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Sample records for growth regulatory mediator

  1. Heparin affin regulatory peptide/pleiotrophin mediates fibroblast growth factor 2 stimulatory effects on human prostate cancer cells.

    PubMed

    Hatziapostolou, Maria; Polytarchou, Christos; Katsoris, Panagiotis; Courty, Jose; Papadimitriou, Evangelia

    2006-10-27

    Fibroblast growth factor 2 (FGF2) is a pleiotropic growth factor that has been implicated in prostate cancer formation and progression. In the present study we found that exogenous FGF2 significantly increased human prostate cancer LNCaP cell proliferation and migration. Heparin affin regulatory peptide (HARP) or pleiotrophin seems to be an important mediator of FGF2 stimulatory effects, since the latter had no effect on stably transfected LNCaP cells that did not express HARP. Moreover, FGF2, through FGF receptors (FGFRs), significantly induced HARP expression and secretion by LNCaP cells and increased luciferase activity of a reporter gene vector carrying the full-length promoter of HARP gene. Using a combination of Western blot analyses, as well as genetic and pharmacological inhibitors, we found that activation of FGFR by FGF2 in LNCaP cells leads to NAD(P)H oxidase-dependent hydrogen peroxide production, phosphorylation of ERK1/2 and p38, activation of AP-1, increased expression and secretion of HARP, and, finally, increased cell proliferation and migration. These results establish the role and the mode of activity of FGF2 in LNCaP cells and support an interventional role of HARP in FGF2 effects, providing new insights on the interplay among growth factor pathways within prostate cancer cells.

  2. Platelet-derived growth factor mediates survival of leukemic large granular lymphocytes via an autocrine regulatory pathway.

    PubMed

    Yang, Jun; Liu, Xin; Nyland, Susan B; Zhang, Ranran; Ryland, Lindsay K; Broeg, Kathleen; Baab, Kendall Thomas; Jarbadan, Nancy Ruth; Irby, Rosalyn; Loughran, Thomas P

    2010-01-07

    Large granular lymphocyte (LGL) leukemia results from chronic expansion of cytotoxic T cells or natural killer (NK) cells. Apoptotic resistance resulting from constitutive activation of survival signaling pathways is a fundamental pathogenic mechanism. Recent network modeling analyses identified platelet-derived growth factor (PDGF) as a key master switch in controlling these survival pathways in T-cell LGL leukemia. Here we show that an autocrine PDGF regulatory loop mediates survival of leukemic LGLs of both T- and NK-cell origin. We found high levels of circulating PDGF-BB in platelet-poor plasma samples from LGL leukemia patients. Production of PDGF-BB by leukemic LGLs was demonstrated by immunocytochemical staining. Leukemic cells expressed much higher levels of PDGFR-beta transcripts than purified normal CD8(+) T cells or NK cells. We observed that phosphatidylinositol-3-kinase (PI3 kinase), Src family kinase (SFK), and downstream protein kinase B (PKB)/AKT pathways were constitutively activated in both T- and NK-LGL leukemia. Pharmacologic blockade of these pathways led to apoptosis of leukemic LGLs. Neutralizing antibody to PDGF-BB inhibited PKB/AKT phosphorylation induced by LGL leukemia sera. These results suggest that targeting of PDGF-BB, a pivotal regulator for the long-term survival of leukemic LGLs, may be an important therapeutic strategy.

  3. Maraviroc decreases CCL8-mediated migration of CCR5+ regulatory T cells and reduces metastatic tumor growth in the lungs

    PubMed Central

    Halvorsen, E. C.; Hamilton, M. J.; Young, A.; Wadsworth, B. J.; LePard, N. E.; Lee, H. N.; Firmino, N.; Collier, J. L.; Bennewith, K. L.

    2016-01-01

    ABSTRACT Regulatory T cells (Tregs) play a crucial physiological role in the regulation of immune homeostasis, although recent data suggest Tregs can contribute to primary tumor growth by suppressing antitumor immune responses. Tregs may also influence the development of tumor metastases, although there is a paucity of information regarding the phenotype and function of Tregs in metastatic target organs. Herein, we demonstrate that orthotopically implanted metastatic mammary tumors induce significant Treg accumulation in the lungs, which is a site of mammary tumor metastasis. Tregs in the primary tumor and metastatic lungs express high levels of C–C chemokine receptor type 5 (CCR5) relative to Tregs in the mammary fat pad and lungs of tumor-free mice, and Tregs in the metastatic lungs are enriched for CCR5 expression in comparison to other immune cell populations. We also identify that C–C chemokine ligand 8 (CCL8), an endogenous ligand of CCR5, is produced by F4/80+ macrophages in the lungs of mice with metastatic primary tumors. Migration of Tregs toward CCL8 ex vivo is reduced in the presence of the CCR5 inhibitor Maraviroc. Importantly, treatment of mice with Maraviroc (MVC) reduces the level of CCR5+ Tregs and metastatic tumor burden in the lungs. This work provides evidence of a CCL8/CCR5 signaling axis driving Treg recruitment to the lungs of mice bearing metastatic primary tumors, representing a potential therapeutic target to decrease Treg accumulation and metastatic tumor growth. PMID:27471618

  4. Maraviroc decreases CCL8-mediated migration of CCR5(+) regulatory T cells and reduces metastatic tumor growth in the lungs.

    PubMed

    Halvorsen, E C; Hamilton, M J; Young, A; Wadsworth, B J; LePard, N E; Lee, H N; Firmino, N; Collier, J L; Bennewith, K L

    2016-06-01

    Regulatory T cells (Tregs) play a crucial physiological role in the regulation of immune homeostasis, although recent data suggest Tregs can contribute to primary tumor growth by suppressing antitumor immune responses. Tregs may also influence the development of tumor metastases, although there is a paucity of information regarding the phenotype and function of Tregs in metastatic target organs. Herein, we demonstrate that orthotopically implanted metastatic mammary tumors induce significant Treg accumulation in the lungs, which is a site of mammary tumor metastasis. Tregs in the primary tumor and metastatic lungs express high levels of C-C chemokine receptor type 5 (CCR5) relative to Tregs in the mammary fat pad and lungs of tumor-free mice, and Tregs in the metastatic lungs are enriched for CCR5 expression in comparison to other immune cell populations. We also identify that C-C chemokine ligand 8 (CCL8), an endogenous ligand of CCR5, is produced by F4/80(+) macrophages in the lungs of mice with metastatic primary tumors. Migration of Tregs toward CCL8 ex vivo is reduced in the presence of the CCR5 inhibitor Maraviroc. Importantly, treatment of mice with Maraviroc (MVC) reduces the level of CCR5(+) Tregs and metastatic tumor burden in the lungs. This work provides evidence of a CCL8/CCR5 signaling axis driving Treg recruitment to the lungs of mice bearing metastatic primary tumors, representing a potential therapeutic target to decrease Treg accumulation and metastatic tumor growth.

  5. Cell Contact–Dependent Immunosuppression by Cd4+Cd25+Regulatory T Cells Is Mediated by Cell Surface–Bound Transforming Growth Factor β

    PubMed Central

    Nakamura, Kazuhiko; Kitani, Atsushi; Strober, Warren

    2001-01-01

    CD4+CD25+ T cells have been identified as a population of immunoregulatory T cells, which mediate suppression of CD4+CD25− T cells by cell–cell contact and not secretion of suppressor cytokines. In this study, we demonstrated that CD4+CD25+ T cells do produce high levels of transforming growth factor (TGF)-β1 and interleukin (IL)-10 compared with CD4+CD25− T cells when stimulated by plate-bound anti-CD3 and soluble anti-CD28 and/or IL-2, and secretion of TGF-β1 (but not other cytokines), is further enhanced by costimulation via cytotoxic T lymphocyte–associated antigen (CTLA)-4. As in prior studies, we found that CD4+CD25+ T cells suppress proliferation of CD4+CD25− T cells; however, we observed here that such suppression is abolished by the presence of anti–TGF-β. In addition, we found that CD4+CD25+ T cells suppress B cell immunoglobulin production and that anti–TGF-β again abolishes such suppression. Finally, we found that stimulated CD4+CD25+ T cells but not CD4+CD25− T cells express high and persistent levels of TGF-β1 on the cell surface. This, plus the fact that we could find no evidence that a soluble factor mediates suppression, strongly suggests that CD4+CD25+ T cells exert immunosuppression by a cell–cell interaction involving cell surface TGF-β1. PMID:11535631

  6. CREB is a regulatory target for the protein kinase Akt/PKB in the differentiation of pancreatic ductal cells into islet {beta}-cells mediated by hepatocyte growth factor

    SciTech Connect

    Li, Xin-Yu; Zhan, Xiao-Rong; Liu, Xiao-Min; Wang, Xiao-Chen

    2011-01-14

    Research highlights: {yields} CREB is a regulatory target for the protein kinase Akt/PKB in pancreatic duct cells. {yields} Activation of the PI3K/AKT/CREB pathway plays a critical role in the HGF-mediated differentiation of pancreatic duct cells in vivo. {yields} CREB was causally linked to the expression of transcription factors during PDEC differentiation induced by HGF. -- Abstract: We have previously reported that the PI3K/Akt signaling pathway is involved in hepatocyte growth factor (HGF)-induced differentiation of adult rat pancreatic ductal epithelial cells (PDECs) into islet {beta}-cells in vitro. The transcription factor CREB is one of the downstream key effectors of the PI3K/Akt signaling pathway. Recent studies showing that CREB is required for the survival of certain cell types prompted us to examine whether CREB is a nuclear target for activation via the HGF-dependent Ser/Thr kinase Akt/PKB in the differentiation of pancreatic duct cell into islet {beta}-cells. In this study, we first attempted to examine whether HGF modulates the Akt-dependent activation of target gene CREB and then investigated whether CREB activity affects the differentiation of HGF-induced PDECs. Finally, we studied the role of CREB in modulating the expression of transcription factors in PDECs during the differentiation of HGF-induced PDECs. Our results demonstrated that CREB is a regulatory target for the protein kinase Akt/PKB in the differentiation of pancreatic ductal cells into islet {beta}-cells mediated by HGF.

  7. Network growth models and genetic regulatory networks

    NASA Astrophysics Data System (ADS)

    Foster, D. V.; Kauffman, S. A.; Socolar, J. E. S.

    2006-03-01

    We study a class of growth algorithms for directed graphs that are candidate models for the evolution of genetic regulatory networks. The algorithms involve partial duplication of nodes and their links, together with the innovation of new links, allowing for the possibility that input and output links from a newly created node may have different probabilities of survival. We find some counterintuitive trends as the parameters are varied, including the broadening of the in-degree distribution when the probability for retaining input links is decreased. We also find that both the scaling of transcription factors with genome size and the measured degree distributions for genes in yeast can be reproduced by the growth algorithm if and only if a special seed is used to initiate the process.

  8. Network growth models and genetic regulatory networks

    NASA Astrophysics Data System (ADS)

    Socolar, Joshua; Foster, David; Kauffman, Stuart

    2006-03-01

    We study a class of growth algorithms for directed graphs that are candidate models for the evolution of genetic regulatory networks. The algorithms involve partial duplication of nodes and their links, together with innovation of new links, allowing for the possibility that input and output links from a newly created node may have different probabilities of survival. We find some counterintuitive trends as parameters are varied, including the broadening of indegree distribution when the probability for retaining input links is decreased. We also find that both the scaling of transcription factors with genome size and the measured degree distributions for genes in yeast can be reproduced by the growth algorithm if and only if a special seed is used to initiate the process.

  9. p85 regulatory subunit of PI3K mediates cAMP-PKA and estrogens biological effects on growth and survival.

    PubMed

    Cosentino, C; Di Domenico, M; Porcellini, A; Cuozzo, C; De Gregorio, G; Santillo, M R; Agnese, S; Di Stasio, R; Feliciello, A; Migliaccio, A; Avvedimento, E V

    2007-03-29

    Cyclic adenosine 3'5' monophosphate (cAMP) and protein kinase A (PKA) cooperate with phosphatidylinositol 3' kinase (PI3K) signals in the control of growth and survival. To determine the molecular mechanism(s) involved, we identified and mutagenized a specific serine (residue 83) in p85alpha(PI3K), which is phosphorylated in vivo and in vitro by PKA. Expression of p85alpha(PI3K) mutants (alanine or aspartic substitutions) significantly altered the biological responses of the cells to cAMP. cAMP protection from anoikis was reduced in cells expressing the alanine version p85alpha(PI3K). These cells did not arrest in G1 in the presence of cAMP, whereas cells expressing the aspartic mutant p85D accumulated in G1 even in the absence of cAMP. S phase was still efficiently inhibited by cAMP in cells expressing both mutants. The binding of PI3K to Ras p21 was greatly reduced in cells expressing p85A in the presence or absence of cAMP. Conversely, expression of the aspartic mutant stimulated robustly the binding of PI3K to p21 Ras in the presence of cAMP. Mutation in the Ser 83 inhibited cAMP, but not PDGF stimulation of PI3K. Conversely, the p85D aspartic mutant amplified cAMP stimulation of PI3K activity. Phosphorylation of Ser 83 by cAMP-PKA in p85alpha(PI3K) was also necessary for estrogen signaling as expression of p85A or p85D mutants inhibited or amplified, respectively, the binding of estrogen receptor to p85alpha and AKT phosphorylation induced by estrogens. The data presented indicate that: (1) phosphorylation of Ser 83 in p85alpha(PI3K) is critical for cAMP-PKA induced G1 arrest and survival in mouse 3T3 fibroblasts; (2) this site is necessary for amplification of estrogen signals by cAMP-PKA and related receptors. Finally, these data suggest a general mechanism of PI3K regulation by cAMP, operating in various cell types and under different conditions.

  10. Regulatory T Cell Immunotherapy in Immune-Mediated Diseases

    PubMed Central

    Pierini, Antonio; Schneidawind, Dominik; Nishikii, Hidekazu; Negrin, Robert S.

    2015-01-01

    Broad clinical interest rapidly followed the recent discovery of different subpopulations of T cells that have immune regulatory properties and a number of studies have been conducted aiming to dissect the translational potential of these promising cells. In this review we will focus on forkhead box P3 (FoxP3) positive regulatory T cells, T regulatory type 1 cells and invariant natural killer T cells (iNKT). We will analyze their ability to correct immune dysregulation in animal models of immune mediated diseases and we will examine the first clinical approaches where these cells have been directly or indirectly employed. We will discuss successes, challenges and limitations that rose in the road to the clinical use of regulatory T cells. PMID:26779417

  11. Fur-mediated global regulatory circuits in pathogenic Neisseria species.

    PubMed

    Yu, Chunxiao; Genco, Caroline Attardo

    2012-12-01

    The ferric uptake regulator (Fur) protein has been shown to function as a repressor of transcription in a number of diverse microorganisms. However, recent studies have established that Fur can function at a global level as both an activator and a repressor of transcription through both direct and indirect mechanisms. Fur-mediated indirect activation occurs via the repression of additional repressor proteins, or small regulatory RNAs, thereby activating transcription of a previously silent gene. Fur mediates direct activation through binding of Fur to the promoter regions of genes. Whereas the repressive mechanism of Fur has been thoroughly investigated, emerging studies on direct and indirect Fur-mediated activation mechanisms have revealed novel global regulatory circuits.

  12. Secretory pattern and regulatory mechanism of growth hormone in cattle

    PubMed Central

    2016-01-01

    Abstract The ultradian rhythm of growth hormone (GH) secretion has been known in several animal species for years and has recently been observed in cattle. Although the physiological significance of the rhythm is not yet fully understood, it appears essential for normal growth. In this review, previous studies concerning the GH secretory pattern in cattle, including its ultradian rhythm, are introduced and the regulatory mechanism is discussed on the basis of recent findings. PMID:26260675

  13. Secretory pattern and regulatory mechanism of growth hormone in cattle.

    PubMed

    Kasuya, Etsuko

    2016-02-01

    The ultradian rhythm of growth hormone (GH) secretion has been known in several animal species for years and has recently been observed in cattle. Although the physiological significance of the rhythm is not yet fully understood, it appears essential for normal growth. In this review, previous studies concerning the GH secretory pattern in cattle, including its ultradian rhythm, are introduced and the regulatory mechanism is discussed on the basis of recent findings.

  14. Multiple regulatory systems coordinate DNA replication with cell growth in Bacillus subtilis.

    PubMed

    Murray, Heath; Koh, Alan

    2014-10-01

    In many bacteria the rate of DNA replication is linked with cellular physiology to ensure that genome duplication is coordinated with growth. Nutrient-mediated growth rate control of DNA replication initiation has been appreciated for decades, however the mechanism(s) that connects these cell cycle activities has eluded understanding. In order to help address this fundamental question we have investigated regulation of DNA replication in the model organism Bacillus subtilis. Contrary to the prevailing view we find that changes in DnaA protein level are not sufficient to account for nutrient-mediated growth rate control of DNA replication initiation, although this regulation does require both DnaA and the endogenous replication origin. We go on to report connections between DNA replication and several essential cellular activities required for rapid bacterial growth, including respiration, central carbon metabolism, fatty acid synthesis, phospholipid synthesis, and protein synthesis. Unexpectedly, the results indicate that multiple regulatory systems are involved in coordinating DNA replication with cell physiology, with some of the regulatory systems targeting oriC while others act in a oriC-independent manner. We propose that distinct regulatory systems are utilized to control DNA replication in response to diverse physiological and chemical changes.

  15. Immune regulatory effects of simvastatin on regulatory T cell-mediated tumour immune tolerance.

    PubMed

    Lee, K J; Moon, J Y; Choi, H K; Kim, H O; Hur, G Y; Jung, K H; Lee, S Y; Kim, J H; Shin, C; Shim, J J; In, K H; Yoo, S H; Kang, K H; Lee, S Y

    2010-08-01

    Statins are potent inhibitors of hydroxyl-3-methylglutaryl co-enzyme A (HMG-CoA) reductase, and have emerged as potential anti-cancer agents based on preclinical evidence. In particular, compelling evidence suggests that statins have a wide range of immunomodulatory properties. However, little is known about the role of statins in tumour immune tolerance. Tumour immune tolerance involves the production of immunosuppressive molecules, such as interleukin (IL)-10, transforming growth factor (TGF)-beta and indoleamine-2,3-dioxygenase (IDO) by tumours, which induce a regulatory T cell (T(reg)) response. In this study, we investigated the effect of simvastatin on the production of IL-10, TGF-beta and IDO production and the proliferation of T(regs) using several cancer cell lines, and Lewis lung cancer (3LL) cells-inoculated mouse tumour model. Simvastatin treatment resulted in a decrease in the number of cancer cells (3LL, A549 and NCI-H292). The production of the immune regulatory markers IL-10, TGF-beta in 3LL and NCI-H292 cells increased after treatment with simvastatin. The expression of IDO and forkhead box P3 (FoxP3) transcription factor was also increased in the presence of simvastatin. In a murine 3LL model, there were no significant differences in tumour growth rate between untreated and simvastatin-treated mice groups. Therefore, while simvastatin had an anti-proliferative effect, it also exhibited immune tolerance-promoting properties during tumour development. Thus, due to these opposing actions, simvastatin had no net effect on tumour growth.

  16. A regulatory role for cAMP in phosphatidylinositol 3-kinase/p70 ribosomal S6 kinase-mediated DNA synthesis in platelet-derived-growth-factor-stimulated bovine airway smooth-muscle cells.

    PubMed Central

    Scott, P H; Belham, C M; al-Hafidh, J; Chilvers, E R; Peacock, A J; Gould, G W; Plevin, R

    1996-01-01

    In bovine airway smooth-muscle cells platelet-derived growth factor (PDGF) and endothelin (Et-1) stimulate sustained and comparable activation of mitogen-activated protein kinase (MAP kinase) but display very different mitogenic efficacies, with PDGF inducing 50 times more DNA synthesis than Et-1. To examine additional signalling pathways which may be involved in this response, we investigated the role of phosphatidylinositol 3-kinase (PtdIns 3-kinase)/p70 ribosomal protein S6 kinase (p70s6k) in mediating PDGF- and Et-1-induced mitogenesis, and whether inhibition of this pathway may underly the ability of cAMP to inhibit cell proliferation. PDGF stimulated an increase in PtdIns 3-kinase activity and a sustained 15-fold increase in p70s6k activity that was abolished by both wortmannin and rapamycin. Et-1, however, stimulated only a 2-fold increase in p70s6k activity that was rapamycin-sensitive but wortmannin-insensitive. DNA synthesis stimulated by PDGF (50-fold) and Et-1 (2-fold) followed a similar pattern of inhibition. Pretreatment with phorbol ester did not affect p70s6k activation in response to PDGF. Raising intracellular cAMP levels using forskolin, however, resulted in a marked time-dependent inhibition of p70s6k activity, a decrease in the tyrosine phosphorylation of the PtdIns 3-kinase p85 subunit and reduced PtdIns 3-kinase activity. Forskolin also inhibited PDGF-stimulated DNA synthesis. These results suggest that PtdIns 3-kinase-dependent activation of p70s6k may determine mitogenic efficacy of agonists that generate comparable MAP kinase signals. Negative regulation of PtdIns 3-kinase by cAMP may play an important role in the inhibition of airway smooth-muscle cell proliferation. PMID:8836145

  17. Regulatory mechanisms underlying the differential growth of dendrites and axons.

    PubMed

    Wang, Xin; Sterne, Gabriella R; Ye, Bing

    2014-08-01

    A typical neuron is comprised of an information input compartment, or the dendrites, and an output compartment, known as the axon. These two compartments are the structural basis for functional neural circuits. However, little is known about how dendritic and axonal growth are differentially regulated. Recent studies have uncovered two distinct types of regulatory mechanisms that differentiate dendritic and axonal growth: dedicated mechanisms and bimodal mechanisms. Dedicated mechanisms regulate either dendritespecific or axon-specific growth; in contrast, bimodal mechanisms direct dendritic and axonal development in opposite manners. Here, we review the dedicated and bimodal regulators identified by recent Drosophila and mammalian studies. The knowledge of these underlying molecular mechanisms not only expands our understanding about how neural circuits are wired, but also provides insights that will aid in the rational design of therapies for neurological diseases.

  18. Myogenic regulatory transcription factors regulate growth in rhabdomyosarcoma

    PubMed Central

    Tenente, Inês M; Hayes, Madeline N; Ignatius, Myron S; McCarthy, Karin; Yohe, Marielle; Sindiri, Sivasish; Gryder, Berkley; Oliveira, Mariana L; Ramakrishnan, Ashwin; Tang, Qin; Chen, Eleanor Y; Petur Nielsen, G; Khan, Javed; Langenau, David M

    2017-01-01

    Rhabdomyosarcoma (RMS) is a pediatric malignacy of muscle with myogenic regulatory transcription factors MYOD and MYF5 being expressed in this disease. Consensus in the field has been that expression of these factors likely reflects the target cell of transformation rather than being required for continued tumor growth. Here, we used a transgenic zebrafish model to show that Myf5 is sufficient to confer tumor-propagating potential to RMS cells and caused tumors to initiate earlier and have higher penetrance. Analysis of human RMS revealed that MYF5 and MYOD are mutually-exclusively expressed and each is required for sustained tumor growth. ChIP-seq and mechanistic studies in human RMS uncovered that MYF5 and MYOD bind common DNA regulatory elements to alter transcription of genes that regulate muscle development and cell cycle progression. Our data support unappreciated and dominant oncogenic roles for MYF5 and MYOD convergence on common transcriptional targets to regulate human RMS growth. DOI: http://dx.doi.org/10.7554/eLife.19214.001 PMID:28080960

  19. Putting theory to the test: which regulatory mechanisms can drive realistic growth of a root?

    PubMed

    De Vos, Dirk; Vissenberg, Kris; Broeckhove, Jan; Beemster, Gerrit T S

    2014-10-01

    In recent years there has been a strong development of computational approaches to mechanistically understand organ growth regulation in plants. In this study, simulation methods were used to explore which regulatory mechanisms can lead to realistic output at the cell and whole organ scale and which other possibilities must be discarded as they result in cellular patterns and kinematic characteristics that are not consistent with experimental observations for the Arabidopsis thaliana primary root. To aid in this analysis, a 'Uniform Longitudinal Strain Rule' (ULSR) was formulated as a necessary condition for stable, unidirectional, symplastic growth. Our simulations indicate that symplastic structures are robust to differences in longitudinal strain rates along the growth axis only if these differences are small and short-lived. Whereas simple cell-autonomous regulatory rules based on counters and timers can produce stable growth, it was found that steady developmental zones and smooth transitions in cell lengths are not feasible. By introducing spatial cues into growth regulation, those inadequacies could be avoided and experimental data could be faithfully reproduced. Nevertheless, a root growth model based on previous polar auxin-transport mechanisms violates the proposed ULSR due to the presence of lateral gradients. Models with layer-specific regulation or layer-driven growth offer potential solutions. Alternatively, a model representing the known cross-talk between auxin, as the cell proliferation promoting factor, and cytokinin, as the cell differentiation promoting factor, predicts the effect of hormone-perturbations on meristem size. By down-regulating PIN-mediated transport through the transcription factor SHY2, cytokinin effectively flattens the lateral auxin gradient, at the basal boundary of the division zone, (thereby imposing the ULSR) to signal the exit of proliferation and start of elongation. This model exploration underlines the value of

  20. Putting Theory to the Test: Which Regulatory Mechanisms Can Drive Realistic Growth of a Root?

    PubMed Central

    De Vos, Dirk; Vissenberg, Kris; Broeckhove, Jan; Beemster, Gerrit T. S.

    2014-01-01

    In recent years there has been a strong development of computational approaches to mechanistically understand organ growth regulation in plants. In this study, simulation methods were used to explore which regulatory mechanisms can lead to realistic output at the cell and whole organ scale and which other possibilities must be discarded as they result in cellular patterns and kinematic characteristics that are not consistent with experimental observations for the Arabidopsis thaliana primary root. To aid in this analysis, a ‘Uniform Longitudinal Strain Rule’ (ULSR) was formulated as a necessary condition for stable, unidirectional, symplastic growth. Our simulations indicate that symplastic structures are robust to differences in longitudinal strain rates along the growth axis only if these differences are small and short-lived. Whereas simple cell-autonomous regulatory rules based on counters and timers can produce stable growth, it was found that steady developmental zones and smooth transitions in cell lengths are not feasible. By introducing spatial cues into growth regulation, those inadequacies could be avoided and experimental data could be faithfully reproduced. Nevertheless, a root growth model based on previous polar auxin-transport mechanisms violates the proposed ULSR due to the presence of lateral gradients. Models with layer-specific regulation or layer-driven growth offer potential solutions. Alternatively, a model representing the known cross-talk between auxin, as the cell proliferation promoting factor, and cytokinin, as the cell differentiation promoting factor, predicts the effect of hormone-perturbations on meristem size. By down-regulating PIN-mediated transport through the transcription factor SHY2, cytokinin effectively flattens the lateral auxin gradient, at the basal boundary of the division zone, (thereby imposing the ULSR) to signal the exit of proliferation and start of elongation. This model exploration underlines the value of

  1. Regulatory mechanisms of growth hormone secretion are sexually dimorphic.

    PubMed Central

    Jaffe, C A; Ocampo-Lim, B; Guo, W; Krueger, K; Sugahara, I; DeMott-Friberg, R; Bermann, M; Barkan, A L

    1998-01-01

    Sexually dimorphic growth hormone (GH) secretory pattern is important in the determination of gender-specific patterns of growth and metabolism in rats. Whether GH secretion in humans is also sexually dimorphic and the neuroendocrine mechanisms governing this potential difference are not fully established. We have compared pulsatile GH secretion profiles in young men and women in the baseline state and during a continuous intravenous infusion of recombinant human insulin-like growth factor I (rhIGF-I). During the baseline study, men had large nocturnal GH pulses and relatively small pulses during the rest of the day. In contrast, women had more continuous GH secretion and more frequent GH pulses that were of more uniform size. The infusion of rhIGF-I (10 microg/kg/h) potently suppressed both spontaneous and growth hormone-releasing hormone (GHRH)-induced GH secretion in men. In women, however, rhIGF-I had less effect on pulsatile GH secretion and did not suppress the GH response to GHRH. These data demonstrate the existence of sexual dimorphism in the regulatory mechanisms involved in GH secretion in humans. The persistence of GH responses to GHRH in women suggests that negative feedback by IGF-I might be expressed, in part, through suppression of hypothalamic GHRH. PMID:9649569

  2. Interleukin-35-mediated induction of a novel regulatory T cell population

    PubMed Central

    Collison, Lauren W.; Chaturvedi, Vandana; Henderson, Abigail L.; Giacomin, Paul R.; Guy, Cliff; Bankoti, Jaishree; Finkelstein, David; Forbes, Karen; Workman, Creg J.; Brown, Scott A.; Rehg, Jerold E.; Jones, Michael L.; Ni, Hsiao-Tzu; Artis, David; Turk, Mary Jo; Vignali, Dario A. A.

