence of any diffusion constraints, cell surface protein interaction rate is extremely high relative to cytoplasmic protein interaction rate even in a large mammalian cell with a receptor abundance of a mere two hundred molecules. Since a larger number of downstream signaling events needs to take place, each occurring at a much slower rate than the initial activation via association of cell surface proteins, we conclude that the role of co-localization is most likely that of cross-talk reduction rather than coupling efficiency enhancement. PMID:16699596

  5. Abscisic acid perception and signaling transduction in strawberry

    PubMed Central

    Li, Chunli; Jia, Haifeng; Chai, Yemao; Shen, Yuanyue

    2011-01-01

    On basis of fruit differential respiration and ethylene effects, climacteric and non-climacteric fruits have been classically defined. Over the past decades, the molecular mechanisms of climacteric fruit ripening were abundantly described and found to focus on ethylene perception and signaling transduction. In contrast, until our most recent breakthroughs, much progress has been made toward understanding the signaling perception and transduction mechanisms for abscisic acid (ABA) in strawberry, a model for non-climacteric fruit ripening. Our reports not only have provided several lines of strong evidences for ABA-regulated ripening of strawberry fruit, but also have demonstrated that homology proteins of Arabidopsis ABA receptors, including PYR/PYL/RCAR and ABAR/CHLH, act as positive regulators of ripening in response to ABA. These receptors also trigger a set of ABA downstream signaling components, and determine significant changes in the expression levels of both sugar and pigment metabolism-related genes that are closely associated with ripening. Soluble sugars, especially sucrose, may act as a signal molecular to trigger ABA accumulation through an enzymatic action of 9-cis-epoxycarotenoid dioxygenase 1 (FaNCED1). This mini-review offers an overview of these processes and also outlines the possible, molecular mechanisms for ABA in the regulation of strawberry fruit ripening through the ABA receptors. PMID:22095148

  6. Analysis and logical modeling of biological signaling transduction networks

    NASA Astrophysics Data System (ADS)

    Sun, Zhongyao

    The study of network theory and its application span across a multitude of seemingly disparate fields of science and technology: computer science, biology, social science, linguistics, etc. It is the intrinsic similarities embedded in the entities and the way they interact with one another in these systems that link them together. In this dissertation, I present from both the aspect of theoretical analysis and the aspect of application three projects, which primarily focus on signal transduction networks in biology. In these projects, I assembled a network model through extensively perusing literature, performed model-based simulations and validation, analyzed network topology, and proposed a novel network measure. The application of network modeling to the system of stomatal opening in plants revealed a fundamental question about the process that has been left unanswered in decades. The novel measure of the redundancy of signal transduction networks with Boolean dynamics by calculating its maximum node-independent elementary signaling mode set accurately predicts the effect of single node knockout in such signaling processes. The three projects as an organic whole advance the understanding of a real system as well as the behavior of such network models, giving me an opportunity to take a glimpse at the dazzling facets of the immense world of network science.

  7. Compensation of the AKT signaling by ERK signaling in transgenic mice hearts overexpressing TRIM72

    SciTech Connect

    Ham, Young-Mi; Mahoney, Sarah Jane

    2013-06-10

    The AKT and ERK signaling pathways are known to be involved in cell hypertrophy, proliferation, survival and differentiation. Although there is evidence for crosstalk between these two signaling pathways in cellulo, there is less evidence for cross talk in vivo. Here, we show that crosstalk between AKT and ERK signaling in the hearts of TRIM72-overexpressing transgenic mice (TRIM72-Tg) with alpha-MHC promoter regulates and maintains their heart size. TRIM72, a heart- and skeletal muscle-specific protein, downregulates AKT-mTOR signaling via IRS-1 degradation and reduces the size of rat cardiomyocytes and the size of postnatal TRIM72-Tg hearts. TRIM72 expression was upregulated by hypertrophic inducers in cardiomyocytes, while IRS-1 was downregulated by IGF-1. TRIM72 specifically regulated IGF-1-dependent AKT-mTOR signaling, resulting in a reduction of the size of cardiomyocytes. Postnatal TRIM72-Tg hearts were smaller than control-treated hearts with inhibition of AKT-mTOR signaling. However, adult TRIM72-Tg hearts were larger than of control despite the suppression of AKT-mTOR signaling. Activation of ERK, PKC-α, and JNK were observed to be elevated in adult TRIM72-Tg, and these signals were mediated by ET-1 via the ET receptors A and B. Altogether, these results suggest that AKT signaling regulates cardiac hypertrophy in physiological conditions, and ERK signaling compensates for the absence of AKT signaling during TRIM72 overexpression, leading to pathological hypertrophy. -- Highlights: • TRIM72 inhibits AKT signaling through ubiquitination of IRS-1 in cardiac cells. • TRIM72 regulates the size of cardiac cells. • TRIM72 regulates size of postnatal TRIM72-overexpressing transgenic mice hearts. • Adult TRIM72-overexpressing transgenic mice hearts showed cardiac dysfunction. • Adult TRIM72 transgenic mice hearts showed higher expression of endothelin receptors.

  8. Fanconi Anemia: A Signal Transduction and DNA Repair Pathway

    PubMed Central

    Kupfer, Gary M.

    2013-01-01

    Fanconi anemia (FA) is a fascinating, rare genetic disorder marked by congenital defects, bone marrow failure, and cancer susceptibility. Research in recent years has led to the elucidation of FA as a DNA repair disorder and involved multiple pathways as well as having wide applicability to common cancers, including breast, ovarian, and head and neck. This review will describe the clinical aspects of FA as well as the current state of its molecular pathophysiology. In particular, work from the Kupfer laboratory will be described that demonstrates how the FA pathway interacts with multiple DNA repair pathways, including the mismatch repair system and signal transduction pathway of the DNA damage response. PMID:24348213

  9. Transduction of nanovolt signals: Limits of electric-field detection

    NASA Astrophysics Data System (ADS)

    Kalmijn, J.

    1989-11-01

    Life scientists discussed the extreme electrical sensitivity of marine sharks, skates, and rays. After reviewing the results of earlier studies on the electric sense at the animal and system levels, the participants discussed the basic process of signal transduction in terms of voltage-sensitive ionic channels. Struck by the small charge displacements needed for excitation, they strongly recommended that sensory biologists, physiologists, and biophysicists join in a concerted effort to initiate new research on the ionic mechanisms of electric field detection. To obtain detailed information on the electroreceptive membrane and its ionic channels, high resolution recording techniques will be mandatory.

  10. Ion channels and the transduction of light signals

    NASA Technical Reports Server (NTRS)

    Spalding, E. P.; Evans, M. L. (Principal Investigator)

    2000-01-01

    Studies of biological light-sensing mechanisms are revealing important roles for ion channels. Photosensory transduction in plants is no exception. In this article, the evidence that ion channels perform such signal-transducing functions in the complex array of mechanisms that bring about plant photomorphogenesis will be reviewed and discussed. The examples selected for discussion range from light-gradient detection in unicellular algae to the photocontrol of stem growth in Arabidopsis. Also included is some discussion of the technical aspects of studies that combine electrophysiology and photobiology.

  11. Olfactory Signal Transduction in the Mouse Septal Organ

    PubMed Central

    Ma, Minghong; Grosmaitre, Xavier; Iwema, Carrie L.; Baker, Harriet; Greer, Charles A.; Shepherd, Gordon M.

    2008-01-01

    The septal organ, a distinct chemosensory organ observed in the mammalian nose, is essentially a small island of olfactory neuroepithelium located bilaterally at the ventral base of the nasal septum. Virtually nothing is known about its physiological properties and function. To understand the nature of the sensory neurons in this area, we studied the mechanisms underlying olfactory signal transduction in these neurons. The majority of the sensory neurons in the septal organ express olfactory-specific G-protein and adenylyl cyclase type III, suggesting that the cAMP signaling pathway plays a critical role in the septal organ as in the main olfactory epithelium (MOE). This is further supported by patch-clamp recordings from individual dendritic knobs of the sensory neurons in the septal organ. Odorant responses can be mimicked by an adenylyl cyclase activator and a phosphodiesterase inhibitor, and these responses can be blocked by an adenylyl cyclase inhibitor. There is a small subset of cells in the septal organ expressing a cGMP-stimulated phosphodiesterase (phosphodiesterase 2), a marker for the guanylyl cyclase-D subtype sensory neurons identified in the MOE. The results indicate that the septal organ resembles the MOE in major olfactory signal transduction pathways, odorant response properties, and projection to the main olfactory bulb. Molecular and functional analysis of the septal organ, which constitutes ~1% of the olfactory epithelium, will provide new insights into the organization of the mammalian olfactory system and the unique function this enigmatic organ may serve. PMID:12514230

  12. [Contractile proteins in chemical signal transduction in plant microspores].

    PubMed

    Roshchina, V V

    2005-01-01

    Involvement of contractile components in chemical signal transduction from the cell surface to the organelles was studied using unicellular systems. Neurotransmitters dopamine and serotonin as well as active forms of oxygen hydrogen peroxide and tert-butyl peroxide were used as chemical signals. Experiments were carried out on vegetative microspores of field horsetail Equisetum arvense and generative microspores (pollen) of amaryllis Hippeastrum hybridum treated with cytochalasin B (an inhibitor of actin polymerization in microfilaments), colchicine, and vinblastine (inhibitors of tubulin polymerization in microtubules). Both types of thus treated microspores demonstrated suppressed development, particularly, for cytochalasin B treatment. At the same time, an increased typical blue fluorescence of certain cell regions (along the cell wall and around nuclei and chloroplasts) where the corresponding contractile proteins could reside was observed. In contrast to anticontractile agents, dopamine, serotonin B, and the peroxides stimulated microspore germination. Microspore pretreatment with cytochalasin B and colchicine followed by the treatment with serotonin, dopamine, or the peroxides decreased the germination rate. Involvement of actin and tubulin in chemical signal transduction from the cell surface to the nucleus is proposed. PMID:16004258

  13. Gravity perception and signal transduction in single cells

    NASA Astrophysics Data System (ADS)

    Block, I.; Wolke, A.; Briegleb, W.; Ivanova, K.

    Cellular signal processing in multi-, as well as in unicellular organisms, has to rely on fundamentally similar mechanisms. Free-living single cells often use the gravity vector for their spatial orientation (gravitaxis) and show distinct gravisensitivities. In this investigation the gravisensitive giant ameboid cell Physarum polycephalum (Myxomycetes, acellular slime molds) is used. Its gravitaxis and the modulation of its intrinsic rhythmic contraction activity by gravity was demonstrated in 180 °turn experiments and in simulated, as well as in actual, near-weightlessness studies (fast-rotating clinostat; Spacelab D1, IML-1). The stimulus perception was addressed in an IML-2 experiment, which provided information on the gravireceptor itself by the determination of the cell's acceleration-sensitivity threshold. Ground-based experiments designed to elucidate the subsequent steps in signal transduction leading to a motor response, suggest that an acceleration stimulus induces changes in the level of second messenger, adenosine 3',5'-cyclic monophosphate (cAMP), indicating also that the acceleration-stimulus signal transduction chain of Physarum uses an ubiquitous second messenger pathway.

  14. Genetic analysis of gravity signal transduction in roots

    NASA Astrophysics Data System (ADS)

    Masson, Patrick; Strohm, Allison; Baldwin, Katherine

    To grow downward into the soil, roots use gravity as a guide. Specialized cells, named stato-cytes, enable this directional growth response by perceiving gravity. Located in the columella region of the cap, these cells sense a reorientation of the root within the gravity field through the sedimentation of, and/or tension/pressure exerted by, dense amyloplasts. This process trig-gers a gravity signal transduction pathway that leads to a fast alkalinization of the cytoplasm and a change in the distribution of the plasma membrane-associated auxin-efflux carrier PIN3. The latter protein is uniformly distributed within the plasma membrane on all sides of the cell in vertically oriented roots. However, it quickly accumulates at the bottom side upon gravis-timulation. This process correlates with a preferential transport of auxin to the bottom side of the root cap, resulting in a lateral gradient across the tip. This gradient is then transported to the elongation zone where it promotes differential cellular elongation, resulting in downward curvature. We isolated mutations that affect gravity signal transduction at a step that pre-cedes cytoplasmic alkalinization and/or PIN3 relocalization and lateral auxin transport across the cap. arg1 and arl2 mutations identify a common genetic pathway that is needed for all three gravity-induced processes in the cap statocytes, indicating these genes function early in the pathway. On the other hand, adk1 affects gravity-induced PIN3 relocalization and lateral auxin transport, but it does not interfere with cytoplasmic alkalinization. ARG1 and ARL2 encode J-domain proteins that are associated with membranes of the vesicular trafficking path-way whereas ADK1 encodes adenosine kinase, an enzyme that converts adenosine derived from nucleic acid metabolism and the AdoMet cycle into AMP, thereby alleviating feedback inhibi-tion of this important methyl-donor cycle. Because mutations in ARG1 (and ARL2) do not completely eliminate

  15. Perspective: Adhesion Mediated Signal Transduction in Bacterial Pathogens

    PubMed Central

    Moorthy, Sudha; Keklak, Julia; Klein, Eric A.

    2016-01-01

    During the infection process, pathogenic bacteria undergo large-scale transcriptional changes to promote virulence and increase intrahost survival. While much of this reprogramming occurs in response to changes in chemical environment, such as nutrient availability and pH, there is increasing evidence that adhesion to host-tissue can also trigger signal transduction pathways resulting in differential gene expression. Determining the molecular mechanisms of adhesion-mediated signaling requires disentangling the contributions of chemical and mechanical stimuli. Here we highlight recent work demonstrating that surface attachment drives a transcriptional response in bacterial pathogens, including uropathogenic Escherichia coli (E. coli), and discuss the complexity of experimental design when dissecting the specific role of adhesion-mediated signaling during infection. PMID:26901228

  16. Perspective: Adhesion Mediated Signal Transduction in Bacterial Pathogens.

    PubMed

    Moorthy, Sudha; Keklak, Julia; Klein, Eric A

    2016-01-01

    During the infection process, pathogenic bacteria undergo large-scale transcriptional changes to promote virulence and increase intrahost survival. While much of this reprogramming occurs in response to changes in chemical environment, such as nutrient availability and pH, there is increasing evidence that adhesion to host-tissue can also trigger signal transduction pathways resulting in differential gene expression. Determining the molecular mechanisms of adhesion-mediated signaling requires disentangling the contributions of chemical and mechanical stimuli. Here we highlight recent work demonstrating that surface attachment drives a transcriptional response in bacterial pathogens, including uropathogenic Escherichia coli (E. coli), and discuss the complexity of experimental design when dissecting the specific role of adhesion-mediated signaling during infection. PMID:26901228

  17. Genetic Analysis of Gravity Signal Transduction in Arabidopsis Roots

    NASA Astrophysics Data System (ADS)

    Masson, Patrick; Strohm, Allison; Barker, Richard; Su, Shih-Heng

    Like most other plant organs, roots use gravity as a directional guide for growth. Specialized cells within the columella region of the root cap (the statocytes) sense the direction of gravity through the sedimentation of starch-filled plastids (amyloplasts). Amyloplast movement and/or pressure on sensitive membranes triggers a gravity signal transduction pathway within these cells, which leads to a fast transcytotic relocalization of plasma-membrane associated auxin-efflux carrier proteins of the PIN family (PIN3 and PIN7) toward the bottom membrane. This leads to a polar transport of auxin toward the bottom flank of the cap. The resulting lateral auxin gradient is then transmitted toward the elongation zones where it triggers a curvature that ultimately leads to a restoration of vertical downward growth. Our laboratory is using strategies derived from genetics and systems biology to elucidate the molecular mechanisms that modulate gravity sensing and signal transduction in the columella cells of the root cap. Our previous research uncovered two J-domain-containing proteins, ARG1 and ARL2, as contributing to this process. Mutations in the corresponding paralogous genes led to alterations of root and hypocotyl gravitropism accompanied by an inability for the statocytes to develop a cytoplasmic alkalinization, relocalize PIN3, and transport auxin laterally, in response to gravistimulation. Both proteins are associated peripherally to membranes belonging to various compartments of the vesicular trafficking pathway, potentially modulating the trafficking of defined proteins between plasma membrane and endosomes. MAR1 and MAR2, on the other end, are distinct proteins of the plastidic outer envelope protein import TOC complex (the transmembrane channel TOC75 and the receptor TOC132, respectively). Mutations in the corresponding genes enhance the gravitropic defects of arg1. Using transformation-rescue experiments with truncated versions of TOC132 (MAR2), we have shown

  18. Influence of Unweighting on Insulin Signal Transduction in Muscle

    NASA Technical Reports Server (NTRS)

    Tischler, Marc E.

    2002-01-01

    Unweighting of the juvenile soleus muscle is characterized by an increased binding capacity for insulin relative to muscle mass due to sparing of the receptors during atrophy. Although carbohydrate metabolism and protein degradation in the unweighted muscle develop increased sensitivity to insulin in vivo, protein synthesis in vivo and system A amino acid transport in vitro do not appear to develop such an enhanced response. The long-term goal is to identify the precise nature of this apparent resistance in the insulin signal transduction pathway and to consider how reduced weight-bearing may elicit this effect, by evaluating specific components of the insulin signalling pathway. Because the insulin-signalling pathway has components in common with the signal transduction pathway for insulin-like growth factor (IGF-1) and potentially other growth factors, the study could have important implications in the role of weight-bearing function on muscle growth and development. Since the insulin signalling pathway diverges following activation of insulin receptor tyrosine kinase, the immediate specific aims will be to study the receptor tyrosine kinase (IRTK) and those branches, which lead to phosphorylation of insulin receptor substrate-1 (IRS-1) and of Shc protein. To achieve these broader objectives, we will test in situ, by intramuscular injection, the responses of glucose transport, system A amino acid transport and protein synthesis to insulin analogues for which the receptor has either a weaker or much stronger binding affinity compared to insulin. Studies will include: (1) estimation of the ED(sub 50) for each analogue for these three processes; (2) the effect of duration (one to four days) of unweighting on the response of each process to all analogues tested; (3) the effect of unweighting and the analogues on IRTK activity; and (4) the comparative effects of unweighting and analogue binding on the tyrosine phosphorylation of IRTK, IRS-1, and Shc protein.

  19. Editorial of the Special Issue: Signaling Molecules and Signal Transduction in Cells

    PubMed Central

    Schlossmann, Jens

    2013-01-01

    In the special issue “Signaling Molecules and Signal Transduction in Cells” authors were invited to submit papers regarding important and novel aspects of extra- and intracellular signaling which have implications on physiological and pathophysiological processes. These aspects included compounds which are involved in these processes, elucidation of signaling pathways, as well as novel techniques for the analysis of signaling pathways. In response, various novel and important topics are elucidated in this special issue. PMID:23759992

  20. Akt and MAPK signaling mediate pregnancy-induced cardiac adaptation.

    PubMed

    Chung, Eunhee; Yeung, Fan; Leinwand, Leslie A

    2012-05-01

    Although the signaling pathways underlying exercise-induced cardiac adaptation have been extensively studied, little is known about the molecular mechanisms that result in the response of the heart to pregnancy. The objective of this study was to define the morphological, functional, and gene expression patterns that define the hearts of pregnant mice, and to identify the signaling pathways that mediate this response. Mice were divided into three groups: nonpregnant diestrus control, midpregnancy, and late pregnancy. Both time points of pregnancy were associated with significant cardiac hypertrophy. The prosurvival signaling cascades of Akt and ERK1/2 were activated in the hearts of pregnant mice, while the stress kinase, p38, was decreased. Given the activation of Akt in pregnancy and its known role in cardiac hypertrophy, the hypertrophic response to pregnancy was tested in mice expressing a cardiac-specific activated (myristoylated) form of Akt (myrAkt) or a cardiac-specific constitutively active (antipathologic hypertrophic) form of its downstream target, glycogen synthase kinase 3β (caGSK3β). The pregnancy-induced hypertrophic responses of hearts from these mice were significantly attenuated. Finally, we tested whether pregnancy-associated sex hormones could induce hypertrophy and alter signaling pathways in isolated neonatal rat ventricular myocytes (NRVMs). In fact, progesterone, but not estradiol treatment increased NRVM cell size via phosphorylation of ERK1/2. Inhibition of MEK1 effectively blocked progesterone-induced cellular hypertrophy. Taken together, our study demonstrates that pregnancy-induced cardiac hypertrophy is mediated by activation of Akt and ERK1/2 pathways. PMID:22345431

  1. PI3K/Akt signalling pathway and cancer.

    PubMed

    Fresno Vara, Juan Angel; Casado, Enrique; de Castro, Javier; Cejas, Paloma; Belda-Iniesta, Cristóbal; González-Barón, Manuel

    2004-04-01

    Phosphatidylinositol-3 kinases, PI3Ks, constitute a lipid kinase family characterized by their ability to phosphorylate inositol ring 3'-OH group in inositol phospholipids to generate the second messenger phosphatidylinositol-3,4,5-trisphosphate (PI-3,4,5-P(3)). RPTK activation results in PI(3,4,5)P(3) and PI(3,4)P(2) production by PI3K at the inner side of the plasma membrane. Akt interacts with these phospholipids, causing its translocation to the inner membrane, where it is phosphorylated and activated by PDK1 and PDK2. Activated Akt modulates the function of numerous substrates involved in the regulation of cell survival, cell cycle progression and cellular growth. In recent years, it has been shown that PI3K/Akt signalling pathway components are frequently altered in human cancers. Cancer treatment by chemotherapy and gamma-irradiation kills target cells primarily by the induction of apoptosis. However, the development of resistance to therapy is an important clinical problem. Failure to activate the apoptotic programme represents an important mode of drug resistance in tumor cells. Survival signals induced by several receptors are mediated mainly by PI3K/Akt, hence this pathway may decisively contribute to the resistant phenotype. Many of the signalling pathways involved in cellular transformation have been elucidated and efforts are underway to develop treatment strategies that target these specific signalling molecules or their downstream effectors. The PI3K/Akt pathway is involved in many of the mechanisms targeted by these new drugs, thus a better understanding of this crossroad can help to fully exploit the potential benefits of these new agents. PMID:15023437

  2. Fetus Sound Stimulation: Cilia Memristor Effect of Signal Transduction

    PubMed Central

    Jankovic-Raznatovic, Svetlana; Dragojevic-Dikic, Svetlana; Rakic, Snezana; Nikolic, Branka; Plesinac, Snezana; Tasic, Lidija; Perisic, Zivko; Sovilj, Mirjana; Adamovic, Tatjana; Koruga, Djuro

    2014-01-01

    Background. This experimental study evaluates fetal middle cerebral artery (MCA) circulation after the defined prenatal acoustical stimulation (PAS) and the role of cilia in hearing and memory and could explain signal transduction and memory according to cilia optical-acoustical properties. Methods. PAS was performed twice on 119 no-risk term pregnancies. We analyzed fetal MCA circulation before, after first and second PAS. Results. Analysis of the Pulsatility index basic (PIB) and before PAS and Pulsatility index reactive after the first PAS (PIR 1) shows high statistical difference, representing high influence on the brain circulation. Analysis of PIB and Pulsatility index reactive after the second PAS (PIR 2) shows no statistical difference. Cilia as nanoscale structure possess magnetic flux linkage that depends on the amount of charge that has passed between two-terminal variable resistors of cilia. Microtubule resistance, as a function of the current through and voltage across the structure, leads to appearance of cilia memory with the “memristor” property. Conclusion. Acoustical and optical cilia properties play crucial role in hearing and memory processes. We suggest that fetuses are getting used to sound, developing a kind of memory patterns, considering acoustical and electromagnetically waves and involving cilia and microtubules and try to explain signal transduction. PMID:24719851

  3. Signal Transduction Model of Magnetic Sensing in Cryptochrome Mediated Photoreception

    NASA Astrophysics Data System (ADS)

    Todd, Phillise Tiffeny

    While migratory birds have long been known to use the Earth's magnetic field for navigation, the precise biophysical mechanism behind this magnetic sense remains unconfirmed. A leading theory of magnetoreception suggests a chemical compass model with a yet undetermined molecular reaction site and unknown magnetically sensitive reactants. The cryptochrome photoreceptor has emerged as a promising candidate site. This investigation numerically models the first order kinetics of cryptochrome mediated photoreception, in order to evaluate its ability to function as a magnetic sensor and transduce orientation information along a neural pathway. A signal-to-noise ratio is defined to quantify the threshold for the functioning of a cryptochrome-based chemical compass. The model suggests that a flavin-superoxide radical pair in cryptochrome functions as the chemical reactants for magnetoreception. Such a cryptochrome-based signal transduction model reasonably predicts the general light intensity and wavelength effects that have been experimentally observed in migratory birds.

  4. PRDM Proteins: Molecular Mechanisms in Signal Transduction and Transcriptional Regulation.

    PubMed

    Di Zazzo, Erika; De Rosa, Caterina; Abbondanza, Ciro; Moncharmont, Bruno

    2013-01-01

    PRDM (PRDI-BF1 and RIZ homology domain containing) protein family members are characterized by the presence of a PR domain and a variable number of Zn-finger repeats. Experimental evidence has shown that the PRDM proteins play an important role in gene expression regulation, modifying the chromatin structure either directly, through the intrinsic methyltransferase activity, or indirectly through the recruitment of chromatin remodeling complexes. PRDM proteins have a dual action: they mediate the effect induced by different cell signals like steroid hormones and control the expression of growth factors. PRDM proteins therefore have a pivotal role in the transduction of signals that control cell proliferation and differentiation and consequently neoplastic transformation. In this review, we describe pathways in which PRDM proteins are involved and the molecular mechanism of their transcriptional regulation. PMID:24832654

  5. PRDM Proteins: Molecular Mechanisms in Signal Transduction and Transcriptional Regulation

    PubMed Central

    Di Zazzo, Erika; De Rosa, Caterina; Abbondanza, Ciro; Moncharmont, Bruno

    2013-01-01

    PRDM (PRDI-BF1 and RIZ homology domain containing) protein family members are characterized by the presence of a PR domain and a variable number of Zn-finger repeats. Experimental evidence has shown that the PRDM proteins play an important role in gene expression regulation, modifying the chromatin structure either directly, through the intrinsic methyltransferase activity, or indirectly through the recruitment of chromatin remodeling complexes. PRDM proteins have a dual action: they mediate the effect induced by different cell signals like steroid hormones and control the expression of growth factors. PRDM proteins therefore have a pivotal role in the transduction of signals that control cell proliferation and differentiation and consequently neoplastic transformation. In this review, we describe pathways in which PRDM proteins are involved and the molecular mechanism of their transcriptional regulation. PMID:24832654

  6. A computational model on the modulation of mitogen-activated protein kinase (MAPK) and Akt pathways in heregulin-induced ErbB signalling.

    PubMed Central

    Hatakeyama, Mariko; Kimura, Shuhei; Naka, Takashi; Kawasaki, Takuji; Yumoto, Noriko; Ichikawa, Mio; Kim, Jae-Hoon; Saito, Kazuki; Saeki, Mihoro; Shirouzu, Mikako; Yokoyama, Shigeyuki; Konagaya, Akihiko

    2003-01-01

    ErbB tyrosine kinase receptors mediate mitogenic signal cascade by binding a variety of ligands and recruiting the different cassettes of adaptor proteins. In the present study, we examined heregulin (HRG)-induced signal transduction of ErbB4 receptor and found that the phosphatidylinositol 3'-kinase (PI3K)-Akt pathway negatively regulated the extracellular signal-regulated kinase (ERK) cascade by phosphorylating Raf-1 on Ser(259). As the time-course kinetics of Akt and ERK activities seemed to be transient and complex, we constructed a mathematical simulation model for HRG-induced ErbB4 receptor signalling to explain the dynamics of the regulation mechanism in this signal transduction cascade. The model reflected well the experimental results observed in HRG-induced ErbB4 cells and in other modes of growth hormone-induced cell signalling that involve Raf-Akt cross-talk. The model suggested that HRG signalling is regulated by protein phosphatase 2A as well as Raf-Akt cross-talk, and protein phosphatase 2A modulates the kinase activity in both the PI3K-Akt and MAPK (mitogen-activated protein kinase) pathways. PMID:12691603

  7. The Hippo signal transduction network for exercise physiologists.

    PubMed

    Gabriel, Brendan M; Hamilton, D Lee; Tremblay, Annie M; Wackerhage, Henning

    2016-05-15

    The ubiquitous transcriptional coactivators Yap (gene symbol Yap1) and Taz (gene symbol Wwtr1) regulate gene expression mainly by coactivating the Tead transcription factors. Being at the center of the Hippo signaling network, Yap and Taz are regulated by the Hippo kinase cassette and additionally by a plethora of exercise-associated signals and signaling modules. These include mechanotransduction, the AKT-mTORC1 network, the SMAD transcription factors, hypoxia, glucose homeostasis, AMPK, adrenaline/epinephrine and angiotensin II through G protein-coupled receptors, and IL-6. Consequently, exercise should alter Hippo signaling in several organs to mediate at least some aspects of the organ-specific adaptations to exercise. Indeed, Tead1 overexpression in muscle fibers has been shown to promote a fast-to-slow fiber type switch, whereas Yap in muscle fibers and cardiomyocytes promotes skeletal muscle hypertrophy and cardiomyocyte adaptations, respectively. Finally, genome-wide association studies in humans have linked the Hippo pathway members LATS2, TEAD1, YAP1, VGLL2, VGLL3, and VGLL4 to body height, which is a key factor in sports. PMID:26940657

  8. Analysis of the gravitaxis signal transduction chain in Euglena gracilis

    NASA Astrophysics Data System (ADS)

    Nasir, Adeel

    Abstract Euglena gracilis is a photosynthetic, eukaryotic flagellate. It can adapt autotrophic and heterotrophic mode of growth and respond to different stimuli, this makes it an organism of choice for different research disciplines. It swims to reach a suitable niche by employing different stimuli such as oxygen, light, gravity and different chemicals. Among these stimuli light and gravity are the most important. Phototaxis (locomotion under light stimulus) and gravitaxis (locomotion under gravity stimulus) synergistically help cells to attain an optimal niche in the environment. However, in the complete absence of light or under scarcity of detectable light, cells can totally depend on gravity to find its swimming path. Therefore gravity has certain advantages over other stimuli.Unlike phototatic signal transduction chain of Euglena gracilis no clear primary gravity receptor has been identified in Euglena cells so far. However, there are some convincing evidence that TRP like channels act as a primary gravity receptor in Euglena gracilis.Use of different inhibitors gave rise to the involvement of protein kinase and calmodulin proteins in signal transduction chain of Euglena gracilis. Recently, specific calmodulin (Calmodulin 2) and protein kinase (PKA) have been identified as potential candidates of gravitactic signal transduction chain. Further characterization and investigation of these candidates was required. Therefore a combination of biochemical and genetic techniques was employed to localize proteins in cells and also to find interacting partners. For localization studies, specific antibodies were raised and characterized. Specificity of antibodies was validated by knockdown mutants, Invitro-translated proteins and heterologously expressed proteins. Cell fractionation studies, involving separation of the cell body and flagella for western blot analysis and confocal immunofluorescence studies were performed for subcellular localization. In order to find

  9. p53 attenuates AKT signaling by modulating membrane phospholipid composition

    PubMed Central

    Rueda-Rincon, Natalia; Bloch, Katarzyna; Derua, Rita; Vyas, Rajesh; Harms, Amy; Hankemeier, Thomas; Khan, Niamat Ali; Dehairs, Jonas; Bagadi, Muralidhararao; Binda, Maria Mercedes; Waelkens, Etienne; Marine, Jean-Christophe; Swinnen, Johannes V.

    2015-01-01

    The p53 tumor suppressor is the central component of a complex network of signaling pathways that protect organisms against the propagation of cells carrying oncogenic mutations. Here we report a previously unrecognized role of p53 in membrane phospholipids composition. By repressing the expression of stearoyl-CoA desaturase 1, SCD, the enzyme that converts saturated to mono-unsaturated fatty acids, p53 causes a shift in the content of phospholipids with mono-unsaturated acyl chains towards more saturated phospholipid species, particularly of the phosphatidylinositol headgroup class. This shift affects levels of phosphatidylinositol phosphates, attenuates the oncogenic AKT pathway, and contributes to the p53-mediated control of cell survival. These findings expand the p53 network to phospholipid metabolism and uncover a new molecular pathway connecting p53 to AKT signaling. PMID:26061814

  10. In search of cellular control: signal transduction in context

    NASA Technical Reports Server (NTRS)

    Ingber, D.

    1998-01-01

    The field of molecular cell biology has experienced enormous advances over the last century by reducing the complexity of living cells into simpler molecular components and binding interactions that are amenable to rigorous biochemical analysis. However, as our tools become more powerful, there is a tendency to define mechanisms by what we can measure. The field is currently dominated by efforts to identify the key molecules and sequences that mediate the function of critical receptors, signal transducers, and molecular switches. Unfortunately, these conventional experimental approaches ignore the importance of supramolecular control mechanisms that play a critical role in cellular regulation. Thus, the significance of individual molecular constituents cannot be fully understood when studied in isolation because their function may vary depending on their context within the structural complexity of the living cell. These higher-order regulatory mechanisms are based on the cell's use of a form of solid-state biochemistry in which molecular components that mediate biochemical processing and signal transduction are immobilized on insoluble cytoskeletal scaffolds in the cytoplasm and nucleus. Key to the understanding of this form of cellular regulation is the realization that chemistry is structure and hence, recognition of the the importance of architecture and mechanics for signal integration and biochemical control. Recent work that has unified chemical and mechanical signaling pathways provides a glimpse of how this form of higher-order cellular control may function and where paths may lie in the future.

  11. AKT signaling in ERBB2-amplified breast cancer.

    PubMed

    Carmona, F Javier; Montemurro, Filippo; Kannan, Srinivasaraghavan; Rossi, Valentina; Verma, Chandra; Baselga, José; Scaltriti, Maurizio

    2016-02-01

    The PI3K/AKT pathway is the focus of several targeted therapeutic agents for a variety of malignancies. In ERBB2-amplified breast cancer, the hyperactivation of this signaling cascade is associated with resistance to ERBB2-targeted therapy. This can occur through gain-of-function alterations or compensatory mechanisms that enter into play upon pharmacological pressure. The strong rationale in combining anti-ERBB2 agents with PI3K/AKT inhibitors, together with the identification of genomic alterations conferring sensitivity to targeted inhibition, are guiding the design of clinical studies aimed at preventing the emergence of drug resistance and achieving more durable response. In the present review, we describe the involvement of this pathway in breast cancer pathogenesis, with an emphasis on AKT kinases, and provide insight into currently available targeted agents for the treatment of ERBB2-amplified breast cancer. Finally, we provide preliminary data on a novel AKT3 mutation detected in the context of resistance to anti-ERBB2 therapy as an example of genomics-based approaches towards uncovering novel actionable targets in this setting. PMID:26645663

  12. Engineering aspects of enzymatic signal transduction: photoreceptors in the retina.

    PubMed Central

    Detwiler, P B; Ramanathan, S; Sengupta, A; Shraiman, B I

    2000-01-01

    Identifying the basic module of enzymatic amplification as an irreversible cycle of messenger activation/deactivation by a "push-pull" pair of opposing enzymes, we analyze it in terms of gain, bandwidth, noise, and power consumption. The enzymatic signal transduction cascade is viewed as an information channel, the design of which is governed by the statistical properties of the input and the noise and dynamic range constraints of the output. With the example of vertebrate phototransduction cascade we demonstrate that all of the relevant engineering parameters are controlled by enzyme concentrations and, from functional considerations, derive bounds on the required protein numbers. Conversely, the ability of enzymatic networks to change their response characteristics by varying only the abundance of different enzymes illustrates how functional diversity may be built from nearly conserved molecular components. PMID:11106590

  13. Interferons, Signal Transduction Pathways, and the Central Nervous System

    PubMed Central

    Nallar, Shreeram C.

    2014-01-01

    The interferon (IFN) family of cytokines participates in the development of innate and acquired immune defenses against various pathogens and pathogenic stimuli. Discovered originally as a proteinaceous substance secreted from virus-infected cells that afforded immunity to neighboring cells from virus infection, these cytokines are now implicated in various human pathologies, including control of tumor development, cell differentiation, and autoimmunity. It is now believed that the IFN system (IFN genes and the genes induced by them, and the factors that regulate these processes) is a generalized alarm of cellular stress, including DNA damage. IFNs exert both beneficial and deleterious effects on the central nervous system (CNS). Our knowledge of the IFN-regulated processes in the CNS is far from being clear. In this article, we reviewed the current understanding of IFN signal transduction pathways and gene products that might have potential relevance to diseases of the CNS. PMID:25084173

  14. Prolactin receptor and signal transduction to milk protein genes

    SciTech Connect

    Djiane, J.; Daniel, N.; Bignon, C.

    1994-06-01

    After cloning of the mammary gland prolactin (PRL) receptor cDNA, a functional assay was established using co-transfection of PRL receptor cDNA together with a milk protein promoter/chloramphenicol acetyl transferase (CAT) construct in Chinese hamster ovary (CHO) cells. Different mutants of the PRL receptor were tested in this CAT assay to delimit the domains in the receptor necessary for signal transduction to milk protein genes. In CHO cells stably transfected with PRL receptor cDNA, high numbers of PRL receptor are expressed. By metabolic labeling and immunoprecipitation, expressed PRL receptor was identified as a single species of 100 kDa. Using these cells, we analyzed the effects of PRL on intracellular free Ca{sup ++} concentration. PRL stimulates Ca{sup ++} entry and induces secondary Ca{sup ++} mobilization. The entry of Ca{sup ++} is a result of an increase in K{sup +} conductance that hyperpolarizes the membranes. We have also analyzed tyrosine phosphorylation induced by PRL. In CHO cells stably transfected with PRL receptor cDNA, PRL induced a very rapid and transient tyrosine phosphorylation of a 100-kDa protein which is most probably the PRL receptor. The same finding was obtained in mammary membranes after PRL injection to lactating rabbits. Whereas tyrosine kinase inhibitors genistein and lavendustin were without effect, PRL stimulation of milk protein gene promoters was partially inhibited by 2 {mu}M herbimycin in CHO cells co-transfected with PRL receptor cDNA and the {Beta} lactoglobulin CAT construct. Taken together these observations indicate that the cytoplasmic domain of the PRL receptor interacts with one or several tyrosine kinases, which may represent early postreceptor events necessary for PRL signal transduction to milk protein genes. 14 refs., 4 figs.

  15. [Recent studies on PI3K/AKT/mTOR signaling pathway in hematopoietic stem cells].

    PubMed

    Zhang, Ying-Chi; Cheng, Tao; Yuan, Wei-Ping

    2013-02-01

    PI3K/AKT/mTOR signaling pathway plays an essential role in the growth, proliferation and survival of various type of cells and also hematopoietic stem cells (HSC). Aberrant activation of PI3K/AKT/mTOR signaling pathway leads to exhaustion of HSC, while the inhibition of PI3K/AKT/mTOR signaling pathway results in blocking of B cell differentiation. This article reviews the latest advances on the role of key components involved in the PI3K/AKT/mTOR signaling pathway, including PI3K, AKT, mTOR, FoxO and GSK-3 in HSC. PMID:23484729

  16. Impaired Striatal Akt Signaling Disrupts Dopamine Homeostasis and Increases Feeding

    PubMed Central

    Davis, Adeola R.; Owens, W. Anthony; Matthies, Heinrich J. G.; Saadat, Sanaz; Kennedy, Jack P.; Vaughan, Roxanne A.; Neve, Rachael L.; Lindsley, Craig W.; Russo, Scott J.; Daws, Lynette C.; Niswender1, Kevin D.; Galli, Aurelio

    2011-01-01

    Background The prevalence of obesity has increased dramatically worldwide. The obesity epidemic begs for novel concepts and therapeutic targets that cohesively address “food-abuse” disorders. We demonstrate a molecular link between impairment of a central kinase (Akt) involved in insulin signaling induced by exposure to a high-fat (HF) diet and dysregulation of higher order circuitry involved in feeding. Dopamine (DA) rich brain structures, such as striatum, provide motivation stimuli for feeding. In these central circuitries, DA dysfunction is posited to contribute to obesity pathogenesis. We identified a mechanistic link between metabolic dysregulation and the maladaptive behaviors that potentiate weight gain. Insulin, a hormone in the periphery, also acts centrally to regulate both homeostatic and reward-based HF feeding. It regulates DA homeostasis, in part, by controlling a key element in DA clearance, the DA transporter (DAT). Upon HF feeding, nigro-striatal neurons rapidly develop insulin signaling deficiencies, causing increased HF calorie intake. Methodology/Principal Findings We show that consumption of fat-rich food impairs striatal activation of the insulin-activated signaling kinase, Akt. HF-induced Akt impairment, in turn, reduces DAT cell surface expression and function, thereby decreasing DA homeostasis and amphetamine (AMPH)-induced DA efflux. In addition, HF-mediated dysregulation of Akt signaling impairs DA-related behaviors such as (AMPH)-induced locomotion and increased caloric intake. We restored nigro-striatal Akt phosphorylation using recombinant viral vector expression technology. We observed a rescue of DAT expression in HF fed rats, which was associated with a return of locomotor responses to AMPH and normalization of HF diet-induced hyperphagia. Conclusions/Significance Acquired disruption of brain insulin action may confer risk for and/or underlie “food-abuse” disorders and the recalcitrance of obesity. This molecular model

  17. Roles of oxidative stress and Akt signaling in doxorubicin cardiotoxicity

    SciTech Connect

    Ichihara, Sahoko . E-mail: saho@gene.mie-u.ac.jp; Yamada, Yoshiji; Kawai, Yoshichika; Osawa, Toshihiko; Furuhashi, Koichi; Duan Zhiwen; Ichihara, Gaku

    2007-07-20

    Cardiotoxicity is a treatment-limiting side effect of the anticancer drug doxorubicin (DOX). We have now investigated the roles of oxidative stress and signaling by the protein kinase Akt in DOX-induced cardiotoxicity as well as the effects on such toxicity both of fenofibrate, an agonist of peroxisome proliferator-activated receptor-{alpha}, and of polyethylene glycol-conjugated superoxide dismutase (PEG-SOD), an antioxidant. Mice injected intraperitoneally with DOX were treated for 4 days with fenofibrate or PEG-SOD. Fenofibrate and PEG-SOD each prevented the induction of cardiac dysfunction by DOX. Both drugs also inhibited the activation of the transcription factor NF-{kappa}B and increase in lipid peroxidation in the left ventricle induced by DOX, whereas only PEG-SOD inhibited the DOX-induced activation of Akt and Akt-regulated gene expression. These results suggest that fenofibrate and PEG-SOD prevented cardiac dysfunction induced by DOX through normalization of oxidative stress and redox-regulated NF-{kappa}B signaling.

  18. Insulin Attenuates Beta-Amyloid-Associated Insulin/Akt/EAAT Signaling Perturbations in Human Astrocytes.

    PubMed

    Han, Xiaojuan; Yang, Liling; Du, Heng; Sun, Qinjian; Wang, Xiang; Cong, Lin; Liu, Xiaohui; Yin, Ling; Li, Shan; Du, Yifeng

    2016-08-01

    The excitatory amino acid transporters 1 and 2 (EAAT1 and EAAT2), mostly located on astrocytes, are the main mediators for glutamate clearance in humans. Malfunctions of these transporters may lead to excessive glutamate accumulation and subsequent excitotoxicity to neurons, which has been implicated in many kinds of neurodegenerative disorders including Alzheimer's disease (AD). Yet, the specific mechanism of the glutamate system dysregulation remains vague. To explore whether the insulin/protein kinase B (Akt)/EAAT signaling in human astrocytes could be disturbed by beta-amyloid protein (Aβ) and be protected by insulin, we incubated HA-1800 cells with varying concentrations of Aβ1-42 oligomers and insulin. Then the alterations of several key substrates in this signal transduction pathway were determined. Our results showed that expressions of insulin receptor, phospho-insulin receptor, phospho-protein kinase B, phospho-mammalian target of rapamycin, and EAAT1 and EAAT2 were decreased by the Aβ1-42 oligomers in a dose-dependent manner (p < 0.05) and this trend could be recovered by insulin treatment (p < 0.05). However, the expressions of total Akt and mTOR were invariant (p > 0.05), and the mRNA levels of EAAT1 and EAAT2 were also unchanged (p > 0.05). Taken together, this study indicates that Aβ1-42 oligomers could cause disturbances in insulin/Akt/EAAT signaling in astrocytes, which might be responsible for AD onset and progression. Additionally, insulin can exert protective functions to the brain by modulating protein modifications or expressions. PMID:26358886

  19. SAM68: Signal Transduction and RNA Metabolism in Human Cancer

    PubMed Central

    Frisone, Paola; Pradella, Davide; Di Matteo, Anna; Belloni, Elisa; Ghigna, Claudia; Paronetto, Maria Paola

    2015-01-01

    Alterations in expression and/or activity of splicing factors as well as mutations in cis-acting splicing regulatory sequences contribute to cancer phenotypes. Genome-wide studies have revealed more than 15,000 tumor-associated splice variants derived from genes involved in almost every aspect of cancer cell biology, including proliferation, differentiation, cell cycle control, metabolism, apoptosis, motility, invasion, and angiogenesis. In the past decades, several RNA binding proteins (RBPs) have been implicated in tumorigenesis. SAM68 (SRC associated in mitosis of 68 kDa) belongs to the STAR (signal transduction and activation of RNA metabolism) family of RBPs. SAM68 is involved in several steps of mRNA metabolism, from transcription to alternative splicing and then to nuclear export. Moreover, SAM68 participates in signaling pathways associated with cell response to stimuli, cell cycle transitions, and viral infections. Recent evidence has linked this RBP to the onset and progression of different tumors, highlighting misregulation of SAM68-regulated splicing events as a key step in neoplastic transformation and tumor progression. Here we review recent studies on the role of SAM68 in splicing regulation and we discuss its contribution to aberrant pre-mRNA processing in cancer. PMID:26273626

  20. A microfluidic platform for regulating signal transduction in single cells

    NASA Astrophysics Data System (ADS)

    Wong, Pak Kin; Yu, Fuqu; Sun, Ren; Ho, Chih-Ming

    2004-11-01

    Recent progress in micro cell culture systems has lead to new approaches in cell biology studies. Using micro devices for cell culturing possesses distinctive advantages over traditional methods. Length scale matching facilitates manipulation and detection at the single cell level. Previously, we have demonstrated generation of various stimulations such as spatial chemical gradient, electric field, and shear stress to study the dynamic responses of individual cells. Dynamic stimulations and continuous monitoring in a microfluidic system can be useful in studying different aspects of cellular process. In this work, we present a microfluidic platform for regulating nuclear factor kappa B (NF-kB) signal transduction in human embryonic kidney 293T cells. Time-varying bio-chemical stimulants, such as interleukin 1 and tumor necrosis factor, are introduced into the microchannel to activate the NF-kB signaling pathway. The dynamic responses of individual cells are monitored with the expression of reporter gene, green fluorescent protein. Regulation of the NF-kB activity is successfully demonstrated. This work is supported by CMISE through NASA URETI program.

  1. NO, nitrotyrosine, and cyclic GMP in signal transduction

    NASA Technical Reports Server (NTRS)

    Hanafy, K. A.; Krumenacker, J. S.; Murad, F.

    2001-01-01

    Over the past 25 years, the role of nitric oxide (NO) in biology has evolved from being recognized as an environmental pollutant to an endogenously produced substance involved in cell communication and signal transduction. NO is produced by a family of enzymes called nitric oxide synthases (NOSs), which can be stimulated by a variety of factors that mediate responses to various stimuli. NO can initiate its biological effects through activation of the heterodimeric enzyme, soluble guanylyl cyclase (sGC), or through several other chemical reactions. Activation of sGC results in the production of 3',5'-cyclic guanosine monophosphate (cGMP), an intracellular second messenger signaling molecule, which can subsequently mediate such diverse physiological events such as vasodilatation and immunomodulation. Chemically reactive NO can affect physiological changes through modifications to cellular proteins, one of which is tyrosine nitration. The demonstration that NO is involved in so many biological pathways indicates the importance of this endogenously produced substance, and suggests that there is much more to be discovered about its role in biology in years to come.

  2. Signal transduction in esophageal and LES circular muscle contraction.

    PubMed Central

    Harnett, K. M.; Cao, W.; Kim, N.; Sohn, U. D.; Rich, H.; Behar, J.; Biancani, P.

    1999-01-01

    Contraction of normal esophageal circular muscle (ESO) in response to acetylcholine (ACh) is linked to M2 muscarinic receptors activating at least three intracellular phospholipases, i.e., phosphatidylcholine-specific phospholipase C (PC-PLC), phospholipase D (PLD), and the high molecular weight (85 kDa) cytosolic phospholipase A2 (cPLA2) to induce phosphatidylcholine (PC) metabolism, production of diacylglycerol (DAG) and arachidonic acid (AA), resulting in activation of a protein kinase C (PKC)-dependent pathway. In contrast, lower esophageal sphincter (LES) contraction induced by maximally effective doses of ACh is mediated by muscarinic M3 receptors, linked to pertussis toxin-insensitive GTP-binding proteins of the G(q/11) type. They activate phospholipase C, which hydrolyzes phosphatidylinositol bisphosphate (PIP2), producing inositol 1,4,5-trisphosphate (IP3) and DAG. IP3 causes release of intracellular Ca++ and formation of a Ca++-calmodulin complex, resulting in activation of myosin light chain kinase and contraction through a calmodulin-dependent pathway. Signal transduction pathways responsible for maintenance of LES tone are quite distinct from those activated during contraction in response to maximally effective doses of agonists (e.g., ACh). Resting LES tone is associated with activity of a low molecular weight (approximately 14 kDa) pancreatic-like (group 1) secreted phospholipase A2 (sPLA2) and production of arachidonic acid (AA), which is metabolized to prostaglandins and thromboxanes. These AA metabolites act on receptors linked to G-proteins to induce activation of PI- and PC-specific phospholipases, and production of second messengers. Resting LES tone is associated with submaximal PI hydrolysis resulting in submaximal levels of inositol trisphosphate (IP3-induced Ca++ release, and interaction with DAG to activate PKC. In an animal model of acute esophagitis, acid-induced inflammation alters the contractile pathway of ESO and LES. In LES circular

  3. Genetic Analysis of Gravity Signal Transduction in Arabidopsis thaliana Seedlings

    NASA Astrophysics Data System (ADS)

    Boonsirichai, K.; Harrison, B.; Stanga, J.; Young, L.-S.; Neal, C.; Sabat, G.; Murthy, N.; Harms, A.; Sedbrook, J.; Masson, P.

    The primary roots of Arabidopsis thaliana seedlings respond to gravity stimulation by developing a tip curvature that results from differential cellular elongation on opposite flanks of the elongation zone. This curvature appears modulated by a lateral gradient of auxin that originates in the gravity-perceiving cells (statocytes) of the root cap through an apparent lateral repositioning of a component the auxin efflux carrier complex within these cells (Friml et al, 2002, Nature 415: 806-809). Unfortunately, little is known about the molecular mechanisms that govern early phases of gravity perception and signal transduction within the root-cap statocytes. We have used a molecular genetic approach to uncover some of these mechanisms. Mutations in the Arabidopsis ARG1 and ARL2 genes, which encode J-domain proteins, resulted in specific alterations in root and hypocotyl gravitropism, without pleiotropic phenotypes. Interestingly, ARG1 and ARL2 appear to function in the same genetic pathway. A combination of molecular genetic, biochemical and cell-biological approaches were used to demonstrate that ARG1 functions in early phases of gravity signal transduction within the root and hypocotyl statocytes, and is needed for efficient lateral auxin transport within the cap. The ARG1 protein is associated with components of the secretory and/or endosomal pathways, suggesting its role in the recycling of components of the auxin efflux carrier complex between plasma membrane and endosome (Boonsirichai et al, 2003, Plant Cell 15:2612-2625). Genetic modifiers of arg1-2 were isolated and shown to enhance the gravitropic defect of arg1-2, while resulting in little or no gravitropic defects in a wild type ARG1 background. A slight tendency for arg1-2;mar1-1 and arg1-2;mar2-1 double-mutant organs to display an opposite gravitropic response compared to wild type suggests that all three genes contribute to the interpretation of the gravity-vector information by seedling organs. The

  4. Activation of PI3K/Akt and ERK signaling pathways antagonized sinomenine-induced lung cancer cell apoptosis.

    PubMed

    Zhou, Liping; Luan, Hong; Liu, Qingpeng; Jiang, Tingshu; Liang, Hongyuan; Dong, Xihua; Shang, Hong

    2012-05-01

    Sinomenine (SIN) is a bioactive component derived from a Chinese medicinal plant. Our previous studies demonstrated that SIN has cytotoxic effects on human lung cancer cells. However, the antitumor molecular mechanisms of SIN have yet to be elucidated in detail. In the present study, we further explored the effects of SIN on NCI-H460 human lung cancer cell viability and apoptosis and investigated the regulation and function of PI3K/Akt and ERK signaling pathways during SIN-induced apoptosis in various lung cancer cell lines. NCI-H460 cells were incubated with 200 µg/ml SIN for the indicated times (0, 24, 48 or 72 h). Cell viability was assessed by MTT assay. Akt, p-Akt, ERK1/2 and p-ERK1/2 protein levels were detected by western blotting, respectively. Two different selective inhibitors (LY294002 for the PI3K pathway; PD98059 for the MEK/ERK pathway) were used to characterize the relative roles of PI3K/Akt and ERK in SIN-induced apoptosis. Apoptosis was determined by flow cytometry. SIN inhibited the proliferation of NCI-H460 cells in a time-dependent manner, which was accompanied with significant activation of pAkt and pERK. LY294002 and PD98059 both significantly increased SIN-induced apoptosis in NCI-H460, NCI-H226 and NCI-H522 cells. Our findings suggest that the activation of the PI3K/Akt and ERK signaling pathways antagonize SIN-induced lung cancer cell apoptosis and molecules that inhibit these pathways should potentiate the effects of SIN. This study represents a significant step forward in our understanding of the signal transduction pathways associated with the apoptosis elicited by SIN. PMID:22367396

  5. Specific cellular signal-transduction responses to in vivo combination therapy with ATRA, valproic acid and theophylline in acute myeloid leukemia.

    PubMed

    Skavland, J; Jørgensen, K M; Hadziavdic, K; Hovland, R; Jonassen, I; Bruserud, O; Gjertsen, B T

    2011-02-01

    Acute myeloid leukemia (AML) frequently comprises mutations in genes that cause perturbation in intracellular signaling pathways, thereby altering normal responses to growth factors and cytokines. Such oncogenic cellular signal transduction may be therapeutic if targeted directly or through epigenetic regulation. We treated 24 selected elderly AML patients with all-trans retinoic acid for 2 days before adding theophylline and the histone deacetylase inhibitor valproic acid (ClinicalTrials.gov NCT00175812; EudraCT no. 2004-001663-22), and sampled 11 patients for peripheral blood at day 0, 2 and 7 for single-cell analysis of basal level and signal-transduction responses to relevant myeloid growth factors (granulocyte-colony-stimulating factor, granulocyte/macrophage-colony-stimulating factor, interleukin-3, Flt3L, stem cell factor, erythropoietin, CXCL-12) on 10 signaling molecules (CREB, STAT1/3/5, p38, Erk1/2, Akt, c-Cbl, ZAP70/Syk and rpS6). Pretreatment analysis by unsupervised clustering and principal component analysis divided the patients into three distinguishable signaling clusters (non-potentiated, potentiated basal and potentiated signaling). Signal-transduction pathways were modulated during therapy and patients moved between the clusters. Patients with multiple leukemic clones demonstrated distinct stimulation responses and therapy-induced modulation. Individual signaling profiles together with clinical and hematological information may be used to early identify AML patients in whom epigenetic and signal-transduction targeted therapy is beneficial. PMID:22829110

  6. Specific cellular signal-transduction responses to in vivo combination therapy with ATRA, valproic acid and theophylline in acute myeloid leukemia

    PubMed Central

    Skavland, J; Jørgensen, K M; Hadziavdic, K; Hovland, R; Jonassen, I; Bruserud, Ø; Gjertsen, B T

    2011-01-01

    Acute myeloid leukemia (AML) frequently comprises mutations in genes that cause perturbation in intracellular signaling pathways, thereby altering normal responses to growth factors and cytokines. Such oncogenic cellular signal transduction may be therapeutic if targeted directly or through epigenetic regulation. We treated 24 selected elderly AML patients with all-trans retinoic acid for 2 days before adding theophylline and the histone deacetylase inhibitor valproic acid (ClinicalTrials.gov NCT00175812; EudraCT no. 2004-001663-22), and sampled 11 patients for peripheral blood at day 0, 2 and 7 for single-cell analysis of basal level and signal-transduction responses to relevant myeloid growth factors (granulocyte-colony-stimulating factor, granulocyte/macrophage-colony-stimulating factor, interleukin-3, Flt3L, stem cell factor, erythropoietin, CXCL-12) on 10 signaling molecules (CREB, STAT1/3/5, p38, Erk1/2, Akt, c-Cbl, ZAP70/Syk and rpS6). Pretreatment analysis by unsupervised clustering and principal component analysis divided the patients into three distinguishable signaling clusters (non-potentiated, potentiated basal and potentiated signaling). Signal-transduction pathways were modulated during therapy and patients moved between the clusters. Patients with multiple leukemic clones demonstrated distinct stimulation responses and therapy-induced modulation. Individual signaling profiles together with clinical and hematological information may be used to early identify AML patients in whom epigenetic and signal-transduction targeted therapy is beneficial. PMID:22829110

  7. Mannotriose regulates learning and memory signal transduction in the hippocampus

    PubMed Central

    Zhang, Lina; Dai, Weiwei; Zhang, Xueli; Gong, Zhangbin; Jin, Guoqin

    2013-01-01

    Rehmannia is a commonly used Chinese herb, which improves learning and memory. However, the crucial components of the signal transduction pathway associated with this effect remain elusive. Pri-mary hippocampal neurons were cultured in vitro, insulted with high-concentration (1 × 10−4 mol/L) cor-ticosterone, and treated with 1 × 10−4 mol/L mannotriose. Thiazolyl blue tetrazolium bromide assay and western blot analysis showed that hippocampal neuron survival rates and protein levels of glucocorti-coid receptor, serum and glucocorticoid-regulated protein kinase, and brain-derived neurotrophic factor were all dramatically decreased after high-concentration corticosterone-induced injury. This effect was reversed by mannotriose, to a similar level as RU38486 and donepezil. Our findings indicate that mannotriose could protect hippocampal neurons from high-concentration corticosterone-induced injury. The mechanism by which this occurred was associated with levels of glucocorticoid receptor protein, serum and glucocorticoid-regulated protein kinase, and brain-derived neurotrophic factor. PMID:25206622

  8. Post-translational modification of PII signal transduction proteins

    PubMed Central

    Merrick, Mike

    2015-01-01

    The PII proteins constitute one of the most widely distributed families of signal transduction proteins in nature. They are pivotal players in the control of nitrogen metabolism in bacteria and archaea, and are also found in the plastids of plants. Quite remarkably PII proteins control the activities of a diverse range of enzymes, transcription factors and membrane transport proteins, and in all known cases they achieve their regulatory effect by direct interaction with their target. PII proteins in the Proteobacteria and the Actinobacteria are subject to post-translational modification by uridylylation or adenylylation respectively, whilst in some Cyanobacteria they can be modified by phosphorylation. In all these cases the protein’s modification state is influenced by the cellular nitrogen status and is thought to regulate its activity. However, in many organisms there is no evidence for modification of PII proteins and indeed the ability of these proteins to respond to the cellular nitrogen status is fundamentally independent of post-translational modification. In this review we explore the role of post-translational modification in PII proteins in the light of recent studies. PMID:25610437

  9. Nitric oxide-mediated bystander signal transduction induced by heavy-ion microbeam irradiation.

    PubMed

    Tomita, Masanori; Matsumoto, Hideki; Funayama, Tomoo; Yokota, Yuichiro; Otsuka, Kensuke; Maeda, Munetoshi; Kobayashi, Yasuhiko

    2015-07-01

    In general, a radiation-induced bystander response is known to be a cellular response induced in non-irradiated cells after receiving bystander signaling factors released from directly irradiated cells within a cell population. Bystander responses induced by high-linear energy transfer (LET) heavy ions at low fluence are an important health problem for astronauts in space. Bystander responses are mediated via physical cell-cell contact, such as gap-junction intercellular communication (GJIC) and/or diffusive factors released into the medium in cell culture conditions. Nitric oxide (NO) is a well-known major initiator/mediator of intercellular signaling within culture medium during bystander responses. In this study, we investigated the NO-mediated bystander signal transduction induced by high-LET argon (Ar)-ion microbeam irradiation of normal human fibroblasts. Foci formation by DNA double-strand break repair proteins was induced in non-irradiated cells, which were co-cultured with those irradiated by high-LET Ar-ion microbeams in the same culture plate. Foci formation was suppressed significantly by pretreatment with an NO scavenger. Furthermore, NO-mediated reproductive cell death was also induced in bystander cells. Phosphorylation of NF-κB and Akt were induced during NO-mediated bystander signaling in the irradiated and bystander cells. However, the activation of these proteins depended on the incubation time after irradiation. The accumulation of cyclooxygenase-2 (COX-2), a downstream target of NO and NF-κB, was observed in the bystander cells 6 h after irradiation but not in the directly irradiated cells. Our findings suggest that Akt- and NF-κB-dependent signaling pathways involving COX-2 play important roles in NO-mediated high-LET heavy-ion-induced bystander responses. In addition, COX-2 may be used as a molecular marker of high-LET heavy-ion-induced bystander cells to distinguish them from directly irradiated cells, although this may depend on the time

  10. Nitric oxide-mediated bystander signal transduction induced by heavy-ion microbeam irradiation

    NASA Astrophysics Data System (ADS)

    Tomita, Masanori; Matsumoto, Hideki; Funayama, Tomoo; Yokota, Yuichiro; Otsuka, Kensuke; Maeda, Munetoshi; Kobayashi, Yasuhiko

    2015-07-01

    In general, a radiation-induced bystander response is known to be a cellular response induced in non-irradiated cells after receiving bystander signaling factors released from directly irradiated cells within a cell population. Bystander responses induced by high-linear energy transfer (LET) heavy ions at low fluence are an important health problem for astronauts in space. Bystander responses are mediated via physical cell-cell contact, such as gap-junction intercellular communication (GJIC) and/or diffusive factors released into the medium in cell culture conditions. Nitric oxide (NO) is a well-known major initiator/mediator of intercellular signaling within culture medium during bystander responses. In this study, we investigated the NO-mediated bystander signal transduction induced by high-LET argon (Ar)-ion microbeam irradiation of normal human fibroblasts. Foci formation by DNA double-strand break repair proteins was induced in non-irradiated cells, which were co-cultured with those irradiated by high-LET Ar-ion microbeams in the same culture plate. Foci formation was suppressed significantly by pretreatment with an NO scavenger. Furthermore, NO-mediated reproductive cell death was also induced in bystander cells. Phosphorylation of NF-κB and Akt were induced during NO-mediated bystander signaling in the irradiated and bystander cells. However, the activation of these proteins depended on the incubation time after irradiation. The accumulation of cyclooxygenase-2 (COX-2), a downstream target of NO and NF-κB, was observed in the bystander cells 6 h after irradiation but not in the directly irradiated cells. Our findings suggest that Akt- and NF-κB-dependent signaling pathways involving COX-2 play important roles in NO-mediated high-LET heavy-ion-induced bystander responses. In addition, COX-2 may be used as a molecular marker of high-LET heavy-ion-induced bystander cells to distinguish them from directly irradiated cells, although this may depend on the time

  11. FIST: a sensory domain for diverse signal transduction pathways in prokaryotes and ubiquitin signaling in eukaryotes

    SciTech Connect

    Borziak, Kirill; Jouline, Igor B

    2007-01-01

    Motivation: Sensory domains that are conserved among Bacteria, Archaea and Eucarya are important detectors of common signals detected by living cells. Due to their high sequence divergence, sensory domains are difficult to identify. We systematically look for novel sensory domains using sensitive profile-based searches initi-ated with regions of signal transduction proteins where no known domains can be identified by current domain models. Results: Using profile searches followed by multiple sequence alignment, structure prediction, and domain architecture analysis, we have identified a novel sensory domain termed FIST, which is present in signal transduction proteins from Bacteria, Archaea and Eucarya. Remote similarity to a known ligand-binding fold and chromosomal proximity of FIST-encoding genes to those coding for proteins involved in amino acid metabolism and transport suggest that FIST domains bind small ligands, such as amino acids.