    2010-01-01

    Regulatory T cells (Tregs) play a critical role in the maintenance of immunological self-tolerance. Naïve human or murine T cell treatment with the inhibitory cytokine IL-35 induces a regulatory population, termed iTR35, that mediates suppression via IL-35, but not IL-10 or TGFβ, neither express nor require Foxp3, are strongly suppressive in five in vivo models, and exhibit in vivo stability. Treg-mediated suppression induces iTR35 generation in an IL-35- and IL-10-dependent manner in vitro, and in inflammatory conditions in vivo in Trichuris-infected intestines and within the tumor microenvironment, where they appear to contribute to the regulatory milieu. iTR35 may constitute a key mediator of infectious tolerance, may contribute to Treg-mediated tumor progression, and ex vivo generated iTR35 may possess therapeutic utility. PMID:20953201

  3. Involvement of microRNA-related regulatory pathways in the glucose-mediated control of Arabidopsis early seedling development

    PubMed Central

    Vincentz, Michel

    2013-01-01

    In plants, sugars such as glucose act as signalling molecules that promote changes in gene expression programmes that impact on growth and development. Recent evidence has revealed the potential importance of controlling mRNA decay in some aspects of glucose-mediated regulatory responses suggesting a role of microRNAs (miRNAs) in these responses. In order to get a better understanding of glucose-mediated development modulation involving miRNA-related regulatory pathways, early seedling development of mutants impaired in miRNA biogenesis (hyl1-2 and dcl1-11) and miRNA activity (ago1-25) was evaluated. All mutants exhibited a glucose hyposensitive phenotype from germination up to seedling establishment, indicating that miRNA regulatory pathways are involved in the glucose-mediated delay of early seedling development. The expression profile of 200 miRNA primary transcripts (pri-miRs) was evaluated by large-scale quantitative real-time PCR profiling, which revealed that 38 pri-miRs were regulated by glucose. For several of them, the corresponding mature miRNAs are known to participate directly or indirectly in plant development, and their accumulation was shown to be co-regulated with the pri-miR by glucose. Furthermore, the expression of several miRNA target genes was found to be deregulated in response to glucose in the miRNA machinery mutants ago1-25, dcl1-11, and hyl1-2. Also, in these mutants, glucose promoted misexpression of genes for the three abscisic acid signalling elements ABI3, ABI4, and ABI5. Thus, miRNA regulatory pathways play a role in the adjustments of growth and development triggered by glucose signalling. PMID:23997203

  4. Regulatory focus and burnout in nurses: The mediating effect of perception of transformational leadership.

    PubMed

    Shi, Rui; Zhang, Shilei; Xu, Hang; Liu, Xufeng; Miao, Danmin

    2015-12-01

    This correlation study investigated the relationship between nurses' regulatory focus and burnout, as mediated by their perceptions of transformational leadership, using a cross-sectional research design with anonymous questionnaires. In July-August 2012, data were collected from 378 nurses from three hospitals in Shaanxi Province, China, using self-report questionnaires for measuring the nurses' regulatory focus, their level of burnout and their perception of whether the leadership of their supervisor was transformational. Structural equation modelling and bootstrapping procedures were used to identify the mediating effect of their perceptions of transformational leadership. The results supported our hypothesized model. The type of regulatory focus emerged as a significant predictor of burnout. Having a perception of transformational leadership partially mediated the relationship between regulatory focus and burnout. Having a promotion focus reduced burnout when the participants perceived transformational leadership, whereas having a prevention focus exhibited the opposite pattern. The mediating effect of the perception of transformational leadership suggests that a promotion focus may help diminish burnout, directly and indirectly. Nurse managers must be aware of the role of a regulatory focus and cultivate promotion focus in their followers.

  5. Compensatory adrenal growth - A neurally mediated reflex

    NASA Technical Reports Server (NTRS)

    Dallman, M. F.; Engeland, W. C.; Shinsako, J.

    1976-01-01

    The responses of young rats to left adrenalectomy or left adrenal manipulation were compared to surgical sham adrenalectomy in which adrenals were observed but not touched. At 12 h right adrenal wet weight, dry weight, DNA, RNA, and protein content were increased (P less than 0.05) after the first two operations. Left adrenal manipulation resulted in increased right adrenal weight at 12 h but no change in left adrenal weight. Sequential manipulation of the left adrenal at time 0 and the right adrenal at 12 h resulted in an enlarged right adrenal at 12 h (P less than 0.01), and an enlarged left adrenal at 24 h (P less than 0.05), showing that the manipulated gland was capable of response. Bilateral adrenal manipulation of the adrenal glands resulted in bilateral enlargement of 12 h (P less than 0.01). Taken together with previous results, these findings strongly suggest that compensatory adrenal growth is a neurally mediated reflex.

  6. Growth-Phase Sterigmatocystin Formation on Lactose Is Mediated via Low Specific Growth Rates in Aspergillus nidulans

    PubMed Central

    Németh, Zoltán; Molnár, Ákos P.; Fejes, Balázs; Novák, Levente; Karaffa, Levente; Keller, Nancy P.; Fekete, Erzsébet

    2016-01-01

    Seed contamination with polyketide mycotoxins such as sterigmatocystin (ST) produced by Aspergilli is a worldwide issue. The ST biosynthetic pathway is well-characterized in A. nidulans, but regulatory aspects related to the carbon source are still enigmatic. This is particularly true for lactose, inasmuch as some ST production mutant strains still synthesize ST on lactose but not on other carbon substrates. Here, kinetic data revealed that on d-glucose, ST forms only after the sugar is depleted from the medium, while on lactose, ST appears when most of the carbon source is still available. Biomass-specified ST production on lactose was significantly higher than on d-glucose, suggesting that ST formation may either be mediated by a carbon catabolite regulatory mechanism, or induced by low specific growth rates attainable on lactose. These hypotheses were tested by d-glucose limited chemostat-type continuous fermentations. No ST formed at a high growth rate, while a low growth rate led to the formation of 0.4 mg·L−1 ST. Similar results were obtained with a CreA mutant strain. We concluded that low specific growth rates may be the primary cause of mid-growth ST formation on lactose in A. nidulans, and that carbon utilization rates likely play a general regulatory role during biosynthesis. PMID:27916804

  7. Toxin-mediated gene regulatory mechanism in Staphylococcus aureus

    PubMed Central

    Joo, Hwang-Soo; Otto, Michael

    2016-01-01

    The dangerous human pathogen Staphylococcus aureus relies heavily on toxins to cause disease, but toxin production can put a strong burden on the bacteria’s energy balance. Thus, controlling the synthesis of proteins solely needed in times of toxin production represents a way for the bacteria to avoid wasting energy. One hypothetical manner to accomplish this sort of regulation is by gene regulatory functions of the toxins themselves. There have been several reports about gene regulation by toxins in S. aureus, but these were never verified on the molecular level. In our study published in MBio [Joo et al., 7(5). pii: e01579-16], we show that phenol-soluble modulins (PSMs), important peptide toxins of S. aureus, release a repressor from the promoter of the operon encoding the toxin export system, thereby enabling toxin secretion. This study describes the first molecular regulatory mechanism exerted by an S. aureus toxin, setting a paradigmatic example of how S. aureus toxins may influence cell functions to adjust them to times of toxin production.

  8. Iron Regulatory Proteins Mediate Host Resistance to Salmonella Infection.

    PubMed

    Nairz, Manfred; Ferring-Appel, Dunja; Casarrubea, Daniela; Sonnweber, Thomas; Viatte, Lydie; Schroll, Andrea; Haschka, David; Fang, Ferric C; Hentze, Matthias W; Weiss, Guenter; Galy, Bruno

    2015-08-12

    Macrophages are essential for systemic iron recycling, and also control iron availability to pathogens. Iron metabolism in mammalian cells is orchestrated posttranscriptionally by iron-regulatory proteins (IRP)-1 and -2. Here, we generated mice with selective and combined ablation of both IRPs in macrophages to investigate the role of IRPs in controlling iron availability. These animals are hyperferritinemic but otherwise display normal clinical iron parameters. However, mutant mice rapidly succumb to systemic infection with Salmonella Typhimurium, a pathogenic bacterium that multiplies within macrophages, with increased bacterial burdens in liver and spleen. Ex vivo infection experiments indicate that IRP function restricts bacterial access to iron via the EntC and Feo bacterial iron-acquisition systems. Further, IRPs contain Salmonella by promoting the induction of lipocalin 2, a host antimicrobial factor that inhibits bacterial uptake of iron-laden siderophores, and by suppressing the ferritin iron pool. This work reveals the importance of the IRPs in innate immunity.

  9. Cpeb4-mediated translational regulatory circuitry controls terminal erythroid differentiation.

    PubMed

    Hu, Wenqian; Yuan, Bingbing; Lodish, Harvey F

    2014-09-29

    While we have considerable understanding of the transcriptional networks controlling mammalian cell differentiation, our knowledge of posttranscriptional regulatory events is very limited. Using differentiation of primary erythroid cells as a model, we show that the sequence-specific mRNA-binding protein Cpeb4 is strongly induced by the erythroid-important transcription factors Gata1 and Tal1 and is essential for terminal erythropoiesis. By interacting with the translation initiation factor eIF3, Cpeb4 represses the translation of a large set of mRNAs, including its own mRNA. Thus, transcriptional induction and translational repression combine to form a negative feedback loop to control Cpeb4 protein levels within a specific range that is required for terminal erythropoiesis. Our study provides an example of how translational control is integrated with transcriptional regulation to precisely control gene expression during mammalian cell differentiation.

  10. Cpeb4-mediated translational regulatory circuitry controls terminal erythroid differentiation

    PubMed Central

    Hu, Wenqian; Yuan, Bingbing; Lodish, Harvey F.

    2014-01-01

    SUMMARY While we have considerable understanding of the transcriptional networks controlling mammalian cell differentiation, our knowledge of post-transcriptional regulatory events is very limited. Using differentiation of primary erythroid cells as a model, we show that the sequence-specific mRNA-binding protein Cpeb4 is strongly induced by the erythroid important transcription factors Gata1 and Tal1 and is essential for terminal erythropoiesis. By interacting with the translation initiation factor eIF3 Cpeb4 represses the translation of a large set of mRNAs, including its own mRNA. Thus transcriptional induction and translational repression combine to form a negative feedback loop to control Cpeb4 protein levels within a specific range that is required for terminal erythropoiesis. Our study provides an example of how translational control is integrated with transcriptional regulation to precisely control gene expression during mammalian cell differentiation. PMID:25220394

  11. Mediators in cell growth and differentiation

    SciTech Connect

    Ford, R.J.; Maizel, A.L.

    1985-01-01

    This book contains papers divided among seven sections. The section headings are: Cell Cycle and Control of Cell Growth, Growth Factors for Nonlymphoid Cells, Colony-Stimulating Factors, Stem Cells and Hematopoiesis, Lymphoid Growth Factors, Growth Factors in Neoplasia, Interferon, and Differentiation in Normal and Neoplastic Cells.

  12. Distinct Glucocorticoid Receptor Transcriptional Regulatory Surfaces Mediate the Cytotoxic and Cytostatic Effects of Glucocorticoids

    PubMed Central

    Rogatsky, Inez; Hittelman, Adam B.; Pearce, David; Garabedian, Michael J.

    1999-01-01

    Glucocorticoids act through the glucocorticoid receptor (GR), which can function as a transcriptional activator or repressor, to elicit cytostatic and cytotoxic effects in a variety of cells. The molecular mechanisms regulating these events and the target genes affected by the activated receptor remain largely undefined. Using cultured human osteosarcoma cells as a model for the GR antiproliferative effect, we demonstrate that in U20S cells, GR activation leads to irreversible growth inhibition, apoptosis, and repression of Bcl2. This cytotoxic effect is mediated by GR’s transcriptional repression function, since transactivation-deficient mutants and ligands still bring about apoptosis and Bcl2 down-regulation. In contrast, the antiproliferative effect of GR in SAOS2 cells is reversible, does not result in apoptosis or repression of Bcl2, and is a function of the receptor’s ability to stimulate transcription. Thus, the cytotoxic versus cytostatic outcome of glucocorticoid treatment is cell context dependent. Interestingly, the cytostatic effect of glucocorticoids in SAOS2 cells involves multiple GR activation surfaces. GR mutants and ligands that disrupt individual transcriptional activation functions (activation function 1 [AF-1] and AF-2) or receptor dimerization fail to fully inhibit cellular proliferation and, remarkably, discriminate between the targets of GR’s cytostatic action, the cyclin-dependent kinase inhibitors p21Cip1 and p27Kip1. Induction of p21Cip1 is agonist dependent and requires AF-2 but not AF-1 or GR dimerization. In contrast, induction of p27Kip1 is agonist independent, does not require AF-2 or AF-1, but depends on GR dimerization. Our findings indicate that multiple GR transcriptional regulatory mechanisms that employ distinct receptor surfaces are used to evoke either the cytostatic or cytotoxic response to glucocorticoids. PMID:10373553

  13. CO/FT regulatory module controls timing of flowering and seasonal growth cessation in trees.

    PubMed

    Böhlenius, Henrik; Huang, Tao; Charbonnel-Campaa, Laurence; Brunner, Amy M; Jansson, Stefan; Strauss, Steven H; Nilsson, Ove

    2006-05-19

    Forest trees display a perennial growth behavior characterized by a multiple-year delay in flowering and, in temperate regions, an annual cycling between growth and dormancy. We show here that the CO/FT regulatory module, which controls flowering time in response to variations in daylength in annual plants, controls flowering in aspen trees. Unexpectedly, however, it also controls the short-day-induced growth cessation and bud set occurring in the fall. This regulatory mechanism can explain the ecogenetic variation in a highly adaptive trait: the critical daylength for growth cessation displayed by aspen trees sampled across a latitudinal gradient spanning northern Europe.

  14. Daylength mediated control of seasonal growth patterns in perennial trees.

    PubMed

    Petterle, Anna; Karlberg, Anna; Bhalerao, Rishikesh P

    2013-06-01

    Daylength is a key regulator of seasonal growth patterns in perennial trees in temperate regions. Cessation of growth is induced by short day signal in these trees before the advent of winter and constitutes a major adaptive developmental program. In this review, we report on the recent progress made in identifying the molecular mechanisms that underlie the daylength mediated control of seasonal growth in perennial trees. A major finding that has emerged from the analysis of this process is that the regulation of growth cessation in perennial trees and flowering time by daylength in annuals such as Arabidopsis thaliana involves identical signalling components.

  15. Identification of bolting-related microRNAs and their targets reveals complex miRNA-mediated flowering-time regulatory networks in radish (Raphanus sativus L.).

    PubMed

    Nie, Shanshan; Xu, Liang; Wang, Yan; Huang, Danqiong; Muleke, Everlyne M; Sun, Xiaochuan; Wang, Ronghua; Xie, Yang; Gong, Yiqin; Liu, Liwang

    2015-09-15

    MicroRNAs (miRNAs) play vital regulatory roles in plant growth and development. The phase transition from vegetative growth to flowering is crucial in the life cycle of plants. To date, miRNA-mediated flowering regulatory networks remain largely unexplored in radish. In this study, two small RNA libraries from radish leaves at vegetative and reproductive stages were constructed and sequenced by Solexa sequencing. A total of 94 known miRNAs representing 21 conserved and 13 non-conserved miRNA families, and 44 potential novel miRNAs, were identified from the two libraries. In addition, 42 known and 17 novel miRNAs were significantly differentially expressed and identified as bolting-related miRNAs. RT-qPCR analysis revealed that some miRNAs exhibited tissue- or developmental stage-specific expression patterns. Moreover, 154 target transcripts were identified for 50 bolting-related miRNAs, which were predominately involved in plant development, signal transduction and transcriptional regulation. Based on the characterization of bolting-related miRNAs and their target genes, a putative schematic model of miRNA-mediated bolting and flowering regulatory network was proposed. These results could provide insights into bolting and flowering regulatory networks in radish, and facilitate dissecting the molecular mechanisms underlying bolting and flowering time regulation in vegetable crops.

  16. Insulin-like growth factor 1: common mediator of multiple enterotrophic hormones and growth factors

    PubMed Central

    Bortvedt, Sarah F.; Lund, P. Kay

    2013-01-01

    Purpose of review To summarize recent evidence that IGF1 mediates growth effects of multiple trophic factors and discuss clinical relevance. Recent findings Recent reviews and original reports indicate benefits of growth hormone (GH) and long-acting glucagon-like peptide 2 (GLP2) analogues in short bowel syndrome and Crohn’s disease. This review highlights evidence that biomarkers of sustained small intestinal growth or mucosal healing and evaluation of intestinal epithelial stem cell biomarkers may improve clinical measures of intestinal growth or response to trophic hormones. Compelling evidence that IGF1 mediates growth effects of GH and GLP2 on intestine or linear growth in preclinical models of resection or Crohn’s disease is presented, along with a concept that these hormones or IGF1 may enhance sustained growth if given early after bowel resection. Evidence that SOCS protein induction by GH or GLP2 in normal or inflamed intestine, may limit IGF1-induced growth, but protect against risk of dysplasia or fibrosis is reviewed. Whether IGF1 receptor mediates IGF1 action and potential roles of insulin receptors are addressed. Summary IGF1 has a central role in mediating trophic hormone action in small intestine. Better understanding of benefits and risks of IGF1, receptors that mediate IGF1 action, and factors that limit undesirable growth are needed. PMID:22241077

  17. Drug-device combination products: regulatory landscape and market growth.

    PubMed

    Bayarri, L

    2015-08-01

    Combination products are therapeutic and diagnostic products that combine drugs, devices and/or biological products, leading to safer and more effective treatments thanks to careful and precise drug targeting, local administration and individualized therapy. These technologies can especially benefit patients suffering from serious diseases and conditions such as cancer, heart disease, multiple sclerosis and diabetes, among others. On the other hand, drug-device combination products have also introduced a new dynamic in medical product development, regulatory approval and corporate interaction. Due to the increasing integration of drugs and devices observed in the latest generation of combination products, regulatory agencies have developed specific competences and regulations over the last decade. Manufacturers are required to fully understand the specific requirements in each country in order to ensure timely and accurate market access of new combination products, and the development of combination products involves a very specific pattern of interactions between manufacturers and regulatory agencies. The increased sophistication of the products brought to market over the last couple of decades has accentuated the need to develop drugs and devices collaboratively using resources from both industries, fostering the need of business partnering and technology licensing. This review will provide a global overview of the market trends, as well as (in the last section) an analysis of the drug-device combination products approved by the FDA during the latest 5 years.

  18. Genetic modification through oligonucleotide-mediated mutagenesis. A GMO regulatory challenge?

    PubMed

    Breyer, Didier; Herman, Philippe; Brandenburger, Annick; Gheysen, Godelieve; Remaut, Erik; Soumillion, Patrice; Van Doorsselaere, Jan; Custers, René; Pauwels, Katia; Sneyers, Myriam; Reheul, Dirk

    2009-01-01

    In the European Union, the definition of a GMO is technology-based. This means that a novel organism will be regulated under the GMO regulatory framework only if it has been developed with the use of defined techniques. This approach is now challenged with the emergence of new techniques. In this paper, we describe regulatory and safety issues associated with the use of oligonucleotide-mediated mutagenesis to develop novel organisms. We present scientific arguments for not having organisms developed through this technique fall within the scope of the EU regulation on GMOs. We conclude that any political decision on this issue should be taken on the basis of a broad reflection at EU level, while avoiding discrepancies at international level.

  19. Deciphering Cis-Regulatory Element Mediated Combinatorial Regulation in Rice under Blast Infected Condition

    PubMed Central

    Deb, Arindam; Kundu, Sudip

    2015-01-01

    Combinations of cis-regulatory elements (CREs) present at the promoters facilitate the binding of several transcription factors (TFs), thereby altering the consequent gene expressions. Due to the eminent complexity of the regulatory mechanism, the combinatorics of CRE-mediated transcriptional regulation has been elusive. In this work, we have developed a new methodology that quantifies the co-occurrence tendencies of CREs present in a set of promoter sequences; these co-occurrence scores are filtered in three consecutive steps to test their statistical significance; and the significantly co-occurring CRE pairs are presented as networks. These networks of co-occurring CREs are further transformed to derive higher order of regulatory combinatorics. We have further applied this methodology on the differentially up-regulated gene-sets of rice tissues under fungal (Magnaporthe) infected conditions to demonstrate how it helps to understand the CRE-mediated combinatorial gene regulation. Our analysis includes a wide spectrum of biologically important results. The CRE pairs having a strong tendency to co-occur often exhibit very similar joint distribution patterns at the promoters of rice. We couple the network approach with experimental results of plant gene regulation and defense mechanisms and find evidences of auto and cross regulation among TF families, cross-talk among multiple hormone signaling pathways, similarities and dissimilarities in regulatory combinatorics between different tissues, etc. Our analyses have pointed a highly distributed nature of the combinatorial gene regulation facilitating an efficient alteration in response to fungal attack. All together, our proposed methodology could be an important approach in understanding the combinatorial gene regulation. It can be further applied to unravel the tissue and/or condition specific combinatorial gene regulation in other eukaryotic systems with the availability of annotated genomic sequences and suitable

  20. T regulatory Cell–mediated Immunotherapy for Solid Organ Transplantation: A Clinical Perspective

    PubMed Central

    Khan, Mohammad Afzal

    2016-01-01

    T regulatory cells (Tregs) play a vital role in suppressing heightened immune response, and thereby promote a state of immunological tolerance. Tregs modulate both innate and adaptive immunity, which makes them potential candidates for cell-based immunotherapy in the suppression of uncontrolled activation of graft-specific inflammatory cells and toxic mediators. Graft-specific inflammatory cells (T effector cells) and other inflammatory mediators (immunoglobulins, active complement mediators) are mainly responsible for graft vascular deterioration followed by acute/chronic rejection. Treg-mediated immunotherapy is under investigation to induce allospecific tolerance in various ongoing clinical trials in organ transplant recipients. Treg immunotherapy shows promising results; however, key issues regarding Treg immunotherapy remain unresolved, including the mechanism of action and specific Treg cell phenotypes responsible for a state of tolerance. This review highlights the involvement of various subsets of Tregs during immune suppression, the novelty of Treg functions, effects on angiogenesis, emerging technologies for effective Treg expansion, and plasticity and safety associated with clinical applications. Altogether, this information will assist in designing single/combined Treg-mediated therapies for successful clinical trials in solid organ transplantations. PMID:27878210

  1. Mediation Analysis in a Latent Growth Curve Modeling Framework

    ERIC Educational Resources Information Center

    von Soest, Tilmann; Hagtvet, Knut A.

    2011-01-01

    This article presents several longitudinal mediation models in the framework of latent growth curve modeling and provides a detailed account of how such models can be constructed. Logical and statistical challenges that might arise when such analyses are conducted are also discussed. Specifically, we discuss how the initial status (intercept) and…

  2. Perception and regulatory principles of microbial growth control.

    PubMed

    Khonsari, Armin S; Kollmann, Markus

    2015-01-01

    Fast growth represents an effective strategy for microbial organisms to survive in competitive environments. To accomplish this task, cells must adapt their metabolism to changing nutrient conditions in a way that maximizes their growth rate. However, the regulation of the growth related metabolic pathways can be fundamentally different among microbes. We therefore asked whether growth control by perception of the cell's intracellular metabolic state can give rise to higher growth than by direct perception of extracellular nutrient availability. To answer this question, we created a simplified dynamical computer model of a cellular metabolic network whose regulation was inferred by an optimization approach. We used this model for a competing species experiment, where a species with extracellular nutrient perception competes against one with intracellular nutrient perception by evaluating their respective average growth rate. We found that the intracellular perception is advantageous under situations where the up and down regulation of pathways cannot follow the fast changing nutrient availability in the environment. In this case, optimal regulation ignores any other nutrients except the most preferential ones, in agreement with the phenomenon of catabolite repression in prokaryotes. The corresponding metabolic pathways remain activated, despite environmental fluctuations. Therefore, the cell can take up preferential nutrients as soon as they are available without any prior regulation. As a result species that rely on intracellular perception gain a relevant fitness advantage in fluctuating nutrient environments, which enables survival by outgrowing competitors.

  3. Regulatory Focus as a Mediator of the Influence of Initiating Structure and Servant Leadership on Employee Behavior

    ERIC Educational Resources Information Center

    Neubert, Mitchell J.; Kacmar, K. Michele; Carlson, Dawn S.; Chonko, Lawrence B.; Roberts, James A.

    2008-01-01

    In this research, the authors test a model in which the regulatory focus of employees at work mediates the influence of leadership on employee behavior. In a nationally representative sample of 250 workers who responded over 2 time periods, prevention focus mediated the relationship of initiating structure to in-role performance and deviant…

  4. Reappraisal and suppression mediate the contribution of regulatory focus to anxiety in healthy adults.

    PubMed

    Llewellyn, Nicole; Dolcos, Sanda; Iordan, Alexandru D; Rudolph, Karen D; Dolcos, Florin

    2013-08-01

    Theory and research link regulatory focus (RF) in the form of promotion and prevention goal orientation with internalizing symptoms (e.g., anxiety), but the relevant mechanisms are not well understood. This study investigated the role of two emotion regulation (ER) strategies (cognitive reappraisal and expressive suppression) as possible mediators. Path analysis using data from 179 healthy young participants (110 women, 69 men) revealed that stronger promotion orientation was significantly associated with less anxiety, and that the use of reappraisal and suppression partially mediated this association. Prevention was associated with more suppression but was not directly associated with anxiety. There were no gender differences in these effects. Collectively, these findings suggest that effective ER, through heightened use of reappraisal and dampened use of suppression, serves as a mechanism through which promotion confers protection against anxiety. This research provides empirical support to initiatives aimed at promoting healthy psychological adjustment and preventing anxiety, by optimizing ER strategies with respect to RF goal orientations.

  5. Advantages of mixing bioinformatics and visualization approaches for analyzing sRNA-mediated regulatory bacterial networks

    PubMed Central

    Bourqui, Romain; Benchimol, William; Gaspin, Christine; Sirand-Pugnet, Pascal; Uricaru, Raluca; Dutour, Isabelle

    2015-01-01

    The revolution in high-throughput sequencing technologies has enabled the acquisition of gigabytes of RNA sequences in many different conditions and has highlighted an unexpected number of small RNAs (sRNAs) in bacteria. Ongoing exploitation of these data enables numerous applications for investigating bacterial transacting sRNA-mediated regulation networks. Focusing on sRNAs that regulate mRNA translation in trans, recent works have noted several sRNA-based regulatory pathways that are essential for key cellular processes. Although the number of known bacterial sRNAs is increasing, the experimental validation of their interactions with mRNA targets remains challenging and involves expensive and time-consuming experimental strategies. Hence, bioinformatics is crucial for selecting and prioritizing candidates before designing any experimental work. However, current software for target prediction produces a prohibitive number of candidates because of the lack of biological knowledge regarding the rules governing sRNA–mRNA interactions. Therefore, there is a real need to develop new approaches to help biologists focus on the most promising predicted sRNA–mRNA interactions. In this perspective, this review aims at presenting the advantages of mixing bioinformatics and visualization approaches for analyzing predicted sRNA-mediated regulatory bacterial networks. PMID:25477348

  6. Understanding microRNA-mediated gene regulatory networks through mathematical modelling

    PubMed Central

    Lai, Xin; Wolkenhauer, Olaf; Vera, Julio

    2016-01-01

    The discovery of microRNAs (miRNAs) has added a new player to the regulation of gene expression. With the increasing number of molecular species involved in gene regulatory networks, it is hard to obtain an intuitive understanding of network dynamics. Mathematical modelling can help dissecting the role of miRNAs in gene regulatory networks, and we shall here review the most recent developments that utilise different mathematical modelling approaches to provide quantitative insights into the function of miRNAs in the regulation of gene expression. Key miRNA regulation features that have been elucidated via modelling include: (i) the role of miRNA-mediated feedback and feedforward loops in fine-tuning of gene expression; (ii) the miRNA–target interaction properties determining the effectiveness of miRNA-mediated gene repression; and (iii) the competition for shared miRNAs leading to the cross-regulation of genes. However, there is still lack of mechanistic understanding of many other properties of miRNA regulation like unconventional miRNA–target interactions, miRNA regulation at different sub-cellular locations and functional miRNA variant, which will need future modelling efforts to deal with. This review provides an overview of recent developments and challenges in this field. PMID:27317695

  7. An assessment of the contribution of NRC (Nuclear Regulatory Commission) regulatory growth to nuclear plant cost growth using engineering scope changes

    SciTech Connect

    Cohen, S.

    1987-03-01

    The purpose of this study is to determine the contribution of NRC regulations to the growth in nuclear power plant capital costs using the case study method. The two plants selected for the case studies are Florida Power and Light Company's (FP and L) St. Lucie Unit 1 (SL1) and St. Lucie Unit 2 (SL2). SL1 was constructed in the early 1970s and was granted an operating license in 1976. SL2 was constructed in the late 1970s and early 1980s, and was granted an operating license in 1983. The information bases were the amendments to the contracts between FP and L and its architect-engineer/constructor, i.e., the ''scope changes''. These were examined and analyzed for causation, i.e., NRC-initiated or utility-initiated, and all of the costs associated with scope changes of each type were aggregated to determine the contribution of each. Although the scope changes accounted for only a small fraction of the total cost growth for either plant, they were still used to determine the relative contribution of regulatory growth to cost growth. Unexpectedly, a significantly higher percentage of out-of-scope work (approximately 84%) was attributable to NRC regulatory requirements for SL1 than SL2 (approximately 47%). These results were unexpected because SL2 was constructed during a period in which regulation was considered to be particularly unstable. However, a more detailed analysis of causation indicates that a shift occurred from an ad-hoc mode of regulation in the early 1970s to a more prescriptive process in the late 1970s. Thus the number of formal NRC requirements may not be a valid measure of regulatory stability.

  8. Intergenic GWAS SNPs are key components of the spatial and regulatory network for human growth.

    PubMed

    Schierding, William; Antony, Jisha; Cutfield, Wayne S; Horsfield, Julia A; O'Sullivan, Justin M

    2016-08-01

    Meta-analysis of genome-wide association studies has resulted in the identification of hundreds of genetic variants associated with growth and stature. Determining how these genetic variants influence growth is important, but most are non-coding, and there is little understanding of how these variants contribute to adult height. To determine the mechanisms by which human variation contributes to growth, we combined spatial genomic connectivity (high-throughput conformation capture) with functional (gene expression, expression Quantitative Trait Loci) data to determine how non-genic loci associated with infant length, pubertal and adult height and contribute to gene regulatory networks. This approach identified intergenic single-nucleotide polymorphisms (SNPs) ∼85 kb upstream of FBXW11 that spatially connect with distant loci. These regulatory connections are reinforced by evidence of SNP-enhancer effects and altered expression in genes influencing the action of human growth hormone. Functional assays provided evidence for enhancer activity of the intergenic region near FBXW11 that harbors SNP rs12153391, which is associated with an expression Quantitative Trait Loci. Our results suggest that variants in this locus have genome-wide effects as key modifiers of growth (both overgrowth and short stature) acting through a regulatory network. We believe that the genes and pathways connected with this regulatory network are potential targets that could be investigated for diagnostic, prenatal and carrier testing for growth disorders. Finally, the regulatory networks we generated illustrate the power of using existing datasets to interrogate the contribution of intergenic SNPs to common syndromes/diseases.

  9. Growth factors as mediators of exercise actions on the brain.

    PubMed

    Llorens-Martín, M; Torres-Alemán, I; Trejo, José L

    2008-01-01

    Physical exercise has long been recognized as highly beneficial for brain and body health. The molecular mechanisms responsible for translation of exercise stimuli in the brain have claimed attention due to mounting evidence for the neuroprotective actions of the exercise and its positive effects in preventing both ageing and neurodegenerative disease. These molecular mediators are currently under investigation with new tools able to yield deep insights into the neurobiology of exercise. In the present work we focus on the evidence pertaining to the mediation of exercise effects by insulin-like growth factor 1 (IGF1), as recent reports suggest that this growth factor shows brain area-specific, temporal rank-sensitive, and behavioural task-dependent features in response to exercise.

  10. Penfluridol suppresses pancreatic tumor growth by autophagy-mediated apoptosis

    PubMed Central

    Ranjan, Alok; Srivastava, Sanjay K.

    2016-01-01

    Pancreatic tumors exhibit enhanced autophagy as compared to any other cancer, making it resistant to chemotherapy. We evaluated the effect of penfluridol against pancreatic cancer. Penfluridol treatment induced apoptosis and inhibited the growth of Panc-1, BxPC-3 and AsPC-1, pancreatic cancer cells with IC50 ranging between 6–7 μM after 24 h of treatment. Significant autophagy was induced by penfluridol treatment in pancreatic cancer cells. Punctate LC3B and autophagosomes staining confirmed autophagy. Inhibiting autophagy by chloroquine, bafilomycin, 3-methyladenine or LC3BsiRNA, significantly blocked penfluridol-induced apoptosis, suggesting that autophagy lead to apoptosis in our model. Penfluridol treatment suppressed the growth of BxPC-3 tumor xenografts by 48% as compared to 17% when treated in combination with chloroquine. Similarly, penfluridol suppressed the growth of AsPC-1 tumors by 40% versus 16% when given in combination with chloroquine. TUNEL staining and caspase-3 cleavage revealed less apoptosis in the tumors from mice treated with penfluridol and chloroquine as compared to penfluridol alone. Penfluridol treatment also suppressed the growth of orthotopically implanted Panc-1 tumors by 80% by inducing autophagy-mediated apoptosis in the tumors. These studies established that penfluridol inhibits pancreatic tumor growth by autophagy-mediated apoptosis. Since penfluridol is already in clinic, positive findings from our study will accelerate its clinical development. PMID:27189859

  11. TGF-β Controls miR-181/ERK Regulatory Network during Retinal Axon Specification and Growth

    PubMed Central

    Carrella, Sabrina; Salierno, Francesco Giuseppe; Manfredi, Anna; Banfi, Sandro; Conte, Ivan

    2015-01-01

    Retinal axon specification and growth are critically sensitive to the dosage of numerous signaling molecules and transcription factors. Subtle variations in the expression levels of key molecules may result in a variety of axonal growth anomalies. miR-181a and miR-181b are two eye-enriched microRNAs whose inactivation in medaka fish leads to alterations of the proper establishment of connectivity and function in the visual system. miR-181a/b are fundamental regulators of MAPK signaling and their role in retinal axon growth and specification is just beginning to be elucidated. Here we demonstrate that miR-181a/b are key nodes in the interplay between TGF-β and MAPK/ERK within the functional pathways that control retinal axon specification and growth. Using a variety of in vivo and in vitro approaches in medaka fish, we demonstrate that TGF-β signaling controls the miR-181/ERK regulatory network, which in turn strengthens the TGF-β-mediated regulation of RhoA degradation. Significantly, these data uncover the role of TGF-β signaling in vivo, for the first time, in defining the correct wiring and assembly of functional retina neural circuits and further highlight miR-181a/b as key factors in axon specification and growth. PMID:26641497

  12. Regulatory focus as a mediator of the influence of initiating structure and servant leadership on employee behavior.

    PubMed

    Neubert, Mitchell J; Kacmar, K Michele; Carlson, Dawn S; Chonko, Lawrence B; Roberts, James A

    2008-11-01

    In this research, the authors test a model in which the regulatory focus of employees at work mediates the influence of leadership on employee behavior. In a nationally representative sample of 250 workers who responded over 2 time periods, prevention focus mediated the relationship of initiating structure to in-role performance and deviant behavior, whereas promotion focus mediated the relationship of servant leadership to helping and creative behavior. The results indicate that even though initiating structure and servant leadership share some variance in explaining other variables, each leadership style incrementally predicts disparate outcomes after controlling for the other style and dispositional tendencies. A new regulatory focus scale, the Work Regulatory Focus (WRF) Scale, also was developed and initially validated for this study. Implications for the results and the WRF Scale are discussed.

  13. PKA regulatory IIα subunit is essential for PGD2-mediated resolution of inflammation.

    PubMed

    Kong, Deping; Shen, Yujun; Liu, Guizhu; Zuo, Shengkai; Ji, Yong; Lu, Ankang; Nakamura, Masataka; Lazarus, Michael; Stratakis, Constantine A; Breyer, Richard M; Yu, Ying

    2016-09-19

    The kinetic participation of macrophages is critical for inflammatory resolution and recovery from myocardial infarction (MI), particularly with respect to the transition from the M1 to the M2 phenotype; however, the underlying mechanisms are poorly understood. In this study, we found that the deletion of prostaglandin (PG) D2 receptor subtype 1 (DP1) in macrophages retarded M2 polarization, antiinflammatory cytokine production, and resolution in different inflammatory models, including the MI model. DP1 deletion up-regulated proinflammatory genes expression via JAK2/STAT1 signaling in macrophages, whereas its activation facilitated binding of the separated PKA regulatory IIα subunit (PRKAR2A) to the transmembrane domain of IFN-γ receptor, suppressed JAK2-STAT1 axis-mediated M1 polarization, and promoted resolution. PRKAR2A deficiency attenuated DP1 activation-mediated M2 polarization and resolution of inflammation. Collectively, PGD2-DP1 axis-induced M2 polarization facilitates resolution of inflammation through the PRKAR2A-mediated suppression of JAK2/STAT1 signaling. These observations indicate that macrophage DP1 activation represents a promising strategy in the management of inflammation-associated diseases, including post-MI healing.

  14. Regulatory Effect of Cinnamaldehyde on Monocyte/Macrophage-Mediated Inflammatory Responses

    PubMed Central

    Kim, Byung Hun; Lee, Yong Gyu; Lee, Jaehwi; Lee, Joo Young; Cho, Jae Youl

    2010-01-01

    Cinnamaldehyde (CA) has been known to exhibit anti-inflammatory and anticancer effects. Although numerous pharmacological effects have been demonstrated, regulatory effect of CA on the functional activation of monocytes and macrophages has not been fully elucidated yet. To evaluate its monocyte/macrophage-mediated immune responses, macrophages activated by lipopolysaccharide (LPS), and monocytes treated with proaggregative antibodies, and extracellular matrix protein fibronectin were employed. CA was able to suppress both the production of nitric oxide (NO) and upregulation of surface levels of costimulatory molecules (CD80 and CD69) and pattern recognition receptors (toll-like receptor 2 (TLR2) and complement receptor (CR3)). In addition, CA also blocked cell-cell adhesion induced by the activation of CD29 and CD43 but not cell-fibronectin adhesion. Immunoblotting analysis suggested that CA inhibition was due to the inhibition of phosphoinositide-3-kinase (PI3K) and phosphoinositide-dependent kinase (PDK)1 as well as nuclear factor-(NF-) κB activation. In particular, thiol compounds with sulphydryl group, L-cysteine and dithiothreitol (DTT), strongly abrogated CA-mediated NO production and NF-κB activation. Therefore, our results suggest that CA can act as a strong regulator of monocyte/macrophage-mediated immune responses by thiolation of target cysteine residues in PI3K or PDK1. PMID:20467561

  15. Regulatory effect of cinnamaldehyde on monocyte/macrophage-mediated inflammatory responses.

    PubMed

    Kim, Byung Hun; Lee, Yong Gyu; Lee, Jaehwi; Lee, Joo Young; Cho, Jae Youl

    2010-01-01

    Cinnamaldehyde (CA) has been known to exhibit anti-inflammatory and anticancer effects. Although numerous pharmacological effects have been demonstrated, regulatory effect of CA on the functional activation of monocytes and macrophages has not been fully elucidated yet. To evaluate its monocyte/macrophage-mediated immune responses, macrophages activated by lipopolysaccharide (LPS), and monocytes treated with proaggregative antibodies, and extracellular matrix protein fibronectin were employed. CA was able to suppress both the production of nitric oxide (NO) and upregulation of surface levels of costimulatory molecules (CD80 and CD69) and pattern recognition receptors (toll-like receptor 2 (TLR2) and complement receptor (CR3)). In addition, CA also blocked cell-cell adhesion induced by the activation of CD29 and CD43 but not cell-fibronectin adhesion. Immunoblotting analysis suggested that CA inhibition was due to the inhibition of phosphoinositide-3-kinase (PI3K) and phosphoinositide-dependent kinase (PDK)1 as well as nuclear factor-(NF-) kappaB activation. In particular, thiol compounds with sulphydryl group, L-cysteine and dithiothreitol (DTT), strongly abrogated CA-mediated NO production and NF-kappaB activation. Therefore, our results suggest that CA can act as a strong regulator of monocyte/macrophage-mediated immune responses by thiolation of target cysteine residues in PI3K or PDK1.

  16. PKA regulatory IIα subunit is essential for PGD2-mediated resolution of inflammation

    PubMed Central

    Shen, Yujun; Zuo, Shengkai; Ji, Yong; Nakamura, Masataka; Stratakis, Constantine A.

    2016-01-01

    The kinetic participation of macrophages is critical for inflammatory resolution and recovery from myocardial infarction (MI), particularly with respect to the transition from the M1 to the M2 phenotype; however, the underlying mechanisms are poorly understood. In this study, we found that the deletion of prostaglandin (PG) D2 receptor subtype 1 (DP1) in macrophages retarded M2 polarization, antiinflammatory cytokine production, and resolution in different inflammatory models, including the MI model. DP1 deletion up-regulated proinflammatory genes expression via JAK2/STAT1 signaling in macrophages, whereas its activation facilitated binding of the separated PKA regulatory IIα subunit (PRKAR2A) to the transmembrane domain of IFN-γ receptor, suppressed JAK2–STAT1 axis–mediated M1 polarization, and promoted resolution. PRKAR2A deficiency attenuated DP1 activation–mediated M2 polarization and resolution of inflammation. Collectively, PGD2–DP1 axis–induced M2 polarization facilitates resolution of inflammation through the PRKAR2A-mediated suppression of JAK2/STAT1 signaling. These observations indicate that macrophage DP1 activation represents a promising strategy in the management of inflammation-associated diseases, including post-MI healing. PMID:27621415

  17. Receptor-mediated control of regulatory volume decrease (RVD) and apoptotic volume decrease (AVD)

    PubMed Central

    Okada, Yasunobu; Maeno, Emi; Shimizu, Takahiro; Dezaki, Katsuya; Wang, Jun; Morishima, Shigeru

    2001-01-01

    A fundamental property of animal cells is the ability to regulate their own cell volume. Even under hypotonic stress imposed by either decreased extracellular or increased intracellular osmolarity, the cells can re-adjust their volume after transient osmotic swelling by a mechanism known as regulatory volume decrease (RVD). In most cell types, RVD is accomplished mainly by KCl efflux induced by parallel activation of K+ and Cl− channels. We have studied the molecular mechanism of RVD in a human epithelial cell line (Intestine 407). Osmotic swelling results in a significant increase in the cytosolic Ca2+ concentration and thereby activates intermediate-conductance Ca2+-dependent K+ (IK) channels. Osmotic swelling also induces ATP release from the cells to the extracellular compartment. Released ATP stimulates purinergic ATP (P2Y2) receptors, thereby inducing phospholipase C-mediated Ca2+ mobilization. Thus, RVD is facilitated by stimulation of P2Y2 receptors due to augmentation of IK channels. In contrast, stimulation of another G protein-coupled Ca2+-sensing receptor (CaR) enhances the activity of volume-sensitive outwardly rectifying Cl− channels, thereby facilitating RVD. Therefore, it is possible that Ca2+ efflux stimulated by swelling-induced and P2Y2 receptor-mediated intracellular Ca2+ mobilization activates the CaR, thereby secondarily upregulating the volume-regulatory Cl− conductance. On the other hand, the initial process towards apoptotic cell death is coupled to normotonic cell shrinkage, called apoptotic volume decrease (AVD). Stimulation of death receptors, such as TNFα receptor and Fas, induces AVD and thereafter biochemical apoptotic events in human lymphoid (U937), human epithelial (HeLa), mouse neuroblastoma × rat glioma hybrid (NG108-15) and rat phaeochromocytoma (PC12) cells. In those cells exhibiting AVD, facilitation of RVD is always observed. Both AVD induction and RVD facilitation as well as succeeding apoptotic events can be

  18. Twin-mediated crystal growth: an enigma resolved

    NASA Astrophysics Data System (ADS)

    Shahani, Ashwin J.; Gulsoy, E. Begum; Poulsen, Stefan O.; Xiao, Xianghui; Voorhees, Peter W.

    2016-06-01

    During crystal growth, faceted interfaces may be perturbed by defects, leading to a rich variety of polycrystalline growth forms. One such defect is the coherent Σ3 {111} twin boundary, which is widely known to catalyze crystal growth. These defects have a profound effect on the properties of many materials: for example, electron-hole recombination rates strongly depend on the character of the twin boundaries in polycrystalline Si photovoltaic cells. However, the morphology of the twinned interface during growth has long been a mystery due to the lack of four-dimensional (i.e., space and time resolved) experiments. Many controversial mechanisms have been proposed for this process, most of which lack experimental verification. Here, we probe the real-time interfacial dynamics of polycrystalline Si particles growing from an Al-Si-Cu liquid via synchrotron-based X-ray tomography. Our novel analysis of the time evolution of the interfacial normals allows us to quantify unambiguously the habit plane and grain boundary orientations during growth. This, when combined with direct measurements of the interfacial morphology provide the first confirmation of twin-mediated growth, proposed over 50 years ago. Using the insights provided by these experiments, we have developed a unified picture of the phenomena responsible for the dynamics of faceted Si growth.

  19. Thyroid hormone mediates otolith growth and development during flatfish metamorphosis.

    PubMed

    Schreiber, A M; Wang, X; Tan, Y; Sievers, Q; Sievers, B; Lee, M; Burrall, K

    2010-11-01

    Flatfish begin life as bilaterally symmetrical larvae that swim up-right, then abruptly metamorphose into asymmetrically shaped juveniles with lateralized swimming postures. Flatfish metamorphosis is mediated entirely by thyroid hormone (TH). Changes in flatfish swim posture are thought to be regulated via vestibular remodeling, although the influence of TH on teleost inner ear development remains unclear. This study addresses the role of TH on the development of the three otolith end-organs (sacculus, utricle, and lagena) during southern flounder (Paralichthys lethostigma) metamorphosis. Compared with pre-metamorphosis, growth rates of the sacculus and utricle otoliths increase dramatically during metamorphosis in a manner that is uncoupled from general somatic growth. Treatment of P. lethostigma larvae with methimazol (a pharmacological inhibitor of endogenous TH production) inhibits growth of the sacculus and utricle, whereas treatment with TH dramatically accelerates their growth. In contrast with the sacculus and utricle otoliths that begin to form and mineralize during embryogenesis, a non-mineralized lagena otolith is first visible 10-12 days after hatching. The lagena grows during pre- and pro-metamorphosis, then abruptly mineralizes during metamorphic climax. Mineralization of the lagena, but not growth, can be induced with TH treatment, whereas treatment with methimazol completely inhibits lagena mineralization without inhibiting its growth. These findings suggest that during southern flounder metamorphosis TH exerts differential effects on growth and development among the three types of otolith.

  20. Twin-mediated crystal growth: an enigma resolved

    PubMed Central

    Shahani, Ashwin J.; Gulsoy, E. Begum; Poulsen, Stefan O.; Xiao, Xianghui; Voorhees, Peter W.

    2016-01-01

    During crystal growth, faceted interfaces may be perturbed by defects, leading to a rich variety of polycrystalline growth forms. One such defect is the coherent Σ3 {111} twin boundary, which is widely known to catalyze crystal growth. These defects have a profound effect on the properties of many materials: for example, electron-hole recombination rates strongly depend on the character of the twin boundaries in polycrystalline Si photovoltaic cells. However, the morphology of the twinned interface during growth has long been a mystery due to the lack of four-dimensional (i.e., space and time resolved) experiments. Many controversial mechanisms have been proposed for this process, most of which lack experimental verification. Here, we probe the real-time interfacial dynamics of polycrystalline Si particles growing from an Al-Si-Cu liquid via synchrotron-based X-ray tomography. Our novel analysis of the time evolution of the interfacial normals allows us to quantify unambiguously the habit plane and grain boundary orientations during growth. This, when combined with direct measurements of the interfacial morphology provide the first confirmation of twin-mediated growth, proposed over 50 years ago. Using the insights provided by these experiments, we have developed a unified picture of the phenomena responsible for the dynamics of faceted Si growth. PMID:27346073

  1. Intrinsic Mathematics Motivation as a Mediator between Regulatory Fit and Mathematics Performance

    ERIC Educational Resources Information Center

    Van Slooten, Courtney

    2013-01-01

    Regulatory Fit Theory research has indicated that the presence of a regulatory fit between an individuals chronic regulatory focus orientation and induced regulatory state can increase motivation, performance, and achievement. The research in support of regulatory fit theory is immense; however, there has been little research that focuses on…

  2. Junction-mediating and regulatory protein (JMY) is essential for early porcine embryonic development.

    PubMed

    Lin, Zi Li; Cui, Xiang-Shun; Namgoong, Suk; Kim, Nam-Hyung

    2015-01-01

    Junction-mediating and regulatory protein (JMY) is a regulator of both transcription and actin filament assembly. JMY is a critical nucleation-promoting factor (NPF); however, its role in the development of mammalian embryos is poorly understood. In the current study, we investigated the functional roles of the NPF JMY in porcine embryos. Porcine embryos expressed JMY mRNA and protein, and JMY protein moved from the cytoplasm to the nucleus at later embryonic developmental stages. Knockdown of JMY by RNA interference markedly decreased the rate of blastocyst development, validating its role in the development of porcine embryos. Furthermore, injection of JMY dsRNA also impaired actin and Arp2 expression, and co-injection of actin and Arp2 mRNA partially rescued blastocyst development. Taken together, our results show that the NPF JMY is involved in the development of porcine embryos by regulating the NPF-Arp2-actin pathway.

  3. Junction-mediating and regulatory protein (JMY) is essential for early porcine embryonic development

    PubMed Central

    LIN, Zi Li; CUI, Xiang-Shun; NAMGOONG, Suk; KIM, Nam-Hyung

    2015-01-01

    Junction-mediating and regulatory protein (JMY) is a regulator of both transcription and actin filament assembly. JMY is a critical nucleation-promoting factor (NPF); however, its role in the development of mammalian embryos is poorly understood. In the current study, we investigated the functional roles of the NPF JMY in porcine embryos. Porcine embryos expressed JMY mRNA and protein, and JMY protein moved from the cytoplasm to the nucleus at later embryonic developmental stages. Knockdown of JMY by RNA interference markedly decreased the rate of blastocyst development, validating its role in the development of porcine embryos. Furthermore, injection of JMY dsRNA also impaired actin and Arp2 expression, and co-injection of actin and Arp2 mRNA partially rescued blastocyst development. Taken together, our results show that the NPF JMY is involved in the development of porcine embryos by regulating the NPF-Arp2-actin pathway. PMID:26052154

  4. A wave of regulatory T cells into neonatal skin mediates tolerance to commensal microbes

    PubMed Central

    Scharschmidt, Tiffany C.; Vasquez, Kimberly S.; Truong, Hong-An; Gearty, Sofia V.; Pauli, Mariela L.; Nosbaum, Audrey; Gratz, Iris K.; Otto, Michael; Moon, James J.; Liese, Jan; Abbas, Abul K.; Fischbach, Michael A.; Rosenblum, Michael D.

    2015-01-01

    Summary The skin is a site of constant dialogue between the immune system and commensal bacteria. However, the molecular mechanisms that allow us to tolerate the presence of skin commensals without eliciting destructive inflammation are unknown. Using a model system to study the antigen-specific response to S. epidermidis, we demonstrated that skin colonization during a defined period of neonatal life was required to establish immune tolerance to commensal microbes. This crucial window was characterized by an abrupt influx of highly activated regulatory T (Treg) cells into neonatal skin. Selective inhibition of this Treg cell wave completely abrogated tolerance. Thus, the host-commensal relationship in the skin relied on a unique Treg cell population that mediated tolerance to bacterial antigens during a defined developmental window. This suggests that the cutaneous microbiome composition in neonatal life is crucial in shaping adaptive immune responses to commensals, and disrupting these interactions may have enduring health implications. PMID:26588783

  5. A multilevel examination of the relationships among training outcomes, mediating regulatory processes, and adaptive performance.

    PubMed

    Chen, Gilad; Thomas, Brian; Wallace, J Craig

    2005-09-01

    This study examined whether cognitive, affective-motivational, and behavioral training outcomes relate to posttraining regulatory processes and adaptive performance similarly at the individual and team levels of analysis. Longitudinal data were collected from 156 individuals composing 78 teams who were trained on and then performed a simulated flight task. Results showed that posttraining regulation processes related similarly to adaptive performance across levels. Also, regulation processes fully mediated the influences of self- and collective efficacy beliefs on individual and team adaptive performance. Finally, knowledge and skill more strongly and directly related to adaptive performance at the individual than the team level of analysis. Implications to theory and practice, limitations, and future directions are discussed.

  6. A Wave of Regulatory T Cells into Neonatal Skin Mediates Tolerance to Commensal Microbes.

    PubMed

    Scharschmidt, Tiffany C; Vasquez, Kimberly S; Truong, Hong-An; Gearty, Sofia V; Pauli, Mariela L; Nosbaum, Audrey; Gratz, Iris K; Otto, Michael; Moon, James J; Liese, Jan; Abbas, Abul K; Fischbach, Michael A; Rosenblum, Michael D

    2015-11-17

    The skin is a site of constant dialog between the immune system and commensal bacteria. However, the molecular mechanisms that allow us to tolerate the presence of skin commensals without eliciting destructive inflammation are unknown. Using a model system to study the antigen-specific response to S. epidermidis, we demonstrated that skin colonization during a defined period of neonatal life was required for establishing immune tolerance to commensal microbes. This crucial window was characterized by an abrupt influx of highly activated regulatory T (Treg) cells into neonatal skin. Selective inhibition of this Treg cell wave completely abrogated tolerance. Thus, the host-commensal relationship in the skin relied on a unique Treg cell population that mediated tolerance to bacterial antigens during a defined developmental window. This suggests that the cutaneous microbiome composition in neonatal life is crucial in shaping adaptive immune responses to commensals, and disrupting these interactions might have enduring health implications.

  7. β-arrestins: regulatory role and therapeutic potential in opioid and cannabinoid receptor-mediated analgesia.

    PubMed

    Raehal, Kirsten M; Bohn, Laura M

    2014-01-01

    Pain is a complex disorder with neurochemical and psychological components contributing to the severity, the persistence, and the difficulty in adequately treating the condition. Opioid and cannabinoids are two classes of analgesics that have been used to treat pain for centuries and are arguably the oldest of "pharmacological" interventions used by man. Unfortunately, they also produce several adverse side effects that can complicate pain management. Opioids and cannabinoids act at G protein-coupled receptors (GPCRs), and much of their effects are mediated by the mu-opioid receptor (MOR) and cannabinoid CB1 receptor (CB1R), respectively. These receptors couple to intracellular second messengers and regulatory proteins to impart their biological effects. In this chapter, we review the role of the intracellular regulatory proteins, β-arrestins, in modulating MOR and CB1R and how they influence the analgesic and side-effect profiles of opioid and cannabinoid drugs in vivo. This review of the literature suggests that the development of opioid and cannabinoid agonists that bias MOR and CB1R toward G protein signaling cascades and away from β-arrestin interactions may provide a novel mechanism by which to produce analgesia with less severe adverse effects.

  8. β-Arrestins: Regulatory Role and Therapeutic Potential in Opioid and Cannabinoid Receptor-Mediated Analgesia

    PubMed Central

    Bohn, Laura M.

    2016-01-01

    Pain is a complex disorder with neurochemical and psychological components contributing to the severity, the persistence, and the difficulty in adequately treating the condition. Opioid and cannabinoids are two classes of analgesics that have been used to treat pain for centuries and are arguably the oldest of “pharmacological” interventions used by man. Unfortunately, they also produce several adverse side effects that can complicate pain management. Opioids and cannabinoids act at G protein-coupled receptors (GPCRs), and much of their effects are mediated by the mu-opioid receptor (MOR) and cannabinoid CB1 receptor (CB1R), respectively. These receptors couple to intracellular second messengers and regulatory proteins to impart their biological effects. In this chapter, we review the role of the intracellular regulatory proteins, β-arrestins, in modulating MOR and CB1R and how they influence the analgesic and side-effect profiles of opioid and cannabinoid drugs in vivo. This review of the literature suggests that the development of opioid and cannabinoid agonists that bias MOR and CB1R toward G protein signaling cascades and away from β-arrestin interactions may provide a novel mechanism by which to produce analgesia with less severe adverse effects. PMID:24292843

  9. Self-Regulatory Processes Mediating between Career Calling and Perceived Employability and Life Satisfaction in Emerging Adults

    ERIC Educational Resources Information Center

    Praskova, Anna; Creed, Peter A.; Hood, Michelle

    2015-01-01

    We tested a cross-sectional, mediation model of career calling, in which career calling was associated positively with life satisfaction and perceptions of future employability, and these relationships were explained by the self-regulatory mechanisms of work effort, career strategies, and emotional regulation. Using a sample of 664 emerging adults…

  10. Growth hormone STAT5-mediated signaling and its modulation in mice liver during the growth period.

    PubMed

    Martinez, Carolina S; Piazza, Verónica G; Ratner, Laura D; Matos, Marina N; González, Lorena; Rulli, Susana B; Miquet, Johanna G; Sotelo, Ana I

    2013-01-01

    Postnatal growth exhibits two instances of rapid growth in mice: the first is perinatal and independent of growth hormone (GH), the second is peripuberal and GH-dependent. Signal transducer and activator of transcription 5b (STAT5b) is the main GH-signaling mediator and it is related to IGF1 synthesis and somatic growth. The aim of this work was to assess differential STAT5 sensitivity to GH during the growth period in mouse liver of both sexes. Three representative ages were selected: 1-week-old animals, in the GH-independent phase of growth; 2.5-week-old mice, at the onset of the GH-dependent phase of growth; and 9-week-old young adults. GH-signaling mediators were assessed by immunoblotting, quantitative RT-PCR and immunohistochemistry. GH-induced STAT5 phosphorylation is low at one-week and maximal at 2.5-weeks of age when compared to young adults, accompanied by higher protein content at the onset of growth. Suppressor CIS and phosphatase PTP1B exhibit high levels in one-week animals, which gradually decline, while SOCS2 and SOCS3 display higher levels at adulthood. Nuclear phosphorylated STAT5 is low in one-week animals while in 2.5-week animals it is similar to 9-week control; expression of SOCS3, an early response GH-target gene, mimics this pattern. STAT5 coactivators glucocorticoid receptor (GR) and hepatic nuclear factor 1 (HNF1) abundance is higher in adulthood. Therefore, GH-induced STAT5 signaling presents age-dependent activity in liver, with its maximum coinciding with the onset of GH-dependent phase of growth, accompanied by an age-dependent variation of modulating factors. This work contributes to elucidate the molecular mechanisms implicated in GH responsiveness during growth.