  12. Matricellular signal transduction involving calmodulin in the social amoebozoan dictyostelium.

    PubMed

    O'Day, Danton H; Huber, Robert J

    2013-01-01

    The social amoebozoan Dictyostelium discoideum undergoes a developmental sequence wherein an extracellular matrix (ECM) sheath surrounds a group of differentiating cells. This sheath is comprised of proteins and carbohydrates, like the ECM of mammalian tissues. One of the characterized ECM proteins is the cysteine-rich, EGF-like (EGFL) repeat-containing, calmodulin (CaM)-binding protein (CaMBP) CyrA. The first EGFL repeat of CyrA increases the rate of random cell motility and cyclic AMP-mediated chemotaxis. Processing of full-length CyrA (~63 kDa) releases two major EGFL repeat-containing fragments (~45 kDa and ~40 kDa) in an event that is developmentally regulated. Evidence for an EGFL repeat receptor also exists and downstream intracellular signaling pathways involving CaM, Ras, protein kinase A and vinculin B phosphorylation have been characterized. In total, these results identify CyrA as a true matricellular protein comparable in function to tenascin C and other matricellular proteins from mammalian cells. Insight into the regulation and processing of CyrA has also been revealed. CyrA is the first identified extracellular CaMBP in this eukaryotic microbe. In keeping with this, extracellular CaM (extCaM) has been shown to be present in the ECM sheath where it binds to CyrA and inhibits its cleavage to release the 45 kDa and 40 kDa EGFL repeat-containing fragments. The presence of extCaM and its role in regulating a matricellular protein during morphogenesis extends our understanding of CaM-mediated signal transduction in eukaryotes. PMID:24705101

  13. Biomechanical Origins of Muscle Stem Cell Signal Transduction.

    PubMed

    Morrissey, James B; Cheng, Richard Y; Davoudi, Sadegh; Gilbert, Penney M

    2016-04-10

    Skeletal muscle, the most abundant and widespread tissue in the human body, contracts upon receiving electrochemical signals from the nervous system to support essential functions such as thermoregulation, limb movement, blinking, swallowing and breathing. Reconstruction of adult muscle tissue relies on a pool of mononucleate, resident muscle stem cells, known as "satellite cells", expressing the paired-box transcription factor Pax7 necessary for their specification during embryonic development and long-term maintenance during adult life. Satellite cells are located around the myofibres in a niche at the interface of the basal lamina and the host fibre plasma membrane (i.e., sarcolemma), at a very low frequency. Upon damage to the myofibres, quiescent satellite cells are activated and give rise to a population of transient amplifying myogenic progenitor cells, which eventually exit the cell cycle permanently and fuse to form new myofibres and regenerate the tissue. A subpopulation of satellite cells self-renew and repopulate the niche, poised to respond to future demands. Harnessing the potential of satellite cells relies on a complete understanding of the molecular mechanisms guiding their regulation in vivo. Over the past several decades, studies revealed many signal transduction pathways responsible for satellite cell fate decisions, but the niche cues driving the activation and silencing of these pathways are less clear. Here we explore the scintillating possibility that considering the dynamic changes in the biophysical properties of the skeletal muscle, namely stiffness, and the stretch and shear forces to which a myofibre can be subjected to may provide missing information necessary to gain a full understanding of satellite cell niche regulation. PMID:26004541

  14. Signal transduction inhibitors in treatment of myelodysplastic syndromes.

    PubMed

    Bachegowda, Lohith; Gligich, Oleg; Mantzaris, Ionnis; Schinke, Carolina; Wyville, Dale; Carrillo, Tatiana; Braunschweig, Ira; Steidl, Ulrich; Verma, Amit

    2013-01-01

    such as Farnesyl transferase inhibitors (Tipifarnib, Lonafarnib) and MEK inhibitor (GSK1120212) have shown acceptable toxicity profiles in small studies and efforts are underway to select mutational subgroups of MDS and AML that may benefit from these inhibitors. Altogether, these studies show that targeting various signal transduction pathways that regulate hematopoiesis offers promising therapeutic potential in this disease. Future studies in combination with high resolution correlative studies will clarify the subgroup specific efficacies of these agents. PMID:23841999

  15. Signal transduction inhibitors in treatment of myelodysplastic syndromes

    PubMed Central

    2013-01-01

    such as Farnesyl transferase inhibitors (Tipifarnib, Lonafarnib) and MEK inhibitor (GSK1120212) have shown acceptable toxicity profiles in small studies and efforts are underway to select mutational subgroups of MDS and AML that may benefit from these inhibitors. Altogether, these studies show that targeting various signal transduction pathways that regulate hematopoiesis offers promising therapeutic potential in this disease. Future studies in combination with high resolution correlative studies will clarify the subgroup specific efficacies of these agents. PMID:23841999

  16. Signal transduction molecules in gliomas of all grades

    PubMed Central

    Ermoian, Ralph P.; Kaprealian, Tania; Lamborn, Kathleen R.; Yang, Xiaodong; Jelluma, Nannette; Arvold, Nils D.; Zeidman, Ruth; Berger, Mitchel S.; Stokoe, David

    2010-01-01

    Purpose To interrogate grade II, III, and IV gliomas and characterize the critical effectors within the PI3-kinase pathway upstream and downstream of mTOR. Experimental design Tissues from 87 patients who were treated at UCSF between 1990 and 2004 were analyzed. Twenty-eight grade II, 17 grade III glioma, 26 grade IV gliomas, and 16 non-tumor brain specimens were analyzed. Protein levels were assessed by immunoblots; RNA levels were determined by polymerase chain reaction amplification. To address the multiple comparisons, first an overall analysis was done comparing the four groups using Spearman’s Correlation Coefficient. Only if this analysis was statistically significant were individual pairwise comparisons done. Results Multiple comparison analyses revealed a significant correlation with grade for all variables examined, except phosphorylated-S6. Expression of phosphorylated-4E-BP1, phosphorylated-PKB/Akt, PTEN, TSC1, and TSC2 correlated with grade (P < 0.01 for all). We extended our analyses to ask whether decreases in TSC proteins levels were due to changes in mRNA levels, or due to changes in post-transcriptional alterations. We found significantly lower levels of TSC1 and TSC2 mRNA in GBMs than in grade II gliomas or non-tumor brain (P < 0.01). Conclusions Expression levels of critical signaling molecules upstream and downstream of mTOR differ between non-tumor brain and gliomas of any grade. The single variable whose expression did not differ between non-tumor brain and gliomas was phosphorylated-S6, suggesting that other protein kinases, in addition to mTOR, contribute significantly to S6 phosphorylation. mTOR provides a rational therapeutic target in gliomas of all grades, and clinical benefit may emerge as mTOR inhibitors are combined with additional agents. PMID:18759130

  17. Changes in gene expression and signal transduction in microgravity

    NASA Technical Reports Server (NTRS)

    Hughes-Fulford, M.

    2001-01-01

    Studies from space flights over the past three decades have demonstrated that basic physiological changes occur in humans during space flight. These changes include cephalic fluid shifts, loss of fluid and electrolytes, loss of muscle mass, space motion sickness, anemia, reduced immune response, and loss of calcium and mineralized bone. The cause of most of these manifestations is not known and until recently, the general approach was to investigate general systemic changes, not basic cellular responses to microgravity. This laboratory has recently studied gene growth and activation of normal osteoblasts (MC3T3-El) during spaceflight. Osteoblast cells were grown on glass coverslips and loaded in the Biorack plunger boxes. The osteoblasts were launched in a serum deprived state, activated in microgravity and collected in microgravity. The osteoblasts were examined for changes in gene expression and signal transduction. Approximately one day after growth activation significant changes were observed in gene expression in 0-G flight samples. Immediate early growth genes/growth factors cox-2, c-myc, bcl2, TGF beta1, bFGF and PCNA showed a significant diminished mRNA induction in microgravity FCS activated cells when compared to ground and 1-G flight controls. Cox-1 was not detected in any of the samples. There were no significant differences in the expression of reference gene mRNA between the ground, 0-G and 1-G samples. The data suggest that quiescent osteoblasts are slower to enter the cell cycle in microgravity and that the lack of gravity itself may be a significant factor in bone loss in spaceflight. Preliminary data from our STS 76 flight experiment support our hypothesis that a basic biological response occurs at the tissue, cellular, and molecular level in 0-G. Here we examine ground-based and space flown data to help us understand the mechanism of bone loss in microgravity.

  18. Second-Chance Signal Transduction Explains Cooperative Flagellar Switching

    PubMed Central

    Zot, Henry G.; Hasbun, Javier E.; Van Minh, Nguyen

    2012-01-01

    The reversal of flagellar motion (switching) results from the interaction between a switch complex of the flagellar rotor and a torque-generating stationary unit, or stator (motor unit). To explain the steeply cooperative ligand-induced switching, present models propose allosteric interactions between subunits of the rotor, but do not address the possibility of a reaction that stimulates a bidirectional motor unit to reverse direction of torque. During flagellar motion, the binding of a ligand-bound switch complex at the dwell site could excite a motor unit. The probability that another switch complex of the rotor, moving according to steady-state rotation, will reach the same dwell site before that motor unit returns to ground state will be determined by the independent decay rate of the excited-state motor unit. Here, we derive an analytical expression for the energy coupling between a switch complex and a motor unit of the stator complex of a flagellum, and demonstrate that this model accounts for the cooperative switching response without the need for allosteric interactions. The analytical result can be reproduced by simulation when (1) the motion of the rotor delivers a subsequent ligand-bound switch to the excited motor unit, thereby providing the excited motor unit with a second chance to remain excited, and (2) the outputs from multiple independent motor units are constrained to a single all-or-none event. In this proposed model, a motor unit and switch complex represent the components of a mathematically defined signal transduction mechanism in which energy coupling is driven by steady-state and is regulated by stochastic ligand binding. Mathematical derivation of the model shows the analytical function to be a general form of the Hill equation (Hill AV (1910) The possible effects of the aggregation of the molecules of haemoglobin on its dissociation curves. J Physiol 40: iv–vii). PMID:22844429

  19. A respiratory chain controlled signal transduction cascade in the mitochondrial intermembrane space mediates hydrogen peroxide signaling

    PubMed Central

    Patterson, Heide Christine; Gerbeth, Carolin; Thiru, Prathapan; Vögtle, Nora F.; Knoll, Marko; Shahsafaei, Aliakbar; Samocha, Kaitlin E.; Huang, Cher X.; Harden, Mark Michael; Song, Rui; Chen, Cynthia; Kao, Jennifer; Shi, Jiahai; Salmon, Wendy; Shaul, Yoav D.; Stokes, Matthew P.; Silva, Jeffrey C.; Bell, George W.; MacArthur, Daniel G.; Ruland, Jürgen; Meisinger, Chris; Lodish, Harvey F.

    2015-01-01

    Reactive oxygen species (ROS) such as hydrogen peroxide (H2O2) govern cellular homeostasis by inducing signaling. H2O2 modulates the activity of phosphatases and many other signaling molecules through oxidation of critical cysteine residues, which led to the notion that initiation of ROS signaling is broad and nonspecific, and thus fundamentally distinct from other signaling pathways. Here, we report that H2O2 signaling bears hallmarks of a regular signal transduction cascade. It is controlled by hierarchical signaling events resulting in a focused response as the results place the mitochondrial respiratory chain upstream of tyrosine-protein kinase Lyn, Lyn upstream of tyrosine-protein kinase SYK (Syk), and Syk upstream of numerous targets involved in signaling, transcription, translation, metabolism, and cell cycle regulation. The active mediators of H2O2 signaling colocalize as H2O2 induces mitochondria-associated Lyn and Syk phosphorylation, and a pool of Lyn and Syk reside in the mitochondrial intermembrane space. Finally, the same intermediaries control the signaling response in tissues and species responsive to H2O2 as the respiratory chain, Lyn, and Syk were similarly required for H2O2 signaling in mouse B cells, fibroblasts, and chicken DT40 B cells. Consistent with a broad role, the Syk pathway is coexpressed across tissues, is of early metazoan origin, and displays evidence of evolutionary constraint in the human. These results suggest that H2O2 signaling is under control of a signal transduction pathway that links the respiratory chain to the mitochondrial intermembrane space-localized, ubiquitous, and ancient Syk pathway in hematopoietic and nonhematopoietic cells. PMID:26438848

  20. A respiratory chain controlled signal transduction cascade in the mitochondrial intermembrane space mediates hydrogen peroxide signaling.

    PubMed

    Patterson, Heide Christine; Gerbeth, Carolin; Thiru, Prathapan; Vögtle, Nora F; Knoll, Marko; Shahsafaei, Aliakbar; Samocha, Kaitlin E; Huang, Cher X; Harden, Mark Michael; Song, Rui; Chen, Cynthia; Kao, Jennifer; Shi, Jiahai; Salmon, Wendy; Shaul, Yoav D; Stokes, Matthew P; Silva, Jeffrey C; Bell, George W; MacArthur, Daniel G; Ruland, Jürgen; Meisinger, Chris; Lodish, Harvey F

    2015-10-20

    Reactive oxygen species (ROS) such as hydrogen peroxide (H2O2) govern cellular homeostasis by inducing signaling. H2O2 modulates the activity of phosphatases and many other signaling molecules through oxidation of critical cysteine residues, which led to the notion that initiation of ROS signaling is broad and nonspecific, and thus fundamentally distinct from other signaling pathways. Here, we report that H2O2 signaling bears hallmarks of a regular signal transduction cascade. It is controlled by hierarchical signaling events resulting in a focused response as the results place the mitochondrial respiratory chain upstream of tyrosine-protein kinase Lyn, Lyn upstream of tyrosine-protein kinase SYK (Syk), and Syk upstream of numerous targets involved in signaling, transcription, translation, metabolism, and cell cycle regulation. The active mediators of H2O2 signaling colocalize as H2O2 induces mitochondria-associated Lyn and Syk phosphorylation, and a pool of Lyn and Syk reside in the mitochondrial intermembrane space. Finally, the same intermediaries control the signaling response in tissues and species responsive to H2O2 as the respiratory chain, Lyn, and Syk were similarly required for H2O2 signaling in mouse B cells, fibroblasts, and chicken DT40 B cells. Consistent with a broad role, the Syk pathway is coexpressed across tissues, is of early metazoan origin, and displays evidence of evolutionary constraint in the human. These results suggest that H2O2 signaling is under control of a signal transduction pathway that links the respiratory chain to the mitochondrial intermembrane space-localized, ubiquitous, and ancient Syk pathway in hematopoietic and nonhematopoietic cells. PMID:26438848

  1. Use of double-stranded RNA interference in Drosophila cell lines to dissect signal transduction pathways

    PubMed Central

    Clemens, James C.; Worby, Carolyn A.; Simonson-Leff, Nancy; Muda, Marco; Maehama, Tomohiko; Hemmings, Brian A.; Dixon, Jack E.

    2000-01-01

    We demonstrate the efficacy of double-stranded RNA-mediated interference (RNAi) of gene expression in generating “knock-out” phenotypes for specific proteins in several Drosophila cell lines. We prove the applicability of this technique for studying signaling cascades by dissecting the well-characterized insulin signal transduction pathway. Specifically, we demonstrate that inhibiting the expression of the DSOR1 (mitogen-activated protein kinase kinase, MAPKK) prevents the activation of the downstream ERK-A (MAPK). In contrast, blocking ERK-A expression results in increased activation of DSOR1. We also show that Drosophila AKT (DAKT) activation depends on the insulin receptor substrate, CHICO (IRS1–4). Finally, we demonstrate that blocking the expression of Drosophila PTEN results in the activation of DAKT. In all cases, the interference of the biochemical cascade by RNAi is consistent with the known steps in the pathway. We extend this powerful technique to study two proteins, DSH3PX1 and Drosophila ACK (DACK). DSH3PX1 is an SH3, phox homology domain-containing protein, and DACK is homologous to the mammalian activated Cdc42 tyrosine kinase, ACK. Using RNAi, we demonstrate that DACK is upstream of DSH3PX1 phosphorylation, making DSH3PX1 an identified downstream target/substrate of ACK-like tyrosine kinases. These experiments highlight the usefulness of RNAi in dissecting complex biochemical signaling cascades and provide a highly effective method for determining the function of the identified genes arising from the Drosophila genome sequencing project. PMID:10823906

  2. Dynamic Modeling and Analysis of the Cross-Talk between Insulin/AKT and MAPK/ERK Signaling Pathways

    PubMed Central

    Arkun, Yaman

    2016-01-01

    Feedback loops play a key role in the regulation of the complex interactions in signal transduction networks. By studying the network of interactions among the biomolecules present in signaling pathways at the systems level, it is possible to understand how the biological functions are regulated and how the diseases emerge from their deregulations. This paper identifies the key feedback loops involved in the cross-talk among the insulin-AKT and MAPK/ERK signaling pathways. We developed a mathematical model that can be used to study the steady-state and dynamic behavior of the interactions among these two important signaling pathways. Modeling analysis and simulation case studies identify the key interaction parameters and the feedback loops that determine the normal and disease phenotypes. PMID:26930065

  3. Control of macrophage metabolism and activation by mTOR and Akt signaling

    PubMed Central

    Covarrubias, Anthony J.; Aksoylar, H. Ibrahim; Horng, Tiffany

    2015-01-01

    Macrophages are pleiotropic cells that assume a variety of functions depending on their tissue of residence and tissue state. They maintain homeostasis as well as coordinate responses to stresses such as infection and metabolic challenge. The ability of macrophages to acquire diverse, context-dependent activities requires their activation (or polarization) to distinct functional states. While macrophage activation is well understood at the level of signal transduction and transcriptional regulation, the metabolic underpinnings are poorly understood. Importantly, emerging studies indicate that metabolic shifts play a pivotal role in control of macrophage activation and acquisition of context-dependent effector activities. The signals that drive macrophage activation impinge on metabolic pathways, allowing for coordinate control of macrophage activation and metabolism. Here we discuss how mTOR and Akt, major metabolic regulators and targets of such activation signals, control macrophage metabolism and activation. Dysregulated macrophage activities contribute to many diseases, including infectious, inflammatory, and metabolic diseases and cancer, thus a better understanding of metabolic control of macrophage activation could pave the way to the development of new therapeutic strategies. PMID:26360589

  4. The gravity persistent signal (gps) Mutants of Arabidopsis: Insights into Gravitropic Signal Transduction

    NASA Astrophysics Data System (ADS)

    Wyatt, S.

    The gravitropic response of Arabidopsis stems is rapid with a visible within 30 min and vertical reorientation within 2 h. However, horizontal gravistimulation for 3 h at 4°C does not cause curvature. When the stems are subsequently placed in the vertical position at RT, they bend in response to the previous, horizontal gravistimulation. These results indicate that the gravity perception step can occur at 4°C, but that part of the response is sensitive to cold. At 4°C, starch-containing amyloplasts in the endodermis of the inflorescence stems sedimented normally but auxin transport was abolished indicating that the cold treatment affected early events of the signal transduction pathway that occur after amyloplast sedimentation but prior to auxin transport. The gps mutants of Arabidopsis are a unique group of mutants that respond abnormally after gravistimulation at 4°C. gps1 shows no response to the cold gravistimulation, gps2 bends the wrong way as compared to wild type and gps3 over responds, bending past the anticipated curvature. The mutants were selected from a T-DNA tagged population. Cloning strategies based on the tag have been employed to identify the genes disrupted. GPS1 was cloned using TAIL PCR and is At3g20130, a cytochrome P450, CYP705A22, of unknown function. GPS1p::GFP fusions are being used to determine temporal and spatial expression of GPS1. The mutation in gps3 appears to disrupt a non-coding region downstream of At1g43950 No function has yet been determined for this region, but it appears that the mutation disrupts transcription of a transcription factor homologous to the DNA binding domain of an auxin response factor (ARF) 9-like protein. The identity of GPS2 is as yet unknown. The gps mutants represent potentially three independent aspects of signal transduction in the gravitropic response: perception or retention of the gravity signal (gps1), determination of the polarity of the response (gps2), and the tissue specificity of the

  5. Physician Education: The Erythropoietin Receptor and Signal Transduction.

    PubMed

    Yoshimura; Arai

    1996-01-01

    differentiation such as the induction of globin synthesis [3, 4]. The remaining half is not required for this signaling, and, conversely, it acts to dampen the signals. It is known that a tyrosine kinase called JAK2 associates with the region near the plasma membrane, undergoes autophosphorylation, and phosphorylates the EPO receptor, and a transcription factor called a STAT [5]. It is thought that JAK2 plays an important role in promoting cellular proliferation. The STAT is activated by the phosphorylation, and it then translocates to the nucleus, recognizes a specific base sequence in the promoter region of its target gene, and initiates transcription. At present, we know that the STAT whose activation is mediated by the EPO receptor is STAT5, and the target genes are CIS [6], which has an SH2 domain (a molecular structure that recognizes a phosphorylated tyrosine) and OSM [7], which is a pleiotropic cytokine. However, activation of STAT5 and activation of the target genes are not unique to the EPO receptor, and they also occur with the IL-2 and IL-3 receptors. Moreover, the JAK2 substrate that is directly linked to cellular proliferation is still unknown. At present, studies are under way to determine the transcription factors specific to EPO and their target genes, as well as the substrates of JAK2. RECEPTOR PHOSPHORYLATION AND CESSATION OF THE SIGNAL: On the other hand, tyrosine phosphorylation of the receptor is necessary at the cytoplasmic tail region far from the plasma membrane, and the signal transduction pathway that originates with this phosphorylated tyrosine and is mediated by proteins with SH2 domains becomes activated. First, a GTP/GDP exchange factor called SOS, which is mediated by Shc and Grb2, migrates to the plasma membrane and converts a ras protein to its GTP form. The activated ras protein then activates the Raf-MAP kinase kinase-MAP kinase cascade, and ultimately initiates the transcription of oncogenes such as c-fos and c-jun. An enzyme called PI3 kinase

  6. FASEB summer research conference on signal transduction in plants. Final report, June 16, 1996--June 21, 1996

    SciTech Connect

    Lomax, T.L.; Quatrano, R.S.

    1996-12-31

    This is the program from the second FASEB conference on Signal Transduction in Plants. Topic areas included the following: environmental signaling; perception and transduction of light signals; signaling in plant microbe interactions; signaling in plant pathogen interactions; cell, cell communication; cytoskeleton, plasma membrane, and cellwall continuum; signaling molecules in plant growth and development I and II. A list of participants is included.

  7. PI3K-C2γ is a Rab5 effector selectively controlling endosomal Akt2 activation downstream of insulin signalling

    PubMed Central

    Braccini, Laura; Ciraolo, Elisa; Campa, Carlo C.; Perino, Alessia; Longo, Dario L.; Tibolla, Gianpaolo; Pregnolato, Marco; Cao, Yanyan; Tassone, Beatrice; Damilano, Federico; Laffargue, Muriel; Calautti, Enzo; Falasca, Marco; Norata, Giuseppe D.; Backer, Jonathan M.; Hirsch, Emilio

    2015-01-01

    In the liver, insulin-mediated activation of the phosphatidylinositol 3-kinase (PI3K)/Akt pathway is at the core of metabolic control. Multiple PI3K and Akt isoenzymes are found in hepatocytes and whether isoform-selective interplays exist is currently unclear. Here we report that insulin signalling triggers the association of the liver-specific class II PI3K isoform γ (PI3K-C2γ) with Rab5-GTP, and its recruitment to Rab5-positive early endosomes. In these vesicles, PI3K-C2γ produces a phosphatidylinositol-3,4-bisphosphate pool specifically required for delayed and sustained endosomal Akt2 stimulation. Accordingly, loss of PI3K-C2γ does not affect insulin-dependent Akt1 activation as well as S6K and FoxO1-3 phosphorylation, but selectively reduces Akt2 activation, which specifically inhibits glycogen synthase activity. As a consequence, PI3K-C2γ-deficient mice display severely reduced liver accumulation of glycogen and develop hyperlipidemia, adiposity as well as insulin resistance with age or after consumption of a high-fat diet. Our data indicate PI3K-C2γ supports an isoenzyme-specific forking of insulin-mediated signal transduction to an endosomal pool of Akt2, required for glucose homeostasis. PMID:26100075

  8. PI3K-C2γ is a Rab5 effector selectively controlling endosomal Akt2 activation downstream of insulin signalling.

    PubMed

    Braccini, Laura; Ciraolo, Elisa; Campa, Carlo C; Perino, Alessia; Longo, Dario L; Tibolla, Gianpaolo; Pregnolato, Marco; Cao, Yanyan; Tassone, Beatrice; Damilano, Federico; Laffargue, Muriel; Calautti, Enzo; Falasca, Marco; Norata, Giuseppe D; Backer, Jonathan M; Hirsch, Emilio

    2015-01-01

    In the liver, insulin-mediated activation of the phosphatidylinositol 3-kinase (PI3K)/Akt pathway is at the core of metabolic control. Multiple PI3K and Akt isoenzymes are found in hepatocytes and whether isoform-selective interplays exist is currently unclear. Here we report that insulin signalling triggers the association of the liver-specific class II PI3K isoform γ (PI3K-C2γ) with Rab5-GTP, and its recruitment to Rab5-positive early endosomes. In these vesicles, PI3K-C2γ produces a phosphatidylinositol-3,4-bisphosphate pool specifically required for delayed and sustained endosomal Akt2 stimulation. Accordingly, loss of PI3K-C2γ does not affect insulin-dependent Akt1 activation as well as S6K and FoxO1-3 phosphorylation, but selectively reduces Akt2 activation, which specifically inhibits glycogen synthase activity. As a consequence, PI3K-C2γ-deficient mice display severely reduced liver accumulation of glycogen and develop hyperlipidemia, adiposity as well as insulin resistance with age or after consumption of a high-fat diet. Our data indicate PI3K-C2γ supports an isoenzyme-specific forking of insulin-mediated signal transduction to an endosomal pool of Akt2, required for glucose homeostasis. PMID:26100075

  9. Cellular context–mediated Akt dynamics regulates MAP kinase signaling thresholds during angiogenesis

    PubMed Central

    Hellesøy, Monica; Lorens, James B.

    2015-01-01

    The formation of new blood vessels by sprouting angiogenesis is tightly regulated by contextual cues that affect angiogeneic growth factor signaling. Both constitutive activation and loss of Akt kinase activity in endothelial cells impair angiogenesis, suggesting that Akt dynamics mediates contextual microenvironmental regulation. We explored the temporal regulation of Akt in endothelial cells during formation of capillary-like networks induced by cell–cell contact with vascular smooth muscle cells (vSMCs) and vSMC-associated VEGF. Expression of constitutively active Akt1 strongly inhibited network formation, whereas hemiphosphorylated Akt1 epi-alleles with reduced kinase activity had an intermediate inhibitory effect. Conversely, inhibition of Akt signaling did not affect endothelial cell migration or morphogenesis in vSMC cocultures that generate capillary-like structures. We found that endothelial Akt activity is transiently blocked by proteasomal degradation in the presence of SMCs during the initial phase of capillary-like structure formation. Suppressed Akt activity corresponded to the increased endothelial MAP kinase signaling that was required for angiogenic endothelial morphogenesis. These results reveal a regulatory principle by which cellular context regulates Akt protein dynamics, which determines MAP kinase signaling thresholds necessary drive a morphogenetic program during angiogenesis. PMID:26023089

  10. Cell Type-Specific Activation of AKT and ERK Signaling Pathways by Small Negatively-Charged Magnetic Nanoparticles

    PubMed Central

    Rauch, Jens; Kolch, Walter; Mahmoudi, Morteza

    2012-01-01

    The interaction of nanoparticles (NPs) with living organisms has become a focus of public and scientific debate due to their potential wide applications in biomedicine, but also because of unwanted side effects. Here, we show that superparamagnetic iron oxide NPs (SPIONs) with different surface coatings can differentially affect signal transduction pathways. Using isogenic pairs of breast and colon derived cell lines we found that the stimulation of ERK and AKT signaling pathways by SPIONs is selectively dependent on the cell type and SPION type. In general, cells with Ras mutations respond better than their non-mutant counterparts. Small negatively charged SPIONs (snSPIONs) activated ERK to a similar extent as epidermal growth factor (EGF), and used the same upstream signaling components including activation of the EGF receptor. Importantly, snSPIONs stimulated the proliferation of Ras transformed breast epithelial cells as efficiently as EGF suggesting that NPs can mimic physiological growth factors. PMID:23162692

  11. Cell Type-Specific Activation of AKT and ERK Signaling Pathways by Small Negatively-Charged Magnetic Nanoparticles

    NASA Astrophysics Data System (ADS)

    Rauch, Jens; Kolch, Walter; Mahmoudi, Morteza

    2012-11-01

    The interaction of nanoparticles (NPs) with living organisms has become a focus of public and scientific debate due to their potential wide applications in biomedicine, but also because of unwanted side effects. Here, we show that superparamagnetic iron oxide NPs (SPIONs) with different surface coatings can differentially affect signal transduction pathways. Using isogenic pairs of breast and colon derived cell lines we found that the stimulation of ERK and AKT signaling pathways by SPIONs is selectively dependent on the cell type and SPION type. In general, cells with Ras mutations respond better than their non-mutant counterparts. Small negatively charged SPIONs (snSPIONs) activated ERK to a similar extent as epidermal growth factor (EGF), and used the same upstream signaling components including activation of the EGF receptor. Importantly, snSPIONs stimulated the proliferation of Ras transformed breast epithelial cells as efficiently as EGF suggesting that NPs can mimic physiological growth factors.

  12. {delta}-Opioid receptor-stimulated Akt signaling in neuroblastoma x glioma (NG108-15) hybrid cells involves receptor tyrosine kinase-mediated PI3K activation

    SciTech Connect

    Heiss, Anika; Ammer, Hermann; Eisinger, Daniela A.

    2009-07-15

    {delta}-Opioid receptor (DOR) agonists possess cytoprotective properties, an effect associated with activation of the 'pro-survival' kinase Akt. Here we delineate the signal transduction pathway by which opioids induce Akt activation in neuroblastoma x glioma (NG108-15) hybrid cells. Exposure of the cells to both [D-Pen{sup 2,5}]enkephalin and etorphine resulted in a time- and dose-dependent increase in Akt activity, as measured by means of an activation-specific antibody recognizing phosphoserine-473. DOR-mediated Akt signaling is blocked by the opioid antagonist naloxone and involves inhibitory G{sub i/o} proteins, because pre-treatment with pertussis toxin, but not over-expression of the G{sub q/11} scavengers EBP50 and GRK2-K220R, prevented this effect. Further studies with Wortmannin and LY294002 revealed that phophoinositol-3-kinase (PI3K) plays a central role in opioid-induced Akt activation. Opioids stimulate Akt activity through transactivation of receptor tyrosine kinases (RTK), because pre-treatment of the cells with inhibitors for neurotrophin receptor tyrosine kinases (AG879) and the insulin-like growth factor receptor IGF-1 (AG1024), but not over-expression of the G{beta}{gamma} scavenger phosducin, abolished this effect. Activated Akt translocates to the nuclear membrane, where it promotes GSK3 phosphorylation and prevents caspase-3 cleavage, two key events mediating inhibition of cell apoptosis and enhancement of cell survival. Taken together, these results demonstrate that in NG108-15 hybrid cells DOR agonists possess cytoprotective properties mediated by activation of the RTK/PI3K/Akt signaling pathway.

  13. Drosophila tribbles antagonizes insulin signaling-mediated growth and metabolism via interactions with Akt kinase.

    PubMed

    Das, Rahul; Sebo, Zachary; Pence, Laramie; Dobens, Leonard L

    2014-01-01

    Drosophila Tribbles (Trbl) is the founding member of the Trib family of kinase-like docking proteins that modulate cell signaling during proliferation, migration and growth. In a wing misexpression screen for Trbl interacting proteins, we identified the Ser/Thr protein kinase Akt1. Given the central role of Akt1 in insulin signaling, we tested the function of Trbl in larval fat body, a tissue where rapid increases in size are exquisitely sensitive to insulin/insulin-like growth factor levels. Consistent with a role in antagonizing insulin-mediated growth, trbl RNAi knockdown in the fat body increased cell size, advanced the timing of pupation and increased levels of circulating triglyceride. Complementarily, overexpression of Trbl reduced fat body cell size, decreased overall larval size, delayed maturation and lowered levels of triglycerides, while circulating glucose levels increased. The conserved Trbl kinase domain is required for function in vivo and for interaction with Akt in a yeast two-hybrid assay. Consistent with direct regulation of Akt, overexpression of Trbl in the fat body decreased levels of activated Akt (pSer505-Akt) while misexpression of trbl RNAi increased phospho-Akt levels, and neither treatment affected total Akt levels. Trbl misexpression effectively suppressed Akt-mediated wing and muscle cell size increases and reduced phosphorylation of the Akt target FoxO (pSer256-FoxO). Taken together, these data show that Drosophila Trbl has a conserved role to bind Akt and block Akt-mediated insulin signaling, and implicate Trib proteins as novel sites of signaling pathway integration that link nutrient availability with cell growth and proliferation. PMID:25329475

  14. Drosophila Tribbles Antagonizes Insulin Signaling-Mediated Growth and Metabolism via Interactions with Akt Kinase

    PubMed Central

    Das, Rahul; Sebo, Zachary; Pence, Laramie; Dobens, Leonard L.

    2014-01-01

    Drosophila Tribbles (Trbl) is the founding member of the Trib family of kinase-like docking proteins that modulate cell signaling during proliferation, migration and growth. In a wing misexpression screen for Trbl interacting proteins, we identified the Ser/Thr protein kinase Akt1. Given the central role of Akt1 in insulin signaling, we tested the function of Trbl in larval fat body, a tissue where rapid increases in size are exquisitely sensitive to insulin/insulin-like growth factor levels. Consistent with a role in antagonizing insulin-mediated growth, trbl RNAi knockdown in the fat body increased cell size, advanced the timing of pupation and increased levels of circulating triglyceride. Complementarily, overexpression of Trbl reduced fat body cell size, decreased overall larval size, delayed maturation and lowered levels of triglycerides, while circulating glucose levels increased. The conserved Trbl kinase domain is required for function in vivo and for interaction with Akt in a yeast two-hybrid assay. Consistent with direct regulation of Akt, overexpression of Trbl in the fat body decreased levels of activated Akt (pSer505-Akt) while misexpression of trbl RNAi increased phospho-Akt levels, and neither treatment affected total Akt levels. Trbl misexpression effectively suppressed Akt-mediated wing and muscle cell size increases and reduced phosphorylation of the Akt target FoxO (pSer256-FoxO). Taken together, these data show that Drosophila Trbl has a conserved role to bind Akt and block Akt-mediated insulin signaling, and implicate Trib proteins as novel sites of signaling pathway integration that link nutrient availability with cell growth and proliferation. PMID:25329475

  15. Dissecting signalling by individual Akt/PKB isoforms, three steps at once.

    PubMed

    Osorio-Fuentealba, Cesar; Klip, Amira

    2015-09-01

    The serine/threonine kinase Akt/PKB (protein kinase B) is key for mammalian cell growth, survival, metabolism and oncogenic transformation. The diverse level and tissue expression of its three isoforms, Akt1/PKBα, Akt2/PKBβ and Akt3/PKBγ, make it daunting to identify isoform-specific actions in vivo and even in isolated tissues/cells. To date, isoform-specific knockout and knockdown have been the best strategies to dissect their individual overall functions. In a recent article in the Biochemical Journal, Kajno et al. reported a new strategy to study isoform selectivity in cell lines. Individual Akt/PKB isoforms in 3T3-L1 pre-adipocytes are first silenced via shRNA and stable cellular clones lacking one or the other isoform are selected. The stably silenced isoform is then replaced by a mutant engineered to be refractory to inhibition by MK-2206 (Akt1(W80A) or Akt2(W80A)). Akt1(W80A) or Akt2(W80A) are functional and effectively recruited to the plasma membrane in response to insulin. The system affords the opportunity to acutely control the activity of the endogenous non-silenced isoform through timely addition of MK-2206. Using this approach, it is confirmed that Akt1/PKBα is the preferred isoform sustaining adipocyte differentiation, but both Akt1/PKBα and Akt2/PKBβ can indistinctly support insulin-dependent FoxO1 (forkhead box O1) nuclear exclusion. Surprisingly, either isoform can also support insulin-dependent glucose transporter (GLUT) 4 translocation to the membrane, in contrast with the preferential role of Akt2/PKBβ assessed by knockdown studies. The new strategy should allow analysis of the plurality of Akt/PKB functions in other cells and in response to other stimuli. It should also be amenable to high-throughput studies to speed up advances in signal transmission by this pivotal kinase. PMID:26348913

  16. Inhibition of clathrin-mediated endocytosis selectively attenuates specific insulin receptor signal transduction pathways.

    PubMed

    Ceresa, B P; Kao, A W; Santeler, S R; Pessin, J E

    1998-07-01

    To examine the role of clathrin-dependent insulin receptor internalization in insulin-stimulated signal transduction events, we expressed a dominant-interfering mutant of dynamin (K44A/dynamin) by using a recombinant adenovirus in the H4IIE hepatoma and 3T3L1 adipocyte cell lines. Expression of K44A/dynamin inhibited endocytosis of the insulin receptor as determined by both cell surface radioligand binding and trypsin protection analysis. The inhibition of the insulin receptor endocytosis had no effect on either the extent of insulin receptor autophosphorylation or insulin receptor substrate 1 (IRS1) tyrosine phosphorylation. In contrast, expression of K44A/dynamin partially inhibited insulin-stimulated Shc tyrosine phosphorylation and activation of the mitogen-activated protein kinases ERK1 and -2. Although there was an approximately 50% decrease in the insulin-stimulated activation of the phosphatidylinositol 3-kinase associated with IRS1, insulin-stimulated Akt kinase phosphorylation and activation were unaffected. The expression of K44A/dynamin increased the basal rate of amino acid transport, which was additive with the effect of insulin but had no effect on the basal or insulin-stimulated DNA synthesis. In 3T3L1 adipocytes, expression of K44A/dynamin increased the basal rate of glucose uptake, glycogen synthesis, and lipogenesis without any significant effect on insulin stimulation. Together, these data demonstrate that the acute actions of insulin are largely independent of insulin receptor endocytosis and are initiated by activation of the plasma membrane-localized insulin receptor. PMID:9632770

  17. Inhibition of Clathrin-Mediated Endocytosis Selectively Attenuates Specific Insulin Receptor Signal Transduction Pathways

    PubMed Central

    Ceresa, Brian P.; Kao, Aimee W.; Santeler, Scott R.; Pessin, Jeffrey E.

    1998-01-01

    To examine the role of clathrin-dependent insulin receptor internalization in insulin-stimulated signal transduction events, we expressed a dominant-interfering mutant of dynamin (K44A/dynamin) by using a recombinant adenovirus in the H4IIE hepatoma and 3T3L1 adipocyte cell lines. Expression of K44A/dynamin inhibited endocytosis of the insulin receptor as determined by both cell surface radioligand binding and trypsin protection analysis. The inhibition of the insulin receptor endocytosis had no effect on either the extent of insulin receptor autophosphorylation or insulin receptor substrate 1 (IRS1) tyrosine phosphorylation. In contrast, expression of K44A/dynamin partially inhibited insulin-stimulated Shc tyrosine phosphorylation and activation of the mitogen-activated protein kinases ERK1 and -2. Although there was an approximately 50% decrease in the insulin-stimulated activation of the phosphatidylinositol 3-kinase associated with IRS1, insulin-stimulated Akt kinase phosphorylation and activation were unaffected. The expression of K44A/dynamin increased the basal rate of amino acid transport, which was additive with the effect of insulin but had no effect on the basal or insulin-stimulated DNA synthesis. In 3T3L1 adipocytes, expression of K44A/dynamin increased the basal rate of glucose uptake, glycogen synthesis, and lipogenesis without any significant effect on insulin stimulation. Together, these data demonstrate that the acute actions of insulin are largely independent of insulin receptor endocytosis and are initiated by activation of the plasma membrane-localized insulin receptor. PMID:9632770

  18. Transcription factor and microRNA-regulated network motifs for cancer and signal transduction networks

    PubMed Central

    2015-01-01

    Abstract Background Molecular networks are the basis of biological processes. Such networks can be decomposed into smaller modules, also known as network motifs. These motifs show interesting dynamical behaviors, in which co-operativity effects between the motif components play a critical role in human diseases. We have developed a motif-searching algorithm, which is able to identify common motif types from the cancer networks and signal transduction networks (STNs). Some of the network motifs are interconnected which can be merged together and form more complex structures, the so-called coupled motif structures (CMS). These structures exhibit mixed dynamical behavior, which may lead biological organisms to perform specific functions. Results In this study, we integrate transcription factors (TFs), microRNAs (miRNAs), miRNA targets and network motifs information to build the cancer-related TF-miRNA-motif networks (TMMN). This allows us to examine the role of network motifs in cancer formation at different levels of regulation, i.e. transcription initiation (TF → miRNA), gene-gene interaction (CMS), and post-transcriptional regulation (miRNA → target genes). Among the cancer networks and STNs we considered, it is found that there is a substantial amount of crosstalking through motif interconnections, in particular, the crosstalk between prostate cancer network and PI3K-Akt STN. Conclusions To validate the role of network motifs in cancer formation, several examples are presented which demonstrated the effectiveness of the present approach. A web-based platform has been set up which can be accessed at: http://ppi.bioinfo.asia.edu.tw/pathway/. It is very likely that our results can supply very specific CMS missing information for certain cancer types, it is an indispensable tool for cancer biology research. PMID:25707690

  19. Calcium specificity signaling mechanisms in abscisic acid signal transduction in Arabidopsis guard cells.

    PubMed

    Brandt, Benjamin; Munemasa, Shintaro; Wang, Cun; Nguyen, Desiree; Yong, Taiming; Yang, Paul G; Poretsky, Elly; Belknap, Thomas F; Waadt, Rainer; Alemán, Fernando; Schroeder, Julian I

    2015-01-01

    A central question is how specificity in cellular responses to the eukaryotic second messenger Ca(2+) is achieved. Plant guard cells, that form stomatal pores for gas exchange, provide a powerful system for in depth investigation of Ca(2+)-signaling specificity in plants. In intact guard cells, abscisic acid (ABA) enhances (primes) the Ca(2+)-sensitivity of downstream signaling events that result in activation of S-type anion channels during stomatal closure, providing a specificity mechanism in Ca(2+)-signaling. However, the underlying genetic and biochemical mechanisms remain unknown. Here we show impairment of ABA signal transduction in stomata of calcium-dependent protein kinase quadruple mutant plants. Interestingly, protein phosphatase 2Cs prevent non-specific Ca(2+)-signaling. Moreover, we demonstrate an unexpected interdependence of the Ca(2+)-dependent and Ca(2+)-independent ABA-signaling branches and the in planta requirement of simultaneous phosphorylation at two key phosphorylation sites in SLAC1. We identify novel mechanisms ensuring specificity and robustness within stomatal Ca(2+)-signaling on a cellular, genetic, and biochemical level. PMID:26192964

  20. Tackling the bottleneck in bacterial signal transduction research: high-throughput identification of signal molecules.

    PubMed

    Krell, Tino

    2015-05-01

    Signal transduction processes are typically initiated by the interaction of signal molecules with sensor domains. The current lack of information on the signal molecules that feed into regulatory circuits forms a major bottleneck that hampers the understanding of regulatory processes. McKellar et al. report a high-throughput approach for the identification of signal molecules, which is based on thermal shift assays of recombinant sensor domains in the absence and presence of compounds from commercially available ligand collections. Initial binding studies with the sensor domain of the PctA chemoreceptor of Pseudomonas aeruginosa showed a close match between thermal shift assay results and microcalorimetric studies reported previously. Using thermal shift assays the authors then identify signals that bind to three chemoreceptors of the kiwifruit pathogen P. syringae pv. Actinidiae NZ-V13. Microcalorimetric binding studies and chemotaxis assays have validated the relevance of these ligands. The power of this technique lies in the combination of a high-throughput analytical tool with commercially available compound collections. The approach reported is universal since it can be employed to identify signal molecules to any type of sensor domain. There is no doubt that this technique will facilitate the identification of many signal molecules in future years. PMID:25708679

  1. Calcium specificity signaling mechanisms in abscisic acid signal transduction in Arabidopsis guard cells

    PubMed Central

    Brandt, Benjamin; Munemasa, Shintaro; Wang, Cun; Nguyen, Desiree; Yong, Taiming; Yang, Paul G; Poretsky, Elly; Belknap, Thomas F; Waadt, Rainer; Alemán, Fernando; Schroeder, Julian I

    2015-01-01

    A central question is how specificity in cellular responses to the eukaryotic second messenger Ca2+ is achieved. Plant guard cells, that form stomatal pores for gas exchange, provide a powerful system for in depth investigation of Ca2+-signaling specificity in plants. In intact guard cells, abscisic acid (ABA) enhances (primes) the Ca2+-sensitivity of downstream signaling events that result in activation of S-type anion channels during stomatal closure, providing a specificity mechanism in Ca2+-signaling. However, the underlying genetic and biochemical mechanisms remain unknown. Here we show impairment of ABA signal transduction in stomata of calcium-dependent protein kinase quadruple mutant plants. Interestingly, protein phosphatase 2Cs prevent non-specific Ca2+-signaling. Moreover, we demonstrate an unexpected interdependence of the Ca2+-dependent and Ca2+-independent ABA-signaling branches and the in planta requirement of simultaneous phosphorylation at two key phosphorylation sites in SLAC1. We identify novel mechanisms ensuring specificity and robustness within stomatal Ca2+-signaling on a cellular, genetic, and biochemical level. DOI: http://dx.doi.org/10.7554/eLife.03599.001 PMID:26192964

  2. Polyunsaturated fatty acids affect the localization and signaling of PIP3/AKT in prostate cancer cells.

    PubMed

    Gu, Zhennan; Wu, Jiansheng; Wang, Shihua; Suburu, Janel; Chen, Haiqin; Thomas, Michael J; Shi, Lihong; Edwards, Iris J; Berquin, Isabelle M; Chen, Yong Q

    2013-09-01

    AKT is a serine-threonine protein kinase that plays important roles in cell growth, proliferation and apoptosis. It is activated after binding to phosphatidylinositol phosphates (PIPs) with phosphate groups at positions 3,4 and 3,4,5 on the inositol ring. In spite of extensive research on AKT, one aspect has been largely overlooked, namely the role of the fatty acid chains on PIPs. PIPs are phospholipids composed of a glycerol backbone with fatty acids at the sn-1 and sn-2 position and inositol at the sn-3 position. Here, we show that polyunsaturated fatty acids (PUFAs) modify phospholipid content. Docosahexaenoic acid (DHA), an ω3 PUFA, can replace the fatty acid at the sn-2 position of the glycerol backbone, thereby changing the species of phospholipids. DHA also inhibits AKT(T308) but not AKT(S473) phosphorylation, alters PI(3,4,5)P3 (PIP3) and phospho-AKT(S473) protein localization, decreases pPDPK1(S241)-AKT and AKT-BAD interaction and suppresses prostate tumor growth. Our study highlights a potential novel mechanism of cancer inhibition by ω3 PUFA through alteration of PIP3 and AKT localization and affecting the AKT signaling pathway. PMID:23633519

  3. Regulation of Akt signaling by Sirtuins: Its implication in cardiac hypertrophy and aging

    PubMed Central

    Pillai, Vinodkumar B.; Sundaresan, Nagalingam R.; Gupta, Mahesh P.

    2014-01-01

    Cardiac hypertrophy is a multifactorial disease characterized by multiple molecular alterations. One of these alterations is change in activity of Akt, which plays a central role in regulating a variety of cellular processes ranging from cell survival to aging. Akt activation is mainly achieved by its binding to phosphatidylinositol 3,4,5 triphosphate (PIP3). This results in a conformational change that exposes the kinase domain of Akt for phosphorylation and activation by its upstream kinase PDK1 in the cell membrane. Recent studies have shown that sirtuin isoforms SIRT1, SIRT3 and SIRT6 play an essential role in the regulation of Akt activation. While SIRT1 deacetylates Akt to promote PIP3 binding and activation, SIRT3 controls ROS-mediated Akt activation and SIRT6 transcriptionally represses Akt at the level of chromatin. In the first part of this review, we discuss the mechanisms by which sirtuins regulate Akt activation and how they influence other post-translational modifications of Akt. In the latter part of the review, we summarize the implications of sirtuin-dependent regulation of Akt signaling in the control of major cellular processes like cellular growth, angiogenesis, apoptosis, autophagy and aging; which are involved in the initiation and progression of several diseases. PMID:24436432

  4. Homocysteine enhances MMP-9 production in murine macrophages via ERK and Akt signaling pathways

    SciTech Connect

    Lee, Seung Jin; Lee, Yi Sle; Seo, Kyo Won; Bae, Jin Ung; Kim, Gyu Hee; Park, So Youn; Kim, Chi Dae

    2012-04-01

    Homocysteine (Hcy) at elevated levels is an independent risk factor of cardiovascular diseases, including atherosclerosis. In the present study, we investigated the effect of Hcy on the production of matrix metalloproteinases (MMP) in murine macrophages. Among the MMP known to regulate the activities of collagenase and gelatinase, Hcy exclusively increased the gelatinolytic activity of MMP-9 in J774A.1 cells as well as in mouse peritoneal macrophages. Furthermore, this activity was found to be correlated with Western blot findings in J774A.1 cells, which showed that MMP-9 expression was concentration- and time-dependently increased by Hcy. Inhibition of the ERK and Akt pathways led to a significant decrease in Hcy-induced MMP-9 expression, and combined treatment with inhibitors of the ERK and Akt pathways showed an additive effects. Activity assays for ERK and Akt showed that Hcy increased the phosphorylation of both, but these phosphorylation were not affected by inhibitors of the Akt and ERK pathways. In line with these findings, the molecular inhibition of ERK and Akt using siRNA did not affect the Hcy-induced phosphorylation of Akt and ERK, respectively. Taken together, these findings suggest that Hcy enhances MMP-9 production in murine macrophages by separately activating the ERK and Akt signaling pathways. -- Highlights: ► Homocysteine (Hcy) induced MMP-9 production in murine macrophages. ► Hcy induced MMP-9 production through ERK and Akt signaling pathways. ► ERK and Akt signaling pathways were activated by Hcy in murine macrophages. ► ERK and Akt pathways were additively act on Hcy-induced MMP-9 production. ► Hcy enhances MMP-9 production in macrophages via activation of ERK and Akt signaling pathways in an independent manner.

  5. [Cellular signal transduction pathways in cardiac hypertrophy and heart failure].

    PubMed

    Lewartowski, Bohdan; Mackiewicz, Urszula

    2006-10-01

    Cardiac hypertrophy and heart failure are characterized by significant changes of expression and function of many proteins. These changes are responsible for arrhythmias and haemodynamic disturbances. They are initiated by increased cardiac load, detected by cellular mechanoreceptors, and by activation of sarcolemmal chemoreceptors in myocytes and fibroblasts. In the present paper the authors describe the structure and function of molecular cellular pathways for transmission of the information generated by receptors to the nucleus, where it modifies the expression of genes coding for cellular proteins. The authors describe in detail: structure and function of Z-discs and integrins working as mechanoreceptors, calcineurin/NFAT pathways, MAP kinases pathways, pathway activated by AT1 receptors: protein kinase C pathways, AKT/mTHOR kinase pathway and EGRF/ERK1,2 pathway. Functional relationships between pathways mentioned and the results of studies analysing their role in cardiac hypertrophy and heart failure are also presented. PMID:20527382

  6. Angiogenin interacts with ribonuclease inhibitor regulating PI3K/AKT/mTOR signaling pathway in bladder cancer cells.

    PubMed

    Peng, Yuan; Li, Lin; Huang, Mengge; Duan, Changzhu; Zhang, Luyu; Chen, Junxia

    2014-12-01

    Angiogenin (ANG), a member of RNase A superfamily, is the only angiogenic factor that possesses ribonucleolytic activity. Recent studies showed that the expression of ANG was elevated in various types of cancers. Accumulating evidence indicates that ANG plays an essential role in cancer progression by stimulating both cancer cell proliferation and tumor angiogenesis. Human ribonuclease inhibitor (RI), a cytoplasmic protein, is constructed almost entirely of leucine rich repeats (LRRs), which are present in a large family of proteins that are distinguished by their display of vast surface areas to foster protein-protein interactions. RI might be involved in unknown biological effects except inhibiting RNase A activity. The experiment demonstrated that RI also could suppress activity of angiogenin (ANG) through closely combining with it in vitro. PI3K/AKT/mTOR signaling pathway exerts a key role in cell growth, survival, proliferation, apoptosis and angiogenesis. We recently reported that up-regulating RI inhibited the growth and induced apoptosis of murine melanoma cells through repression of angiogenin and PI3K/AKT signaling pathway. However, ANG receptors have not yet been identified to date, its related signal transduction pathways are not fully clear and underlying interacting mechanisms between RI and ANG remain largely unknown. Therefore, we hypothesize that RI might combine with intracellular ANG to block its nuclear translocation and regulate PI3K/AKT/mTOR signaling pathway to inhibit biological functions of ANG. Here, we reported for the first time that ANG could interact with RI endogenously and exogenously by using co-immunoprecipitation (Co-IP) and GST pull-down. Furthermore, we observed the colocalization of ANG and RI in cells with immunofluorescence staining under laser confocal microscope. Moreover, through fluorescence resonance energy transfer (FRET) assay, we further confirmed that these two proteins have a physical interaction in living cells

  7. VEGF-A isoforms program differential VEGFR2 signal transduction, trafficking and proteolysis

    PubMed Central

    Fearnley, Gareth W.; Smith, Gina A.; Abdul-Zani, Izma; Yuldasheva, Nadira; Mughal, Nadeem A.; Homer-Vanniasinkam, Shervanthi; Kearney, Mark T.; Zachary, Ian C.; Tomlinson, Darren C.; Harrison, Michael A.; Wheatcroft, Stephen B.; Ponnambalam, Sreenivasan

    2016-01-01

    ABSTRACT Vascular endothelial growth factor A (VEGF-A) binding to the receptor tyrosine kinase VEGFR2 triggers multiple signal transduction pathways, which regulate endothelial cell responses that control vascular development. Multiple isoforms of VEGF-A can elicit differential signal transduction and endothelial responses. However, it is unclear how such cellular responses are controlled by isoform-specific VEGF-A–VEGFR2 complexes. Increasingly, there is the realization that the membrane trafficking of receptor–ligand complexes influences signal transduction and protein turnover. By building on these concepts, our study shows for the first time that three different VEGF-A isoforms (VEGF-A165, VEGF-A121 and VEGF-A145) promote distinct patterns of VEGFR2 endocytosis for delivery into early endosomes. This differential VEGFR2 endocytosis and trafficking is linked to VEGF-A isoform-specific signal transduction events. Disruption of clathrin-dependent endocytosis blocked VEGF-A isoform-specific VEGFR2 activation, signal transduction and caused substantial depletion in membrane-bound VEGFR1 and VEGFR2 levels. Furthermore, such VEGF-A isoforms promoted differential patterns of VEGFR2 ubiquitylation, proteolysis and terminal degradation. Our study now provides novel insights into how different VEGF-A isoforms can bind the same receptor tyrosine kinase and elicit diverse cellular outcomes. PMID:27044325

  8. VEGF-A isoforms program differential VEGFR2 signal transduction, trafficking and proteolysis.

    PubMed

    Fearnley, Gareth W; Smith, Gina A; Abdul-Zani, Izma; Yuldasheva, Nadira; Mughal, Nadeem A; Homer-Vanniasinkam, Shervanthi; Kearney, Mark T; Zachary, Ian C; Tomlinson, Darren C; Harrison, Michael A; Wheatcroft, Stephen B; Ponnambalam, Sreenivasan

    2016-01-01

    Vascular endothelial growth factor A (VEGF-A) binding to the receptor tyrosine kinase VEGFR2 triggers multiple signal transduction pathways, which regulate endothelial cell responses that control vascular development. Multiple isoforms of VEGF-A can elicit differential signal transduction and endothelial responses. However, it is unclear how such cellular responses are controlled by isoform-specific VEGF-A-VEGFR2 complexes. Increasingly, there is the realization that the membrane trafficking of receptor-ligand complexes influences signal transduction and protein turnover. By building on these concepts, our study shows for the first time that three different VEGF-A isoforms (VEGF-A165, VEGF-A121 and VEGF-A145) promote distinct patterns of VEGFR2 endocytosis for delivery into early endosomes. This differential VEGFR2 endocytosis and trafficking is linked to VEGF-A isoform-specific signal transduction events. Disruption of clathrin-dependent endocytosis blocked VEGF-A isoform-specific VEGFR2 activation, signal transduction and caused substantial depletion in membrane-bound VEGFR1 and VEGFR2 levels. Furthermore, such VEGF-A isoforms promoted differential patterns of VEGFR2 ubiquitylation, proteolysis and terminal degradation. Our study now provides novel insights into how different VEGF-A isoforms can bind the same receptor tyrosine kinase and elicit diverse cellular outcomes. PMID:27044325

  9. Akt1 signaling coordinates BMP signaling and β-catenin activity to regulate second heart field progenitor development.

    PubMed

    Luo, Wen; Zhao, Xia; Jin, Hengwei; Tao, Lichan; Zhu, Jingai; Wang, Huijuan; Hemmings, Brian A; Yang, Zhongzhou

    2015-02-15

    Second heart field (SHF) progenitors exhibit continued proliferation and delayed differentiation, which are modulated by FGF4/8/10, BMP and canonical Wnt/β-catenin signaling. PTEN-Akt signaling regulates the stem cell/progenitor cell homeostasis in several systems, such as hematopoietic stem cells, intestinal stem cells and neural progenitor cells. To address whether PTEN-Akt signaling is involved in regulating cardiac progenitors, we deleted Pten in SHF progenitors. Deletion of Pten caused SHF expansion and increased the size of the SHF derivatives, the right ventricle and the outflow tract. Cell proliferation of cardiac progenitors was enhanced, whereas cardiac differentiation was unaffected by Pten deletion. Removal of Akt1 rescued the phenotype and early lethality of Pten deletion mice, suggesting that Akt1 was the key downstream target that was negatively regulated by PTEN in cardiac progenitors. Furthermore, we found that inhibition of FOXO by Akt1 suppressed the expression of the gene encoding the BMP ligand (BMP7), leading to dampened BMP signaling in the hearts of Pten deletion mice. Cardiac activation of Akt also increased the Ser552 phosphorylation of β-catenin, thus enhancing its activity. Reducing β-catenin levels could partially rescue heart defects of Pten deletion mice. We conclude that Akt signaling regulates the cell proliferation of SHF progenitors through coordination of BMP signaling and β-catenin activity. PMID:25670795

  10. Increased Akt signaling in the mosquito fat body increases adult survivorship.

    PubMed

    Arik, Anam J; Hun, Lewis V; Quicke, Kendra; Piatt, Michael; Ziegler, Rolf; Scaraffia, Patricia Y; Badgandi, Hemant; Riehle, Michael A

    2015-04-01

    Akt signaling regulates diverse physiologies in a wide range of organisms. We examine the impact of increased Akt signaling in the fat body of 2 mosquito species, the Asian malaria mosquito Anopheles stephensi and the yellow fever mosquito Aedes aegypti. Overexpression of a myristoylated and active form of A. stephensi and Ae. aegypti Akt in the fat body of transgenic mosquitoes led to activation of the downstream signaling molecules forkhead box O (FOXO) and p70 S6 kinase in a tissue and blood meal-specific manner. In both species, increased Akt signaling in the fat body after blood feeding significantly increased adult survivorship relative to nontransgenic sibling controls. In A. stephensi, survivorship was increased by 15% to 45%, while in Ae. aegypti, it increased 14% to 47%. Transgenic mosquitoes fed only sugar, and thus not expressing active Akt, had no significant difference in survivorship relative to nontransgenic siblings. Expression of active Akt also increased expression of fat body vitellogenin, but the number of viable eggs did not differ significantly between transgenic and nontransgenic controls. This work demonstrates a novel mechanism of enhanced survivorship through increased Akt signaling in the fat bodies of multiple mosquito genera and provides new tools to unlock the molecular underpinnings of aging in eukaryotic organisms. PMID:25550465

  11. AKT-independent signaling downstream of oncogenic PIK3CA mutations in human cancer.

    PubMed

    Vasudevan, Krishna M; Barbie, David A; Davies, Michael A; Rabinovsky, Rosalia; McNear, Chontelle J; Kim, Jessica J; Hennessy, Bryan T; Tseng, Hsiuyi; Pochanard, Panisa; Kim, So Young; Dunn, Ian F; Schinzel, Anna C; Sandy, Peter; Hoersch, Sebastian; Sheng, Qing; Gupta, Piyush B; Boehm, Jesse S; Reiling, Jan H; Silver, Serena; Lu, Yiling; Stemke-Hale, Katherine; Dutta, Bhaskar; Joy, Corwin; Sahin, Aysegul A; Gonzalez-Angulo, Ana Maria; Lluch, Ana; Rameh, Lucia E; Jacks, Tyler; Root, David E; Lander, Eric S; Mills, Gordon B; Hahn, William C; Sellers, William R; Garraway, Levi A

    2009-07-01

    Dysregulation of the phosphatidylinositol 3-kinase (PI3K) signaling pathway occurs frequently in human cancer. PTEN tumor suppressor or PIK3CA oncogene mutations both direct PI3K-dependent tumorigenesis largely through activation of the AKT/PKB kinase. However, here we show through phosphoprotein profiling and functional genomic studies that many PIK3CA mutant cancer cell lines and human breast tumors exhibit only minimal AKT activation and a diminished reliance on AKT for anchorage-independent growth. Instead, these cells retain robust PDK1 activation and membrane localization and exhibit dependency on the PDK1 substrate SGK3. SGK3 undergoes PI3K- and PDK1-dependent activation in PIK3CA mutant cancer cells. Thus, PI3K may promote cancer through both AKT-dependent and AKT-independent mechanisms. Knowledge of differential PI3K/PDK1 signaling could inform rational therapeutics in cancers harboring PIK3CA mutations. PMID:19573809

  12. Bacterial signal transduction network in a genomic perspective†

    PubMed Central

    Galperin, Michael Y.

    2005-01-01

    Summary Bacterial signalling network includes an array of numerous interacting components that monitor environmental and intracellular parameters and effect cellular response to changes in these parameters. The complexity of bacterial signalling systems makes comparative genome analysis a particularly valuable tool for their studies. Comparative studies revealed certain general trends in the organization of diverse signalling systems. These include (i) modular structure of signalling proteins; (ii) common organization of signalling components with the flow of information from N-terminal sensory domains to the C-terminal transmitter or signal output domains (N-to-C flow); (iii) use of common conserved sensory domains by different membrane receptors; (iv) ability of some organisms to respond to one environmental signal by activating several regulatory circuits; (v) abundance of intracellular signalling proteins, typically consisting of a PAS or GAF sensor domains and various output domains; (vi) importance of secondary messengers, cAMP and cyclic diguanylate; and (vii) crosstalk between components of different signalling pathways. Experimental characterization of the novel domains and domain combinations would be needed for achieving a better understanding of the mechanisms of signalling response and the intracellular hierarchy of different signalling pathways. PMID:15142243

  13. Analysis of a signal transduction pathway involved in leaf epidermis differentiation.

    SciTech Connect

    Philip W. Becraft

    2005-05-23

    The major objective of this study was to identify and analyze signal transduction factors that function with the CR4 receptor kinase. We pursued this analysis in Arabidopsis. Analysis of other members of the ACR4 related receptor (CRR) family produced biochemical evidence consistent with some of them functioning in ACR4 signal transduction. Yeast 2-hybrid identified six proteins that interact with the cytoplasmic domain of ACR4, representing putative downstream signal transduction components. The interactions for all 6 proteins were verified by in vitro pull down assays. Five of the interacting proteins were phosphorylated by ACR4. We also identified candidate interactors with the extracellular TNFR domain. We hypothesize this may be the ligand binding domain for ACR4. In one approach, yeast 2-hybrid was again used and five candidate proteins identified. Nine additional candidates were identified in a genome wide scan of Arabidopsis amino acid sequences that threaded onto the TNF structure.

  14. An integrated signal transduction network of macrophage migration inhibitory factor.

    PubMed

    Subbannayya, Tejaswini; Variar, Prathyaksha; Advani, Jayshree; Nair, Bipin; Shankar, Subramanian; Gowda, Harsha; Saussez, Sven; Chatterjee, Aditi; Prasad, T S Keshava

    2016-06-01

    Macrophage migration inhibitory factor (MIF) is a glycosylated multi-functional protein that acts as an enzyme as well as a cytokine. MIF mediates its actions through a cell surface class II major histocompatibility chaperone, CD74 and co-receptors such as CD44, CXCR2, CXCR4 or CXCR7. MIF has been implicated in the pathogenesis of several acute and chronic inflammatory diseases. Although MIF is a molecule of biomedical importance, a public resource of MIF signaling pathway is currently lacking. In view of this, we carried out detailed data mining and documentation of the signaling events pertaining to MIF from published literature and developed an integrated reaction map of MIF signaling. This resulted in the cataloguing of 68 molecules belonging to MIF signaling pathway, which includes 24 protein-protein interactions, 44 post-translational modifications, 11 protein translocation events and 8 activation/inhibition events. In addition, 65 gene regulation events at the mRNA levels induced by MIF signaling have also been catalogued. This signaling pathway has been integrated into NetPath ( http://www.netpath.org ), a freely available human signaling pathway resource developed previously by our group. The MIF pathway data is freely available online in various community standard data exchange formats. We expect that data on signaling events and a detailed signaling map of MIF will provide the scientific community with an improved platform to facilitate further molecular as well as biomedical investigations on MIF. PMID:27139435

  15. Signal Transduction Pathways that Regulate CAB Gene Expression

    SciTech Connect

    Chory, Joanne

    2006-01-16

    The process of chloroplast differentiation, involves the coordinate regulation of many nuclear and chloroplast genes. The cues for the initiation of this developmental program are both extrinsic (e.g., light) and intrinsic (cell-type and plastid signals). During this project period, we utilized a molecular genetic approach to select for Arabidopsis mutants that did not respond properly to environmental light conditions, as well as mutants that were unable to perceive plastid damage. These latter mutants, called gun mutants, define two retrograde signaling pathways that regulate nuclear gene expression in response to chloroplasts. A major finding was to identify a signal from chloroplasts that regulates nuclear gene transcription. This signal is the build-up of Mg-Protoporphyrin IX, a key intermediate of the chlorophyll biosynthetic pathway. The signaling pathways downstream of this signal are currently being studied. Completion of this project has provided an increased understanding of the input signals and retrograde signaling pathways that control nuclear gene expression in response to the functional state of chloroplasts. These studies should ultimately influence our abilities to manipulate plant growth and development, and will aid in the understanding of the developmental control of photosynthesis.

  16. Signal Transduction Pathways that Regulate CAB Gene Expression

    SciTech Connect

    Chory, Joanne

    2004-12-31

    The process of chloroplast differentiation, involves the coordinate regulation of many nuclear and chloroplast genes. The cues for the initiation of this developmental program are both extrinsic (e.g., light) and intrinsic (cell-type and plastid signals). During this project period, we utilized a molecular genetic approach to select for Arabidopsis mutants that did not respond properly to environmental light conditions, as well as mutants that were unable to perceive plastid damage. These latter mutants, called gun mutants, define two retrograde signaling pathways that regulate nuclear gene expression in response to chloroplasts. A major finding was to identify a signal from chloroplasts that regulates nuclear gene transcription. This signal is the build-up of Mg-Protoporphyrin IX, a key intermediate of the chlorophyll biosynthetic pathway. The signaling pathways downstream of this signal are currently being studied. Completion of this project has provided an increased understanding of the input signals and retrograde signaling pathways that control nuclear gene expression in response to the functional state of chloroplasts. These studies should ultimately influence our abilities to manipulate plant growth and development, and will aid in the understanding of the developmental control of photosynthesis.

  17. Molecular mechanisms of gravity perception and signal transduction in plants.

    PubMed

    Kolesnikov, Yaroslav S; Kretynin, Serhiy V; Volotovsky, Igor D; Kordyum, Elizabeth L; Ruelland, Eric; Kravets, Volodymyr S

    2016-07-01

    Gravity is one of the environmental cues that direct plant growth and development. Recent investigations of different gravity signalling pathways have added complexity to how we think gravity is perceived. Particular cells within specific organs or tissues perceive gravity stimulus. Many downstream signalling events transmit the perceived information into subcellular, biochemical, and genomic responses. They are rapid, non-genomic, regulatory, and cell-specific. The chain of events may pass by signalling lipids, the cytoskeleton, intracellular calcium levels, protein phosphorylation-dependent pathways, proteome changes, membrane transport, vacuolar biogenesis mechanisms, or nuclear events. These events culminate in changes in gene expression and auxin lateral redistribution in gravity response sites. The possible integration of these signalling events with amyloplast movements or with other perception mechanisms is discussed. Further investigation is needed to understand how plants coordinate mechanisms and signals to sense this important physical factor. PMID:26215561

  18. Structural mechanism for signal transduction in RXR nuclear receptor heterodimers

    PubMed Central

    Kojetin, Douglas J.; Matta-Camacho, Edna; Hughes, Travis S.; Srinivasan, Sathish; Nwachukwu, Jerome C.; Cavett, Valerie; Nowak, Jason; Chalmers, Michael J.; Marciano, David P.; Kamenecka, Theodore M.; Shulman, Andrew I.; Rance, Mark; Griffin, Patrick R.; Bruning, John B.; Nettles, Kendall W.

    2015-01-01

    A subset of nuclear receptors (NRs) function as obligate heterodimers with retinoid X receptor (RXR), allowing integration of ligand-dependent signals across the dimer interface via an unknown structural mechanism. Using nuclear magnetic resonance (NMR) spectroscopy, x-ray crystallography and hydrogen/deuterium exchange (HDX) mass spectrometry, here we show an allosteric mechanism through which RXR co-operates with a permissive dimer partner, peroxisome proliferator-activated receptor (PPAR)-γ, while rendered generally unresponsive by a non-permissive dimer partner, thyroid hormone (TR) receptor. Amino acid residues that mediate this allosteric mechanism comprise an evolutionarily conserved network discovered by statistical coupling analysis (SCA). This SCA network acts as a signalling rheostat to integrate signals between dimer partners, ligands and coregulator-binding sites, thereby affecting signal transmission in RXR heterodimers. These findings define rules guiding how NRs integrate two ligand-dependent signalling pathways into RXR heterodimer-specific responses. PMID:26289479

  19. Signal Transduction in the Chronic Leukemias: Implications for Targeted Therapies

    PubMed Central

    Ahmed, Wesam; Van Etten, Richard A.

    2013-01-01

    The chronic leukemias, including chronic myeloid leukemia (CML), the Philadelphia-negative myeloproliferative neoplasms (MPNs), and chronic lymphocytic leukemia (CLL), have been characterized extensively for abnormalities of cellular signaling pathways. This effort has led to the elucidation of the central role of dysregulated tyrosine kinase signaling in the chronic myeloid neoplasms and of constitutive B-cell receptor signaling in CLL. This, in turn, has stimulated the development of small molecule inhibitors of these signaling pathways for therapy of chronic leukemia. Although the field is still in its infancy, the clinical results with these agents have ranged from encouraging (CLL) to spectacular (CML). In this review, we summarize recent studies that have helped to define the signaling pathways critical to the pathogenesis of the chronic leukemias. We also discuss correlative studies emerging from clinical trials of drugs targeting these pathways. PMID:23307472

  20. Regulation of neutrophil apoptosis by modulation of PKB/Akt activation.

    PubMed

    Rane, Madhavi J; Klein, Jon B

    2009-01-01

    The serine/threonine kinase, Akt, also known as PKB (Protein Kinase B) is one important signal transduction pathway that mediates the delay of neutrophil apoptosis caused by inflammatory mediators. Proteins controlled by the PKB/Akt pathway have been reported to prevent or reverse apoptotic-signaling pathways and regulate cell survival. In this review we discuss the role of PKB/Akt activation in the regulation of neutrophil activation during inflammation, and the importance of resolving the inflammatory response by inhibiting PKB/Akt activation and neutrophil survival. Furthermore, we introduce the concept of a dynamic Akt signal complex that is altered when an extracellular signal is initiated such that changes in protein-protein interactions within the Akt signal complex regulates Akt activity and cell survival. Various substrates of PKB/Akt as well as positive and negative regulators of PKB/Akt activation are discussed which in turn inhibit or enhance cellular survival. PMID:19273208

  1. Exploring signal transduction in heteromultimeric protein based on energy dissipation model.

    PubMed

    Ma, Cheng-Wei; Xiu, Zhi-Long; Zeng, An-Ping

    2015-01-01

    Dynamic intersubunit interactions are key elements in the regulation of many biological systems. A better understanding of how subunits interact with each other and how their interactions are related to dynamic protein structure is a fundamental task in biology. In this paper, a heteromultimeric allosteric protein, Corynebacterium glutamicum aspartokinase, is used as a model system to explore the signal transduction involved in intersubunit interactions and allosteric communication with an emphasis on the intersubunit signaling process. For this purpose, energy dissipation simulation and network construction are conducted for each subunit and the whole protein. Comparison with experimental results shows that the new approach is able to predict all the mutation sites that have been experimentally proved to desensitize allosteric regulation of the enzyme. Additionally, analysis revealed that the function of the effector threonine is to facilitate the binding of the two subunits without contributing to the allosteric communication. During the allosteric regulation upon the binding of the effector lysine, signals can be transferred from the β-subunit to the catalytic site of the α-subunit through both a direct way of intersubunit signal transduction, and an indirect way: first, to the regulatory region of the α-subunit by intersubunit signal transduction and then to the catalytic region by intramolecular signal transduction. Therefore, the new approach is able to illustrate the diversity of the underlying mechanisms when the strength of feedback inhibition by the effector(s) is modulated, providing useful information that has potential applications in engineering heteromultimeric allosteric regulation. PMID:24279729

  2. 17β-estradiol activates mTOR in chondrocytes by AKT-dependent and AKT-independent signaling pathways

    PubMed Central

    Tao, Yulei; Sun, Haibiao; Sun, Hongyan; Qiu, Xianxing; Xu, Changbo; Shi, Changxiu; Du, Jiahui

    2015-01-01

    To confirm whether 17β-estradiol (E2) activates mammalian target of rapamycin (mTOR) signaling pathway in chondrocytes and in what way activates mTOR. Human immortalized chondrocytes cell lines TC28a2 and C28/I2 were subjected to incubate with or without E2, LY294002 (the inhibitor of PI3K), rapamycin (the inhibitor of mTOR), or E2 in combination with LY294002 or rapamycin. Thereafter, protein levels of S6K1, p-S6K1, protein kinase B (AKT), and p-AKT were determined by Western blot analysis. Matrix metallopeptidase (MMP) 3 or MMP13 mRNA levels were evaluated by quantitative real-time PCR (qRT-PCR). Co-immunoprecipitation and Western blot analysis were performed to verify the interaction between ERα and mTOR. Both p-S6K1 and p-AKT protein levels in TC28a2 and C28/I2E2 cells were significantly increased by incubation with E2 (0.5 h and 1 h) (P < 0.05). Rapamycin did not affect the levels of p-AKT, but were significantly reduced by LY294002 or E2 in combination with LY294002. The levels of p-S6K1 were significantly decreased by incubation with LY294002, but the effect could be reversed by E2 in combination with LY294002. Rabbit anti-mTOR antibody was able to immunoprecipitate ERα after incubation with E2. Moreover, E2 inhibited the mRNA levels of MMP3 and MMP13 by mTOR pathway. E2 actives mTOR in chondrocytes through AKT-dependent and independent ways. PMID:26884863

  3. Naringenin suppresses TPA-induced tumor invasion by suppressing multiple signal transduction pathways in human hepatocellular carcinoma cells.

    PubMed

    Yen, Hung-Rong; Liu, Ching-Ju; Yeh, Chia-Chou

    2015-06-25

    Naringenin, a common dietary flavonoid abundantly present in fruits and vegetables, is believed to possess strong anti-proliferative properties and the ability to induce apoptosis in hepatoma cell lines. However, there are no reports describing its effects on the invasion and metastasis of hepatoma cell lines, and the detailed molecular mechanisms of its effects are still unclear. In this study, we investigated the mechanisms underlying naringenin-mediated inhibition of 12-O-Tetradecanoylphorbol-13-acetate (TPA)-induced cell invasion and inhibition of secreted and cytosolic MMP-9 production in human hepatoma cells (HepG2, Huh-7, and HA22T) and murine embryonic liver cells (BNL CL2). Naringenin suppressed MMP-9 transcription by inhibiting activator protein (AP)-1 and nuclear factor-κB (NF-κB) activity. It suppressed TPA-induced AP-1 activity through inhibiting the phosphorylation of the extracellular signal-related kinase (ERK) and c-Jun N-terminal kinase (JNK) signaling pathways, and it suppressed TPA-induced inhibition of NF-κB nuclear translocation through IκB. Additionally, it suppressed TPA-induced activation of ERK/phosphatidylinositol 3-kinase/Akt upstream of NF-κB and AP-1. These data suggest that naringenin suppresses the invasiveness and metastatic potential of hepatocellular carcinoma (HCC) by inhibiting multiple signal transduction pathways. PMID:25866363

  4. Signal Transduction: Turning a Switch into a Rheostat

    PubMed Central

    Bardwell, Lee

    2010-01-01

    MAP kinase cascades are inherently switch-like, but, during yeast mating, MAPK signaling is graded. A new study suggests that the Ste5 scaffold protein is responsible for making this switch less switch-like. PMID:18957235

  5. The MiST2 database: a comprehensive genomics resource on microbial signal transduction

    PubMed Central

    Ulrich, Luke E.; Zhulin, Igor B.

    2010-01-01

    The MiST2 database (http://mistdb.com) identifies and catalogs the repertoire of signal transduction proteins in microbial genomes. Signal transduction systems regulate the majority of cellular activities including the metabolism, development, host-recognition, biofilm production, virulence, and antibiotic resistance of human pathogens. Thus, knowledge of the proteins and interactions that comprise these communication networks is an essential component to furthering biomedical discovery. These are identified by searching protein sequences for specific domain profiles that implicate a protein in signal transduction. Compared to the previous version of the database, MiST2 contains a host of new features and improvements including the following: draft genomes; extracytoplasmic function (ECF) sigma factor protein identification; enhanced classification of signaling proteins; novel, high-quality domain models for identifying histidine kinases and response regulators; neighboring two-component genes; gene cart; better search capabilities; enhanced taxonomy browser; advanced genome browser; and a modern, biologist-friendly web interface. MiST2 currently contains 966 complete and 157 draft bacterial and archaeal genomes, which collectively contain more than 245 000 signal transduction proteins. The majority (66%) of these are one-component systems, followed by two-component proteins (26%), chemotaxis (6%), and finally ECF factors (2%). PMID:19900966

  6. The relation of signal transduction to the sensitivity and dynamic range of bacterial chemotaxis.

    PubMed

    Namba, Toshinori; Nishikawa, Masatoshi; Shibata, Tatsuo

    2012-09-19

    Complex networks of interacting molecular components of living cells are responsible for many important processes, such as signal processing and transduction. An important challenge is to understand how the individual properties of these molecular interactions and biochemical transformations determine the system-level properties of biological functions. Here, we address the issue of the accuracy of signal transduction performed by a bacterial chemotaxis system. The chemotaxis sensitivity of bacteria to a chemoattractant gradient has been measured experimentally from bacterial aggregation in a chemoattractant-containing capillary. The observed precision of the chemotaxis depended on environmental conditions such as the concentration and molecular makeup of the chemoattractant. In a quantitative model, we derived the chemotactic response function, which is essential to describing the signal transduction process involved in bacterial chemotaxis. In the presence of a gradient, an analytical solution is derived that reveals connections between the chemotaxis sensitivity and the characteristics of the signaling system, such as reaction rates. These biochemical parameters are integrated into two system-level parameters: one characterizes the efficiency of gradient sensing, and the other is related to the dynamic range of chemotaxis. Thus, our approach explains how a particular signal transduction property affects the system-level performance of bacterial chemotaxis. We further show that the two parameters can be derived from published experimental data from a capillary assay, which successfully characterizes the performance of bacterial chemotaxis. PMID:22995512

  7. Phospholipase D2 Mediates Survival Signaling through Direct Regulation of Akt in Glioblastoma Cells*♦

    PubMed Central

    Bruntz, Ronald C.; Taylor, Harry E.; Lindsley, Craig W.; Brown, H. Alex

    2014-01-01

    The lack of innovative drug targets for glioblastoma multiforme (GBM) limits patient survival to approximately 1 year following diagnosis. The pro-survival kinase Akt provides an ideal target for the treatment of GBM as Akt signaling is frequently activated in this cancer type. However, the central role of Akt in physiological processes limits its potential as a therapeutic target. In this report, we show that the lipid-metabolizing enzyme phospholipase D (PLD) is a novel regulator of Akt in GBM. Studies using a combination of small molecule PLD inhibitors and siRNA knockdowns establish phosphatidic acid, the product of the PLD reaction, as an essential component for the membrane recruitment and activation of Akt. Inhibition of PLD enzymatic activity and subsequent Akt activation decreases GBM cell viability by specifically inhibiting autophagic flux. We propose a mechanism whereby phosphorylation of beclin1 by Akt prevents binding of Rubicon (RUN domain cysteine-rich domain containing beclin1-interacting protein), an interaction known to inhibit autophagic flux. These findings provide a novel framework through which Akt inhibition can be achieved without directly targeting the kinase. PMID:24257753

  8. Hipk proteins dually regulate Wnt/Wingless signal transduction.

    PubMed

    Verheyen, Esther M; Swarup, Sharan; Lee, Wendy

    2012-01-01

    The Wnt/Wingless (Wg) pathway is an evolutionarily conserved signaling system that is used reiteratively, both spatially and temporally, to control the development of multicellular animals. The stability of cytoplasmic β-catenin/Armadillo, the transcriptional effector of the pathway, is controlled by sequential N-terminal phosphorylation and ubiquitination that targets it for proteasome-mediated degradation. Orthologous members of the Homeodomain-interacting protein kinase family from Drosophila to vertebrates have been implicated in the regulation of Wnt/Wingless signaling. In Drosophila, as a consequence of Hipk activity, cells accumulate stabilized Armadillo that directs the expression of Wg-specific target genes. Hipk promotes the stabilization of Armadillo by inhibiting its ubiquitination (and hence subsequent degradation) by the SCF(Slimb) E3 ubiquitin ligase complex. Vertebrate Hipk2 impedes β-catenin ubiquitination to promote its stability and the Wnt signal in a mechanism that is functionally conserved. Moreover, we describe here that Hipk proteins have a role independent of their effect on β-catenin/Armadillo stability to enhance Wnt/Wingless signaling. PMID:22634475

  9. Signal Transduction Pathways of EMT Induced by TGF-β, SHH, and WNT and Their Crosstalks

    PubMed Central

    Zhang, Jingyu; Tian, Xiao-Jun; Xing, Jianhua

    2016-01-01

    Epithelial-to-mesenchymal transition (EMT) is a key step in development, wound healing, and cancer development. It involves cooperation of signaling pathways, such as transformation growth factor-β (TGF-β), Sonic Hedgehog (SHH), and WNT pathways. These signaling pathways crosstalk to each other and converge to key transcription factors (e.g., SNAIL1) to initialize and maintain the process of EMT. The functional roles of multi-signaling pathway crosstalks in EMT are sophisticated and, thus, remain to be explored. In this review, we focused on three major signal transduction pathways that promote or regulate EMT in carcinoma. We discussed the network structures, and provided a brief overview of the current therapy strategies and drug development targeted to these three signal transduction pathways. Finally, we highlighted systems biology approaches that can accelerate the process of deconstructing complex networks and drug discovery. PMID:27043642

  10. Essential role of Her3 in two signaling transduction patterns: Her2/Her3 and MET/Her3 in proliferation of human gastric cancer.

    PubMed

    Yun, Chen; Gang, Li; Rongmin, Gu; Xu, Wen; Xuezhi, Ming; Huanqiu, Chen

    2015-12-01

    Various receptor tyrosine kinase (RTK) pathways were verified in many cancers including gastric cancer (GC), We sought to investigate the expression of RTKs including Her2, Her3, and Met and their transduction patterns in human GC. Over-expression of Her2, Her3, and c-Met in human GC was verified by immunohistochemistry leading to constitutive activation of RTK signaling pathways. Combined RTKs expression was valuable indicators for poor prognosis of GC patients. Using ErbB2 specific inhibitor Lapatinib and c-Met specific inhibitor PHA-665752, we further demonstrated that this constitutive activation of RTK signaling is necessary for the survival of GC cells. However, various RTK pattern: Her3/Her2 and Met/Her3 were verified in the transduction growth stimulus from outside via both AKT and MAPK signaling. Moreover, the essential roles of Her3 in both two heterodimers were obtained which showed significantly attenuated growth effect due to Her3 knockdown both in vitro and in vivo. In conclusion, various molecular transduction patterns: Her2/Her3 and Met/Her3 were verified in human GC, and Her3 could serve as a potential target in GC treatment. PMID:25400108

  11. Notch1 endocytosis is induced by ligand and is required for signal transduction.

    PubMed

    Chapman, G; Major, J A; Iyer, K; James, A C; Pursglove, S E; Moreau, J L M; Dunwoodie, S L

    2016-01-01

    The Notch signalling pathway is widely utilised during embryogenesis in situations where cell-cell interactions are important for cell fate specification and differentiation. DSL ligand endocytosis into the ligand-expressing cell is an important aspect of Notch signalling because it is thought to supply the force needed to separate the Notch heterodimer to initiate signal transduction. A functional role for receptor endocytosis during Notch signal transduction is more controversial. Here we have used live-cell imaging to examine trafficking of the Notch1 receptor in response to ligand binding. Contact with cells expressing ligands induced internalisation and intracellular trafficking of Notch1. Notch1 endocytosis was accompanied by transendocytosis of ligand into the Notch1-expressing signal-receiving cell. Ligand caused Notch1 endocytosis into SARA-positive endosomes in a manner dependent on clathrin and dynamin function. Moreover, inhibition of endocytosis in the receptor-expressing cell impaired ligand-induced Notch1 signalling. Our findings resolve conflicting observations from mammalian and Drosophila studies by demonstrating that ligand-dependent activation of Notch1 signalling requires receptor endocytosis. Endocytosis of Notch1 may provide a force on the ligand:receptor complex that is important for potent signal transduction. PMID:26522918

  12. Reactive Oxygen Species via Redox Signaling to PI3K/AKT Pathway Contribute to the Malignant Growth of 4-Hydroxy Estradiol-Transformed Mammary Epithelial Cells

    PubMed Central

    Okoh, Victor O.; Felty, Quentin; Parkash, Jai; Poppiti, Robert; Roy, Deodutta

    2013-01-01

    The purpose of this study was to investigate the effects of 17-β-estradiol (E2)-induced reactive oxygen species (ROS) on the induction of mammary tumorigenesis. We found that ROS-induced by repeated exposures to 4-hydroxy-estradiol (4-OH-E2), a predominant catechol metabolite of E2, caused transformation of normal human mammary epithelial MCF-10A cells with malignant growth in nude mice. This was evident from inhibition of estrogen-induced breast tumor formation in the xenograft model by both overexpression of catalase as well as by co-treatment with Ebselen. To understand how 4-OH-E2 induces this malignant phenotype through ROS, we investigated the effects of 4-OH-E2 on redox-sensitive signal transduction pathways. During the malignant transformation process we observed that 4-OH-E2 treatment increased AKT phosphorylation through PI3K activation. The PI3K-mediated phosphorylation of AKT in 4-OH-E2-treated cells was inhibited by ROS modifiers as well as by silencing of AKT expression. RNA interference of AKT markedly inhibited 4-OH-E2-induced in vitro tumor formation. The expression of cell cycle genes, cdc2, PRC1 and PCNA and one of transcription factors that control the expression of these genes – nuclear respiratory factor-1 (NRF-1) was significantly up-regulated during the 4-OH-E2-mediated malignant transformation process. The increased expression of these genes was inhibited by ROS modifiers as well as by silencing of AKT expression. These results indicate that 4-OH-E2-induced cell transformation may be mediated, in part, through redox-sensitive AKT signal transduction pathways by up-regulating the expression of cell cycle genes cdc2, PRC1 and PCNA, and the transcription factor – NRF-1. In summary, our study has demonstrated that: (i) 4-OH-E2 is one of the main estrogen metabolites that induce mammary tumorigenesis and (ii) ROS-mediated signaling leading to the activation of PI3K/AKT pathway plays an important role in the generation of 4-OH-E2-induced

  13. Chemical approaches for investigating phosphorylation in signal transduction networks.

    PubMed

    Rothman, Deborah M; Shults, Melissa D; Imperiali, Barbara

    2005-09-01

    The power and scope of chemical synthesis offer considerable opportunities to broaden the lexicon of chemical tools that can be implemented for the study of complex biological systems. To investigate individual signaling proteins and pathways, chemical tools provide a powerful complement to existing genetic, chemical genetic and immunologic methods. In particular, understanding phosphorylation-mediated signaling in real time yields important information about the regulation of cellular function and insights into the origin of disease. Recent advances in the development of photolabile caged analogs of bioactive species and fluorescence-based sensors of protein kinase activities are useful for investigating protein phosphorylation and the roles of phosphoproteins. Photolabile caged analogs allow spatial and temporal control over the release of a compound, while fluorescence-based sensors allow the real-time visualization of kinase activity. Here, we discuss recent advances that have increased the specificity and availability of these tools. PMID:16084095

  14. Signal Transduction Molecules as Targets for Cancer Prevention

    PubMed Central

    Bode, Ann M.; Dong, Zigang

    2010-01-01

    Environmental and life-style aspects are major contributors to human carcinogenesis and, therefore, many human cancers may be preventable. Cancer is the end result of defects in cellular signaling processes that play a key role in the control of cell growth, survival, division, and differentiation. Therefore, identifying molecular and cellular targets critical in cancer development and prevention is an area of intensive research, driving the development of highly specific small-molecule inhibitors. A major idea today is that cancer may be prevented or treated by targeting the products of specific cancer-related genes, frequently encoding signaling proteins or transcription factors. Participants in these joint conferences discussed their latest findings in the identification of promising molecular targets and the development of agents directed against these targets with the goal of effectively transitioning these into the clinical setting. PMID:19244209

  15. Signal transduction regulating meristem development in Arabidopsis. Final report

    SciTech Connect

    Cark, Steven E.

    2003-09-10

    Research support by DE-FG02-96ER20227 focused on the CLV loci and their regulation of organ formation at the Arabidopsis shoot meristem. Shoot meristem function is central to plant development as all of the above-ground organs and tissues of the plant are derived post-embryonically from the shoot meristem. At the shoot meristem, stem cells are maintained, and progeny cells undergo a switch toward differentiation and organ formation. The CLV loci, represented by three genes CLV1, CLV2 and CLV3 are key regulators of meristem development. Each of the CLV loci encode a putative receptor-mediated signaling component. When this work began, virtually nothing was known about receptor-mediated signaling in plants. Thus, our goal was to both characterize these genes and the proteins they encode as regulators of meristem development, and to investigate how receptor-mediated signaling might function in plants. Our work lead to several major publications that were significant contributions to understanding this system.

  16. Network Features and Pathway Analyses of a Signal Transduction Cascade

    PubMed Central

    Yanashima, Ryoji; Kitagawa, Noriyuki; Matsubara, Yoshiya; Weatheritt, Robert; Oka, Kotaro; Kikuchi, Shinichi; Tomita, Masaru; Ishizaki, Shun

    2008-01-01

    The scale-free and small-world network models reflect the functional units of networks. However, when we investigated the network properties of a signaling pathway using these models, no significant differences were found between the original undirected graphs and the graphs in which inactive proteins were eliminated from the gene expression data. We analyzed signaling networks by focusing on those pathways that best reflected cellular function. Therefore, our analysis of pathways started from the ligands and progressed to transcription factors and cytoskeletal proteins. We employed the Python module to assess the target network. This involved comparing the original and restricted signaling cascades as a directed graph using microarray gene expression profiles of late onset Alzheimer's disease. The most commonly used method of shortest-path analysis neglects to consider the influences of alternative pathways that can affect the activation of transcription factors or cytoskeletal proteins. We therefore introduced included k-shortest paths and k-cycles in our network analysis using the Python modules, which allowed us to attain a reasonable computational time and identify k-shortest paths. This technique reflected results found in vivo and identified pathways not found when shortest path or degree analysis was applied. Our module enabled us to comprehensively analyse the characteristics of biomolecular networks and also enabled analysis of the effects of diseases considering the feedback loop and feedforward loop control structures as an alternative path. PMID:19543432

  17. The effect of photoinitiators on intracellular AKT signaling pathway in tissue engineering application

    PubMed Central

    Xu, Leyuan; Sheybani, Natasha; Yeudall, W. Andrew; Yang, Hu

    2015-01-01

    Free-radical photopolymerization initiated by photoinitiators is an important method to make tissue engineering scaffolds. To advance understanding of photoinitiator cytocompatibility, we examined three photoinitiators including 2,2-dimethoxy-2-phenylacetophenone (DMPA), Irgacure 2959 (I-2959), and eosin Y photoinitiating system (EY) in terms of their effects on viability of HN4 cells and expression levels of intracellular AKT and its phosphorylated form p-AKT. Our results show that the photoinitiators and their UV-exposed counterparts affect intracellular AKT signaling, which can be used in conjunction with cell viability for cytocompatibility assessment of photoinitiators. PMID:25709809

  18. Metformin improves hepatic IRS2/PI3K/Akt signaling in insulin-resistant rats of NASH and cirrhosis.

    PubMed

    Xu, Hong; Zhou, Yang; Liu, Yongxia; Ping, Jian; Shou, Qiyang; Chen, Fangming; Ruo, Ru

    2016-05-01

    Nonalcoholic fatty liver disease and cirrhosis are strongly associated with insulin resistance and glucose intolerance. To date, the influence of metformin on glycogen synthesis in the liver is controversial. Limited studies have evaluated the effect of metformin on hepatic insulin signaling pathway in vivo In this study, an insulin-resistant rat model of nonalcoholic steatohepatitis and cirrhosis was developed by high-fat and high-sucrose diet feeding in combination with subcutaneous injection of carbon tetrachloride. Liver tissues of the model rats were featured with severe steatosis and cirrhosis, accompanied by impaired liver function and antioxidant capacity. The glucose tolerance was impaired, and the index of insulin resistance was increased significantly compared with the control. The content of hepatic glycogen was dramatically decreased. The expression of insulin receptor β (IRβ); phosphorylations of IRβ, insulin receptor substrate 2 (IRS2), and Akt; and activities of phosphatidylinositol 3-kinase (PI3K) and glycogen synthase (GS) in the liver were significantly decreased, whereas the activities of glycogen synthase kinase 3α (GSK3α) and glycogen phosphorylase a (GPa) were increased. Metformin treatment remarkably improved liver function, alleviated lipid peroxidation and histological damages of the liver, and ameliorated glucose intolerance and insulin resistance. Metfromin also significantly upregulated the expression of IRβ; increased the phosphorylations of IRβ, IRS2, and Akt; increased the activities of PI3K and GS; and decreased GSK3α and GPa activities. In conclusion, our study suggests that metformin upregulates IRβ expression and the downstream IRS2/PI3K/Akt signaling transduction, therefore, to increase hepatic glycogen storage and improve insulin resistance. These actions may be attributed to the improved liver histological alterations by metformin. PMID:26941037

  19. Transduction of Redox Signaling by Electrophile-Protein Reactions

    PubMed Central

    Rudolph, Tanja K.; Freeman, Bruce A.

    2014-01-01

    Over the last 50 years, the posttranslational modification (PTM) of proteins has emerged as a central mechanism for cells to regulate metabolism, growth, differentiation, cell-cell interactions, and immune responses. By influencing protein structure and function, PTM leads to a multiplication of proteome diversity. Redox-dependent PTMs, mediated by environmental and endogenously generated reactive species, induce cell signaling responses and can have toxic effects in organisms. PTMs induced by the electrophilic by-products of redox reactions most frequently occur at protein thiols; other nucleophilic amino acids serve as less favorable targets. Advances in mass spectrometry and affinity-chemistry strategies have improved the detection of electrophile-induced protein modifications both in vitro and in vivo and have revealed a high degree of amino acid and protein selectivity of electrophilic PTM. The identification of biological targets of electrophiles has motivated further study of the functional impact of various PTM reactions on specific signaling pathways and how this might affect organisms. PMID:19797270

  20. Exercise-Induced Signal Transduction and Gene Regulation In Skeletal Muscle

    PubMed Central

    Wackerhage, Henning; Woods, Niall M.

    2002-01-01

    Skeletal muscle adapts to various forms of exercise depending on the force, speed and duration characteristics of the contraction pattern. The stresses and signals associated with each contraction pattern are likely to specifically activate a network of signal transduction pathways that integrate this information. These pathways include the calcineurin, Calcium/calmodulin-dependent protein kinase (CaMK), mitogen-activated protein kinase (MAPK), protein kinase C (PKC), nuclear factor kappa B (NF-κB), AMP-dependent protein kinase (AMPK), insulin signalling and developmental pathways. Activated signal transduction pathways activate or increase the expression of transcription factors via various mechanisms. Skeletal muscle genes are usually regulated by combinatorial control exerted by several transcription factors and possibly other mechanisms. In addition, adaptations such as an increase in mitochondrial biogenesis or the activation of satellite cell proliferation involve distinct regulatory mechanisms. PMID:24748841

  1. [Application of bio-mass spectrometry in cellular signal transduction].

    PubMed

    Zhang, Xue-Min; Wei, Kai-Hua; Yang, Song-Cheng

    2002-09-01

    The quickly developing techniques of biological mass spectrometry (bio-MS) in recent years realized the high throughput identification of proteins by determining the accurate mass values of trypsin digested peptides and the randomly selected peptide sequence tags, and have been successfully used in the studies of protein interactions and post-translational modification such as the phosphorylation. Compared to the conventional approaches, the above techniques can identify all the phosphorylated proteins (including their phosphorylated amino acid sites) involved in a multi-signal pathway in a single experiment, and they have been developed into a hot-spot of proteomics. The three strategies for the application of bio-MS in the above fields are briefly reviewed. PMID:12198553

  2. Mitogen Activated Protein kinase signal transduction pathways in the prostate

    PubMed Central

    Maroni, Paul D; Koul, Sweaty; Meacham, Randall B; Koul, Hari K

    2004-01-01

    The biochemistry of the mitogen activated protein kinases ERK, JNK, and p38 have been studied in prostate physiology in an attempt to elucidate novel mechanisms and pathways for the treatment of prostatic disease. We reviewed articles examining mitogen-activated protein kinases using prostate tissue or cell lines. As with other tissue types, these signaling modules are links/transmitters for important pathways in prostate cells that can result in cellular survival or apoptosis. While the activation of the ERK pathway appears to primarily result in survival, the roles of JNK and p38 are less clear. Manipulation of these pathways could have important implications for the treatment of prostate cancer and benign prostatic hypertrophy. PMID:15219238

  3. Succinate, an intermediate in metabolism, signal transduction, ROS, hypoxia, and tumorigenesis.

    PubMed

    Tretter, Laszlo; Patocs, Attila; Chinopoulos, Christos

    2016-08-01

    Succinate is an important metabolite at the cross-road of several metabolic pathways, also involved in the formation and elimination of reactive oxygen species. However, it is becoming increasingly apparent that its realm extends to epigenetics, tumorigenesis, signal transduction, endo- and paracrine modulation and inflammation. Here we review the pathways encompassing succinate as a metabolite or a signal and how these may interact in normal and pathological conditions.(1). PMID:26971832

  4. Reactivation of AKT signaling following treatment of cancer cells with PI3K inhibitors attenuates their antitumor effects

    SciTech Connect

    Dufour, Marc; Dormond-Meuwly, Anne; Pythoud, Catherine; Demartines, Nicolas; Dormond, Olivier

    2013-08-16

    Highlights: •PI3K inhibitors inhibit AKT only transiently. •Re-activation of AKT limits the anti-cancer effect of PI3K inhibitors. •The results suggest to combine PI3K and AKT inhibitors in cancer therapy. -- Abstract: Targeting the phosphatidylinositol-3-kinase (PI3K) is a promising approach in cancer therapy. In particular, PI3K blockade leads to the inhibition of AKT, a major downstream effector responsible for the oncogenic activity of PI3K. However, we report here that small molecule inhibitors of PI3K only transiently block AKT signaling. Indeed, treatment of cancer cells with PI3K inhibitors results in a rapid inhibition of AKT phosphorylation and signaling which is followed by the reactivation of AKT signaling after 48 h as observed by Western blot. Reactivation of AKT signaling occurs despite effective inhibition of PI3K activity by PI3K inhibitors. In addition, wortmannin, a broad range PI3K inhibitor, did not block AKT reactivation suggesting that AKT signals independently of PI3K. In a therapeutical perspective, combining AKT and PI3K inhibitors exhibit stronger anti-proliferative and pro-apoptotic effects compared to AKT or PI3K inhibitors alone. Similarly, in a tumor xenograft mouse model, concomitant PI3K and AKT blockade results in stronger anti-cancer activity compared with either blockade alone. This study shows that PI3K inhibitors only transiently inhibit AKT which limits their antitumor activities. It also provides the proof of concept to combine PI3K inhibitors with AKT inhibitors in cancer therapy.

  5. Control of cancer-related signal transduction networks

    NASA Astrophysics Data System (ADS)

    Albert, Reka

    2013-03-01

    Intra-cellular signaling networks are crucial to the maintenance of cellular homeostasis and for cell behavior (growth, survival, apoptosis, movement). Mutations or alterations in the expression of elements of cellular signaling networks can lead to incorrect behavioral decisions that could result in tumor development and/or the promotion of cell migration and metastasis. Thus, mitigation of the cascading effects of such dysregulations is an important control objective. My group at Penn State is collaborating with wet-bench biologists to develop and validate predictive models of various biological systems. Over the years we found that discrete dynamic modeling is very useful in molding qualitative interaction information into a predictive model. We recently demonstrated the effectiveness of network-based targeted manipulations on mitigating the disease T cell large granular lymphocyte (T-LGL) leukemia. The root of this disease is the abnormal survival of T cells which, after successfully fighting an infection, should undergo programmed cell death. We synthesized the relevant network of within-T-cell interactions from the literature, integrated it with qualitative knowledge of the dysregulated (abnormal) states of several network components, and formulated a Boolean dynamic model. The model indicated that the system possesses a steady state corresponding to the normal cell death state and a T-LGL steady state corresponding to the abnormal survival state. For each node, we evaluated the restorative manipulation consisting of maintaining the node in the state that is the opposite of its T-LGL state, e.g. knocking it out if it is overexpressed in the T-LGL state. We found that such control of any of 15 nodes led to the disappearance of the T-LGL steady state, leaving cell death as the only potential outcome from any initial condition. In four additional cases the probability of reaching the T-LGL state decreased dramatically, thus these nodes are also possible control

  6. Crystal structure of Pseudomonas aeruginosa bacteriophytochrome: Photoconversion and signal transduction

    SciTech Connect

    Yang, Xiaojing; Kuk, Jane; Moffat, Keith

    2008-11-12

    Phytochromes are red-light photoreceptors that regulate light responses in plants, fungi, and bacteria via reversible photoconversion between red (Pr) and far-red (Pfr) light-absorbing states. Here we report the crystal structure at 2.9 {angstrom} resolution of a bacteriophytochrome from Pseudomonas aeruginosa with an intact, fully photoactive photosensory core domain in its dark-adapted Pfr state. This structure reveals how unusual interdomain interactions, including a knot and an 'arm' structure near the chromophore site, bring together the PAS (Per-ARNT-Sim), GAF (cGMP phosphodiesterase/adenyl cyclase/FhlA), and PHY (phytochrome) domains to achieve Pr/Pfr photoconversion. The PAS, GAF, and PHY domains have topologic elements in common and may have a single evolutionary origin. We identify key interactions that stabilize the chromophore in the Pfr state and provide structural and mutational evidence to support the essential role of the PHY domain in efficient Pr/Pfr photoconversion. We also identify a pair of conserved residues that may undergo concerted conformational changes during photoconversion. Modeling of the full-length bacteriophytochrome structure, including its output histidine kinase domain, suggests how local structural changes originating in the photosensory domain modulate interactions between long, cross-domain signaling helices at the dimer interface and are transmitted to the spatially distant effector domain, thereby regulating its histidine kinase activity.

  7. Crystal structure of Pseudomonas aeruginosa bacteriophytochrome: photoconversion and signal transduction.

    PubMed

    Yang, Xiaojing; Kuk, Jane; Moffat, Keith

    2008-09-23

    Phytochromes are red-light photoreceptors that regulate light responses in plants, fungi, and bacteria via reversible photoconversion between red (Pr) and far-red (Pfr) light-absorbing states. Here we report the crystal structure at 2.9 A resolution of a bacteriophytochrome from Pseudomonas aeruginosa with an intact, fully photoactive photosensory core domain in its dark-adapted Pfr state. This structure reveals how unusual interdomain interactions, including a knot and an "arm" structure near the chromophore site, bring together the PAS (Per-ARNT-Sim), GAF (cGMP phosphodiesterase/adenyl cyclase/FhlA), and PHY (phytochrome) domains to achieve Pr/Pfr photoconversion. The PAS, GAF, and PHY domains have topologic elements in common and may have a single evolutionary origin. We identify key interactions that stabilize the chromophore in the Pfr state and provide structural and mutational evidence to support the essential role of the PHY domain in efficient Pr/Pfr photoconversion. We also identify a pair of conserved residues that may undergo concerted conformational changes during photoconversion. Modeling of the full-length bacteriophytochrome structure, including its output histidine kinase domain, suggests how local structural changes originating in the photosensory domain modulate interactions between long, cross-domain signaling helices at the dimer interface and are transmitted to the spatially distant effector domain, thereby regulating its histidine kinase activity. PMID:18799746

  8. Modulation of signal transduction pathways by natural compounds in cancer.

    PubMed

    Ranjan, Alok; Fofaria, Neel M; Kim, Sung-Hoon; Srivastava, Sanjay K

    2015-10-01

    Cancer is generally regarded as the result of abnormal growth of cells. According to World Health Organization, cancer is the leading cause of mortality worldwide. Mother nature provides a large source of bioactive compounds with excellent therapeutic efficacy. Numerous phytochemicals from nature have been investigated for anticancer properties. In this review article, we discuss several natural compounds, which have shown anti-cancer activity. Natural compounds induce cell cycle arrest, activate intrinsic and extrinsic apoptosis pathways, generate Reactive Oxygen Species (ROS), and down-regulate activated signaling pathways, resulting in inhibition of cell proliferation, progression and metastasis of cancer. Several preclinical studies have suggested that natural compounds can also increase the sensitivity of resistant cancers to available chemotherapy agents. Furthermore, combining FDA approved anti-cancer drugs with natural compounds results in improved efficacy. On the basis of these exciting outcomes of natural compounds against several cancer types, several agents have already advanced to clinical trials. In conclusion, preclinical results and clinical outcomes against cancer suggest promising anticancer efficacy of agents from natural sources. PMID:26481373

  9. Role of relaxation time scale in noisy signal transduction.

    PubMed

    Maity, Alok Kumar; Chaudhury, Pinaki; Banik, Suman K

    2015-01-01

    Intra-cellular fluctuations, mainly triggered by gene expression, are an inevitable phenomenon observed in living cells. It influences generation of phenotypic diversity in genetically identical cells. Such variation of cellular components is beneficial in some contexts but detrimental in others. To quantify the fluctuations in a gene product, we undertake an analytical scheme for studying few naturally abundant linear as well as branched chain network motifs. We solve the Langevin equations associated with each motif under the purview of linear noise approximation and derive the expressions for Fano factor and mutual information in close analytical form. Both quantifiable expressions exclusively depend on the relaxation time (decay rate constant) and steady state population of the network components. We investigate the effect of relaxation time constraints on Fano factor and mutual information to indentify a time scale domain where a network can recognize the fluctuations associated with the input signal more reliably. We also show how input population affects both quantities. We extend our calculation to long chain linear motif and show that with increasing chain length, the Fano factor value increases but the mutual information processing capability decreases. In this type of motif, the intermediate components act as a noise filter that tune up input fluctuations and maintain optimum fluctuations in the output. For branched chain motifs, both quantities vary within a large scale due to their network architecture and facilitate survival of living system in diverse environmental conditions. PMID:25955500

  10. Role of Relaxation Time Scale in Noisy Signal Transduction

    PubMed Central

    Maity, Alok Kumar; Chaudhury, Pinaki; Banik, Suman K

    2015-01-01

    Intra-cellular fluctuations, mainly triggered by gene expression, are an inevitable phenomenon observed in living cells. It influences generation of phenotypic diversity in genetically identical cells. Such variation of cellular components is beneficial in some contexts but detrimental in others. To quantify the fluctuations in a gene product, we undertake an analytical scheme for studying few naturally abundant linear as well as branched chain network motifs. We solve the Langevin equations associated with each motif under the purview of linear noise approximation and derive the expressions for Fano factor and mutual information in close analytical form. Both quantifiable expressions exclusively depend on the relaxation time (decay rate constant) and steady state population of the network components. We investigate the effect of relaxation time constraints on Fano factor and mutual information to indentify a time scale domain where a network can recognize the fluctuations associated with the input signal more reliably. We also show how input population affects both quantities. We extend our calculation to long chain linear motif and show that with increasing chain length, the Fano factor value increases but the mutual information processing capability decreases. In this type of motif, the intermediate components act as a noise filter that tune up input fluctuations and maintain optimum fluctuations in the output. For branched chain motifs, both quantities vary within a large scale due to their network architecture and facilitate survival of living system in diverse environmental conditions. PMID:25955500

  11. Discovery of GPCR ligands for probing signal transduction pathways

    PubMed Central

    Brogi, Simone; Tafi, Andrea; Désaubry, Laurent; Nebigil, Canan G.

    2014-01-01

    G protein-coupled receptors (GPCRs) are seven integral transmembrane proteins that are the primary targets of almost 30% of approved drugs and continue to represent a major focus of pharmaceutical research. All of GPCR targeted medicines were discovered by classical medicinal chemistry approaches. After the first GPCR crystal structures were determined, the docking screens using these structures lead to discovery of more novel and potent ligands. There are over 360 pharmaceutically relevant GPCRs in the human genome and to date about only 30 of structures have been determined. For these reasons, computational techniques such as homology modeling and molecular dynamics simulations have proven their usefulness to explore the structure and function of GPCRs. Furthermore, structure-based drug design and in silico screening (High Throughput Docking) are still the most common computational procedures in GPCRs drug discovery. Moreover, ligand-based methods such as three-dimensional quantitative structure–selectivity relationships, are the ideal molecular modeling approaches to rationalize the activity of tested GPCR ligands and identify novel GPCR ligands. In this review, we discuss the most recent advances for the computational approaches to effectively guide selectivity and affinity of ligands. We also describe novel approaches in medicinal chemistry, such as the development of biased agonists, allosteric modulators, and bivalent ligands for class A GPCRs. Furthermore, we highlight some knockout mice models in discovering biased signaling selectivity. PMID:25506327

  12. New signal transduction paradigms in anthracycline-induced cardiotoxicity.

    PubMed

    Ghigo, Alessandra; Li, Mingchuan; Hirsch, Emilio

    2016-07-01

    Anthracyclines, such as doxorubicin, are the most potent and widely used chemotherapeutic agents for the treatment of a variety of human cancers, including solid tumors and hematological malignancies. However, their clinical use is hampered by severe cardiotoxic side effects and cancer therapy-related heart disease has become a leading cause of morbidity and mortality among cancer survivors. The identification of therapeutic strategies limiting anthracycline cardiotoxicity with preserved antitumor efficacy thus represents the current challenge of cardio-oncologists. Anthracycline cardiotoxicity has been originally ascribed to the ability of this class of drugs to disrupt iron metabolism and generate excess of reactive oxygen species (ROS). However, small clinical trials with iron chelators and anti-oxidants failed to provide any benefit and suggested that doxorubicin cardiotoxicity is not solely due to redox cycling. New emerging explanations include anthracycline-dependent regulation of major signaling pathways controlling DNA damage response, cardiomyocyte survival, cardiac inflammation, energetic stress and gene expression modulation. This review will summarize recent studies unraveling the complex web of mechanisms of doxorubicin-mediated cardiotoxicity, and identifying new druggable players for the prevention of heart disease in cancer patients. This article is part of a Special Issue entitled: Cardiomyocyte Biology: Integration of Developmental and Environmental Cues in the Heart edited by Marcus Schaub and Hughes Abriel. PMID:26828775

  13. Expansion of Signal Transduction Pathways in Fungi by Extensive Genome Duplication.

    PubMed

    Corrochano, Luis M; Kuo, Alan; Marcet-Houben, Marina; Polaino, Silvia; Salamov, Asaf; Villalobos-Escobedo, José M; Grimwood, Jane; Álvarez, M Isabel; Avalos, Javier; Bauer, Diane; Benito, Ernesto P; Benoit, Isabelle; Burger, Gertraud; Camino, Lola P; Cánovas, David; Cerdá-Olmedo, Enrique; Cheng, Jan-Fang; Domínguez, Angel; Eliáš, Marek; Eslava, Arturo P; Glaser, Fabian; Gutiérrez, Gabriel; Heitman, Joseph; Henrissat, Bernard; Iturriaga, Enrique A; Lang, B Franz; Lavín, José L; Lee, Soo Chan; Li, Wenjun; Lindquist, Erika; López-García, Sergio; Luque, Eva M; Marcos, Ana T; Martin, Joel; McCluskey, Kevin; Medina, Humberto R; Miralles-Durán, Alejandro; Miyazaki, Atsushi; Muñoz-Torres, Elisa; Oguiza, José A; Ohm, Robin A; Olmedo, María; Orejas, Margarita; Ortiz-Castellanos, Lucila; Pisabarro, Antonio G; Rodríguez-Romero, Julio; Ruiz-Herrera, José; Ruiz-Vázquez, Rosa; Sanz, Catalina; Schackwitz, Wendy; Shahriari, Mahdi; Shelest, Ekaterina; Silva-Franco, Fátima; Soanes, Darren; Syed, Khajamohiddin; Tagua, Víctor G; Talbot, Nicholas J; Thon, Michael R; Tice, Hope; de Vries, Ronald P; Wiebenga, Ad; Yadav, Jagjit S; Braun, Edward L; Baker, Scott E; Garre, Victoriano; Schmutz, Jeremy; Horwitz, Benjamin A; Torres-Martínez, Santiago; Idnurm, Alexander; Herrera-Estrella, Alfredo; Gabaldón, Toni; Grigoriev, Igor V

    2016-06-20

    Plants and fungi use light and other signals to regulate development, growth, and metabolism. The fruiting bodies of the fungus Phycomyces blakesleeanus are single cells that react to environmental cues, including light, but the mechanisms are largely unknown [1]. The related fungus Mucor circinelloides is an opportunistic human pathogen that changes its mode of growth upon receipt of signals from the environment to facilitate pathogenesis [2]. Understanding how these organisms respond to environmental cues should provide insights into the mechanisms of sensory perception and signal transduction by a single eukaryotic cell, and their role in pathogenesis. We sequenced the genomes of P. blakesleeanus and M. circinelloides and show that they have been shaped by an extensive genome duplication or, most likely, a whole-genome duplication (WGD), which is rarely observed in fungi [3-6]. We show that the genome duplication has expanded gene families, including those involved in signal transduction, and that duplicated genes have specialized, as evidenced by differences in their regulation by light. The transcriptional response to light varies with the developmental stage and is still observed in a photoreceptor mutant of P. blakesleeanus. A phototropic mutant of P. blakesleeanus with a heterozygous mutation in the photoreceptor gene madA demonstrates that photosensor dosage is important for the magnitude of signal transduction. We conclude that the genome duplication provided the means to improve signal transduction for enhanced perception of environmental signals. Our results will help to understand the role of genome dynamics in the evolution of sensory perception in eukaryotes. PMID:27238284

  14. IL-8 induces the epithelial-mesenchymal transition of renal cell carcinoma cells through the activation of AKT signaling

    PubMed Central

    Zhou, Nan; Lu, Fuding; Liu, Cheng; Xu, Kewei; Huang, Jian; Yu, Dexin; Bi, Liangkuan

    2016-01-01

    The epithelial-mesenchymal transition (EMT) process has increasingly been examined due to its role in the progression of human tumors. Renal cell carcinoma (RCC) is one of the most common urological tumors that results in patient mortality. Previous studies have demonstrated that the EMT process is closely associated with the metastasis of RCC; however, the underlying molecular mechanism has not been determined yet. The present study revealed that interleukin (IL)-8 was highly expressed in metastatic RCC. IL-8 could induce the EMT of an RCC cell line by enhancing N-cadherin expression and decreasing E-cadherin expression. Furthermore, IL-8 could induce AKT phosphorylation, and the phosphatidylinositol-4,5-bisphosphate 3-kinase inhibitor LY294002 could inhibit the EMT of RCC cells that was induced by IL-8. Therefore, these results suggest that IL-8 is able to promote the EMT of RCC through the activation of the AKT signal transduction pathway, and this may provide a possible molecular mechanism for RCC metastasis. PMID:27588140

  15. Signal Transduction and Molecular Targets of Selected Flavonoids

    PubMed Central

    Bode, Ann M.

    2013-01-01

    Abstract Significance: Diet exerts a major influence on the risk for developing cancer and heart disease. Food factors such as flavonoids are alleged to protect cells from premature aging and disease by shielding DNA, proteins, and lipids from oxidative damage. Recent Advances: Our work has focused on clarifying the effects of dietary components on cancer cell proliferation and tumor growth, discovering mechanisms to explain the effects, and identifying the specific molecular targets of these compounds. Our strategy for identifying specific molecular targets of phytochemicals involves the use of supercomputer technology combined with protein crystallography, molecular biology, and experimental laboratory verification. Critical Issues: One of the greatest challenges for scientists is to reduce the accumulation of distortion and half truths reported in the popular media regarding the health benefits of certain foods or food supplements. The use of these is not new, but interest has increased dramatically because of perceived health benefits that are presumably acquired without unpleasant side effects. Flavonoids are touted to exert many beneficial effects in vitro. However, whether they can produce these effects in vivo is disputed. Future Directions: The World Health Organization indicates that one third of all cancer deaths are preventable and that diet is closely linked to prevention. Based on this idea and epidemiological findings, attention has centered on dietary phytochemicals as an effective intervention in cancer development. However, an unequivocal link between diet and cancer has not been established. Thus, identifying cancer preventive dietary agents with specific molecular targets is essential to move forward toward successful cancer prevention. Antioxid. Redox Signal. 19, 163–180. PMID:23458437

  16. Fangchinoline suppresses the growth and invasion of human glioblastoma cells by inhibiting the kinase activity of Akt and Akt-mediated signaling cascades.

    PubMed

    Guo, Bingyu; Xie, Peng; Su, Jingyuan; Zhang, Tingting; Li, Xiaoming; Liang, Guobiao

    2016-02-01

    Glioblastoma multiforme (GBM) is one of the most palindromic and malignant central nervous system neoplasms, and the current treatment is not effectual for GBM. Research of specific medicine for GBM is significant. Fangchinoline possesses a wide range of pharmacological activities and attracts more attentions due to its anti-tumor effects. In this study, two WHO grade IV human GBM cell lines (U87 MG and U118 MG) were exposed to fangchinoline, and we found that fangchinoline specifically inhibits the kinase activity of Akt and markedly suppresses the phosphorylation of Thr308 and Ser473 of Akt in human GBM cells. We also observed that fangchinoline inhibits tumor cell proliferation and invasiveness and induces apoptosis through suppressing the Akt-mediated signaling cascades, including Akt/p21, Akt/Bad, and Akt/matrix metalloproteinases (MMPs). These data demonstrated that fangchinoline exerts its anti-tumor effects in human glioblastoma cells, at least partly by inhibiting the kinase activity of Akt and suppressing Akt-mediated signaling cascades. PMID:26408176

  17. MYOCARDIAL AKT: THE OMNIPRESENT NEXUS

    PubMed Central

    Sussman, Mark A.; Völkers, Mirko; Fischer, Kimberlee; Bailey, Brandi; Cottage, Christopher T.; Din, Shabana; Gude, Natalie; Avitabile, Daniele; Alvarez, Roberto; Sundararaman, Balaji; Quijada, Pearl; Mason, Matt; Konstandin, Mathias H.; Malhowski, Amy; Cheng, Zhaokang; Khan, Mohsin; McGregor, Michael

    2013-01-01

    One of the greatest examples of integrated signal transduction is revealed by examination of effects mediated by AKT kinase in myocardial biology. Positioned at the intersection of multiple afferent and efferent signals, AKT exemplifies a molecular sensing node that coordinates dynamic responses of the cell in literally every aspect of biological responses. The balanced and nuanced nature of homeostatic signaling is particularly essential within the myocardial context, where regulation of survival, energy production, contractility, and response to pathological stress all flow through the nexus of AKT activation or repression. Equally important, the loss of regulated AKT activity is primarily the cause or consequence of pathological conditions leading to remodeling of the heart and eventual decompensation. This review presents an overview compendium of the complex world of myocardial AKT biology gleaned from more than a decade of research. Summarization of the widespread influence that AKT exerts upon myocardial responses leaves no doubt that the participation of AKT in molecular signaling will need to be reckoned with as a seemingly omnipresent regulator of myocardial molecular biological responses. PMID:21742795

  18. Signal Integration by Akt Regulates CD8 TCell Effector and Memory Differentiation

    PubMed Central

    Kim, Eui Ho; Sullivan, Jeremy A.; Plisch, Erin H.; Tejera, Melba Marie; Jatzek, Anna; Choi, Kwan Yong; Suresh, M.

    2012-01-01

    During a T cell response, the effector CTL pool contains two cellular subsets: short-lived effector cells (SLECs), a majority of which are destined for apoptosis, and the memory precursor effector cells (MPECs) that differentiate into memory cells. Understanding the mechanisms that govern the differentiation of memory CD8 T cells is of fundamental importance in the development of effective CD8 T cell-based vaccines. The strength and nature of TCR signaling along with signals delivered by cytokines like IL-2 and IL-12influence differentiation of SLECs and MPECs. A central question is, how are signals emanating from multiple receptors integrated and interpreted to define the fate of effector CTLs? Using genetic and pharmacological tools, we have identified Akt as a signal integrator that links distinct facets of CTL differentiation to the specific signaling pathways of FOXO, mTOR, and Wnt/β-catenin. Sustained Akt activation triggered by convergent extracellular signals evokes a transcription program that enhances effector functions, drives differentiation of terminal effectors, and diminishes the CTLs’ potential to survive and differentiate into memory cells. While sustained Akt activation severely impaired CD8 T cell memory and protective immunity, in vivo inhibition of Akt rescued SLECs from deletion and increased the number of memory CD8 T cells. Thus, the cumulative strength of convergent signals from signaling molecules such as TCR, costimulatory molecules, and cytokine receptors governs the magnitude of Akt activation, which in turn controls the generation of long-lived memory cells. These findings suggest that therapeutic modulation of Akt might be a strategy to augment vaccine-induced immunity. PMID:22467649

  19. [Current understanding of signaling transduction pathway and biological functions of Karrikins].

    PubMed

    Luo, Xiaofeng; Qi, Ying; Meng, Yongjie; Shuai, Haiwei; Chen, Feng; Yang, Wenyu; Shu, Kai

    2016-01-01

    Karrikins are a class of signaling molecules discovered in wildfire smoke, which can significantly promote seed germination in some species (such as Arabidopsis and Avena fatua). The structures of Karrikins were first elucidated in 2004. At present, six different types of Karrikins have been documented, and their biological activities vary significantly. So far, studies for Karrikins have become a hot spot in the plant molecular biology field. Recent advances demonstrate that Karrikins regulate plant photomorphogenesis and leaf differentiation effectively, in addition to the effect on seed germination. Furthermore, Karrikins share highly similar molecular structures and signaling transduction pathways with strigolactone. In this review, we summarize the history of discovery, signaling transduction pathways, physiological functions and ecological significance of Karrikins, and further discuss the future research directions. PMID:26787523

  20. Signal transduction pathways and transcription factors triggered by arsenic trioxide in leukemia cells

    SciTech Connect

    Sumi, Daigo; Shinkai, Yasuhiro; Kumagai, Yoshito

    2010-05-01

    Arsenic trioxide (As{sub 2}O{sub 3}) is widely used to treat acute promyelocytic leukemia (APL). Several lines of evidence have indicated that As{sub 2}O{sub 3} affects signal transduction and transactivation of transcription factors, resulting in the stimulation of apoptosis in leukemia cells, because some transcription factors are reported to associate with the redox condition of the cells, and arsenicals cause oxidative stress. Thus, the disturbance and activation of the cellular signaling pathway and transcription factors due to reactive oxygen species (ROS) generation during arsenic exposure may explain the ability of As{sub 2}O{sub 3} to induce a complete remission in relapsed APL patients. In this report, we review recent findings on ROS generation and alterations in signal transduction and in transactivation of transcription factors during As{sub 2}O{sub 3} exposure in leukemia cells.

  1. Assembly and function of a quaternary signal transduction complex monitored by surface plasmon resonance

    NASA Astrophysics Data System (ADS)

    Schuster, Stephan C.; Swanson, Ronald V.; Alex, Lisa A.; Bourret, Robert B.; Simon, Melvin I.

    1993-09-01

    WE have used surface plasmon resonance biosensor technology to monitor the assembly and dynamics of a signal transduction complex which controls chemotaxis in Escherichia coli A quaternary complex formed which consisted of the response regulator CheY, the histidine protein kinase CheA, a coupling protein CheW and a membrane-bound chemoreceptor Tar. Using various experimental conditions and mutant proteins, we have shown that the complex dissociates under conditions that favour phosphorylation of CheY. Direct physical analysis of interactions among proteins in this signal transduction pathway provides evidence for a previously unrecognized binding interaction between the kinase and its substrate. This interaction may be important for enhancing substrate specificity and preventing 'crosstalk' with other systems. The approach is generally applicable to furthering our understanding of how signalling complexes transduce intracellular messages.

  2. Protein Tyrosine Phosphatases: From Housekeeping Enzymes to Master-Regulators of Signal Transduction

    PubMed Central

    Tonks, Nicholas K.

    2013-01-01

    There are many misconceptions surrounding the roles of protein phosphatases in the regulation of signal transduction, perhaps the most damaging of which is the erroneous view that these enzymes exert their effects merely as constitutively active housekeeping enzymes. On the contrary, the phosphatases are critical, specific regulators of signaling in their own right and serve an essential function, in a coordinated manner with the kinases, to determine the response to a physiological stimulus. This review is a personal perspective on the development of our understanding of the protein tyrosine phosphatase (PTP) family of enzymes. I have discussed various aspects of the structure, regulation and function of the PTP family, which I hope will illustrate the fundamental importance of these enzymes to the control of signal transduction. PMID:23176256

  3. Connective tissue growth factor induces cardiac hypertrophy through Akt signaling

    SciTech Connect

    Hayata, Nozomi; Fujio, Yasushi; Yamamoto, Yasuhiro; Iwakura, Tomohiko; Obana, Masanori; Takai, Mika; Mohri, Tomomi; Nonen, Shinpei; Maeda, Makiko; Azuma, Junichi

    2008-05-30

    In the process of cardiac remodeling, connective tissue growth factor (CTGF/CCN2) is secreted from cardiac myocytes. Though CTGF is well known to promote fibroblast proliferation, its pathophysiological effects in cardiac myocytes remain to be elucidated. In this study, we examined the biological effects of CTGF in rat neonatal cardiomyocytes. Cardiac myocytes stimulated with full length CTGF and its C-terminal region peptide showed the increase in cell surface area. Similar to hypertrophic ligands for G-protein coupled receptors, such as endothelin-1, CTGF activated amino acid uptake; however, CTGF-induced hypertrophy is not associated with the increased expression of skeletal actin or BNP, analyzed by Northern-blotting. CTGF treatment activated ERK1/2, p38 MAPK, JNK and Akt. The inhibition of Akt by transducing dominant-negative Akt abrogated CTGF-mediated increase in cell size, while the inhibition of MAP kinases did not affect the cardiac hypertrophy. These findings indicate that CTGF is a novel hypertrophic factor in cardiac myocytes.

  4. Mutual inhibition of insulin signaling and PHLPP-1 determines cardioprotective efficiency of Akt in aged heart.

    PubMed

    Xing, Yuan; Sun, Wanqing; Wang, Yishi; Gao, Feng; Ma, Heng

    2016-05-01

    Insulin protects cardiomyocytes from myocardial ischemia/reperfusion (MI/R) injury through activating Akt. However, phosphatase PHLPP-1 (PH domain leucine-rich repeat protein phosphatase-1) dephosphorylates and inactivates Akt. The balanced competitive interaction of insulin and PHLPP-1 has not been directly examined. In this study, we have identified the effect of mutual inhibition of insulin signaling and PHLPP-1 on the cardioprotective efficiency of Akt in aged heart. Young (3 months) and aged (20 months) Sprague Dawley (SD) rats were subjected to MI/Rin vivo. The PHLPP-1 level was higher in aged vs. young hearts at base. But, insulin treatment failed to decrease PHLPP-1 level during reperfusion in the aged hearts. Consequently, the cardioprotection of insulin-induced Akt activation was impaired in aged hearts, resulting in more susceptible to MI/R injury. In cultured rat ventricular myocytes, PHLPP-1 knockdown significantly enhanced insulin-induced Akt phosphorylation and reduced simulated hypoxia/reoxygenation-induced apoptosis. Contrary, PHLPP-1 overexpression terminated Akt phosphorylation and deteriorated myocytes apoptosis. Using in vivo aged animal models, we confirmed that cardiac PHLPP-1 knockdown or enhanced insulin sensitivity by exercise training dramatically increased insulin-induced Akt phosphorylation. Specifically, MI/R-induced cardiomyocyte apoptosis and infarct size were decreased and cardiac function was increased. More importantly, we found that insulin regulated the degradation of PHLPP-1 and insulin treatment could enhance the binding between PHLPP-1 and β-transducin repeat-containing protein (β-TrCP) to target for ubiquitin-dependent degradation. Altogether, we have identified a new mechanism by which insulin suppresses PHLPP-1 to enhance Akt activation. But, aged heart possesses lower insulin effectiveness and fails to decrease PHLPP-1 during MI/R, which subsequently limited Akt activity and cardioprotection. PHLPP-1 could be a

  5. Mutual inhibition of insulin signaling and PHLPP-1 determines cardioprotective efficiency of Akt in aged heart

    PubMed Central

    Xing, Yuan; Sun, Wanqing; Wang, Yishi; Gao, Feng; Ma, Heng

    2016-01-01

    Insulin protects cardiomyocytes from myocardial ischemia/reperfusion (MI/R) injury through activating Akt. However, phosphatase PHLPP-1 (PH domain leucine-rich repeat protein phosphatase-1) dephosphorylates and inactivates Akt. The balanced competitive interaction of insulin and PHLPP-1 has not been directly examined. In this study, we have identified the effect of mutual inhibition of insulin signaling and PHLPP-1 on the cardioprotective efficiency of Akt in aged heart. Young (3 mon) and aged (20 mon) Sprague Dawley (SD) rats were subjected to MI/R in vivo. The PHLPP-1 level was higher in aged vs. young hearts at base. But, insulin treatment failed to decrease PHLPP-1 level during reperfusion in the aged hearts. Consequently, the cardioprotection of insulin-induced Akt activation was impaired in aged hearts, resulting in more susceptible to MI/R injury. In cultured rat ventricular myocytes, PHLPP-1 knockdown significantly enhanced insulin-induced Akt phosphorylation and reduced simulated hypoxia/reoxygenation-induced apoptosis. Contrary, PHLPP-1 overexpression terminated Akt phosphorylation and deteriorated myocytes apoptosis. Using in vivo aged animal models, we confirmed that cardiac PHLPP-1 knockdown or enhanced insulin sensitivity by exercise training dramatically increased insulin-induced Akt phosphorylation. Specifically, MI/R-induced cardiomyocyte apoptosis and infarct size were decreased and cardiac function was increased. More importantly, we found that insulin regulated the degradation of PHLPP-1 and insulin treatment could enhance the binding between PHLPP-1 and β-transducin repeat-containing protein (β-TrCP) to target for ubiquitin-dependent degradation. Altogether, we have identified a new mechanism by which insulin suppresses PHLPP-1 to enhance Akt activation. But, aged heart possesses lower insulin effectiveness and fails to decrease PHLPP-1 during MI/R, which subsequently limited Akt activity and cardioprotection. PHLPP-1 could be a promising

  6. Interference with Akt signaling pathway contributes curcumin-induced adipocyte insulin resistance.

    PubMed

    Zhang, Deling; Zhang, Yemin; Ye, Mao; Ding, Youming; Tang, Zhao; Li, Mingxin; Zhou, Yu; Wang, Changhua

    2016-07-01

    Previous study has shown that curcumin directly or indirectly suppresses insulin signaling in 3T3-L1 adipocytes. However, the underlying mechanism remains unclear. Here we experimentally demonstrate that curcumin inhibited the ubiquitin-proteasome system (UPS) function, activated autophagy, and reduced protein levels of protein kinase B (Akt) in a dose- and time-dependent manner in 3T3-L1 adipocytes, accompanied with attenuation of insulin-stimulated Akt phosphorylation, plasma membrane translocation of glucose transporter type 4 (GLUT4), and glucose uptake. These in vitro inhibitory effects of curcumin on Akt protein expression and insulin action were reversed by pharmacological and genetic inhibition of autophagy but not by inhibition of the UPS and caspases. In addition, Akt reduction in adipose tissues of mice treated with curcumin could be recovered by administration of autophagy inhibitor bafilomycin A1 (BFA). This new finding provides a novel mechanism by which curcumin induces insulin resistance in adipocytes. PMID:27113027

  7. Modulators of Stomatal Lineage Signal Transduction Alter Membrane Contact Sites and Reveal Specialization among ERECTA Kinases.

    PubMed

    Ho, Chin-Min Kimmy; Paciorek, Tomasz; Abrash, Emily; Bergmann, Dominique C

    2016-08-22

    Signal transduction from a cell's surface to its interior requires dedicated signaling elements and a cellular environment conducive to signal propagation. Plant development, defense, and homeostasis rely on plasma membrane receptor-like kinases to perceive endogenous and environmental signals, but little is known about their immediate downstream targets and signaling modifiers. Using genetics, biochemistry, and live-cell imaging, we show that the VAP-RELATED SUPPRESSOR OF TMM (VST) family is required for ERECTA-mediated signaling in growth and cell-fate determination and reveal a role for ERECTA-LIKE2 in modulating signaling by its sister kinases. We show that VSTs are peripheral plasma membrane proteins that can form complexes with integral ER-membrane proteins, thereby potentially influencing the organization of the membrane milieu to promote efficient and differential signaling from the ERECTA-family members to their downstream intracellular targets. PMID:27554856

  8. Insulin reveals Akt signaling as a novel regulator of norepinephrine transporter trafficking and norepinephrine homeostasis.