  11. FOXA and master transcription factors recruit Mediator and Cohesin to the core transcriptional regulatory circuitry of cancer cells

    NASA Astrophysics Data System (ADS)

    Fournier, Michèle; Bourriquen, Gaëlle; Lamaze, Fabien C.; Côté, Maxime C.; Fournier, Éric; Joly-Beauparlant, Charles; Caron, Vicky; Gobeil, Stéphane; Droit, Arnaud; Bilodeau, Steve

    2016-10-01

    Controlling the transcriptional program is essential to maintain the identity and the biological functions of a cell. The Mediator and Cohesin complexes have been established as central cofactors controlling the transcriptional program in normal cells. However, the distribution, recruitment and importance of these complexes in cancer cells have not been fully investigated. Here we show that FOXA and master transcription factors are part of the core transcriptional regulatory circuitry of cancer cells and are essential to recruit M ediator and Cohesin. Indeed, Mediator and Cohesin occupied the enhancer and promoter regions of actively transcribed genes and maintained the proliferation and colony forming potential. Through integration of publically available ChIP-Seq datasets, we predicted the core transcriptional regulatory circuitry of each cancer cell. Unexpectedly, for all cells investigated, the pioneer transcription factors FOXA1 and/or FOXA2 were identified in addition to cell-specific master transcription factors. Loss of both types of transcription factors phenocopied the loss of Mediator and Cohesin. Lastly, the master and pioneer transcription factors were essential to recruit Mediator and Cohesin to regulatory regions of actively transcribed genes. Our study proposes that maintenance of the cancer cell state is dependent on recruitment of Mediator and Cohesin through FOXA and master transcription factors.

  12. FOXA and master transcription factors recruit Mediator and Cohesin to the core transcriptional regulatory circuitry of cancer cells.

    PubMed

    Fournier, Michèle; Bourriquen, Gaëlle; Lamaze, Fabien C; Côté, Maxime C; Fournier, Éric; Joly-Beauparlant, Charles; Caron, Vicky; Gobeil, Stéphane; Droit, Arnaud; Bilodeau, Steve

    2016-10-14

    Controlling the transcriptional program is essential to maintain the identity and the biological functions of a cell. The Mediator and Cohesin complexes have been established as central cofactors controlling the transcriptional program in normal cells. However, the distribution, recruitment and importance of these complexes in cancer cells have not been fully investigated. Here we show that FOXA and master transcription factors are part of the core transcriptional regulatory circuitry of cancer cells and are essential to recruit M ediator and Cohesin. Indeed, Mediator and Cohesin occupied the enhancer and promoter regions of actively transcribed genes and maintained the proliferation and colony forming potential. Through integration of publically available ChIP-Seq datasets, we predicted the core transcriptional regulatory circuitry of each cancer cell. Unexpectedly, for all cells investigated, the pioneer transcription factors FOXA1 and/or FOXA2 were identified in addition to cell-specific master transcription factors. Loss of both types of transcription factors phenocopied the loss of Mediator and Cohesin. Lastly, the master and pioneer transcription factors were essential to recruit Mediator and Cohesin to regulatory regions of actively transcribed genes. Our study proposes that maintenance of the cancer cell state is dependent on recruitment of Mediator and Cohesin through FOXA and master transcription factors.

  13. FOXA and master transcription factors recruit Mediator and Cohesin to the core transcriptional regulatory circuitry of cancer cells

    PubMed Central

    Fournier, Michèle; Bourriquen, Gaëlle; Lamaze, Fabien C.; Côté, Maxime C.; Fournier, Éric; Joly-Beauparlant, Charles; Caron, Vicky; Gobeil, Stéphane; Droit, Arnaud; Bilodeau, Steve

    2016-01-01

    Controlling the transcriptional program is essential to maintain the identity and the biological functions of a cell. The Mediator and Cohesin complexes have been established as central cofactors controlling the transcriptional program in normal cells. However, the distribution, recruitment and importance of these complexes in cancer cells have not been fully investigated. Here we show that FOXA and master transcription factors are part of the core transcriptional regulatory circuitry of cancer cells and are essential to recruit M ediator and Cohesin. Indeed, Mediator and Cohesin occupied the enhancer and promoter regions of actively transcribed genes and maintained the proliferation and colony forming potential. Through integration of publically available ChIP-Seq datasets, we predicted the core transcriptional regulatory circuitry of each cancer cell. Unexpectedly, for all cells investigated, the pioneer transcription factors FOXA1 and/or FOXA2 were identified in addition to cell-specific master transcription factors. Loss of both types of transcription factors phenocopied the loss of Mediator and Cohesin. Lastly, the master and pioneer transcription factors were essential to recruit Mediator and Cohesin to regulatory regions of actively transcribed genes. Our study proposes that maintenance of the cancer cell state is dependent on recruitment of Mediator and Cohesin through FOXA and master transcription factors. PMID:27739523

  14. Regulatory B cells preferentially accumulate in tumor-draining lymph nodes and promote tumor growth.

    PubMed

    Ganti, Sheila N; Albershardt, Tina C; Iritani, Brian M; Ruddell, Alanna

    2015-07-20

    Our previous studies found that B16-F10 melanoma growth in the rear footpad of immunocompetent mice induces marked B cell accumulation within tumor-draining popliteal lymph nodes (TDLN). This B cell accumulation drives TDLN remodeling that precedes and promotes metastasis, indicating a tumor-promoting role for TDLN B cells. Here we show that phenotypic characterization of lymphocytes in mice bearing B16-F10 melanomas identifies preferential accumulation of T2-MZP B cells in the TDLN. Comparison of non-draining LNs and spleens of tumor-bearing mice with LNs and spleens from naïve mice determined that this pattern of B cell accumulation was restricted to the TDLN. B cell-deficient and immunocompetent mice reconstituted with T2-MZP B cells but not with other B cell subsets displayed accelerated tumor growth, demonstrating that T2-MZP B cells possess regulatory activity in tumor-bearing mice. Unlike splenic regulatory B cells, however, these TDLN B cells did not exhibit increased IL-10 production, nor did they promote Treg generation in the TDLN. These findings demonstrate that tumors initially signal via the lymphatic drainage to stimulate the preferential accumulation of T2-MZP regulatory B cells. This local response may be an early and critical step in generating an immunosuppressive environment to permit tumor growth and metastasis.

  15. Constitutive Smad linker phosphorylation in melanoma: a mechanism of resistance to transforming growth factor-β-mediated growth inhibition.

    PubMed

    Cohen-Solal, Karine A; Merrigan, Kim T; Chan, Joseph L-K; Goydos, James S; Chen, Wenjin; Foran, David J; Liu, Fang; Lasfar, Ahmed; Reiss, Michael

    2011-06-01

    Melanoma cells are resistant to transforming growth factor-β (TGFβ)-induced cell-cycle arrest. In this study, we investigated a mechanism of resistance involving a regulatory domain, called linker region, in Smad2 and Smad3, main downstream effectors of TGFβ. Melanoma cells in culture and tumor samples exhibited constitutive Smad2 and Smad3 linker phosphorylation. Treatment of melanoma cells with the MEK1/2 inhibitor, U0126, or the two pan-CDK and GSK3 inhibitors, Flavopiridol and R547, resulted in decreased linker phosphorylation of Smad2 and Smad3. Overexpression of the linker phosphorylation-resistant Smad3 EPSM mutant in melanoma cells resulted in an increase in expression of p15(INK4B) and p21(WAF1) , as compared with cells transfected with wild-type (WT) Smad3. In addition, the cell numbers of EPSM Smad3-expressing melanoma cells were significantly reduced compared with WT Smad3-expressing cells. These results suggest that the linker phosphorylation of Smad3 contributes to the resistance of melanoma cells to TGFβ-mediated growth inhibition.

  16. Genome-wide analysis of the regulatory function mediated by the small regulatory psm-mec RNA of methicillin-resistant Staphylococcus aureus.

    PubMed

    Cheung, Gordon Y C; Villaruz, Amer E; Joo, Hwang-Soo; Duong, Anthony C; Yeh, Anthony J; Nguyen, Thuan H; Sturdevant, Daniel E; Queck, S Y; Otto, M

    2014-07-01

    Several methicillin resistance (SCCmec) clusters characteristic of hospital-associated methicillin-resistant Staphylococcus aureus (MRSA) strains harbor the psm-mec locus. In addition to encoding the cytolysin, phenol-soluble modulin (PSM)-mec, this locus has been attributed gene regulatory functions. Here we employed genome-wide transcriptional profiling to define the regulatory function of the psm-mec locus. The immune evasion factor protein A emerged as the primary conserved and strongly regulated target of psm-mec, an effect we show is mediated by the psm-mec RNA. Furthermore, the psm-mec locus exerted regulatory effects that were more moderate in extent. For example, expression of PSM-mec limited expression of mecA, thereby decreasing methicillin resistance. Our study shows that the psm-mec locus has a rare dual regulatory RNA and encoded cytolysin function. Furthermore, our findings reveal a specific mechanism underscoring the recently emerging concept that S. aureus strains balance pronounced virulence and high expression of antibiotic resistance.

  17. Regulatory Factor X (RFX)-mediated transcriptional rewiring of ciliary genes in animals.

    PubMed

    Piasecki, Brian P; Burghoorn, Jan; Swoboda, Peter

    2010-07-20

    Cilia were present in the last eukaryotic common ancestor (LECA) and were retained by most organisms spanning all extant eukaryotic lineages, including organisms in the Unikonta (Amoebozoa, fungi, choanoflagellates, and animals), Archaeplastida, Excavata, Chromalveolata, and Rhizaria. In certain animals, including humans, ciliary gene regulation is mediated by Regulatory Factor X (RFX) transcription factors (TFs). RFX TFs bind X-box promoter motifs and thereby positively regulate >50 ciliary genes. Though RFX-mediated ciliary gene regulation has been studied in several bilaterian animals, little is known about the evolutionary conservation of ciliary gene regulation. Here, we explore the evolutionary relationships between RFX TFs and cilia. By sampling the genome sequences of >120 eukaryotic organisms, we show that RFX TFs are exclusively found in unikont organisms (whether ciliated or not), but are completely absent from the genome sequences of all nonunikont organisms (again, whether ciliated or not). Sampling the promoter sequences of 12 highly conserved ciliary genes from 23 diverse unikont and nonunikont organisms further revealed that phylogenetic footprints of X-box promoter motif sequences are found exclusively in ciliary genes of certain animals. Thus, there is no correlation between cilia/ciliary genes and the presence or absence of RFX TFs and X-box promoter motifs in nonanimal unikont and in nonunikont organisms. These data suggest that RFX TFs originated early in the unikont lineage, distinctly after cilia evolved. The evolutionary model that best explains these observations indicates that the transcriptional rewiring of many ciliary genes by RFX TFs occurred early in the animal lineage.

  18. A simple model for dislocation emission mediated dynamic nanovoid growth

    NASA Astrophysics Data System (ADS)

    Wilkerson, Justin; Ramesh, K. T.

    2015-06-01

    Failure of ductile metals has long been attributed to the microscopic processes of void nucleation, growth, and finally coalescence leading to fracture. Our traditional view of void nucleation is associated with interface debonding at second-phase particles. However, much of this understanding has been gleaned from observations of quasi-static fracture surfaces. Under more extreme dynamic loading conditions second-phase particles may not necessarily be the dominant source of void nucleating material defects, and a few key experimental observations of laser spall surfaces seem to support this assertion. Here, we motivate an alternative mechanism to the traditional view, namely shock-induced vacancy generation and clustering followed by nanovoid growth mediated by dislocation emission. This mechanism only becomes active at very large stresses, and thus it is desirable to establish a closed-form criterion for the macroscopic stress required to activate dislocation emission in porous materials. Following an approach similar to Lubarda and co-workers, we make use of stability arguments applied to the analytic solutions of the elastic interactions of dislocations and voids to derive the desired criterion. We then propose a dynamic nanovoid growth law that is motivated by the kinetics of dislocation emission. The resulting failure model is validated against a number of molecular dynamics simulations with favorable agreement. Lastly, we make use of our simple model to predict some interesting anomalous behaviors associated with high surface energies and nonlinear elasticity.

  19. Histamine suppresses regulatory T cells mediated by TGF-β in murine chronic allergic contact dermatitis.

    PubMed

    Tamaka, Kyoko; Seike, Masahiro; Hagiwara, Tamio; Sato, Atsushi; Ohtsu, Hiroshi

    2015-04-01

    Regulatory T cells (Tregs) suppress effector T cells and ameliorate contact hypersensitivity (CH); however, the role of Tregs in chronic allergic contact dermatitis (CACD) has not been assessed. Repeated elicitation of CH has been used to produce CACD models in mice. We previously showed that the presence of histamine facilitates the creation of eczematous lesions in this model using histidine decarboxylase (HDC) (-/-) mice. Therefore, the effects of histamine on Tregs in the CACD model were investigated in this study. CACD was developed by repeated epicutaneous application of 2, 4, 6-trinitro-1-chlorobenzene (TNCB) on HDC (+/+) and HDC (-/-) murine skin to assess the effects of histamine in CACD. Histamine aggravated CACD in the murine model and suppressed the number of Tregs in the skin. Histamine also suppressed the level of TGF-β1 in this model. Recombinant TGF-β1 or anti-TGF-β1 antibody was injected into the dorsal dermis of HDC (+/+) mice daily just before TNCB challenge to determine the effects of histamine-regulated TGF-β on the Treg population in CACD. Recombinant TGF-β1 injection promoted the infiltration of Tregs in the skin and the production of IL-10; however, anti-TGF-β1 antibody injection suppressed the number of Tregs in the skin and the production of IL-10. Histamine suppresses the number of Tregs in CACD, and this effect is mediated by TGF-β.

  20. Regulatory T cells participate in CD39-mediated protection from renal injury.

    PubMed

    Wang, Yuan Min; McRae, Jennifer L; Robson, Simon C; Cowan, Peter J; Zhang, Geoff Yu; Hu, Min; Polhill, Tania; Wang, Yiping; Zheng, Guoping; Wang, Ya; Lee, Vincent W S; Unwin, Robert J; Harris, David C H; Dwyer, Karen M; Alexander, Stephen I

    2012-09-01

    CD39 is an ecto-enzyme that degrades extracellular nucleotides, such as ATP, and is highly expressed on by the vasculature and circulating cells including Foxp3+ regulatory T (Treg) cells. To study the role of purinergic regulation in renal disease, we used the adriamycin nephropathy (AN) mouse model of chronic renal injury, using human CD39-transgenic (hCD39Tg) and wild-type (WT) BALB/c mice. Effects of CD39 expression by Treg cells were assessed in AN by adoptive transfer of CD4(+) CD25(+) and CD4(+) CD25(-) T cells isolated from hCD39Tg and WT mice. hCD39Tg mice were protected from renal injury in AN with decreased urinary protein and serum creatinine, and significantly less renal injury compared with WT mice. While WT CD25(+) and hCD39Tg CD25(-) T cells conferred some protection against AN, hCD39Tg CD25(+) Treg cells offered greater protection. In vitro studies showed direct pro-apoptotic effects of ATP on renal tubular cells. In conclusion, hCD39 expressed by circulating leukocytes and intrinsic renal cells limits innate AN injury. Specifically, CD39 expression by Treg cells contributes to its protective role in renal injury. These findings suggest that extracellular nucleotides mediate AN kidney injury and that CD39, expressed by Treg cells and other cells, is protective in this model.

  1. Polymer-mediated growth of crystals and mesocrystals.

    PubMed

    Cölfen, Helmut

    2013-01-01

    Polymers are important additives for the control of mineralization reactions in both biological and bioinspired mineralization. The reason is that they allow for a number of interactions with the growing crystals and even amorphous minerals. These can substantially influence the way the mineral grows on several levels. Already in the prenucleation phase, polymers can control the formation of prenucleation clusters and subsequently the nucleation event. Also, polymers can control whether the further crystallization follows a classical or nonclassical particle-mediated growth path. In this chapter, the main ways in which polymers can be used to control a crystallization reaction will be highlighted. In addition, polymers that are useful for this purpose and the experimental conditions suitable for directing a crystallization reaction into the desired direction through the use of polymers will be described.

  2. Study of surfactant mediated growth of Ni/V superlattices

    SciTech Connect

    Amir, S. M.; Gupta, Mukul; Potdar, Satish; Gupta, Ajay; Stahn, Jochen

    2013-07-14

    The Ni/V multilayers are useful as soft x-ray mirrors, polarizers, and phase retarders. For these applications, it is necessary that the interfaces roughness and interdiffusion must be as small as possible. The V-on-Ni and Ni-on-V interfaces are asymmetric due to the difference in the surface free energy of Ni and V. In this work, we report Ag surfactant mediated growth of Ni/V superlattices prepared using ion beam sputter deposition technique. These superlattices were studied using x-ray and neutron scattering techniques. It was found that when added in an optimum amount, Ag surfactant results in reduced interface roughness and interdiffusion across the interfaces. Obtained results can be understood with the surfactant floating-off mechanism leading to a balance in the surface free energy of Ni and V.

  3. Sphingosine kinase-1 mediates androgen-induced osteoblast cell growth

    SciTech Connect

    Martin, Claire; Lafosse, Jean-Michel; Malavaud, Bernard; Cuvillier, Olivier

    2010-01-01

    Herein we report that the lipid kinase sphingosine kinase-1 (SphK1) is instrumental in mediating androgen-induced cell proliferation in osteoblasts. Dihydrotestosterone (DHT) triggered cell growth in steroid-deprived MC3T3 cells, which was associated with a rapid stimulation of SphK1 and activation of both Akt and ERK signaling pathways. This mechanism relied on functional androgen receptor/PI3K/Akt nongenotropic signaling as pharmacological antagonists could block SphK1 stimulation by DHT and its consequences. Finally, SphK1 inhibition not only abrogated DHT-induced ERK activation but also blocked cell proliferation, while ERK inhibition had no impact, suggesting that SphK1 was critical for DHT signaling yet independently of the ERK.

  4. Impulsivity, perceived self-regulatory success in dieting, and body mass in children and adolescents: A moderated mediation model.

    PubMed

    Meule, Adrian; Hofmann, Johannes; Weghuber, Daniel; Blechert, Jens

    2016-12-01

    Impulsivity has been suggested to contribute to overeating and obesity. However, findings are inconsistent and it appears that only specific facets of impulsivity are related to eating-related variables and to body mass. In the current study, relationships between self-reported impulsivity, perceived self-regulatory success in dieting, and objectively measured body mass were examined in N = 122 children and adolescents. Scores on attentional and motor impulsivity interactively predicted perceived self-regulatory success in dieting, but not body mass: Higher attentional impulsivity was associated with lower perceived self-regulatory success at high levels of motor impulsivity, but not at low levels of motor impulsivity. A moderated mediation model revealed an indirect effect of attentional and motor impulsivity on body mass, which was mediated by perceived self-regulatory success in dieting. Thus, results show that only specific facets of impulsivity are relevant in eating- and weight-regulation and interact with each other in the prediction of these variables. These facets of impulsivity, however, are not directly related to higher body mass, but indirectly via lower success in eating-related self-regulation in children and adolescents.

  5. Axl as a mediator of cellular growth and survival

    PubMed Central

    Axelrod, Haley; Pienta, Kenneth J.

    2014-01-01

    The control of cellular growth and proliferation is key to the maintenance of homeostasis. Survival, proliferation, and arrest are regulated, in part, by Growth Arrest Specific 6 (Gas6) through binding to members of the TAM receptor tyrosine kinase family. Activation of the TAM receptors leads to downstream signaling through common kinases, but the exact mechanism within each cellular context varies and remains to be completely elucidated. Deregulation of the TAM family, due to its central role in mediating cellular proliferation, has been implicated in multiple diseases. Axl was cloned as the first TAM receptor in a search for genes involved in the progression of chronic to acute-phase leukemia, and has since been established as playing a critical role in the progression of cancer. The oncogenic nature of Axl is demonstrated through its activation of signaling pathways involved in proliferation, migration, inhibition of apoptosis, and therapeutic resistance. Despite its recent discovery, significant progress has been made in the development of effective clinical therapeutics targeting Axl. In order to accurately define the role of Axl in normal and diseased processes, it must be analyzed in a cell type-specific context. PMID:25344858

  6. Parthenolide suppresses pancreatic cell growth by autophagy-mediated apoptosis

    PubMed Central

    Liu, Weifeng; Wang, Xinshuai; Sun, Junjun; Yang, Yanhui; Li, Wensheng; Song, Junxin

    2017-01-01

    Pancreatic cancer is an aggressive malignancy and is unresponsive to conventional chemotherapies. Parthenolide, a sesquiterpene lactone isolated from feverfew, has exhibited potent anticancer effects against various cancers. The purpose of this report was to investigate the effect and underlying mechanism of parthenolide in human pancreatic cancer Panc-1 and BxPC3 cells. The results demonstrated that parthenolide suppressed the growth and induced apoptosis of Panc-1 and BxPC3 pancreatic cancer cells with the half maximal inhibitory concentration (IC50) ranging between 7 and 9 μM after 24 h of treatment. Significant autophagy was induced by parthenolide treatment in pancreatic cancer cells. Parthenolide treatment concentration-dependently increased the percentage of autophagic cells and significantly increased the expression levels of p62/SQSTM1, Beclin 1, and LC3II in Panc-1 cells. Punctate LC3II staining confirmed autophagy. Furthermore, inhibiting autophagy by chloroquine, 3-methyladenine, or LC3II siRNA significantly blocked parthenolide-induced apoptosis, suggesting that parthenolide induced apoptosis through autophagy in this study. In conclusion, these studies established that parthenolide inhibits pancreatic cell growth by autophagy-mediated apoptosis. Data of the present study suggest that parthenolide can serve as a potential chemotherapeutic agent for pancreatic cancer. PMID:28176967

  7. Identification of miRNAs and miRNA-mediated regulatory pathways in Carica papaya.

    PubMed

    Liang, Gang; Li, Yang; He, Hua; Wang, Fang; Yu, Diqiu

    2013-10-01

    Plant microRNAs (miRNAs) post-transcriptionally regulate target gene expression to modulate growth and development and biotic and abiotic stress responses. By analyzing small RNA deep sequencing data in combination with the genome sequence, we identified 75 conserved miRNAs and 11 novel miRNAs. Their target genes were also predicted. For most conserved miRNAs, the miRNA-target pairs were conserved across plant species. In addition to these conserved miRNA-target pairs, we also identified some papaya-specific miRNA-target regulatory pathways. Both miR168 and miR530 target the Argonaute 1 gene, indicating a second autoregulatory mechanism for miRNA regulation. A non-conserved miRNA was mapped within an intron of Dicer-like 1 (DCL1), suggesting a conserved homeostatic autoregulatory mechanism for DCL1 expression. A 21-nt miRNA triggers secondary siRNA production from its target genes, nucleotide-binding site leucine-rich repeat protein genes. Certain phased-miRNAs were processed from their conserved miRNA precursors, indicating a putative miRNA evolution mechanism. In addition, we identified a Carica papaya-specific miRNA that targets an ethylene receptor gene, implying its function in the ethylene signaling pathway. This work will also advance our understanding of miRNA functions and evolution in plants.

  8. Syntrophic growth via quinone-mediated interspecies electron transfer

    PubMed Central

    Smith, Jessica A.; Nevin, Kelly P.; Lovley, Derek R.