    PubMed

    Robertson, Sabrina D; Matthies, Heinrich J G; Owens, W Anthony; Sathananthan, Vidiya; Christianson, Nicole S Bibus; Kennedy, J Phillip; Lindsley, Craig W; Daws, Lynette C; Galli, Aurelio

    2010-08-25

    Noradrenergic signaling in the CNS plays an essential role in circuits involving attention, mood, memory, and stress as well as providing pivotal support for autonomic function in the peripheral nervous system. The high-affinity norepinephrine (NE) transporter (NET) is the primary mechanism by which noradrenergic synaptic transmission is terminated. Data indicate that NET function is regulated by insulin, a hormone critical for the regulation of metabolism. Given the high comorbidity of metabolic disorders such as diabetes and obesity with mental disorders such as depression and schizophrenia, we sought to determine how insulin signaling regulates NET function and thus noradrenergic homeostasis. Here, we show that acute insulin treatment, through the downstream kinase protein kinase B (Akt), significantly decreases NET surface expression in mouse hippocampal slices and superior cervical ganglion neuron boutons (sites of synaptic NE release). In vivo manipulation of insulin/Akt signaling, with streptozotocin, a drug that induces a type 1-like diabetic state in mice, also results in aberrant NET function and NE homeostasis. Notably, we also demonstrate that Akt inhibition or stimulation, independent of insulin, is capable of altering NET surface availability. These data suggest that aberrant states of Akt signaling such as in diabetes and obesity have the potential to alter NET function and noradrenergic tone in the brain. Furthermore, they provide one potential molecular mechanism by which Akt, a candidate gene for mood disorders such as schizophrenia and depression, can impact brain monoamine homeostasis. PMID:20739551

  9. Mechanisms of signal transduction by ethylene: overlapping and non-overlapping signalling roles in a receptor family

    PubMed Central

    Shakeel, Samina N.; Wang, Xiaomin; Binder, Brad M.; Schaller, G. Eric

    2013-01-01

    The plant hormone ethylene regulates growth and development as well as responses to biotic and abiotic stresses. Over the last few decades, key elements involved in ethylene signal transduction have been identified through genetic approaches, these elements defining a pathway that extends from initial ethylene perception at the endoplasmic reticulum to changes in transcriptional regulation within the nucleus. Here, we present our current understanding of ethylene signal transduction, focusing on recent developments that support a model with overlapping and non-overlapping roles for members of the ethylene receptor family. We consider the evidence supporting this model for sub-functionalization within the receptor family, and then discuss mechanisms by which such a sub-functionalization may occur. To this end, we consider the importance of receptor interactions in modulating their signal output and how such interactions vary in the receptor family. In addition, we consider evidence indicating that ethylene signal output by the receptors involves both phosphorylation-dependent and phosphorylation-independent mechanisms. We conclude with a current model for signalling by the ethylene receptors placed within the overall context of ethylene signal transduction. PMID:23543258

  10. Global Transcription Profiling Reveals Multiple Sugar Signal Transduction Mechanisms in ArabidopsisW⃞

    PubMed Central

    Price, John; Laxmi, Ashverya; St. Martin, Steven K.; Jang, Jyan-Chyun

    2004-01-01

    Complex and interconnected signaling networks allow organisms to control cell division, growth, differentiation, or programmed cell death in response to metabolic and environmental cues. In plants, it is known that sugar and nitrogen are critical nutrient signals; however, our understanding of the molecular mechanisms underlying nutrient signal transduction is very limited. To begin unraveling complex sugar signaling networks in plants, DNA microarray analysis was used to determine the effects of glucose and inorganic nitrogen source on gene expression on a global scale in Arabidopsis thaliana. In whole seedling tissue, glucose is a more potent signal in regulating transcription than inorganic nitrogen. In fact, other than genes associated with nitrate assimilation, glucose had a greater effect in regulating nitrogen metabolic genes than nitrogen itself. Glucose also regulated a broader range of genes, including genes associated with carbohydrate metabolism, signal transduction, and metabolite transport. In addition, a large number of stress responsive genes were also induced by glucose, indicating a role of sugar in environmental responses. Cluster analysis revealed significant interaction between glucose and nitrogen in regulating gene expression because glucose can modulate the effects of nitrogen and vise versa. Intriguingly, cycloheximide treatment appeared to disrupt glucose induction more than glucose repression, suggesting that de novo protein synthesis is an intermediary event required before most glucose induction can occur. Cross talk between sugar and ethylene signaling may take place on the transcriptional level because several ethylene biosynthetic and signal transduction genes are repressed by glucose, and the repression is largely unaffected by cycloheximide. Collectively, our global expression data strongly support the idea that glucose and inorganic nitrogen act as both metabolites and signaling molecules. PMID:15273295

  11. Image informatics for studying signal transduction in cells interacting with 3D matrices

    NASA Astrophysics Data System (ADS)

    Tzeranis, Dimitrios S.; Guo, Jin; Chen, Chengpin; Yannas, Ioannis V.; Wei, Xunbin; So, Peter T. C.

    2014-03-01

    Cells sense and respond to chemical stimuli on their environment via signal transduction pathways, complex networks of proteins whose interactions transmit chemical information. This work describes an implementation of image informatics, imaging-based methodologies for studying signal transduction networks. The methodology developed focuses on studying signal transduction networks in cells that interact with 3D matrices. It utilizes shRNA-based knock down of network components, 3D high-content imaging of cells inside the matrix by spectral multi-photon microscopy, and single-cell quantification using features that describe both cell morphology and cell-matrix adhesion pattern. The methodology is applied in a pilot study of TGFβ signaling via the SMAD pathway in fibroblasts cultured inside porous collagen-GAG scaffolds, biomaterials similar to the ones used clinically to induce skin regeneration. Preliminary results suggest that knocking down all rSMAD components affects fibroblast response to TGFβ1 and TGFβ3 isoforms in different ways, and suggest a potential role for SMAD1 and SMAD5 in regulating TGFβ isoform response. These preliminary results need to be verified with proteomic results that can provide solid evidence about the particular role of individual components of the SMAD pathway.

  12. HC toxin (a HDAC inhibitor) enhances IRS1-Akt signalling and metabolism in mouse myotubes.

    PubMed

    Tan, Hayden Weng Siong; Sim, Arthur Yi Loong; Huang, Su Ling; Leng, Ying; Long, Yun Chau

    2015-12-01

    Exercise enhances numerous signalling pathways and activates substrate metabolism in skeletal muscle. Small molecule compounds that activate these cellular responses have been shown to recapitulate the metabolic benefits of exercise. In this study, a histone deacetylase (HDAC) inhibitor, HC toxin, was investigated as a small molecule compound that activates exercise-induced adaptations. In C2C12 myotubes, HC toxin treatment activated two exercise-stimulated pathways: AMP-activated protein kinase (AMPK) and Akt pathways. HC toxin increased the protein content and phosphorylation of insulin receptor substrate 1 as well as the activation of downstream Akt signalling. The effects of HC toxin on IRS1-Akt signalling were PI3K-dependent as wortmannin abolishes its effects on IRS1 protein accumulation and Akt phosphorylation. HC toxin-induced Akt activation was sufficient to enhance downstream mTOR complex 1 (mTORC1) signalling including p70S6K and S6, which were consistently abolished by PI3K inhibition. Insulin-stimulated glucose uptake, glycolysis, mitochondrial respiration and fatty acid oxidation were also enhanced in HC toxin-treated myotubes. When myotubes were challenged with serum starvation for the induction of atrophy, HC toxin treatment prevented the induction of genes that are involved in autophagy and proteasomal proteolysis. Conversely, IRS1-Akt signalling was not induced by HC toxin in several hepatoma cell lines, providing evidence for a favourable safety profile of this small molecule. These data highlight the potential of HDAC inhibitors as a novel class of small molecules for the induction of exercise-like signalling pathways and metabolism. PMID:26373795

  13. A hypothesis-effect of T cell epitope fusion peptide specific immunotherapy on signal transduction

    PubMed Central

    Li, Chao-Pin; Yang, Bang-He

    2015-01-01

    Asthma is a chronic nonspecific inflammatory disease of the airway primarily mediated by different inflammatory cells, including mast cells, eosinophils and T cells. We hereby specially focused on a signal pathway for Janus kinase-signal transducer and activators of transduction (JAK-STATs), which has been the interest of study in asthma since it more likely regulates cellular proliferation and differentiation, and consequently modulates immune system. In our consideration, knowledge on this signal pathway may provide an avenue for rational options in treatment of asthma on control of immune response basis. PMID:26770626

  14. GD2 ganglioside specific antibody treatment downregulates PI3K/Akt/mTOR signaling network in human neuroblastoma cell lines.

    PubMed

    Durbas, Małgorzata; Horwacik, Irena; Boratyn, Elżbieta; Kamycka, Elżbieta; Rokita, Hanna

    2015-09-01

    Mechanisms leading to inhibitory effects of an anti-GD2 ganglioside (GD2) 14G2a mouse monoclonal antibody (mAb) and PI3K/Akt/mTOR pathway inhibitors on human neuroblastoma cell survival were studied in vitro. We have recently shown on IMR-32, CHP‑134, and LA-N-1 neuroblastoma cells that targeting GD2 with the mAb decreases cell viability of the cell lines. In this study we used cytotoxicity assays, proteomic arrays and immunoblotting to evaluate the response of the three cell lines to the anti‑GD2 14G2a mAb and specific PI3K/Akt/mTOR pathway inhibitors. We show here that the mAb modulates intracellular signal transduction through changes in several kinases and their substrates phosphorylation. More detailed analysis of the PI3K/Akt/mTOR pathway showed significant decrease in activity of Akt, mTOR, p70 S6 and 4E-BP1 proteins and transient increase in PTEN (a suppressor of the pathway), leading to inhibition of the signaling network responsible for stimulation of translation and proliferation. Additionally, combining the GD2-specific 14G2a mAb with an Akt inhibitor (perifosine), dual mTOR/PI3K inhibitors (BEZ-235 and SAR245409), and a pan-PI3K inhibitor (LY294002) was shown to enhance cytotoxic effects against IMR-32, CHP‑134 and LA-N-1 cells. Our study extends knowledge on mechanisms of action of the 14G2a mAb on the neuroblastoma cells. Also, it stresses the need for further delineation of molecular signal orchestration aimed at more reasonable selection of drugs to target key cellular pathways in quest for better cure for neuroblastoma patients. PMID:26134970

  15. Role of functionality in two-component signal transduction: A stochastic study

    NASA Astrophysics Data System (ADS)

    Maity, Alok Kumar; Bandyopadhyay, Arnab; Chaudhury, Pinaki; Banik, Suman K.

    2014-03-01

    We present a stochastic formalism for signal transduction processes in a bacterial two-component system. Using elementary mass action kinetics, the proposed model takes care of signal transduction in terms of a phosphotransfer mechanism between the cognate partners of a two-component system, viz., the sensor kinase and the response regulator. Based on the difference in functionality of the sensor kinase, the noisy phosphotransfer mechanism has been studied for monofunctional and bifunctional two-component systems using the formalism of the linear noise approximation. Steady-state analysis of both models quantifies different physically realizable quantities, e.g., the variance, the Fano factor (variance/mean), and mutual information. The resultant data reveal that both systems reliably transfer information of extracellular environment under low external stimulus and in a high-kinase-and-phosphatase regime. We extend our analysis further by studying the role of the two-component system in downstream gene regulation.

  16. Role of functionality in two-component signal transduction: a stochastic study.

    PubMed

    Maity, Alok Kumar; Bandyopadhyay, Arnab; Chaudhury, Pinaki; Banik, Suman K

    2014-03-01

    We present a stochastic formalism for signal transduction processes in a bacterial two-component system. Using elementary mass action kinetics, the proposed model takes care of signal transduction in terms of a phosphotransfer mechanism between the cognate partners of a two-component system, viz., the sensor kinase and the response regulator. Based on the difference in functionality of the sensor kinase, the noisy phosphotransfer mechanism has been studied for monofunctional and bifunctional two-component systems using the formalism of the linear noise approximation. Steady-state analysis of both models quantifies different physically realizable quantities, e.g., the variance, the Fano factor (variance/mean), and mutual information. The resultant data reveal that both systems reliably transfer information of extracellular environment under low external stimulus and in a high-kinase-and-phosphatase regime. We extend our analysis further by studying the role of the two-component system in downstream gene regulation. PMID:24730880

  17. The AKT/mTOR signaling pathway plays a key role in statin-induced myotoxicity.

    PubMed

    Bonifacio, Annalisa; Sanvee, Gerda M; Bouitbir, Jamal; Krähenbühl, Stephan

    2015-08-01

    Statins are drugs that lower blood cholesterol levels and reduce cardiovascular morbidity and mortality. They are generally well-tolerated, but myopathy is a potentially severe adverse reaction of these compounds. The mechanisms by which statins induce myotoxicity are not completely understood, but may be related to inhibition of the AKT signaling pathway. The current studies were performed to explore the down-stream effects of the statin-associated inhibition of AKT within the AKT signaling pathway and on myocyte biology and morphology in C2C12 myotubes and in mice in vivo. We exposed C2C12 myotubes to 10 μM or 50 μM simvastatin, atorvastatin or rosuvastatin for 24 h. Simvastatin and atorvastatin inhibited AKT phosphorylation and were cytotoxic starting at 10 μM, whereas similar effects were observed for rosuvastatin at 50 μM. Inhibition of AKT phosphorylation was associated with impaired phosphorylation of S6 kinase, ribosomal protein S6, 4E-binding protein 1 and FoxO3a, resulting in reduced protein synthesis, accelerated myofibrillar degradation and atrophy of C2C12 myotubes. Furthermore, impaired AKT phosphorylation was associated with activation of caspases and PARP, reflecting induction of apoptosis. Similar findings were detected in skeletal muscle of mice treated orally with 5 mg/kg/day simvastatin for 3 weeks. In conclusion, this study highlights the importance of the AKT/mTOR signaling pathway in statin-induced myotoxicity and reveals potential drug targets for treatment of patients with statin-associated myopathies. PMID:25913013

  18. miR-302 Attenuates Amyloid-β-Induced Neurotoxicity through Activation of Akt Signaling.

    PubMed

    Li, Hsin-Hua; Lin, Shi-Lung; Huang, Chien-Ning; Lu, Fung-Jou; Chiu, Pai-Yi; Huang, Wen-Nung; Lai, Te-Jen; Lin, Chih-Li

    2016-01-01

    Deficiency of insulin signaling has been linked to diabetes and ageing-related neurodegenerative diseases such as Alzheimer's disease (AD). In this regard, brains exhibit defective insulin receptor substrate-1 (IRS-1) and hence result in alteration of insulin signaling in progression of AD, the most common cause of dementia. Consequently, dysregulation of insulin signaling plays an important role in amyloid-β (Aβ)-induced neurotoxicity. As the derivation of induced pluripotent stem cells (iPSC) involves cell reprogramming, it may provide a means for regaining the control of ageing-associated dysfunction and neurodegeneration via affecting insulin-related signaling. To this, we found that an embryonic stem cell (ESC)-specific microRNA, miR-302, silences phosphatase and tensin homolog (PTEN) to activate Akt signaling, which subsequently stimulates nuclear factor erythroid 2-related factor 2 (Nrf2)/heme oxygenase-1 (HO-1) elevation and hence inhibits Aβ-induced neurotoxicity. miR-302 is predominantly expressed in iPSCs and is known to regulate several important biological processes of anti-oxidative stress, anti-apoptosis, and anti-aging through activating Akt signaling. In addition, we also found that miR-302-mediated Akt signaling further stimulates Nanog expression to suppress Aβ-induced p-Ser307 IRS-1 expression and thus enhances tyrosine phosphorylation and p-Ser 473-Akt/p-Ser 9-GSK3β formation. Furthermore, our in vivo studies revealed that the mRNA expression levels of both Nanog and miR-302-encoding LARP7 genes were significantly reduced in AD patients' blood cells, providing a novel diagnosis marker for AD. Taken together, our findings demonstrated that miR-302 is able to inhibit Aβ-induced cytotoxicity via activating Akt signaling to upregulate Nrf2 and Nanog expressions, leading to a marked restoration of insulin signaling in AD neurons. PMID:26890744

  19. Escin activates AKT-Nrf2 signaling to protect retinal pigment epithelium cells from oxidative stress.

    PubMed

    Wang, Kaijun; Jiang, Yiqian; Wang, Wei; Ma, Jian; Chen, Min

    2015-12-25

    Here we explored the anti-oxidative and cytoprotective potentials of escin, a natural triterpene-saponin, against hydrogen peroxide (H2O2) in retinal pigment epithelium (RPE) cells. We showed that escin remarkably attenuated H2O2-induced death and apoptosis of established (ARPE-19) and primary murine RPE cells. Meanwhile, ROS production and lipid peroxidation by H2O2 were remarkably inhibited by escin. Escin treatment in RPE cells resulted in NF-E2-related factor 2 (Nrf2) signaling activation, evidenced by transcription of anti-oxidant-responsive element (ARE)-regulated genes, including HO-1, NQO-1 and SRXN-1. Knockdown of Nrf2 through targeted shRNAs/siRNAs alleviated escin-mediated ARE gene transcription, and almost abolished escin-mediated anti-oxidant activity and RPE cytoprotection against H2O2. Reversely, escin was more potent against H2O2 damages in Nrf2-over-expressed ARPE-19 cells. Further studies showed that escin-induced Nrf2 activation in RPE cells required AKT signaling. AKT inhibitors (LY294002 and perifosine) blocked escin-induced AKT activation, and dramatically inhibited Nrf2 phosphorylation, its cytosol accumulation and nuclear translocation in RPE cells. Escin-induced RPE cytoprotection against H2O2 was also alleviated by the AKT inhibitors. Together, these results demonstrate that escin protects RPE cells from oxidative stress possibly through activating AKT-Nrf2 signaling. PMID:26505797

  20. Antitumor Effects of Fucoidan on Human Colon Cancer Cells via Activation of Akt Signaling

    PubMed Central

    Han, Yong-seok; Lee, Jun Hee; Lee, Sang Hun

    2015-01-01

    We identified a novel Akt signaling mechanism that mediates fucoidan-induced suppression of human colon cancer cell (HT29) proliferation and anticancer effects. Fucoidan treatment significantly inhibited growth, induced G1-phase-associated upregulation of p21WAF1 expression, and suppressed cyclin and cyclin-dependent kinase expression in HT29 colon cancer cells. Additionally, fucoidan treatment activated the Akt signaling pathway, which was inhibited by treatment with an Akt inhibitor. The inhibition of Akt activation reversed the fucoidan-induced decrease in cell proliferation, the induction of G1-phase-associated p21WAF1 expression, and the reduction in cell cycle regulatory protein expression. Intraperitoneal injection of fucoidan reduced tumor volume; this enhanced antitumor efficacy was associated with induction of apoptosis and decreased angiogenesis. These data suggest that the activation of Akt signaling is involved in the growth inhibition of colon cancer cells treated with fucoidan. Thus, fucoidan may serve as a potential therapeutic agent for colon cancer. PMID:25995820

  1. Signal transduction in human pancreatic cancer: roles of transforming growth factor beta, somatostatin receptors, and other signal intermediates.

    PubMed

    Li, Min; Becnel, Lauren S; Li, Wei; Fisher, William E; Chen, Changyi; Yao, Qizhi

    2005-01-01

    Pancreatic cancer is a devastating disease because of the lack of early detection markers and effective treatments. It is the fourth leading cause of cancer-related death in western countries, including the United States. The mechanisms of pancreatic cancer progression remain unknown. Transforming growth factor beta (TGF-beta), a multifunctional cytokine, regulates cell growth and differentiation in healthy tissues, yet fails to do so in pancreatic cancer. Alterations of the TGF-beta and TGF-beta receptor/Smad signal transduction pathway have been implicated in pancreatic cancer. Furthermore, both the TGF-beta receptor and Smad proteins interact with a variety of cellular signal pathways, such as the somatostatin receptors (SSTRs), ERK1/2, and Wnt signal transduction cascades. This suggests that pancreatic cancer is a multi-gene-controlled malignancy and that effective treatments for pancreatic cancer should be aimed at multiple targets. In this review, we summarized the major signal intermediates involved in pancreatic cancer signal transduction pathways and specifically discussed how alterations in the regulatory functions of TGF-beta and Smad proteins allow for pancreatic carcinogenesis. PMID:16314822

  2. Role of Glycolytic Intermediates in Global Regulation and Signal Transduction. Final Report

    SciTech Connect

    Liao, J.C.

    2000-05-08

    The goal of this project is to determine the role of glycolytic intermediates in regulation of cell physiology. It is known that many glycolytic intermediates are involved in regulation of enzyme activities at the kinetic level. However, little is known regarding the role of these metabolites in global regulation and signal transduction. This project aims to investigate the role of glycolytic intermediates in the regulation of gene expression.

  3. Structure of the signal transduction protein TRAP (target of RNAIII-activating protein)

    PubMed Central

    Henrick, Kim; Hirshberg, Miriam

    2012-01-01

    The crystal structure of the signal transduction protein TRAP is reported at 1.85 Å resolution. The structure of TRAP consists of a central eight-stranded β-­barrel flanked asymmetrically by helices and is monomeric both in solution and in the crystal structure. A formate ion was found bound to TRAP identically in all four molecules in the asymmetric unit. PMID:22750855

  4. Calcium and protein phosphorylation in the transduction of gravity signal in corn roots

    NASA Technical Reports Server (NTRS)

    Friedmann, M.; Poovaiah, B. W.

    1991-01-01

    The involvement of calcium and protein phosphorylation in the transduction of gravity signal was studied using corn roots of a light-insensitive variety (Zea mays L., cv. Patriot). The gravitropic response was calcium-dependent. Horizontal placement of roots preloaded with 32P for three minutes resulted in changes in protein phosphorylation of polypeptides of 32 and 35 kD. Calcium depletion resulted in decreased phosphorylation of these phosphoproteins and replenishment of calcium restored the phosphorylation.

  5. Myocardial ischemic post-conditioning attenuates ischemia reperfusion injury via PTEN/Akt signal pathway

    PubMed Central

    Li, Chun-Mei; Shen, Shu-Wen; Wang, Tao; Zhang, Xing-Hua

    2015-01-01

    Objectives: To investigate whether myocardial ischemic post-conditioning attenuates ischemia reperfusion injury via PTEN/Akt signal pathway. Design: Forty-five male Sprague-Dawley rats were randomly divided into three groups: Sham, Ischemia reperfusion (I/R) and Ischemic post-conditioning (IPost) group. After the experiment finished, myocardial infarction area was examined. Serum creatine phosphokinase and lactate dehydrogenase activity were detected at baseline and the end of reperfusion. The protein levels of PTEN, Akt, p-Akt, Bax and Bcl-2 were measured by Western blot method. Results: Myocardial infarct size was significantly reduced in IPost as compared to I/R. Results were confirmed by serum creatine phosphokinase and lactate dehydrogenase activity. In addition, PTEN and Bax protein expression were inhibited and the p-Akt and bcl-2 protein expression were enhanced in IPost compared with I/R (P < 0.05). At the same time, the ratio of Bax and Bcl-2 was decreased in IPost (P < 0.05). However, ischemic post conditioning did not affect the total Akt level (P > 0.05). Conclusions: We confirmed that ischemic post-conditioning protects the heart against reperfusion injury. It is important that we demonstrated that the cardioprotective effect of ischemic post-conditioning was involved in the inhibition of PTEN, activation of the PI3K/Akt pathway and reduction of the cardiomyocyte apoptosis. PMID:26629079

  6. O-GlcNAcylation regulates ischemia-induced neuronal apoptosis through AKT signaling

    PubMed Central

    Shi, Jianhua; Gu, Jin-hua; Dai, Chun-ling; Gu, Jianlan; Jin, Xiaoxia; Sun, Jianming; Iqbal, Khalid; Liu, Fei; Gong, Cheng-Xin

    2015-01-01

    Apoptosis plays an important role in neural development and neurological disorders. In this study, we found that O-GlcNAcylation, a unique protein posttranslational modification with O-linked β-N-acetylglucosamine (GlcNAc), promoted apoptosis through attenuating phosphorylation/activation of AKT and Bad. By using co-immunoprecipitation and mutagenesis techniques, we identified O-GlcNAc modification at both Thr308 and Ser473 of AKT. O-GlcNAcylation-induced apoptosis was attenuated by over-expression of AKT. We also found a dynamic elevation of protein O-GlcNAcylation during the first four hours of cerebral ischemia, followed by continuous decline after middle cerebral artery occlusion (MCAO) in the mouse brain. The elevation of O-GlcNAcylation coincided with activation of cell apoptosis. Finally, we found a negative correlation between AKT phosphorylation and O-GlcNAcylation in ischemic brain tissue. These results indicate that cerebral ischemia induces a rapid increase of O-GlcNAcylation that promotes apoptosis through down-regulation of AKT activity. These findings provide a novel mechanism through which O-GlcNAcylation regulates ischemia-induced neuronal apoptosis through AKT signaling. PMID:26412745

  7. Influence of arsenate and arsenite on signal transduction pathways: an update

    PubMed Central

    Druwe, Ingrid L.

    2010-01-01

    Arsenic has been a recognized contaminant and toxicant, as well as a medicinal compound throughout human history. Populations throughout the world are exposed to arsenic and these exposures have been associated with a number of human cancers. Not much is known about the role of arsenic as a human carcinogen and more recently its role in non-cancerous diseases, such as cardiovascular disease, hypertension and diabetes mellitus have been uncovered. The health effects associated with arsenic are numerous and the association between arsenic exposure and human disease has intensified the search for molecular mechanisms that describe the biological activity of arsenic in humans and leads to the aforementioned disease states. Arsenic poses a human health risk due in part to the regulation of cellular signal transduction pathways and over the last few decades, some cellular mechanisms that account for arsenic toxicity, as well as, signal transduction pathways have been discovered. However, given the ubiquitous nature of arsenic in the environment, making sense of all the data remains a challenge. This review will focus on our knowledge of signal transduction pathways that are regulated by arsenic. PMID:20502880

  8. Differential effects of ethanol on glial signal transduction initiated by lipopolysaccharide and interferon-gamma.

    PubMed

    Choi, Dong-Kug; Lee, Heasuk; Jeong, Jaeyoon; Lim, Beongou; Suk, Kyoungho

    2005-10-15

    Although the pathogenic effects of alcohol abuse on brain are well established, its specific effects on the intracellular signal transduction pathways of glial cells in the central nervous system (CNS) are poorly understood. In this study, we evaluated how ethanol affects the glial signal transduction associated with inflammatory activation. Lipopolysaccharide (LPS), gangliosides, and interferon (IFN)-gamma induced the inflammatory activation of glia, which was differentially influenced by ethanol: 1) ethanol inhibited LPS- or gangliosides-induced, but not IFNgamma-induced, glial activation as demonstrated by the production of nitric oxide and the expression of inflammatory genes such as interleukin-1beta, tumor necrosis factor-alpha, IP-10, and CD86; 2) nuclear factor (NF)-kappaB or JAK/STAT1 pathway was necessary for LPS- or IFNgamma-induced glial activation, respectively; 3) ethanol inhibited LPS-induced NF-kappaB activation; and 4) ethanol did not significantly affect IFNgamma-induced STAT1/IRF-1 activation. Based on these results, ethanol seems to inhibit selectively some parts of the glial signal transduction pathways that are associated with inflammatory activation, which may lead to the deregulation of CNS inflammatory responses. PMID:16175582

  9. Receptor clustering affects signal transduction at the membrane level in the reaction-limited regime

    NASA Astrophysics Data System (ADS)

    Caré, Bertrand R.; Soula, Hédi A.

    2013-01-01

    Many types of membrane receptors are found to be organized as clusters on the cell surface. We investigate the potential effect of such receptor clustering on the intracellular signal transduction stage. We consider a canonical pathway with a membrane receptor (R) activating a membrane-bound intracellular relay protein (G). We use Monte Carlo simulations to recreate biochemical reactions using different receptor spatial distributions and explore the dynamics of the signal transduction. Results show that activation of G by R is severely impaired by R clustering, leading to an apparent blunted biological effect compared to control. Paradoxically, this clustering decreases the half maximal effective dose (ED50) of the transduction stage, increasing the apparent affinity. We study an example of inter-receptor interaction in order to account for possible compensatory effects of clustering and observe the parameter range in which such interactions slightly counterbalance the loss of activation of G. The membrane receptors’ spatial distribution affects the internal stages of signal amplification, suggesting a functional role for membrane domains and receptor clustering independently of proximity-induced receptor-receptor interactions.

  10. PI3K/Akt/mTOR signaling pathway in cancer stem cells: from basic research to clinical application

    PubMed Central

    Xia, Pu; Xu, Xiao-Yan

    2015-01-01

    Cancer stem cells (CSCs) are a subpopulation of tumor cells that possess unique self-renewal activity and mediate tumor initiation and propagation. The PI3K/Akt/mTOR signaling pathway can be considered as a master regulator for cancer. More and more recent studies have shown the links between PI3K/Akt/mTOR signaling pathway and CSC biology. Herein, we provide a comprehensive review on the role of signaling components upstream and downstream of PI3K/Akt/mTOR signaling in CSC. In addition, we also summarize various classes of small molecule inhibitors of PI3K/Akt/mTOR signaling pathway and their clinical potential in CSC. Overall, the current available data suggest that the PI3K/Akt/mTOR signaling pathway could be a promising target for development of CSC-target drugs. PMID:26175931

  11. Gravity persistent signal 1 reveals a novel cytochrome P450 involved in gravitropic signal transduction

    NASA Astrophysics Data System (ADS)

    Wyatt, Sarah

    is involved in gravitropic signal transduction. (Partially support by NSF: 0618506 to SEW)

  12. Inhibition of TYRO3/Akt signaling participates in hypoxic injury in hippocampal neurons

    PubMed Central

    Zhu, Yan-zhen; Wang, Wei; Xian, Na; Wu, Bing

    2016-01-01

    In this study, we investigated the role of the TYRO3/Akt signaling pathway in hypoxic injury to hippocampal neurons. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay showed that hypoxia inhibited the proliferation and viability of hippocampal neurons. Terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling assay demonstrated that hypoxia induced neuronal apoptosis in a time-dependent manner, with a greater number of apoptotic cells with longer hypoxic exposure. Immunofluorescence labeling revealed that hypoxia suppressed TYRO3 expression. Western blot assay showed that hypoxia decreased Akt phosphorylation levels in a time-dependent manner. Taken together, these findings suggest that hypoxia inhibits the proliferation of hippocampal neurons and promotes apoptosis, and that the inhibition of the TYRO3/Akt signaling pathway plays an important role in hypoxia-induced neuronal injury. PMID:27335558

  13. The Stimulatory Gαs Protein Is Involved in Olfactory Signal Transduction in Drosophila

    PubMed Central

    Deng, Ying; Zhang, Weiyi; Farhat, Katja; Oberland, Sonja; Gisselmann, Günter; Neuhaus, Eva M.

    2011-01-01

    Seven-transmembrane receptors typically mediate olfactory signal transduction by coupling to G-proteins. Although insect odorant receptors have seven transmembrane domains like G-protein coupled receptors, they have an inverted membrane topology, constituting a key difference between the olfactory systems of insects and other animals. While heteromeric insect ORs form ligand-activated non-selective cation channels in recombinant expression systems, the evidence for an involvement of cyclic nucleotides and G-proteins in odor reception is inconsistent. We addressed this question in vivo by analyzing the role of G-proteins in olfactory signaling using electrophysiological recordings. We found that Gαs plays a crucial role for odorant induced signal transduction in OR83b expressing olfactory sensory neurons, but not in neurons expressing CO2 responsive proteins GR21a/GR63a. Moreover, signaling of Drosophila ORs involved Gαs also in a heterologous expression system. In agreement with these observations was the finding that elevated levels of cAMP result in increased firing rates, demonstrating the existence of a cAMP dependent excitatory signaling pathway in the sensory neurons. Together, we provide evidence that Gαs plays a role in the OR mediated signaling cascade in Drosophila. PMID:21490930

  14. Regulation of drug-induced liver injury by signal transduction pathways: critical role of mitochondria.

    PubMed

    Han, Derick; Dara, Lily; Win, Sanda; Than, Tin Aung; Yuan, Liyun; Abbasi, Sadeea Q; Liu, Zhang-Xu; Kaplowitz, Neil

    2013-04-01

    Drugs that cause liver injury often 'stress' mitochondria and activate signal transduction pathways important in determining cell survival or death. In most cases, hepatocytes adapt to the drug-induced stress by activating adaptive signaling pathways, such as mitochondrial adaptive responses and nuclear factor erythroid 2-related factor 2 (Nrf-2), a transcription factor that upregulates antioxidant defenses. Owing to adaptation, drugs alone rarely cause liver injury, with acetaminophen (APAP) being the notable exception. Drug-induced liver injury (DILI) usually involves other extrinsic factors, such as the adaptive immune system, that cause 'stressed' hepatocytes to become injured, leading to idiosyncratic DILI, the rare and unpredictable adverse drug reaction in the liver. Hepatocyte injury, due to drug and extrinsic insult, causes a second wave of signaling changes associated with adaptation, cell death, and repair. If the stress and injury reach a critical threshold, then death signaling pathways such as c-Jun N-terminal kinase (JNK) become dominant and hepatocytes enter a failsafe mode to undergo self-destruction. DILI can be seen as an active process involving recruitment of death signaling pathways that mediate cell death rather than a passive process due to overwhelming biochemical injury. In this review, we highlight the role of signal transduction pathways, which frequently involve mitochondria, in the development of DILI. PMID:23453390

  15. The Role of PI3K/Akt/mTOR Signaling in Gastric Carcinoma

    PubMed Central

    Matsuoka, Tasuku; Yashiro, Masakazu

    2014-01-01

    The phosphatidylinositol 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) pathway is one of the key signaling pathways induced by various receptor-tyrosine kinases. Accumulating evidence shows that this pathway is an important promoter of cell growth, metabolism, survival, metastasis, and resistance to chemotherapy. Genetic alterations in the PI3K/Akt/mTOR pathway in gastric carcinoma have often been demonstrated. Many kinds of molecular targeting therapies are currently undergoing clinical testing in patients with solid tumors. However, with the exception of the ErbB2-targeting antibody, targeting agents, including PI3K/Akt/mTOR inhibitors, have not been approved for treatment of patients with gastric carcinoma. This review summarizes the current knowledge on PI3K/Akt/mTOR signaling in the pathogenesis of gastric carcinoma and the possible therapeutic targets for gastric carcinoma. Improved knowledge of the PI3K/Akt/mTOR pathway in gastric carcinoma will be useful in understanding the mechanisms of tumor development and for identifying ideal targets of anticancer therapy for gastric carcinoma. PMID:25003395

  16. Prognostic significance of XIAP expression in DLBCL and effect of its inhibition on AKT signalling.

    PubMed

    Hussain, Azhar R; Uddin, Shahab; Ahmed, Maqbool; Bu, Rong; Ahmed, Saeeda O; Abubaker, Jehad; Sultana, Mehar; Ajarim, Dahish; Al-Dayel, Fouad; Bavi, Prashant P; Al-Kuraya, Khawla S

    2010-10-01

    The inhibitor of apoptosis protein (IAP) family member X-linked inhibitor of apoptosis protein (XIAP) is essential for cell survival in lymphoma. However, the role of XIAP overexpression in diffuse large B-cell lymphoma (DLBCL) is not fully elucidated. Therefore, we analysed the expression of XIAP protein and its clinicopathological correlation in a large cohort of DLBCLs by immunohistochemistry in a tissue micro-array format. XIAP was found to be overexpressed in 55% of DLBCLs and significantly associated with poor clinical outcome (p = 0.0421). To further elucidate the role of XIAP in DLBCL and the inter-relationship with PI3-kinase/AKT signalling, we conducted several in vitro studies using a panel of DLBCL cell lines. We found that pharmacological inhibition of XIAP led to caspase-dependent apoptosis in DLBCL cells. We also detected an inter-relationship between XIAP expression and activated AKT in DLBCL cells that may explain cellular resistance to PI3-kinase/AKT inhibition-mediated apoptosis. Finally, this anti-apoptotic effect was overcome by simultaneous pharmacological inhibition of XIAP and PI3-kinase/AKT signalling leading to a more potent synergistically induced apoptosis. In summary, our data suggest that XIAP expression is a poor prognostic factor in DLBCL and the XIAP-AKT relationship should be explored further as a potential therapeutic target in DLBCL. PMID:20632385

  17. The emerging role of phosphoinositide clustering in intracellular trafficking and signal transduction

    PubMed Central

    Picas, Laura; Gaits-Iacovoni, Frederique; Goud, Bruno

    2016-01-01

    Phosphoinositides are master regulators of multiple cellular processes: from vesicular trafficking to signaling, cytoskeleton dynamics, and cell growth. They are synthesized by the spatiotemporal regulated activity of phosphoinositide-metabolizing enzymes. The recent observation that some protein modules are able to cluster phosphoinositides suggests that alternative or complementary mechanisms might operate to stabilize the different phosphoinositide pools within cellular compartments. Herein, we discuss the different known and potential molecular players that are prone to engage phosphoinositide clustering and elaborate on how such a mechanism might take part in the regulation of intracellular trafficking and signal transduction. PMID:27092250

  18. New Signal Transduction Principles for Amperometric Enzyme and Antibody based Sensors

    NASA Astrophysics Data System (ADS)

    Warsinke, Axel

    2008-10-01

    The way of how the signal transfer from the analyte recognizing biocomponent to the sensor surface is performed influences strongly the characteristics of a biosensor e.g. response time, sensitivity and specificity. Most of the described amperometric enzyme sensors are using oxidases. The signal transduction is carried out simply by electrochemical indication of the produced hydrogen peroxide or via a sensor-immobilized redox polymer. However, due to the limited number of appropriate oxidases the range of detectable analytes is restricted. Hence, we have developed a new general principle for the sensitive transduction of the more than 400 different NAD(P) dependent dehydrogenase reactions. The transduction is based on a hydroxylase reaction which produces an electrochemically active substance under the consumption of NAD(P)H. The principle should be applicable to miniaturized sensor configuration and could be the basis for a new generation of point-of-care devices. For other analytes where no oxidases and dehydrogenases are available antibodies can be used as specific recognition element. We have developed a new principle of redox-labeled immunoassays called size exclusion redox-labeled immunoassay (SERI), where after the antigen antibody binding reaction the indication is carried out amperometrically without a washing step in between. The principle was proved for measurement of creatinine. At the moment the assay needs a relatively high amount of antibodies. However, in future it should be possible to reduce the amount of antibodies by using miniaturized microfluidic chips.

  19. Prion-like Polymerization Underlies Signal Transduction in Antiviral Immune Defense and Inflammasome Activation

    PubMed Central

    Cai, Xin; Chen, Jueqi; Xu, Hui; Liu, Siqi; Jiang, Qiu-Xing; Halfmann, Randal; Chen, Zhijian J.

    2014-01-01

    SUMMARY Pathogens and cellular danger signals activate sensors such as RIG-I and NLRP3 to produce robust immune and inflammatory responses through respective adaptor proteins MAVS and ASC, which harbor essential N-terminal CARD and PYRIN domains, respectively. Here, we show that CARD and PYRIN function as bona fide prions in yeast and their prion forms are inducible by their respective upstream activators. Likewise, a yeast prion domain can functionally replace CARD and PYRIN in mammalian cell signaling. Mutations in MAVS and ASC that disrupt their prion activities in yeast also abrogate their ability to signal in mammalian cells. Furthermore, fibers of recombinant PYRIN can convert ASC into functional polymers capable of activating caspase-1. Remarkably, a conserved fungal NOD-like receptor and prion pair can functionally reconstitute signaling of NLRP3 and ASC PYRINs in mammalian cells. These results indicate that prion-like polymerization is a conserved signal transduction mechanism in innate immunity and inflammation. PMID:24630723

  20. beta3-Adrenergic-dependent and -independent mechanisms participate in cold-induced modulation of insulin signal transduction in brown adipose tissue of rats.

    PubMed

    Gasparetti, Alessandra L; Alvarez-Rojas, Fernanda; de Araujo, Eliana P; Hirata, Aparecida E; Saad, Mário J A; Velloso, Lício A

    2005-03-01

    During cold exposure, homeothermic animals mobilize glucose with higher efficiency than at thermoneutrality. An interaction between the insulin signal transduction machinery and high sympathetic tonus is thought to play an important role in this phenomenon. In the present study, rats were exposed to cold during 8 days and treated, or not, with a beta3-adrenergic agonist, BRL37344 sodium 4-2-2-(3-chlorophenyl)-2-hydroxyethyl amino propyl phenoxy-acetic acid sodium (BRL37344), or antagonist, SR59230A 3-(2-ethylphenoxy)-[(1S)-1,2,3,4-tetrahydronaphth-1-ylamino]-(2S)-2-propanol oxalate (SR59230A), to evaluate the cross-talk between insulin and beta3-adrenergic intracellular signaling in brown adipose tissue. The drugs did not modify food ingestion, body temperature, and body weight in control and cold-exposed rats. Treatment of control rats with BRL37344 led to higher insulin-induced tyrosine phosphorylation of the insulin receptors, insulin receptor substrate (IRS)-1 and ERK, higher insulin-induced IRS-1/PI3-kinase association, and higher [Ser(473)] phosphorylation of Akt. Cold exposure alone promoted higher insulin-induced tyrosine phosphorylation of the insulin receptors, IRS-1, IRS-2, and ERK, and higher insulin-induced IRS-1 and IRS-2/PI3-kinase association. Except for the regulation of ERK, SR59230A abolished all the cold-induced effects upon the insulin signal transduction pathway. However, this antagonist only partially inhibited the cold-induced increase of glucose uptake. Thus, the sympathetic tonus generated during cold-exposure acts, in brown adipose tissue, through the beta3-adrenergic receptor and modulates insulin signal transduction, with the exception of ERK. However, insulin-independent mechanisms other than beta3-adrenergic activation participate in cold-induced glucose uptake in brown adipose tissue of rats. PMID:15750837

  1. A thermal signal generator probe for the study of neural thermal transduction.

    PubMed

    Maluf, N I; McNutt, E L; Monroe, S; Tanelian, D L; Kovacs, G T

    1994-07-01

    The study of thermal transduction in neural tissues has been impeded by the lack of instrumentation able to generate complex, focal temperature variations. Specifically, we are interested in the study of neural thermal transduction within the cornea, with its homogeneous thermal conductivity and avascularity. We present a thermal signal generator probe that is capable of producing arbitrarily shaped bipolar (heating or cooling) thermal swings in a small volume of corneal tissue with which it is in contact. Heating and cooling of the probe tip are achieved by means of a Peltier effect thermoelectric device. The probe temperature, measured directly at the tip, is controlled using closed-loop control circuitry and waveform generation software on a host computer. Response characteristics of thermally sensitive C-fibers were investigated in an in vitro preparation of the rabbit cornea. PMID:7927385

  2. The Clickable Guard Cell, Version II: Interactive Model of Guard Cell Signal Transduction Mechanisms and Pathways.

    PubMed

    Kwak, June M; Mäser, Pascal; Schroeder, Julian I

    2008-01-01

    Guard cells are located in the leaf epidermis and pairs of guard cells surround and form stomatal pores, which regulate CO(2) influx from the atmosphere into leaves for photosynthetic carbon fixation. Stomatal guard cells also regulate water loss of plants via transpiration to the atmosphere. Signal transduction mechanisms in guard cells integrate a multitude of different stimuli to modulate stomatal apertures. Stomata open in response to light. Stomata close in response to drought stress, elevated CO(2), ozone and low humidity. In response to drought, plants synthesize the hormone abscisic acid (ABA) that triggers closing of stomatal pores. Guard cells have become a highly developed model system for dissecting signal transduction mechanisms in plants and for elucidating how individual signaling mechanisms can interact within a network in a single cell. Many new findings have been made in the last few years. This chapter is an update of an electronic interactive chapter in the previous edition of The Arabidopsis Book (Mäser et al. 2003). Here we focus on mechanisms for which genes and mutations have been characterized, including signaling components for which there is substantial signaling, biochemical and genetic evidence. Ion channels have been shown to represent targets of early signal transduction mechanisms and provide functional signaling and quantitative analysis points to determine where and how mutations affect branches within the guard cell signaling network. Although a substantial number of genes and proteins that function in guard cell signaling have been identified in recent years, there are many more left to be identified and the protein-protein interactions within this network will be an important subject of future research. A fully interactive clickable electronic version of this publication can be accessed at the following web site: http://www-biology.ucsd.edu/labs/schroeder/clickablegc2/. The interactive clickable version includes the following

  3. Genome-Wide RNAi Screening to Dissect the TGF-β Signal Transduction Pathway.

    PubMed

    Chen, Xiaochu; Xu, Lan

    2016-01-01

    The transforming growth factor-β (TGF-β) family of cytokines figures prominently in regulation of embryonic development and adult tissue homeostasis from Drosophila to mammals. Genetic defects affecting TGF-β signaling underlie developmental disorders and diseases such as cancer in human. Therefore, delineating the molecular mechanism by which TGF-β regulates cell biology is critical for understanding normal biology and disease mechanisms. Forward genetic screens in model organisms and biochemical approaches in mammalian tissue culture were instrumental in initial characterization of the TGF-β signal transduction pathway. With complete sequence information of the genomes and the advent of RNA interference (RNAi) technology, genome-wide RNAi screening emerged as a powerful functional genomics approach to systematically delineate molecular components of signal transduction pathways. Here, we describe a protocol for image-based whole-genome RNAi screening aimed at identifying molecules required for TGF-β signaling into the nucleus. Using this protocol we examined >90 % of annotated Drosophila open reading frames (ORF) individually and successfully uncovered several novel factors serving critical roles in the TGF-β pathway. Thus cell-based high-throughput functional genomics can uncover new mechanistic insights on signaling pathways beyond what the classical genetics had revealed. PMID:26520138

  4. Stability and function of adult vasculature is sustained by Akt/Jagged1 signalling axis in endothelium

    PubMed Central

    Kerr, Bethany A.; West, Xiaoxia Z.; Kim, Young-Woong; Zhao, Yongzhong; Tischenko, Miroslava; Cull, Rebecca M.; Phares, Timothy W.; Peng, Xiao-Ding; Bernier-Latmani, Jeremiah; Petrova, Tatiana V.; Adams, Ralf H.; Hay, Nissim; Naga Prasad, Sathyamangla V.; Byzova, Tatiana V.

    2016-01-01

    The signalling pathways operational in quiescent, post-development vasculature remain enigmatic. Here we show that unlike neovascularization, endothelial Akt signalling in established vasculature is crucial not for endothelial cell (EC) survival, but for sustained interactions with pericytes and vascular smooth muscle cells (VSMCs) regulating vascular stability and function. Inducible endothelial-specific Akt1 deletion in adult global Akt2KO mice triggers progressive VSMC apoptosis. In hearts, this causes a loss of arteries and arterioles and, despite a high capillary density, diminished vascular patency and severe cardiac dysfunction. Similarly, endothelial Akt deletion induces retinal VSMC loss and basement membrane deterioration resulting in vascular regression and retinal atrophy. Mechanistically, the Akt/mTOR axis controls endothelial Jagged1 expression and, thereby, Notch signalling regulating VSMC maintenance. Jagged1 peptide treatment of Akt1ΔEC;Akt2KO mice and Jagged1 re-expression in Akt-deficient endothelium restores VSMC coverage. Thus, sustained endothelial Akt1/2 signalling is critical in maintaining vascular stability and homeostasis, thereby preserving tissue and organ function. PMID:26971877

  5. Stability and function of adult vasculature is sustained by Akt/Jagged1 signalling axis in endothelium.

    PubMed

    Kerr, Bethany A; West, Xiaoxia Z; Kim, Young-Woong; Zhao, Yongzhong; Tischenko, Miroslava; Cull, Rebecca M; Phares, Timothy W; Peng, Xiao-Ding; Bernier-Latmani, Jeremiah; Petrova, Tatiana V; Adams, Ralf H; Hay, Nissim; Naga Prasad, Sathyamangla V; Byzova, Tatiana V

    2016-01-01

    The signalling pathways operational in quiescent, post-development vasculature remain enigmatic. Here we show that unlike neovascularization, endothelial Akt signalling in established vasculature is crucial not for endothelial cell (EC) survival, but for sustained interactions with pericytes and vascular smooth muscle cells (VSMCs) regulating vascular stability and function. Inducible endothelial-specific Akt1 deletion in adult global Akt2KO mice triggers progressive VSMC apoptosis. In hearts, this causes a loss of arteries and arterioles and, despite a high capillary density, diminished vascular patency and severe cardiac dysfunction. Similarly, endothelial Akt deletion induces retinal VSMC loss and basement membrane deterioration resulting in vascular regression and retinal atrophy. Mechanistically, the Akt/mTOR axis controls endothelial Jagged1 expression and, thereby, Notch signalling regulating VSMC maintenance. Jagged1 peptide treatment of Akt1ΔEC;Akt2KO mice and Jagged1 re-expression in Akt-deficient endothelium restores VSMC coverage. Thus, sustained endothelial Akt1/2 signalling is critical in maintaining vascular stability and homeostasis, thereby preserving tissue and organ function. PMID:26971877

  6. Excitation and Adaptation in Bacteria–a Model Signal Transduction System that Controls Taxis and Spatial Pattern Formation

    PubMed Central

    Othmer, Hans G.; Xin, Xiangrong; Xue, Chuan

    2013-01-01

    The machinery for transduction of chemotactic stimuli in the bacterium E. coli is one of the most completely characterized signal transduction systems, and because of its relative simplicity, quantitative analysis of this system is possible. Here we discuss models which reproduce many of the important behaviors of the system. The important characteristics of the signal transduction system are excitation and adaptation, and the latter implies that the transduction system can function as a “derivative sensor” with respect to the ligand concentration in that the DC component of a signal is ultimately ignored if it is not too large. This temporal sensing mechanism provides the bacterium with a memory of its passage through spatially- or temporally-varying signal fields, and adaptation is essential for successful chemotaxis. We also discuss some of the spatial patterns observed in populations and indicate how cell-level behavior can be embedded in population-level descriptions. PMID:23624608

  7. Activated AKT/PKB signaling in C. elegans uncouples temporally distinct outputs of DAF-2/insulin-like signaling

    PubMed Central

    Gami, Minaxi S; Iser, Wendy B; Hanselman, Keaton B; Wolkow, Catherine A

    2006-01-01

    Background In the nematode, Caenorhabditis elegans, a conserved insulin-like signaling pathway controls larval development, stress resistance and adult lifespan. AGE-1, a homolog of the p110 catalytic subunit of phosphoinositide 3-kinases (PI3K) comprises the major known effector pathway downstream of the insulin receptor, DAF-2. Phospholipid products of AGE-1/PI3K activate AKT/PKB kinase signaling via PDK-1. AKT/PKB signaling antagonizes nuclear translocation of the DAF-16/FOXO transcription factor. Reduced AGE-1/PI3K signaling permits DAF-16 to direct dauer larval arrest and promote long lifespan in adult animals. In order to study the downstream effectors of AGE-1/PI3K signaling in C. elegans, we conducted a genetic screen for mutations that suppress the constitutive dauer arrest phenotype of age-1(mg109) animals. Results This report describes mutations recovered in a screen for suppressors of the constitutive dauer arrest (daf-C) phenotype of age-1(mg109). Two mutations corresponded to alleles of daf-16. Two mutations were gain-of-function alleles in the genes, akt-1 and pdk-1, encoding phosphoinositide-dependent serine/threonine kinases. A fifth mutation, mg227, located on chromosome X, did not correspond to any known dauer genes, suggesting that mg227 may represent a new component of the insulin pathway. Genetic epistasis analysis by RNAi showed that reproductive development in age-1(mg109);akt-1(mg247) animals was dependent on the presence of pdk-1. Similarly, reproductive development in age-1(mg109);pdk-1(mg261) animals was dependent on akt-1. However, reproductive development in age-1(mg109); mg227 animals required only akt-1, and pdk-1 activity was dispensable in this background. Interestingly, while mg227 suppressed dauer arrest in age-1(mg109) animals, it enhanced the long lifespan phenotype. In contrast, akt-1(mg247) and pdk-1(mg261) did not affect lifespan or stress resistance, while both daf-16 alleles fully suppressed these phenotypes. Conclusion A

  8. Subcutaneous Adipocytes Promote Melanoma Cell Growth by Activating the Akt Signaling Pathway

    PubMed Central

    Kwan, Hiu Yee; Fu, Xiuqiong; Liu, Bin; Chao, Xiaojuan; Chan, Chi Leung; Cao, Huihui; Su, Tao; Tse, Anfernee Kai Wing; Fong, Wang Fun; Yu, Zhi-Ling

    2014-01-01

    Tumorigenesis involves constant communication between tumor cells and neighboring normal cells such as adipocytes. The canonical function of adipocytes is to store triglyceride and release fatty acids for other tissues. This study was aimed to find out if adipocytes promoted melanoma cell growth and to investigate the underlying mechanism. Here we isolated adipocytes from inguinal adipose tissue in mice and co-cultured with melanoma cells. We found that the co-cultured melanoma had higher lipid accumulation compared with mono-cultured melanoma. In addition, fluorescently labeled fatty acid BODIPY® FLC16 signal was detected in melanoma co-cultured with the adipocytes that had been loaded with the fluorescent dye, suggesting that the adipocytes provide fatty acids to melanoma cells. Compared with mono-cultured melanoma, co-cultured melanoma cells had a higher proliferation and phospho-Akt (Ser-473 and Thr-450) expression. Overexpression of Akt mutants in melanoma cells reduced the co-culture-enhanced proliferation. A lipidomic study showed that the co-cultured melanoma had an elevated palmitic acid level. Interestingly, we found that palmitic acid stimulated melanoma cell proliferation, changed the cell cycle distribution, and increased phospho-Akt (Ser-473 and Thr-450) and PI3K but not phospho-PTEN (phosphophosphatase and tensin homolog) expressions. More importantly, the palmitic acid-stimulated proliferation was further enhanced in the Akt-overexpressed melanoma cells and was reduced by LY294002 or knockdown of endogenous Akt or overexpression of Akt mutants. We also found that palmitic acid-pretreated B16F10 cells were grown to a significantly larger tumor in mice compared with control cells. Taken together, we suggest that adipocytes may serve as an exogenous source of palmitic acid that promotes melanoma cell growth by activating Akt. PMID:25228694

  9. Do signal transduction cascades influence survival in triple-negative breast cancer? A preliminary study

    PubMed Central

    Mumm, Jan-Niclas; Kölbl, Alexandra C; Jeschke, Udo; Andergassen, Ulrich

    2016-01-01

    Background Triple-negative breast cancer (TNBC) is a rather aggressive form of breast cancer, comprised by early metastasis formation and reduced overall survival of the affected patients. Steroid hormone receptors and the human epidermal growth factor receptor 2 are not overexpressed, limiting therapeutic options. Therefore, new treatment options have to be investigated. The aim of our preliminary study was to detect coherences between some molecules of intracellular signal transduction pathways and survival of patients with TNBC, in order to obtain some hints for new therapeutical solutions. Methods Thirty-one paraffin-embedded tumor tissue samples, which were determined to be negative for steroid hormone receptors as well as human epidermal growth factor receptor 2, were immunohistochemically stained for a number of signal transduction molecules from several signaling pathways. β-Catenin, HIF1α, MCL, Notch1, LRP6, XBP1, and FOXP3 were stained with specific antibodies, and their staining was correlated with patient survival by Kaplan–Meier analyses. Results Only two of the investigated molecules have shown correlation with overall survival. Cytoplasmic staining of HIF1α and centro-tumoral lymphocyte FOXP3 staining showed statistically significant correlations with survival. Conclusion The coherence of signal transduction molecules with survival of patients with TNBC is still controversially discussed in the literature. Our study comprises one more mosaic stone in the elucidation of these intracellular processes and their influences on patient outcome. Lots of research still has to be done in this field, but it would be worthwhile as it may offer new therapeutic targets for a group of patients with breast cancer, which is still hard to treat. PMID:27307757

  10. Oxidative Stress in Fungi: Its Function in Signal Transduction, Interaction with Plant Hosts, and Lignocellulose Degradation

    PubMed Central

    Breitenbach, Michael; Weber, Manuela; Rinnerthaler, Mark; Karl, Thomas; Breitenbach-Koller, Lore

    2015-01-01

    In this review article, we want to present an overview of oxidative stress in fungal cells in relation to signal transduction, interaction of fungi with plant hosts, and lignocellulose degradation. We will discuss external oxidative stress which may occur through the interaction with other microorganisms or plant hosts as well as internally generated oxidative stress, which can for instance originate from NADPH oxidases or “leaky” mitochondria and may be modulated by the peroxiredoxin system or by protein disulfide isomerases thus contributing to redox signaling. Analyzing redox signaling in fungi with the tools of molecular genetics is presently only in its beginning. However, it is already clear that redox signaling in fungal cells often is linked to cell differentiation (like the formation of perithecia), virulence (in plant pathogens), hyphal growth and the successful passage through the stationary phase. PMID:25854186

  11. Spatial and Temporal Regulation of Receptor Tyrosine Kinase Activation and Intracellular Signal Transduction.

    PubMed

    Bergeron, John J M; Di Guglielmo, Gianni M; Dahan, Sophie; Dominguez, Michel; Posner, Barry I

    2016-06-01

    Epidermal growth factor (EGF) and insulin receptor tyrosine kinases (RTKs) exemplify how receptor location is coupled to signal transduction. Extracellular binding of ligands to these RTKs triggers their concentration into vesicles that bud off from the cell surface to generate intracellular signaling endosomes. On the exposed cytosolic surface of these endosomes, RTK autophosphorylation selects the downstream signaling proteins and lipids to effect growth factor and polypeptide hormone action. This selection is followed by the recruitment of protein tyrosine phosphatases that inactivate the RTKs and deliver them by membrane fusion and fission to late endosomes. Coincidentally, proteinases inside the endosome cleave the EGF and insulin ligands. Subsequent inward budding of the endosomal membrane generates multivesicular endosomes. Fusion with lysosomes then results in RTK degradation and downregulation. Through the spatial positioning of RTKs in target cells for EGF and insulin action, the temporal extent of signaling, attenuation, and downregulation is regulated. PMID:27023845

  12. Apoptosis prediction via inhibition of AKT signaling pathway by neogrifolin

    PubMed Central

    Chen, Yang; Peng, Guo-Fang; Han, Xiang-Zhen; Wang, Wei; Zhang, Guo-Qiang; Li, Xiao

    2015-01-01

    Neogrifolin, a natural biologically active substance isolated from the edible bodies of the mushroom Albatrellus confluens, has been shown to possess several pharmacological properties. No studies were investigated against osteosarcoma cancer. Hence, in this study, we investigated the apoptosis-inducing effects and the mechanisms of neogrifolin on human osteosarcoma cells. Our results demonstrated that neogrifolin induced concentration- and time-dependent suppression of proliferation. Further, induction of apoptosis in U2OS and MG63 osteosarcoma cell lines were also observed. Neogrifolin induced the release of cytochrome c accompanied by activation of caspase-9, caspase-3 and cleavage of poly (ADP-ribose) polymerase (PARP). In addition, z-VAD-fmk, a universal inhibitor of caspases, prevented caspase-3 activation and PARP cleavage and inhibited neogrifolin-induced cell growth inhibition. Furthermore, neogrifolin treatment resulted in a reduction of phosphorylated AKT level, FOXO transcription factor, and glycogen synthase kinase 3 (GSK3). Knockdown of GSK3 with siRNA inhibited the apoptotic effects of neogrifolin. On the other hand, neogrifolin treatment also down-regulated the expression of the inhibitor of apoptosis protein (IAP) in both osteosarcoma cells. Collectively, our results suggested that neogrifolin is a potential candidate for osteosarcoma. PMID:25973001

  13. Hesperidin Inhibits Vascular Formation by Blocking the AKT/mTOR Signaling Pathways.

    PubMed

    Kim, Gi Dae

    2015-12-01

    Hesperidin has been shown to possess a potential inhibitory effect on vascular formation in endothelial cells. However, the fundamental mechanism for the anti-angiogenic activity of hesperidin is not fully understood. In the present study, we evaluated whether hesperidin has anti-angiogenic effects in mouse embryonic stem cell (mES)-derived endothelial-like cells, and human umbilical vascular endothelial cells (HUVECs), and evaluated their mechanism via the AKT/mammalian target of rapamycin (mTOR) signaling pathway. The endothelial cells were treated with several doses of hesperidin (12.5, 25, 50, and 100 μM) for 24 h. Cell viability and vascular formation were analyzed using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and tube formation assay, respectively. Alteration of the AKT/mTOR signaling in vascular formation was analyzed by western blot. In addition, a mouse aortic ring assay was used to determine the effect of hesperidin on vascular formation. There were no differences between the viability of mES-derived endothelial-like cells and HUVECs after hesperidin treatment. However, hesperidin significantly inhibited cell migration and tube formation of HUVECs (P<0.05) and suppressed sprouting of microvessels in the mouse aortic ring assay. Moreover, hesperidin suppressed the expression of AKT and mTOR in HUVECs. Taken together, these findings suggest that hesperidin inhibits vascular formation by blocking the AKT/mTOR signaling pathways. PMID:26770908

  14. Grouper (Epinephelus coioides) MyD88 and Tollip: intracellular localization and signal transduction function.

    PubMed

    Li, Yan-Wei; Wang, Zheng; Mo, Ze-Quan; Li, Xia; Luo, Xiao-Chun; Dan, Xue-Ming; Li, An-Xing

    2015-01-01

    Myeloid differentiation factor 88 (MyD88) and Toll-interacting protein (Tollip) are two important regulatory proteins of the Toll-like receptor (TLR) signaling pathways. In this paper, a Tollip sequence of grouper (Epinephelus coioides) was identified and the signal transduction functions of Tollip and MyD88 were studied. The full length of E. coioides Tollip (EcTollip) cDNA with an open reading frame (ORF) of 1734 nucleotides encoded a putative protein of 274 amino acid residues. The EcTollip protein had conservative domains with mammalian homologous proteins, and high identity (78%-95%) with other vertebrates. MyD88 and Tollip were distributed in the HeLa cytoplasm in a highly condensed form. Over-expression of MyD88 could activate nuclear factor-κB (NF-κB) and its function was dependent on the death domain and ID domain on the N-terminal. Some important functional sites of mammalian MyD88 also affected fish MyD88 signal transduction. Tollip impaired NF-κB signals activated by MyD88, and its activity was dependent on the coupling of ubiquitin to the endoplasmic reticulum degradation (CUE) domain on the C-terminal. These results suggest that MyD88 and Tollip of fish and mammals are conservative on function during evolution. PMID:25449381

  15. Linear models of activation cascades: analytical solutions and coarse-graining of delayed signal transduction.

    PubMed

    Beguerisse-Díaz, Mariano; Desikan, Radhika; Barahona, Mauricio

    2016-08-01

    Cellular signal transduction usually involves activation cascades, the sequential activation of a series of proteins following the reception of an input signal. Here, we study the classic model of weakly activated cascades and obtain analytical solutions for a variety of inputs. We show that in the special but important case of optimal gain cascades (i.e. when the deactivation rates are identical) the downstream output of the cascade can be represented exactly as a lumped nonlinear module containing an incomplete gamma function with real parameters that depend on the rates and length of the cascade, as well as parameters of the input signal. The expressions obtained can be applied to the non-identical case when the deactivation rates are random to capture the variability in the cascade outputs. We also show that cascades can be rearranged so that blocks with similar rates can be lumped and represented through our nonlinear modules. Our results can be used both to represent cascades in computational models of differential equations and to fit data efficiently, by reducing the number of equations and parameters involved. In particular, the length of the cascade appears as a real-valued parameter and can thus be fitted in the same manner as Hill coefficients. Finally, we show how the obtained nonlinear modules can be used instead of delay differential equations to model delays in signal transduction. PMID:27581482

  16. Transmembrane signal transduction by peptide hormones via family B G protein-coupled receptors

    PubMed Central

    Culhane, Kelly J.; Liu, Yuting; Cai, Yingying; Yan, Elsa C. Y.

    2015-01-01

    Although family B G protein-coupled receptors (GPCRs) contain only 15 members, they play key roles in transmembrane signal transduction of hormones. Family B GPCRs are drug targets for developing therapeutics for diseases ranging from metabolic to neurological disorders. Despite their importance, the molecular mechanism of activation of family B GPCRs remains largely unexplored due to the challenges in expression and purification of functional receptors to the quantity for biophysical characterization. Currently, there is no crystal structure available of a full-length family B GPCR. However, structures of key domains, including the extracellular ligand binding regions and seven-helical transmembrane regions, have been solved by X-ray crystallography and NMR, providing insights into the mechanisms of ligand recognition and selectivity, and helical arrangements within the cell membrane. Moreover, biophysical and biochemical methods have been used to explore functions, key residues for signaling, and the kinetics and dynamics of signaling processes. This review summarizes the current knowledge of the signal transduction mechanism of family B GPCRs at the molecular level and comments on the challenges and outlook for mechanistic studies of family B GPCRs. PMID:26594176

  17. Linear models of activation cascades: analytical solutions and coarse-graining of delayed signal transduction

    PubMed Central

    Desikan, Radhika

    2016-01-01

    Cellular signal transduction usually involves activation cascades, the sequential activation of a series of proteins following the reception of an input signal. Here, we study the classic model of weakly activated cascades and obtain analytical solutions for a variety of inputs. We show that in the special but important case of optimal gain cascades (i.e. when the deactivation rates are identical) the downstream output of the cascade can be represented exactly as a lumped nonlinear module containing an incomplete gamma function with real parameters that depend on the rates and length of the cascade, as well as parameters of the input signal. The expressions obtained can be applied to the non-identical case when the deactivation rates are random to capture the variability in the cascade outputs. We also show that cascades can be rearranged so that blocks with similar rates can be lumped and represented through our nonlinear modules. Our results can be used both to represent cascades in computational models of differential equations and to fit data efficiently, by reducing the number of equations and parameters involved. In particular, the length of the cascade appears as a real-valued parameter and can thus be fitted in the same manner as Hill coefficients. Finally, we show how the obtained nonlinear modules can be used instead of delay differential equations to model delays in signal transduction. PMID:27581482

  18. Extracellular vesicles including exosomes are mediators of signal transduction: Are they protective or pathogenic?

    PubMed Central

    Gangoda, Lahiru; Boukouris, Stephanie; Liem, Michael; Kalra, Hina; Mathivanan, Suresh

    2015-01-01

    Extracellular vesicles are signaling organelles that are released by many cell types and is highly conserved in both prokaryotes and eukaryotes. Based on the mechanism of biogenesis, these membranous vesicles can be classified as exosomes, shedding microvesicles and apoptotic blebs. It is becoming clearer that these extracellular vesicles mediate signal transduction in both autocrine and paracrine fashion by the transfer of proteins and RNA. Whilst the role of extracellular vesicles including exosomes in pathogenesis is well established, very little is known about their function in normal physiological conditions. Recent evidences allude that extracellular vesicles can mediate both protective and pathogenic effects depending on the precise state. In this review, we discuss the involvement of extracellular vesicle as mediators of signal transduction in neurodegenerative diseases and cancer. In addition, the role of extracellular vesicles in mediating Wnt and PI3K signaling pathways is also discussed. Additional findings on the involvement of extracellular vesicles in homeostasis and disease progression will promote a better biological understanding, advance future therapeutic and diagnostic applications. PMID:25307053

  19. Identification of specific gravity sensitive signal transduction pathways in human A431 carcinoma cells

    NASA Astrophysics Data System (ADS)

    Rijken, P. J.; de Groot, R. P.; Kruijer, W.; de Laat, S. W.; Verkleij, A. J.; Boonstra, J.

    Epidermal growth factor (EGF) activates a well characterized signal transduction cascade in human A431 epidermoid carcinoma cells. The influence of gravity on EGF-induced EGF-receptor clustering and early gene expression as well as on actin polymerization and actin organization have been investigated. Different signalling pathways induced by the agents TPA, forskolin and A23187 that activate gene expression were tested for sensitivity to gravity. EGF-induced c-fos and c-jun expression were decreased in microgravity. However, constitutive β-2 microglobulin expression remained unaltered. Under simulated weightlessness conditions EGF- and TPA-induced c-fos expression was decreased, while forskolin- and A23187-induced c-fos expression was independent of the gravity conditions. These results suggest that gravity affects specific signalling pathways. Preliminary results indicate that EGF-induced EGF-receptor clustering remained unaltered irrespective of the gravity conditions. Furthermore, the relative filamentous actin content of steady state A431 cells was enhanced under microgravity conditions and actin filament organization was altered. Under simulated weightlessness actin filament organization in steady state cells as well as in EGF-treated cells was altered as compared to the 1 G reference experiment. Interestingly the microtubule and keratin organization in untreated cells showed no difference with the normal gravity samples. This indicates that gravity may affect specific components of the signal transduction circuitry.

  20. Rhizobium nod factor signaling. Evidence for a g protein-mediated transduction mechanism

    PubMed Central

    Pingret, JL; Journet, EP; Barker, DG

    1998-01-01

    Rhizobium nodulation (Nod) factors are lipochitooligosaccharide signals that elicit key symbiotic developmental responses in the host legume root. In this study, we have investigated Nod factor signal transduction in the Medicago root epidermis by using a pharmacological approach in conjunction with transgenic plants expressing the Nod factor-responsive reporter construct pMtENOD12-GUS. Evidence for the participation of heterotrimeric G proteins in Nod factor signaling has come from three complementary observations: (1) the amphiphilic peptides mastoparan and Mas7, known G protein agonists, are able to mimic Nod factor-induced epidermal MtENOD12 expression; (2) growth of plants in nodulation-inhibiting conditions (10 mM NH4NO3) leads to a dramatic reduction in both Nod factor- and mastoparan-elicited gene expression; and (3) bacterial pertussis toxin, a well-characterized G protein antagonist, blocks the activities of both the Nod factor and mastoparan. In addition, we have found that antagonists that interfere with phospholipase C activity (neomycin and U73122) and Ca2+ influx/release (EGTA, La3+, and ruthenium red) block Nod factor/mastoparan activity. Taken together, these results are consistent with a Nod factor signal transduction mechanism involving G protein mediation coupled to the activation of both phosphoinositide and Ca2+ second messenger pathways. PMID:9596628

  1. Reactive oxygen species and PI3K/Akt signaling in cancer.

    PubMed

    Jin, Seo Yeon; Lee, Hye Sun; Kim, Eun Kyoung; Ha, Jung Min; Kim, Young Whan; Bae, SunSik

    2014-10-01

    Reactive oxygen species (ROS) are chemically reactive molecules containing oxygen and associates with multiple cellular functions such as cell proliferation, differentiation, and apoptosis. In the present study, we showed that Insulin-like growth factor-1(IGF-1) modulates SKOV-3 ovarian cancer cell by regulation of generation of ROS. Akt mediates cellular signaling pathways in association with mammalian target of rapamycin complex (mTOR) and Rac small G protein. Insulin-like growth factor-1 (IGF-1)-induced generation of ROS was completely abolished by phosphatidylinositol 3-kinase (PI3K) (LY294002, 10?µM) or Akt inhibitors (SH-5, 50?µM), whereas inhibition of extracellular-regulated kinase by an ERK inhibitor (PD98059, 10?µM) or inhibition of mammalian target of rapamycin complex 1 (mTORC1) by an mTORC1 inhibitor (Rapamycin, 100?nM) did not affect IGF-1-induced generation of ROS. Inactivation of mTORC2 by silencing Rapamycin-insensitive companion of mTOR (Rictor), abolished IGF-1-induced SKOV-3 cell migration as well as activation of Akt. However, inactivation of mTORC1 by silencing of Raptor had no effect. Silencing of Akt1 but not Akt2 attenuated IGF-1-induced generation of ROS. Expression of PIP3-dependent Rac exchanger1 (P-Rex1), a Rac guanosine exchange factor and a component of the mTOR complex. Silencing of P-Rex1 abolished IGF-1-induced generation of ROS. Finally, inhibition of NADPH oxidase system completely blunted IGF-1-induced generation of ROS, whereas inhibition of xanthine oxiase,cyclooxygenase, and mitochondrial respiratory chain complex was not effective. Given these results, we suggest that IGF-1 induces ROS generation through the PI3K/Akt/ mTOR2/NADPH oxidase signaling axis. PMID:26461347

  2. Testosterone regulation of Akt/mTORC1/FoxO3a Signaling in Skeletal Muscle

    PubMed Central

    White, James P.; Gao, Song; Puppa, Melissa J.; Sato, Shuichi; Welle, Stephen L.; Carson, James A.

    2012-01-01

    Low endogenous testosterone production, known as hypogonadism is commonly associated with conditions inducing muscle wasting. Akt signaling can control skeletal muscle mass through mTOR regulation of protein synthesis and FoxO regulation of protein degradation, and this pathway has been previously identified as a target of androgen signaling. However, the testosterone sensitivity of Akt/mTOR signaling requires further understanding in order to grasp the significance of varied testosterone levels seen with wasting disease on muscle protein turnover regulation. Therefore, the purpose of this study is to determine the effect of androgen availability on muscle Akt/mTORC1/FoxO3a regulation in skeletal muscle and cultured C2C12 myotubes. C57BL/6 mice were either castrated for 42 days or castrated and treated with the nandrolone decanoate (ND) (6 mg/kg bw/wk). Testosterone loss (TL) significantly decreased volitional grip strength, body weight, and gastrocnemius (GAS) muscle mass, and ND reversed these changes. Related to muscle mass regulation, TL decreased muscle IGF-1 mRNA, the rate of myofibrillar protein synthesis, Akt phosphorylation, and the phosphorylation of Akt targets, GSK3β, PRAS40 and FoxO3a. TL induced expression of FoxO transcriptional targets, MuRF1, atrogin1 and REDD1. Muscle AMPK and raptor phosphorylation, mTOR inhibitors, were not altered by low testosterone. ND restored IGF-1 expression and Akt/mTORC1 signaling while repressing expression of FoxO transcriptional targets. Testosterone (T) sensitivity of Akt/mTORC1 signaling was examined in C2C12 myotubes, and mTOR phosphorylation was induced independent of Akt activation at low T concentrations, while a higher T concentration was required to activate Akt signaling. Interestingly, low concentration T was sufficient to amplify myotube mTOR and Akt signaling after 24h of T withdrawal, demonstrating the potential in cultured myotubes for a T initiated positive feedback mechanism to amplify Akt

  3. Testosterone regulation of Akt/mTORC1/FoxO3a signaling in skeletal muscle.

    PubMed

    White, James P; Gao, Song; Puppa, Melissa J; Sato, Shuichi; Welle, Stephen L; Carson, James A

    2013-01-30

    Low endogenous testosterone production, known as hypogonadism is commonly associated with conditions inducing muscle wasting. Akt signaling can control skeletal muscle mass through mTOR regulation of protein synthesis and FoxO regulation of protein degradation, and this pathway has been previously identified as a target of androgen signaling. However, the testosterone sensitivity of Akt/mTOR signaling requires further understanding in order to grasp the significance of varied testosterone levels seen with wasting disease on muscle protein turnover regulation. Therefore, the purpose of this study is to determine the effect of androgen availability on muscle Akt/mTORC1/FoxO3a regulation in skeletal muscle and cultured C(2)C(12) myotubes. C57BL/6 mice were either castrated for 42 days or castrated and treated with the nandrolone decanoate (ND) (6 mg/kg bw/wk). Testosterone loss (TL) significantly decreased volitional grip strength, body weight, and gastrocnemius (GAS) muscle mass, and ND reversed these changes. Related to muscle mass regulation, TL decreased muscle IGF-1 mRNA, the rate of myofibrillar protein synthesis, Akt phosphorylation, and the phosphorylation of Akt targets, GSK3β, PRAS40 and FoxO3a. TL induced expression of FoxO transcriptional targets, MuRF1, atrogin1 and REDD1. Muscle AMPK and raptor phosphorylation, mTOR inhibitors, were not altered by low testosterone. ND restored IGF-1 expression and Akt/mTORC1 signaling while repressing expression of FoxO transcriptional targets. Testosterone (T) sensitivity of Akt/mTORC1 signaling was examined in C(2)C(12) myotubes, and mTOR phosphorylation was induced independent of Akt activation at low T concentrations, while a higher T concentration was required to activate Akt signaling. Interestingly, low concentration T was sufficient to amplify myotube mTOR and Akt signaling after 24 h of T withdrawal, demonstrating the potential in cultured myotubes for a T initiated positive feedback mechanism to amplify Akt

  4. DNA Synthesis during Endomitosis Is Stimulated by Insulin via the PI3K/Akt and TOR Signaling Pathways in the Silk Gland Cells of Bombyx mori

    PubMed Central

    Li, Yaofeng; Chen, Xiangyun; Tang, Xiaofang; Zhang, Chundong; Wang, La; Chen, Peng; Pan, Minhui; Lu, Cheng

    2015-01-01

    Silk gland cells undergo multiple endomitotic cell cycles during silkworm larval ontogeny. Our previous study demonstrated that feeding is required for continued endomitosis in the silk gland cells of silkworm larvae. Furthermore, the insulin signaling pathway is closely related to nutritional signals. To investigate whether the insulin signaling pathway is involved in endomitosis in silk gland cells, in this study, we initially analyzed the effects of bovine insulin on DNA synthesis in endomitotic silk gland cells using 5-bromo-2'-deoxyuridine (BrdU) labeling technology, and found that bovine insulin can stimulate DNA synthesis. Insulin signal transduction is mainly mediated via phosphoinositide 3-kinase (PI3K)/Akt, the target of rapamycin (TOR) and the extracellular signal-regulated kinase (ERK) pathways in vertebrates. We ascertained that these three pathways are involved in DNA synthesis in endomitotic silk gland cells using specific inhibitors against each pathway. Moreover, we investigated whether these three pathways are involved in insulin-stimulated DNA synthesis in endomitotic silk gland cells, and found that the PI3K/Akt and TOR pathways, but not the ERK pathway, are involved in this process. These results provide an important theoretical foundation for the further investigations of the mechanism underlying efficient endomitosis in silk gland cells. PMID:25794286

  5. DNA synthesis during endomitosis is stimulated by insulin via the PI3K/Akt and TOR signaling pathways in the silk gland cells of Bombyx mori.

    PubMed

    Li, Yaofeng; Chen, Xiangyun; Tang, Xiaofang; Zhang, Chundong; Wang, La; Chen, Peng; Pan, Minhui; Lu, Cheng

    2015-01-01

    Silk gland cells undergo multiple endomitotic cell cycles during silkworm larval ontogeny. Our previous study demonstrated that feeding is required for continued endomitosis in the silk gland cells of silkworm larvae. Furthermore, the insulin signaling pathway is closely related to nutritional signals. To investigate whether the insulin signaling pathway is involved in endomitosis in silk gland cells, in this study, we initially analyzed the effects of bovine insulin on DNA synthesis in endomitotic silk gland cells using 5-bromo-2'-deoxyuridine (BrdU) labeling technology, and found that bovine insulin can stimulate DNA synthesis. Insulin signal transduction is mainly mediated via phosphoinositide 3-kinase (PI3K)/Akt, the target of rapamycin (TOR) and the extracellular signal-regulated kinase (ERK) pathways in vertebrates. We ascertained that these three pathways are involved in DNA synthesis in endomitotic silk gland cells using specific inhibitors against each pathway. Moreover, we investigated whether these three pathways are involved in insulin-stimulated DNA synthesis in endomitotic silk gland cells, and found that the PI3K/Akt and TOR pathways, but not the ERK pathway, are involved in this process. These results provide an important theoretical foundation for the further investigations of the mechanism underlying efficient endomitosis in silk gland cells. PMID:25794286

  6. Conformational transition in signal transduction: metastable states and transition pathways in the activation of a signaling protein.

    PubMed

    Banerjee, Rahul; Yan, Honggao; Cukier, Robert I

    2015-06-01

    Signal transduction is of vital importance to the growth and adaptation of living organisms. The key to understand mechanisms of biological signal transduction is elucidation of the conformational dynamics of its signaling proteins, as the activation of a signaling protein is fundamentally a process of conformational transition from an inactive to an active state. A predominant form of signal transduction for bacterial sensing of environmental changes in the wild or inside their hosts is a variety of two-component systems, in which the conformational transition of a response regulator (RR) from an inactive to an active state initiates responses to the environmental changes. Here, RR activation has been investigated using RR468 as a model system by extensive unbiased all-atom molecular dynamics (MD) simulations in explicit solvent, starting from snapshots along a targeted MD trajectory that covers the conformational transition. Markov state modeling, transition path theory, and geometric analyses of the wealth of the MD data have provided a comprehensive description of the RR activation. It involves a network of metastable states, with one metastable state essentially the same as the inactive state and another very similar to the active state that are connected via a small set of intermediates. Five major pathways account for >75% of the fluxes of the conformational transition from the inactive to the active-like state. The thermodynamic stability of the states and the activation barriers between states are found, to identify rate-limiting steps. The conformal transition is initiated predominantly by movements of the β3α3 loop, followed by movements of the β4α4-loop and neighboring α4 helix region, and capped by additional movements of the β3α3 loop. A number of transient hydrophobic and hydrogen bond interactions are revealed, and they may be important for the conformational transition. PMID:25945797

  7. HAMP domain-mediated signal transduction probed with a mycobacterial adenylyl cyclase as a reporter.

    PubMed

    Mondéjar, Laura García; Lupas, Andrei; Schultz, Anita; Schultz, Joachim E

    2012-01-01

    HAMP domains, ∼55 amino acid motifs first identified in histidine kinases, adenylyl cyclases, methyl-accepting chemotaxis proteins, and phosphatases, operate as signal mediators in two-component signal transduction proteins. A bioinformatics study identified a coevolving signal-accepting network of 10 amino acids in membrane-delimited HAMP proteins. To probe the functionality of this network we used a HAMP containing mycobacterial adenylyl cyclase, Rv3645, as a reporter enzyme in which the membrane anchor was substituted by the Escherichia coli chemotaxis receptor for serine (Tsr receptor) and the HAMP domain alternately with that from the protein Af1503 of the archaeon Archaeoglobus fulgidus or the Tsr receptor. In a construct with the Tsr-HAMP, cyclase activity was inhibited by serine, whereas in a construct with the HAMP domain from A. fulgidus, enzyme activity was not responsive to serine. Amino acids of the signal-accepting network were mutually swapped between both HAMP domains, and serine signaling was examined. The data biochemically tentatively established the functionality of the signal-accepting network. Based on a two-state gearbox model of rotation in HAMP domain-mediated signal propagation, we characterized the interaction between permanent and transient core residues in a coiled coil HAMP structure. The data are compatible with HAMP rotation in signal propagation but do not exclude alternative models for HAMP signaling. Finally, we present data indicating that the connector, which links the α-helices of HAMP domains, plays an important structural role in HAMP function. PMID:22094466

  8. Expression and functional analyses of the Arabidopsis QUA1 gene in light signal transduction.

    PubMed

    Zhaojin, Chen; Chuanyu, Ding; Yuan, Zheng

    2016-05-01

    Plants not only use light as an energy source for photosynthesis, but also have to monitor the light quality and quantity input to execute appropriate physiological and developmental responses, such as cell differentiation, structural and functional changes, as well as the formation of tissues and organs. The process is referred to as photomorphogenesis. Arabidopsis QUA1 (QUASIMODO1), which functions in pectin synthesis, is identified as a member of glycosyltransferases. Previously, the hypocotyl elongation of the qua1-1 mutant was shown to be inhibited under dark conditions. In this study, we used the qua1-1/cry1 and qua1-1/phyB double mutants as the materials to study the function of the QUA1 gene in light signal transduction. The results showed that QUA1 not only participated in hypocotyl elongation under dark conditions, but also in blue light, red light and far red light conditions. In qua1-1 mutant seedlings, both the cell length of hypocotyl and the light-regulated gene expression were affected. Compared with cry1 and phyB mutants, qua1-1/cry1 and qua1-1/phyB double mutants had the shorter hypocotyl. Light-regulated gene expression was also affected in the double mutants. These data indicated that QUA1 might participate in the light signal transduction regulated by CRY1 and PHYB. Hence, the QUA1 gene may play multiple roles in light signal transduction by regulating the cell elongation and light-regulated gene expression. PMID:27232492

  9. Plant gravitropic signal transduction: A network analysis leads to gene discovery

    NASA Astrophysics Data System (ADS)

    Wyatt, Sarah

    Gravity plays a fundamental role in plant growth and development. Although a significant body of research has helped define the events of gravity perception, the role of the plant growth regulator auxin, and the mechanisms resulting in the gravity response, the events of signal transduction, those that link the biophysical action of perception to a biochemical signal that results in auxin redistribution, those that regulate the gravitropic effects on plant growth, remain, for the most part, a “black box.” Using a cold affect, dubbed the gravity persistent signal (GPS) response, we developed a mutant screen to specifically identify components of the signal transduction pathway. Cloning of the GPS genes have identified new proteins involved in gravitropic signaling. We have further exploited the GPS response using a multi-faceted approach including gene expression microarrays, proteomics analysis, and bioinformatics analysis and continued mutant analysis to identified additional genes, physiological and biochemical processes. Gene expression data provided the foundation of a regulatory network for gravitropic signaling. Based on these gene expression data and related data sets/information from the literature/repositories, we constructed a gravitropic signaling network for Arabidopsis inflorescence stems. To generate the network, both a dynamic Bayesian network approach and a time-lagged correlation coefficient approach were used. The dynamic Bayesian network added existing information of protein-protein interaction while the time-lagged correlation coefficient allowed incorporation of temporal regulation and thus could incorporate the time-course metric from the data set. Thus the methods complemented each other and provided us with a more comprehensive evaluation of connections. Each method generated a list of possible interactions associated with a statistical significance value. The two networks were then overlaid to generate a more rigorous, intersected

  10. A model for signal transduction during quorum sensing in Vibrio harveyi

    NASA Astrophysics Data System (ADS)

    Banik, Suman K.; Fenley, Andrew T.; Kulkarni, Rahul V.

    2009-12-01

    We present a framework for analyzing luminescence regulation during quorum sensing in the bioluminescent bacterium Vibrio harveyi. Using a simplified model for signal transduction in the quorum sensing pathway, we identify key dimensionless parameters that control the system's response. These parameters are estimated using experimental data on luminescence phenotypes for different mutant strains. The corresponding model predictions are consistent with results from other experiments which did not serve as input for determining model parameters. Furthermore, the proposed framework leads to novel testable predictions for luminescence phenotypes and for responses of the network to different perturbations.

  11. Cellerator: extending a computer algebra system to include biochemical arrows for signal transduction simulations

    NASA Technical Reports Server (NTRS)

    Shapiro, Bruce E.; Levchenko, Andre; Meyerowitz, Elliot M.; Wold, Barbara J.; Mjolsness, Eric D.

    2003-01-01

    Cellerator describes single and multi-cellular signal transduction networks (STN) with a compact, optionally palette-driven, arrow-based notation to represent biochemical reactions and transcriptional activation. Multi-compartment systems are represented as graphs with STNs embedded in each node. Interactions include mass-action, enzymatic, allosteric and connectionist models. Reactions are translated into differential equations and can be solved numerically to generate predictive time courses or output as systems of equations that can be read by other programs. Cellerator simulations are fully extensible and portable to any operating system that supports Mathematica, and can be indefinitely nested within larger data structures to produce highly scaleable models.

  12. Structure-function relationships in the IL-17 receptor: Implications for signal transduction and therapy

    PubMed Central

    Shen, Fang; Gaffen, Sarah L.

    2008-01-01

    IL-17 is the defining cytokine of a newly-described “Th17” population that plays critical roles in mediating inflammation and autoimmunity. The IL-17/IL-17 receptor superfamily is the most recent class of cytokines and receptors to be described, and until recently very little was known about its function or molecular biology. However, in the last year important new insights into the composition and dynamics of the receptor complex and mechanisms of downstream signal transduction have been made, which will be reviewed here. PMID:18178098

  13. Nuclear Akt2 opposes limbal keratinocyte stem cell self-renewal by repressing a FOXO-mTORC1 signaling pathway.

    PubMed

    Saoncella, Stefania; Tassone, Beatrice; Deklic, Erika; Avolio, Fabio; Jon, Cristina; Tornillo, Giusy; De Luca, Elisa; Di Iorio, Enzo; Piva, Roberto; Cabodi, Sara; Turco, Emilia; Pandolfi, Pier Paolo; Calautti, Enzo

    2014-03-01

    Signals downstream of Akt can either favor or oppose stem cell (SC) maintenance, but how this dual role can be achieved is still undefined. Using human limbal keratinocyte stem cells (LKSCs), a SC type used in transplantation therapies for corneal regeneration, we show that Akt signaling is prominent in SC populations both in vivo and in vitro, and that Akt1 promotes while Akt2 opposes SC self-renewal. Noteworthy, loss of Akt2 signaling enhances LKSC maintenance ex vivo, whereas Akt1 depletion anticipates SC exhaustion. Mechanistically, the antagonistic functions of Akt1 and Akt2 in SC control are mainly dictated by their differential subcellular distribution, being nuclear Akt2 selectively implicated in FOXO inhibition. Akt2 downregulation favors LKSC maintenance as a result of a gain of FOXO functions, which attenuates the mechanistic target of rapamycin complex one signaling via tuberous sclerosis one gene induction, and promotes growth factor signaling through Akt1. Consistently, Akt2 deficiency also enhances limbal SCs in vivo. Thus, our findings reveal distinct roles for nuclear versus cytosolic Akt signaling in normal epithelial SC control and suggest that the selective Akt2 inhibition may provide novel pharmacological strategies for human LKSC expansion in therapeutic settings and mechanistic research. PMID:24123662

  14. Evaluation of signal transduction pathways after transient cutaneous adenoviral gene delivery

    PubMed Central

    2011-01-01

    Background Adenoviral vectors have provided effective methods for in vivo gene delivery in therapeutic applications. However, these vectors can induce immune responses that may severely affect the ability of vector re-application. There is limited information about the mechanisms and signal transduction pathways involved in adenoviral recognition. For optimization of cutaneous gene therapy it is necessary to investigate molecular mechanisms of virus recognition in epidermal cells. The aim of this study was to investigate the signal transduction of the innate immunity after adenoviral DNA internalization in keratinocytes. Methods In vitro, keratinocytes were transfected with DNA, in the presence and absence of inhibitors for signalling molecules. In vivo, immunocompetent and athymic mice (n = 3 per group) were twice transduced with an Ad-vector. Results The results show an acute induction of type-I-interferon after in vitro transfection. Inhibition of PI3K, p38 MAPK, JNK and NFkappaB resulted in a decreased expression of type-I-interferon. In contrast to immunocompetent mice, athymic mice demonstrated a constant transgene expression and reduced inflammatory response in vivo. Conclusion The results suggest an induction of the innate immunity triggered by cytoplasm localised DNA which is mediated by PI3K-, p38 MAPK-, JNK-, NFkappaB-, JAK/STAT- and ERK1/2-dependent pathways. A stable transgene expression and a reduced inflammatory response in immunodeficient mice have been observed. These results provide potential for an effective adenoviral gene delivery into immunosupressed skin. PMID:21255430

  15. Transmembrane signal transduction in bacterial chemotaxis involves ligand-dependent activation of phosphate group transfer.

    PubMed Central

    Borkovich, K A; Kaplan, N; Hess, J F; Simon, M I

    1989-01-01

    Signal transduction in Escherichia coli involves the interaction of transmembrane receptor proteins such as the aspartate receptor, Tar, and the products of four chemotaxis genes, cheA, cheY, cheW, and cheZ. It was previously shown that the cheA gene product is an autophosphorylating protein kinase that transfers phosphate to CheY, whereas the cheZ gene product acts as a specific CheY phosphatase. Here we report that the system can be reconstituted in vitro and receptor function can be coupled to CheY phosphorylation. Coupling requires the presence of the CheW protein, the appropriate form of the receptor, and the CheA and CheY proteins. Under these conditions the accumulation of CheY-phosphate is enhanced approximately 300-fold. This rate enhancement is seen in reactions using wild-type and "tumble" mutant receptors but not "smooth" mutant receptors. The increased accumulation of phosphoprotein was inhibited by micromolar concentrations of aspartate, using wild-type, but not tumble, receptors. These results provide evidence that the signal transduction pathway in bacterial chemotaxis involves receptor-mediated alteration of the levels of phosphorylated proteins. They suggest that CheW acts as the coupling factor between receptor and phosphorylation. The results also support the suggestion that CheY-phosphate is the tumble signal. Images PMID:2645576

  16. Signal Transduction: From the Atomic Age to the Post-Genomic Era

    PubMed Central

    Thorner, Jeremy; Hunter, Tony; Cantley, Lewis C.; Sever, Richard

    2016-01-01

    SUMMARY We have come a long way in the 55 years since Edmond Fischer and the late Edwin Krebs discovered that the activity of glycogen phosphorylase is regulated by reversible protein phosphorylation. Many of the fundamental molecular mechanisms that operate in biological signaling have since been characterized and the vast web of interconnected pathways that make up the cellular signaling network has been mapped in considerable detail. Nonetheless, it is important to consider how fast this field is still moving and the issues at the current boundaries of our understanding. One must also appreciate what experimental strategies have allowed us to attain our present level of knowledge. We summarize here some key issues (both conceptual and methodological), raise unresolved questions, discuss potential pitfalls, and highlight areas in which our understanding is still rudimentary. We hope these wide-ranging ruminations will be useful to investigators who carry studies of signal transduction forward during the rest of the 21st century. PMID:25359498

  17. Endocytosis of pro-inflammatory cytokine receptors and its relevance for signal transduction.

    PubMed

    Hermanns, Heike M; Wohlfahrt, Julia; Mais, Christine; Hergovits, Sabine; Jahn, Daniel; Geier, Andreas

    2016-08-01

    The pro-inflammatory cytokines tumor necrosis factor (TNF), interleukin-1 (IL-1) and interleukin-6 (IL-6) are key players of the innate and adaptive immunity. Their activity needs to be tightly controlled to allow the initiation of an appropriate immune response as defense mechanism against pathogens or tissue injury. Excessive or sustained signaling of either of these cytokines leads to severe diseases, including rheumatoid arthritis, inflammatory bowel diseases (Crohn's disease, ulcerative colitis), steatohepatitis, periodic fevers and even cancer. Studies carried out in the last 30 years have emphasized that an elaborate control system for each of these cytokines exists. Here, we summarize what is currently known about the involvement of receptor endocytosis in the regulation of these pro-inflammatory cytokines' signaling cascades. Particularly in the last few years it was shown that this cellular process is far more than a mere feedback mechanism to clear cytokines from the circulation and to shut off their signal transduction. PMID:27071147

  18. G-Protein-Coupled Lysophosphatidic Acid Receptors and Their Regulation of AKT Signaling

    PubMed Central

    Riaz, Anjum; Huang, Ying; Johansson, Staffan

    2016-01-01

    A hallmark of G-protein-coupled receptors (GPCRs) is their ability to recognize and respond to chemically diverse ligands. Lysophospholipids constitute a relatively recent addition to these ligands and carry out their biological functions by activating G-proteins coupled to a large family of cell-surface receptors. This review aims to highlight salient features of cell signaling by one class of these receptors, known as lysophosphatidic acid (LPA) receptors, in the context of phosphatidylinositol 3-kinase (PI3K)–AKT pathway activation. LPA moieties efficiently activate AKT phosphorylation and activation in a multitude of cell types. The interplay between LPA, its receptors, the associated Gαi/o subunits, PI3K and AKT contributes to the regulation of cell survival, migration, proliferation and confers chemotherapy-resistance in certain cancers. However, detailed information on the regulation of PI3K–AKT signals induced by LPA receptors is missing from the literature. Here, some urgent issues for investigation are highlighted. PMID:26861299

  19. G-Protein-Coupled Lysophosphatidic Acid Receptors and Their Regulation of AKT Signaling.

    PubMed

    Riaz, Anjum; Huang, Ying; Johansson, Staffan

    2016-01-01

    A hallmark of G-protein-coupled receptors (GPCRs) is their ability to recognize and respond to chemically diverse ligands. Lysophospholipids constitute a relatively recent addition to these ligands and carry out their biological functions by activating G-proteins coupled to a large family of cell-surface receptors. This review aims to highlight salient features of cell signaling by one class of these receptors, known as lysophosphatidic acid (LPA) receptors, in the context of phosphatidylinositol 3-kinase (PI3K)-AKT pathway activation. LPA moieties efficiently activate AKT phosphorylation and activation in a multitude of cell types. The interplay between LPA, its receptors, the associated Gαi/o subunits, PI3K and AKT contributes to the regulation of cell survival, migration, proliferation and confers chemotherapy-resistance in certain cancers. However, detailed information on the regulation of PI3K-AKT signals induced by LPA receptors is missing from the literature. Here, some urgent issues for investigation are highlighted. PMID:26861299

  20. Activation of Akt Signaling in Prostate Induces a TGFβ Mediated Restraint on Cancer Progression and Metastasis

    PubMed Central

    Bjerke, Glen A.; Yang, Chun-Song; Frierson, Henry F.; Paschal, Bryce M.; Wotton, David

    2014-01-01

    Mutations in the PTEN tumor suppressor gene are found in a high proportion of human prostate cancers, and in mice, Pten deletion induces high-grade prostate intra-epithelial neoplasia (HGPIN). However, progression from HGPIN to invasive cancer occurs slowly, suggesting that tumorigenesis is subject to restraint. We show that Pten deletion, or constitutive activation of the downstream kinase AKT, activates the transforming growth factor (TGF) β pathway in prostate epithelial cells. TGFβ signaling is known to play a tumor suppressive role in many cancer types, and reduced expression of TGFβ receptors correlates with advanced human prostate cancer. We demonstrate that in combination either with loss of Pten, or expression of constitutively active AKT1, inactivation of TGFβ signaling by deletion of the TGFβ type II receptor gene relieves a restraint on tumorigenesis. This results in rapid progession to lethal prostate cancer, including metastasis to lymph node and lung. In prostate epithelium, inactivation of TGFβ signaling alone is insufficient to initiate tumorigenesis, but greatly accelerates cancer progression. The activation of TGFβ signaling by Pten loss or AKT activation suggests that the same signaling events that play key roles in tumor initiation also induce the activity of a pathway that restrains disease progression. PMID:23995785

  1. mTORC2-Akt signaling axis is implicated in myocardial compensation and fibrosis.

    PubMed

    Dany, M; Rimmani, H H; Matar, S A; Hajj Hussein, I

    2015-01-01

    mTOR signaling has long been implicated in the mechanisms of cardiomyocyte survival in response to volume or pressure overload. Several studies have focused on the significance of mTORC1 in cardiomyocyte survival, questioning the role of mTORC2. mTORC2–Akt signaling is an emerging axis implicated in cardiomyocyte compensation and thus heart failure. Upon being subjected to chronic stress, cardiomyocytes activate mTORC2–Akt signaling pathway to promote survival by activating the ubiquitin proteasome system, inducing the degradation of pro-apoptotic proteins, and altering the actin cytoskeleton. Given the importance of mTORC2 in cardiomyocyte survival, studies suggest that loss of mTORC2 signaling would result in loss of cardiomyocytes, fibrosis, and heart failure. This review serves to elaborate on how mTORC2-Akt signaling plays a role in cardiomyocyte growth and survival under stress and how the loss of the axis would result in fibrosis and heart failure. PMID:26753634

  2. PI3K/AKT Signaling Pathway Is Essential for Survival of Induced Pluripotent Stem Cells

    PubMed Central

    Hossini, Amir M.; Quast, Annika S.; Plötz, Michael; Grauel, Katharina; Exner, Tarik; Küchler, Judit; Stachelscheid, Harald; Eberle, Jürgen; Rabien, Anja

    2016-01-01

    Apoptosis is a highly conserved biochemical mechanism which is tightly controlled in cells. It contributes to maintenance of tissue homeostasis and normally eliminates highly proliferative cells with malignant properties. Induced pluripotent stem cells (iPSCs) have recently been described with significant functional and morphological similarities to embryonic stem cells. Human iPSCs are of great hope for regenerative medicine due to their broad potential to differentiate into specialized cell types in culture. They may be useful for exploring disease mechanisms and may provide the basis for future cell-based replacement therapies. However, there is only poor insight into iPSCs cell signaling as the regulation of apoptosis. In this study, we focused our attention on the apoptotic response of Alzheimer fibroblast-derived iPSCs and two other Alzheimer free iPSCs to five biologically relevant kinase inhibitors as well as to the death ligand TRAIL. To our knowledge, we are the first to report that the relatively high basal apoptotic rate of iPSCs is strongly suppressed by the pancaspase inhibitor QVD-Oph, thus underlining the dependency on proapoptotic caspase cascades. Furthermore, wortmannin, an inhibitor of phosphoinositid-3 kinase / Akt signaling (PI3K-AKT), dramatically and rapidly induced apoptosis in iPSCs. In contrast, parental fibroblasts as well as iPSC-derived neuronal cells were not responsive. The resulting condensation and fragmentation of DNA and decrease of the membrane potential are typical features of apoptosis. Comparable effects were observed with an AKT inhibitor (MK-2206). Wortmannin resulted in disappearance of phosphorylated AKT and activation of the main effector caspase-3 in iPSCs. These results clearly demonstrate for the first time that PI3K-AKT represents a highly essential survival signaling pathway in iPSCs. The findings provide improved understanding on the underlying mechanisms of apoptosis regulation in iPSCs. PMID:27138223

  3. PI3K/AKT Signaling Pathway Is Essential for Survival of Induced Pluripotent Stem Cells.

    PubMed

    Hossini, Amir M; Quast, Annika S; Plötz, Michael; Grauel, Katharina; Exner, Tarik; Küchler, Judit; Stachelscheid, Harald; Eberle, Jürgen; Rabien, Anja; Makrantonaki, Evgenia; Zouboulis, Christos C

    2016-01-01

    Apoptosis is a highly conserved biochemical mechanism which is tightly controlled in cells. It contributes to maintenance of tissue homeostasis and normally eliminates highly proliferative cells with malignant properties. Induced pluripotent stem cells (iPSCs) have recently been described with significant functional and morphological similarities to embryonic stem cells. Human iPSCs are of great hope for regenerative medicine due to their broad potential to differentiate into specialized cell types in culture. They may be useful for exploring disease mechanisms and may provide the basis for future cell-based replacement therapies. However, there is only poor insight into iPSCs cell signaling as the regulation of apoptosis. In this study, we focused our attention on the apoptotic response of Alzheimer fibroblast-derived iPSCs and two other Alzheimer free iPSCs to five biologically relevant kinase inhibitors as well as to the death ligand TRAIL. To our knowledge, we are the first to report that the relatively high basal apoptotic rate of iPSCs is strongly suppressed by the pancaspase inhibitor QVD-Oph, thus underlining the dependency on proapoptotic caspase cascades. Furthermore, wortmannin, an inhibitor of phosphoinositid-3 kinase / Akt signaling (PI3K-AKT), dramatically and rapidly induced apoptosis in iPSCs. In contrast, parental fibroblasts as well as iPSC-derived neuronal cells were not responsive. The resulting condensation and fragmentation of DNA and decrease of the membrane potential are typical features of apoptosis. Comparable effects were observed with an AKT inhibitor (MK-2206). Wortmannin resulted in disappearance of phosphorylated AKT and activation of the main effector caspase-3 in iPSCs. These results clearly demonstrate for the first time that PI3K-AKT represents a highly essential survival signaling pathway in iPSCs. The findings provide improved understanding on the underlying mechanisms of apoptosis regulation in iPSCs. PMID:27138223

  4. Influence of zinc deficiency on AKT-MDM2-P53 signaling axes in normal and malignant human prostate cells

    Technology Transfer Automated Retrieval System (TEKTRAN)

    With prostate being the highest zinc-accumulating tissue before the onset of cancer, the effects of physiologic levels of zinc on Akt-Mdm2-p53 and Akt-p21 signaling axes in human normal prostate epithelial cells (PrEC) and malignant prostate LNCaP cells were examined. Cells were cultured for 6 d in...

  5. Molecular characterization of the Akt-TOR signaling pathway in rainbow trout: potential role in muscle growth/degradation

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The Akt-TOR signaling pathway plays a key role in cellular metabolism and muscle growth. Hormone, nutrition and stress factors affect the Akt-TOR pathway by regulating gene transcription, protein synthesis and degradation. In addition, we previously showed that energetic demands elevate during vit...

  6. Diversity training for signal transduction: leveraging cell-to-cell variability to dissect cellular signaling, differentiation and death

    PubMed Central

    Cotari, Jesse W.; Voisinne, Guillaume; Altan-Bonnet, Grégoire

    2013-01-01

    Populations of “identical” cells are rarely truly identical. Even when in the same state of differentiation, isogenic cells may vary in expression of key signaling regulators, activate signal transduction at different thresholds, and consequently respond heterogeneously to a given stimulus. Here, we review how new experimental and analytical techniques are suited to connect these different levels of variability, quantitatively mapping the effects of cell-to-cell variability on cellular decision-making. In particular, we summarize how this helps classify signaling regulators according to the impact of their variability on biological functions. We further discuss how variability can also be leveraged to shed light on the molecular mechanisms regulating cellular signaling, from the individual cell to the population of cells as a whole. PMID:23747193

  7. Crosstalking between Androgen and PI3K/AKT Signaling Pathways in Prostate Cancer Cells*

    PubMed Central

    Lee, Suk Hyung; Johnson, Daniel; Luong, Richard; Sun, Zijie

    2015-01-01

    Both androgen action and PI3K medicated signaling pathways have been implicated in prostate tumorigenesis. Our androgen receptor (AR) conditional transgenic mice developed murine prostatic intraepithelial neoplasia (mPIN) and prostatic adenocarcinoma lesions recapitulating human prostate cancer development and progression. Role of transgenic AR contributing to malignancy was demonstrated by high degree of transgenic AR expression in atypical and tumor cells in mPIN as well as prostatic adenocarcinoma lesions of the transgenic mice, but not in adjacent normal tissue. Interestingly, reduced PI3K/Akt activation also appeared in these mouse atypical and tumor cells, suggesting an interaction between androgen and PI3K/AKT pathways. In this study, we further investigated this interaction. We showed that the androgen depletion or knockdown of AR expression results in elevated levels of active phosphorylated AKT in prostate cancer cells. Castration of conditional Pten knock-out mice showed increased Akt, phosphorylated Akt, and pS6 expression in the mouse prostate. Using a series of newly generated Ar reporter and Pten knock-out compound mice, we showed that Pten loss directly represses endogenous Ar expression in prostatic epithelial cells. Moreover, Pten loss and PI3K/Akt activation reduced Ar-mediated transcription in purified Pten-null cells. This study provides novel evidence demonstrating interplay between androgen and PI3K pathways, as well as introduces unique and relevant mouse models for further studies of PI3K and AR pathways in the context of prostate tumorigenesis. PMID:25527506

  8. Polydatin improves glucose and lipid metabolism in experimental diabetes through activating the Akt signaling pathway.

    PubMed

    Hao, Jie; Chen, Cheng; Huang, Kaipeng; Huang, Junying; Li, Jie; Liu, Peiqing; Huang, Heqing

    2014-12-15

    Recently, the effect of polydatin on lipid regulation has gained considerable attention. And previous study has demonstrated that polydatin has hypoglycemic effect on experimental diabetic rats. Repressed Akt pathway contributes to glucose and lipid disorders in diabetes. Thus, whether polydatin regulates glucose and lipid metabolism in experimental diabetic models through the Akt pathway arouses interest. The purpose was to explore the regulatory mechanism of polydain on glucose and lipid through Akt pathway. We used a diabetic rat model induced by high-fat and -sugar diet with low-dose of streptozocin and an insulin resistant HepG2 cell model induced by palmitic acid to clarify the role of polydatin on glucose and lipid metabolism. Here, we found that polydatin significantly attenuated fasting blood–glucose, glycosylated hemoglobin, glycosylated serum protein, total cholesterol, triglyceride, and low-density lipoprotein cholesterol in diabetic rats. Furthermore, polydatin significantly increased glucose uptake and consumption and decreased lipid accumulation in insulin resistant HepG2 cells. Polydatin markedly increased serum insulin levels in diabetic rats, and obviously activated the Akt signaling pathway in diabetic rat livers and insulin resistant HepG2 cells. Polydatin markedly increased phosphorylated GSK-3β, decreased the protein levels of G6Pase and SREBP-1c, and increased protein levels of GCK, LDLR, and phosphorylated IRS in livers and HepG2 cells. Overall, the results indicate that polydatin regulates glucose and lipid metabolism in experimental diabetic models, the underlying mechanism is probably associated with regulating the Akt pathway. The effect of polydatin on increased Akt phosphorylation is independent of prompting insulin secretion, but dependent of increasing IRS phosphorylation. PMID:25310908

  9. Phosphatidylinositol 3-kinase/Akt signaling enhances nuclear localization and transcriptional activity of BRCA1

    SciTech Connect

    Hinton, Cimona V.; Fitzgerald, Latricia D.; Thompson, Marilyn E. . E-mail: methompson@mmc.edu

    2007-05-15

    Signaling pathways involved in regulating nuclear-cytoplasmic distribution of BRCA1 have not been previously reported. Here, we provide evidence that heregulin {beta}1-induced activation of the Akt pathway increases the nuclear content of BRCA1. First, treatment of T47D breast cancer cells with heregulin {beta}1 results in a two-fold increase in nuclear BRCA1 as assessed by FACS analysis, immunoblotting and immunofluorescence. This heregulin-induced increase in nuclear BRCA1 is blocked by siRNA-mediated down-regulation of Akt. Second, mutation of threonine 509 in BRCA1, the site of Akt phosphorylation, to an alanine, attenuates the ability of heregulin to induce BRCA1 nuclear accumulation. These data suggest that Akt-catalyzed phosphorylation of BRCA1 is required for the heregulin-regulated nuclear concentration of BRCA1. Because most functions ascribed to BRCA1 occur within the nucleus, we postulated that phosphorylation-dependent nuclear accumulation of BRCA1 would result in enhanced nuclear activity, specifically transcriptional activity, of BRCA1. This postulate is affirmed by our observation that the ability of BRCA1 to transactivate GADD45 promoter constructs was enhanced in T47D cells treated with heregulin {beta}1. Furthermore, the heterologous expression of BRCA1 in HCC1937 human breast cancer cells, which have constitutively active Akt, also induces GADD45 promoter activity, whereas the expression of BRCA1 in which threonine 509 has been mutated to an alanine is able to only minimally induce promoter activity. These findings implicate Akt in upstream events leading to BRCA1 nuclear localization and function.

  10. Phosphatidylinositol 3-kinase/Akt signaling enhances nuclear localization and transcriptional activity of BRCA1

    PubMed Central

    Hinton, Cimona V.; Fitzgerald, Latricia D.; Thompson, Marilyn E.

    2007-01-01

    Signaling pathways involved in regulating nuclear-cytoplasmic distribution of BRCA1 have not been previously reported. Here, we provide evidence that heregulin β1-induced activation of the Akt pathway increases the nuclear content of BRCA1. First, treatment of T47D breast cancer cells with heregulin β1 results in a two-fold increase in nuclear BRCA1 as assessed by FACS analysis, immunoblotting and immunofluorescence. This heregulin-induced increase in nuclear BRCA1 is blocked by siRNA-mediated down-regulation of Akt. Second, mutation of threonine 509 in BRCA1, the site of Akt phosphorylation, to an alanine, attenuates the ability of heregulin to induce BRCA1 nuclear accumulation. These data suggest that Akt-catalyzed phosphorylation of BRCA1 is required for the heregulin-regulated nuclear concentration of BRCA1. Because most functions ascribed to BRCA1 occur within the nucleus, we postulated that phosphorylation-dependent nuclear accumulation of BRCA1 would result in enhanced nuclear activity, specifically transcriptional activity, of BRCA1. This postulate is affirmed by our observation that the ability of BRCA1 to transactivate GADD45 promoter constructs was enhanced in T47D cells treated with heregulin β1. Furthermore, the heterologous expression of BRCA1 in HCC1937 human breast cancer cells, which have constitutively active Akt, also induces GADD45 promoter activity, whereas the expression of BRCA1 in which threonine 509 has been mutated to an alanine is able to only minimally induce promoter activity. These findings implicate Akt in upstream events leading to BRCA1 nuclear localization and function. PMID:17428466

  11. Cadmium induces carcinogenesis in BEAS-2B cells through ROS-dependent activation of PI3K/AKT/GSK-3β/β-catenin signaling

    SciTech Connect

    Son, Young-Ok; Wang, Lei; Poyil, Pratheeshkumar; Budhraja, Amit; Hitron, J. Andrew; Zhang, Zhuo; Lee, Jeong-Chae; Shi, Xianglin

    2012-10-15

    Cadmium has been widely used in industry and is known to be carcinogenic to humans. Although it is widely accepted that chronic exposure to cadmium increases the incidence of cancer, the mechanisms underlying cadmium-induced carcinogenesis are unclear. The main aim of this study was to investigate the role of reactive oxygen species (ROS) in cadmium-induced carcinogenesis and the signal transduction pathways involved. Chronic exposure of human bronchial epithelial BEAS-2B cells to cadmium induced cell transformation, as evidenced by anchorage-independent growth in soft agar and clonogenic assays. Chronic cadmium treatment also increased the potential of these cells to invade and migrate. Injection of cadmium-stimulated cells into nude mice resulted in the formation of tumors. In contrast, the cadmium-mediated increases in colony formation, cell invasion and migration were prevented by transfection with catalase, superoxide dismutase-1 (SOD1), or SOD2. In particular, chronic cadmium exposure led to activation of signaling cascades involving PI3K, AKT, GSK-3β, and β-catenin and transfection with each of the above antioxidant enzymes markedly inhibited cadmium-mediated activation of these signaling proteins. Inhibitors specific for AKT or β-catenin almost completely suppressed the cadmium-mediated increase in total and active β-catenin proteins and colony formation. Moreover, there was a marked induction of AKT, GSK-3β, β-catenin, and carcinogenic markers in tumor tissues formed in mice after injection with cadmium-stimulated cells. Collectively, our findings suggest a direct involvement of ROS in cadmium-induced carcinogenesis and implicate a role of AKT/GSK-3β/β-catenin signaling in this process. -- Highlights: ► Chronic exposure to cadmium induces carcinogenic properties in BEAS-2B cells. ► ROS involved in cadmium-induced tumorigenicity of BEAS-2B cells. ► Cadmium activates ROS-dependent AKT/GSK-3β/β-catenin-mediated signaling. ► ROS

  12. New therapeutic strategies targeting transmembrane signal transduction in the immune system

    PubMed Central

    2010-01-01

    Single-chain receptors and multi-chain immune recognition receptors (SRs and MIRRs, respectively) represent families of structurally related but functionally different surface receptors expressed on different cells. In contrast to SRs, a distinctive and common structural characteristic of MIRR family members is that the extracellular recognition domains and intracellular signaling domains are located on separate subunits. How extracellular ligand binding triggers MIRRs and initiates intracellular signal transduction processes is not clear. A novel model of immune signaling, the Signaling Chain HOmoOLigomerization (SCHOOL) model, suggests that the homooligomerization of receptor intracellular signaling domains represents a necessary and sufficient condition for receptor triggering. In this review, I demonstrate striking similarities between a consensus model of SR signaling and the SCHOOL model of MIRR signaling and show how these models, together with the lessons learned from viral pathogenesis, provide a molecular basis for novel pharmacological approaches targeting inter- and intrareceptor transmembrane interactions as universal therapeutic targets for a diverse variety of immune and other disorders. PMID:20519929

  13. A Functional Role for Anopheles gambiae Arrestin1 in Olfactory Signal Transduction

    PubMed Central

    Walker, William B.; Smith, Elaine M.; Jan, Taha; Zwiebel, L.J.

    2008-01-01

    Insect sensory arrestins act to desensitize visual and olfactory signal transduction pathways, as evidenced by the phenotypic effects of mutations in the genes encoding both Arr1 and Arr2 in Drosophila melanogaster. To assess whether such arrestins play similar roles in other, more medically relevant dipterans, we examined the ability of Anopheles gambiae sensory arrestin homologues AgArr1 and AgArr2 to rescue phenotypes associated with an olfactory deficit observed in D. melanogaster arrestin mutants. Of these, only AgArr1 facilitated significant phenotypic rescue of the corresponding Drosophila arr mutant olfactory phenotype, consistent with the view that functional orthology is shared by these Arr1 homologues. These results represent the first step in the functional characterization of AgArr1, which is highly expressed in olfactory appendages of An. gambiae in which it is likely to play an essential role in olfactory signal transduction. In addition to providing insight into the common elements of the peripheral olfactory system of dipterans, this work validates the importance of AgArr1 as a potential target for novel anti-malaria strategies that focus on olfactory-based behaviors in An. gambiae. PMID:18328499

  14. Amyloid beta-peptide disrupts carbachol-induced muscarinic cholinergic signal transduction in cortical neurons.

    PubMed Central

    Kelly, J F; Furukawa, K; Barger, S W; Rengen, M R; Mark, R J; Blanc, E M; Roth, G S; Mattson, M P

    1996-01-01

    Cholinergic pathways serve important functions in learning and memory processes, and deficits in cholinergic transmission occur in Alzheimer disease (AD). A subset of muscarinic cholinergic receptors are linked to G-proteins that activate phospholipase C, resulting in the liberation of inositol trisphosphate and Ca2+ release from intracellular stores. We now report that amyloid beta-peptide (Abeta), which forms plaques in the brain in AD, impairs muscarinic receptor activation of G proteins in cultured rat cortical neurons. Exposure of rodent fetal cortical neurons to Abeta25-35 and Abeta1-40 resulted in a concentration and time-dependent attenuation of carbachol-induced GTPase activity without affecting muscarinic receptor ligand binding parameters. Downstream events in the signal transduction cascade were similarly attenuated by Abeta. Carbachol-induced accumulation of inositol phosphates (IP, IP2, IP3, and IP4) was decreased and calcium imaging studies revealed that carbachol-induced release of calcium was severely impaired in neurons pretreated with Abeta. Muscarinic cholinergic signal transduction was disrupted with subtoxic levels of exposure to AP. The effects of Abeta on carbachol-induced GTPase activity and calcium release were attenuated by antioxidants, implicating free radicals in the mechanism whereby Abeta induced uncoupling of muscarinic receptors. These data demonstrate that Abeta disrupts muscarinic receptor coupling to G proteins that mediate induction of phosphoinositide accumulation and calcium release, findings that implicate Abeta in the impairment of cholinergic transmission that occurs in AD. PMID:8692890

  15. An integrated image analysis platform to quantify signal transduction in single cells.

    PubMed

    Pelet, Serge; Dechant, Reinhard; Lee, Sung Sik; van Drogen, Frank; Peter, Matthias

    2012-10-01

    Microscopy can provide invaluable information about biological processes at the single cell level. It remains a challenge, however, to extract quantitative information from these types of datasets. We have developed an image analysis platform named YeastQuant to simplify data extraction by offering an integrated method to turn time-lapse movies into single cell measurements. This platform is based on a database with a graphical user interface where the users can describe their experiments. The database is connected to the engineering software Matlab, which allows extracting the desired information by automatically segmenting and quantifying the microscopy images. We implemented three different segmentation methods that recognize individual cells under different conditions, and integrated image analysis protocols that allow measuring and analyzing distinct cellular readouts. To illustrate the power and versatility of YeastQuant, we investigated dynamic signal transduction processes in yeast. First, we quantified the expression of fluorescent reporters induced by osmotic stress to study noise in gene expression. Second, we analyzed the dynamic relocation of endogenous proteins from the cytoplasm to the cell nucleus, which provides a fast measure of pathway activity. These examples demonstrate that YeastQuant provides a versatile and expandable database and an experimental framework that improves image analysis and quantification of diverse microscopy-based readouts. Such dynamic single cell measurements are highly needed to establish mathematical models of signal transduction pathways. PMID:22976484

  16. P2CS: a two-component system resource for prokaryotic signal transduction research

    PubMed Central

    Barakat, Mohamed; Ortet, Philippe; Jourlin-Castelli, Cécile; Ansaldi, Mireille; Méjean, Vincent; Whitworth, David E

    2009-01-01

    Background With the escalation of high throughput prokaryotic genome sequencing, there is an ever-increasing need for databases that characterise, catalogue and present data relating to particular gene sets and genomes/metagenomes. Two-component system (TCS) signal transduction pathways are the dominant mechanisms by which micro-organisms sense and respond to external as well as internal environmental changes. These systems respond to a wide range of stimuli by triggering diverse physiological adjustments, including alterations in gene expression, enzymatic reactions, or protein-protein interactions. Description We present P2CS (Prokaryotic 2-Component Systems), an integrated and comprehensive database of TCS signal transduction proteins, which contains a compilation of the TCS genes within 755 completely sequenced prokaryotic genomes and 39 metagenomes. P2CS provides detailed annotation of each TCS gene including family classification, sequence features, functional domains, as well as genomic context visualization. To bypass the generic problem of gene underestimation during genome annotation, we also constituted and searched an ORFeome, which improves the recovery of TCS proteins compared to searches on the equivalent proteomes. Conclusion P2CS has been developed for computational analysis of the modular TCSs of prokaryotic genomes and metagenomes. It provides a complete overview of information on TCSs, including predicted candidate proteins and probable proteins, which need further curation/validation. The database can be browsed and queried with a user-friendly web interface at . PMID:19604365

  17. Exploitation of host signal transduction pathways and cytoskeletal functions by invasive bacteria.

    PubMed

    Rosenshine, I; Finlay, B B

    1993-01-01

    Many bacteria that cause disease have the capacity to enter into and live within eukaryotic cells such as epithelial cells and macrophages. The mechanisms used by these organisms to achieve and maintain this intracellular lifestyle vary considerably, but most mechanisms involve subversion and exploitation of host cell functions. Entry into non-phagocytic cells involves triggering host signal transduction mechanisms to induce rearrangement of the host cytoskeleton, thereby facilitating bacterial uptake. Once inside the host cell, intracellular pathogens either remain within membrane bound inclusions or escape to the cytoplasm. Those living in the cytoplasm can further pirate the host actin system, using actin as a mechanism to facilitate movement within and between host cells. Organisms remaining within the vacuole have specialized mechanisms for intracellular survival and growth which involve additional communication with the host cell. Some of the processes involved in the various steps of facultative intracellular parasitism are discussed in the context of subverting the host cell cytoskeleton and signal transduction pathways for bacterial benefit. PMID:8466472

  18. Signal transduction in endothelial cells by the angiogenesis inhibitor histidine-rich glycoprotein targets focal adhesions

    SciTech Connect

    Lee, Chunsik; Dixelius, Johan; Thulin, Asa; Kawamura, Harukiyo; Claesson-Welsh, Lena; Olsson, Anna-Karin . E-mail: Anna-Karin.Olsson@genpat.uu.se

    2006-08-01

    Histidine-rich glycoprotein (HRGP) is an abundant heparin-binding plasma protein. We have shown that a fragment released from the central histidine/proline-rich (His/Pro-rich) domain of HRGP blocks endothelial cell migration in vitro and vascularization and growth of murine fibrosarcoma in vivo. The minimal active HRGP domain exerting the anti-angiogenic effect was recently narrowed down to a 35 amino acid peptide, HRGP330, derived from the His/Pro-rich domain of HRGP. By use of a signal transduction antibody array representing 400 different signal transduction molecules, we now show that HRGP and the synthetic peptide HRGP330 specifically induce tyrosine phosphorylation of focal adhesion kinase and its downstream substrate paxillin in endothelial cells. HRGP/HRGP330 treatment of endothelial cells induced disruption of actin stress fibers, a process reversed by treatment of cells with the FAK inhibitor geldanamycin. In addition, VEGF-mediated endothelial cell tubular morphogenesis in a three-dimensional collagen matrix was inhibited by HRGP and HRGP330. In contrast, VEGF-induced proliferation was not affected by HRGP or HRGP330, demonstrating the central role of cell migration during tube formation. In conclusion, our data show that HRGP targets focal adhesions in endothelial cells, thereby disrupting the cytoskeletal organization and the ability of endothelial cells to assemble into vessel structures.

  19. [Cyclic Guanosine Monophosphate as a Mediator in Processes of Stress Signaling Transduction in Higher Plants].

    PubMed

    Dubovskaya, L V; Bakakina, Y S; Volotovski, I D

    2015-01-01

    Currently, biophysical mechanisms of stress signaling transduction became an object of consideration of researchers in connection with the urgent necessity to develop new techniques to enhance the sustainability and productivity of agricultural crops. The development of sensitive methods for the determination of cyclic guanosine monophosphate (cGMP) and comparative analysis of cGMP-dependent events in biological systems has contributed to progress in the understanding of the functioning of cGMP in plant cells. Currently, it is shown that cGMP as a secondary mediator is involved in such vital processes of growth and development of plants as seed germination, cell division, development of chloroplasts, flowering and regulation of stomatal movements. This review summarizes the available data in the literature about the role of cGMP in the responses of plant organisms to the action of stress factors of abiotic and biotic nature and its interaction with other intracellular mediators. With the use of existing ideas about the biophysical mechanisms of stress in plants, the basic elements of cGMP-dependent signal transduction system in a plant cell are considered. PMID:26394467

  20. Amyloid beta-peptide disrupts carbachol-induced muscarinic cholinergic signal transduction in cortical neurons.

    PubMed

    Kelly, J F; Furukawa, K; Barger, S W; Rengen, M R; Mark, R J; Blanc, E M; Roth, G S; Mattson, M P

    1996-06-25

    Cholinergic pathways serve important functions in learning and memory processes, and deficits in cholinergic transmission occur in Alzheimer disease (AD). A subset of muscarinic cholinergic receptors are linked to G-proteins that activate phospholipase C, resulting in the liberation of inositol trisphosphate and Ca2+ release from intracellular stores. We now report that amyloid beta-peptide (Abeta), which forms plaques in the brain in AD, impairs muscarinic receptor activation of G proteins in cultured rat cortical neurons. Exposure of rodent fetal cortical neurons to Abeta25-35 and Abeta1-40 resulted in a concentration and time-dependent attenuation of carbachol-induced GTPase activity without affecting muscarinic receptor ligand binding parameters. Downstream events in the signal transduction cascade were similarly attenuated by Abeta. Carbachol-induced accumulation of inositol phosphates (IP, IP2, IP3, and IP4) was decreased and calcium imaging studies revealed that carbachol-induced release of calcium was severely impaired in neurons pretreated with Abeta. Muscarinic cholinergic signal transduction was disrupted with subtoxic levels of exposure to AP. The effects of Abeta on carbachol-induced GTPase activity and calcium release were attenuated by antioxidants, implicating free radicals in the mechanism whereby Abeta induced uncoupling of muscarinic receptors. These data demonstrate that Abeta disrupts muscarinic receptor coupling to G proteins that mediate induction of phosphoinositide accumulation and calcium release, findings that implicate Abeta in the impairment of cholinergic transmission that occurs in AD. PMID:8692890

  1. The application of multiple biophysical cues to engineer functional neocartilage for treatment of osteoarthritis. Part II: signal transduction.

    PubMed

    Brady, Mariea A; Waldman, Stephen D; Ethier, C Ross

    2015-02-01

    The unique mechanoelectrochemical environment of cartilage has motivated researchers to investigate the effect of multiple biophysical cues, including mechanical, magnetic, and electrical stimulation, on chondrocyte biology. It is well established that biophysical stimuli promote chondrocyte proliferation, differentiation, and maturation within "biological windows" of defined dose parameters, including mode, frequency, magnitude, and duration of stimuli (see companion review Part I: Cellular Response). However, the underlying molecular mechanisms and signal transduction pathways activated in response to multiple biophysical stimuli remain to be elucidated. Understanding the mechanisms of biophysical signal transduction will deepen knowledge of tissue organogenesis, remodeling, and regeneration and aiding in the treatment of pathologies such as osteoarthritis. Further, this knowledge will provide the tissue engineer with a potent toolset to manipulate and control cell fate and subsequently develop functional replacement cartilage. The aim of this article is to review chondrocyte signal transduction pathways in response to mechanical, magnetic, and electrical cues. Signal transduction does not occur along a single pathway; rather a number of parallel pathways appear to be activated, with calcium signaling apparently common to all three types of stimuli, though there are different modes of activation. Current tissue engineering strategies, such as the development of "smart" functionalized biomaterials that enable the delivery of growth factors or integration of conjugated nanoparticles, may further benefit from targeting known signal transduction pathways in combination with external biophysical cues. PMID:25065615

  2. Excessive training impairs the insulin signal transduction in mice skeletal muscles.

    PubMed

    Pereira, Bruno C; da Rocha, Alisson L; Pinto, Ana P; Pauli, José R; de Moura, Leandro P; Mekary, Rania A; de Freitas, Ellen C; da Silva, Adelino S R

    2016-07-01

    The main aim of this investigation was to verify the effects of overtraining (OT) on the insulin and inflammatory signaling pathways in mice skeletal muscles. Rodents were divided into control (CT), overtrained by downhill running (OTR/down), overtrained by uphill running (OTR/up), and overtrained by running without inclination (OTR) groups. Rotarod, incremental load, exhaustive, and grip force tests were used to evaluate performance. Thirty-six hours after the grip force test, the extensor digitorum longus (EDL) and soleus were extracted for subsequent protein analyses. The three OT protocols led to similar responses of all performance evaluation tests. The phosphorylation of insulin receptor beta (pIRβ; Tyr), protein kinase B (pAkt; Ser473), and the protein levels of plasma membrane glucose transporter-4 (GLUT4) were lower in the EDL and soleus after the OTR/down protocol and in the soleus after the OTR/up and OTR protocols. While the pIRβ was lower after the OTR/up and OTR protocols, the pAkt was higher after the OTR/up in the EDL. The phosphorylation of IκB kinase alpha and beta (pIKKα/β; Ser180/181), stress-activated protein kinases/Jun amino-terminal kinases (pSAPK-JNK; Thr183/Tyr185), factor nuclear kappa B (pNFκB p65; Ser536), and insulin receptor substrate 1 (pIRS1; Ser307) were higher after the OTR/down protocol, but were not altered after the two other OT protocols. In summary, these data suggest that OT may lead to skeletal muscle insulin signaling pathway impairment, regardless of the predominance of eccentric contractions, although the insulin signal pathway impairment induced in OTR/up and OTR appeared to be muscle fiber-type specific. PMID:27189188

  3. Structure-Function of CD36 and Importance of Fatty Acid Signal Transduction in Fat Metabolism

    PubMed Central

    Pepino, Marta Yanina; Kuda, Ondrej; Samovski, Dmitri; Abumrad, Nada A

    2015-01-01

    CD36 is a scavenger receptor that functions in high affinity tissue uptake of long chain fatty acids (FA) and contributes under excessive fat supply to lipid accumulation and metabolic dysfunction. This review describes recent evidence regarding the CD36 FA binding site and a potential mechanism for FA transfer. It also presents the view that CD36 and FA signaling coordinate fat utilization based on newly identified CD36 actions that involve oral fat perception, intestinal fat absorption, secretion of the peptides cholecystokinin and secretin, regulation of hepatic lipoprotein output, activation of beta oxidation by muscle and regulation of the production of the FA derived bioactive eicosanoids. Thus abnormalities of fat metabolism and the associated pathology might involve dysfunction of CD36-mediated signal transduction in addition to the changes of FA uptake. PMID:24850384

  4. An intimate link: two-component signal transduction systems and metal transport systems in bacteria

    PubMed Central

    Singh, Kamna; Senadheera, Dilani B; Cvitkovitch, Dennis G

    2014-01-01

    Bacteria have evolved various strategies to contend with high concentrations of environmental heavy metal ions for rapid, adaptive responses to maintain cell viability. Evidence gathered in the past two decades suggests that bacterial two-component signal transduction systems (TCSTSs) are intimately involved in monitoring cation accumulation, and can regulate the expression of related metabolic and virulence genes to elicit adaptive responses to changes in the concentration of these ions. Using examples garnered from recent studies, we summarize the cross-regulatory relationships between metal ions and TCSTSs. We present evidence of how bacterial TCSTSs modulate metal ion homeostasis and also how metal ions, in turn, function to control the activities of these signaling systems linked with bacterial survival and virulence. PMID:25437189

  5. A CRISPR-Based Toolbox for Studying T Cell Signal Transduction

    PubMed Central

    Chi, Shen; Weiss, Arthur; Wang, Haopeng

    2016-01-01

    CRISPR/Cas9 system is a powerful technology to perform genome editing in a variety of cell types. To facilitate the application of Cas9 in mapping T cell signaling pathways, we generated a toolbox for large-scale genetic screens in human Jurkat T cells. The toolbox has three different Jurkat cell lines expressing distinct Cas9 variants, including wild-type Cas9, dCas9-KRAB, and sunCas9. We demonstrated that the toolbox allows us to rapidly disrupt endogenous gene expression at the DNA level and to efficiently repress or activate gene expression at the transcriptional level. The toolbox, in combination with multiple currently existing genome-wide sgRNA libraries, will be useful to systematically investigate T cell signal transduction using both loss-of-function and gain-of-function genetic screens. PMID:27057542

  6. Dissecting single-molecule signal transduction in carbon nanotube circuits with protein engineering

    PubMed Central

    Choi, Yongki; Olsen, Tivoli J.; Sims, Patrick C.; Moody, Issa S.; Corso, Brad L.; Dang, Mytrang N.; Weiss, Gregory A.; Collins, Philip G.

    2013-01-01

    Single molecule experimental methods have provided new insights into biomolecular function, dynamic disorder, and transient states that are all invisible to conventional measurements. A novel, non-fluorescent single molecule technique involves attaching single molecules to single-walled carbon nanotube field-effective transistors (SWNT FETs). These ultrasensitive electronic devices provide long-duration, label-free monitoring of biomolecules and their dynamic motions. However, generalization of the SWNT FET technique first requires design rules that can predict the success and applicability of these devices. Here, we report on the transduction mechanism linking enzymatic processivity to electrical signal generation by a SWNT FET. The interaction between SWNT FETs and the enzyme lysozyme was systematically dissected using eight different lysozyme variants synthesized by protein engineering. The data prove that effective signal generation can be accomplished using a single charged amino acid, when appropriately located, providing a foundation to widely apply SWNT FET sensitivity to other biomolecular systems. PMID:23323846

  7. Involvement of the second messenger cAMP in gravity-signal transduction in physarum

    NASA Astrophysics Data System (ADS)

    Block, I.; Rabien, H.; Ivanova, K.

    The aim of the investigation was to clarify, whether cellular signal processing following graviperception involves second messenger pathways. The test object was a most gravisensitive free-living ameboid cell, the myxomycete (acellular slime mold) Physarum polycephalum. It was demonstrated that the motor response is related to acceleration-dependent changes in the levels of the cellular second messenger cyclic adenosine monophosphate (cAMP). Rotating Physarum plasmodia in the gravity field of the Earth about a horizontal axis increased their cAMP concentration. Depriving the cells for a few days of the acceleration stimulus (near weightlessness in a space experiment on STS-69) slightly lowered plasmodial cAMP levels. Thus, the results provide first indications that the acceleration-stimulus signal transduction chain of Physarum uses an ubiquitous second messenger pathway.

  8. Computational Modeling of PI3K/AKT and MAPK Signaling Pathways in Melanoma Cancer

    PubMed Central

    Pappalardo, Francesco; Russo, Giulia; Candido, Saverio; Pennisi, Marzio; Cavalieri, Salvatore; Motta, Santo; McCubrey, James A.; Nicoletti, Ferdinando; Libra, Massimo

    2016-01-01

    Background Malignant melanoma is an aggressive tumor of the skin and seems to be resistant to current therapeutic approaches. Melanocytic transformation is thought to occur by sequential accumulation of genetic and molecular alterations able to activate the Ras/Raf/MEK/ERK (MAPK) and/or the PI3K/AKT (AKT) signalling pathways. Specifically, mutations of B-RAF activate MAPK pathway resulting in cell cycle progression and apoptosis prevention. According to these findings, MAPK and AKT pathways may represent promising therapeutic targets for an otherwise devastating disease. Result Here we show a computational model able to simulate the main biochemical and metabolic interactions in the PI3K/AKT and MAPK pathways potentially involved in melanoma development. Overall, this computational approach may accelerate the drug discovery process and encourages the identification of novel pathway activators with consequent development of novel antioncogenic compounds to overcome tumor cell resistance to conventional therapeutic agents. The source code of the various versions of the model are available as S1 Archive. PMID:27015094

  9. miR-21 promotes human nucleus pulposus cell proliferation through PTEN/AKT signaling.