    2015-01-01

    The mechanisms by which microbial species exchange electrons are of interest because interspecies electron transfer can expand the metabolic capabilities of microbial communities. Previous studies with the humic substance analog anthraquinone-2,6-disulfonate (AQDS) suggested that quinone-mediated interspecies electron transfer (QUIET) is feasible, but it was not determined if sufficient energy is available from QUIET to support the growth of both species. Furthermore, there have been no previous studies on the mechanisms for the oxidation of anthrahydroquinone-2,6-disulfonate (AHQDS). A co-culture of Geobacter metallireducens and G. sulfurreducens metabolized ethanol with the reduction of fumarate much faster in the presence of AQDS, and there was an increase in cell protein. G. sulfurreducens was more abundant, consistent with G. sulfurreducens obtaining electrons from acetate that G. metallireducens produced from ethanol, as well as from AHQDS. Co-cultures initiated with a citrate synthase-deficient strain of G. sulfurreducens that was unable to use acetate as an electron donor also metabolized ethanol with the reduction of fumarate and cell growth, but acetate accumulated over time. G. sulfurreducens and G. metallireducens were equally abundant in these co-cultures reflecting the inability of the citrate synthase-deficient strain of G. sulfurreducens to metabolize acetate. Evaluation of the mechanisms by which G. sulfurreducens accepts electrons from AHQDS demonstrated that a strain deficient in outer-surface c-type cytochromes that are required for AQDS reduction was as effective at QUIET as the wild-type strain. Deletion of additional genes previously implicated in extracellular electron transfer also had no impact on QUIET. These results demonstrate that QUIET can yield sufficient energy to support the growth of both syntrophic partners, but that the mechanisms by which electrons are derived from extracellular hydroquinones require further investigation. PMID

  9. Glucitol induction in Bacillus subtilis is mediated by a regulatory factor, GutR.

    PubMed Central

    Ye, R; Rehemtulla, S N; Wong, S L

    1994-01-01

    Expression of the glucitol dehydrogenase gene (gutB) is suggested to be regulated both positively and negatively in Bacillus subtilis. A mutation in the gutR locus results in the constitutive expression of gutB. The exact nature of this mutation and the function of gutR are still unknown. Cloning and characterization of gutR indicated that this gene is located immediately upstream of gutB and is transcribed in the opposite direction relative to gutB. GutR is suggested to be a 95-kDa protein with a putative helix-turn-helix motif and a nucleotide binding domain at the N-terminal region. At the C-terminal region, a short sequence of GutR shows homology with two proteins, Cyc8 (glucose repression mediator protein) and GsiA (glucose starvation-inducible protein), known to be directly or indirectly involved in catabolite repression. Part of the C-terminal conserved sequence from these proteins shows all the features observed in the tetratricopeptide motif found in many eucaryotic proteins. To study the functional role of gutR, chromosomal gutR was insertionally inactivated. A total loss of glucitol inducibility was observed. Reintroduction of a functional gutR to the GutR-deficient strain through integration at the amyE locus restores the inducibility. Therefore, GutR serves as a regulatory factor to modulate glucitol induction. The nature of the gutR1 mutation was also determined. A single amino acid change (serine-289 to arginine-289) near the putative nucleotide binding motif B in GutR is responsible for the observed phenotype. Possible models for the action of GutR are discussed. Images PMID:8195087

  10. A regulatory network of two galectins mediates the earliest steps of avian limb skeletal morphogenesis

    PubMed Central

    2011-01-01

    Background The skeletal elements of vertebrate embryonic limbs are prefigured by rod- and spot-like condensations of precartilage mesenchymal cells. The formation of these condensations depends on cell-matrix and cell-cell interactions, but how they are initiated and patterned is as yet unresolved. Results Here we provide evidence that galectins, β-galactoside-binding lectins with β-sandwich folding, play fundamental roles in these processes. We show that among the five chicken galectin (CG) genes, two, CG-1A, and CG-8, are markedly elevated in expression at prospective sites of condensation in vitro and in vivo, with their protein products appearing earlier in development than any previously described marker. The two molecules enhance one another's gene expression but have opposite effects on condensation formation and cartilage development in vivo and in vitro: CG-1A, a non-covalent homodimer, promotes this process, while the tandem-repeat-type CG-8 antagonizes it. Correspondingly, knockdown of CG-1A inhibits the formation of skeletal elements while knockdown of CG-8 enhances it. The apparent paradox of mutual activation at the gene expression level coupled with antagonistic roles in skeletogenesis is resolved by analysis of the direct effect of the proteins on precartilage cells. Specifically, CG-1A causes their aggregation, whereas CG-8, which has no adhesive function of its own, blocks this effect. The developmental appearance and regulation of the unknown cell surface moieties ("ligands") to which CG-1A and CG-8 bind were indicative of specific cognate- and cross-regulatory interactions. Conclusion Our findings indicate that CG-1A and CG-8 constitute a multiscale network that is a major mediator, earlier-acting than any previously described, of the formation and patterning of precartilage mesenchymal condensations in the developing limb. This network functions autonomously of limb bud signaling centers or other limb bud positional cues. PMID:21284876

  11. The Transcriptional and Gene Regulatory Network of Lactococcus lactis MG1363 during Growth in Milk

    PubMed Central

    de Jong, Anne; Hansen, Morten E.; Kuipers, Oscar P.; Kilstrup, Mogens; Kok, Jan

    2013-01-01

    In the present study we examine the changes in the expression of genes of Lactococcus lactis subspecies cremoris MG1363 during growth in milk. To reveal which specific classes of genes (pathways, operons, regulons, COGs) are important, we performed a transcriptome time series experiment. Global analysis of gene expression over time showed that L. lactis adapted quickly to the environmental changes. Using upstream sequences of genes with correlated gene expression profiles, we uncovered a substantial number of putative DNA binding motifs that may be relevant for L. lactis fermentative growth in milk. All available novel and literature-derived data were integrated into network reconstruction building blocks, which were used to reconstruct and visualize the L. lactis gene regulatory network. This network enables easy mining in the chrono-transcriptomics data. A freely available website at http://milkts.molgenrug.nl gives full access to all transcriptome data, to the reconstructed network and to the individual network building blocks. PMID:23349698

  12. Mediation of growth factor induced DNA synthesis and calcium mobilization by Gq and Gi2

    PubMed Central

    1993-01-01

    A newly identified subclass of the heterotrimeric GTP binding regulatory protein family, Gq, has been found to be expressed in a diverse range of cell types. We investigated the potential role of this protein in growth factor signal transduction pathways and its potential relationship to the function of other G alpha subclasses. Recent biochemical studies have suggested that Gq regulates the beta 1 isozyme of phospholipase C (PLC beta 1), an effector for some growth factors. By microinjection of inhibitory antibodies specific to distinct G alpha subunits into living cells, we have determined that G alpha q transduces bradykinin- and thrombin-stimulated intracellular calcium transients which are likely to be mediated by PLC beta 1. Moreover, we found that G alpha q function is required for the mitogenic action of both of these growth factors. These results indicate that both thrombin and bradykinin utilize Gq to couple to increases in intracellular calcium, and that Gq is a necessary component of the mitogenic action of these factors. While microinjection of antibodies against G alpha i2 did not abolish calcium transients stimulated by either of these factors, such microinjection prevented DNA synthesis in response to thrombin but not to bradykinin. These data suggest that thrombin- induced mitogenesis requires both Gq and Gi2, whereas bradykinin needs only the former. Thus, different growth factors operating upon the same cell type use overlapping yet distinct sets of G alpha subtypes in mitogenic signal transduction pathways. The direct identification of the coupling of both a pertussis toxin sensitive and insensitive G protein subtype in the mitogenic pathways utilized by thrombin offers an in vivo biochemical clarification of previous results obtained by pharmacologic studies. PMID:8458876

  13. Innovation of a Regulatory Mechanism Modulating Semi-determinate Stem Growth through Artificial Selection in Soybean

    PubMed Central

    Ping, Jieqing; Li, Shuai; Chen, Zhixiang; Ma, Jianxin

    2016-01-01

    It has been demonstrated that Terminal Flowering 1 (TFL1) in Arabidopsis and its functional orthologs in other plants specify indeterminate stem growth through their specific expression that represses floral identity genes in shoot apical meristems (SAMs), and that the loss-of-function mutations at these functional counterparts result in the transition of SAMs from the vegetative to reproductive state that is essential for initiation of terminal flowering and thus formation of determinate stems. However, little is known regarding how semi-determinate stems, which produce terminal racemes similar to those observed in determinate plants, are specified in any flowering plants. Here we show that semi-determinacy in soybean is modulated by transcriptional repression of Dt1, the functional ortholog of TFL1, in SAMs. Such repression is fulfilled by recently enabled spatiotemporal expression of Dt2, an ancestral form of the APETALA1/FRUITFULL orthologs, which encodes a MADS-box factor directly binding to the regulatory sequence of Dt1. In addition, Dt2 triggers co-expression of the putative SUPPRESSOR OF OVEREXPRESSION OF CONSTANS 1 (GmSOC1) in SAMs, where GmSOC1 interacts with Dt2, and also directly binds to the Dt1 regulatory sequence. Heterologous expression of Dt2 and Dt1 in determinate (tfl1) Arabidopsis mutants enables creation of semi-determinacy, but the same forms of the two genes in the tfl1 and soc1 background produce indeterminate stems, suggesting that Dt2 and SOC1 both are essential for transcriptional repression of Dt1. Nevertheless, the expression of Dt2 is unable to repress TFL1 in Arabidopsis, further demonstrating the evolutionary novelty of the regulatory mechanism underlying stem growth in soybean. PMID:26807727

  14. Spatio-temporal sequence of cross-regulatory events in root meristem growth

    PubMed Central

    Scacchi, Emanuele; Salinas, Paula; Gujas, Bojan; Santuari, Luca; Krogan, Naden; Ragni, Laura; Berleth, Thomas; Hardtke, Christian S.

    2010-01-01

    A central question in developmental biology is how multicellular organisms coordinate cell division and differentiation to determine organ size. In Arabidopsis roots, this balance is controlled by cytokinin-induced expression of SHORT HYPOCOTYL 2 (SHY2) in the so-called transition zone of the meristem, where SHY2 negatively regulates auxin response factors (ARFs) by protein–protein interaction. The resulting down-regulation of PIN-FORMED (PIN) auxin efflux carriers is considered the key event in promoting differentiation of meristematic cells. Here we show that this regulation involves additional, intermediary factors and is spatio-temporally constrained. We found that the described cytokinin–auxin crosstalk antagonizes BREVIS RADIX (BRX) activity in the developing protophloem. BRX is an auxin-responsive target of the prototypical ARF MONOPTEROS (MP), a key promoter of vascular development, and transiently enhances PIN3 expression to promote meristem growth in young roots. At later stages, cytokinin induction of SHY2 in the vascular transition zone restricts BRX expression to down-regulate PIN3 and thus limit meristem growth. Interestingly, proper SHY2 expression requires BRX, which could reflect feedback on the auxin responsiveness of SHY2 because BRX protein can directly interact with MP, likely acting as a cofactor. Thus, cross-regulatory antagonism between BRX and SHY2 could determine ARF activity in the protophloem. Our data suggest a model in which the regulatory interactions favor BRX expression in the early proximal meristem and SHY2 prevails because of supplementary cytokinin induction in the later distal meristem. The complex equilibrium of this regulatory module might represent a universal switch in the transition toward differentiation in various developmental contexts. PMID:21149702

  15. Shape-dependent control of cell growth, differentiation, and apoptosis: switching between attractors in cell regulatory networks

    NASA Technical Reports Server (NTRS)

    Huang, S.; Ingber, D. E.

    2000-01-01

    Development of characteristic tissue patterns requires that individual cells be switched locally between different phenotypes or "fates;" while one cell may proliferate, its neighbors may differentiate or die. Recent studies have revealed that local switching between these different gene programs is controlled through interplay between soluble growth factors, insoluble extracellular matrix molecules, and mechanical forces which produce cell shape distortion. Although the precise molecular basis remains unknown, shape-dependent control of cell growth and function appears to be mediated by tension-dependent changes in the actin cytoskeleton. However, the question remains: how can a generalized physical stimulus, such as cell distortion, activate the same set of genes and signaling proteins that are triggered by molecules which bind to specific cell surface receptors. In this article, we use computer simulations based on dynamic Boolean networks to show that the different cell fates that a particular cell can exhibit may represent a preprogrammed set of common end programs or "attractors" which self-organize within the cell's regulatory networks. In this type of dynamic network model of information processing, generalized stimuli (e.g., mechanical forces) and specific molecular cues elicit signals which follow different trajectories, but eventually converge onto one of a small set of common end programs (growth, quiescence, differentiation, apoptosis, etc.). In other words, if cells use this type of information processing system, then control of cell function would involve selection of preexisting (latent) behavioral modes of the cell, rather than instruction by specific binding molecules. Importantly, the results of the computer simulation closely mimic experimental data obtained with living endothelial cells. The major implication of this finding is that current methods used for analysis of cell function that rely on characterization of linear signaling pathways or

  16. Nucleation and growth mechanisms of ion etching and surfactant-mediated growth

    NASA Astrophysics Data System (ADS)

    Chan, Lana Hiului

    2001-07-01

    Nucleation and growth mechanisms of ion etching of Cu(100) and surfactant (Sb) mediated Ag growth on Ge(100) were studied in ultra-high vacuum (UHV) using scanning tunneling microscopy (STM), low energy electron diffraction (LEED), temperature programmed desorption (TPD), and low energy ion scattering spectroscopy (LEISS). The formation of vacancy clusters from ion etching was characterized and could be modeled and understood as the reverse of film growth. The results suggest that a barrier exists for vacancies to ascend steps. This additional barrier also contributes to pit ordering. The interaction of the Sb with Ge was investigated. Antimony was found to form intermixed surface with Ge. The intermixing, however, could be controlled by deposition and annealing temperatures and Sb coverage. The growth of Ag with and without Sb on Ge(100) was studied and compared. Without Sb, Ag formed elongated 2D islands and 3D clusters. A new metastable Ag-Ge surface alloy phase was found when Ag was deposited at 470 K. This alloy phase can only be obtained by deposition at 470 K. Annealing to 470 K gave a surface with 3D clusters only. When Sb was used as a surfactant for Ag growth, no surface alloy was observed and the cluster shape was isotropic instead of rectangular. On the Sb covered surface, nucleation appeared to be dominated by defects at low temperatures. At higher temperatures, Sb increased the diffusion barrier for Ag. Other effects of using Sb on Ag growth included increase in cluster density and decrease in cluster size. Antimony did not work as a conventional surfactant since it did not segregate to the top of the film at elevated temperatures.

  17. Methionine enkephalin (MENK) inhibits tumor growth through regulating CD4+Foxp3+ regulatory T cells (Tregs) in mice.

    PubMed

    Li, Xuan; Meng, Yiming; Plotnikoff, Nicolas P; Youkilis, Gene; Griffin, Noreen; Wang, Enhua; Lu, Changlong; Shan, Fengping

    2015-01-01

    Methionine enkephalin (MENK), an endogenous neuropeptide, plays an crucial role in both neuroendocrine and immune systems. CD4+Foxp3+ regulatory T cells (Tregs) are identified as a major subpopulation of T lymphocytes in suppressing immune system to keep balanced immunity. The aim of this research work was to elucidate the mechanisms via which MENK interacts with Tregs in cancer situation. The influence of MENK on transforming growth factor-β (TGF-β) mediated conversion from naïve CD4+CD25- T cells to CD4+CD25+ Tregs was determined and the data from flow cytometry (FCM) analysis indicated that MENK effectively inhibited the expression of Foxp3 during the process of TGF-βinduction. Furthermore, this inhibiting process was accompanied by diminishing phosphorylation and nuclear translocation of Smad2/3, confirmed by western blot (WB) analysis and immunofluorescence (IF) at molecular level. We established sarcoma mice model with S180 to investigate whether MENK could modulate Tregs in tumor circumstance. Our findings showed that MENK delayed the development of tumor in S180 tumor bearing mice and down-regulated level of Tregs. Together, these novel findings reached a conclusion that MENK could inhibit Tregs activity directly and retard tumor development through down-regulating Tregs in mice. This work advances the deepening understanding of the influence of MENK on Tregs in cancer situation, and relation of MENK with immune system, supporting the implication of MENK as a new strategy for cancer immunotherapy.

  18. Ligand Receptor-Mediated Regulation of Growth in Plants.

    PubMed

    Haruta, Miyoshi; Sussman, Michael R

    2017-01-01

    Growth and development of multicellular organisms are coordinately regulated by various signaling pathways involving the communication of inter- and intracellular components. To form the appropriate body patterns, cellular growth and development are modulated by either stimulating or inhibiting these pathways. Hormones and second messengers help to mediate the initiation and/or interaction of the various signaling pathways in all complex multicellular eukaryotes. In plants, hormones include small organic molecules, as well as larger peptides and small proteins, which, as in animals, act as ligands and interact with receptor proteins to trigger rapid biochemical changes and induce the intracellular transcriptional and long-term physiological responses. During the past two decades, the availability of genetic and genomic resources in the model plant species, Arabidopsis thaliana, has greatly helped in the discovery of plant hormone receptors and the components of signal transduction pathways and mechanisms used by these immobile but highly complex organisms. Recently, it has been shown that two of the most important plant hormones, auxin and abscisic acid (ABA), act through signaling pathways that have not yet been recognized in animals. For example, auxins stimulate cell elongation by bringing negatively acting transcriptional repressor proteins to the proteasome to be degraded, thus unleashing the gene expression program required for increasing cell size. The "dormancy" inducing hormone, ABA, binds to soluble receptor proteins and inhibits a specific class of protein phosphatases (PP2C), which activates phosphorylation signaling leading to transcriptional changes needed for the desiccation of the seeds prior to entering dormancy. While these two hormone receptors have no known animal counterparts, there are also many similarities between animal and plant signaling pathways. For example, in plants, the largest single gene family in the genome is the protein kinase

  19. Small Regulatory RNA-Induced Growth Rate Heterogeneity of Bacillus subtilis

    PubMed Central

    Mars, Ruben A. T.; Nicolas, Pierre; Ciccolini, Mariano; Reilman, Ewoud; Reder, Alexander; Schaffer, Marc; Mäder, Ulrike; Völker, Uwe; van Dijl, Jan Maarten; Denham, Emma L.

    2015-01-01

    Isogenic bacterial populations can consist of cells displaying heterogeneous physiological traits. Small regulatory RNAs (sRNAs) could affect this heterogeneity since they act by fine-tuning mRNA or protein levels to coordinate the appropriate cellular behavior. Here we show that the sRNA RnaC/S1022 from the Gram-positive bacterium Bacillus subtilis can suppress exponential growth by modulation of the transcriptional regulator AbrB. Specifically, the post-transcriptional abrB-RnaC/S1022 interaction allows B. subtilis to increase the cell-to-cell variation in AbrB protein levels, despite strong negative autoregulation of the abrB promoter. This behavior is consistent with existing mathematical models of sRNA action, thus suggesting that induction of protein expression noise could be a new general aspect of sRNA regulation. Importantly, we show that the sRNA-induced diversity in AbrB levels generates heterogeneity in growth rates during the exponential growth phase. Based on these findings, we hypothesize that the resulting subpopulations of fast- and slow-growing B. subtilis cells reflect a bet-hedging strategy for enhanced survival of unfavorable conditions. PMID:25790031

  20. Small regulatory RNA-induced growth rate heterogeneity of Bacillus subtilis.

    PubMed

    Mars, Ruben A T; Nicolas, Pierre; Ciccolini, Mariano; Reilman, Ewoud; Reder, Alexander; Schaffer, Marc; Mäder, Ulrike; Völker, Uwe; van Dijl, Jan Maarten; Denham, Emma L

    2015-03-01

    Isogenic bacterial populations can consist of cells displaying heterogeneous physiological traits. Small regulatory RNAs (sRNAs) could affect this heterogeneity since they act by fine-tuning mRNA or protein levels to coordinate the appropriate cellular behavior. Here we show that the sRNA RnaC/S1022 from the Gram-positive bacterium Bacillus subtilis can suppress exponential growth by modulation of the transcriptional regulator AbrB. Specifically, the post-transcriptional abrB-RnaC/S1022 interaction allows B. subtilis to increase the cell-to-cell variation in AbrB protein levels, despite strong negative autoregulation of the abrB promoter. This behavior is consistent with existing mathematical models of sRNA action, thus suggesting that induction of protein expression noise could be a new general aspect of sRNA regulation. Importantly, we show that the sRNA-induced diversity in AbrB levels generates heterogeneity in growth rates during the exponential growth phase. Based on these findings, we hypothesize that the resulting subpopulations of fast- and slow-growing B. subtilis cells reflect a bet-hedging strategy for enhanced survival of unfavorable conditions.

  1. Complex quorum-sensing regulatory systems regulate bacterial growth and symbiotic nodulation in Mesorhizobium tianshanense.

    PubMed

    Cao, Huijuan; Yang, Menghua; Zheng, Huiming; Zhang, Jiang; Zhong, Zengtao; Zhu, Jun

    2009-03-01

    LuxR/LuxI-type quorum-sensing systems have been shown to be important for symbiotic interactions between a number of rhizobium species and host legumes. In this study, we found that different isolates of Mesorhizobium tianshanense, a moderately-growing Rhizobium that forms nodules on a number of types of licorice plants, produces several different N-acyl homoserine lactone-like molecules. In M. tianshanense CCBAU060A, we performed a genetic screen and identified a network of regulatory components including a set of LuxI/LuxR-family regulators as well as a MarR-family regulator that is required for quorum-sensing regulation. Furthermore, compared with the wild-type strains, quorum-sensing deficient mutants showed a reduced growth rate and were defective in nodule formation on their host plant Glycyrrhiza uralensis. These data suggest that different M. tianshanense strains may use diverse quorum-sensing systems to regulate symbiotic process.

  2. HuD-mediated distinct BDNF regulatory pathways promote regeneration after nerve injury.

    PubMed

    Sanna, Maria Domenica; Ghelardini, Carla; Galeotti, Nicoletta

    2017-03-15

    Up-regulation of brain-derived neurotrophic factor (BDNF) synthesis is an important mechanism of peripheral nerve regeneration after injury. However, the cellular and molecular mechanisms underlying this process are not fully understood. This study examines the role of BDNF in the spared nerve injury (SNI) mice model. Protein expression and cellular localization were investigated in the dorsal root ganglia (DRG) and spinal cord by western blotting and immunofluorescence experiments respectively. BDNF protein was markedly increased 3 and 7days post-injury in the spinal cord and DRG. Following nerve injury sensory neurons produce molecules to promote regeneration, such as growth-associated protein 43 (GAP-43) and cytoskeletal proteins. Our results show that the expression of GAP-43 was increased in the DRG and spinal cord while, an increased of p-NFH content was detected in the spinal cord, with no modification in the DRG. Both events were counteracted by the administration of an anti-BDNF antibody. In DRG of SNI mice we also detected an increase of HuD expression, a RNA-binding protein known to stabilize BDNF and GAP-43 mRNA. Silencing of HuD prevented the nerve injury-induced BDNF and GAP-43 enhanced expression in the DRG. HuD-mediated BDNF synthesis in the primary sensory neurons, is followed by an anterograde transport of the neurotrophin to the central terminals of the primary afferents in the spinal dorsal horn, to modulate GAP-43 and NFH activation. Our data suggest that BDNF, GAP-43 and p-NFH proteins increase are linked events required for the enhanced regeneration after nerve injury.

  3. Transcriptome-wide analysis of chromium-stress responsive microRNAs to explore miRNA-mediated regulatory networks in radish (Raphanus sativus L.).

    PubMed

    Liu, Wei; Xu, Liang; Wang, Yan; Shen, Hong; Zhu, Xianwen; Zhang, Keyun; Chen, Yinglong; Yu, Rugang; Limera, Cecilia; Liu, Liwang

    2015-09-11

    MicroRNAs (miRNAs) are small noncoding RNAs that play pivotal roles in plant growth, development and stress response. Chromium (Cr) is one of common environmental contaminants possessing potential health hazards to living organisms. To date, little is known about the regulatory roles of miRNAs in response to Cr stress in radish. To systematically identify Cr-responsive miRNAs and their targets in radish, two sRNA libraries derived from Cr-free (CK) and Cr-treated (Cr200) roots were constructed. With Solexa sequencing, 81 known and 72 novel miRNAs were identified, from which 54 known and 16 novel miRNAs were significantly differentially expressed under Cr stress. Several target genes for Cr-responsive miRNAs encode different transcription factor (TF) families, including SPLs, MYBs, ERFs and bZIPs, might regulate corresponding HM-related transcriptional processes in plants. Notably, a few key responsive enzymes or proteins, including HMA, YSL1 and ABC transporter protein were involved in Cr uptake and homeostasis process. Furthermore, the expression patterns of some Cr-responsive miRNAs and their targets were validated by RT-qPCR. This study represents the first characterization of Cr-responsive miRNAs and their targets in radish. The outcomes of this study could provide novel insights into miRNA-mediated regulatory mechanisms underlying plant response to Cr stress in root vegetable crops.

  4. Catecholamine Stress Hormones Regulate Cellular Iron Homeostasis by a Posttranscriptional Mechanism Mediated by Iron Regulatory Protein

    PubMed Central

    Tapryal, Nisha; Vivek G, Vishnu; Mukhopadhyay, Chinmay K.

    2015-01-01

    Adequate availability of iron is important for cellular energy metabolism. Catecholamines such as epinephrine and norepinephrine promote energy expenditure to adapt to conditions that arose due to stress. To restore the energy balance, epinephrine/norepinephrine-exposed cells may face higher iron demand. So far, no direct role of epinephrine/norepinephrine in cellular iron homeostasis has been reported. Here we show that epinephrine/norepinephrine regulates iron homeostasis components such as transferrin receptor-1 and ferritin-H in hepatic and skeletal muscle cells by promoting the binding of iron regulatory proteins to iron-responsive elements present in the UTRs of transferrin receptor-1 and ferritin-H transcripts. Increased transferrin receptor-1, decreased ferritin-H, and increased iron-responsive element-iron regulatory protein interaction are also observed in liver and muscle tissues of epinephrine/norepinephrine-injected mice. We demonstrate the role of epinephrine/norepinephrine-induced generation of reactive oxygen species in converting cytosolic aconitase (ACO1) into iron regulatory protein-1 to bind iron-responsive elements present in UTRs of transferrin receptor-1 and ferritin-H. Our study further reveals that mitochondrial iron content and mitochondrial aconitase (ACO2) activity are elevated by epinephrine/norepinephrine that are blocked by the antioxidant N-acetyl cysteine and iron regulatory protein-1 siRNA, suggesting involvement of reactive oxygen species and iron regulatory protein-1 in this mechanism. This study reveals epinephrine and norepinephrine as novel regulators of cellular iron homeostasis. PMID:25572399

  5. A regulatory governance perspective on health technology assessment (HTA) in France: the contextual mediation of common functional pressures.

    PubMed

    Barron, Anthony J G; Klinger, Corinna; Shah, Sara Mehmood Birchall; Wright, John S F

    2015-02-01

    The new regulatory governance perspective has introduced several insights to the study of health technology assessment (HTA): it has broadened the scope for the analysis of HTA; it has provided a more sophisticated account of national diversity and the potential for cross-border policy learning; and, it has dissolved the distinction between HTA assessment and appraisal processes. In this paper, we undertake a qualitative study of the French process for HTA with a view to introducing a fourth insight: that the emergence and continuing function of national agencies for HTA follows a broadly evolutionary pattern in which contextual factors play an important mediating role. We demonstrate that the French process for HTA is characterised by distinctive institutions, processes and evidential requirements. Consistent with the mediating role of this divergent policy context, we argue that even initiatives for the harmonisation of national approaches to HTA are likely to meet with divergent national policy responses.

  6. Vasoactive intestinal peptide (VIP)-mediated expression and function of steroidogenic acute regulatory protein (StAR) in granulosa cells.

    PubMed

    Kowalewski, Mariusz P; Dyson, Matthew T; Boos, Alois; Stocco, Douglas M

    2010-10-26

    VIP is a peptide hormone capable of activating the cAMP/PKA pathway and modifying gonadal steroidogenic capacity. Less is known about the molecular mechanisms of VIP-mediated steroidogenesis and its role in regulating the steroidogenic acute regulatory protein (STAR). We examined the impact of VIP on STAR expression and function in immortalized (KK1) and primary mouse granulosa cells, where VIP strongly upregulated STAR expression and steroidogenesis. Inhibitors of the PKA and PKC pathways suggested that both are activated by VIP. VIP did not efficiently phosphorylate STAR (P-STAR); however, VIP together with cAMP-analogs that activate Type II PKA increased P-STAR and further increased steroidogenesis. Our results suggest that VIP-induced STAR expression and function in granulosa cells result from the preferential activation of Type I PKA. Furthermore, the PKA and PKC pathways appear to converge at regulating VIP-mediated Star transcription and translation.

  7. Identification and characterization of a retinoid-induced class II tumor suppressor/growth regulatory gene.

    PubMed

    DiSepio, D; Ghosn, C; Eckert, R L; Deucher, A; Robinson, N; Duvic, M; Chandraratna, R A; Nagpal, S

    1998-12-08

    Retinoids, synthetic and natural analogs of retinoic acid, exhibit potent growth inhibitory and cell differentiation activities that account for their beneficial effects in treating hyperproliferative diseases such as psoriasis, actinic keratosis, and certain neoplasias. Tazarotene is a synthetic retinoid that is used in the clinic for the treatment of psoriasis. To better understand the mechanism of retinoid action in the treatment of hyperproliferative diseases, we used a long-range differential display-PCR to isolate retinoid-responsive genes from primary human keratinocytes. We have identified a cDNA, tazarotene-induced gene 3 (TIG3; Retinoic Acid Receptor Responder 3) showing significant homology to the class II tumor suppressor gene, H-rev 107. Tazarotene treatment increases TIG3 expression in primary human keratinocytes and in vivo in psoriatic lesions. Increased TIG3 expression is correlated with decreased proliferation. TIG3 is expressed in a number of tissues, and expression is reduced in cancer cell lines and some primary tumors. In breast cancer cell lines, retinoid-dependent TIG3 induction is observed in lines that are growth suppressed by retinoids but not in nonresponsive lines. Transient over-expression of TIG3 in T47D or Chinese hamster ovary cells inhibits colony expansion. Finally, studies in 293 cells expressing TIG3 linked to an inducible promoter demonstrated decreased proliferation with increased TIG3 levels. These studies suggest that TIG3 may be a growth regulator that mediates some of the growth suppressive effects of retinoids.

  8. Antioxidant and insect growth regulatory activities of stilbenes and extracts from Yucca periculosa.

    PubMed

    Torres, Patricio; Avila, J Guillermo; Romo de Vivar, Alfonso; García, Ana M; Marín, Juan C; Aranda, Eduardo; Céspedes, Carlos L

    2003-09-01

    The methanol extract from the bark of Yucca periculosa F. Baker afforded 4,4'-dihydroxstilbene, resveratrol and 3,3',5,5'-tetrahydroxy-4-methoxystilbene and had growth regulatory activity against the Fall Army worm (Spodoptera frugiperda J.E. Smith, Lepidoptera:Noctuidae) an insect pest of corn. The most active compound was 3,3',5,5'-tetrahydroxy-4-methoxystilbene which had significant effects at 3 microg/g in diets. In addition to the inhibitory activity on bleaching of crocin induced by alkoxyl radicals, these compounds also demonstrated scavenging properties toward 2,2-diphenyl-1-picrylhydrazyl in TLC autographic and spectrophotometric assays. Our results indicate that these compounds could be involved in interference of sclerotization and moulting. These compounds appear to have selective effects on the pre-emergence metabolism of the insect. The results were fully comparable to known natural insect growth inhibitors such as gedunin and Cedrela extracts and have had a possible role as natural insecticidal agents.

  9. Integrative identification of deregulated miRNA/TF-mediated gene regulatory loops and networks in prostate cancer.

    PubMed

    Afshar, Ali Sobhi; Xu, Joseph; Goutsias, John

    2014-01-01

    MicroRNAs (miRNAs) have attracted a great deal of attention in biology and medicine. It has been hypothesized that miRNAs interact with transcription factors (TFs) in a coordinated fashion to play key roles in regulating signaling and transcriptional pathways and in achieving robust gene regulation. Here, we propose a novel integrative computational method to infer certain types of deregulated miRNA-mediated regulatory circuits at the transcriptional, post-transcriptional and signaling levels. To reliably predict miRNA-target interactions from mRNA/miRNA expression data, our method collectively utilizes sequence-based miRNA-target predictions obtained from several algorithms, known information about mRNA and miRNA targets of TFs available in existing databases, certain molecular structures identified to be statistically over-represented in gene regulatory networks, available molecular subtyping information, and state-of-the-art statistical techniques to appropriately constrain the underlying analysis. In this way, the method exploits almost every aspect of extractable information in the expression data. We apply our procedure on mRNA/miRNA expression data from prostate tumor and normal samples and detect numerous known and novel miRNA-mediated deregulated loops and networks in prostate cancer. We also demonstrate instances of the results in a number of distinct biological settings, which are known to play crucial roles in prostate and other types of cancer. Our findings show that the proposed computational method can be used to effectively achieve notable insights into the poorly understood molecular mechanisms of miRNA-mediated interactions and dissect their functional roles in cancer in an effort to pave the way for miRNA-based therapeutics in clinical settings.