    PubMed

    Liu, Hongzhe; Huang, Xiangwang; Liu, Xiangyang; Xiao, Sheng; Zhang, Yi; Xiang, Tiecheng; Shen, Xiongjie; Wang, Guoping; Sheng, Bin

    2014-01-01

    The precise role of nucleus pulposus cell proliferation in the pathogenesis of intervertebral disc degeneration remains to be elucidated. Recent findings have revealed that microRNAs, a class of small noncoding RNAs, may regulate cell proliferation in many pathological conditions. Here, we showed that miR-21 was significantly upregulated in degenerative nucleus pulposus tissues when compared with nucleus pulposus tissues that were isolated from patients with idiopathic scoliosis and that miR-10b levels were associated with disc degeneration grade. Moreover, bioinformatics target prediction identified PTEN as a putative target of miR-21. miR-21 inhibited PTEN expression by directly targeting the 3'UTR, and this inhibition was abolished through miR-21 binding site mutations. miR-21 overexpression stimulated cell proliferation and AKT signaling pathway activation, which led to cyclin D1 translation. Additionally, the increase in proliferation and cyclin D1 expression induced by miR-21 overexpression was almost completely blocked by Ly294002, an AKT inhibitor. Taken together, aberrant miR-21 upregulation in intervertebral disc degeneration could target PTEN, which would contribute to abnormal nucleus pulposus cell proliferation through derepressing the Akt pathway. Our study also underscores the potential of miR-21 and the PTEN/Akt pathway as novel therapeutic targets in intervertebral disc degeneration. PMID:24603539

  10. miR-21 Promotes Human Nucleus Pulposus Cell Proliferation through PTEN/AKT Signaling

    PubMed Central

    Liu, Hongzhe; Huang, Xiangwang; Liu, Xiangyang; Xiao, Sheng; Zhang, Yi; Xiang, Tiecheng; Shen, Xiongjie; Wang, Guoping; Sheng, Bin

    2014-01-01

    The precise role of nucleus pulposus cell proliferation in the pathogenesis of intervertebral disc degeneration remains to be elucidated. Recent findings have revealed that microRNAs, a class of small noncoding RNAs, may regulate cell proliferation in many pathological conditions. Here, we showed that miR-21 was significantly upregulated in degenerative nucleus pulposus tissues when compared with nucleus pulposus tissues that were isolated from patients with idiopathic scoliosis and that miR-10b levels were associated with disc degeneration grade. Moreover, bioinformatics target prediction identified PTEN as a putative target of miR-21. miR-21 inhibited PTEN expression by directly targeting the 3′UTR, and this inhibition was abolished through miR-21 binding site mutations. miR-21 overexpression stimulated cell proliferation and AKT signaling pathway activation, which led to cyclin D1 translation. Additionally, the increase in proliferation and cyclin D1 expression induced by miR-21 overexpression was almost completely blocked by Ly294002, an AKT inhibitor. Taken together, aberrant miR-21 upregulation in intervertebral disc degeneration could target PTEN, which would contribute to abnormal nucleus pulposus cell proliferation through derepressing the Akt pathway. Our study also underscores the potential of miR-21 and the PTEN/Akt pathway as novel therapeutic targets in intervertebral disc degeneration. PMID:24603539

  11. New paradigms in chemokine receptor signal transduction: Moving beyond the two-site model.

    PubMed

    Kleist, Andrew B; Getschman, Anthony E; Ziarek, Joshua J; Nevins, Amanda M; Gauthier, Pierre-Arnaud; Chevigné, Andy; Szpakowska, Martyna; Volkman, Brian F

    2016-08-15

    Chemokine receptor (CKR) signaling forms the basis of essential immune cellular functions, and dysregulated CKR signaling underpins numerous disease processes of the immune system and beyond. CKRs, which belong to the seven transmembrane domain receptor (7TMR) superfamily, initiate signaling upon binding of endogenous, secreted chemokine ligands. Chemokine-CKR interactions are traditionally described by a two-step/two-site mechanism, in which the CKR N-terminus recognizes the chemokine globular core (i.e. site 1 interaction), followed by activation when the unstructured chemokine N-terminus is inserted into the receptor TM bundle (i.e. site 2 interaction). Several recent studies challenge the structural independence of sites 1 and 2 by demonstrating physical and allosteric links between these supposedly separate sites. Others contest the functional independence of these sites, identifying nuanced roles for site 1 and other interactions in CKR activation. These developments emerge within a rapidly changing landscape in which CKR signaling is influenced by receptor PTMs, chemokine and CKR dimerization, and endogenous non-chemokine ligands. Simultaneous advances in the structural and functional characterization of 7TMR biased signaling have altered how we understand promiscuous chemokine-CKR interactions. In this review, we explore new paradigms in CKR signal transduction by considering studies that depict a more intricate architecture governing the consequences of chemokine-CKR interactions. PMID:27106080

  12. S6K Promotes Dopaminergic Neuronal Differentiation Through PI3K/Akt/mTOR-Dependent Signaling Pathways in Human Neural Stem Cells.

    PubMed

    Lee, Jeong Eun; Lim, Mi Sun; Park, Jae Hyun; Park, Chang Hwan; Koh, Hyun Chul

    2016-08-01

    It has recently been reported that the phosphoinositide 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) signaling pathway regulates neuronal differentiation of neural stem cells (NSCs) derived from rats or mice and is essential for the self-renewal of human embryonic stem cells (hESCs). However, the roles of PI3K/Akt/mTOR signaling pathways during proliferation and dopaminergic neuronal differentiation of human neural stem cells (hNSCs) are poorly understood. In this study, we examined the effect of regulation of these intracellular signaling pathways in hNSCs on the potential to maintain proliferation and induce dopaminergic neuronal differentiation. Dopaminergic neuronal differentiation depended on the concentration of insulin in our culture system. Inhibition of PI3K/Akt with LY294002 reduced proliferation and inhibited dopaminergic neuronal differentiation of these cells. We also found that rapamycin, a specific inhibitor of mTOR, significantly reduced neuronal differentiation without affecting proliferation. Inhibition of the Akt/mTOR signaling pathway led to inhibition of p70 ribosomal S6 kinase (S6K) signaling, which reduced dopaminergic neuronal differentiation in hNSCs. Inhibition of S6K by a specific chemical inhibitor, PF-4708671 inhibited dopaminergic neuronal differentiation of hNSCs. As expected, transduction with a dominant negative S6K1 (S6K1-DN) construct impaired dopaminergic neuronal differentiation of hNSCs. Conversely, overexpression of constitutively active S6K1 (S6K1-CA) promoted dopaminergic neuronal differentiation of these cells. In a survival study, 4 weeks after transplantation, no or very few donor cells were viable in striata grafted with S6K1-DN-transduced hNSCs. In contrast, S6K1-CA-transduced hNSCs survived, integrated into striata to generate tubular masses of grafts and differentiated toward TH-positive cells. Taken together, these data demonstrated that insulin promotes dopaminergic neuronal differentiation through a PI

  13. IGF-I binding and receptor signal transduction in primary cell culture of muscle cells of gilthead sea bream: changes throughout in vitro development.

    PubMed

    Montserrat, N; Sánchez-Gurmaches, J; García de la Serrana, D; Navarro, M I; Gutiérrez, J

    2007-12-01

    We examined the possibility of culturing muscle cells of gilthead sea bream in vitro and assessed variations in insulin-like growth factor-I (IGF-I) binding during myocyte development. The viability of the cell culture was determined by fluorescence-activated cell-sorting analysis, which showed that the percentage of dead cells decreased with cell differentiation. The intracellular reduction of MTT into formazan pigment was preferentially carried out as cells differentiated (from day 4) indicating an increase in metabolic activity. IGF-I-binding assays demonstrated that the number of receptors increased from 190 +/- 0.09 fmol/mg protein in myocytes at day 5 to 360 +/- 0.09 fmol/mg protein in myotubes at day 12. The affinity of IGF-I receptors did not change significantly during cell development (from 0.89 +/- 0.09 to 0.98 +/- 0.09 nM). The activation of various kinase (ERK 1/2 MAPK and Akt/PKB) proteins by IGFs and insulin was studied by means of Western blot analysis. Levels of MAPK-P increased after IGF and insulin treatment during the first stages of cell culture, with a low response being observed at day 15, whereas IGFs displayed a stimulatory effect on Akt-P throughout the cell culture period, even on day 15. This study thus shows that (1) gilthead sea bream myocytes can be cultured, (2) they express functional IGF-I receptors that increase in number as they differentiate in vitro; (3) IGF signalling transduction through IGF-I receptors stimulates the MAPK and Akt pathways, depending on the development stage of the muscle cell culture. PMID:17940802

  14. The Roles of Peroxiredoxin and Thioredoxin in Hydrogen Peroxide Sensing and in Signal Transduction

    PubMed Central

    Netto, Luis E. S.; Antunes, Fernando

    2016-01-01

    A challenge in the redox field is the elucidation of the molecular mechanisms, by which H2O2 mediates signal transduction in cells. This is relevant since redox pathways are disturbed in some pathologies. The transcription factor OxyR is the H2O2 sensor in bacteria, whereas Cys-based peroxidases are involved in the perception of this oxidant in eukaryotic cells. Three possible mechanisms may be involved in H2O2 signaling that are not mutually exclusive. In the simplest pathway, H2O2 signals through direct oxidation of the signaling protein, such as a phosphatase or a transcription factor. Although signaling proteins are frequently observed in the oxidized state in biological systems, in most cases their direct oxidation by H2O2 is too slow (101 M−1s−1 range) to outcompete Cys-based peroxidases and glutathione. In some particular cellular compartments (such as vicinity of NADPH oxidases), it is possible that a signaling protein faces extremely high H2O2 concentrations, making the direct oxidation feasible. Alternatively, high H2O2 levels can hyperoxidize peroxiredoxins leading to local building up of H2O2 that then could oxidize a signaling protein (floodgate hypothesis). In a second model, H2O2 oxidizes Cys-based peroxidases that then through thiol-disulfide reshuffling would transmit the oxidized equivalents to the signaling protein. The third model of signaling is centered on the reducing substrate of Cys-based peroxidases that in most cases is thioredoxin. Is this model, peroxiredoxins would signal by modulating the thioredoxin redox status. More kinetic data is required to allow the identification of the complex network of thiol switches. PMID:26813662

  15. Epidermal Growth Factor Receptor in Glioma: Signal Transduction, Neuropathology, Imaging, and Radioresistance1

    PubMed Central

    Hatanpaa, Kimmo J; Burma, Sandeep; Zhao, Dawen; Habib, Amyn A

    2010-01-01

    Aberrant epidermal growth factor receptor (EGFR) signaling is common in cancer. Increased expression of wild type and mutant EGFR is a widespread feature of diverse types of cancer. EGFR signaling in cancer has been the focus of intense investigation for decades primarily for two reasons. First, aberrant EGFR signaling is likely to play an important role in the pathogenesis of cancer, and therefore, the mechanisms of EGFR-mediated oncogenic signaling are of interest. Second, the EGFR signaling system is an attractive target for therapeutic intervention. EGFR gene amplification and overexpression are a particularly striking feature of glioblastoma (GBM), observed in approximately 40% of tumors. GBM is the most common primary malignant tumor of the central nervous system in adults. In approximately 50% of tumors with EGFR amplification, a specific EGFR mutant (EGFRvIII, also known as EGFR type III, de2-7, ΔEGFR) can be detected. This mutant is highly oncogenic and is generated from a deletion of exons 2 to 7 of the EGFR gene, which results in an in-frame deletion of 267 amino acids from the extracellular domain of the receptor. EGFRvIII is unable to bind ligand, and it signals constitutively. Although EGFRvIII has the same signaling domain as the wild type receptor, it seems to generate a distinct set of downstream signals that may contribute to an increased tumorigenicity. In this review, we discuss recent progress in key aspects of EGFR signaling in GBM, focusing on neuropathology, signal transduction, imaging of the EGFR, and the role of the EGFR in mediating resistance to radiation therapy in GBM. PMID:20824044

  16. Dioscin inhibits osteoclast differentiation and bone resorption though down-regulating the Akt signaling cascades

    SciTech Connect

    Qu, Xinhua; Zhai, Zanjing; Liu, Xuqiang; Li, Haowei; Ouyang, Zhengxiao; Wu, Chuanlong; Liu, Guangwang; Fan, Qiming; Tang, Tingting; Qin, An; Dai, Kerong

    2014-01-10

    Highlights: •A natural-derived compound, dioscin, suppresses osteoclast formation and bone resorption. •Dioscin inhibits osteolytic bone loss in vivo. •Dioscin impairs the Akt signaling cascades pathways during osteoclastogenesis. •Dioscin have therapeutic value in treating osteoclast-related diseases. -- Abstract: Bone resorption is the unique function of osteoclasts (OCs) and is critical for both bone homeostasis and pathologic bone diseases including osteoporosis, rheumatoid arthritis and tumor bone metastasis. Thus, searching for natural compounds that may suppress osteoclast formation and/or function is promising for the treatment of osteoclast-related diseases. In this study, we for the first time demonstrated that dioscin suppressed RANKL-mediated osteoclast differentiation and bone resorption in vitro in a dose-dependent manner. The suppressive effect of dioscin is supported by the reduced expression of osteoclast-specific markers. Further molecular analysis revealed that dioscin abrogated AKT phosphorylation, which subsequently impaired RANKL-induced nuclear factor-kappaB (NF-κB) signaling pathway and inhibited NFATc1 transcriptional activity. Moreover, in vivo studies further verified the bone protection activity of dioscin in osteolytic animal model. Together our data demonstrate that dioscin suppressed RANKL-induced osteoclast formation and function through Akt signaling cascades. Therefore, dioscin is a potential natural agent for the treatment of osteoclast-related diseases.

  17. Deoxycholyltaurine rescues human colon cancer cells from apoptosis by activating EGFR-dependent PI3K/Akt signaling.

    PubMed

    Raufman, Jean-Pierre; Shant, Jasleen; Guo, Chang Yue; Roy, Sanjit; Cheng, Kunrong

    2008-05-01

    Recent studies indicate that secondary bile acids promote colon cancer cell proliferation but their role in maintaining cell survival has not been explored. We found that deoxycholyltaurine (DCT) markedly attenuated both unstimulated and TNF-alpha-stimulated programmed cell death in colon cancer cells by a phosphatidylinositol 3-kinase (PI3K)-dependent mechanism. To examine the role of bile acids and PI3K signaling in maintaining colon cancer cell survival, we explored the role of signaling downstream of bile acid-induced activation of the epidermal growth factor receptor (EGFR) in regulating both apoptosis and proliferation of HT-29 and H508 human colon cancer cells. DCT caused dose- and time-dependent Akt (Ser(473)) phosphorylation, a commonly used marker of activated PI3K/Akt signaling. Both EGFR kinase and PI3K inhibitors attenuated DCT-induced Akt phosphorylation and Akt activation, as demonstrated by reduced phosphorylation of a GSK-3-paramyosin substrate. Transfection of HT-29 cells with kinase-dead EGFR (K721M) reduced DCT-induced Akt phosphorylation. In HT-29 cells, EGFR and PI3K inhibitors as well as transfection with dominant negative AKT attenuated DCT-induced cell proliferation. DCT-induced PI3K/Akt activation resulted in downstream phosphorylation of GSK-3 (Ser(21/9)) and BAD (Ser(136)), and nuclear translocation (activation) of NF-kappaB, thereby confirming that DCT-induced activation of PI3K/Akt signaling regulates both proproliferative and prosurvival signals. Collectively, these results indicate that DCT-induced activation of post-EGFR PI3K/Akt signaling stimulates both colon cancer cell survival and proliferation. PMID:18064605

  18. Mitochondrial reactive oxygen species perturb AKT/cyclin D1 cell cycle signaling via oxidative inactivation of PP2A in lowdose irradiated human fibroblasts

    PubMed Central

    Shimura, Tsutomu; Sasatani, Megumi; Kamiya, Kenji; Kawai, Hidehiko; Inaba, Yohei; Kunugita, Naoki

    2016-01-01

    Here we investigated the cellular response of normal human fibroblasts to repeated exposure to low-dose radiation. In contrast to acute single radiation, low-dose fractionated radiation (FR) with 0.01 Gy/fraction or 0.05 Gy/fraction for 31 days increased in mitochondrial mass, decreased cellular levels of the antioxidant glutathione and caused persistent accumulation of mitochondrial reactive oxygen species (ROS). Excess ROS promoted oxidative inactivation of protein phosphatase PP2A which in turn led to disruption of normal negative feed-back control of AKT/cyclin D1 signaling in cells treated with long-term FR. The resulting abnormal nuclear accumulation of cyclin D1 causes growth retardation, cellular senescence and genome instability in low-dose irradiated cells. Thus, loss of redox control and subsequently elevated levels of ROS perturb signal transduction as a result of oxidative stress. Our study highlights a specific role of mitochondrial ROS in perturbation of AKT/cyclin D1 cell cycle signaling after low-dose long-term FR. The antioxidants N-acetyl-L-cysteine, TEMPO and mitochondrial-targeted antioxidant Mito-TEMPO provided protection against the harmful cell cycle perturbations induced by low-dose long-term FR. PMID:26657292

  19. Molecular Insights into Toluene Sensing in the TodS/TodT Signal Transduction System*

    PubMed Central

    Koh, Serry; Hwang, Jungwon; Guchhait, Koushik; Lee, Eun-Gyeong; Kim, Sang-Yoon; Kim, Sujin; Lee, Sangmin; Chung, Jeong Min; Jung, Hyun Suk; Lee, Sang Jun; Ryu, Choong-Min; Lee, Seung-Goo; Oh, Tae-Kwang; Kwon, Ohsuk; Kim, Myung Hee

    2016-01-01

    TodS is a sensor kinase that responds to various monoaromatic compounds, which either cause an agonistic or antagonistic effect on phosphorylation of its cognate response regulator TodT, and controls tod operon expression in Pseudomonas putida strains. We describe a molecular sensing mechanism of TodS that is activated in response to toluene. The crystal structures of the TodS Per-Arnt-Sim (PAS) 1 sensor domain (residues 43–164) and its complex with toluene (agonist) or 1,2,4-trimethylbenzene (antagonist) show a typical β2α3β3 PAS fold structure (residues 45–149), forming a hydrophobic ligand-binding site. A signal transfer region (residues 150–163) located immediately after the canonical PAS fold may be intrinsically flexible and disordered in both apo-PAS1 and antagonist-bound forms and dramatically adapt an α-helix upon toluene binding. This structural change in the signal transfer region is proposed to result in signal transmission to activate the TodS/TodT two-component signal transduction system. Site-directed mutagenesis and β-galactosidase assays using a P. putida reporter strain system verified the essential residues involved in ligand sensing and signal transfer and suggest that the Phe46 residue acts as a ligand-specific switch. PMID:26903514

  20. Molecular Insights into Toluene Sensing in the TodS/TodT Signal Transduction System.

    PubMed

    Koh, Serry; Hwang, Jungwon; Guchhait, Koushik; Lee, Eun-Gyeong; Kim, Sang-Yoon; Kim, Sujin; Lee, Sangmin; Chung, Jeong Min; Jung, Hyun Suk; Lee, Sang Jun; Ryu, Choong-Min; Lee, Seung-Goo; Oh, Tae-Kwang; Kwon, Ohsuk; Kim, Myung Hee

    2016-04-15

    TodS is a sensor kinase that responds to various monoaromatic compounds, which either cause an agonistic or antagonistic effect on phosphorylation of its cognate response regulator TodT, and controls tod operon expression in Pseudomonas putida strains. We describe a molecular sensing mechanism of TodS that is activated in response to toluene. The crystal structures of the TodS Per-Arnt-Sim (PAS) 1 sensor domain (residues 43-164) and its complex with toluene (agonist) or 1,2,4-trimethylbenzene (antagonist) show a typical β2α3β3 PAS fold structure (residues 45-149), forming a hydrophobic ligand-binding site. A signal transfer region (residues 150-163) located immediately after the canonical PAS fold may be intrinsically flexible and disordered in both apo-PAS1 and antagonist-bound forms and dramatically adapt an α-helix upon toluene binding. This structural change in the signal transfer region is proposed to result in signal transmission to activate the TodS/TodT two-component signal transduction system. Site-directed mutagenesis and β-galactosidase assays using a P. putida reporter strain system verified the essential residues involved in ligand sensing and signal transfer and suggest that the Phe(46) residue acts as a ligand-specific switch. PMID:26903514

  1. Signal transduction during C. elegans vulval development: a NeverEnding story.

    PubMed

    Schmid, Tobias; Hajnal, Alex

    2015-06-01

    The Caenorhabditis elegans hermaphrodite vulva is one of the best studied models for signal transduction and cell fate determination during organogenesis. Systematic forward genetic screens have identified a complex and highly interconnected signaling network formed by the conserved EGFR, NOTCH, and WNT signaling pathways that specifies an invariant pattern of cell fates among the six vulval precursor cells (VPCs). Multiple inhibitory interactions between the EGFR and NOTCH pathways ensure the selection of a single 1° VPC that is always flanked by two 2° VPCs thanks to lateral NOTCH signaling. Building on this 'central dogma' of cell fate specification, scientists have investigated a broad spectrum of novel questions that are summarized in this review. For example, vulval development is a unique model to study the intracellular trafficking of signaling molecules, such as NOTCH or EGFR, to investigate the interactions between the cell cycle and cell fate specification pathways, and to observe epithelial tube morphogenesis and cell invasion at single-cell resolution. Finally, computer scientists have integrated the experimental data into mathematical and state-based 'in silico' models of vulval development, allowing them to test the completeness and limits of our current understanding. PMID:25677930

  2. Non Linear Programming (NLP) formulation for quantitative modeling of protein signal transduction pathways.

    PubMed

    Mitsos, Alexander; Melas, Ioannis N; Morris, Melody K; Saez-Rodriguez, Julio; Lauffenburger, Douglas A; Alexopoulos, Leonidas G

    2012-01-01

    Modeling of signal transduction pathways plays a major role in understanding cells' function and predicting cellular response. Mathematical formalisms based on a logic formalism are relatively simple but can describe how signals propagate from one protein to the next and have led to the construction of models that simulate the cells response to environmental or other perturbations. Constrained fuzzy logic was recently introduced to train models to cell specific data to result in quantitative pathway models of the specific cellular behavior. There are two major issues in this pathway optimization: i) excessive CPU time requirements and ii) loosely constrained optimization problem due to lack of data with respect to large signaling pathways. Herein, we address both issues: the former by reformulating the pathway optimization as a regular nonlinear optimization problem; and the latter by enhanced algorithms to pre/post-process the signaling network to remove parts that cannot be identified given the experimental conditions. As a case study, we tackle the construction of cell type specific pathways in normal and transformed hepatocytes using medium and large-scale functional phosphoproteomic datasets. The proposed Non Linear Programming (NLP) formulation allows for fast optimization of signaling topologies by combining the versatile nature of logic modeling with state of the art optimization algorithms. PMID:23226239

  3. Functional Analysis of the Mycobacterium tuberculosis MprAB Two-Component Signal Transduction System

    PubMed Central

    Zahrt, Thomas C.; Wozniak, Christopher; Jones, Denise; Trevett, Andrea

    2003-01-01

    The mechanisms utilized by Mycobacterium tuberculosis to establish, maintain, or reactivate from latent infection in the host are largely unknown but likely include genes that mediate adaptation to conditions encountered during persistence. Previously, a two-component signal transduction system, mprAB, was found to be required in M. tuberculosis for establishment and maintenance of persistent infection in a tissue- and stage-specific fashion. To begin to characterize the role of this system in M. tuberculosis physiology and virulence, a functional analysis of the mprA and mprB gene products was initiated. Here, evidence is presented demonstrating that sensor kinase MprB and response regulator MprA function as an intact signal-transducing pair in vitro and in vivo. Sensor kinase MprB can be autophosphorylated, can donate phosphate to MprA, and can act as a phospho-MprA phosphatase in vitro. Correspondingly, response regulator MprA can accept phosphate from MprB or from small phosphodonors including acetyl phosphate. Mutagenesis of residues His249 in MprB and Asp48 in MprA abolished the ability of these proteins to be phosphorylated in vitro. Introduction of these alleles into Mycobacterium bovis BCGattenuated virulence in macrophages in vivo. Together, these results support a role for the mprAB two-component system in M. tuberculosis physiology and pathogenesis. Characterization of two-component signal transduction systems will enhance our understanding of processes regulated by M. tuberculosis during acute and/or persistent infection in the host. PMID:14638785

  4. Molecular Analysis of the Graviperception Signal Transduction in the Flagellate Euglena

    NASA Astrophysics Data System (ADS)

    Häder, Donat; Daiker, Viktor; Richter, Peter; Lebert, Michael

    The unicellular flagellate Euglena gracilis perceives and reacts to the gravitational vector of the Earth. Recent results of experiments on parabolic rocket flights have revealed that the orientation can be explained by passive orientation only to a small extend while the remainder relies on an active physiological sensor and an internal sensory transduction chain. Our current working hypothesis is based on the fact that the cellular contents is heavier than the surrounding medium and consequently exerts pressure onto the lower membrane where it activates mechano-sensitive ion channels located at the front end under the trailing flagellum. We recently succeeded in identifying these channels as gene products of the TRP family. RNAi of the corresponding gene abolished graviperception. These channels allow a gated influx of calcium which depolarizes the internal electrical potential and eventually causes a course correction by the flagellar beating. The inwardly gated calcium binds to a specific calmodulin which is likewise an intrinsic element of the signal transduction chain. RNAi of the related mRNA also stopped graviperception. This calmodulin is thought to activate an adenylyl cyclase which generates cyclic AMP which in turn modulates the beating pattern of the flagellum.

  5. Interferon-gamma regulates nucleoside transport systems in macrophages through signal transduction and activator of transduction factor 1 (STAT1)-dependent and -independent signalling pathways.

    PubMed

    Soler, Concepció; Felipe, Antonio; García-Manteiga, José; Serra, Maria; Guillén-Gómez, Elena; Casado, F Javier; MacLeod, Carol; Modolell, Manuel; Pastor-Anglada, Marçal; Celada, Antonio

    2003-11-01

    The expressions of CNT and ENT (concentrative and equilibrative nucleoside transporters) in macrophages are differentially regulated by IFN-gamma (interferon-gamma). This cytokine controls gene expression through STAT1-dependent and/or -independent pathways (where STAT1 stands for signal transduction and activator of transcription 1). In the present study, the role of STAT1 in the response of nucleoside transporters to IFN-gamma was studied using macrophages from STAT1 knockout mice. IFN-gamma triggered an inhibition of ENT1-related nucleoside transport activity through STAT1-dependent mechanisms. Such inhibition of macrophage growth and ENT1 activity by IFN-gamma is required for DNA synthesis. Interestingly, IFN-gamma led to an induction of the CNT1- and CNT2-related nucleoside transport activities independent of STAT1, thus ensuring the supply of extracellular nucleosides for the STAT1-independent RNA synthesis. IFN-gamma up-regulated CNT2 mRNA and CNT1 protein levels and down-regulated ENT1 mRNA in both wild-type and STAT1 knockout macrophages. This is consistent with a STAT1-independent, long-term-mediated, probably transcription-dependent, regulation of nucleoside transporter genes. Moreover, STAT1-dependent post-transcriptional mechanisms are implicated in the regulation of ENT1 activity. Although nitric oxide is involved in the regulation of ENT1 activity in B-cells at a post-transcriptional level, our results show that STAT1-dependent induction of nitric oxide by IFN-gamma is not implicated in the regulation of ENT1 activity in macrophages. Our results indicate that both STAT1-dependent and -independent pathways are involved in the regulation of nucleoside transporters by IFN-gamma in macrophages. PMID:12868960

  6. Imbalanced signal transduction in regulatory T cells expressing the transcription factor FoxP3.

    PubMed

    Yan, Dapeng; Farache, Julia; Mingueneau, Michael; Mathis, Diane; Benoist, Christophe

    2015-12-01

    FoxP3(+) T regulatory (Treg) cells have a fundamental role in immunological tolerance, with transcriptional and functional phenotypes that demarcate them from conventional CD4(+) T cells (Tconv). Differences between these two lineages in the signaling downstream of T-cell receptor-triggered activation have been reported, and there are different requirements for some signaling factors. Seeking a comprehensive view, we found that Treg cells have a broadly dampened activation of several pathways and signaling nodes upon TCR-mediated activation, with low phosphorylation of CD3ζ, SLP76, Erk1/2, AKT, or S6 and lower calcium flux. In contrast, STAT phosphorylation triggered by interferons, IL2 or IL6, showed variations between Treg and Tconv in magnitude or choice of preferential STAT activation but no general Treg signaling defect. Much, but not all, of the Treg/Tconv difference in TCR-triggered responses could be attributed to lower responsiveness of antigen-experienced cells with CD44(hi) or CD62L(lo) phenotypes, which form a greater proportion of the Treg pool. Candidate regulators were tested, but the Treg/Tconv differential could not be explained by overexpression in Treg cells of the signaling modulator CD5, the coinhibitors PD-1 and CTLA4, or the regulatory phosphatase DUSP4. However, transcriptome profiling in Dusp4-deficient mice showed that DUSP4 enhances the expression of a segment of the canonical Treg transcriptional signature, which partially overlaps with the TCR-dependent Treg gene set. Thus, Treg cells, likely because of their intrinsically higher reactivity to self, tune down TCR signals but seem comparatively more attuned to cytokines or other intercellular signals. PMID:26627244

  7. Targeting the PI3K/AKT/mTOR Signaling Axis in Children with Hematologic Malignancies

    PubMed Central

    Barrett, David; Brown, Valerie I.; Grupp, Stephan A.; Teachey, David T.

    2014-01-01

    The phosphatidylinositiol 3-kinase (PI3K), AKT, mammalian target of rapamycin (mTOR) signaling pathway (PI3K/AKT/mTOR) is frequently dysregulated in disorders of cell growth and survival, including a number of pediatric hematologic malignancies. The pathway can be abnormally activated in childhood acute lymphoblastic leukemia (ALL), acute myelogenous leukemia (AML), and chronic myelogenous leukemia (CML), as well as in some pediatric lymphomas and lymphoproliferative disorders. Most commonly, this abnormal activation occurs as a consequence of constitutive activation of AKT, providing a compelling rationale to target this pathway in many of these conditions. A variety of agents, beginning with the rapamycin analogue (rapalog) sirolimus, have been used successfully to target this pathway in a number of pediatric hematologic malignancies. Rapalogs demonstrate significant preclinical activity against ALL, which has led to a number of clinical trials. Moreover, rapalogs can synergize with a number of conventional cytotoxic agents and overcome pathways of chemotherapeutic resistance for drugs commonly used in ALL treatment, including methotrexate and corticosteroids. Based on preclinical data, rapalogs are also being studied in AML, CML, and non-Hodgkin’s lymphoma. Recently, significant progress has been made using rapalogs to treat pre-malignant lymphoproliferative disorders, including the autoimmune lymphoproliferative syndrome (ALPS); complete remissions in children with otherwise therapy-resistant disease have been seen. Rapalogs only block one component of the pathway (mTORC1), and newer agents are under preclinical and clinical development that can target different and often multiple protein kinases in the PI3K/AKT/mTOR pathway. Most of these agents have been tolerated in early-phase clinical trials. A number of PI3K inhibitors are under investigation. Of note, most of these also target other protein kinases. Newer agents are under development that target both m

  8. Association between insulin resistance and impairment of FGF21 signal transduction in skeletal muscles.

    PubMed

    Jeon, Ja Young; Choi, Sung-E; Ha, Eun Suk; Kim, Tae Ho; Jung, Jong Gab; Han, Seung Jin; Kim, Hae Jin; Kim, Dae Jung; Kang, Yup; Lee, Kwan-Woo

    2016-07-01

    Fibroblast growth factor (FGF) 21, was identified as a potent metabolic regulator of glucose and lipid metabolism. We investigated whether the levels and signalings of FGF21 changed in the skeletal muscle of type 2 diabetes mellitus (T2DM) patients, participants with impaired glucose tolerance (IGT), human skeletal muscle myotubes (HSMMs) under insulin-resistant conditions, and mice with diet-induced obesity (DIO). A percutaneous biopsy sample of the vastus lateralis muscle of T2DM patients, IGT subjects, and participants with normal glucose tolerance was obtained and the levels and signalings of FGF21 were assessed. We determined whether the expression and signalings of FGF21 in HSMMs altered according to palmitate concentrations and exposure time. Also, we confirmed whether changes of FGF21 signal transduction resulted in the alteration of FGF21 functions. DIO mice were treated intravenously with recombinant FGF21, and the levels and signalings of FGF21 were assessed in their soleus muscles. We checked whether or not FGF21 played a role in the gene transcription related to lipid oxidation. Levels of FGF21 increased, whereas levels of phosphorylated FGF receptor (p-FGFR), phosphorylated FGFR substrates 2α (p-FRS2α), and phosphorylated extracellular signal-regulated kinases (p-ERK) decreased in the skeletal muscle of both T2DM patients and IGT subjects. In vitro, palmitate increased the levels of FGF21 and significantly reduced the levels of β-klotho, p-FGFR, p-FRS2α, and p-ERK1/2 in HSMMs exposed to palmitate. Palmitate also decreased glucose uptake and glycogen contents of FGF21. Consistently, the levels of FGF21 were significantly higher and the levels of β-klotho and p-FGFR were lower in the DIO mice than in normal lean mice. The levels of FGF21 increased but its signal transduction and actions were impaired in skeletal muscles of T2DM patients, IGT subjects, in insulin-resistant HSMMs, and DIO mice. PMID:26758997

  9. An adaptive coarse graining method for signal transduction in three dimensions

    PubMed Central

    Archuleta, Michelle N.; McDermott, Jason E.; Edwards, Jeremy S.; Resat, Haluk

    2013-01-01

    The spatio-temporal landscape of the plasma membrane regulates activation and signal transduction of membrane bound receptors by restricting their two-dimensional mobility and by inducing receptor clustering. This regulation also extends to complex formation between receptors and adaptor proteins, which are the intermediate signaling molecules involved in cellular signaling that relay the received cues from cell surface to cytoplasm and eventually to the nucleus. Although their investigation poses challenging technical difficulties, there is a crucial need to understand the impact of the receptor diffusivity, clustering, and spatial heterogeneity, and of receptor-adaptor protein complex formation on the cellular signal transduction patterns. Building upon our earlier studies, we have developed an adaptive coarse-grained Monte Carlo method that can be used to investigate the role of diffusion, clustering and membrane corralling on receptor association and receptor-adaptor protein complex formation dynamics in three dimensions. The new Monte Carlo lattice based approach allowed us to introduce spatial resolution on the 2-D plasma membrane and to model the cytoplasm in three-dimensions. Being a multi-resolution approach, our new method makes it possible to represent various parts of the cellular system at different levels of detail and enabled us to utilize the locally homogeneous assumption when justified (e.g., cytoplasmic region away from the cell membrane) and avoid its use when high spatial resolution is needed (e.g., cell membrane and cytoplasmic region near the membrane) while keeping the required computational complexity manageable. Our results have shown that diffusion has a significant impact on receptor-receptor dimerization and receptor-adaptor protein complex formation kinetics. We have observed an “adaptor protein hopping” mechanism where the receptor binding proteins may hop between receptors to form short-lived transient complexes. This increased

  10. Low dose of IGF-I increases cell size of skeletal muscle satellite cells via Akt/S6K signaling pathway.

    PubMed

    Gao, Chun-qi; Zhi, Rui; Yang, Zhou; Li, Hai-chang; Yan, Hui-chao; Wang, Xiu-qi

    2015-11-01

    The objective of this study was to investigate the effect of insulin growth factor-I (IGF-I) on the size of pig skeletal muscle satellite cells (SCs). Using microarray, real-time RT-PCR, radioimmunoassay analysis and western blot, we first showed that supplementation of low-dose of IGF-I in culture medium resulted in enlarged cell size of Lantang SCs, only Akt and S6K were up-regulated at both the mRNA and protein levels among almost all of the mTOR pathway key genes, but had no effect on cell number. To elucidate the signaling mechanisms responsible for regulating cell size under low-dose of IGF-I treatment, we blocked Akt and S6K activity with the specific inhibitors MK2206 and PF4708671, respectively. Both inhibitors caused a decrease in cell size. In addition, MK2206 lowered the protein level of p-Akt (Ser473), p-S6K (Thr389), and p-rpS6 (Ser235/236), whereas PF4708671 lowered the protein level of p-S6K (Thr389) and p-rpS6 (Ser235/236). However, low dose of IGF-I didn't affect the protein level of p-mTOR (Ser2448) and p-mTOR (Ser2481). When both inhibitors were applied simultaneously, the effect was the same as that of the Akt inhibition alone. Taken together, we report for the first time that low-dose of IGF-I treatment increases cell size via Akt/S6K signaling pathway. PMID:25923195

  11. A novel computer simulation method for simulating the multiscale transduction dynamics of signal proteins

    NASA Astrophysics Data System (ADS)

    Peter, Emanuel; Dick, Bernhard; Baeurle, Stephan A.

    2012-03-01

    Signal proteins are able to adapt their response to a change in the environment, governing in this way a broad variety of important cellular processes in living systems. While conventional molecular-dynamics (MD) techniques can be used to explore the early signaling pathway of these protein systems at atomistic resolution, the high computational costs limit their usefulness for the elucidation of the multiscale transduction dynamics of most signaling processes, occurring on experimental timescales. To cope with the problem, we present in this paper a novel multiscale-modeling method, based on a combination of the kinetic Monte-Carlo- and MD-technique, and demonstrate its suitability for investigating the signaling behavior of the photoswitch light-oxygen-voltage-2-Jα domain from Avena Sativa (AsLOV2-Jα) and an AsLOV2-Jα-regulated photoactivable Rac1-GTPase (PA-Rac1), recently employed to control the motility of cancer cells through light stimulus. More specifically, we show that their signaling pathways begin with a residual re-arrangement and subsequent H-bond formation of amino acids near to the flavin-mononucleotide chromophore, causing a coupling between β-strands and subsequent detachment of a peripheral α-helix from the AsLOV2-domain. In the case of the PA-Rac1 system we find that this latter process induces the release of the AsLOV2-inhibitor from the switchII-activation site of the GTPase, enabling signal activation through effector-protein binding. These applications demonstrate that our approach reliably reproduces the signaling pathways of complex signal proteins, ranging from nanoseconds up to seconds at affordable computational costs.

  12. A novel computer simulation method for simulating the multiscale transduction dynamics of signal proteins.

    PubMed

    Peter, Emanuel; Dick, Bernhard; Baeurle, Stephan A

    2012-03-28

    Signal proteins are able to adapt their response to a change in the environment, governing in this way a broad variety of important cellular processes in living systems. While conventional molecular-dynamics (MD) techniques can be used to explore the early signaling pathway of these protein systems at atomistic resolution, the high computational costs limit their usefulness for the elucidation of the multiscale transduction dynamics of most signaling processes, occurring on experimental timescales. To cope with the problem, we present in this paper a novel multiscale-modeling method, based on a combination of the kinetic Monte-Carlo- and MD-technique, and demonstrate its suitability for investigating the signaling behavior of the photoswitch light-oxygen-voltage-2-Jα domain from Avena Sativa (AsLOV2-Jα) and an AsLOV2-Jα-regulated photoactivable Rac1-GTPase (PA-Rac1), recently employed to control the motility of cancer cells through light stimulus. More specifically, we show that their signaling pathways begin with a residual re-arrangement and subsequent H-bond formation of amino acids near to the flavin-mononucleotide chromophore, causing a coupling between β-strands and subsequent detachment of a peripheral α-helix from the AsLOV2-domain. In the case of the PA-Rac1 system we find that this latter process induces the release of the AsLOV2-inhibitor from the switchII-activation site of the GTPase, enabling signal activation through effector-protein binding. These applications demonstrate that our approach reliably reproduces the signaling pathways of complex signal proteins, ranging from nanoseconds up to seconds at affordable computational costs. PMID:22462840

  13. Signal transduction pathways leading to the production of IL-8 by human monocytes are differentially regulated by dexamethasone.

    PubMed Central

    Anttila, H S; Reitamo, S; Ceska, M; Hurme, M

    1992-01-01

    Previous studies have shown that IL-8 gene expression is enhanced by various stimuli, which induce different signal transduction pathways. A lipopolysaccharide (LPS)-induced pathway has been reported to be inhibited by glucocorticoids in monocytes. We have now examined the effect of dexamethasone on the LPS-induced and other signal transduction pathways leading to the production of IL-8 by human monocytes. Dexamethasone inhibited the production of IL-8 stimulated with a cyclic adenosine monophosphate analog or LPS. In contrast, dexamethasone had no significant effect on a phorbol ester (PMA)-stimulated IL-8 production. These results suggest that the signal transduction pathways leading to the production of IL-8 by human monocytes are differentially regulated by dexamethasone. PMID:1325308

  14. Twist plays an essential role in FGF and SHH signal transduction during mouse limb development.

    PubMed

    O'Rourke, Meredith P; Soo, Kenneth; Behringer, Richard R; Hui, Chi-Chung; Tam, Patrick P L

    2002-08-01

    Loss of Twist gene function arrests the growth of the limb bud shortly after its formation. In the Twist(-/-) forelimb bud, Fgf10 expression is reduced, Fgf4 is not expressed, and the domain of Fgf8 and Fgfr2 expression is altered. This is accompanied by disruption of the expression of genes (Shh, Gli1, Gli2, Gli3, and Ptch) associated with SHH signalling in the limb bud mesenchyme, the down-regulation of Bmp4 in the apical ectoderm, the absence of Alx3, Alx4, Pax1, and Pax3 activity in the mesenchyme, and a reduced potency of the limb bud tissues to differentiate into osteogenic and myogenic tissues. Development of the hindlimb buds in Twist(-/-) embryos is also retarded. The overall activity of genes involved in SHH signalling is reduced.Fgf4 and Fgf8 expression is lost or reduced in the apical ectoderm, but other genes (Fgf10, Fgfr2) involved with FGF signalling are expressed in normal patterns. Twist(+/-);Gli3(+/XtJ) mice display more severe polydactyly than that seen in either Twist(+/-) or Gli3(+/XtJ) mice, suggesting that there is genetic interaction between Twist and Gli3 activity. Twist activity is therefore essential for the growth and differentiation of the limb bud tissues as well as regulation of tissue patterning via the modulation of SHH and FGF signal transduction. PMID:12142027

  15. TCR Microclusters pre-exist and contain molecules necessary for TCR signal transduction.

    PubMed

    Crites, Travis J; Padhan, Kartika; Muller, James; Krogsgaard, Michelle; Gudla, Prabhakar R; Lockett, Stephen J; Varma, Rajat

    2014-07-01

    TCR-dependent signaling events have been observed to occur in TCR microclusters. We found that some TCR microclusters are present in unstimulated murine T cells, indicating that the mechanisms leading to microcluster formation do not require ligand binding. These pre-existing microclusters increase in absolute number following engagement by low-potency ligands. This increase is accompanied by an increase in cell spreading, with the result that the density of TCR microclusters on the surface of the T cell is not a strong function of ligand potency. In characterizing their composition, we observed a constant number of TCRs in a microcluster, constitutive exclusion of the phosphatase CD45, and preassociation with the signaling adapters linker for activation of T cells and growth factor receptor-bound protein 2. The existence of TCR microclusters prior to ligand binding in a state that is conducive for the initiation of downstream signaling could explain, in part, the rapid kinetics with which TCR signal transduction occurs. PMID:24860189

  16. Uropathogenic E. coli adhesin-induced host cell receptor conformational changes: implications in transmembrane signaling transduction

    PubMed Central

    Wang, Huaibin; Min, Guangwei; Glockshuber, Rudi; Sun, Tung-Tien; Kong, Xiang-Peng

    2009-01-01

    Urinary tract infection (UTI) is the second most common infectious disease, and is caused predominantly by type 1-fimbriated uropathogenic E. coli (UPEC). UPEC initiates infection by attaching to uroplakin Ia, its urothelial surface receptor, via the FimH adhesins capping the distal end of its fimbriae. Uroplakin Ia, together with uroplakins Ib, II and IIIa, forms a 16 nm receptor complex that is assembled into hexagonally packed two-dimensional crystals (urothelial plaques) covering >90% of the urothelial apical surface. Recent studies indicate that FimH is the invasin of UPEC as its attachment to the urothelial surface can induce cellular signaling events including calcium elevation and the phosphorylation of the uroplakin IIIa cytoplasmic tail, leading to cytoskeletal rearrangements and bacterial invasion. However, it remains unknown how the binding of FimH to the uroplakin receptor triggers a signal that can be transmitted through the highly impermeable urothelial apical membrane. We show here by cryo-electron microscopy that FimH-binding to the extracellular domain of UPIa induces global conformational changes in the entire uroplakin receptor complex, including a coordinated movement of the tightly bundled transmembrane helices. This movement of the transmembrane helix bundles can cause a corresponding lateral translocation of the uroplakin cytoplasmic tails, which can be sufficient to trigger downstream signaling events. Our results suggest a novel pathogen-induced transmembrane signal transduction mechanism that plays a key role in the initial stages of UPEC invasion and receptor-mediated bacterial invasion in general. PMID:19577575

  17. Role of peroxynitrite in the redox regulation of cell signal transduction pathways

    PubMed Central

    Liaudet, Lucas; Vassalli, Giuseppe; Pacher, Pal

    2009-01-01

    Peroxynitrite is a potent oxidant and nitrating species formed from the reaction between the free radicals nitric oxide and superoxide. An excessive formation of peroxynitrite represents an important mechanism contributing to cell death and dysfunction in multiple cardiovascular pathologies, such as myocardial infarction, heart failure and atherosclerosis. Whereas initial works focused on direct oxidative biomolecular damage as the main route of peroxynitrite toxicity, more recent evidence, mainly obtained in vitro, indicates that peroxynitrite also behaves as a potent modulator of various cell signal transduction pathways. Due to its ability to nitrate tyrosine residues, peroxynitrite affects cellular processes dependent on tyrosine phosphorylation. Peroxynitrite also exerts complex effects on the activity of various kinases and phosphatases, resulting in the up- or downregulation of signalling cascades, in a concentration- and cell-dependent manner. Such roles of peroxynitrite in the redox regulation of key signalling pathways for cardiovascular homeostasis, including protein kinase B and C, the MAP kinases, Nuclear Factor Kappa B, as well as signalling dependent on insulin and the sympatho-adrenergic system are presented in detail in this review. PMID:19273391

  18. Cadmium Induces Apoptosis in Freshwater Crab Sinopotamon henanense through Activating Calcium Signal Transduction Pathway

    PubMed Central

    Wang, Jinxiang; Zhang, Pingping; Liu, Na; Wang, Qian; Luo, Jixian; Wang, Lan

    2015-01-01

    Calcium ion (Ca2+) is one of the key intracellular signals, which is implicated in the regulation of cell functions such as impregnation, cell proliferation, differentiation and death. Cadmium (Cd) is a toxic environmental pollutant that can disturb cell functions and even lead to cell death. Recently, we have found that Cd induced apoptosis in gill cells of the freshwater crab Sinopotamon henanense via caspase activation. In the present study, we further investigated the role of calcium signaling in the Cd-induced apoptosis in the animals. Our data showed that Cd triggered gill cell apoptosis which is evidenced by apoptotic DNA fragmentation, activations of caspases-3, -8 and -9 and the presence of apoptotic morphological features. Moreover, Cd elevated the intracellular concentration of Ca2+, the protein concentration of calmodulin (CaM) and the activity of Ca2+-ATPase in the gill cells of the crabs. Pretreatment of the animals with ethylene glycol-bis-(b-aminoethyl ether)-N,N,N’,N’-tetraacetic acid (EGTA), Ca2+ chelator, inhibited Cd-induced activation of caspases-3, -8 and -9 as well as blocked the Cd-triggered apoptotic DNA fragmentation. The apoptotic morphological features were no longer observed in gill cells pretreated with the Ca2+ signaling inhibitors before Cd treatment. Our results indicate that Cd evokes gill cell apoptosis through activating Ca2+-CaM signaling transduction pathway. PMID:26714174

  19. CYLD negatively regulates transforming growth factor-β-signalling via deubiquitinating Akt

    PubMed Central

    Lim, Jae Hyang; Jono, Hirofumi; Komatsu, Kensei; Woo, Chang-Hoon; Lee, Jiyun; Miyata, Masanori; Matsuno, Takashi; Xu, Xiangbin; Huang, Yuxian; Zhang, Wenhong; Park, Soo Hyun; Kim, Yu-Il; Choi, Yoo-Duk; Shen, Huahao; Heo, Kyung-Sun; Xu, Haodong; Bourne, Patricia; Koga, Tomoaki; Xu, Haidong; Yan, Chen; Wang, Binghe; Chen, Lin-Feng; Feng, Xin-Hua; Li, Jian-Dong

    2012-01-01

    Lung injury, whether induced by infection or caustic chemicals, initiates a series of complex wound-healing responses. If uncontrolled, these responses may lead to fibrotic lung diseases and loss of function. Thus, resolution of lung injury must be tightly regulated. The key regulatory proteins required for tightly controlling the resolution of lung injury have yet to be identified. Here we show that loss of deubiquitinase CYLD led to the development of lung fibrosis in mice after infection with Streptococcus pneumoniae. CYLD inhibited transforming growth factor-β-signalling and prevented lung fibrosis by decreasing the stability of Smad3 in an E3 ligase carboxy terminus of Hsc70-interacting protein-dependent manner. Moreover, CYLD decreases Smad3 stability by deubiquitinating K63-polyubiquitinated Akt. Together, our results unveil a role for CYLD in tightly regulating the resolution of lung injury and preventing fibrosis by deubiquitinating Akt. These studies may help develop new therapeutic strategies for preventing lung fibrosis. PMID:22491319

  20. Matrix metalloproteinase 14 modulates signal transduction and angiogenesis in the cornea.

    PubMed

    Chang, Jin-Hong; Huang, Yu-Hui; Cunningham, Christy M; Han, Kyu-Yeon; Chang, Michael; Seiki, Motoharu; Zhou, Zhongjun; Azar, Dimitri T

    2016-01-01

    The cornea is transparent and avascular, and retention of these characteristics is critical to maintaining vision clarity. Under normal conditions, wound healing in response to corneal injury occurs without the formation of new blood vessels; however, neovascularization may be induced during corneal wound healing when the balance between proangiogenic and antiangiogenic mediators is disrupted to favor angiogenesis. Matrix metalloproteinases (MMPs), which are key factors in extracellular matrix remodeling and angiogenesis, contribute to the maintenance of this balance, and in pathologic instances, can contribute to its disruption. Here, we elaborate on the facilitative role of MMPs, specifically MMP-14, in corneal neovascularization. MMP-14 is a transmembrane MMP that is critically involved in extracellular matrix proteolysis, exosome transport, and cellular migration and invasion, processes that are critical for angiogenesis. To aid in developing efficacious therapies that promote healing without neovascularization, it is important to understand and further investigate the complex pathways related to MMP-14 signaling, which can also involve vascular endothelial growth factor, basic fibroblast growth factor, Wnt/β-catenin, transforming growth factor, platelet-derived growth factor, hepatocyte growth factor or chemokines, epidermal growth factor, prostaglandin E2, thrombin, integrins, Notch, Toll-like receptors, PI3k/Akt, Src, RhoA/RhoA kinase, and extracellular signal-related kinase. The involvement and potential contribution of these signaling molecules or proteins in neovascularization are the focus of the present review. PMID:26647161

  1. Cysteines in the neuropilin-2 MAM domain modulate receptor homooligomerization and signal transduction.

    PubMed

    Barton, Rachael; Driscoll, Alyssa; Flores, Samuel; Mudbhari, Durlav; Collins, Theresa; Iovine, M Kathryn; Berger, Bryan W

    2015-07-01

    Neuropilins (NRPs) are transmembrane receptors involved in angiogenesis, lymphangiogenesis, and neuronal development as well as in cancer metastasis. Previous studies suggest that NRPs exist in heteromeric complexes with vascular endothelial growth factors (VEGFs) and VEGF receptors as well as plexins and semaphorins. We determined via site-directed mutagenesis and bioluminescent resonance energy transfer assays that a conserved cysteine (C711) in the Danio rerio NRP2a MAM (meprin, A-5 protein, and protein tyrosine phosphatase μ) domain modulates NRP2a homomeric interactions. Mutation of this residue also disrupts semaphorin-3F binding in NRP2a-transfected COS-7 cells and prevents the NRP2a overexpression effects in a zebrafish vascular model. Collectively, our results indicate the MAM domain plays an important role in defining the NRP2 homodimer structure, which is important for semaphorin-dependent signal transduction via NRP2. PMID:25656526

  2. Novel optical methodologies in studying mechanical signal transduction in mammalian cells

    NASA Technical Reports Server (NTRS)

    Stamatas, G. N.; McIntire, L. V.

    1999-01-01

    For the last 3 decades evidence has been accumulating that some types of mammalian cells respond to their mechanically active environment by altering their morphology, growth rate, and metabolism. The study of such responses is very important in understanding, physiological and pathological conditions ranging from bone formation to atherosclerosis. Obtaining this knowledge has been the goal for an active research area in bioengineering termed cell mechanotransduction. The advancement of optical methodologies used in cell biology research has given the tools to elucidate cellular mechanisms that would otherwise be impossible to visualize. Combined with molecular biology techniques, they give engineers invaluable tools in understanding the chemical pathways involved in mechanotransduction. Herein we briefly review the current knowledge on mechanical signal transduction in mammalian cells, focusing on the application of novel optical techniques in the ongoing research.

  3. Lessons in Fundamental Mechanisms and Diverse Adaptations from the 2015 Bacterial Locomotion and Signal Transduction Meeting

    PubMed Central

    Prüβ, Birgit M.; Liu, Jun; Higgs, Penelope I.

    2015-01-01

    In response to rapid changes in their environment, bacteria control a number of processes, including motility, cell division, biofilm formation, and virulence. Research presented in January 2015 at the biennial Bacterial Locomotion and Signal Transduction (BLAST) meeting in Tucson, AZ, illustrates the elegant complexity of the nanoarrays, nanomachines, and networks of interacting proteins that mediate such processes. Studies employing an array of biophysical, genetic, cell biology, and mathematical methods are providing an increasingly detailed understanding of the mechanisms of these systems within well-studied bacteria. Furthermore, comparisons of these processes in diverse bacterial species are providing insight into novel regulatory and functional mechanisms. This review summarizes research presented at the BLAST meeting on these fundamental mechanisms and diverse adaptations, including findings of importance for applications involving bacteria of medical or agricultural relevance. PMID:26195592

  4. Smartphone Operated Signal Transduction by Ion Nanogating (STING) Amplifier for Nanopore Sensors: Design and Analytical Application

    PubMed Central

    Özel, Rıfat Emrah; Kahnemouyi, Sina; Fan, Hsinwen; Mak, Wai Han; Lohith, Akshar; Seger, Adam; Teodorescu, Mircea; Pourmand, Nader

    2016-01-01

    In this report, we demonstrated a handheld wireless voltage-clamp amplifier for current measurement of nanopore sensors. This amplifier interfaces a sensing probe and connects wirelessly with a computer or smartphone for the required stimulus input, data processing and storage. To test the proposed Signal Transduction by Ion Nanogating (STING) wireless amplifier, in the current study the system was tested with a nano-pH sensor to measure pH of standard buffer solutions and the performance was compared against the commercial voltage-clamp amplifier. To our best knowledge, STING amplifier is the first miniaturized wireless voltage-clamp platform operated with a customized smart-phone application (app). PMID:27602408

  5. Bead Assembly Magnetorotation as a Signal Transduction Method for Protein Detection

    PubMed Central

    Hecht, Ariel; Commiskey, Patrick; Shah, Nicholas; Kopelman, Raoul

    2013-01-01

    This paper demonstrates a proof-of-principle for a new signal transduction method for protein detection called Bead Assembly Magnetorotation (BAM). In this paper, we chose to focus on the protein thrombin, a popular choice for proof-of-principle work in this field. BAM is based on using the protein target to mediate the formation of aptamer-coated 1 μm magnetic beads into a bead assembly, formed at the bottom of a 1 μL hanging droplet. The size, shape and fractal dimension of this bead assembly all depend on the protein concentration. The protein concentration can be measured in two ways: by magnetorotation, in which the rotational period of the assembly correlates with the protein concentration, or by fractal analysis. Additionally, a microscope-free magnetorotation detection method is introduced, based on a simple laser apparatus built from standard laboratory components. PMID:23639345

  6. Comprehensive analysis of signal transduction in three-dimensional ECM-based tumor cell cultures

    PubMed Central

    Eke, Iris; Hehlgans, Stephanie; Zong, Yaping; Cordes, Nils

    2015-01-01

    Analysis of signal transduction and protein phosphorylation is fundamental to understanding physiological and pathological cell behavior and identifying novel therapeutic targets. Despite the fact that the use of physiological three-dimensional cell culture assays is increasing, 3D proteomics and phosphoproteomics remain challenging due to difficulties with easy, robust and reproducible sample preparation. Here, we present an easy-to-perform, reliable and time-efficient method for the production of 3D cell lysates that does not compromise cell adhesion before cell lysis. The samples can be used for western blotting as well as phosphoproteome array technology. This technique will be of interest for researchers working in all fields of biology and drug development. PMID:26618185

  7. Phase separation of signaling molecules promotes T cell receptor signal transduction.

    PubMed

    Su, Xiaolei; Ditlev, Jonathon A; Hui, Enfu; Xing, Wenmin; Banjade, Sudeep; Okrut, Julia; King, David S; Taunton, Jack; Rosen, Michael K; Vale, Ronald D

    2016-04-29

    Activation of various cell surface receptors triggers the reorganization of downstream signaling molecules into micrometer- or submicrometer-sized clusters. However, the functional consequences of such clustering have been unclear. We biochemically reconstituted a 12-component signaling pathway on model membranes, beginning with T cell receptor (TCR) activation and ending with actin assembly. When TCR phosphorylation was triggered, downstream signaling proteins spontaneously separated into liquid-like clusters that promoted signaling outputs both in vitro and in human Jurkat T cells. Reconstituted clusters were enriched in kinases but excluded phosphatases and enhanced actin filament assembly by recruiting and organizing actin regulators. These results demonstrate that protein phase separation can create a distinct physical and biochemical compartment that facilitates signaling. PMID:27056844

  8. Role of Cdc42p in Pheromone-Stimulated Signal Transduction in Saccharomyces cerevisiae

    PubMed Central

    Moskow, John J.; Gladfelter, Amy S.; Lamson, Rachel E.; Pryciak, Peter M.; Lew, Daniel J.

    2000-01-01

    CDC42 encodes a highly conserved GTPase of the Rho family that is best known for its role in regulating cell polarity and actin organization. In addition, various studies of both yeast and mammalian cells have suggested that Cdc42p, through its interaction with p21-activated kinases (PAKs), plays a role in signaling pathways that regulate target gene transcription. However, recent studies of the yeast pheromone response pathway suggested that prior results with temperature-sensitive cdc42 mutants were misleading and that Cdc42p and the Cdc42p-PAK interaction are not involved in signaling. To clarify this issue, we have identified and characterized novel viable pheromone-resistant cdc42 alleles that retain the ability to perform polarity-related functions. Mutation of the Cdc42p residue Val36 or Tyr40 caused defects in pheromone signaling and in the localization of the Ste20p PAK in vivo and affected binding to the Ste20p Cdc42p-Rac interactive binding (CRIB) domain in vitro. Epistasis analysis suggested that they affect the signaling step at which Ste20p acts, and overproduction of Ste20p rescued the defect. These results suggest that Cdc42p is in fact required for pheromone response and that interaction with the PAK Ste20p is critical for that role. Furthermore, the ste20ΔCRIB allele, previously used to disrupt the Cdc42p-Ste20p interaction, behaved as an activated allele, largely bypassing the signaling defect of the cdc42 mutants. Additional observations lead us to suggest that Cdc42p collaborates with the SH3-domain protein Bem1p to facilitate signal transduction, possibly by providing a cell surface scaffold that aids in the local concentration of signaling kinases, thus promoting activation of a mitogen-activated protein kinase cascade by Ste20p. PMID:11003652

  9. Role of Cdc42p in pheromone-stimulated signal transduction in Saccharomyces cerevisiae.

    PubMed

    Moskow, J J; Gladfelter, A S; Lamson, R E; Pryciak, P M; Lew, D J

    2000-10-01

    CDC42 encodes a highly conserved GTPase of the Rho family that is best known for its role in regulating cell polarity and actin organization. In addition, various studies of both yeast and mammalian cells have suggested that Cdc42p, through its interaction with p21-activated kinases (PAKs), plays a role in signaling pathways that regulate target gene transcription. However, recent studies of the yeast pheromone response pathway suggested that prior results with temperature-sensitive cdc42 mutants were misleading and that Cdc42p and the Cdc42p-PAK interaction are not involved in signaling. To clarify this issue, we have identified and characterized novel viable pheromone-resistant cdc42 alleles that retain the ability to perform polarity-related functions. Mutation of the Cdc42p residue Val36 or Tyr40 caused defects in pheromone signaling and in the localization of the Ste20p PAK in vivo and affected binding to the Ste20p Cdc42p-Rac interactive binding (CRIB) domain in vitro. Epistasis analysis suggested that they affect the signaling step at which Ste20p acts, and overproduction of Ste20p rescued the defect. These results suggest that Cdc42p is in fact required for pheromone response and that interaction with the PAK Ste20p is critical for that role. Furthermore, the ste20DeltaCRIB allele, previously used to disrupt the Cdc42p-Ste20p interaction, behaved as an activated allele, largely bypassing the signaling defect of the cdc42 mutants. Additional observations lead us to suggest that Cdc42p collaborates with the SH3-domain protein Bem1p to facilitate signal transduction, possibly by providing a cell surface scaffold that aids in the local concentration of signaling kinases, thus promoting activation of a mitogen-activated protein kinase cascade by Ste20p. PMID:11003652

  10. Syntrophin proteins as Santa Claus: role(s) in cell signal transduction.

    PubMed

    Bhat, Hina F; Adams, Marvin E; Khanday, Firdous A

    2013-07-01

    Syntrophins are a family of cytoplasmic membrane-associated adaptor proteins, characterized by the presence of a unique domain organization comprised of a C-terminal syntrophin unique (SU) domain and an N-terminal pleckstrin homology (PH) domain that is split by insertion of a PDZ domain. Syntrophins have been recognized as an important component of many signaling events, and they seem to function more like the cell's own personal 'Santa Claus' that serves to 'gift' various signaling complexes with precise proteins that they 'wish for', and at the same time care enough for the spatial, temporal control of these signaling events, maintaining overall smooth functioning and general happiness of the cell. Syntrophins not only associate various ion channels and signaling proteins to the dystrophin-associated protein complex (DAPC), via a direct interaction with dystrophin protein but also serve as a link between the extracellular matrix and the intracellular downstream targets and cell cytoskeleton by interacting with F-actin. They play an important role in regulating the postsynaptic signal transduction, sarcolemmal localization of nNOS, EphA4 signaling at the neuromuscular junction, and G-protein mediated signaling. In our previous work, we reported a differential expression pattern of alpha-1-syntrophin (SNTA1) protein in esophageal and breast carcinomas. Implicated in several other pathologies, like cardiac dys-functioning, muscular dystrophies, diabetes, etc., these proteins provide a lot of scope for further studies. The present review focuses on the role of syntrophins in membrane targeting and regulation of cellular proteins, while highlighting their relevance in possible development and/or progression of pathologies including cancer which we have recently demonstrated. PMID:23263165

  11. GDF15 regulates Kv2.1-mediated outward K+ current through the Akt/mTOR signalling pathway in rat cerebellar granule cells

    PubMed Central

    Wang, Chang-Ying; Huang, An-Qi; Zhou, Meng-Hua; Mei, Yan-Ai

    2014-01-01

    GDF15 (growth/differentiation factor 15), a novel member of the TGFβ (transforming growth factor β) superfamily, plays critical roles in the central and peripheral nervous systems, but the signal transduction pathways and receptor subtypes involved are not well understood. In the present paper, we report that GDF15 specifically increases the IK (delayed-rectifier outward K+ current) in rat CGNs (cerebellar granule neurons) in time- and concentration-dependent manners. The GDF15-induced amplification of the IK is mediated by the increased expression and reduced lysosome-dependent degradation of the Kv2.1 protein, the main α-subunit of the IK channel. Exposure of CGNs to GDF15 markedly induced the phosphorylation of ERK (extracellular-signal-regulated kinase), Akt and mTOR (mammalian target of rapamycin), but the GDF15-induced IK densities and increased expression of Kv2.1 were attenuated only by Akt and mTOR, and not ERK, inhibitors. Pharmacological inhibition of the Src-mediated phosphorylation of TGFβR2 (TGFβ receptor 2), not TGFβR1, abrogated the effect of GDF15 on IK amplification and Kv2.1 induction. Immunoprecipitation assays showed that GDF15 increased the tyrosine phosphorylation of TGFβRII in the CGN lysate. The results of the present study reveal a novel regulation of Kv2.1 by GDF15 mediated through the TGFβRII-activated Akt/mTOR pathway, which is a previously uncharacterized Smad-independent mechanism of GDF15 signalling. PMID:24597762

  12. Gibberellin biosynthesis and signal transduction is essential for internode elongation in deepwater rice

    PubMed Central

    Ayano, Madoka; Kani, Takahiro; Kojima, Mikiko; Sakakibara, Hitoshi; Kitaoka, Takuya; Kuroha, Takeshi; Angeles-Shim, Rosalyn B; Kitano, Hidemi; Nagai, Keisuke; Ashikari, Motoyuki

    2014-01-01

    Under flooded conditions, the leaves and internodes of deepwater rice can elongate above the water surface to capture oxygen and prevent drowning. Our previous studies showed that three major quantitative trait loci (QTL) regulate deepwater-dependent internode elongation in deepwater rice. In this study, we investigated the age-dependent internode elongation in deepwater rice. We also investigated the relationship between deepwater-dependent internode elongation and the phytohormone gibberellin (GA) by physiological and genetic approach using a QTL pyramiding line (NIL-1 + 3 + 12). Deepwater rice did not show internode elongation before the sixth leaf stage under deepwater condition. Additionally, deepwater-dependent internode elongation occurred on the sixth and seventh internodes during the sixth leaf stage. These results indicate that deepwater rice could not start internode elongation until the sixth leaf stage. Ultra-performance liquid chromatography tandem mass-spectrometry (UPLC-MS/MS) method for the phytohormone contents showed a deepwater-dependent GA1 and GA4 accumulation in deepwater rice. Additionally, a GA inhibitor abolished deepwater-dependent internode elongation in deepwater rice. On the contrary, GA feeding mimicked internode elongation under ordinary growth conditions. However, mutations in GA biosynthesis and signal transduction genes blocked deepwater-dependent internode elongation. These data suggested that GA biosynthesis and signal transduction are essential for deepwater-dependent internode elongation in deepwater rice. Deepwater rice obtained the ability for rapid internode elongation to avoid drowning and adapt to flooded condition. How does it regulate internode elongation? Using both physiological and genetic approach, this paper shows that the plant hormone, gibberellin (GA) regulates internode elongation. PMID:24891164

  13. Signal transduction for taurocholic acid in the olfactory system of Atlantic salmon.

    PubMed

    Lo, Y H; Bellis, S L; Cheng, L J; Pang, J; Bradley, T M; Rhoads, D E

    1994-10-01

    Conjugated bile acids such as taurocholic acid (TChA) are potent olfactory stimuli for Atlantic salmon (Salmo salar). A plasma membrane rich fraction was derived from salmon olfactory rosettes and used to investigate TChA signal transduction and receptor binding. In the presence of GTP gamma S, TChA caused dose-dependent stimulation of phosphatidylinositol 4,5-bisphosphate (PIP2) breakdown, half maximal at less than 10(-7) M TChA. Stimulation of PIP2 breakdown by TChA required GTP gamma S, was blocked by GDP beta S, and was mimicked by A1F4-, consistent with a G protein requirement. A1F4- and Ca2+ stimulated breakdown of PIP2, but not phosphatidylcholine, arguing against a non-specific lipase activation. Stimulation of PIP2 breakdown by TChA was maximal at low Ca2+ concentration, < or = 10 nM. Conventional binding analysis with 3H-TChA was inconclusive due to a high degree of non-specific binding and to lack of tissue specificity expected for an olfactory receptor. Analysis of odorant amino acid binding indicated possible interaction of TChA with a putative acidic amino acid receptor but no interaction of TChA with a putative neutral amino acid receptor. We conclude that olfactory discrimination between amino acids and bile acids occurs in part at the receptor level while both classes of odors appear to use the same signal transduction mechanism, G protein mediated activation of phosphoinositide specific phospholipase C (PLC). PMID:7881971

  14. Sensor-response regulator interactions in a cross-regulated signal transduction network.

    PubMed

    Huynh, TuAnh Ngoc; Chen, Li-Ling; Stewart, Valley

    2015-07-01

    Two-component signal transduction involves phosphoryl transfer between a histidine kinase sensor and a response regulator effector. The nitrate-responsive two-component signal transduction systems in Escherichia coli represent a paradigm for a cross-regulation network, in which the paralogous sensor-response regulator pairs, NarX-NarL and NarQ-NarP, exhibit both cognate (e.g. NarX-NarL) and non-cognate (e.g. NarQ-NarL) interactions to control output. Here, we describe results from bacterial adenylate cyclase two-hybrid (BACTH) analysis to examine sensor dimerization as well as interaction between sensor-response regulator cognate and non-cognate pairs. Although results from BACTH analysis indicated that the NarX and NarQ sensors interact with each other, results from intragenic complementation tests demonstrate that they do not form functional heterodimers. Additionally, intragenic complementation shows that both NarX and NarQ undergo intermolecular autophosphorylation, deviating from the previously reported correlation between DHp (dimerization and histidyl phosphotransfer) domain loop handedness and autophosphorylation mode. Results from BACTH analysis revealed robust interactions for the NarX-NarL, NarQ-NarL and NarQ-NarP pairs but a much weaker interaction for the NarX-NarP pair. This demonstrates that asymmetrical cross-regulation results from differential binding affinities between different sensor-regulator pairs. Finally, results indicate that the NarL effector (DNA-binding) domain inhibits NarX-NarL interaction. Missense substitutions at receiver domain residue Ser-80 enhanced NarX-NarL interaction, apparently by destabilizing the NarL receiver-effector domain interface. PMID:25873583

  15. Nonselective block by La3+ of Arabidopsis ion channels involved in signal transduction

    NASA Technical Reports Server (NTRS)

    Lewis, B. D.; Spalding, E. P.; Evans, M. L. (Principal Investigator)

    1998-01-01

    Lanthanide ions such as La3+ are frequently used as blockers to test the involvement of calcium channels in plant and animal signal transduction pathways. For example, the large rise in cytoplasmic Ca2+ concentration triggered by cold shock in Arabidopsis seedlings is effectively blocked by 10 mM La3+ and we show here that the simultaneous large membrane depolarization is similarly blocked. However, a pharmacological tool is only as useful as it is selective and the specificity of La3+ for calcium channels was brought into question by our finding that it also blocked a blue light (BL)-induced depolarization that results from anion channel activation and believed not to involve calcium channels. This unexpected inhibitory effect of La3+ on the BL-induced depolarization is explained by our finding that 10 mM La3+ directly and completely blocked the BL-activated anion channel when applied to excised patches. We have investigated the ability of La3+ to block noncalcium channels in Arabidopsis. In addition to the BL-activated anion channel, 10 mM La3+ blocked a cation channel and a stretch-activated channel in patches of plasma membrane excised from hypocotyl cells. In root cells, 10 mM La3+ inhibited the activity of an outward-rectifying potassium channel at the whole cell and single-channel level by 47% and 58%, respectively. We conclude that La3+ is a nonspecific blocker of multiple ionic conductances in Arabidopsis and may disrupt signal transduction processes independently of any effect on Ca2+ channels.