  10. Integrative Identification of Deregulated MiRNA/TF-Mediated Gene Regulatory Loops and Networks in Prostate Cancer

    PubMed Central

    Afshar, Ali Sobhi; Xu, Joseph; Goutsias, John

    2014-01-01

    MicroRNAs (miRNAs) have attracted a great deal of attention in biology and medicine. It has been hypothesized that miRNAs interact with transcription factors (TFs) in a coordinated fashion to play key roles in regulating signaling and transcriptional pathways and in achieving robust gene regulation. Here, we propose a novel integrative computational method to infer certain types of deregulated miRNA-mediated regulatory circuits at the transcriptional, post-transcriptional and signaling levels. To reliably predict miRNA-target interactions from mRNA/miRNA expression data, our method collectively utilizes sequence-based miRNA-target predictions obtained from several algorithms, known information about mRNA and miRNA targets of TFs available in existing databases, certain molecular structures identified to be statistically over-represented in gene regulatory networks, available molecular subtyping information, and state-of-the-art statistical techniques to appropriately constrain the underlying analysis. In this way, the method exploits almost every aspect of extractable information in the expression data. We apply our procedure on mRNA/miRNA expression data from prostate tumor and normal samples and detect numerous known and novel miRNA-mediated deregulated loops and networks in prostate cancer. We also demonstrate instances of the results in a number of distinct biological settings, which are known to play crucial roles in prostate and other types of cancer. Our findings show that the proposed computational method can be used to effectively achieve notable insights into the poorly understood molecular mechanisms of miRNA-mediated interactions and dissect their functional roles in cancer in an effort to pave the way for miRNA-based therapeutics in clinical settings. PMID:24968068

  11. Circumvention of regulatory CD4(+) T cell activity during cross-priming strongly enhances T cell-mediated immunity.

    PubMed

    Heit, Antje; Gebhardt, Friedemann; Lahl, Katharina; Neuenhahn, Michael; Schmitz, Frank; Anderl, Florian; Wagner, Hermann; Sparwasser, Tim; Busch, Dirk H; Kastenmüller, Kathrin

    2008-06-01

    Immunization with purified antigens is a safe and practical vaccination strategy but is generally unable to induce sustained CD8(+) T cell-mediated protection against intracellular pathogens. Most efforts to improve the CD8(+) T cell immunogenicity of these vaccines have focused on co-administration of adjuvant to support cross-presentation and dendritic cell maturation. In addition, it has been shown that CD4(+) T cell help during the priming phase contributes to the generation of protective CD8(+) memory T cells. In this report we demonstrate that the depletion of CD4(+) T cells paradoxically enhances long-lasting CD8-mediated protective immunity upon protein vaccination. Functional and genetic in vivo inactivation experiments attribute this enhancement primarily to MHC class II-restricted CD4(+) regulatory T cells (Treg), which appear to physiologically suppress the differentiation process towards long-living effector memory T cells. Since, in functional terms, this suppression by Treg largely exceeds the positive effects of conventional CD4(+) T cell help, even the absence of all CD4(+) T cells or lack of MHC class II-mediated interactions on priming dendritic cells result in enhanced CD8(+) T cell immunogenicity. These findings have important implications for the improvement of vaccines against intracellular pathogens or tumors, especially in patients with highly active Treg.

  12. Glucagon-like peptide 2 may mediate growth and development of the bovine gastrointestinal tract

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Glucagon-like peptide 2 (GLP-2), secreted by enteroendocrine cells, promotes growth, reduces apoptosis, and enhances blood flow, nutrient absorption, and barrier function in intestinal epithelium of monogastric species. Regulatory functions of GLP-2 in the ruminant gastrointestinal tract (GIT) are u...

  13. Epidermal growth factor receptor protects proliferating cell nuclear antigen from cullin 4A protein-mediated proteolysis.

    PubMed

    Lo, Yuan-Hung; Ho, Po-Chun; Wang, Shao-Chun

    2012-08-03

    Proliferating cell nuclear antigen (PCNA) is an essential component for DNA synthesis upon growth stimulation. It has been shown that phosphorylation of PCNA at Tyr-211 by the EGF receptor (EGFR) protects PCNA from polyubiquitylation and degradation, whereas blocking phosphorylation induces ubiquitylation-mediated degradation of the chromatin-bound, but not the -unbound, PCNA, and suppresses cell proliferation. However, the ubiquitin E3 ligase linking growth signaling to the proteolysis of PCNA and the underlying regulatory mechanism remain to be identified. Here we show that, in the absence of Tyr-211 phosphorylation, PCNA is subject to polyubiquitylation at Lys-164 by the CUL4A E3 ligase, resulting in the degradation of PCNA. Mutation of Lys-164 to arginine prevents PCNA ubiquitylation and rescues the degradation of the K164R/Y211F PCNA double mutant. Activation of EGFR inhibits the interaction of PCNA with CUL4A, whereas inhibition of EGFR leads to increased CUL4A-PCNA interaction and CUL4A-dependent ubiquitin-mediated degradation of PCNA. Substitution of endogenous PCNA with the Y211F mutant PCNA conveys enhanced sensitization to EGFR inhibition. Our findings identify CUL4A as the ubiquitin ligase linking the down-regulation of cell surface receptor tyrosine kinase to the nuclear DNA replication machinery in cancer cells.

  14. USP17 is required for clathrin mediated endocytosis of epidermal growth factor receptor

    PubMed Central

    Jaworski, Jakub; de la Vega, Michelle; Fletcher, Sarah J.; McFarlane, Cheryl; Greene, Michelle K.; Smyth, Andrew W.; Van Schaeybroeck, Sandra; Johnston, James A.; Scott, Christopher J.; Rappoport, Joshua Z.; Burrows, James F.

    2014-01-01

    Previously we have shown that expression of the deubiquitinating enzyme USP17 is required for cell proliferation and motility. More recently we reported that USP17 deubiquitinates RCE1 isoform 2 and thus regulates the processing of ‘CaaX’ motif proteins. Here we now show that USP17 expression is induced by epidermal growth factor and that USP17 expression is required for clathrin mediated endocytosis of epidermal growth factor receptor. In addition, we show that USP17 is required for the endocytosis of transferrin, an archetypal substrate for clathrin mediated endocytosis, and that USP17 depletion impedes plasma membrane recruitment of the machinery required for clathrin mediated endocytosis. Thus, our data reveal that USP17 is necessary for epidermal growth factor receptor and transferrin endocytosis via clathrin coated pits, indicate this is mediated via the regulation of the recruitment of the components of the endocytosis machinery and suggest USP17 may play a general role in receptor endocytosis. PMID:25026282

  15. The transcriptional regulator BZR1 mediates trade-off between plant innate immunity and growth

    PubMed Central

    Lozano-Durán, Rosa; Macho, Alberto P; Boutrot, Freddy; Segonzac, Cécile; Somssich, Imre E; Zipfel, Cyril

    2013-01-01

    The molecular mechanisms underlying the trade-off between plant innate immunity and steroid-mediated growth are controversial. Here, we report that activation of the transcription factor BZR1 is required and sufficient for suppression of immune signaling by brassinosteroids (BR). BZR1 induces the expression of several WRKY transcription factors that negatively control early immune responses. In addition, BZR1 associates with WRKY40 to mediate the antagonism between BR and immune signaling. We reveal that BZR1-mediated inhibition of immunity is particularly relevant when plant fast growth is required, such as during etiolation. Thus, BZR1 acts as an important regulator mediating the trade-off between growth and immunity upon integration of environmental cues. DOI: http://dx.doi.org/10.7554/eLife.00983.001 PMID:24381244

  16. The transcriptional regulator BZR1 mediates trade-off between plant innate immunity and growth.

    PubMed

    Lozano-Durán, Rosa; Macho, Alberto P; Boutrot, Freddy; Segonzac, Cécile; Somssich, Imre E; Zipfel, Cyril

    2013-12-31

    The molecular mechanisms underlying the trade-off between plant innate immunity and steroid-mediated growth are controversial. Here, we report that activation of the transcription factor BZR1 is required and sufficient for suppression of immune signaling by brassinosteroids (BR). BZR1 induces the expression of several WRKY transcription factors that negatively control early immune responses. In addition, BZR1 associates with WRKY40 to mediate the antagonism between BR and immune signaling. We reveal that BZR1-mediated inhibition of immunity is particularly relevant when plant fast growth is required, such as during etiolation. Thus, BZR1 acts as an important regulator mediating the trade-off between growth and immunity upon integration of environmental cues. DOI: http://dx.doi.org/10.7554/eLife.00983.001.

  17. Underpotential deposition-mediated layer-by-layer growth of thin films

    DOEpatents

    Wang, Jia Xu; Adzic, Radoslav R.

    2015-05-19

    A method of depositing contiguous, conformal submonolayer-to-multilayer thin films with atomic-level control is described. The process involves the use of underpotential deposition of a first element to mediate the growth of a second material by overpotential deposition. Deposition occurs between a potential positive to the bulk deposition potential for the mediating element where a full monolayer of mediating element forms, and a potential which is less than, or only slightly greater than, the bulk deposition potential of the material to be deposited. By cycling the applied voltage between the bulk deposition potential for the mediating element and the material to be deposited, repeated desorption/adsorption of the mediating element during each potential cycle can be used to precisely control film growth on a layer-by-layer basis. This process is especially suitable for the formation of a catalytically active layer on core-shell particles for use in energy conversion devices such as fuel cells.

  18. Industry growth, work role characteristics, and job satisfaction: a cross-level mediation model.

    PubMed

    Ford, Michael T; Wooldridge, Jessica D

    2012-10-01

    The associations between industry revenue growth, individual work role characteristics, and job satisfaction were examined in this cross-level mediation analysis. Work roles were expected to be more autonomous, involve greater skill variety, and offer more opportunities for growth and development for workers in growing industries than for workers in declining industries. Supervisor support was also hypothesized to be stronger for workers in high-growth industries. Results from a nationally representative (U.S.) sample of service industry workers, using multilevel modeling, supported these propositions and suggest that job enrichment mediates relations between industry growth and job satisfaction. Associations between industry growth and autonomy were also stronger among workers in occupations that are less normatively autonomous, suggesting that industry growth fosters a weakening, and industry decline a strengthening, of traditional differences in autonomy across work roles. These results contribute to a multilevel perspective on organizational environments, individual work roles, and worker attitudes and well-being.

  19. Regulatory T cells attenuate Th17 cell-mediated nigrostriatal dopaminergic neurodegeneration in a model of Parkinson's disease

    PubMed Central

    Reynolds, Ashley D.; Stone, David K.; Hutter, Jessica A.L.; Benner, Eric J.; Mosley, R. Lee; Gendelman, Howard E.

    2010-01-01

    Nitrated alpha synuclein (N-α-syn) immunization elicits adaptive immune responses to novel antigenic epitopes that exacerbate neuroinflammation and nigrostriatal degeneration in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) model of Parkinson's disease (PD). We show that such neuroimmune degenerative activities, in significant measure, are Th17 cell-mediated with CD4+CD25+ regulatory T cell (Treg) dysfunction seen amongst populations of N-α-syn induced T cells. In contrast, purified vasoactive intestinal peptide (VIP)-induced and natural Treg reversed N-α-syn T cell nigrostriatal degeneration. Combinations of adoptively transferred N-α-syn and VIP immunocytes or natural Treg administered to MPTP mice attenuated microglial inflammatory responses and led to robust nigrostriatal protection. Taken together, these results demonstrate a putative mechanism for Treg control of N-α-syn-induced neurodestructive immunity and as such provide a sound rationale for future PD immunization strategies. PMID:20118279

  20. The simple neuroendocrine-immune regulatory network in oyster Crassostrea gigas mediates complex functions

    PubMed Central

    Liu, Zhaoqun; Wang, Lingling; Zhou, Zhi; Sun, Ying; Wang, Mengqiang; Wang, Hao; Hou, Zhanhui; Gao, Dahai; Gao, Qiang; Song, Linsheng

    2016-01-01

    The neuroendocrine-immune (NEI) regulatory network is a complex system, which plays an indispensable role in the immunity of the host. In the present study, the bioinformatical analysis of the transcriptomic data from oyster Crassostrea gigas and further biological validation revealed that oyster TNF (CgTNF-1 CGI_10018786) could activate the transcription factors NF-κB and HSF (heat shock transcription factor) through MAPK signaling pathway, and then regulate apoptosis, redox reaction, neuro-regulation and protein folding in oyster haemocytes. The activated immune cells then released neurotransmitters including acetylcholine, norepinephrine and [Met5]-enkephalin to regulate the immune response by arising the expression of three TNF (CGI_10005109, CGI_10005110 and CGI_10006440) and translocating two NF-κB (Cgp65, CGI_10018142 and CgRel, CGI_10021567) between the cytoplasm and nuclei of haemocytes. Neurotransmitters exhibited the immunomodulation effects by influencing apoptosis and phagocytosis of oyster haemocytes. Acetylcholine and norepinephrine could down-regulate the immune response, while [Met5]-enkephalin up-regulate the immune response. These results suggested that the simple neuroendocrine-immune regulatory network in oyster might be activated by oyster TNF and then regulate the immune response by virtue of neurotransmitters, cytokines and transcription factors. PMID:27193598

  1. The simple neuroendocrine-immune regulatory network in oyster Crassostrea gigas mediates complex functions

    NASA Astrophysics Data System (ADS)

    Liu, Zhaoqun; Wang, Lingling; Zhou, Zhi; Sun, Ying; Wang, Mengqiang; Wang, Hao; Hou, Zhanhui; Gao, Dahai; Gao, Qiang; Song, Linsheng

    2016-05-01

    The neuroendocrine-immune (NEI) regulatory network is a complex system, which plays an indispensable role in the immunity of the host. In the present study, the bioinformatical analysis of the transcriptomic data from oyster Crassostrea gigas and further biological validation revealed that oyster TNF (CgTNF-1 CGI_10018786) could activate the transcription factors NF-κB and HSF (heat shock transcription factor) through MAPK signaling pathway, and then regulate apoptosis, redox reaction, neuro-regulation and protein folding in oyster haemocytes. The activated immune cells then released neurotransmitters including acetylcholine, norepinephrine and [Met5]-enkephalin to regulate the immune response by arising the expression of three TNF (CGI_10005109, CGI_10005110 and CGI_10006440) and translocating two NF-κB (Cgp65, CGI_10018142 and CgRel, CGI_10021567) between the cytoplasm and nuclei of haemocytes. Neurotransmitters exhibited the immunomodulation effects by influencing apoptosis and phagocytosis of oyster haemocytes. Acetylcholine and norepinephrine could down-regulate the immune response, while [Met5]-enkephalin up-regulate the immune response. These results suggested that the simple neuroendocrine-immune regulatory network in oyster might be activated by oyster TNF and then regulate the immune response by virtue of neurotransmitters, cytokines and transcription factors.

  2. Regulatory Mode and Risk-Taking: The Mediating Role of Anticipated Regret

    PubMed Central

    Panno, Angelo; Lauriola, Marco; Pierro, Antonio

    2015-01-01

    We propose that decision maker’s regulatory mode affects risk-taking through anticipated regret. In the Study 1 either a locomotion or an assessment orientation were experimentally induced, and in the Studies 2 and 3 these different orientations were assessed as chronic individual differences. To assess risk-taking we used two behavioral measures of risk: BART and hot-CCT. The results show that experimentally induced assessment orientation–compared to locomotion–leads to decreased risk-taking through increased anticipated regret (Study 1). People chronically predisposed to be in the assessment state take less risk through increased anticipated regret (Study 2 and Study 3). Study 2 results also show a marginally non-significant indirect effect of chronic locomotion mode on BART through anticipated regret. Differently, Study 3 shows that people chronically predisposed to be in the locomotion state take greater risk through decreased anticipated regret, when play a dynamic risk task triggering stronger emotional arousal. Through all three studies, the average effect size for the relationship of assessment with anticipated regret was in the moderate-large range, whereas for risk-taking was in the moderate range. The average effect size for the relationship of locomotion with anticipated regret was in the moderate range, whereas for risk-taking was in the small-moderate range. These results increase our understanding of human behavior under conditions of risk obtaining novel insights into regulatory mode theory and decision science. PMID:26580960

  3. The simple neuroendocrine-immune regulatory network in oyster Crassostrea gigas mediates complex functions.

    PubMed

    Liu, Zhaoqun; Wang, Lingling; Zhou, Zhi; Sun, Ying; Wang, Mengqiang; Wang, Hao; Hou, Zhanhui; Gao, Dahai; Gao, Qiang; Song, Linsheng

    2016-05-19

    The neuroendocrine-immune (NEI) regulatory network is a complex system, which plays an indispensable role in the immunity of the host. In the present study, the bioinformatical analysis of the transcriptomic data from oyster Crassostrea gigas and further biological validation revealed that oyster TNF (CgTNF-1 CGI_10018786) could activate the transcription factors NF-κB and HSF (heat shock transcription factor) through MAPK signaling pathway, and then regulate apoptosis, redox reaction, neuro-regulation and protein folding in oyster haemocytes. The activated immune cells then released neurotransmitters including acetylcholine, norepinephrine and [Met(5)]-enkephalin to regulate the immune response by arising the expression of three TNF (CGI_10005109, CGI_10005110 and CGI_10006440) and translocating two NF-κB (Cgp65, CGI_10018142 and CgRel, CGI_10021567) between the cytoplasm and nuclei of haemocytes. Neurotransmitters exhibited the immunomodulation effects by influencing apoptosis and phagocytosis of oyster haemocytes. Acetylcholine and norepinephrine could down-regulate the immune response, while [Met(5)]-enkephalin up-regulate the immune response. These results suggested that the simple neuroendocrine-immune regulatory network in oyster might be activated by oyster TNF and then regulate the immune response by virtue of neurotransmitters, cytokines and transcription factors.

  4. Lactose inhibits regulatory T-cell-mediated suppression of effector T-cell interferon-γ and IL-17 production.

    PubMed

    Paasela, Monika; Kolho, Kaija-Leena; Vaarala, Outi; Honkanen, Jarno

    2014-12-14

    Our interest in lactose as an immunomodulatory molecule results from studies showing that lactose binds to galectin-9, which has been shown to have various regulatory functions in the immune system including regulation of T-cell responses. Impaired regulation of T helper (Th)1 and Th17 type immune responses and dysfunction of regulatory T cells (Treg) have been implicated in many human immune-mediated diseases. In the present study, we investigated the effects of lactose on immune regulation using co-cultures of human peripheral blood mononuclear cell (PBMC)-derived Treg and effector T cells (Teff) obtained from twenty healthy adults. Treg, i.e. CD4+CD25+CD127-, were isolated from PBMC by immunomagnetic separation. The fraction of CD4+CD127- cells that was depleted of CD25+ cells was used as Teff. Treg and Teff at a ratio 1:5 were activated and the effects of lactose on the secretion of interferon-γ (IFN-γ) and IL-17 were analysed using ELISA for protein and quantitative RT-PCR for mRNA. Treg down-regulated the secretion of both IFN-γ (8.8-3.9 ng/ml, n 20, P= 0.003) and IL-17 (0.83-0.64 ng/ml, n 15, P= 0.04) in co-cultures, while in the presence of lactose the levels of secreted IFN-γ and IL-17 remained high and no down-regulation was observed (16.4 v. 3.99 ng/ml, n 20, P< 0.0001, and 0.74 v. 0.64 ng/ml, n 15, P= 0.005, respectively). We showed that lactose inhibits human Treg-mediated suppression of Th1 and Th17 immune responses in vitro.

  5. PD-L1 is a critical mediator of regulatory B cells and T cells in invasive breast cancer

    PubMed Central

    Guan, Honggeng; Wan, Yuqiu; Lan, Jing; Wang, Qin; Wang, Zhangyu; Li, Yecheng; Zheng, Jiqing; Zhang, Xueguang; Wang, Zemin; Shen, Yueping; Xie, Fang

    2016-01-01

    Regulatory T cells (Tregs), a key mediator in regulating anti-tumor immune suppression, tumor immune escape, metastasis and relapse, are considered an important therapeutic target in immunotherapy of human cancers. In the present investigation, elevated CD19+ CD24+ CD38+ regulatory B cells (Bregs) were observed in PBMCs of invasive carcinoma of breast (IBCa) patients compared with that in patients with fibroadenoma (FIBma) or healthy individuals, and the positive correlation existed between Bregs and CD4+ CD25+ CD127− Tregs (r = 0.316, P = 0.001). We found that PD-L1 expression was higher on Bregs in IBCa patients compared with patients with FIBma or healthy individuals (P < 0.05, respectively), and that a tight correlation exists between CD19+ CD24+ CD38+ PD-L1+ Bregs and CD19+ CD24+ CD38+ Bregs (r = 0.267, P = 0.007), poor TNM phases and up-regulated expression of PD-L1 on Bregs. The pattern of PD-1 expression on CD4+ T cells indicated that high level of PD-1hi expressed on CD4+ CD25+ CD127+ effector T cells (P < 0.001). More importantly, the presence of PD-L1 on Bregs was positively correlated with Tregs (r = 0.299, P = 0.003), but negatively correlated with PD-1hi effector T cells (r = −0.22, P = 0.031). Together, results of the present study indicated that PD-L1 is an important molecule on Bregs, mediated the generation of Tregs in IBCa. PMID:27762298

  6. The Yin and Yang of the Opioid Growth Regulatory System: Focus on Diabetes—The Lorenz E. Zimmerman Tribute Lecture

    PubMed Central

    Mc Laughlin, Patricia J.; Zagon, Ian S.

    2016-01-01

    The Opioid Growth Regulatory System consists of opioid growth factor (OGF), [Met5]-enkephalin, and its unique receptor (OGFr). OGF inhibits cell division when bound to OGFr. Conversely, blockade of the interaction of OGF and OGFr, using the potent, long-acting opioid receptor antagonist, naltrexone (NTX), results in increased DNA synthesis and cell division. The authors have demonstrated both in vitro and in vivo that the addition of exogenous OGF or an increase in available OGFr decreases corneal epithelial cell division and wound healing. Conversely, blockade of the OGF-OGFr interaction by NTX or a decrease in the production of the OGFr increases corneal epithelial cell division and facilitates corneal epithelial wound healing. The authors also have demonstrated that depressed corneal and cutaneous wound healing, dry eye, and abnormal corneal sensitivity in type 1 and type 2 diabetes in animals can be reversed by OGF-OGFr blockade by NTX. Thus, the function of the Opioid Growth Regulatory System appears to be disordered in diabetic animals, and its function can be restored with NTX treatment. These studies suggest a fundamental role for the Opioid Growth Regulatory System in the pathobiology of diabetic complications and a need for studies to elucidate this role further. PMID:27703986

  7. IL-21 restricts T follicular regulatory T cell proliferation through Bcl-6 mediated inhibition of responsiveness to IL-2.

    PubMed

    Jandl, Christoph; Liu, Sue M; Cañete, Pablo F; Warren, Joanna; Hughes, William E; Vogelzang, Alexis; Webster, Kylie; Craig, Maria E; Uzel, Gulbu; Dent, Alexander; Stepensky, Polina; Keller, Bärbel; Warnatz, Klaus; Sprent, Jonathan; King, Cecile

    2017-03-17

    T follicular regulatory (Tfr) cells control the magnitude and specificity of the germinal centre reaction, but how regulation is contained to ensure generation of high-affinity antibody is unknown. Here we show that this balance is maintained by the reciprocal influence of interleukin (IL)-2 and IL-21. The number of IL-2-dependent FoxP3(+) regulatory T cells is increased in the peripheral blood of human patients with loss-of-function mutations in the IL-21 receptor (IL-21R). In mice, IL-21:IL-21R interactions influence the phenotype of T follicular cells, reducing the expression of CXCR4 and inhibiting the expansion of Tfr cells after T-cell-dependent immunization. The negative effect of IL-21 on Tfr cells in mice is cell intrinsic and associated with decreased expression of the high affinity IL-2 receptor (CD25). Bcl-6, expressed in abundance in Tfr cells, inhibits CD25 expression and IL-21-mediated inhibition of CD25 is Bcl-6 dependent. These findings identify a mechanism by which IL-21 reinforces humoral immunity by restricting Tfr cell proliferation.

  8. IL-21 restricts T follicular regulatory T cell proliferation through Bcl-6 mediated inhibition of responsiveness to IL-2

    PubMed Central

    Jandl, Christoph; Liu, Sue M.; Cañete, Pablo F.; Warren, Joanna; Hughes, William E.; Vogelzang, Alexis; Webster, Kylie; Craig, Maria E.; Uzel, Gulbu; Dent, Alexander; Stepensky, Polina; Keller, Bärbel; Warnatz, Klaus; Sprent, Jonathan; King, Cecile

    2017-01-01

    T follicular regulatory (Tfr) cells control the magnitude and specificity of the germinal centre reaction, but how regulation is contained to ensure generation of high-affinity antibody is unknown. Here we show that this balance is maintained by the reciprocal influence of interleukin (IL)-2 and IL-21. The number of IL-2-dependent FoxP3+ regulatory T cells is increased in the peripheral blood of human patients with loss-of-function mutations in the IL-21 receptor (IL-21R). In mice, IL-21:IL-21R interactions influence the phenotype of T follicular cells, reducing the expression of CXCR4 and inhibiting the expansion of Tfr cells after T-cell-dependent immunization. The negative effect of IL-21 on Tfr cells in mice is cell intrinsic and associated with decreased expression of the high affinity IL-2 receptor (CD25). Bcl-6, expressed in abundance in Tfr cells, inhibits CD25 expression and IL-21-mediated inhibition of CD25 is Bcl-6 dependent. These findings identify a mechanism by which IL-21 reinforces humoral immunity by restricting Tfr cell proliferation. PMID:28303891

  9. Social support influences on eating awareness in children and adolescents: the mediating effect of self-regulatory strategies.

    PubMed

    Gaspar de Matos, Margarida; Palmeira, Antonio L; Gaspar, Tania; De Wit, John B F; Luszczynska, Aleksandra

    2016-01-01

    The impact of the social environment on healthy eating awareness results from complex interactions among physical, economic, cultural, interpersonal and individual characteristics. This study investigated the impact of social support and social influence on healthy eating awareness, controlling for socio-economic status, gender and age. Additionally, the mediating effect of self-regulation strategies was examined. A total of 2764 children and adolescents aged 10-17 from four European countries completed self-report measures on healthy eating awareness, social influence and the use of self-regulation strategies. Healthy eating awareness and the use of self-regulation strategies were more likely to occur among younger participants. An interaction between gender and age was related to the use of some self-regulation strategies; compared to girls, boys decreased the use of self-regulation strategies more from pre-adolescence to adolescence. Peer social influence was associated with more unhealthy eating in older participants. Results suggest a need to promote self-regulatory competences among young people in order to assist them with regulating their eating behaviours, especially in the presence of peers. Both school-based interventions and family-based interventions, focusing on self-regulation cognitions and social (peer) influence, could help children and adolescents to use self-regulatory strategies which are essential to eat healthier.

  10. P-type ATPase TAT-2 negatively regulates monomethyl branched-chain fatty acid mediated function in post-embryonic growth and development in C. elegans.

    PubMed

    Seamen, Emylie; Blanchette, Jennifer M; Han, Min

    2009-08-01

    Monomethyl branched-chain fatty acids (mmBCFAs) are essential for Caenorhabditis elegans growth and development. To identify factors acting downstream of mmBCFAs for their function in growth regulation, we conducted a genetic screen for suppressors of the L1 arrest that occurs in animals depleted of the 17-carbon mmBCFA C17ISO. Three of the suppressor mutations defined an unexpected player, the P-type ATPase TAT-2, which belongs to the flippase family of proteins that are implicated in mediating phospholipid bilayer asymmetry. We provide evidence that TAT-2, but not other TAT genes, has a specific role in antagonizing the regulatory activity of mmBCFAs in intestinal cells. Interestingly, we found that mutations in tat-2 also suppress the lethality caused by inhibition of the first step in sphingolipid biosynthesis. We further showed that the fatty acid side-chains of glycosylceramides contain 20%-30% mmBCFAs and that this fraction is greatly diminished in the absence of mmBCFA biosynthesis. These results suggest a model in which a C17ISO-containing sphingolipid may mediate the regulatory functions of mmBCFAs and is negatively regulated by TAT-2 in intestinal cells. This work indicates a novel connection between a P-type ATPase and the critical regulatory function of a specific fatty acid.