  16. Manumycin inhibits ras signal transduction pathway and induces apoptosis in COLO320-DM human colon tumourcells

    PubMed Central

    Paolo, A Di; Danesi, R; Nardini, D; Bocci, G; Innocenti, F; Fogli, S; Barachini, S; Marchetti, A; Bevilacqua, G; Tacca, M Del

    2000-01-01

    The aim of the present study was to assess the cytotoxicity of manumycin, a specific inhibitor of farnesyl:protein transferase, as well as its effects on protein isoprenylation and kinase-dependent signal transduction in COLO320-DM human colon adenocarcinoma which harbours a wild-type K- ras gene. Immunoblot analysis of isolated cell membranes and total cellular lysates of COLO320-DM cells demonstrated that manumycin dose-dependently reduced p21 ras farnesylation with a 50% inhibitory concentration (IC50) of 2.51 ± 0.11 μM and 2.68 ± 0.20 μM, respectively, while the geranylgeranylation of p21 rhoA and p21 rap1 was not affected. Manumycin dose-dependently inhibited (IC50= 2.40 ± 0.67 μM) the phosphorylation of the mitogen-activated protein kinase/extracellular-regulated kinase 2 (p42MAPK/ERK2), the main cytoplasmic effector of p21 ras, as well as COLO320-DM cell growth (IC50= 3.58 ± 0.27 μM) without affecting the biosynthesis of cholesterol. Mevalonic acid (MVA, 100 μM), a substrate of the isoprenoid synthesis, was unable to protect COLO320-DM cells from manumycin cytotoxicity. Finally, manumycin 1–25 μM for 24–72 h induced oligonucleosomal fragmentation in a dose- and time-dependent manner and MVA did not protect COLO320-DM cells from undergoing DNA cleavage. The present findings indicate that the inhibition of p21 ras processing and signal transduction by manumycin is associated with marked inhibition of cell proliferation and apoptosis in colon cancer cells and the effect on cell growth does not require the presence of a mutated ras gene for maximal expression of chemotherapeutic activity. © 2000 Cancer Research Campaign PMID:10732765

  17. Chemotactic signal transduction and phosphate metabolism as adaptive strategies during citrus canker induction by Xanthomonas citri.

    PubMed

    Moreira, Leandro Marcio; Facincani, Agda Paula; Ferreira, Cristiano Barbalho; Ferreira, Rafael Marine; Ferro, Maria Inês Tiraboshi; Gozzo, Fabio Cesar; de Oliveira, Julio Cezar Franco; Ferro, Jesus Aparecido; Soares, Márcia Regina

    2015-03-01

    The genome of Xanthomonas citri subsp. Citri strain 306 pathotype A (Xac) was completely sequenced more than 10 years; to date, few studies involving functional genomics Xac and its host compatible have been developed, specially related to adaptive events that allow the survival of Xac within the plant. Proteomic analysis of Xac showed that the processes of chemotactic signal transduction and phosphate metabolism are key adaptive strategies during the interaction of a pathogenic bacterium with its plant host. The results also indicate the importance of a group of proteins that may not be directly related to the classical virulence factors, but that are likely fundamental to the success of the initial stages of the infection, such as methyl-accepting chemotaxis protein (Mcp) and phosphate specific transport (Pst). Furthermore, the analysis of the mutant of the gene pstB which codifies to an ABC phosphate transporter subunit revealed a complete absence of citrus canker symptoms when inoculated in compatible hosts. We also conducted an in silico analysis which established the possible network of genes regulated by two-component systems PhoPQ and PhoBR (related to phosphate metabolism), and possible transcriptional factor binding site (TFBS) motifs of regulatory proteins PhoB and PhoP, detaching high degree of conservation of PhoB TFBS in 84 genes of Xac genome. This is the first time that chemotaxis signal transduction and phosphate metabolism were therefore indicated to be fundamental to the process of colonization of plant tissue during the induction of disease associated with Xanthomonas genus bacteria. PMID:25403594

  18. Apelin/APJ signaling promotes hypoxia-induced proliferation of endothelial progenitor cells via phosphoinositide-3 kinase/Akt signaling.

    PubMed

    Zhang, Jingchang; Liu, Qiming; Hu, Xinqun; Fang, Zhenfei; Huang, Feng; Tang, Liang; Zhou, Shenghua

    2015-09-01

    Endothelial progenitor cells (EPCs) can adhere to the endothelium at sites of hypoxia/ischemia and participate in the formation of novel vessels through differentiating into endothelial cells (ECs). Apelin is an endogenous ligand for the G protein‑coupled receptor APJ, and apelin/APJ signaling has a role in cardiovascular function. The present study aimed to investigate the role of apelin/APJ signaling in the regulation of EPC proliferation under hypoxia. The results showed that hypoxia was able to induce EPC proliferation, accompanied with an upregulation of hypoxia‑inducible factor (HIF)‑1α as well as apelin/APJ signaling. Further investigation indicated that siRNA‑mediated knockdown of apelin or APJ expression attenuated the hypoxia‑induced proliferation of EPCs, suggesting that apelin/APJ signaling has an important role in hypoxia‑induced EPC proliferation. Moreover, the phosphoinositide‑3 kinase (PI3K)/Akt signaling pathway was found to be involved in the apelin/APJ‑mediated EPC proliferation under hypoxia. Based on these findings, the present study suggested that hypoxia‑induced upregulation of HIF‑1α promotes the expression of apelin and APJ, which further activate the downstream PI3K/Akt signaling pathway, a key promoter of EPC proliferation. In conclusion, the present study highlighted the role of apelin/APJ in the regulation of EPC proliferation, and apelin/APJ may therefore serve as a potential target for the prevention of hypoxic ischemic injury. PMID:26018184

  19. The cellular response to vascular endothelial growth factors requires co-ordinated signal transduction, trafficking and proteolysis

    PubMed Central

    Smith, Gina A.; Fearnley, Gareth W.; Tomlinson, Darren C.; Harrison, Michael A.; Ponnambalam, Sreenivasan

    2015-01-01

    VEGFs (vascular endothelial growth factors) are a family of conserved disulfide-linked soluble secretory glycoproteins found in higher eukaryotes. VEGFs mediate a wide range of responses in different tissues including metabolic homoeostasis, cell proliferation, migration and tubulogenesis. Such responses are initiated by VEGF binding to soluble and membrane-bound VEGFRs (VEGF receptor tyrosine kinases) and co-receptors. VEGF and receptor splice isoform diversity further enhances complexity of membrane protein assembly and function in signal transduction pathways that control multiple cellular responses. Different signal transduction pathways are simultaneously activated by VEGFR–VEGF complexes with membrane trafficking along the endosome–lysosome network further modulating signal output from multiple enzymatic events associated with such pathways. Balancing VEGFR–VEGF signal transduction with trafficking and proteolysis is essential in controlling the intensity and duration of different intracellular signalling events. Dysfunction in VEGF-regulated signal transduction is important in chronic disease states including cancer, atherosclerosis and blindness. This family of growth factors and receptors is an important model system for understanding human disease pathology and developing new therapeutics for treating such ailments. PMID:26285805

  20. MicroRNA-145 suppresses hepatocellular carcinoma by targeting IRS1 and its downstream Akt signaling

    SciTech Connect

    Wang, Yelin; Hu, Chen; Cheng, Jun; Chen, Binquan; Ke, Qinghong; Lv, Zhen; Wu, Jian; Zhou, Yanfeng

    2014-04-18

    Highlights: • MiR-145 expression is down-regulated in HCC tissues and inversely related with IRS1 levels. • MiR-145 directly targets IRS1 in HCC cells. • Restored expression of miR-145 suppressed HCC cell proliferation and growth. • MiR-145 induced IRS1 under-expression potentially reduced downstream AKT signaling. - Abstract: Accumulating evidences have proved that dysregulation of microRNAs (miRNAs) is involved in cancer initiation and progression. In this study, we showed that miRNA-145 level was significantly decreased in hepatocellular cancer (HCC) tissues and cell lines, and its low expression was inversely associated with the abundance of insulin receptor substrate 1 (IRS1), a key mediator in oncogenic insulin-like growth factor (IGF) signaling. We verified IRS1 as a direct target of miR-145 using Western blotting and luciferase reporter assay. Further, the restoration of miR-145 in HCC cell lines suppressed cancer cell growth, owing to down-regulated IRS1 expression and its downstream Akt/FOXO1 signaling. Our results demonstrated that miR-145 could inhibit HCC through targeting IRS1 and its downstream signaling, implicating the loss of miR-145 regulation may be a potential molecular mechanism causing aberrant oncogenic signaling in HCC.

  1. Sam68 affects cell proliferation and apoptosis of human adult T-acute lymphoblastic leukemia cells via AKT/mTOR signal pathway.

    PubMed

    Wang, Qi; Li, Yuanye; Cheng, Jingying; Chen, Long; Xu, Hua; Li, Qinghua; Pang, Tianxiang

    2016-07-01

    Sam68 (Src associated in mitosis, 68kDa) belongs to the signal,transduction and activation of RNA (STAR) family and its function has been linked to the onset and progression of many tumors. However, the role of Sam68 in T-acute lymphoblastic leukemia (T-ALL) remains unclear. This present study aimed to investigate whether and how Sam68 involved in T-ALL. Our results showed high expression of Sam68 in adult T-ALL cases, Jurkat and CCRF-CEM cell lines. Knockdown of Sam68 repressed cell proliferation, increased apoptosis, induced S arrest along with upregulation of p21, Bad, cleaved caspase-9, caspase-3, PARP and downregulation of CDK2 and Bcl-xl. Furthermore, the data indicated that the expression change of Sam68 went with the changes of AKT/mTOR signaling pathway in T-ALL cell lines. Our findings demonstrated that Sam68 possibly participated in the progresses of T-ALL at least partially via AKT/mTOR signaling pathway. PMID:27107742

  2. Recombinant adeno-associated virus-delivered anginex inhibits angiogenesis and growth of HUVECs by regulating the Akt, JNK and NF-κB signaling pathways.

    PubMed

    Ma, Ke; Wang, Chuying; Geng, Qianqian; Fan, Yangwei; Ning, Jing; Yang, Haixia; Dong, Xuyuan; Dong, Danfeng; Guo, Yuyan; Wei, Xin; Li, Enxiao; Wu, Yinying

    2016-06-01

    Anginex is an artificial synthetic small molecule β-sheet-forming peptide shown to have anti-angiogenesis and antitumor effects in various solid tumors. However, its molecular mechanism remains largely unclear and efficient delivery methods for anginex remains to be developed. We report on the development of recombinant adeno-associated virus (rAAV2)-delivered anginex and the underlying mechanism of anti-angiogenesis and antitumor effects of anginex. We have successfully developed the rAAV2 vector to efficiently express anginex (rAAV2‑anginex). Transduction of rAAV2-anginex significantly induced apoptosis, and inhibited the proliferation, migration, invasion and tube formation of human umbilical vein endothelial cells in vitro. Western blot analysis revealed that rAAV2‑anginex inhibited the phosphorylation of Akt, while inducing the phosphorylation of JNK and activation of the NF-κB signaling pathway. In an in vivo CAM assay and xenograft model of SKOV3, rAAV2-anginex significantly reduced microvessel density (MVD) and vascular endothelial growth factor 165 (VEGF165), as demonstrated by immunohistochemistry analysis. Importantly, rAAV2-anginex inhibited tumor growth in an ovarian cancer SKOV3 cell nude mouse xenograft model. Our results suggest that rAAV2-anginex may inhibit tumor angiogenesis and growth through regulating Akt, JNK and NF-κB signaling pathways. PMID:27035232

  3. Disruption of Akt signaling decreases dopamine sensitivity in modulation of inhibitory synaptic transmission in rat prefrontal cortex.

    PubMed

    Li, Yan-Chun; Yang, Sha-Sha; Gao, Wen-Jun

    2016-09-01

    Akt is a serine/threonine kinase, which is dramatically reduced in the prefrontal cortex (PFC) of patients with schizophrenia, and a deficiency in Akt1 results in PFC function abnormalities. Although the importance of Akt in dopamine (DA) transmission is well established, how impaired Akt signaling affects the DA modulation of synaptic transmission in the PFC has not been characterized. Here we show that Akt inhibitors significantly decreased receptor sensitivity to DA by shifting DA modulation of GABAA receptor-mediated inhibitory postsynaptic currents (IPSCs) in prefrontal cortical neurons. Akt inhibition caused a significant decrease in synaptic dopamine D2 receptor (D2R) levels with high-dose DA exposure. In addition, Akt inhibition failed to affect DA modulation of IPSCs after blockade of β-arrestin 2. β-arrestin 2-mediated interaction of clathrin with D2R was enhanced by co-application of a Akt inhibitor and DA. Taken together, the reduced response in DA modulation of inhibitory transmission mainly involved β-arrestin 2-dependent D2R desensitization. PMID:27163190

  4. Quantitative Analysis of Robustness of Dynamic Response and Signal Transfer in Insulin mediated PI3K/AKT Pathway

    PubMed Central

    Mathew, Shibin; Banerjee, Ipsita

    2014-01-01

    Robustness is a critical feature of signaling pathways ensuring signal propagation with high fidelity in the event of perturbations. Here we present a detailed quantitative analysis of robustness in insulin mediated PI3K/AKT pathway, a critical signaling pathway maintaining self-renewal in human embryonic stem cells. Using global sensitivity analysis, we identified robustness promoting mechanisms that ensure (1) maintenance of a first order or overshoot dynamics of self-renewal molecule, p-AKT and (2) robust transfer of signals from oscillatory insulin stimulus to p-AKT in the presence of noise. Our results indicate that negative feedback controls the robustness to most perturbations. Faithful transfer of signal from the stimulating ligand to p-AKT occurs even in the presence of noise, albeit with signal attenuation and high frequency cut-off. Negative feedback contributes to signal attenuation, while positive regulators upstream of PIP3 contribute to signal amplification. These results establish precise mechanisms to modulate self-renewal molecules like p-AKT. PMID:25506104

  5. Mutations in the gravity persistence signal loci in Arabidopsis disrupt the perception and/or signal transduction of gravitropic stimuli

    NASA Technical Reports Server (NTRS)

    Wyatt, Sarah E.; Rashotte, Aaron M.; Shipp, Matthew J.; Robertson, Dominique; Muday, Gloria K.; Brown, C. S. (Principal Investigator)

    2002-01-01

    Gravity plays a fundamental role in plant growth and development, yet little is understood about the early events of gravitropism. To identify genes affected in the signal perception and/or transduction phase of the gravity response, a mutant screen was devised using cold treatment to delay the gravity response of inflorescence stems of Arabidopsis. Inflorescence stems of Arabidopsis show no response to gravistimulation at 4 degrees C for up to 3 h. However, when gravistimulated at 4 degrees C and then returned to vertical at room temperature (RT), stems bend in response to the previous, horizontal gravistimulation (H. Fukaki, H. Fujisawa, M. Tasaka [1996] Plant Physiology 110: 933-943). This indicates that gravity perception, but not the gravitropic response, occurs at 4 degrees C. Recessive mutations were identified at three loci using this cold effect on gravitropism to screen for gravity persistence signal (gps) mutants. All three mutants had an altered response after gravistimulation at 4 degrees C, yet had phenotypically normal responses to stimulations at RT. gps1-1 did not bend in response to the 4 degrees C gravity stimulus upon return to RT. gps2-1 responded to the 4 degrees C stimulus but bent in the opposite direction. gps3-1 over-responded after return to RT, continuing to bend to an angle greater than wild-type plants. At 4 degrees C, starch-containing statoliths sedimented normally in both wild-type and the gps mutants, but auxin transport was abolished at 4 degrees C. These results are consistent with GPS loci affecting an aspect of the gravity signal perception/transduction pathway that occurs after statolith sedimentation, but before auxin transport.

  6. Signal transduction by VEGF receptors in regulation of angiogenesis and lymphangiogenesis

    SciTech Connect

    Shibuya, Masabumi . E-mail: shibuya@ims.u-tokyo.ac.jp; Claesson-Welsh, Lena . E-mail: lena.welsh@genpat.uu.se

    2006-03-10

    The VEGF/VPF (vascular endothelial growth factor/vascular permeability factor) ligands and receptors are crucial regulators of vasculogenesis, angiogenesis, lymphangiogenesis and vascular permeability in vertebrates. VEGF-A, the prototype VEGF ligand, binds and activates two tyrosine kinase receptors: VEGFR1 (Flt-1) and VEGFR2 (KDR/Flk-1). VEGFR1, which occurs in transmembrane and soluble forms, negatively regulates vasculogenesis and angiogenesis during early embryogenesis, but it also acts as a positive regulator of angiogenesis and inflammatory responses, playing a role in several human diseases such as rheumatoid arthritis and cancer. The soluble VEGFR1 is overexpressed in placenta in preeclampsia patients. VEGFR2 has critical functions in physiological and pathological angiogenesis through distinct signal transduction pathways regulating proliferation and migration of endothelial cells. VEGFR3, a receptor for the lymphatic growth factors VEGF-C and VEGF-D, but not for VEGF-A, regulates vascular and lymphatic endothelial cell function during embryogenesis. Loss-of-function variants of VEGFR3 have been identified in lymphedema. Formation of tumor lymphatics may be stimulated by tumor-produced VEGF-C, allowing increased spread of tumor metastases through the lymphatics. Mapping the signaling system of these important receptors may provide the knowledge necessary to suppress specific signaling pathways in major human diseases.

  7. Enhanced fluorescence cyanide detection at physiologically lethal levels: reduced ICT-based signal transduction.

    PubMed

    Badugu, Ramachandram; Lakowicz, Joseph R; Geddes, Chris D

    2005-03-16

    Three water-soluble fluorescent probes have been specifically designed to determine free cyanide concentrations up to physiologically lethal levels, >20 microM. The probes have been designed in such a way as to afford many notable sensing features, which render them unique with regard to signal transduction, photophysical characteristics, and their application to physiological cyanide determination and safeguard. The probes are readily able to reversibly bind free aqueous cyanide with dissociation constants around 4 microM3. Subsequent cyanide binding modulates the intramolecular charge transfer within the probes, a change in the electronic properties within the probes, resulting in enhanced fluorescence optical signals as a function of increased solution cyanide concentration. The ground-state chelation with cyanide produces wavelength shifts, which also enable the probes to sense cyanide in both an excitation and emission ratiometric manner, in addition to enhanced fluorescence signaling. This has enabled a generic cyanide sensing platform to be realized that is not dependent on fluorescent probe concentration, probe photodegradation, or fluctuations in the intensity of any employed excitation sources, ideal for remote cyanide sensing applications. Further, the >600 nm fluorescence emission of the probes potentially allows for enhanced fluorescence ratiometric cyanide sensing in the optical window of tissues and blood, facilitating their use for the transdermal monitoring of cyanide for mammalian safeguard or postmortem in fire victims, both areas of active research. PMID:15755185

  8. An Agent-Based Model of Signal Transduction in Bacterial Chemotaxis

    PubMed Central

    Miller, Jameson; Parker, Miles; Bourret, Robert B.; Giddings, Morgan C.

    2010-01-01

    We report the application of agent-based modeling to examine the signal transduction network and receptor arrays for chemotaxis in Escherichia coli, which are responsible for regulating swimming behavior in response to environmental stimuli. Agent-based modeling is a stochastic and bottom-up approach, where individual components of the modeled system are explicitly represented, and bulk properties emerge from their movement and interactions. We present the Chemoscape model: a collection of agents representing both fixed membrane-embedded and mobile cytoplasmic proteins, each governed by a set of rules representing knowledge or hypotheses about their function. When the agents were placed in a simulated cellular space and then allowed to move and interact stochastically, the model exhibited many properties similar to the biological system including adaptation, high signal gain, and wide dynamic range. We found the agent based modeling approach to be both powerful and intuitive for testing hypotheses about biological properties such as self-assembly, the non-linear dynamics that occur through cooperative protein interactions, and non-uniform distributions of proteins in the cell. We applied the model to explore the role of receptor type, geometry and cooperativity in the signal gain and dynamic range of the chemotactic response to environmental stimuli. The model provided substantial qualitative evidence that the dynamic range of chemotactic response can be traced to both the heterogeneity of receptor types present, and the modulation of their cooperativity by their methylation state. PMID:20485527

  9. PII signal transduction proteins are ATPases whose activity is regulated by 2-oxoglutarate

    PubMed Central

    Radchenko, Martha V.; Thornton, Jeremy; Merrick, Mike

    2013-01-01

    PII proteins are one of the most widespread families of signal transduction proteins in nature, being ubiquitous throughout bacteria, archaea, and plants. In all these organisms, PII proteins coordinate many facets of nitrogen metabolism by interacting with and regulating the activities of enzymes, transcription factors, and membrane transport proteins. The primary mode of signal perception by PII proteins derives from their ability to bind the effector molecules 2-oxoglutarate (2-OG) and ATP or ADP. The role of 2-OG as an indicator of cellular nitrogen status is well understood, but the function of ATP/ADP binding has remained unresolved. We have now shown that the Escherichia coli PII protein, GlnK, has an ATPase activity that is inhibited by 2-OG. Hence, when a drop in the cellular 2-OG pool signals nitrogen sufficiency, 2-OG depletion of GlnK causes bound ATP to be hydrolyzed to ADP, leading to a conformational change in the protein. We propose that the role of ATP/ADP binding in E. coli GlnK is to effect a 2-OG-dependent molecular switch that drives a conformational change in the T loops of the PII protein. We have further shown that two other PII proteins, Azospirillum brasilense GlnZ and Arabidopsis thaliana PII, have a similar ATPase activity, and we therefore suggest that this switch mechanism is likely to be a general property of most members of the PII protein family. PMID:23818625

  10. The role of calcium in hypoxia-induced signal transduction and gene expression.

    PubMed

    Seta, Karen A; Yuan, Yong; Spicer, Zachary; Lu, Gang; Bedard, James; Ferguson, Tsuneo K; Pathrose, Peterson; Cole-Strauss, Allyson; Kaufhold, Alexa; Millhorn, David E

    2004-01-01

    Mammalian cells require a constant supply of oxygen in order to maintain adequate energy production, which is essential for maintaining normal function and for ensuring cell survival. Sustained hypoxia can result in cell death. Sophisticated mechanisms have therefore evolved which allow cells to respond and adapt to hypoxia. Specialized oxygen-sensing cells have the ability to detect changes in oxygen tension and transduce this signal into organ system functions that enhance the delivery of oxygen to tissue in a wide variety of different organisms. An increase in intracellular calcium levels is a primary response of many cell types to hypoxia/ischemia. The response to hypoxia is complex and involves the regulation of multiple signaling pathways and coordinated expression of perhaps hundreds of genes. This review discusses the role of calcium in hypoxia-induced regulation of signal transduction pathways and gene expression. An understanding of the molecular events initiated by changes in intracellular calcium will lead to the development of therapeutic approaches toward the treatment of hypoxic/ischemic diseases and tumors. PMID:15261489

  11. Isoforms of the CD79 signal transduction component of the macropod B-cell receptor.

    PubMed

    Suthers, Amy N; Young, Lauren J

    2014-12-01

    B cell responses and their concomitant signal transduction pathways are not well understood in marsupial mammals, despite the availability of gene expression data for key immunoglobulin genes and for elements of the CD79a/CD79b heterodimer signalling complex for two model marsupials. Broader studies of factors that influence B cell responses are still hampered by a lack of species-specific reagents and there are few reports of other factors that influence gene expression such as the potential for splice variants in BCR components, which may influence immune signalling pathways. In this study, we characterise CD79a and CD79b genes in the endangered macropod marsupial, Onychogalea fraenata (the bridled nailtail wallaby) and show that domains and residues important for the structural and functional integrity of both monomers are conserved in this species, consistent with results previously reported for the closely-related macropod, Macropus eugenii (the tammar wallaby). We extend this work to report the detection of splice variants for CD79a and CD79b in wallaby species; three CD79a isoforms and one CD79b isoform. Of these, two CD79a isoforms and the CD79b isoform have not been reported in any other mammalian species. PMID:25064685

  12. Costal2 Functions as a Microtubule-Dependent Motor in the Hedgehog Signal Transduction Pathway

    PubMed Central

    Farzan, Shohreh F.; Ascano, Manuel; Ogden, Stacey K.; Sanial, Matthieu; Brigui, Amira; Plessis, Anne; Robbins, David J.

    2009-01-01

    SUMMARY The Hedgehog (Hh) signaling pathway initiates an evolutionarily conserved developmental program required for the proper patterning of many tissues. Costal2 (Cos2) is a requisite component of the Hh pathway, whose mechanistic role is not well understood. Cos2 was initially predicted, based on its primary sequence, to function as a microtubule-associated (MT) molecular motor. However, despite being identified over a decade ago, evidence showing that Cos2 function might require kinesin-like properties has for the most part been lacking. Thus the prevailing dogma in the field is that Cos2 functions solely as a scaffolding protein during Hh signal transduction. Here, we provide the first evidence that Cos2 motility is required for its biological function, and that this motility may be Hh regulated. We show that Cos2 motility requires an active motor domain, ATP and microtubules. Additionally, Cos2 recruits and transports other components of the Hh signaling pathway, including the transcription factor Cubitus interruptus (Ci), throughout the cell. Drosophila expressing cos2 mutations that encode proteins that lack motility are attenuated in their ability to regulate Ci activity and exhibit phenotypes consistent with attenuated Cos2 function. Combined, these results demonstrate that Cos2 motility plays an important role in its function, regulating the amounts and activity of Ci that ultimately interpret the level of Hh to which cells are exposed. PMID:18691888

  13. Cross-talk between the Akt and NF-κB Signaling Pathways Inhibits MEHP-Induced Germ Cell Apoptosis

    PubMed Central

    Rogers, Rachel; Ouellet, Gregory; Moyer, Ben; Rasoulpour, Teresa; Hixon, Mary

    2008-01-01

    Phthalates are ubiquitous contaminants that target the testis during in utero and postnatal development. The PI3K/Akt and nuclear factor kappa B (NF-κB) signaling pathways have been implicated in germ cell survival following testicular injury. Here we observe that Akt kinase activity increases in the testes of postnatal day 28 wild-type mice following exposure to 500 mg/kg mono-(2-ethylhexyl) phthalate (MEHP), and that loss of Akt1 results in the premature onset of germ cell apoptosis. To further determine the basis for this sensitivity, we investigated the potential for cross-talk between the PI3K/Akt and NF-κB signaling pathways. We found a twofold increase in Akt1-dependent phosphorylation of the IκBα subunit following exposure to 500 mg/kg MEHP and decreased levels of the total IκBα protein. Examination of the expression of the NF-κB subunits, p50 and p65, in Akt1 wild-type testes following MEHP exposure revealed a twofold increase in p50 mRNA at 6 h. Interestingly, in Akt1-deficient testes, basal expression of both the p50 and p65 subunits was elevated 1.6- and 4-fold, respectively. This was due, at least in part, to increased levels of oxidative stress as measured by both superoxide anion formation and increased expression of SMAC/DIABLO, a proapoptotic mitochondrial protein. In wild-type testes, MEHP-induced Akt1-dependent transcription of the antiapoptotic mitochondrial target gene, Bcl-xL. Together, these results indicate that Akt1 plays a role in the initial protection of germ cells following MEHP-induced germ cell apoptosis and that this response is partially mediated by cross-talk with the NF-κB signaling pathway and an increased sensitivity to oxidative stress. PMID:18755736

  14. Targeting FAK Radiosensitizes 3-Dimensional Grown Human HNSCC Cells Through Reduced Akt1 and MEK1/2 Signaling

    SciTech Connect

    Hehlgans, Stephanie; Department of Radiotherapy and Oncology, University of Frankfurt, Frankfurt am Main; Institute of Radiopharmacy, Helmholtz Center Dresden-Rossendorf, Dresden ; Eke, Iris; Cordes, Nils; Institute of Radiopharmacy, Helmholtz Center Dresden-Rossendorf, Dresden; Department of Radiation Oncology, University Hospital and Medical Faculty Carl Gustav Carus, Dresden University of Technology, Dresden

    2012-08-01

    Purpose: Focal adhesion kinase (FAK), a main regulator of integrin signaling and cell migration, is frequently overexpressed and hyperphosphorylated in human head-and-neck squamous cell carcinoma (HNSCC). We have previously shown that pharmacologic FAK inhibition leads to radiosensitization of 3-dimensionally grown HNSCC cell lines. To further evaluate the role of FAK in radioresistance and as a potential cancer target, we examined FAK and FAK downstream signaling in HNSCC cell lines grown in more physiologic extracellular matrix-based 3-dimensional cell cultures. Methods and Materials: Seven HNSCC cell lines were grown in 3-dimensional extracellular matrix and the clonogenic radiation survival, expression, and phosphorylation of FAK, paxillin, Akt1, extracellular signal-regulated kinase (ERK)1/2, and MEK1/2 were analyzed after siRNA-mediated knockdown of FAK, Akt1, MEK1, FAK+Akt1, or FAK+MEK1 compared with controls or stable overexpression of FAK. The role of MEK1/2 for clonogenic survival and signaling was investigated using the MEK inhibitor U0126 with or without irradiation. Results: FAK knockdown moderately or significantly enhanced the cellular radiosensitivity of 3-dimensionally grown HNSCC cells. The FAK downstream targets paxillin, Akt1, and ERK1/2 were substantially dephosphorylated under FAK depletion. FAK overexpression, in contrast, increased radiation survival and paxillin, Akt1, and ERK1/2 phosphorylation. The degree of radiosensitization upon Akt1, ERK1/2, or MEK1 depletion or U0126 was superimposable to FAK knockdown. Combination knockdown conditions (ie, Akt1/FAK, MEK1/FAK, or U0126/FAK) failed to provide additional radiosensitization. Conclusions: Our data provide further evidence for FAK as important determinant of radiation survival, which acts in the same signaling axis as Akt1 and ERK1/2. These data strongly support our hypothesis that FAK is a relevant molecular target for HNSCC radiotherapy.

  15. Statistics of cellular signal transduction as a race to the nucleus by multiple random walkers in compartment/phosphorylation space

    PubMed Central

    Lu, Ting; Shen, Tongye; Zong, Chenghang; Hasty, Jeff; Wolynes, Peter G.

    2006-01-01

    Cellular signal transduction often involves a reaction network of phosphorylation and transport events arranged with a ladder topology. If we keep track of the location of the phosphate groups describing an abstract state space, a simple model of signal transduction involving enzymes can be mapped on to a problem of how multiple biased random walkers compete to reach their target in the nucleus yielding a signal. Here, the first passage time probability and the survival probability for multiple walkers can be used to characterize the response of the network. The statistics of the first passage through the network has an asymmetric distribution with a long tail arising from the hierarchical structure of the network. This distribution implies a significant difference between the mean and the most probable signal transduction time. The response patterns for various external inputs generated by our model agree with recent experiments. In addition, the model predicts that there is an optimal phosphorylation enzyme concentration for rapid signal transduction. PMID:17071742

  16. Roles of Intracellular Cyclic AMP Signal Transduction in the Capacitation and Subsequent Hyperactivation of Mouse and Boar Spermatozoa

    PubMed Central

    HARAYAMA, Hiroshi

    2013-01-01

    It is not until accomplishment of a variety of molecular changes during the transit through the female reproductive tract that mammalian spermatozoa are capable of exhibiting highly activated motility with asymmetric whiplash beating of the flagella (hyperactivation) and undergoing acrosomal exocytosis in the head (acrosome reaction). These molecular changes of the spermatozoa are collectively termed capacitation and promoted by bicarbonate, calcium and cholesterol acceptors. Such capacitation-promoting factors can stimulate intracellular cyclic AMP (cAMP) signal transduction in the spermatozoa. Meanwhile, hyperactivation and the acrosome reaction are essential to sperm fertilization with oocytes and are apparently triggered by a sufficient increase of intracellular Ca2+ in the sperm flagellum and head, respectively. Thus, it is necessary to investigate the relationship between cAMP signal transduction and calcium signaling cascades in the spermatozoa for the purpose of understanding the molecular basis of capacitation. In this review, I cover updated insights regarding intracellular cAMP signal transduction, the acrosome reaction and flagellar motility in mammalian spermatozoa and then account for possible roles of intracellular cAMP signal transduction in the capacitation and subsequent hyperactivation of mouse and boar spermatozoa. PMID:24162806

  17. Inhibition of Akt/mTOR signaling by the dietary flavonoid fisetin.

    PubMed

    Syed, Deeba N; Adhami, Vaqar M; Khan, Mohammad Imran; Mukhtar, Hasan

    2013-09-01

    Plants have long been providing mankind with remedies of different ailments. Flavonoids, a family of naturally occurring polyphenolic compounds are ubiquitous in plants. Development of polyphenol-based drugs has not attracted much attention by researchers and drug companies. Therefore, despite extensive studies on polyphenols, this vast group of compounds is underrepresented in clinical medicine. Fisetin (3,7,3',4'-tetrahydroxyflavone) belongs to the flavonol subgroup of flavonoids together with quercetin, myricetin and kaempferol and is found in several fruits and vegetables including strawberries, apples, persimmons and onions. Fisetin is showing promise as a useful natural agent against cancer and has been evaluated for its potential inhibitory role against cancer in several in vitro and in vivo studies. The Akt/mTOR pathway is known to play a central role in various cellular processes that contribute to the malignant phenotype. Accordingly, inhibition of this signaling cascade has been a focus of recent therapeutic studies. Novel inhibitors of PI3-K, Akt, and mTOR are now passing through early phase clinical trials. Herein, we review the effect of fisetin on the PI3- K/Akt/mTOR pathway as studied in different cancer cell models. PMID:23293889

  18. Inhibition of Akt/mTOR Signaling by the Dietary Flavonoid Fisetin

    PubMed Central

    Syed, Deeba N.; Adhami, Vaqar M.; Khan, Mohammad Imran; Mukhtar, Hasan

    2014-01-01

    Plants have long been providing mankind with remedies of different ailments. Flavonoids, a family of naturally occurring polyphenolic compounds are ubiquitous in plants. Development of polyphenol-based drugs has not attracted much attention by researchers and drug companies. Therefore, despite extensive studies on polyphenols, this vast group of compounds is underrepresented in clinical medicine. Fisetin (3,7,3’,4’-tetrahydroxyflavone) belongs to the flavonol subgroup of flavonoids together with quercetin, myricetin and kaempferol and is found in several fruits and vegetables including strawberries, apples, persimmons and onions. Fisetin is showing promise as a useful natural agent against cancer and has been evaluated for its potential inhibitory role against cancer in several in vitro and in vivo studies. The Akt/mTOR pathway is known to play a central role in various cellular processes that contribute to the malignant phenotype. Accordingly, inhibition of this signaling cascade has been a focus of recent therapeutic studies. Novel inhibitors of PI3-K, Akt, and mTOR are now passing through early phase clinical trials. Herein, we review the effect of fisetin on the PI3-K/Akt/mTOR pathway as studied in different cancer cell models. PMID:23293889

  19. p38 MAPK and PI3K/AKT Signalling Cascades inParkinson’s Disease

    PubMed Central

    Jha, Saurabh Kumar; Jha, Niraj Kumar; Kar, Rohan; Ambasta, Rashmi K; Kumar, Pravir

    2015-01-01

    Parkinson's disease (PD) is a chronic neurodegenerative condition which has the second largest incidence rate among all other neurodegenerative disorders barring Alzheimer's disease (AD). Currently there is no cure and researchers continue to probe the therapeutic prospect in cell cultures and animal models of PD. Out of the several factors contributing to PD prognosis, the role of p38 MAPK (Mitogen activated protein-kinase) and PI3K/AKT signalling module in PD brains is crucial because the impaired balance between the pro- apoptotic and anti-apoptotic pathways trigger unwanted phenotypes such as microglia activation, neuroinflammation, oxidative stress and apoptosis. These factors continue challenging the brain homeostasis in initial stages thereby essentially assisting the dopaminergic (DA) neurons towards progressive degeneration in PD. Neurotherapeutics against PD shall then be targeted against the misregulated accomplices of the p38 and PI3K/AKT cascades. In this review, we have outlined many such established mechanisms involving the p38 MAPK and PI3K/AKT pathways which can offer therapeutic windows for the rectification of aberrant DA neuronal dynamics in PD brains. PMID:26261796

  20. Adiponectin Induces Oncostatin M Expression in Osteoblasts through the PI3K/Akt Signaling Pathway

    PubMed Central

    Su, Chen-Ming; Lee, Wei-Lin; Hsu, Chin-Jung; Lu, Ting-Ting; Wang, Li-Hong; Xu, Guo-Hong; Tang, Chih-Hsin

    2015-01-01

    Rheumatoid arthritis (RA), a common autoimmune disorder, is associated with a chronic inflammatory response and unbalanced bone metabolism within the articular microenvironment. Adiponectin, an adipokine secreted by adipocytes, is involved in multiple functions, including lipid metabolism and pro-inflammatory activity. However, the mechanism of adiponectin performance within arthritic inflammation remains unclear. In this study, we observed the effect of adiponectin on the expression of oncostatin M (OSM), a pro-inflammatory cytokine, in human osteoblastic cells. Pretreatment of cells with inhibitors of phosphatidylinositol 3-kinase (PI3K), Akt, and nuclear factor (NF)-κB reduced the adiponectin-induced OSM expression in osteoblasts. Stimulation of the cells with adiponectin increased phosphorylation of PI3K, Akt, and p65. Adiponectin treatment of osteoblasts increased OSM-luciferase activity and p65 binding to NF-κB on the OSM promoter. Our results indicate that adiponectin increased OSM expression via the PI3K, Akt, and NF-κB signaling pathways in osteoblastic cells, suggesting that adiponectin is a novel target for arthritis treatment. PMID:26712749

  1. Breviscapine attenuatted contrast medium-induced nephropathy via PKC/Akt/MAPK signalling in diabetic mice

    PubMed Central

    Jiang, Wenbin; Li, Zhengwei; Zhao, Wei; Chen, Hao; Wu, Youyang; Wang, Yi; Shen, Zhida; He, Jialin; Chen, Shengyu; Zhang, Jiefang; Fu, Guosheng

    2016-01-01

    Contrast medium-induced nephropathy (CIN) remains a major cause of iatrogenic, drug-induced renal injury. Recent studies reveal that Breviscapine can ameliorate diabetic nephropathy in mice. Yet it remains unknown if Breviscapine could reduce CIN in diabetic mice. In this study, male C57/BL6J mice were randomly divided into 7 groups: control, diabetes mellitus, CIN, diabetes mellitus+CIN, diabetes mellitus+Breviscapine, CIN+Breviscapine and diabetes mellitus+CIN+Breviscapine. Model of CIN was induced by tail intravenous administration of iopromide and model of diabetes mellitus was induced by Streptozotocin intraperitoneally. Breviscapine was administered intragastrically for 4 weeks. Renal function parameters, kidney histology, markers of renal fibrosis, phosphorylation of protein kinase C/Akt/mitogen activated protein kinases were measured by western blot. We found out that diabetes mellitus aggravated CIN damage. Renal histological analysis showed Breviscapine reduced of renal fibrosis and tubular damage. Breviscapine was also shown markedly to ameliorate CIN fibrotic markers expression, reduced proteinuria and serum creatinine. Furthermore, Breviscapine decreased phosphorylation of PKCβII, Akt, JNK1/2 and p38. Therefore, Breviscapine treatment could ameliorate the development of CIN in diabetic mice, which was partly attributed to its suppression of renal fibrosis via phosphorylation of PKCβII/Akt/JNK1/2/p38 signalling. PMID:27158329

  2. Singularity analysis of the AKT signaling pathway reveals connections between cancer and metabolic diseases

    NASA Astrophysics Data System (ADS)

    Wang, Guanyu

    2010-12-01

    Connections between cancer and metabolic diseases may consist in the complex network of interactions among a common set of biomolecules. By applying singularity and bifurcation analysis, the phenotypes constrained by the AKT signaling pathway are identified and mapped onto the parameter space, which include cancer and certain metabolic diseases. By considering physiologic properties (sensitivity, robustness and adaptivity) the AKT pathway must possess in order to efficiently sense growth factors and nutrients, the region of normal responses is located. To optimize these properties, the intracellular concentration of the AKT protein must be sufficiently high to saturate its enzymes; the strength of the positive feedback must be stronger than that of the negative feedback. The analysis illuminates the parameter space and reveals system-level mechanisms in regulating biological functions (cell growth, survival, proliferation and metabolism) and how their deregulation may lead to the development of diseases. The analytical expressions summarize the synergistic interactions among many molecules, which provides valuable insights into therapeutic interventions. In particular, a strategy for overcoming the limitations of mTOR inhibition is proposed for cancer therapy.

  3. Peroxiredoxins in Regulation of MAPK Signalling Pathways; Sensors and Barriers to Signal Transduction

    PubMed Central

    Latimer, Heather R.; Veal, Elizabeth A.

    2016-01-01

    Peroxiredoxins are highly conserved and abundant peroxidases. Although the thioredoxin peroxidase activity of peroxiredoxin (Prx) is important to maintain low levels of endogenous hydrogen peroxide, Prx have also been shown to promote hydrogen peroxide-mediated signalling. Mitogen activated protein kinase (MAPK) signalling pathways mediate cellular responses to a variety of stimuli, including reactive oxygen species (ROS). Here we review the evidence that Prx can act as both sensors and barriers to the activation of MAPK and discuss the underlying mechanisms involved, focusing in particular on the relationship with thioredoxin. PMID:26813660

  4. Peroxiredoxins in Regulation of MAPK Signalling Pathways; Sensors and Barriers to Signal Transduction.

    PubMed

    Latimer, Heather R; Veal, Elizabeth A

    2016-01-01

    Peroxiredoxins are highly conserved and abundant peroxidases. Although the thioredoxin peroxidase activity of peroxiredoxin (Prx) is important to maintain low levels of endogenous hydrogen peroxide, Prx have also been shown to promote hydrogen peroxide-mediated signalling. Mitogen activated protein kinase (MAPK) signalling pathways mediate cellular responses to a variety of stimuli, including reactive oxygen species (ROS). Here we review the evidence that Prx can act as both sensors and barriers to the activation of MAPK and discuss the underlying mechanisms involved, focusing in particular on the relationship with thioredoxin. PMID:26813660

  5. Elucidating the Functional Roles of Spatial Organization in Cross-Membrane Signal Transduction by a Hybrid Simulation Method.

    PubMed

    Chen, Jiawen; Xie, Zhong-Ru; Wu, Yinghao

    2016-07-01

    The ligand-binding of membrane receptors on cell surfaces initiates the dynamic process of cross-membrane signal transduction. It is an indispensable part of the signaling network for cells to communicate with external environments. Recent experiments revealed that molecular components in signal transduction are not randomly mixed, but spatially organized into distinctive patterns. These patterns, such as receptor clustering and ligand oligomerization, lead to very different gene expression profiles. However, little is understood about the molecular mechanisms and functional impacts of this spatial-temporal regulation in cross-membrane signal transduction. In order to tackle this problem, we developed a hybrid computational method that decomposes a model of signaling network into two simulation modules. The physical process of binding between receptors and ligands on cell surfaces are simulated by a diffusion-reaction algorithm, while the downstream biochemical reactions are modeled by stochastic simulation of Gillespie algorithm. These two processes are coupled together by a synchronization framework. Using this method, we tested the dynamics of a simple signaling network in which the ligand binding of cell surface receptors triggers the phosphorylation of protein kinases, and in turn regulates the expression of target genes. We found that spatial aggregation of membrane receptors at cellular interfaces is able to either amplify or inhibit downstream signaling outputs, depending on the details of clustering mechanism. Moreover, by providing higher binding avidity, the co-localization of ligands into multi-valence complex modulates signaling in very different ways that are closely related to the binding affinity between ligand and receptor. We also found that the temporal oscillation of the signaling pathway that is derived from genetic feedback loops can be modified by the spatial clustering of membrane receptors. In summary, our method demonstrates the functional

  6. Phosphatase of Regenerating Liver 3 (PRL3) Provokes a Tyrosine Phosphoproteome to Drive Prometastatic Signal Transduction*

    PubMed Central

    Walls, Chad D.; Iliuk, Anton; Bai, Yunpeng; Wang, Mu; Tao, W. Andy; Zhang, Zhong-Yin

    2013-01-01

    Phosphatase of regenerating liver 3 (PRL3) is suspected to be a causative factor toward cellular metastasis when in excess. To date, the molecular basis for PRL3 function remains an enigma, making efforts at distilling a concerted mechanism for PRL3-mediated metastatic dissemination very difficult. We previously discovered that PRL3 expressing cells exhibit a pronounced increase in protein tyrosine phosphorylation. Here we take an unbiased mass spectrometry-based approach toward identifying the phosphoproteins exhibiting enhanced levels of tyrosine phosphorylation with a goal to define the “PRL3-mediated signaling network.” Phosphoproteomic data support intracellular activation of an extensive signaling network normally governed by extracellular ligand-activated transmembrane growth factor, cytokine, and integrin receptors in the PRL3 cells. Additionally, data implicate the Src tyrosine kinase as the major intracellular kinase responsible for “hijacking” this network and provide strong evidence that aberrant Src activation is a major consequence of PRL3 overexpression. Importantly, the data support a PDGF(α/β)-, Eph (A2/B3/B4)-, and Integrin (β1/β5)-receptor array as being the predominant network coordinator in the PRL3 cells, corroborating a PRL3-induced mesenchymal-state. Within this network, we find that tyrosine phosphorylation is increased on a multitude of signaling effectors responsible for Rho-family GTPase, PI3K-Akt, STAT, and ERK activation, linking observations made by the field as a whole under Src as a primary signal transducer. Our phosphoproteomic data paint the most comprehensive picture to date of how PRL3 drives prometastatic molecular events through Src activation. PMID:24030100

  7. Chromium stress response effect on signal transduction and expression of signaling genes in rice.

    PubMed

    Trinh, Ngoc-Nam; Huang, Tsai-Lien; Chi, Wen-Chang; Fu, Shih-Feng; Chen, Chi-Chien; Huang, Hao-Jen

    2014-02-01

    Hexavalent chromium [Cr(VI)] is a non-essential metal for normal plants and is toxic to plants at high concentrations. However, signaling pathways and molecular mechanisms of its action on cell function and gene expression remain elusive. In this study, we found that Cr(VI) induced endogenous reactive oxygen species (ROS) generation and Ca(2+) accumulation and activated NADPH oxidase and calcium-dependent protein kinase. We investigated global transcriptional changes in rice roots by microarray analysis. Gene expression profiling indicated activation of abscisic acid-, ethylene- and jasmonic acid-mediated signaling and inactivation of gibberellic acid-related pathways in Cr(VI) stress-treated rice roots. Genes encoding signaling components such as the protein kinases domain of unknown function 26, receptor-like cytoplasmic kinase, LRK10-like kinase type 2 and protein phosphatase 2C, as well as transcription factors WRKY and apetala2/ethylene response factor were predominant during Cr(VI) stress. Genes involved in vesicle trafficking were subjected to functional characterization. Pretreating rice roots with a vesicle trafficking inhibitor, brefeldin A, effectively reduced Cr(VI)-induced ROS production. Suppression of the vesicle trafficking gene, Exo70, by virus-induced gene silencing strategies revealed that vesicle trafficking is required for mediation of Cr(VI)-induced ROS production. Taken together, these findings shed light on the molecular mechanisms in signaling pathways and transcriptional regulation in response to Cr stress in plants. PMID:24033343

  8. Design principles of nuclear receptor signaling: how complex networking improves signal transduction

    PubMed Central

    Kolodkin, Alexey N; Bruggeman, Frank J; Plant, Nick; Moné, Martijn J; Bakker, Barbara M; Campbell, Moray J; van Leeuwen, Johannes P T M; Carlberg, Carsten; Snoep, Jacky L; Westerhoff, Hans V

    2010-01-01

    The topology of nuclear receptor (NR) signaling is captured in a systems biological graphical notation. This enables us to identify a number of ‘design' aspects of the topology of these networks that might appear unnecessarily complex or even functionally paradoxical. In realistic kinetic models of increasing complexity, calculations show how these features correspond to potentially important design principles, e.g.: (i) cytosolic ‘nuclear' receptor may shuttle signal molecules to the nucleus, (ii) the active export of NRs may ensure that there is sufficient receptor protein to capture ligand at the cytoplasmic membrane, (iii) a three conveyor belts design dissipating GTP-free energy, greatly aids response, (iv) the active export of importins may prevent sequestration of NRs by importins in the nucleus and (v) the unspecific nature of the nuclear pore may ensure signal-flux robustness. In addition, the models developed are suitable for implementation in specific cases of NR-mediated signaling, to predict individual receptor functions and differential sensitivity toward physiological and pharmacological ligands. PMID:21179018

  9. Flotillin-2 promotes metastasis of nasopharyngeal carcinoma by activating NF-κB and PI3K/Akt3 signaling pathways

    PubMed Central

    Liu, Jie; Huang, Wei; Ren, Caiping; Wen, Qiuyuan; Liu, Weidong; Yang, Xuyu; Wang, Lei; Zhu, Bin; Zeng, Liang; Feng, Xiangling; Zhang, Chang; Chen, Huan; Jia, Wei; Zhang, Lihua; Xia, Xiaomeng; Chen, Yuxiang

    2015-01-01

    Lipid raft proteins have been confirmed to be important in cell signal transduction. Some reports have shown that the aberrant expression of lipid raft proteins is associated with malignant phenotypes in some cancers. However, the role of the lipid raft protein flotillin-2 (Flot-2) in nasopharyngeal carcinoma (NPC) remains to be comprehensively characterized. Here, overexpression of Flot-2 in NPC tissues and cell lines was detected by immunostaining, and Flot-2 expression was found to be positively associated with NPC metastasis. Furthermore, inhibiting Flot-2 expression impaired the malignancy of the highly metastatic NPC cell line 5-8F by constraining its growth and proliferation, mobility and migration, and decreasing the capacity of 5-8F cells to metastasize in nude mice. In contrast, forced overexpression of Flot-2 increased the malignancy of 6-10B, a non-metastatic NPC cell line that weakly expresses Flot-2. Moreover, in 5-8F-shFlot-2 cells, which have inhibited Flot-2 expression, the NF-κB and PI3K/Akt3 pathways were inactivated. Subsequently, MMPs expression were decreased, and Foxo1 activity was increased. In addition, enhanced NF-κB and PI3K/Akt3 activities were observed in Flot-2 overexpressing 6-10B cells. Thus, Flot-2 exerts a pro-neoplastic role in NPC and is involved in tumor progression and metastasis. Moreover, Flot-2 exerts its role through NF-κB and PI3K/Akt3 signaling. PMID:26206082

  10. NFV, an HIV-1 protease inhibitor, induces growth arrest, reduced Akt signalling, apoptosis and docetaxel sensitisation in NSCLC cell lines.

    PubMed

    Yang, Y; Ikezoe, T; Nishioka, C; Bandobashi, K; Takeuchi, T; Adachi, Y; Kobayashi, M; Takeuchi, S; Koeffler, H P; Taguchi, H

    2006-12-18

    HIV-1 protease inhibitor (PI), nelfinavir (NFV) induced growth arrest and apoptosis of NCI-H460 and -H520, A549, EBC-1 and ABC-1 non-small-cell lung cancer (NSCLC) cells in association with upregulation of p21waf1, p27kip1 and p53, and downregulation of Bcl-2 and matrix metalloproteinase (MMP)-2 proteins. We found that NFV blocked Akt signalling in these cells as measured by Akt kinase assay with glycogen synthase kinase-3alpha/beta (GSK-3alpha/beta) as a substrate. To explore the role of Akt signalling in NFV-mediated growth inhibition of NSCLC cells, we blocked this signal pathway by transfection of Akt small interfering RNA (siRNA) in these cells; transient transfection of Akt siRNA in NCI-H460 cells decreased the level of Bcl-2 protein and slowed their proliferation compared to the nonspecific siRNA-transfected cells. Conversely, forced-expression of Akt partially reversed NFV-mediated growth inhibition of these cells, suggesting that Akt may be a molecular target of NFV in NSCLC cells. Also, we found that inhibition of Akt signalling by NFV enhanced the ability of docetaxel to inhibit the growth of NCI-H460 and -H520 cells, as measured by MTT assay. Importantly, NFV slowed the proliferation and induced apoptosis of NCI-H460 cells present as tumour xenografts in nude mice without adverse systemic effects. Taken together, this family of compounds might be useful for the treatment of individuals with NSCLC. PMID:17133272

  11. Glucocorticoid Suppresses Connexin 43 Expression by Inhibiting the Akt/mTOR Signaling Pathway in Osteoblasts.

    PubMed

    Shen, Chen; Kim, Mi Ran; Noh, Jeong Mi; Kim, Su Jin; Ka, Sun-O; Kim, Ji Hye; Park, Byung-Hyun; Park, Ji Hyun

    2016-07-01

    The inhibition of proliferation or functional alteration of osteoblasts by glucocorticoids (GCs) has been recognized as an important etiology of GC-induced osteoporosis (GIO). Connexin 43 (Cx43) is the most abundant connexin isoform in bone cells and plays important roles in bone remodeling. Despite the important role of Cx43 in bone homeostasis and the prevalence of GIO, the direct action of GCs on Cx43 expression in osteoblasts has been poorly described. The aim of the present study was to evaluate how GCs affect Cx43 expression in osteoblasts. Dexamethasone (Dex) treatment decreased expression of Cx43 RNA and protein in MC3T3-E1 mouse osteoblastic cells. Reduction of Cx43 expression by Dex was dependent on the glucocorticoid receptor (GR), as it was abolished by pretreatment with a GR blocker. Treatment with PTH (1-34), a medication used for GIO management, counteracted the suppression of Cx43 by Dex. Akt or mTOR signaling modulators revealed the involvement of the Akt/mTOR signaling pathway in Dex-induced reduction of Cx43 expression. Moreover, overexpression of Cx43 significantly attenuated Dex-inhibited cell viability and proliferation, as evidenced by MTT and bromodeoxyuridine (BrdU) incorporation assay of MC3T3-E1 cells. To account for possible species or cell type differences, human primary osteoblasts were treated with Dex and similar downregulation of Cx43 by Dex was observed. In addition, immunofluorescent staining for Cx43 further demonstrated an apparent decrease in Dex-treated human osteoblasts, while analysis of lucifer yellow propagation revealed reduced gap junction intercellular communication by Dex. Collectively, these findings indicate that GCs suppress Cx43 expression in osteoblasts via GR and the Akt/mTOR signaling pathway and overexpression of Cx43 may, at least in part, rescue osteoblasts from GC-induced reductions in proliferation. PMID:26914606

  12. [Progress of studies on acu-moxibustion stimulation-induced cellular transmembrane signal transduction of the target-organs].

    PubMed

    Yi, Shou-Xiang; Peng, Yan

    2009-10-01

    Abundant research results have shown that multiple levels and links of cellular transmembrane signal transduction pathways in the target organs were involved in the efficacy of acupuncture. For instance, 1) various extra-cellular growth factors for initiating signal transduction by activating tyrosine protein kinase and non-receptor tyrosine kinase, 2) G protein-coupled protein kinase-second signal messengers, 3) ligands acting on intra-nuclear receptors to activate transduction pathway of nuclear transcription factors of the target genes, have been demonstrated in the favorable regulating process of acupuncture and moxibustion in different pathological animal models. In the present paper, the authors review the progress of studies on the abovementioned mechanism of acu-moxibustion underlying improving some disorders as 1) pain, cerebral ischemia, and senile dementia, 2) inflammation and tumor, and 3) myocardial ischemia. Moreover, the authors also analyze the extant problems and make a prospect on the future studies about the cellular transmembrane signal transduction pathways involving the effects of acupuncture and moxibustion. PMID:20128298

  13. Inquiry into Chemotherapy-Induced P53 Activation in Cancer Cells as a Model for Teaching Signal Transduction

    ERIC Educational Resources Information Center

    Srougi, Melissa C.; Carson, Susan

    2013-01-01

    Intracellular and extracellular communication is conducted through an intricate and interwoven network of signal transduction pathways. The mechanisms for how cells speak with one another are of significant biological importance to both basic and industrial scientists from a number of different disciplines. We have therefore developed and…

  14. Activation of gene expression by metal-responsive signal transduction pathways.

    PubMed Central

    Adams, Timothy K; Saydam, Nurten; Steiner, Florian; Schaffner, Walter; Freedman, Jonathan H

    2002-01-01

    Metallothioneins are small, cysteine-rich, metal-binding proteins that play important roles in maintaining intracellular metal homeostasis and in transition metal detoxification. MTF-1 (metal transcription factor-1) plays a central role in regulating the metal-inducible, transcriptional activation of metallothionein. Here we report that the phosphorylation of MTF-1 plays a critical role in the activation of MTF-1/metal-responsive element-mediated transcription. Inhibitor studies indicate that signal transduction cascades, including those mediated by protein kinase C, tyrosine kinase, and casein kinase II, are essential for zinc- and cadmium-inducible transcription. In addition, calcium signaling is also involved in regulating transcription. In contrast, cAMP-dependent protein kinase may not be directly involved in the metal response. Contrary to what has been reported for other transcription factors, the inhibition of transcriptional activation does not impair the binding of MTF-1 to DNA, suggesting that phosphorylation is not regulating DNA binding. Elevated phosphorylation of MTF-1 is observed under conditions of protein kinase C inhibition, suggesting that dephosphorylation of this transcription factor mediates its activation. PMID:12426137

  15. Information theory and signal transduction systems: from molecular information processing to network inference.

    PubMed

    Mc Mahon, Siobhan S; Sim, Aaron; Filippi, Sarah; Johnson, Robert; Liepe, Juliane; Smith, Dominic; Stumpf, Michael P H

    2014-11-01

    Sensing and responding to the environment are two essential functions that all biological organisms need to master for survival and successful reproduction. Developmental processes are marshalled by a diverse set of signalling and control systems, ranging from systems with simple chemical inputs and outputs to complex molecular and cellular networks with non-linear dynamics. Information theory provides a powerful and convenient framework in which such systems can be studied; but it also provides the means to reconstruct the structure and dynamics of molecular interaction networks underlying physiological and developmental processes. Here we supply a brief description of its basic concepts and introduce some useful tools for systems and developmental biologists. Along with a brief but thorough theoretical primer, we demonstrate the wide applicability and biological application-specific nuances by way of different illustrative vignettes. In particular, we focus on the characterisation of biological information processing efficiency, examining cell-fate decision making processes, gene regulatory network reconstruction, and efficient signal transduction experimental design. PMID:24953199

  16. Immunological detection of potential signal-transduction proteins expressed during wheat somatic tissue culture.

    PubMed

    Nato, A; Mirshahi, A; Tichtinsky, G; Mirshahi, M; Faure, J P; Lavergne, D; De Buyser, J; Jean, C; Ducreux, G; Henry, Y

    1997-03-01

    An immunochemical approach was used to detect the expression of putative guanine nucleotide-binding proteins (G-proteins), arrestin, and nucleoside diphosphate kinases during wheat (Triticum aestivum) tissue culture initiated from immature embryos. Both the soluble and membrane extracts from the immature embryos revealed bands of 58, 40, and 16 kD with antibodies to G-protein (alpha subunit), arrestin, and nucleoside diphosphate kinase, respectively. These proteins were overexpressed in vitro in both nonembryogenic callus and embryogenic cultures. An additional soluble protein (32 kD) was detected by anti-G alpha antibodies in cultured tissues but not in immature embryos, suggesting a possible function in cell multiplication. Moreover, somatic embryogenesis was associated with the appearance of a 29-kD protein reactive with anti-arrstin antibodies, both in soluble and membrane fractions. Tissue-cultured genetic stocks of Chinese Spring wheat, including the disomic, 36 ditelosomic, and 6 nullisomic-tetrasomic wheat lines, were used to ascertain the chromosomal location of the genes encoding the 29-kD arrestin-like protein. The lack of a signal with the nonembryogenic ditelosomic 3 D short chromosome arm line suggests that the 3 D long chromosome arm possesses at least one gene involved in the expression of the 29-kD protein. The putative role of the 29-kD protein in signal-transduction regulating embryogenesis is discussed. PMID:9085574

  17. Mechanisms of Cardiovascular Homeostasis and Pathophysiology--From Gene Expression, Signal Transduction to Cellular Communication.

    PubMed

    Akazawa, Hiroshi

    2015-01-01

    During embryogenesis, progenitor cells are specified and differentiated into mature cardiomyocytes. Soon after birth, the ability of cardiomyocytes to proliferate is strongly restrained, and thereafter, they grow in size without cell division. Under pathological conditions, cardiomyocytes show adaptive and maladaptive responses through complex intracellular signaling pathways and cross-talking networks of intercellular and inter-tissue communications, but ultimately, they become dysfunctional and undergo cell death or degeneration. Cardiovascular diseases remain the most prevalent, costly, disabling, and deadly medical conditions. To develop novel therapies for them, it is important to elucidate the underlying mechanisms that govern gene expression, signal transduction to cellular communication. In this review article for the 2014 SATO Memorial Award, an approach to uncover molecular and cellular pathophysiology is summarized, focusing on homeobox transcription factor Nkx2-5 in the transcriptional regulation of the cardiac gene program, 3-phosphoinositide-dependent kinase-1, in the regulation of postnatal cardiomyocyte growth, survival, and function, angiotensin II type 1 receptor in the development of pathological hypertrophy and remodeling, and mast cell infiltration in the pathogenesis of atrial remodeling and fibrillation. PMID:26538467

  18. Chemosensory and photosensory perception in purple photosynthetic bacteria utilize common signal transduction components.

    PubMed Central

    Jiang, Z Y; Gest, H; Bauer, C E

    1997-01-01

    The chemotaxis gene cluster from the photosynthetic bacterium Rhodospirillum centenum contains five open reading frames (ORFs) that have significant sequence homology to chemotaxis genes from other bacteria. To elucidate the functions of each ORF, we have made various mutations in the gene cluster and analyzed their phenotypic defects. Deletion of the entire che operon (delta che), as well as nonpolar disruptions of cheAY, cheW, and cheR, resulted in a smooth-swimming phenotype, whereas disruption of cheB resulted in a locked tumbly phenotype. Each of these mutants was defective in chemotactic response. Interestingly, disruption of cheY resulted in a slight increase in the frequency of tumbling/reversal with no obvious defects in chemotactic response. In contrast to observations with Escherichia coli and several other bacteria, we found that all of the che mutant cells were capable of differentiating into hyperflagellated swarmer cells when plated on a solid agar surface. When viewed microscopically, the smooth-swimming che mutants exhibited active surface motility but were unable to respond to a step-down in light intensity. Both positive and negative phototactic responses were abolished in all che mutants, including the cheY mutant. These results indicate that eubacterial photosensory perception is mediated by light-generated signals that are transmitted through the chemotaxis signal transduction cascade. PMID:9294427

  19. The effect of rotation on function and signal transduction in immune cell

    NASA Astrophysics Data System (ADS)

    Song, J. P.; Zhong, P.; Li, Y. H.; Yang, F.

    Objective Both spaceflight and modeled weightlessness on ground could compromise immune function especially cellular immunity In turn astrouants would not resist to external pathogen effectually the health status and work ability of astrounants were perhaps affected but the cellular and molecular mechanisms by which spaceflight alters human immune functions are poorly understood The aim this trial was to using high aspect rotation vessal HARV investigate the functional changes of immune cell rotated for virous time period in vitro and explore mechanisms in which space weightlessness affect immune function through cell signal transduction Methods Using high aspect rotation vessal HARV as simulated weightlessness model mouse splenic lymphocyte and Jurkat E6 1 as cell model the effects of rotation on cell proliferation cytokine secretion expression and activation of signal molecule ZAP-70 were studied Results After rotation T lymphocytic proliferation in mouse splenocyte were inhibited and the concentration of IL-2 and IFN- A secreted were reduced markly and all this happen within 6 hours after T cell were activated The activity of ZAP-70 in Jurkat cell were repressed significantly Conclusion Incapable activation of ZAP-70 might be one cause of depressed lymphocyte function under weightlessness

  20. Immunological detection of potential signal-transduction proteins expressed during wheat somatic tissue culture.

    PubMed Central

    Nato, A; Mirshahi, A; Tichtinsky, G; Mirshahi, M; Faure, J P; Lavergne, D; De Buyser, J; Jean, C; Ducreux, G; Henry, Y

    1997-01-01

    An immunochemical approach was used to detect the expression of putative guanine nucleotide-binding proteins (G-proteins), arrestin, and nucleoside diphosphate kinases during wheat (Triticum aestivum) tissue culture initiated from immature embryos. Both the soluble and membrane extracts from the immature embryos revealed bands of 58, 40, and 16 kD with antibodies to G-protein (alpha subunit), arrestin, and nucleoside diphosphate kinase, respectively. These proteins were overexpressed in vitro in both nonembryogenic callus and embryogenic cultures. An additional soluble protein (32 kD) was detected by anti-G alpha antibodies in cultured tissues but not in immature embryos, suggesting a possible function in cell multiplication. Moreover, somatic embryogenesis was associated with the appearance of a 29-kD protein reactive with anti-arrstin antibodies, both in soluble and membrane fractions. Tissue-cultured genetic stocks of Chinese Spring wheat, including the disomic, 36 ditelosomic, and 6 nullisomic-tetrasomic wheat lines, were used to ascertain the chromosomal location of the genes encoding the 29-kD arrestin-like protein. The lack of a signal with the nonembryogenic ditelosomic 3 D short chromosome arm line suggests that the 3 D long chromosome arm possesses at least one gene involved in the expression of the 29-kD protein. The putative role of the 29-kD protein in signal-transduction regulating embryogenesis is discussed. PMID:9085574

  1. Signal transduction in light–oxygen–voltage receptors lacking the adduct-forming cysteine residue

    PubMed Central

    Yee, Estella F.; Diensthuber, Ralph P.; Vaidya, Anand T.; Borbat, Peter P.; Engelhard, Christopher; Freed, Jack H.; Bittl, Robert; Möglich, Andreas; Crane, Brian R.

    2015-01-01

    Light–oxygen–voltage (LOV) receptors sense blue light through the photochemical generation of a covalent adduct between a flavin-nucleotide chromophore and a strictly conserved cysteine residue. Here we show that, after cysteine removal, the circadian-clock LOV-protein Vivid still undergoes light-induced dimerization and signalling because of flavin photoreduction to the neutral semiquinone (NSQ). Similarly, photoreduction of the engineered LOV histidine kinase YF1 to the NSQ modulates activity and downstream effects on gene expression. Signal transduction in both proteins hence hinges on flavin protonation, which is common to both the cysteinyl adduct and the NSQ. This general mechanism is also conserved by natural cysteine-less, LOV-like regulators that respond to chemical or photoreduction of their flavin cofactors. As LOV proteins can react to light even when devoid of the adduct-forming cysteine, modern LOV photoreceptors may have arisen from ancestral redox-active flavoproteins. The ability to tune LOV reactivity through photoreduction may have important implications for LOV mechanism and optogenetic applications. PMID:26648256

  2. Small Molecule Inhibition of Ligand-Stimulated RAGE-DIAPH1 Signal Transduction

    PubMed Central

    Manigrasso, Michaele B.; Pan, Jinhong; Rai, Vivek; Zhang, Jinghua; Reverdatto, Sergey; Quadri, Nosirudeen; DeVita, Robert J.; Ramasamy, Ravichandran; Shekhtman, Alexander; Schmidt, Ann Marie

    2016-01-01

    The receptor for advanced glycation endproducts (RAGE) binds diverse ligands linked to chronic inflammation and disease. NMR spectroscopy and x-ray crystallization studies of the extracellular domains of RAGE indicate that RAGE ligands bind by distinct charge- and hydrophobicity-dependent mechanisms. The cytoplasmic tail (ct) of RAGE is essential for RAGE ligand-mediated signal transduction and consequent modulation of gene expression and cellular properties. RAGE signaling requires interaction of ctRAGE with the intracellular effector, mammalian diaphanous 1 or DIAPH1. We screened a library of 58,000 small molecules and identified 13 small molecule competitive inhibitors of ctRAGE interaction with DIAPH1. These compounds, which exhibit in vitro and in vivo inhibition of RAGE-dependent molecular processes, present attractive molecular scaffolds for the development of therapeutics against RAGE-mediated diseases, such as those linked to diabetic complications, Alzheimer’s disease, and chronic inflammation, and provide support for the feasibility of inhibition of protein-protein interaction (PPI). PMID:26936329

  3. Control of Streptococcus pyogenes virulence: modeling of the CovR/S signal transduction system.

    PubMed

    Mitrophanov, Alexander Y; Churchward, Gordon; Borodovsky, Mark

    2007-05-01

    The CovR/S system in Streptococcus pyogenes (Group A Streptococcus, or GAS), a two-component signal transduction/transcription regulation system, controls the expression of major virulence factors. The presence of a negative feedback loop distinguishes the CovR/S system from the majority of bacterial two-component systems. We developed a deterministic model of the CovR/S system consisting of eight delay differential equations. Computational experiments showed that the system possessed a unique stable steady state. The dynamical behavior of the system showed a tendency for oscillations becoming more pronounced for longer but still biochemically realistic delays resulting from reductions in the rates of translation elongation. We have devised an efficient procedure for computing the system's steady state. Further, we have shown that the signal-response curves are hyperbolic for the default parameter values. However, in experiments with randomized parameters we demonstrated that sigmoidality of signal-response curves, implying a response threshold, is not only possible, but seems to be rather typical for CovR/S-like systems even when binding of the CovR response regulator protein to a promoter is non-cooperative. We used sensitivity analysis to simplify the model in order to make it analytically tractable. The existence and uniqueness of the steady state and hyperbolicity of signal-response curves for the majority of the variables was proved for the simplified model. Also, we found that provided CovS was active, the system was insensitive to changes in the concentration of any other phosphoryl donor such as acetyl phosphate. PMID:17240398

  4. Converting mid-infrared signals to near-infrared through optomechanical transduction

    NASA Astrophysics Data System (ADS)

    Kapsalis, A.; Mesaritakis, C.; Bogris, A.; Syvridis, D.

    2015-01-01

    Mid-infrared silicon photonics emerge as the dominant technology to bridge photonics and electronics in multifunctional high-speed integrated chips. The transmission and processing of optical signals lying at the mid-infrared wavelength region is ideal for sensing, absorption-spectroscopy and free-space communications and the use of group IV materials becomes principally promising as the vehicle towards their realization. In parallel, optical forces originating from modes and cavities can reach to outstandingly large values when sizes drop into the nanoscale. In this work, we propose the exploitation of large gradient optical forces generated between suspended silicon beams and optomechanical transduction as a means of converting signals from the mid-infrared to the near-infrared region. A midinfrared signal is injected into the waveguide system so as to excite the fundamental symmetric mode. In the 2-5μm wavelength range, separation gaps in the 100nm order and waveguide widths ranging from 300-600nm, the mode is mostly guided in the air slot between the waveguides which maximizes the optomechanical coupling coefficient and optical force. The resulting attractive force deflects the waveguides and the deflection is linearly dependent on the midinfrared optical power. A simple read-out technique using 1.55μm signals with conventional waveguiding in the directional coupler formed by the two beams is analyzed. A positive conversion efficiency (<0dB) is foreseen for waveguides with suspending lengths up to 150μm. The converter could be ideal for use in sensing and spectroscopy rendering the inefficient mid-infrared detectors obsolete. The low-index unconventional guiding in mid-infrared could be a key component towards multifunctional lab-on-a-chip devices.

  5. Sustained nitric oxide (NO)-releasing compound reverses dysregulated NO signal transduction in priapism

    PubMed Central

    Lagoda, Gwen; Sezen, Sena F.; Hurt, K. Joseph; Cabrini, Marcelo R.; Mohanty, Dillip K.; Burnett, Arthur L.

    2014-01-01

    We evaluated the therapeutic potential of a sustained nitric oxide (NO)-releasing compound to correct the molecular hallmarks and pathophysiology of priapism, an important but poorly characterized erectile disorder. 1,5-Bis-(dihexyl-N-nitrosoamino)-2,4-dinitrobenzene (C6′) and an inactive form of the compound [1,5-bis-(dihexylamino)-2,4-dinitrobenzene (C6)] were tested in neuronal cell cultures and penile lysates for NO release (Griess assay) and biological activity (cGMP production). The effect of local depot C6′ or C6 was evaluated in mice with a priapic phenotype due to double neuronal and endothelial NO synthase deletion (dNOS−/−) or human sickle hemoglobin transgenic expression (Sickle). Changes in NO signaling molecules and reactive oxygen species (ROS) surrogates were assessed by Western blot. The physiological response after C6′ treatment was assessed using an established model of electrically stimulated penile erection. C6′ generated NO, increased cGMP, and dose dependently increased NO metabolites. C6′ treatment reversed abnormalities in key penile erection signaling molecules, including phosphodiesterase type 5, phosphorylated endothelial nitric oxide synthase, and phosphorylated vasodilator-stimulated phosphoprotein. In Sickle mice, C6′ also attenuated the increased ROS markers gp91phox, 4-hydroxynonenal, and 3-nitrotyrosine. Finally, C6′ corrected the excessive priapic erection response of dNOS−/− mice. Exogenous sustained NO release from C6′ corrects pathological erectile signaling in mouse models of priapism and suggests novel approaches to human therapy.—Lagoda, G., Sezen, S. F., Hurt, K. J., Cabrini, M. R., Mohanty, D. K., Burnett, A. L. Sustained nitric oxide (NO)-releasing compound reverses dysregulated NO signal transduction in priapism. PMID:24076963

  6. Signal transduction pathways in mast cell granule-mediated endothelial cell activation.

    PubMed Central

    Chi, Luqi; Stehno-Bittel, Lisa; Smirnova, Irina; Stechschulte, Daniel J; Dileepan, Kottarappat N

    2003-01-01

    BACKGROUND: We have previously shown that incubation of human endothelial cells with mast cell granules results in potentiation of lipopolysaccharide-induced production of interleukin-6 and interleukin-8. AIMS: The objective of the present study was to identify candidate molecules and signal transduction pathways involved in the synergy between mast cell granules and lipopolysaccharide on endothelial cell activation. METHODS: Human umbilical vein endothelial cells were incubated with rat mast cell granules in the presence and absence of lipopolysaccharide, and IL-6 production was quantified. The status of c-Jun amino-terminal kinase and extracellular signal-regulated kinase 1/2 activation, nuclear factor-kappaB translocation and intracellular calcium levels were determined to identify the mechanism of synergy between mast cell granules and lipopolysaccaride. RESULTS: Mast cell granules induced low levels of interleukin-6 production by endothelial cells, and this effect was markedly enhanced by lipopolysaccharide. The results revealed that both serine proteases and histamine present in mast cell granules were involved in this activation process. Mast cell granules increased intracellular calcium, and activated c-Jun amino-terminal kinase and extracellular signal-regulated kinase 1/2. The combination of lipopolysaccharide and mast cell granules prolonged c-Jun amino-terminal kinase activity beyond the duration of induction by either stimulant alone and was entirely due to active proteases. However, both proteases and histamine contributed to calcium mobilization and extracellular signal-regulated kinase 1/2 activation. The nuclear translocation of nuclear factor-kappaB proteins was of greater magnitude in endothelial cells treated with the combination of mast cell granules and lipopolysaccharide. CONCLUSIONS:Mast cell granule serine proteases and histamine can amplify lipopolysaccharide-induced endothelial cell activation, which involves calcium mobilization, mitogen

  7. Crosstalk Between MAPK/ERK and PI3K/AKT Signal Pathways During Brain Ischemia/Reperfusion

    PubMed Central

    Zhou, Jing; Du, Ting; Li, Baoman; Rong, Yan; Verkhratsky, Alexei

    2015-01-01

    The epidermal growth factor receptor (EGFR) is linked to the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) and Raf/mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK1/2) signaling pathways. During brain ischemia/reperfusion, EGFR could be transactivated, which stimulates these intracellular signaling cascades that either protect cells or potentiate cell injury. In the present study, we investigated the activation of EGFR, PI3K/AKT, and Raf/MAPK/ERK1/2 during ischemia or reperfusion of the brain using the middle cerebral artery occlusion model. We found that EGFR was phosphorylated and transactivated during both ischemia and reperfusion periods. During ischemia, the activity of PI3K/AKT pathway was significantly increased, as judged from the strong phosphorylation of AKT; this activation was suppressed by the inhibitors of EGFR and Zn-dependent metalloproteinase. Ischemia, however, did not induce ERK1/2 phosphorylation, which was dependent on reperfusion. Coimmunoprecipitation of Son of sevenless 1 (SOS1) with EGFR showed increased association between the receptor and SOS1 in ischemia, indicating the inhibitory node downstream of SOS1. The inhibitory phosphorylation site of Raf-1 at Ser259, but not its stimulatory phosphorylation site at Ser338, was phosphorylated during ischemia. Furthermore, ischemia prompted the interaction between Raf-1 and AKT, while both the inhibitors of PI3K and AKT not only abolished AKT phosphorylation but also restored ERK1/2 phosphorylation. All these findings suggest that Raf/MAPK/ERK1/2 signal pathway is inhibited by AKT via direct phosphorylation and inhibition at Raf-1 node during ischemia. During reperfusion, we observed a significant increase of ERK1/2 phosphorylation but no change in AKT phosphorylation. Inhibitors of reactive oxygen species and phosphatase and tensin homolog restored AKT phosphorylation but abolished ERK1/2 phosphorylation, suggesting that the reactive oxygen species

  8. Effects of RAF inhibitors on PI3K/AKT signalling depend on mutational status of the RAS/RAF signalling axis

    PubMed Central

    Fritsche-Guenther, Raphaela; Witzel, Franziska; Kempa, Stefan; Brummer, Tilman; Sers, Christine; Blüthgen, Nils

    2016-01-01

    Targeted therapies within the RAS/RAF/MEK/ERK signalling axis become increasingly popular, yet cross-talk and feedbacks in the signalling network lead to unexpected effects. Here we look systematically into how inhibiting RAF and MEK with clinically relevant inhibitors result in changes in PI3K/AKT activation. We measure the signalling response using a bead-based ELISA, and use a panel of three cell lines, and isogenic cell lines that express mutant forms of the oncogenes KRAS and BRAF to interrogate the effects of the MEK and RAF inhibitors on signalling. We find that treatment with the RAF inhibitors have opposing effects on AKT phosphorylation depending on the mutational status of two important oncogenes, KRAS and BRAF. If these two genes are in wildtype configuration, RAF inhibitors reduce AKT phosphorylation. In contrast, if BRAF or KRAS are mutant, RAF inhibitors will leave AKT phosphorylation unaffected or lead to an increase of AKT phosphorylation. Down-regulation of phospho-AKT by RAF inhibitors also extends to downstream transcription factors, and correlates with apoptosis induction. Our results show that oncogenes rewire signalling such that targeted therapies can have opposing effects on parallel pathways, which depend on the mutational status of the cell. PMID:26799289

  9. Effects of RAF inhibitors on PI3K/AKT signalling depend on mutational status of the RAS/RAF signalling axis.

    PubMed

    Fritsche-Guenther, Raphaela; Witzel, Franziska; Kempa, Stefan; Brummer, Tilman; Sers, Christine; Blüthgen, Nils

    2016-02-16

    Targeted therapies within the RAS/RAF/MEK/ERK signalling axis become increasingly popular, yet cross-talk and feedbacks in the signalling network lead to unexpected effects. Here we look systematically into how inhibiting RAF and MEK with clinically relevant inhibitors result in changes in PI3K/AKT activation. We measure the signalling response using a bead-based ELISA, and use a panel of three cell lines, and isogenic cell lines that express mutant forms of the oncogenes KRAS and BRAF to interrogate the effects of the MEK and RAF inhibitors on signalling. We find that treatment with the RAF inhibitors have opposing effects on AKT phosphorylation depending on the mutational status of two important oncogenes, KRAS and BRAF. If these two genes are in wildtype configuration, RAF inhibitors reduce AKT phosphorylation. In contrast, if BRAF or KRAS are mutant, RAF inhibitors will leave AKT phosphorylation unaffected or lead to an increase of AKT phosphorylation. Down-regulation of phospho-AKT by RAF inhibitors also extends to downstream transcription factors, and correlates with apoptosis induction. Our results show that oncogenes rewire signalling such that targeted therapies can have opposing effects on parallel pathways, which depend on the mutational status of the cell. PMID:26799289

  10. Akt-mTORC1 signaling regulates Acly to integrate metabolic input to control of macrophage activation

    PubMed Central

    Covarrubias, Anthony J; Aksoylar, Halil Ibrahim; Yu, Jiujiu; Snyder, Nathaniel W; Worth, Andrew J; Iyer, Shankar S; Wang, Jiawei; Ben-Sahra, Issam; Byles, Vanessa; Polynne-Stapornkul, Tiffany; Espinosa, Erika C; Lamming, Dudley; Manning, Brendan D; Zhang, Yijing; Blair, Ian A; Horng, Tiffany

    2016-01-01

    Macrophage activation/polarization to distinct functional states is critically supported by metabolic shifts. How polarizing signals coordinate metabolic and functional reprogramming, and the potential implications for control of macrophage activation, remains poorly understood. Here we show that IL-4 signaling co-opts the Akt-mTORC1 pathway to regulate Acly, a key enzyme in Ac-CoA synthesis, leading to increased histone acetylation and M2 gene induction. Only a subset of M2 genes is controlled in this way, including those regulating cellular proliferation and chemokine production. Moreover, metabolic signals impinge on the Akt-mTORC1 axis for such control of M2 activation. We propose that Akt-mTORC1 signaling calibrates metabolic state to energetically demanding aspects of M2 activation, which may define a new role for metabolism in supporting macrophage activation. DOI: http://dx.doi.org/10.7554/eLife.11612.001 PMID:26894960

  11. Differential regulation of mTOR signaling determines sensitivity to AKT inhibition in diffuse large B cell lymphoma.

    PubMed

    Ezell, Scott A; Wang, Suping; Bihani, Teeru; Lai, Zhongwu; Grosskurth, Shaun E; Tepsuporn, Suprawee; Davies, Barry R; Huszar, Dennis; Byth, Kate F

    2016-02-23

    Agents that target components of the PI3K/AKT/mTOR pathway are under investigation for the treatment of diffuse large B cell lymphoma (DLBCL). Given the highly heterogeneous nature of DLBCL, it is not clear whether all subtypes of DLBCL will be susceptible to PI3K pathway inhibition, or which kinase within this pathway is the most favorable target. Pharmacological profiling of a panel of DLBCL cell lines revealed a subset of DLBCL that was resistant to AKT inhibition. Strikingly, sensitivity to AKT inhibitors correlated with the ability of these inhibitors to block phosphorylation of S6K1 and ribosomal protein S6. Cell lines resistant to AKT inhibition activated S6K1 independent of AKT either through upregulation of PIM2 or through activation by B cell receptor (BCR) signaling components. Finally, combined inhibition of AKT and BTK, PIM2, or S6K1 proved to be an effective strategy to overcome resistance to AKT inhibition in DLBCL. PMID:26824321

  12. Differential regulation of mTOR signaling determines sensitivity to AKT inhibition in diffuse large B cell lymphoma

    PubMed Central

    Ezell, Scott A.; Wang, Suping; Bihani, Teeru; Lai, Zhongwu; Grosskurth, Shaun E.; Tepsuporn, Suprawee; Davies, Barry R.; Huszar, Dennis; Byth, Kate F.

    2016-01-01

    Agents that target components of the PI3K/AKT/mTOR pathway are under investigation for the treatment of diffuse large B cell lymphoma (DLBCL). Given the highly heterogeneous nature of DLBCL, it is not clear whether all subtypes of DLBCL will be susceptible to PI3K pathway inhibition, or which kinase within this pathway is the most favorable target. Pharmacological profiling of a panel of DLBCL cell lines revealed a subset of DLBCL that was resistant to AKT inhibition. Strikingly, sensitivity to AKT inhibitors correlated with the ability of these inhibitors to block phosphorylation of S6K1 and ribosomal protein S6. Cell lines resistant to AKT inhibition activated S6K1 independent of AKT either through upregulation of PIM2 or through activation by B cell receptor (BCR) signaling components. Finally, combined inhibition of AKT and BTK, PIM2, or S6K1 proved to be an effective strategy to overcome resistance to AKT inhibition in DLBCL. PMID:26824321

  13. Pelota Regulates Epidermal Differentiation by Modulating BMP and PI3K/AKT Signaling Pathways.

    PubMed

    Elkenani, Manar; Nyamsuren, Gunsmaa; Raju, Priyadharsini; Liakath-Ali, Kifayathullah; Hamdaoui, Aicha; Kata, Aleksandra; Dressel, Ralf; Klonisch, Thomas; Watt, Fiona M; Engel, Wolfgang; Thliveris, James A; Krishna Pantakani, D V; Adham, Ibrahim M

    2016-08-01

    The depletion of evolutionarily conserved pelota protein causes impaired differentiation of embryonic and spermatogonial stem cells. In this study, we show that temporal deletion of pelota protein before epidermal barrier acquisition leads to neonatal lethality due to perturbations in permeability barrier formation. Further analysis indicated that this phenotype is a result of failed processing of profilaggrin into filaggrin monomers, which promotes the formation of a protective epidermal layer. Molecular analyses showed that pelota protein negatively regulates the activities of bone morphogenetic protein and phosphoinositide 3-kinase (PI3K)/protein kinase B (AKT) signaling pathways in the epidermis. To address whether elevated activities of bone morphogenetic protein and PI3K/AKT signaling pathways were the cause for the perturbed epidermal barrier in Pelo-deficient mice, we made use of organotypic cultures of skin explants from control and mutant embryos at embryonic day 15.5. Inhibition of PI3K/AKT signaling did not significantly affect the bone morphogenetic protein activity. However, inhibition of bone morphogenetic protein signaling caused a significant attenuation of PI3K/AKT activity in mutant skin and, more interestingly, the restoration of profilaggrin processing and normal epidermal barrier function. Therefore, increased activity of the PI3K/AKT signaling pathway in Pelo-deficient skin might conflict with the dephosphorylation of profilaggrin and thereby affect its proper processing into filaggrin monomers and ultimately the epidermal differentiation. PMID:27164299

  14. Leucine minimizes denervation-induced skeletal muscle atrophy of rats through akt/mtor signaling pathways

    PubMed Central

    Ribeiro, Carolina B.; Christofoletti, Daiane C.; Pezolato, Vitor A.; de Cássia Marqueti Durigan, Rita; Prestes, Jonato; Tibana, Ramires A.; Pereira, Elaine C. L.; de Sousa Neto, Ivo V.; Durigan, João L. Q.; da Silva, Carlos A.

    2015-01-01

    The aim of the present study was to evaluate the effect of leucine treatment (0.30 mM) on muscle weight and signaling of myoproteins related to synthesis and degradation pathways of soleus muscle following seven days of complete sciatic nerve lesion. Wistar rats (n = 24) of 3–4 months of age (192 ± 23 g) were used. The animals were randomly distributed into four experimental groups (n = 6/group): control, treated with leucine (L), denervated (D) and denervated treated with leucine (DL). Dependent measures were proteins levels of AKT, AMPK, mTOR, and ACC performed by Western blot. Leucine induced a reduction in the phosphorylation of AMPK (p < 0.05) by 16% in the L and by 68% in the DL groups as compared with control group. Denervation increased AMPK by 24% in the D group as compared with the control group (p < 0.05). AKT was also modulated by denervation and leucine treatment, highlighted by the elevation of AKT phosphorylation in the D (65%), L (98%) and DL (146%) groups as compared with the control group (p < 0.05). AKT phosphorylation was 49% higher in the D group as compared with the DL group. Furthermore, denervation decreased mTOR phosphorylation by 29% in the D group as compared with the control group. However, leucine treatment induced an increase of 49% in the phosphorylation of mTOR in the L group as compared with the control group, and an increase of 154% in the DL as compared with the D group (p < 0.05). ACC phosphorylation was 20% greater in the D group than the control group. Furthermore, ACC in the soleus was 22% lower in the in the L group and 50% lower in the DL group than the respective control group (p < 0.05). In conclusion, leucine treatment minimized the deleterious effects of denervation on rat soleus muscle by increasing anabolic (AKT and mTOR) and decreasing catabolic (AMPK) pathways. These results may be interesting for muscle recovery following acute denervation, which may contribute to musculoskeletal rehabilitation after denervation

  15. Coprinus comatus Cap Inhibits Adipocyte Differentiation via Regulation of PPARγ and Akt Signaling Pathway

    PubMed Central

    Jang, Sun-Hee; Kang, Suk Nam; Jeon, Beong-Sam; Ko, Yeoung-Gyu; Kim, Hong-Duck; Won, Chung-Kil; Kim, Gon-Sup; Cho, Jae-Hyeon

    2014-01-01

    This study assessed the effects of Coprinus comatus cap (CCC) on adipogenesis in 3T3-L1 adipocytes and the effects of CCC on the development of diet-induced obesity in rats. Here, we showed that the CCC has an inhibitory effect on the adipocyte differentiation of 3T3-L1 cells, resulting in a significant decrease in lipid accumulation through the downregulation of several adipocyte specific-transcription factors, including CCAAT/enhancer binding protein β, C/EBPδ, and peroxisome proliferator-activated receptor gamma (PPARγ). Moreover, treatment with CCC during adipocyte differentiation induced a significant down-regulation of PPARγ and adipogenic target genes, including adipocyte protein 2, lipoprotein lipase, and adiponectin. Interestingly, the CCC treatment of the 3T3-L1 adipocytes suppressed the insulin-stimulated Akt and GSK3β phosphorylation, and these effects were stronger in the presence of an inhibitor of Akt phosphorylation, LY294002, suggesting that CCC inhibited adipocyte differentiation through the down-regulation of Akt signaling. In the animal study, CCC administration significantly reduced the body weight and adipose tissue weight of rats fed a high fat diet (HFD) and attenuated lipid accumulation in the adipose tissues of the HFD-induced obese rats. The size of the adipocyte in the epididymal fat of the CCC fed rats was significantly smaller than in the HFD rats. CCC treatment significantly reduced the total cholesterol and triglyceride levels in the serum of HFD rats. These results strongly indicated that the CCC-mediated decrease in body weight was due to a reduction in adipose tissue mass. The expression level of PPARγ and phospho-Akt was significantly lower in the CCC-treated HFD rats than that in the HFD obesity rats. These results suggested that CCC inhibited adipocyte differentiation by the down-regulation of major transcription factor involved in the adipogenesis pathway including PPARγ through the regulation of the Akt pathway in 3T3

  16. Coprinus comatus cap inhibits adipocyte differentiation via regulation of PPARγ and Akt signaling pathway.

    PubMed

    Park, Hyoung Joon; Yun, Jisoo; Jang, Sun-Hee; Kang, Suk Nam; Jeon, Beong-Sam; Ko, Yeoung-Gyu; Kim, Hong-Duck; Won, Chung-Kil; Kim, Gon-Sup; Cho, Jae-Hyeon

    2014-01-01

    This study assessed the effects of Coprinus comatus cap (CCC) on adipogenesis in 3T3-L1 adipocytes and the effects of CCC on the development of diet-induced obesity in rats. Here, we showed that the CCC has an inhibitory effect on the adipocyte differentiation of 3T3-L1 cells, resulting in a significant decrease in lipid accumulation through the downregulation of several adipocyte specific-transcription factors, including CCAAT/enhancer binding protein β, C/EBPδ, and peroxisome proliferator-activated receptor gamma (PPARγ). Moreover, treatment with CCC during adipocyte differentiation induced a significant down-regulation of PPARγ and adipogenic target genes, including adipocyte protein 2, lipoprotein lipase, and adiponectin. Interestingly, the CCC treatment of the 3T3-L1 adipocytes suppressed the insulin-stimulated Akt and GSK3β phosphorylation, and these effects were stronger in the presence of an inhibitor of Akt phosphorylation, LY294002, suggesting that CCC inhibited adipocyte differentiation through the down-regulation of Akt signaling. In the animal study, CCC administration significantly reduced the body weight and adipose tissue weight of rats fed a high fat diet (HFD) and attenuated lipid accumulation in the adipose tissues of the HFD-induced obese rats. The size of the adipocyte in the epididymal fat of the CCC fed rats was significantly smaller than in the HFD rats. CCC treatment significantly reduced the total cholesterol and triglyceride levels in the serum of HFD rats. These results strongly indicated that the CCC-mediated decrease in body weight was due to a reduction in adipose tissue mass. The expression level of PPARγ and phospho-Akt was significantly lower in the CCC-treated HFD rats than that in the HFD obesity rats. These results suggested that CCC inhibited adipocyte differentiation by the down-regulation of major transcription factor involved in the adipogenesis pathway including PPARγ through the regulation of the Akt pathway in 3T3

  17. Mechanism of 2-oxoglutarate signaling by the Synechococcus elongatus PII signal transduction protein

    PubMed Central

    Fokina, Oleksandra; Chellamuthu, Vasuki-Ranjani; Forchhammer, Karl; Zeth, Kornelius

    2010-01-01

    PII proteins control key processes of nitrogen metabolism in bacteria, archaea, and plants in response to the central metabolites ATP, ADP, and 2-oxoglutarate (2-OG), signaling cellular energy and carbon and nitrogen abundance. This metabolic information is integrated by PII and transmitted to regulatory targets (key enzymes, transporters, and transcription factors), modulating their activity. In oxygenic phototrophs, the controlling enzyme of arginine synthesis, N-acetyl-glutamate kinase (NAGK), is a major PII target, whose activity responds to 2-OG via PII. Here we show structures of the Synechococcus elongatus PII protein in complex with ATP, Mg2+, and 2-OG, which clarify how 2-OG affects PII–NAGK interaction. PII trimers with all three sites fully occupied were obtained as well as structures with one or two 2-OG molecules per PII trimer. These structures identify the site of 2-OG located in the vicinity between the subunit clefts and the base of the T loop. The 2-OG is bound to a Mg2+ ion, which is coordinated by three phosphates of ATP, and by ionic interactions with the highly conserved residues K58 and Q39 together with B- and T-loop backbone interactions. These interactions impose a unique T-loop conformation that affects the interactions with the PII target. Structures of PII trimers with one or two bound 2-OG molecules reveal the basis for anticooperative 2-OG binding and shed light on the intersubunit signaling mechanism by which PII senses effectors in a wide range of concentrations. PMID:21041661

  18. Mechanism of 2-oxoglutarate signaling by the Synechococcus elongatus PII signal transduction protein.

    PubMed

    Fokina, Oleksandra; Chellamuthu, Vasuki-Ranjani; Forchhammer, Karl; Zeth, Kornelius

    2010-11-16

    P(II) proteins control key processes of nitrogen metabolism in bacteria, archaea, and plants in response to the central metabolites ATP, ADP, and 2-oxoglutarate (2-OG), signaling cellular energy and carbon and nitrogen abundance. This metabolic information is integrated by P(II) and transmitted to regulatory targets (key enzymes, transporters, and transcription factors), modulating their activity. In oxygenic phototrophs, the controlling enzyme of arginine synthesis, N-acetyl-glutamate kinase (NAGK), is a major P(II) target, whose activity responds to 2-OG via P(II). Here we show structures of the Synechococcus elongatus P(II) protein in complex with ATP, Mg(2+), and 2-OG, which clarify how 2-OG affects P(II)-NAGK interaction. P(II) trimers with all three sites fully occupied were obtained as well as structures with one or two 2-OG molecules per P(II) trimer. These structures identify the site of 2-OG located in the vicinity between the subunit clefts and the base of the T loop. The 2-OG is bound to a Mg(2+) ion, which is coordinated by three phosphates of ATP, and by ionic interactions with the highly conserved residues K58 and Q39 together with B- and T-loop backbone interactions. These interactions impose a unique T-loop conformation that affects the interactions with the P(II) target. Structures of P(II) trimers with one or two bound 2-OG molecules reveal the basis for anticooperative 2-OG binding and shed light on the intersubunit signaling mechanism by which P(II) senses effectors in a wide range of concentrations. PMID:21041661

  19. Phosphodiesterase 4D acts downstream of Neuropilin to control Hedgehog signal transduction and the growth of medulloblastoma.

    PubMed

    Ge, Xuecai; Milenkovic, Ljiljana; Suyama, Kaye; Hartl, Tom; Purzner, Teresa; Winans, Amy; Meyer, Tobias; Scott, Matthew P

    2015-01-01

    Alterations in Hedgehog (Hh) signaling lead to birth defects and cancers including medulloblastoma, the most common pediatric brain tumor. Although inhibitors targeting the membrane protein Smoothened suppress Hh signaling, acquired drug resistance and tumor relapse call for additional therapeutic targets. Here we show that phosphodiesterase 4D (PDE4D) acts downstream of Neuropilins to control Hh transduction and medulloblastoma growth. PDE4D interacts directly with Neuropilins, positive regulators of Hh pathway. The Neuropilin ligand Semaphorin3 enhances this interaction, promoting PDE4D translocation to the plasma membrane and cAMP degradation. The consequent inhibition of protein kinase A (PKA) enhances Hh transduction. In the developing cerebellum, genetic removal of Neuropilins reduces Hh signaling activity and suppresses proliferation of granule neuron precursors. In mouse medulloblastoma allografts, PDE4D inhibitors suppress Hh transduction and inhibit tumor growth. Our findings reveal a new regulatory mechanism of Hh transduction, and highlight PDE4D as a promising target to treat Hh-related tumors. PMID:26371509

  20. Phosphodiesterase 4D acts downstream of Neuropilin to control Hedgehog signal transduction and the growth of medulloblastoma

    PubMed Central

    Ge, Xuecai; Milenkovic, Ljiljana; Suyama, Kaye; Hartl, Tom; Purzner, Teresa; Winans, Amy; Meyer, Tobias; Scott, Matthew P

    2015-01-01

    Alterations in Hedgehog (Hh) signaling lead to birth defects and cancers including medulloblastoma, the most common pediatric brain tumor. Although inhibitors targeting the membrane protein Smoothened suppress Hh signaling, acquired drug resistance and tumor relapse call for additional therapeutic targets. Here we show that phosphodiesterase 4D (PDE4D) acts downstream of Neuropilins to control Hh transduction and medulloblastoma growth. PDE4D interacts directly with Neuropilins, positive regulators of Hh pathway. The Neuropilin ligand Semaphorin3 enhances this interaction, promoting PDE4D translocation to the plasma membrane and cAMP degradation. The consequent inhibition of protein kinase A (PKA) enhances Hh transduction. In the developing cerebellum, genetic removal of Neuropilins reduces Hh signaling activity and suppresses proliferation of granule neuron precursors. In mouse medulloblastoma allografts, PDE4D inhibitors suppress Hh transduction and inhibit tumor growth. Our findings reveal a new regulatory mechanism of Hh transduction, and highlight PDE4D as a promising target to treat Hh-related tumors. DOI: http://dx.doi.org/10.7554/eLife.07068.001 PMID:26371509

  1. Cerebral Artery Signal Transduction Mechanisms: Developmental Changes in Dynamics and Ca2+ Sensitivity

    PubMed Central

    Longo, Lawrence D.; Goyal, Ravi

    2012-01-01

    As compared to the adult, the developing fetus and newborn infant are at much greater risk for dysregulation of cerebral blood flow (CBF), with complications such as intraventricular and germinal matrix hemorrhage with resultant neurologic sequelae. To minimize this dysregulation and its consequences presents a major challenge. Although in many respects the fundamental signal transduction mechanisms that regulate relaxation and contraction pathways, and thus cerebrovascular tone and CBF in the immature organism are similar to those of the adult, the individual elements, pathways, and roles differ greatly. Here, we review aspects of these maturational changes of relaxation/contraction mechanisms in terms of both electro-mechanical and pharmaco-mechanical coupling, their biochemical pathways and signaling networks. In contrast to the adult cerebrovasculature, in addition to attenuated structure with differences in multiple cytoskeletal elements, developing cerebrovasculature of fetus and newborn differs in many respects, such as a strikingly increased sensitivity to [Ca2+]i and requirement for extracellular Ca2+ for contraction. In essence, the immature cerebrovasculature demonstrates both “hyper-relaxation” and “hypo-contraction”. A challenge is to unravel the manner in which these mechanisms are integrated, particularly in terms of both Ca2+-dependent and Ca2+-independent pathways to increase Ca2+ sensitivity. Gaining an appreciation of these significant age-related differences in signal mechanisms also will be critical to understanding more completely the vulnerability of the developing cerebral vasculature to hypoxia and other stresses. Of vital importance, a more complete understanding of these mechanisms promises hope for improved strategies for therapeutic intervention and clinical management of intensive care of the premature newborn. PMID:24063382

  2. Programmable Ligand Detection System in Plants through a Synthetic Signal Transduction Pathway

    PubMed Central

    Smith, J. Jeff; Albrecht, Kirk D.; Bowen, Tessa A.; Zdunek, Jeffrey K.; Troupe, Jared F.; Cuneo, Matthew J.; Webb, Colleen T.; Hellinga, Homme W.; Medford, June I.

    2011-01-01

    Background There is an unmet need to monitor human and natural environments for substances that are intentionally or unintentionally introduced. A long-sought goal is to adapt plants to sense and respond to specific substances for use as environmental monitors. Computationally re-designed periplasmic binding proteins (PBPs) provide a means to design highly sensitive and specific ligand sensing capabilities in receptors. Input from these proteins can be linked to gene expression through histidine kinase (HK) mediated signaling. Components of HK signaling systems are evolutionarily conserved between bacteria and plants. We previously reported that in response to cytokinin-mediated HK activation in plants, the bacterial response regulator PhoB translocates to the nucleus and activates transcription. Also, we previously described a plant visual response system, the de-greening circuit, a threshold sensitive reporter system that produces a visual response which is remotely detectable and quantifiable. Methodology/Principal Findings We describe assembly and function of a complete synthetic signal transduction pathway in plants that links input from computationally re-designed PBPs to a visual response. To sense extracellular ligands, we targeted the computational re-designed PBPs to the apoplast. PBPs bind the ligand and develop affinity for the extracellular domain of a chemotactic protein, Trg. We experimentally developed Trg fusions proteins, which bind the ligand-PBP complex, and activate intracellular PhoR, the HK cognate of PhoB. We then adapted Trg-PhoR fusions for function in plants showing that in the presence of an external ligand PhoB translocates to the nucleus and activates transcription. We linked this input to the de-greening circuit creating a detector plant. Conclusions/Significance Our system is modular and PBPs can theoretically be designed to bind most small molecules. Hence our system, with improvements, may allow plants to serve as a simple and

  3. Backbone NMR reveals allosteric signal transduction networks in the β1-adrenergic receptor.

    PubMed

    Isogai, Shin; Deupi, Xavier; Opitz, Christian; Heydenreich, Franziska M; Tsai, Ching-Ju; Brueckner, Florian; Schertler, Gebhard F X; Veprintsev, Dmitry B; Grzesiek, Stephan

    2016-02-11

    G protein-coupled receptors (GPCRs) are physiologically important transmembrane signalling proteins that trigger intracellular responses upon binding of extracellular ligands. Despite recent breakthroughs in GPCR crystallography, the details of ligand-induced signal transduction are not well understood owing to missing dynamical information. In principle, such information can be provided by NMR, but so far only limited data of functional relevance on few side-chain sites of eukaryotic GPCRs have been obtained. Here we show that receptor motions can be followed at virtually any backbone site in a thermostabilized mutant of the turkey β1-adrenergic receptor (β1AR). Labelling with [(15)N]valine in a eukaryotic expression system provides over twenty resolved resonances that report on structure and dynamics in six ligand complexes and the apo form. The response to the various ligands is heterogeneous in the vicinity of the binding pocket, but gets transformed into a homogeneous readout at the intracellular side of helix 5 (TM5), which correlates linearly with ligand efficacy for the G protein pathway. The effect of several pertinent, thermostabilizing point mutations was assessed by reverting them to the native sequence. Whereas the response to ligands remains largely unchanged, binding of the G protein mimetic nanobody NB80 and G protein activation are only observed when two conserved tyrosines (Y227 and Y343) are restored. Binding of NB80 leads to very strong spectral changes throughout the receptor, including the extracellular ligand entrance pocket. This indicates that even the fully thermostabilized receptor undergoes activating motions in TM5, but that the fully active state is only reached in presence of Y227 and Y343 by stabilization with a G protein-like partner. The combined analysis of chemical shift changes from the point mutations and ligand responses identifies crucial connections in the allosteric activation pathway, and presents a general experimental

  4. Ubiquitin-Related Roles of β-Arrestins in Endocytic Trafficking and Signal Transduction.

    PubMed

    Jean-Charles, Pierre-Yves; Rajiv, Vishwaesh; Shenoy, Sudha K

    2016-10-01

    The non-visual arrestins, β-arrestin1, and β-arrestin2 were originally identified as proteins that bind to seven-transmembrane receptors (7TMRs, also called G protein-coupled receptors, GPCRs) and block heterotrimeric G protein activation, thus leading to desensitization of transmembrane signaling. However, as subsequent discoveries have continually demonstrated, their functionality is not constrained to desensitization. They are now recognized for their critical roles in mediating intracellular trafficking of 7TMRs, growth factor receptors, ion transporters, ion channels, nuclear receptors, and non-receptor proteins. Additionally, they function as crucial mediators of ubiquitination of 7TMRs as well as other receptors and non-receptor proteins. Recently, emerging studies suggest that a class of proteins with predicted structural features of β-arrestins regulate substrate ubiquitination in yeast and higher mammals, lending support to the idea that the adaptor role of β-arrestins in protein ubiquitination is evolutionarily conserved. β-arrestins also function as scaffolds for kinases and transduce signals from 7TMRs through pathways that do not require G protein activation. Remarkably, the endocytic and scaffolding functions of β-arrestin are intertwined with its ubiquitination status; the dynamic and site specific ubiquitination on β-arrestin plays a critical role in stabilizing β-arrestin-7TMR association and the formation of signalosomes. This review summarizes the current findings on ubiquitin-dependent regulation of 7TMRs as well as β-arrestins and the potential role of reversible ubiquitination as a "biological switch" in signal transduction. J. Cell. Physiol. 231: 2071-2080, 2016. © 2016 Wiley Periodicals, Inc. PMID:26790995

  5. Cerebral artery signal transduction mechanisms: developmental changes in dynamics and Ca2+ sensitivity.

    PubMed

    Longo, Lawrence D; Goyal, Ravi

    2013-09-01

    As compared to the adult, the developing fetus and newborn infant are at much greater risk for dysregulation of cerebral blood flow (CBF), with complications such as intraventricular and germinal matrix hemorrhage with resultant neurologic sequelae. To minimize this dysregulation and its consequences presents a major challenge. Although in many respects the fundamental signal transduction mechanisms that regulate relaxation and contraction pathways, and thus cerebrovascular tone and CBF in the immature organism are similar to those of the adult, the individual elements, pathways, and roles differ greatly. Here, we review aspects of these maturational changes of relaxation/contraction mechanisms in terms of both electro-mechanical and pharmaco-mechanical coupling, their biochemical pathways and signaling networks. In contrast to the adult cerebrovasculature, in addition to attenuated structure with differences in multiple cytoskeletal elements, developing cerebrovasculature of fetus and newborn differs in many respects, such as a strikingly increased sensitivity to [Ca(2+)]i and requirement for extracellular Ca(2+) for contraction. In essence, the immature cerebrovasculature demonstrates both "hyper-relaxation" and "hypo-contraction". A challenge is to unravel the manner in which these mechanisms are integrated, particularly in terms of both Ca(2+)-dependent and Ca(2+)-independent pathways to increase Ca(2+) sensitivity. Gaining an appreciation of these significant age-related differences in signal mechanisms also will be critical to understanding more completely the vulnerability of the developing cerebral vasculature to hypoxia and other stresses. Of vital importance, a more complete understanding of these mechanisms promises hope for improved strategies for therapeutic intervention and clinical management of intensive care of the premature newborn. PMID:24063382

  6. The allosteric behavior of Fur mediates oxidative stress signal transduction in Helicobacter pylori.

    PubMed

    Pelliciari, Simone; Vannini, Andrea; Roncarati, Davide; Danielli, Alberto

    2015-01-01

    The microaerophilic gastric pathogen Helicobacter pylori is exposed to oxidative stress originating from the aerobic environment, the oxidative burst of phagocytes and the formation of reactive oxygen species, catalyzed by iron excess. Accordingly, the expression of genes involved in oxidative stress defense have been repeatedly linked to the ferric uptake regulator Fur. Moreover, mutations in the Fur protein affect the resistance to metronidazole, likely due to loss-of-function in the regulation of genes involved in redox control. Although many advances in the molecular understanding of HpFur function were made, little is known about the mechanisms that enable Fur to mediate the responses to oxidative stress. Here we show that iron-inducible, apo-Fur repressed genes, such as pfr and hydA, are induced shortly after oxidative stress, while their oxidative induction is lost in a fur knockout strain. On the contrary, holo-Fur repressed genes, such as frpB1 and fecA1, vary modestly in response to oxidative stress. This indicates that the oxidative stress signal specifically targets apo-Fur repressed genes, rather than impairing indiscriminately the regulatory function of Fur. Footprinting analyses showed that the oxidative signal strongly impairs the binding affinity of Fur toward apo-operators, while the binding toward holo-operators is less affected. Further evidence is presented that a reduced state of Fur is needed to maintain apo-repression, while oxidative conditions shift the preferred binding architecture of Fur toward the holo-operator binding conformation, even in the absence of iron. Together the results demonstrate that the allosteric regulation of Fur enables transduction of oxidative stress signals in H. pylori, supporting the concept that apo-Fur repressed genes can be considered oxidation inducible Fur regulatory targets. These findings may have important implications in the study of H. pylori treatment and resistance to antibiotics. PMID:26347726

  7. Regulation of Early Steps of GPVI Signal Transduction by Phosphatases: A Systems Biology Approach

    PubMed Central

    Dunster, Joanne L.; Mazet, Francoise; Fry, Michael J.; Gibbins, Jonathan M.; Tindall, Marcus J.

    2015-01-01

    We present a data-driven mathematical model of a key initiating step in platelet activation, a central process in the prevention of bleeding following Injury. In vascular disease, this process is activated inappropriately and causes thrombosis, heart attacks and stroke. The collagen receptor GPVI is the primary trigger for platelet activation at sites of injury. Understanding the complex molecular mechanisms initiated by this receptor is important for development of more effective antithrombotic medicines. In this work we developed a series of nonlinear ordinary differential equation models that are direct representations of biological hypotheses surrounding the initial steps in GPVI-stimulated signal transduction. At each stage model simulations were compared to our own quantitative, high-temporal experimental data that guides further experimental design, data collection and model refinement. Much is known about the linear forward reactions within platelet signalling pathways but knowledge of the roles of putative reverse reactions are poorly understood. An initial model, that includes a simple constitutively active phosphatase, was unable to explain experimental data. Model revisions, incorporating a complex pathway of interactions (and specifically the phosphatase TULA-2), provided a good description of the experimental data both based on observations of phosphorylation in samples from one donor and in those of a wider population. Our model was used to investigate the levels of proteins involved in regulating the pathway and the effect of low GPVI levels that have been associated with disease. Results indicate a clear separation in healthy and GPVI deficient states in respect of the signalling cascade dynamics associated with Syk tyrosine phosphorylation and activation. Our approach reveals the central importance of this negative feedback pathway that results in the temporal regulation of a specific class of protein tyrosine phosphatases in controlling the rate

  8. Regulation of Early Steps of GPVI Signal Transduction by Phosphatases: A Systems Biology Approach.

    PubMed

    Dunster, Joanne L; Mazet, Francoise; Fry, Michael J; Gibbins, Jonathan M; Tindall, Marcus J

    2015-11-01

    We present a data-driven mathematical model of a key initiating step in platelet activation, a central process in the prevention of bleeding following Injury. In vascular disease, this process is activated inappropriately and causes thrombosis, heart attacks and stroke. The collagen receptor GPVI is the primary trigger for platelet activation at sites of injury. Understanding the complex molecular mechanisms initiated by this receptor is important for development of more effective antithrombotic medicines. In this work we developed a series of nonlinear ordinary differential equation models that are direct representations of biological hypotheses surrounding the initial steps in GPVI-stimulated signal transduction. At each stage model simulations were compared to our own quantitative, high-temporal experimental data that guides further experimental design, data collection and model refinement. Much is known about the linear forward reactions within platelet signalling pathways but knowledge of the roles of putative reverse reactions are poorly understood. An initial model, that includes a simple constitutively active phosphatase, was unable to explain experimental data. Model revisions, incorporating a complex pathway of interactions (and specifically the phosphatase TULA-2), provided a good description of the experimental data both based on observations of phosphorylation in samples from one donor and in those of a wider population. Our model was used to investigate the levels of proteins involved in regulating the pathway and the effect of low GPVI levels that have been associated with disease. Results indicate a clear separation in healthy and GPVI deficient states in respect of the signalling cascade dynamics associated with Syk tyrosine phosphorylation and activation. Our approach reveals the central importance of this negative feedback pathway that results in the temporal regulation of a specific class of protein tyrosine phosphatases in controlling the rate

  9. Focal adhesions are foci for tyrosine-based signal transduction via GIV/Girdin and G proteins

    PubMed Central

    Lopez-Sanchez, Inmaculada; Kalogriopoulos, Nicholas; Lo, I-Chung; Kabir, Firooz; Midde, Krishna K.; Wang, Honghui; Ghosh, Pradipta

    2015-01-01

    GIV/Girdin is a multimodular signal transducer and a bona fide metastasis-related protein. As a guanidine exchange factor (GEF), GIV modulates signals initiated by growth factors (chemical signals) by activating the G protein Gαi. Here we report that mechanical signals triggered by the extracellular matrix (ECM) also converge on GIV-GEF via β1 integrins and that focal adhesions (FAs) serve as the major hubs for mechanochemical signaling via GIV. GIV interacts with focal adhesion kinase (FAK) and ligand-activated β1 integrins. Phosphorylation of GIV by FAK enhances PI3K-Akt signaling, the integrity of FAs, increases cell–ECM adhesion, and triggers ECM-induced cell motility. Activation of Gαi by GIV-GEF further potentiates FAK-GIV-PI3K-Akt signaling at the FAs. Spatially restricted signaling via tyrosine phosphorylated GIV at the FAs is enhanced during cancer metastasis. Thus GIV-GEF serves as a unifying platform for integration and amplification of adhesion (mechanical) and growth factor (chemical) signals during cancer progression. PMID:26446841

  10. Immunohistochemical analysis of the Akt/mTOR/4E-BP1 signalling pathway in canine haemangiomas and haemangiosarcomas.

    PubMed

    Murai, A; Abou Asa, S; Kodama, A; Sakai, H; Hirata, A; Yanai, T

    2012-11-01

    The specific signalling pathways that are deregulated in canine endothelial tumours have not yet fully elucidated. Therefore, the aim of the present study was to examine activation of the Akt/mammalian target of rapamycin (mTOR)/eukaryotic initiation factor 4E-binding protein 1 (4E-BP1) signalling pathway in spontaneously arising canine haemangiomas (HAs) and haemangiosarcomas (HSAs) in order to identify novel molecular targets for treatment. Surgically-resected samples of HA (n = 27), HSA (n = 37), granulation tissue (n = 4) and normal skin (n = 4) were investigated by immunohistochemistry. Approximately 80% of the HSA samples had moderate to intense expression of phosphorylated Akt at Ser473 (p-Akt Ser473), p-Akt Thr308, p-4E-BP1 Thr37/46 and eukaryotic initiation factor 4E, which was significantly higher than in the HAs and was similar to the expression in activated endothelial cells (ECs). Although p-mTOR complex1 (p-mTORC1) Ser2448 was expressed by most of the activated ECs, only 35% of the HSA samples had weak to moderate expression. Because mTORC2 and phosphorylates Akt Ser473 was activated in HSA samples, the present findings suggest that the mTORC2/Akt/4E-BP1 pathway, regulated independently of mTORC1, may be important for targeting therapy in canine HSAs. PMID:22789858

  11. Titration of signalling output: insights into clinical combinations of MEK and AKT inhibitors

    PubMed Central

    Stewart, A.; Thavasu, P.; de Bono, J. S.; Banerji, U.

    2015-01-01

    Background We aimed to understand the relative contributions of inhibiting MEK and AKT on cell growth to guide combinations of these agents. Materials and methods A panel of 20 cell lines was exposed to either the MEK inhibitor, PD0325901, or AKT inhibitor, AKT 1/2 inhibitor. p-ERK and p-S6 ELISAs were used to define degrees of MEK and AKT inhibition, respectively. Growth inhibition to different degrees of MEK and AKT inhibition, either singly or in combination using 96-h sulphorhodamine assays was then studied. Results A significantly greater growth inhibition was seen in BRAFM and PIK3CAM cells upon maximal MEK (P = 0.004) and AKT inhibition (P = 0.038), respectively. KRASM and BRAF/PIK3CA/KRASWT cells were not significantly more likely to be sensitive to MEK or AKT inhibition. Significant incremental growth inhibition of the combination of MEK + AKT over either MEK or AKT inhibition alone was seen when MEK + AKT was inhibited maximally and not when sub-maximal inhibition of both MEK + AKT was used (11/20 cell lines versus 1/20 cell lines; P = 0.0012). Conclusions KRASM cells are likely to benefit from combinations of MEK and AKT inhibitors. Sub-maximally inhibiting both MEK and AKT within a combination, in a majority of instances, does not significantly increase growth inhibition compared with maximally inhibiting MEK or AKT alone and alternative phase I trial designs are needed to clinically evaluate such combinations. PMID:25908604

  12. PI3K/Akt signaling pathway is involved in the neurotrophic effect of senegenin.

    PubMed

    Pi, Ting; Zhou, Xiao-Wen; Cai, Liang; Zhang, Wei; Su, Chao-Fen; Wu, Wu-Tian; Ren, Xiao-Ming; Luo, Huan-Min

    2016-02-01

    Neurodegenerative diseases are frequently associated with the loss of synapses and neurons. Senegenin, extracted from the Chinese herb Polygala tenuifolia Willd, was previously found to promote neurite outgrowth and neuronal survival in primary cultured rat cortical neurons. The aim of the present study was to investigate the underlying mechanisms of senegenin-induced neurotrophic effects on rat cortical neurons. Primary cortical rat neurons were treated with various pharmacological antagonists and with or without senegenin, and subjected to MTT and western blot analysis to explore the effects of senegenin on cell survival as well as the activation of signaling pathways. Neurite outgrowth and neuronal survival induced by senegenin were significantly inhibited by A2A receptor antagonist ZM241385 and specific phosphoinositide-3 kinase (PI3K) inhibitor LY294002, but not by tropomyosin receptor kinase A receptor inhibitor K252a, mitogen-activated protein kinase kinase inhibitor PD98059 or protein kinase C inhibitor GÖ6976. Furthermore, senegenin enhanced the phosphorylation of Akt, which was blocked by LY294002. The present study revealed that the PI3K/Akt signaling pathway may be involved in the neurotrophic effects of senegenin. PMID:26647727

  13. Progranulin promotes colorectal cancer proliferation and angiogenesis through TNFR2/Akt and ERK signaling pathways

    PubMed Central

    Yang, Dong; Wang, Lin-Lin; Dong, Tao-Tao; Shen, Yi-Hang; Guo, Xiao-Sun; Liu, Chuan-Yong; Liu, Jie; Zhang, Pei; Li, Juan; Sun, Yu-Ping

    2015-01-01

    Progranulin (PGRN) has been shown to be involved in the process of inflammation, wound healing, and cartilage development; and its role in the progression of breast and ovarian cancer is also well established. However, the expression status of PGRN in colorectal cancers (CRCs) and its molecular mechanisms responsible for tumorigenesis have not been addressed so far. Herein, we demonstrated that PGRN was highly expressed and had clinical relevance with CRCs since its overexpression was associated with advanced stages of CRCs, poorer patients’ prognosis, and increased expression of proliferation and angiogenesis markers. PGRN up-regulation significantly promoted the expression of Ki67 and vascular endothelial growth factor A (VEGF-A) as well as the growth rate in CRC cell lines, while PGRN down-regulation had the opposite effects. Strikingly, PGRN derived from CRCs could directly induce proliferation, migration, tubule formation, as well as VEGF-A expression in human umbilical vein endothelial cells (HUVECs). Furthermore, we provided mechanistic evidences that the regulation of Ki67 and VEGF-A expression by PGRN was mediated by tumor necrosis factor receptor 2 (TNFR2)/Akt and the ERK signaling pathways in both CRC cells and HUVECs. Taken together, these findings suggested that PGRN could promote proliferation and angiogenesis through TNFR2/Akt and ERK signaling pathways in CRCs, providing the new insight into the mechanism of PGRN in tumor proliferation and angiogenesis. PMID:26693061

  14. Shikonin Derivative DMAKO-05 Inhibits Akt Signal Activation and Melanoma Proliferation.

    PubMed

    Yang, Yao-Yao; He, Hui-Qiong; Cui, Jia-Hua; Nie, Yun-Juan; Wu, Ya-Xian; Wang, Rui; Wang, Gang; Zheng, Jun-Nian; Ye, Richard D; Wu, Qiong; Li, Shao-Shun; Qian, Feng

    2016-06-01

    DMAKO-05((S)-1-((5E,8E)-5,8-bis(hydroxyimino)-1,4-dimethoxy-5,8-dihydronaphthalen-2-yl)-4-methylpent-3-enyl 3-methylbutanoate) is a novel oxime derivative of shikonin, the major component extracted from Chinese herb Lithospermun erythrorhizon. Here, we report that DMAKO-05 had an antitumor activity against mouse melanoma cell line B16F0. Our studies indicated that DMAKO-05 not only inhibited B16F0 proliferation and migration but also led to cell cycle arrest at G1 phase and cell apoptosis, in which DMAKO-05 triggered mitochondrial-mediated apoptosis signal including caspase-9/3 and PARP. In response to DMAKO-05 treatment, the Akt-mediated survival signals were remarkably attenuated in B16F0 cells. Collectively, DMAKO-05 has a strong cytotoxicity in B16F0 cells via inhibiting Akt activation, inducing G1 arrest, and promoting B16F0 cell apoptosis. DMAKO-05 might serve as a potential candidate lead compound for melanoma. PMID:26804061

  15. Akt signaling-associated metabolic effects of dietary gold nanoparticles in Drosophila

    NASA Astrophysics Data System (ADS)

    Wang, Bin; Chen, Nan; Wei, Yingliang; Li, Jiang; Sun, Li; Wu, Jiarui; Huang, Qing; Liu, Chang; Fan, Chunhai; Song, Haiyun

    2012-08-01

    Gold nanoparticles (AuNPs) are often used as vehicles to deliver drugs or biomolecules, due to their mild effect on cell survival and proliferation. However, little is known about their effect on cellular metabolism. Here we examine the in vivo effect of AuNPs on metabolism using Drosophila as a model. Drosophila and vertebrates possess similar basic metabolic functions, and a highly conserved PI3K/Akt/mTOR signaling pathway plays a central role in the regulation of energy metabolism in both organisms. We show that dietary AuNPs enter the fat body, a key metabolic tissue in Drosophila larvae. Significantly, larvae fed with AuNP show increased lipid levels without triggering stress responses. In addition, activities of the PI3K/Akt/mTOR signaling pathway and fatty acids synthesis are increased in these larvae. This study thus reveals a novel function of AuNPs in influencing animal metabolism and suggests its potential therapeutic applications for metabolic disorders.

  16. Dioscin inhibits osteoclast differentiation and bone resorption though down-regulating the Akt signaling cascades.

    PubMed

    Qu, Xinhua; Zhai, Zanjing; Liu, Xuqiang; Li, Haowei; Ouyang, Zhengxiao; Wu, Chuanlong; Liu, Guangwang; Fan, Qiming; Tang, Tingting; Qin, An; Dai, Kerong

    2014-01-10

    Bone resorption is the unique function of osteoclasts (OCs) and is critical for both bone homeostasis and pathologic bone diseases including osteoporosis, rheumatoid arthritis and tumor bone metastasis. Thus, searching for natural compounds that may suppress osteoclast formation and/or function is promising for the treatment of osteoclast-related diseases. In this study, we for the first time demonstrated that dioscin suppressed RANKL-mediated osteoclast differentiation and bone resorption in vitro in a dose-dependent manner. The suppressive effect of dioscin is supported by the reduced expression of osteoclast-specific markers. Further molecular analysis revealed that dioscin abrogated AKT phosphorylation, which subsequently impaired RANKL-induced nuclear factor-kappaB (NF-κB) signaling pathway and inhibited NFATc1 transcriptional activity. Moreover, in vivo studies further verified the bone protection activity of dioscin in osteolytic animal model. Together our data demonstrate that dioscin suppressed RANKL-induced osteoclast formation and function through Akt signaling cascades. Therefore, dioscin is a potential natural agent for the treatment of osteoclast-related diseases. PMID:24333429

  17. Remodeling the clock: coactivators and signal transduction in the circadian clockworks

    NASA Astrophysics Data System (ADS)

    Weber, Frank

    2009-03-01

    Most organisms on earth such as cyanobacteria, fungi, plants, insects, animals, and humans synchronize their physiological and behavioral activities with the environmental cycles of day and night. Significant progress has been made in unraveling the genetic components that constitute a molecular circadian clock, which facilitates the temporal control of physiology and behavior. Clock genes assemble interlocked transcriptional/translational feedback loops that underlie the circadian oscillations. Recent investigations revealed that posttranslational regulation of clock proteins is crucial for functioning of the molecular oscillator and for precise temporal control of circadian transcription. This review provides an overview of the homologous clockworks in Drosophila and mammals, with a special focus on recent insights in the posttranslational regulation of clock proteins as well as the role of coactivators, repressors, and signal transduction for circadian controlled genome-wide transcription. The emerging mechanisms of clock gene regulation provide an understanding of the temporal control of transcription in general and the circadian orchestration of physiology and behavior in particular.

  18. Functional characterization of WalRK: A two-component signal transduction system from Bacillus anthracis.

    PubMed

    Dhiman, Alisha; Bhatnagar, Sonika; Kulshreshtha, Parul; Bhatnagar, Rakesh

    2014-01-01

    Two-component signal transduction systems (TCS), consisting of a sensor histidine protein kinase and its cognate response regulator, are an important mode of environmental sensing in bacteria. Additionally, they have been found to regulate virulence determinants in several pathogens. Bacillus anthracis, the causative agent of anthrax and a bioterrorism agent, harbours 41 pairs of TCS. However, their role in its pathogenicity has remained largely unexplored. Here, we show that WalRK of B. anthracis forms a functional TCS which exhibits some species-specific functions. Biochemical studies showed that domain variants of WalK, the histidine kinase, exhibit classical properties of autophosphorylation and phosphotransfer to its cognate response regulator WalR. Interestingly, these domain variants also show phosphatase activity towards phosphorylated WalR, thereby making WalK a bifunctional histidine kinase/phosphatase. An in silico regulon determination approach, using a consensus binding sequence from Bacillus subtilis, provided a list of 30 genes that could form a putative WalR regulon in B. anthracis. Further, electrophoretic mobility shift assay was used to show direct binding of purified WalR to the upstream regions of three putative regulon candidates, an S-layer protein EA1, a cell division ABC transporter FtsE and a sporulation histidine kinase KinB3. Our work lends insight into the species-specific functions and mode of action of B. anthracis WalRK. PMID:24490131

  19. Activation of signal transduction in platelets by the tyrosine phosphatase inhibitor pervanadate (vanadyl hydroperoxide).

    PubMed Central

    Pumiglia, K M; Lau, L F; Huang, C K; Burroughs, S; Feinstein, M B

    1992-01-01

    The protein tyrosine phosphatase (PTPase) inhibitor pervanadate (vanadyl hydroperoxide) stimulated protein tyrosine phosphorylation 29-fold more than did thrombin in intact and saponin-permeabilized platelets. Increased tyrosine phosphorylation preceded, or was coincident with, a fall in PtdIns(4,5)P2 levels, production of PtdIns(3,4)P2 and phosphatidic acid, mobilization of intracellular Ca2+, stimulation of protein kinase C-dependent protein phosphorylation, secretion of dense and alpha-granules, increased actin polymerization, shape change and aggregation which required fibrinogen and was mediated by increased surface expression of GPIIb-IIIa. The tyrosine kinase inhibitor RG 50864 totally prevented induction of tyrosine phosphorylation by pervanadate, as well as all other responses measured; in contrast, the inactive structural analogue, tyrphostin #1, had no effect. Dense-granule secretion induced by pervanadate required protein kinase C activity; however, aggregation and alpha-granule secretion were independent of protein kinase C. In saponin-permeabilized platelets pervanadate and thrombin stimulated phospholipase C activity by GTP-independent and GTP-dependent mechanisms respectively. We conclude that PTPases are important regulators of signal transduction in platelets. Images Fig. 1. Fig. 2. PMID:1530576

  20. Signal transduction pathways mediating parathyroid hormone regulation of osteoblastic gene expression

    NASA Technical Reports Server (NTRS)

    Partridge, N. C.; Bloch, S. R.; Pearman, A. T.

    1994-01-01

    Parathyroid hormone (PTH) plays a central role in regulation of calcium metabolism. For example, excessive or inappropriate production of PTH or the related hormone, parathyroid hormone related protein (PTHrP), accounts for the majority of the causes of hypercalcemia. Both hormones act through the same receptor on the osteoblast to elicit enhanced bone resorption by the osteoclast. Thus, the osteoblast mediates the effect of PTH in the resorption process. In this process, PTH causes a change in the function and phenotype of the osteoblast from a cell involved in bone formation to one directing the process of bone resorption. In response to PTH, the osteoblast decreases collagen, alkaline phosphatase, and osteopontin expression and increases production of osteocalcin, cytokines, and neutral proteases. Many of these changes have been shown to be due to effects on mRNA abundance through either transcriptional or post-transcriptional mechanisms. However, the signal transduction pathway for the hormone to cause these changes is not completely elucidated in any case. Binding of PTH and PTHrP to their common receptor has been shown to result in activation of protein kinases A and C and increases in intracellular calcium. The latter has not been implicated in any changes in mRNA of osteoblastic genes. On the other hand activation of PKA can mimic all the effects of PTH; protein kinase C may be involved in some responses. We will discuss possible mechanisms linking PKA and PKC activation to changes in gene expression, particularly at the nuclear level.