  11. The miR-199-dynamin regulatory axis controls receptor-mediated endocytosis.

    PubMed

    Aranda, Juan F; Canfrán-Duque, Alberto; Goedeke, Leigh; Suárez, Yajaira; Fernández-Hernando, Carlos

    2015-09-01

    Small non-coding RNAs (microRNAs) are important regulators of gene expression that modulate many physiological processes; however, their role in regulating intracellular transport remains largely unknown. Intriguingly, we found that the dynamin (DNM) genes, a GTPase family of proteins responsible for endocytosis in eukaryotic cells, encode the conserved miR-199a and miR-199b family of miRNAs within their intronic sequences. Here, we demonstrate that miR-199a and miR-199b regulate endocytic transport by controlling the expression of important mediators of endocytosis such as clathrin heavy chain (CLTC), Rab5A, low-density lipoprotein receptor (LDLR) and caveolin-1 (Cav-1). Importantly, miR-199a-5p and miR-199b-5p overexpression markedly inhibits CLTC, Rab5A, LDLR and Cav-1 expression, thus preventing receptor-mediated endocytosis in human cell lines (Huh7 and HeLa). Of note, miR-199a-5p inhibition increases target gene expression and receptor-mediated endocytosis. Taken together, our work identifies a new mechanism by which microRNAs regulate intracellular trafficking. In particular, we demonstrate that the DNM, miR-199a-5p and miR-199b-5p genes act as a bifunctional locus that regulates endocytosis, thus adding an unexpected layer of complexity in the regulation of intracellular trafficking.

  12. Theory of surfactant-mediated growth on semiconductor surfaces

    NASA Astrophysics Data System (ADS)

    Kaxiras, Efthimios; Kandel, Daniel

    1996-08-01

    The surfactant effect, first demonstrated by Copel et al. [Phys. Rev. Lett. 63 (1989) 632] by using As to promote epitaxial growth of Ge on Si(100), has now been studied in a wide variety of systems, thus making systematic studies possible. We present theoretical models that account for the observed behavior of various surfactants on semiconductor surfaces, including homo-epitaxial and hetero-epitaxial growth. The theoretical models include first-principles calculations of the relative energy of different structures associated with surfactant layers and the activation energies for diffusion and exchange mechanisms, as well as solid-on-solid Monte Carlo simulations.

  13. VEGF and colon cancer growth beyond angiogenesis: does VEGF directly mediate colon cancer growth via a non-angiogenic mechanism?

    PubMed

    Ahluwalia, Amrita; Jones, Michael K; Matysiak-Budnik, Tamara; Tarnawski, Andrzej S

    2014-01-01

    In this article we review the role of vascular endothelial growth factor (VEGF) in colon cancer growth and the underlying mechanisms. Angiogenesis, the growth of new capillary blood vessels in the body, is critical for tissue injury healing and cancer growth. In 1971, Judah Folkman proposed the concept that tumor growth beyond 2 mm is critically dependent on angiogenesis. Tumors including colon cancers release angiogenic growth factors that stimulate blood vessels to grow into the tumors thus providing oxygen and nutrients that enable exponential growth. VEGF is the most potent angiogenic growth factor. Several studies have highlighted the role of VEGF in colon cancer, specifically in the stimulation of angiogenesis. This role of VEGF is strongly supported by studies showing that inhibition of VEGF using the blocking antibody, bevacizumab, results in decreased angiogenesis and abrogation of cancer growth. In the United States, bevacizumab in combination with chemotherapy is FDA approved for the treatment of metastatic colon cancer. However, the source of VEGF in colon cancer tissue, the mechanisms of VEGF generation in colon cancer cells and the molecular pathways involved in VEGF mediated angiogenesis in colon cancer are not fully known. The possibility that VEGF directly stimulates cancer cell growth in an autocrine manner has not been explored in depth.

  14. Cognitive and Emotion-Regulatory Mediators of the Relationship Between Behavioral Approach System Sensitivity and Nonsuicidal Self-Injury Frequency.

    PubMed

    Burke, Taylor A; Stange, Jonathan P; Hamilton, Jessica L; Cohen, Jonah N; O'Garro-Moore, Jared; Daryanani, Issar; Abramson, Lyn Y; Alloy, Lauren B

    2015-08-01

    Nonsuicidal self-injury (NSSI) is highly prevalent among late adolescents and predicts the onset of suicidal ideation and behavior. Although research has established an association between the behavioral approach system (BAS) and NSSI, less research has explored mechanisms underlying this relationship. The authors examined negative and positive emotion regulation patterns, as well as the BAS-relevant cognitive style of self-criticism, as potential mechanisms through which a hypersensitive BAS might be related to NSSI frequency. Late adolescents (N = 177) with high and moderate BAS levels completed measures of self-criticism, positive emotion regulation, brooding, and both lifetime and last-year frequency of NSSI. Results indicated that self-criticism and positive emotion dampening independently mediated the relationship between BAS and last-year frequency of NSSI. Self-criticism also mediated the relationship between BAS and lifetime frequency of NSSI. Results suggest that cognitive and emotion-regulatory styles may help to explain why high BAS individuals are likely to engage in NSSI.

  15. Cutting edge: antigen-specific TGF beta-induced regulatory T cells suppress Th17-mediated autoimmune disease.

    PubMed

    Huter, Eva N; Stummvoll, Georg H; DiPaolo, Richard J; Glass, Deborah D; Shevach, Ethan M

    2008-12-15

    CD4(+) T cells from the TCR transgenic TxA23 mouse recognize a peptide from the H/K-ATPase alpha-chain. When TxA23 CD4(+) thymocytes are differentiated into Th1, Th2, and Th17 lines, all three subpopulations induced autoimmune gastritis (AIG) upon transfer into nu/nu recipients. The induction of AIG by naive T cells or by Th1 or Th2 cell lines could be prevented by the cotransfer of polyclonal Foxp3(+) T regulatory cells (nTreg), whereas Th17-induced AIG was resistant to suppression. We compared the capacity of different types of Treg to suppress Th17-mediated AIG. Cotransfer of either nTreg or polyclonal TGFbeta-induced Treg (iTreg) did not prevent AIG, while cotransfer of TGFbeta-induced Ag-specific TxA23 iTreg completely prevented the development of disease. Ag-specific iTreg were able to suppress Th17-mediated disease when injected 6 days after the Th17 effectors. The implications of these results for the use of Treg for the cellular biotherapy of autoimmune disease are discussed.

  16. Cutting Edge: Antigen-specific TGFβ-induced Regulatory T cells Suppress Th17-Mediated Autoimmune Disease1

    PubMed Central

    Huter, Eva N.; Stummvoll, Georg H.; DiPaolo, Richard J.; Glass, Deborah D.; Shevach, Ethan M.

    2009-01-01

    CD4+ T cells from the TCR transgenic TxA23 mouse recognize a peptide from the H/K-ATPase α-chain. When TxA23 CD4+ thymocytes are differentiated into Th1, Th2, and Th17 lines, all three subpopulations induced autoimmune gastritis (AIG) upon transfer into nu/nu recipients. The induction of AIG by naïve T cells or by Th1, or Th2 cell lines could be prevented by co-transfer of polyclonal Foxp3+ T regulatory cells (nTreg), while Th17-induced AIG was resistant to suppression. We compared the capacity of different types of Treg to suppress Th17-mediated AIG. Co-transfer of either nTreg or polyclonal TGFβ-induced Treg (iTreg) did not prevent AIG, while co-transfer of TGFβ-induced antigen-specific TxA23 iTreg completely prevented development of disease. Antigen-specific iTreg were able to suppress Th17-mediated disease when injected 6 days after the Th17 effectors. The implications of these results for the use of Treg for the cellular biotherapy of autoimmune disease are discussed. PMID:19050237

  17. IL-7 inhibits tumor growth by promoting T cell-mediated antitumor immunity in Meth A model.

    PubMed

    Tang, Jian-Cai; Shen, Guo-Bo; Wang, Shi-Min; Wan, Yong-Sheng; Wei, Yu-Quan

    2014-01-01

    Immune suppression is well documented during tumor progression, which includes loss of effect of T cells and expansion of T regulatory (Treg) cells. IL-7 plays a key role in the proliferation, survival and homeostasis of T cells and displays a potent antitumor activity in vivo. In the present study, we investigated the antitumor effect of IL-7 in Meth A model. IL-7 inhibited tumor growth and prolonged the survival of tumor-bearing mice with corresponding increases in the frequency of CD4 and CD8 T cells, Th1 (CD4(+)IFN-γ(+)), Tc1 (CD8(+)IFN-γ(+)) and T cells cytolytic activity against Meth A cells. Neutralization of CD4 or CD8 T cells reversed the antitumor benefit of IL-7. Furthermore, IL-7 decreased regulatory T Foxp3 as well as cells suppressive activity with a reciprocal increase in SMAD7. In addition, we observed an increase of the serum concentrations of IL-6 and IFN-γ, and a significant decrease of TGF-β and IL-10 after IL-7 treatment. Taken together, these results indicate that IL-7 augments T cell-mediated antitumor immunity and improves the effect of antitumor in Meth A model.

  18. Cell cycle regulatory effects of retinoic Acid and forskolin are mediated by the cyclin C gene.

    PubMed

    Makkonen, Katri M; Malinen, Marjo; Ropponen, Antti; Väisänen, Sami; Carlberg, Carsten

    2009-10-23

    As a partner of cyclin-dependent kinase (CDK) 3, Cyclin C controls cellular proliferation and, together with CDK8, represses gene transcription. In this study, we showed that the highly expressed Cyclin C gene is a direct target of the nuclear hormone all-trans retinoic acid (RA) in HEK293 human embryonal kidney cells. The RA receptor (RAR) gamma associates with a Cyclin C promoter region containing two RAR binding sites. The Cyclin C gene also directly responds to the cAMP activator Forskolin via the transcription factor CREB1 (cAMP response element-binding protein 1), for which we identified four binding sites within the first 2250 bp of its promoter. RARgamma and CREB1 show functional convergence via the corepressor NCoR1, which controls in particular the Forskolin response of Cyclin C. The histone deacetylases 1, 5, 6, 7 and 11 are involved in the basal expression of Cyclin C, but in HEK293 and MCF-7 human breast carcinoma cells the antiproliferative effects of the histone deacetylase inhibitor SAHA (suberoylanilide hydroxamic acid) are not mediated by Cyclin C. However, cell cycle progressing effects of all-trans RA and Forskolin are dependent on Cyclin C expression levels. This suggests that the primary regulation of Cyclin C by all-trans RA and Forskolin mediates some of the cell cycle control actions of these compounds.

  19. Plasmodesmata-mediated intercellular signaling during plant growth and development

    PubMed Central

    Yadav, Shri R.; Yan, Dawei; Sevilem, Iris; Helariutta, Ykä

    2014-01-01

    Plasmodesmata (PD) are cytoplasmic channels that connect neighboring cells for cell-to-cell communication. PD structure and function vary temporally and spatially to allow formation of symplastic domains during different stages of plant development. Reversible deposition of callose at PD plays an important role in controlling molecular trafficking through PD by regulating their size exclusion limit. Previously, we reported several semi-dominant mutants for CALLOSE SYNTHASE 3 (CALS3) gene, which overproduce callose at PD in Arabidopsis. By combining two of these mutations in a LexA-VP16-ER (XVE)-based estradiol inducible vector system, a tool known as the “icals3m system” was developed to temporally obstruct the symplastic connections in a specified spatial domain. The system has been successfully tested and used, in combination with other methods, to investigate the route for mobile signals such as the SHR protein, microRNA165/6, and cytokinins in Arabidopsis roots, and also to understand the role of symplastic domain formation during lateral root development. We envision that this tool may also be useful for identifying tissue-specific symplastic regulatory networks and to analyze symplastic movement of metabolites. PMID:24596574

  20. Plasmodesmata-mediated intercellular signaling during plant growth and development.

    PubMed

    Yadav, Shri R; Yan, Dawei; Sevilem, Iris; Helariutta, Ykä

    2014-01-01

    Plasmodesmata (PD) are cytoplasmic channels that connect neighboring cells for cell-to-cell communication. PD structure and function vary temporally and spatially to allow formation of symplastic domains during different stages of plant development. Reversible deposition of callose at PD plays an important role in controlling molecular trafficking through PD by regulating their size exclusion limit. Previously, we reported several semi-dominant mutants for CALLOSE SYNTHASE 3 (CALS3) gene, which overproduce callose at PD in Arabidopsis. By combining two of these mutations in a LexA-VP16-ER (XVE)-based estradiol inducible vector system, a tool known as the "icals3m system" was developed to temporally obstruct the symplastic connections in a specified spatial domain. The system has been successfully tested and used, in combination with other methods, to investigate the route for mobile signals such as the SHR protein, microRNA165/6, and cytokinins in Arabidopsis roots, and also to understand the role of symplastic domain formation during lateral root development. We envision that this tool may also be useful for identifying tissue-specific symplastic regulatory networks and to analyze symplastic movement of metabolites.

  1. GAGE12 mediates human gastric carcinoma growth and metastasis.

    PubMed

    Lee, Eun Kyung; Song, Kyung-A; Chae, Ji-Hye; Kim, Kyoung-Mee; Kim, Seok-Hyung; Kang, Myung-Soo

    2015-05-15

    The spontaneous metastasis from human gastric carcinoma (GC) remains poorly reproduced in animal models. Here, we established an experimental mouse model in which GC progressively developed in the orthotopic stomach wall and metastasized to multiple organs; the tumors colonized in the ovary exhibited typical characteristics of Krukenberg tumor. The expression of mesenchymal markers was low in primary tumors and high in those in intravasating and extravasating veins. However, the expression of epithelial markers did not differ, indicating that the acquisition of mesenchymal markers without a concordant loss of typical epithelial markers was associated with metastasis. We identified 35 differentially expressed genes (DEGs) in GC cells metastasized to ovary, among which overexpression of GAGE12 family genes, the top-ranked DEGs, were validated. In addition, knockdown of the GAGE12 gene family affected transcription of many of the aforementioned 35 DEGs and inhibited trans-well migration, tumor sphere formation in vitro and tumor growth in vivo. In accordance, GAGE12 overexpression augmented migration, tumor sphere formation and sustained in vivo tumor growth. Taken together, the GAGE12 gene family promotes GC growth and metastasis by modulating the expression of GC metastasis-related genes.

  2. Downregulation of Lnc-Spry1 mediates TGF-β-induced epithelial-mesenchymal transition by transcriptional and posttranscriptional regulatory mechanisms.

    PubMed

    Rodríguez-Mateo, Cristina; Torres, Belén; Gutiérrez, Gabriel; Pintor-Toro, José A

    2017-02-10

    Long non-coding RNAs (lncRNAs) are a class of regulatory genes that participate in a wide range of biological processes, including proliferation, differentiation and development, as well as in a broad spectrum of diseases. Although the role of lncRNAs in TGF-β-induced epithelial-to-mesenchymal transition (EMT) has been well established, little is known about the role of lncRNAs as immediate-early regulators of EMT. Here lnc-Spry1 is identified as an immediate-early regulator of EMT that is downregulated by TGF-β. It is also found that knockdown of lnc-Spry1 promotes a mesenchymal-like phenotype and results in increased cell migration and invasion. In addition, it is shown that lnc-Spry1 depletion preferentially affects the expression of TGF-β-regulated gene targets. Moreover, lnc-Spry1 associates with U2AF65 splicing factor, suggesting a role in alternative splicing. Depletion of lnc-Spry1 induces, as TGF-β, isoform switching of fibroblast growth factor receptors, resulting in FGF-2-sensitive cells. Taken together, these results show that lnc-Spry1 could act as an early mediator of TGF-β signaling and reveal different roles for a lncRNA in modulating transcriptional and posttranscriptional gene expressionCell Death and Differentiation advance online publication, 10 February 2017; doi:10.1038/cdd.2017.9.

  3. Gene Targets in Prostate Tumor Cells that Mediate Aberrant Growth and Invasiveness

    DTIC Science & Technology

    2005-02-01

    Craig A. Hauser , Ph.D. Gabriele Foos, Ph.D. CONTRACTING ORGANIZATION: The Burnham Institute La Jolla, California 92037 REPORT DATE: February 2005 TYPE...NUMBERS Gene Targets in Prostate Tumor Cells that Mediate DAMD17-02-1-0019 Aberrant Growth and Invasiveness 6. AUTHOR(S) Craig A. Hauser , Ph.D. Gabriele...REPORTABLE OUTCOMES Foos G, Hauser CA (2004) The role of Ets transcription factors in mediating cellular transformation. In: Handbook of Experimental

  4. Positive and negative regulatory elements mediating transcription from the Drosophila melanogaster actin 5C distal promoter.

    PubMed Central

    Chung, Y T; Keller, E B

    1990-01-01

    The major cytoskeletal actin gene of Drosophila melanogaster, the actin 5C gene, has two promoters, the distal one of which controls synthesis of actin in a tissue- and developmental stage-specific manner. This very strong promoter has widely been used for expression of heterologous genes in cultured cells. To locate functional regulatory elements in this distal promoter, mutants of the promoter were fused to the bacterial chloramphenicol acetyltransferase gene and assayed for transient expression activity in cultured Drosophila embryonic Schneider line 2 cells. The results showed that the upstream end of the promoter extends to 522 bp from the transcription start site. In addition, there are two remote activating regions about 2 kb upstream. Between -522 and -379 are two regions that exert a strong negative effect. Downstream from these negative regions are at least six positive regions and a TATA element. The strongest positive determinant of the promoter was identified at -320 as AAAATGTG by footprinting and by a replacement experiment. When the relevant region was replaced by a synthetic sequence containing this element in a random context, the transient expression activity was restored. The sequence TGTATG located at -355 was also identified as a positive element by a similar replacement approach. Apparently the very high activity of this promoter is the result of the combined activities of multiple factors. Images PMID:2123290

  5. Alterations in CD46-mediated Tr1 regulatory T cells in patients with multiple sclerosis

    PubMed Central

    Astier, Anne L.; Meiffren, Gregory; Freeman, Samuel; Hafler, David A.

    2006-01-01

    Loss of Treg function appears to be a critical factor in the pathogenesis of human autoimmune diseases. Attention has focused on defects of CD4+CD25high Tregs, and techniques have been developed to determine their function. In contrast, the role of Tr1 regulatory T cells, which secrete the antiinflammatory cytokine IL-10, in autoimmune disease has not been well assessed. CD46 is a newly defined costimulatory molecule for T cell activation, and CD46-costimulated human T cells induce a Tr1 Treg phenotype with considerable amounts of IL-10 secretion. Here, we examined the role of Tr1 cells in patients with multiple sclerosis (MS) by stimulating CD4+ T cells with anti-CD3 and -CD46 mAbs and measuring IL-10 secretion. There were striking defects in the induction of Tr1 cells with CD46 costimulation as measured by IL-10 but not IFN-γ secretion in patients with MS compared with healthy subjects. This loss of Tr1 cell–associated IL-10 secretion was specific to CD46 and not CD28 costimulation and was associated with an altered regulation of the CD46-Cy2 isoform that differentially regulates T cell function in a CD46-transgenic murine model. These data demonstrate a second major Treg defect in human autoimmune disease associated with the CD46 pathway. PMID:17099776

  6. PRMT1 mediated methylation of TAF15 is required for its positive gene regulatory function

    SciTech Connect

    Jobert, Laure; Argentini, Manuela; Tora, Laszlo

    2009-04-15

    TAF15 (formerly TAF{sub II}68) is a nuclear RNA-binding protein that is associated with a distinct population of TFIID and RNA polymerase II complexes. TAF15 harbours an N-terminal activation domain, an RNA recognition motif (RRM) and many Arg-Gly-Gly (RGG) repeats at its C-terminal end. The N-terminus of TAF15 serves as an essential transforming domain in the fusion oncoprotein created by chromosomal translocation in certain human chondrosarcomas. Post-transcriptional modifications (PTMs) of proteins are known to regulate their activity, however, nothing is known on how PTMs affect TAF15 function. Here we demonstrate that endogenous human TAF15 is methylated in vivo at its numerous RGG repeats. Furthermore, we identify protein arginine N-methyltransferase 1 (PRMT1) as a TAF15 interactor and the major PRMT responsible for its methylation. In addition, the RGG repeat-containing C-terminus of TAF15 is responsible for the shuttling between the nucleus and the cytoplasm and the methylation of RGG repeats affects the subcellular localization of TAF15. The methylation of TAF15 by PRMT1 is required for the ability of TAF15 to positively regulate the expression of the studied endogenous TAF15-target genes. Our findings demonstrate that arginine methylation of TAF15 by PRMT1 is a crucial event determining its proper localization and gene regulatory function.

  7. Differential potency of regulatory T cell-mediated immunosuppression in kidney tumors compared to subcutaneous tumors

    PubMed Central

    Devaud, Christel; Westwood, Jennifer A; Teng, Michele WL; John, Liza B; Yong, Carmen SM; Duong, Connie PM; Smyth, Mark J; Darcy, Phillip K; Kershaw, Michael H

    2014-01-01

    In many cancers, regulatory T cells (Treg) play a crucial role in suppressing the effector immune response thereby permitting tumor development. Indeed, in mouse models, their depletion can promote the regression of tumors of various origins, including renal cell carcinoma when located subcutaneous (SC). In the present study, we aimed to assess the importance of Treg immunosuppression in the physiologic context of metastatic renal carcinoma (Renca) disease. To that purpose we inoculated renal tumors orthotopically, intra-kidney (IK), in mice. Treg depletions were performed using anti-CD4 antibody in wild type mice or diphtheria toxin (DT) in Foxp3DTR transgenic mice. Our main observation was that Treg were not the key immunosuppressive component of the IK tumoral microenvironment, compared to the same tumors located SC. We demonstrated that the CD8+ effector immune response was still suppressed in IK tumors when compared to SC tumors, following Treg depletion. Furthermore, the level of program cell death protein (PD)-1 was increased on the surface of CD4+ T cells infiltrating IK tumors compared to SC tumors. Finally, the Treg-independent immunosuppression, occurring in IK tumors, was potent enough to inhibit regression of concomitant SC tumors, normally responsive to Treg depletion. Our findings provide further insight into the immunosuppressive nature of the immune response generated in the kidney microenvironment, suggesting that it can have additional mechanisms in addition to Treg. These observations might help to identify better targets from the kidney tumor microenvironment for future cancer therapies. PMID:25941590

  8. Differential potency of regulatory T cell-mediated immunosuppression in kidney tumors compared to subcutaneous tumors.

    PubMed

    Devaud, Christel; Westwood, Jennifer A; Teng, Michele Wl; John, Liza B; Yong, Carmen Sm; Duong, Connie Pm; Smyth, Mark J; Darcy, Phillip K; Kershaw, Michael H

    2014-11-01

    In many cancers, regulatory T cells (Treg) play a crucial role in suppressing the effector immune response thereby permitting tumor development. Indeed, in mouse models, their depletion can promote the regression of tumors of various origins, including renal cell carcinoma when located subcutaneous (SC). In the present study, we aimed to assess the importance of Treg immunosuppression in the physiologic context of metastatic renal carcinoma (Renca) disease. To that purpose we inoculated renal tumors orthotopically, intra-kidney (IK), in mice. Treg depletions were performed using anti-CD4 antibody in wild type mice or diphtheria toxin (DT) in Foxp3(DTR) transgenic mice. Our main observation was that Treg were not the key immunosuppressive component of the IK tumoral microenvironment, compared to the same tumors located SC. We demonstrated that the CD8(+) effector immune response was still suppressed in IK tumors when compared to SC tumors, following Treg depletion. Furthermore, the level of program cell death protein (PD)-1 was increased on the surface of CD4(+) T cells infiltrating IK tumors compared to SC tumors. Finally, the Treg-independent immunosuppression, occurring in IK tumors, was potent enough to inhibit regression of concomitant SC tumors, normally responsive to Treg depletion. Our findings provide further insight into the immunosuppressive nature of the immune response generated in the kidney microenvironment, suggesting that it can have additional mechanisms in addition to Treg. These observations might help to identify better targets from the kidney tumor microenvironment for future cancer therapies.

  9. Protective role of interferon regulatory factor 3-mediated signaling against prion infection.

    PubMed

    Ishibashi, Daisuke; Atarashi, Ryuichiro; Fuse, Takayuki; Nakagaki, Takehiro; Yamaguchi, Naohiro; Satoh, Katsuya; Honda, Kenya; Nishida, Noriyuki

    2012-05-01

    Abnormal prion protein (PrP(Sc)) generated from the cellular isoform of PrP (PrP(C)) is assumed to be the main or sole component of the pathogen, called prion, of transmissible spongiform encephalopathies (TSE). Because PrP is a host-encoded protein, acquired immune responses are not induced in TSE. Meanwhile, activation of the innate immune system has been suggested to partially block the progression of TSE; however, the mechanism is not well understood. To further elucidate the role of the innate immune system in prion infection, we investigated the function of interferon regulatory factor 3 (IRF3), a key transcription factor of the MyD88-independent type I interferon (IFN) production pathway. We found that IRF3-deficient mice exhibited significantly earlier onset with three murine TSE strains, namely, 22L, FK-1, and murine bovine spongiform encephalopathy (mBSE), following intraperitoneal transmission, than with wild-type controls. Moreover, overexpression of IRF3 attenuated prion infection in the cell culture system, while PrP(Sc) was increased in prion-infected cells treated with small interfering RNAs (siRNAs) against IRF3, suggesting that IRF3 negatively regulates PrP(Sc) formation. Our findings provide new insight into the role of the host innate immune system in the pathogenesis of prion diseases.

  10. Protective Role of Interferon Regulatory Factor 3-Mediated Signaling against Prion Infection

    PubMed Central

    Atarashi, Ryuichiro; Fuse, Takayuki; Nakagaki, Takehiro; Yamaguchi, Naohiro; Satoh, Katsuya; Honda, Kenya; Nishida, Noriyuki

    2012-01-01

    Abnormal prion protein (PrPSc) generated from the cellular isoform of PrP (PrPC) is assumed to be the main or sole component of the pathogen, called prion, of transmissible spongiform encephalopathies (TSE). Because PrP is a host-encoded protein, acquired immune responses are not induced in TSE. Meanwhile, activation of the innate immune system has been suggested to partially block the progression of TSE; however, the mechanism is not well understood. To further elucidate the role of the innate immune system in prion infection, we investigated the function of interferon regulatory factor 3 (IRF3), a key transcription factor of the MyD88-independent type I interferon (IFN) production pathway. We found that IRF3-deficient mice exhibited significantly earlier onset with three murine TSE strains, namely, 22L, FK-1, and murine bovine spongiform encephalopathy (mBSE), following intraperitoneal transmission, than with wild-type controls. Moreover, overexpression of IRF3 attenuated prion infection in the cell culture system, while PrPSc was increased in prion-infected cells treated with small interfering RNAs (siRNAs) against IRF3, suggesting that IRF3 negatively regulates PrPSc formation. Our findings provide new insight into the role of the host innate immune system in the pathogenesis of prion diseases. PMID:22379081

  11. Nitric Oxide Mediated Transcriptome Profiling Reveals Activation of Multiple Regulatory Pathways in Arabidopsis thaliana

    PubMed Central

    Hussain, Adil; Mun, Bong-Gyu; Imran, Qari M.; Lee, Sang-Uk; Adamu, Teferi A.; Shahid, Muhammad; Kim, Kyung-Min; Yun, Byung-Wook

    2016-01-01

    Imbalance between the accumulation and removal of nitric oxide and its derivatives is a challenge faced by all plants at the cellular level, and is especially important under stress conditions. Exposure of plants to various biotic and abiotic stresses causes rapid changes in cellular redox tone potentiated by the rise in reactive nitrogen species that serve as signaling molecules in mediating defensive responses. To understand mechanisms mediated by these signaling molecules, we performed a large-scale analysis of the Arabidopsis transcriptome induced by nitrosative stress. We generated an average of 84 and 91 million reads from three replicates each of control and 1 mM S-nitrosocysteine (CysNO)-infiltrated Arabidopsis leaf samples, respectively. After alignment, more than 95% of all reads successfully mapped to the reference and 32,535 genes and 55,682 transcripts were obtained. CysNO infiltration caused differential expression of 6436 genes (3448 up-regulated and 2988 down-regulated) and 6214 transcripts (3335 up-regulated and 2879 down-regulated) 6 h post-infiltration. These differentially expressed genes were found to be involved in key physiological processes, including plant defense against various biotic and abiotic stresses, hormone signaling, and other developmental processes. After quantile normalization of the FPKM values followed by student's T-test (P < 0.05) we identified 1165 DEGs (463 up-regulated and 702 down-regulated) with at least 2-folds change in expression after CysNO treatment. Expression patterns of selected genes involved in various biological pathways were verified using quantitative real-time PCR. This study provides comprehensive information about plant responses to nitrosative stress at transcript level and would prove helpful in understanding and incorporating mechanisms associated with nitrosative stress responses in plants. PMID:27446194

  12. Mediator of ERBB2-driven cell motility (MEMO) promotes extranuclear estrogen receptor signaling involving the growth factor receptors IGF1R and ERBB2.

    PubMed

    Jiang, Kai; Yang, Zhihong; Cheng, Long; Wang, Shibin; Ning, Kang; Zhou, Lei; Lin, Jing; Zhong, Hui; Wang, Lisheng; Li, Yang; Huang, Junjian; Zhang, Hao; Ye, Qinong

    2013-08-23

    In addition to nuclear estrogen receptor (ER) acting as a transcription factor, extranuclear ER also plays an important role in cancer cell growth regulation through activation of kinase cascades. However, the molecular mechanisms by which extranuclear ER exerts its function are still poorly understood. Here, we report that mediator of ERBB2-driven cell motility (MEMO) regulates extranuclear functions of ER. MEMO physically and functionally interacted with ER. Through its interaction with the growth factor receptors IGF1R and ERBB2, MEMO mediated extranuclear functions of ER, including activation of mitogen-activated protein kinase (MAPK) and protein kinase B/AKT, two important growth regulatory protein kinases, and integration of function with nuclear ER. Activation of MAPK and AKT was responsible for MEMO modulation of ER phosphorylation and estrogen-responsive gene expression. Moreover, MEMO increased anchorage-dependent and -independent growth of ER-positive breast cancer cells in vitro and was required for estrogen-induced breast tumor growth in nude mice. Together, our studies identified MEMO as a new component of extranuclear ER signalosome and suggest an essential role for MEMO in the regulation of ER-positive breast cancer cell growth.

  13. Prolactin mediates psychological stress-induced dysfunction of regulatory T cells to facilitate intestinal inflammation

    PubMed Central

    Wu, Wei; Sun, Mingming; Zhang, Huan-Ping; Chen, Tengfei; Wu, Ruijin; Liu, Changqin; Yang, Gui; Geng, Xiao-Rui; Feng, Bai-Sui; Liu, Zhigang; Liu, Zhanju; Yang, Ping-Chang

    2014-01-01

    Objective The dysfunction of immune regulation plays a critical role in the pathogenesis of a number of chronic inflammatory disorders, such as IBD. A close relationship between psychological stress and intestinal inflammation has been noted; the underlying mechanism remains elusive. This study aims to elucidate a pathological pathway between psychological stress and the dysfunction of regulatory T cells (Treg), and its effect on facilitating intestinal inflammation. Design A restraint stress model was employed to induce psychological stress in mice. The functions of Tregs were determined by assessing the immune suppressor effects in the intestine. A mouse model of intestinal inflammation was established using a low dose of trinitrobenzene sulfonic acid (TNBS) or dextran sulfate sodium (DSS) together with the challenge of chronic stress. Results After treating mice with restraint stress, the suppressor function of intestinal Treg was compromised, although the frequency of Treg was not changed in the intestine. Further observation revealed that stress induced Tregs in the intestine to differentiate into foxhead box P3+ interleukin (IL)-17+ tumour necrosis factor (TNF)-α+ T cells. We also observed that exposure to stress-derived prolactin induced dendritic cells (DC) to produce IL-6 and IL-23 in vitro and in vivo, which played a critical role in altering Treg's phenotypes. Treating mice with chronic stress facilitated the initiation of intestinal inflammation by a low dose of TNBS or DSS, which was abolished by pretreatment with an inhibitor of prolactin, the cabergoline. Conclusions Psychological stress-derived prolactin alters DC and Treg's properties to contribute to intestinal inflammation. PMID:24550371

  14. Dynamic designing of microstructures by chemical gradient-mediated growth

    PubMed Central

    Shim, Tae Soup; Yang, Seung-Man; Kim, Shin-Hyun

    2015-01-01

    Shape is one of the most important determinants of the properties of microstructures. Despite of a recent progress on microfabrication techniques, production of three-dimensional micro-objects are yet to be fully achieved. Nature uses reaction–diffusion process during bottom-up self-assembly to create functional shapes and patterns with high complexity. Here we report a method to produce polymeric microstructures by using a dynamic reaction–diffusion process during top-down photolithography, providing unprecedented control over shape and composition. In radical polymerization, oxygen inhibits reaction, and therefore diffusion of oxygen significantly alters spatial distribution of growth rate. Therefore, growth pathways of the microstructures can be controlled by engineering a concentration gradient of oxygen. Moreover, stepwise control of chemical gradients enables the creation of highly complex microstructures. The ease of use and high controllability of this technology provide new opportunities for microfabrication and for fundamental studies on the relationships between shape and function for the materials. PMID:25766762

  15. Autophagy is required for IL-2-mediated fibroblast growth

    SciTech Connect

    Kang, Rui; Tang, Daolin; Lotze, Michael T.; Zeh III, Herbert J.

    2013-02-15

    Autophagy is an evolutionarily conserved pathway responsible for delivery of cytoplasmic material into the lysosomal degradation pathway to enable vesicular exocytosis. Interleukin (IL)-2 is produced by T-cells and its activity is important for immunoregulation. Fibroblasts are an immune competent cell type, playing a critical role in wound healing, chronic inflammation, and tumor development. Although autophagy plays an important role in each of these processes, whether it regulates IL-2 activity in fibroblasts is unknown. Here, we show that autophagy is required for IL-2-induced cell growth in fibroblasts. IL-2 significantly induced autophagy in mouse embryonic fibroblasts (MEFs) and primary lung fibroblasts. Autophagy inhibitors (e.g., 3-methylamphetamine and bafilomycin A1) or knockdown of ATG5 and beclin 1 blocked clinical grade IL-2-induced autophagy. Moreover, IL-2 induced HMGB1 cytoplasmic translocation in MEFs and promoted interaction between HMGB1 and beclin1, which is required for autophagy induction. Pharmacological and genetic inhibition of autophagy inhibited IL-2-induced cell proliferation and enhanced IL-2-induced apoptosis. These findings suggest that autophagy is an important pro-survival regulator for IL-2-induced cell growth in fibroblasts.

  16. The kinases MEKK2 and MEKK3 regulate transforming growth factor-β-mediated helper T cell differentiation.

    PubMed

    Chang, Xing; Liu, Fang; Wang, Xiaofang; Lin, Aiping; Zhao, Hongyu; Su, Bing

    2011-02-25

    Mitogen-activated protein kinases (MAPKs) are key mediators of the T cell receptor (TCR) signals but their roles in T helper (Th) cell differentiation are unclear. Here we showed that the MAPK kinase kinases MEKK2 (encoded by Map3k2) and MEKK3 (encoded by Map3k3) negatively regulated transforming growth factor-β (TGF-β)-mediated Th cell differentiation. Map3k2(-/-)Map3k3(Lck-Cre/-) mice showed an abnormal accumulation of regulatory T (Treg) and Th17 cells in the periphery, consistent with Map3k2(-/-)Map3k3(Lck-Cre/-) naive CD4(+) T cells' differentiation into Treg and Th17 cells with a higher frequency than wild-type (WT) cells after TGF-β stimulation in vitro. In addition, Map3k2(-/-)Map3k3(Lck-Cre/-) mice developed more severe experimental autoimmune encephalomyelitis. Map3k2(-/-)Map3k3(Lck-Cre/-) T cells exhibited impaired phosphorylation of SMAD2 and SMAD3 proteins at their linker regions, which negatively regulated the TGF-β responses in T cells. Thus, the crosstalk between TCR-induced MAPK and the TGF-β signaling pathways is important in regulating Th cell differentiation.

  17. Requirement of Stat3 but not Stat1 activation for epidermal growth factor receptor- mediated cell growth In vitro.

    PubMed Central

    Grandis, J R; Drenning, S D; Chakraborty, A; Zhou, M Y; Zeng, Q; Pitt, A S; Tweardy, D J

    1998-01-01

    Stimulation of epidermal growth factor receptor (EGFR) by ligand(s) leads to activation of signaling molecules including Stat1 and Stat3, two members of the signal transducers and activators of transcription (STAT) protein family. Activation of Stat1 and Stat3 was constitutive in transformed squamous epithelial cells, which produce elevated levels of TGF-alpha, and was enhanced by the addition of exogenous TGF-alpha. Targeting of Stat3 using antisense oligonucleotides directed against the translation initiation site, resulted in significant growth inhibition. In addition, cells stably transfected with dominant negative mutant Stat3 constructs failed to proliferate in vitro. In contrast, targeting of Stat1 using either antisense or dominant-negative strategies had no effect on cell growth. Thus, TGF-alpha/EGFR-mediated autocrine growth of transformed epithelial cells is dependent on activation of Stat3 but not Stat1. PMID:9769331

  18. Mullerian Inhibiting Substance (MIS) Augments IFN-Gamma Mediated Inhibition of Breast Cancer Cell Growth

    DTIC Science & Technology

    2005-06-01

    demonstrated that MIS promotes IFN-y-induced apoptosis demonstrating a functional interaction between these two classes of signaling molecules in regulation...mammary tumors which do not express ER and have functional inactivation of Rb and p53. In agreement with results observed in the transgenic mice, MIS...Interferon regulatory factor-1 (IRF-1), a gene known for its growth inhibitory functions in breast cancer cells is induced by MIS and IFN-y through a NFkB

  19. Mediating role of coping in the dispositional optimism-posttraumatic growth relation in breast cancer patients.

    PubMed

    Büyükaşik-Colak, Canan; Gündoğdu-Aktürk, Elçin; Bozo, Ozlem

    2012-01-01

    The aim of the present study was to examine if coping strategies mediate dispositional optimism-posttraumatic growth relation in postoperative breast cancer patients. The data were collected from 90 patients in two hospitals. Regression analyses revealed that problem-focused coping fully mediated dispositional optimism-posttraumatic growth relation, but emotion-focused coping did not. That is, postoperative breast cancer patients who were optimistic were more likely to use problem-focused coping strategies that, in turn, led to the development of posttraumatic growth. The findings were congruent with the literature in which problem-focused coping was mostly highlighted as compared to emotion-focused coping, and in which optimism and problem-focused coping relationship was emphasized in the path of posttraumatic growth.

  20. Hormone stimulation of androgen receptor mediates dynamic changes in DNA methylation patterns at regulatory elements

    PubMed Central

    Dhiman, Vineet K.; Attwood, Kristopher; Campbell, Moray J.; Smiraglia, Dominic J.

    2015-01-01

    DNA methylation is an epigenetic modification that contributes to stable gene silencing by interfering with the ability of transcriptional regulators to bind to DNA. Recent findings have revealed that hormone stimulation of certain nuclear receptors induces rapid, dynamic changes in DNA methylation patterns alongside transcriptional responses at a subset of target loci, over time. However, the ability of androgen receptor (AR) to dynamically regulate gene transcription is relatively under-studied and its role in the regulation of DNA methylation patterns remains to be elucidated. Here we demonstrate in normal prostate cells that hormone stimulated AR activity results in dynamic changes in the transcription rate and DNA methylation patterns at the AR target genes, TIPARP and SGK1. Time-resolved chromatin immunoprecipitation experiments on the SGK1 locus reveals dynamic recruitment of AR and RNA Polymerase II, as well as the recruitment of proteins involved in the DNA demethylation process, TET1 and TDG. Furthermore, the presence of DNA methylation at dynamic regions inhibits protein binding and transcriptional activity of SGK1. These findings establish AR activity as a contributing factor to the dynamic regulation of DNA methylation patterns at target genes in prostate biology and infer further complexity involved in nuclear receptor mediation of transcriptional regulation. PMID:26646795

  1. Alternative splicing and nonsense-mediated decay modulate expression of important regulatory genes in Arabidopsis

    PubMed Central

    Kalyna, Maria; Simpson, Craig G.; Syed, Naeem H.; Lewandowska, Dominika; Marquez, Yamile; Kusenda, Branislav; Marshall, Jacqueline; Fuller, John; Cardle, Linda; McNicol, Jim; Dinh, Huy Q.; Barta, Andrea; Brown, John W. S.

    2012-01-01

    Alternative splicing (AS) coupled to nonsense-mediated decay (NMD) is a post-transcriptional mechanism for regulating gene expression. We have used a high-resolution AS RT–PCR panel to identify endogenous AS isoforms which increase in abundance when NMD is impaired in the Arabidopsis NMD factor mutants, upf1-5 and upf3-1. Of 270 AS genes (950 transcripts) on the panel, 102 transcripts from 97 genes (32%) were identified as NMD targets. Extrapolating from these data around 13% of intron-containing genes in the Arabidopsis genome are potentially regulated by AS/NMD. This cohort of naturally occurring NMD-sensitive AS transcripts also allowed the analysis of the signals for NMD in plants. We show the importance of AS in introns in 5′ or 3′UTRs in modulating NMD-sensitivity of mRNA transcripts. In particular, we identified upstream open reading frames overlapping the main start codon as a new trigger for NMD in plants and determined that NMD is induced if 3′-UTRs were >350 nt. Unexpectedly, although many intron retention transcripts possess NMD features, they are not sensitive to NMD. Finally, we have shown that AS/NMD regulates the abundance of transcripts of many genes important for plant development and adaptation including transcription factors, RNA processing factors and stress response genes. PMID:22127866

  2. Webinar Presentation: Exposures to Polycyclic Aromatic Hydrocarbons and Childhood Growth Trajectories and Body Composition: Linkages to Disrupted Self-Regulatory Processes

    EPA Pesticide Factsheets

    This presentation, Exposures to Polycyclic Aromatic Hydrocarbons and Childhood Growth Trajectories and Body Composition: Linkages to Disrupted Self-Regulatory Processes, was given at the NIEHS/EPA Children's Centers 2016 Webinar Series: Childhood Obesity

  3. Mammalian mediator 19 mediates H1299 lung adenocarcinoma cell clone conformation, growth, and metastasis.

    PubMed

    Xu, Lu-Lu; Guo, Shu-Liang; Ma, Su-Ren; Luo, Yong-Ai

    2012-01-01

    Mammalian mediator (MED) is a multi-protein coactivator that has been identified by several research groups. The involvement of the MED complex subunit 19 (MED 19) in the metastasis of lung adenocarcinoma cell line (H1299), which expresses the MED 19 subunit, was here investigated. When MED 19 expression was decreased by RNA interference H1299 cells demonstrated reduced clone formation, arrest in the S phase of the cell cycle, and lowered metastatic capacity. Thus, MED 19 appears to play important roles in the biological behavior of non-small cell lung carcinoma cells. These findings may be important for the development of novel lung carcinoma treatments.

  4. ECL-cell histamine mobilization in conscious rats: effects of locally applied regulatory peptides, candidate neurotransmitters and inflammatory mediators.

    PubMed

    Norlén, P; Bernsand, M; Konagaya, T; Håkanson, R

    2001-12-01

    1. The ECL cells control gastric acid secretion by mobilizing histamine in response to circulating gastrin. In addition, the ECL cells are thought to operate under nervous control and to be influenced by local inflammatory processes. 2. The purpose of the present study was to monitor histamine mobilization from ECL cells in conscious rats in response to locally applied regulatory peptides, candidate neurotransmitters and inflammatory mediators. 3. Microdialysis probes were implanted in the submucosa of the acid-producing part of the rat stomach. Three days later, the agents to be tested were administered via the microdialysis probe and their effects on basal (48 h fast) and stimulated (intravenous infusion of gastrin-17, 3 nmol kg(-1) h(-1)) mobilization of ECL-cell histamine was monitored by continuous measurement of histamine in the perfusate (radioimmunoassay). 4. Locally administered gastrin-17 and sulfated cholecystokinin-8 mobilized histamine as did pituitary adenylate cyclase-activating peptide-27, vasoactive intestinal peptide, peptide YY, met-enkephalin, endothelin and noradrenaline, adrenaline and isoprenaline. 5. While gastrin, sulfated-cholecystokinin-8, met-enkephalin and isoprenaline induced a sustained elevation of the submucosal histamine concentration, endothelin, peptide YY, pituitary adenylate cyclase activating peptide, vasoactive intestinal peptide, noradrenaline and adrenaline induced a transient elevation. 6. Calcitonin gene-related peptide, galanin, somatostatin and the prostanoid misoprostol inhibited gastrin-stimulated histamine mobilization. 7. The gut hormones neurotensin and secretin and the neuropeptides gastrin-releasing peptide, neuropeptide Y and substance P failed to affect ECL-cell histamine mobilization, while motilin and neuromedin U-25 had weak stimulatory effects. Also acetylcholine, carbachol, serotonin and the amino acid neurotransmitters aspartate, gamma-aminobutyric acid, glutamate and glycine were inactive or weakly

  5. Selective Photoreceptor Gene Knock-out Reveals a Regulatory Role for the Growth Behavior of Pseudomonas syringae.

    PubMed

    Shah, Rashmi; Pathak, Gopal; Drepper, Thomas; Gärtner, Wolfgang

    2016-07-01

    The plant pathogen Pseudomonas syringae (Ps) is a well-established model organism for bacterial infection of plants. The genome sequences of two pathovars, pv. syringae and pv. tomato, revealed one gene encoding a blue and two genes encoding red/far red light-sensing photoreceptors. Continuing former molecular characterization of the photoreceptor proteins, we here report selective photoreceptor gene disruption for pv. tomato aiming at identification of potentially regulatory functions of these photoreceptors. Transformation of Ps cells with linear DNA constructs yielded interposon mutations of the corresponding genes. Cell growth studies of the generated photoreceptor knock-out mutants revealed their role in light-dependent regulation of cell growth and motility. Disruption of the blue-light (BL) receptor gene caused a growth deregulation, in line with an observed increased virulence of this mutant (Moriconi et al., Plant J., 2013, 76, 322). Bacterial phytochrome-1 (BphP1) deletion mutant caused unaltered cell growth, but a stronger swarming capacity. Inactivation of its ortholog, BphP2, however, caused reduced growth and remarkably altered dendritic swarming behavior. Combined knock-out of both bacteriophytochromes reproduced the swarming pattern observed for the BphP2 mutant alone. A triple knock-out mutant showed a growth rate between that of the BL (deregulation) and the phytochrome-2 mutant (growth reduction).

  6. Ag nanoparticle mediated growth of CdS nanobelts

    NASA Astrophysics Data System (ADS)

    Sreejith, K.; Nuwad, J.; Thinaharan, C.; Dey, G. K.; Pillai, C. G. S.

    2007-06-01

    Catalytic growth of CdS have been carried out on large scale by evaporation of bulk CdS on Ag deposited Si (1 1 1) at atmospheric pressure. The as prepared CdS had wurtzite structure as evidenced by X-ray diffraction. The nanostructures were beltlike with several tens of micrometers length, several micrometers width and few nanometers to tens of nanometers thick as seen by scanning electron microscope and confirmed by TEM studies. The nanobelts were single crystalline in nature and showed reflection corresponding to (1 1 2) and (0 0 2) planes in SAED. The PL studies revealed the green band due to band gap emission and red band due to emission from the surface states. The higher intensity of the defect emission indicated the presence of considerable concentration of surface defects in the as prepared sample. The deposition of CdS could be explained on the basis of catalyst assisted vapor-liquid-solid and vapor-solid mechanism.

  7. Herpesvirus-mediated systemic delivery of nerve growth factor.

    PubMed

    Wolfe, D; Goins, W F; Kaplan, T J; Capuano, S V; Fradette, J; Murphey-Corb, M; Robbins, P D; Cohen, J B; Glorioso, J C

    2001-01-01

    Sustained systemic dissemination of therapeutic proteins from peripheral sites is an attractive prospect for gene therapy applications. Replication-defective genomic herpes simplex virus type 1 (HSV-1) vectors were evaluated for their ability to express nerve growth factor (NGF) as a model gene product both locally and systemically. Intra-articular inoculation of NGF expression vectors in rabbits resulted in significant increases in joint lavage and blood plasma NGF that persisted for 1 year. A rhesus macaque injected intra-articularly displayed a comparable increase in plasma NGF for at least 6 months, at which time the serum NGF levels of this animal were sufficient to cause differentiation of PC12 cells in culture, but not to increase footpad epidermis innervation. Long-term reporter transgene expression was observed primarily in ligaments, a finding confirmed by direct inoculation of patellar ligament. Patellar ligament inoculation with a NGF vector resulted in elevated levels of circulating NGF similar to those observed following intra-articular vector delivery. These results represent the first demonstration of sustained systemic release of a transgene product using HSV vectors, raising the prospect of new applications for HSV-1 vectors in the treatment of systemic disease.

  8. MECP2 promotes the growth of gastric cancer cells by suppressing miR-338-mediated antiproliferative effect

    PubMed Central

    Tong, Dongdong; Zhao, Lingyu; He, Kang; Sun, Hongfei; Cai, Donghui; Ni, Lei; Sun, Ruifang; Chang, Su'e; Song, Tusheng; Huang, Chen

    2016-01-01

    The methyl-CpG-binding protein 2 (MECP2), a transcriptional suppressor, is involved in gene regulation by binding to methylated promoters. We found that MECP2 is overexpressed in gastric cancer (GC), and that Mecp2 knockdown affects the growth of GC cells both in vitro and in vivo. MECP2 can directly bind to the methylated-CpG island of miR-338 promoter and suppress the expression of two mature microRNAs, namely, miR-338-3p and miR-338-5p. Furthermore, miR-338-5p can suppress GC cell growth by targeting BMI1 (B lymphoma Mo-MLV insertion region 1 homolog). We additionally found that decreased miR-338-5p expression in GC tissues, relative to normal tissues, was significantly negatively correlated with increased BMI1 expression. Silencing MECP2 can indirectly lead to reduced expression of P-REX2, which has been identified as the miR-338-3p target, as well as BMI1 and increasing expression of P16 or P21 both in vitro and in vivo. Altogether, our results indicate that MECP2 promote the proliferation of GC cells via miR-338 (miR-338-3p and miR-338-5p)-mediated antitumor and gene regulatory effect. PMID:27166996

  9. Hypoxia-induced CCL28 promotes recruitment of regulatory T cells and tumor growth in liver cancer

    PubMed Central

    Wang, Li; Zhu, Zhifeng; Lu, Rong; Yao, Zhi

    2016-01-01

    Tumor cells craft microenvironment to overcome growth disadvantages and adjust to escape the immunosurveillance during tumorigenesis and metastasis. The evolving adaption to the changing microenvironment is exemplified by the development of strategies to deal with hypoxia resulted from fast proliferation of the tumor cells. In this study, we found that hypoxia hepatocellular carcinoma (HCC) cells recruited Regulatory T cells (Tregs) and expressed more Chemokine (C-C motif) ligand 28 (CCL28). Deletion of CCL28 inhibited Treg recruitment. Furthermore, overexpression of CCL28 promoted tumor growth and Treg infiltration in vivo. Enhanced angiogenesis and VEGF expression was also observed. Moreover, inhibition of HIF1α reversed hypoxia-induced CCL28 upregulation. Taken together, our results demonstrate that HCC recruits Tregs to promote angiogenesis under hypoxic condition by upregulating CCL28 expression. These findings establish a link between Tregs and hypoxia in HCC growth and may provide a new potential therapeutic target for treating HCC. PMID:27716621

  10. Investigation of Mediational Processes Using Parallel Process Latent Growth Curve Modeling.

    ERIC Educational Resources Information Center

    Cheong, JeeWon; MacKinnon, David P.; Khoo, Siek Toon

    2003-01-01

    Investigated a method to evaluate mediational processes using latent growth curve modeling and tested it with empirical data from a longitudinal steroid use prevention program focusing on 1,506 high school football players over 4 years. Findings suggest the usefulness of the approach. (SLD)

  11. Identification of microRNAs and Their Target Genes Explores miRNA-Mediated Regulatory Network of Cytoplasmic Male Sterility Occurrence during Anther Development in Radish (Raphanus sativus L.)

    PubMed Central

    Zhang, Wei; Xie, Yang; Xu, Liang; Wang, Yan; Zhu, Xianwen; Wang, Ronghua; Zhang, Yang; Muleke, Everlyne M.; Liu, Liwang

    2016-01-01

    MicroRNAs (miRNAs) are a type of endogenous non-coding small RNAs that play critical roles in plant growth and developmental processes. Cytoplasmic male sterility (CMS) is typically a maternally inherited trait and widely used in plant heterosis utilization. However, the miRNA-mediated regulatory network of CMS occurrence during anther development remains largely unknown in radish. In this study, a comparative small RNAome sequencing was conducted in floral buds of CMS line ‘WA’ and its maintainer line ‘WB’ by high-throughput sequencing. A total of 162 known miRNAs belonging to 25 conserved and 24 non-conserved miRNA families were isolated and 27 potential novel miRNA families were identified for the first time in floral buds of radish. Of these miRNAs, 28 known and 14 potential novel miRNAs were differentially expressed during anther development. Several target genes for CMS occurrence-related miRNAs encode important transcription factors and functional proteins, which might be involved in multiple biological processes including auxin signaling pathways, signal transduction, miRNA target silencing, floral organ development, and organellar gene expression. Moreover, the expression patterns of several CMS occurrence-related miRNAs and their targets during three stages of anther development were validated by qRT-PCR. In addition, a potential miRNA-mediated regulatory network of CMS occurrence during anther development was firstly proposed in radish. These findings could contribute new insights into complex miRNA-mediated genetic regulatory network of CMS occurrence and advance our understanding of the roles of miRNAs during CMS occurrence and microspore formation in radish and other crops. PMID:27499756

  12. Identification of microRNAs and Their Target Genes Explores miRNA-Mediated Regulatory Network of Cytoplasmic Male Sterility Occurrence during Anther Development in Radish (Raphanus sativus L.).

    PubMed

    Zhang, Wei; Xie, Yang; Xu, Liang; Wang, Yan; Zhu, Xianwen; Wang, Ronghua; Zhang, Yang; Muleke, Everlyne M; Liu, Liwang

    2016-01-01

    MicroRNAs (miRNAs) are a type of endogenous non-coding small RNAs that play critical roles in plant growth and developmental processes. Cytoplasmic male sterility (CMS) is typically a maternally inherited trait and widely used in plant heterosis utilization. However, the miRNA-mediated regulatory network of CMS occurrence during anther development remains largely unknown in radish. In this study, a comparative small RNAome sequencing was conducted in floral buds of CMS line 'WA' and its maintainer line 'WB' by high-throughput sequencing. A total of 162 known miRNAs belonging to 25 conserved and 24 non-conserved miRNA families were isolated and 27 potential novel miRNA families were identified for the first time in floral buds of radish. Of these miRNAs, 28 known and 14 potential novel miRNAs were differentially expressed during anther development. Several target genes for CMS occurrence-related miRNAs encode important transcription factors and functional proteins, which might be involved in multiple biological processes including auxin signaling pathways, signal transduction, miRNA target silencing, floral organ development, and organellar gene expression. Moreover, the expression patterns of several CMS occurrence-related miRNAs and their targets during three stages of anther development were validated by qRT-PCR. In addition, a potential miRNA-mediated regulatory network of CMS occurrence during anther development was firstly proposed in radish. These findings could contribute new insights into complex miRNA-mediated genetic regulatory network of CMS occurrence and advance our understanding of the roles of miRNAs during CMS occurrence and microspore formation in radish and other crops.

  13. MICAL-like1 mediates epidermal growth factor receptor endocytosis

    PubMed Central

    Abou-Zeid, Nancy; Pandjaitan, Rudy; Sengmanivong, Lucie; David, Violaine; Le Pavec, Gwenaelle; Salamero, Jean; Zahraoui, Ahmed

    2011-01-01

    Small GTPase Rabs are required for membrane protein sorting/delivery to precise membrane domains. Rab13 regulates epithelial tight junction assembly and polarized membrane transport. Here we report that Molecule Interacting with CasL (MICAL)-like1 (MICAL-L1) interacts with GTP-Rab13 and shares a similar domain organization with MICAL. MICAL-L1 has a calponin homology (CH), LIM, proline rich and coiled-coil domains. It is associated with late endosomes. Time-lapse video microscopy shows that green fluorescent protein–Rab7 and mcherry-MICAL-L1 are present within vesicles that move rapidly in the cytoplasm. Depletion of MICAL-L1 by short hairpin RNA does not alter the distribution of a late endosome/lysosome-associated protein but affects the trafficking of epidermal growth factor receptor (EGFR). Overexpression of MICAL-L1 leads to the accumulation of EGFR in the late endosomal compartment. In contrast, knocking down MICAL-L1 results in the distribution of internalized EGFR in vesicles spread throughout the cytoplasm and promotes its degradation. Our data suggest that the N-terminal CH domain associates with the C-terminal Rab13 binding domain (RBD) of MICAL-L1. The binding of Rab13 to RBD disrupts the CH/RBD interaction, and may induce a conformational change in MICAL-L1, promoting its activation. Our results provide novel insights into the MICAL-L1/Rab protein complex that can regulate EGFR trafficking at late endocytic pathways. PMID:21795389

  14. How, with whom and when: an overview of CD147-mediated regulatory networks influencing matrix metalloproteinase activity

    PubMed Central

    Grass, G. Daniel; Toole, Bryan P.

    2015-01-01

    Matrix metalloproteinases (MMPs) comprise a family of 23 zinc-dependent enzymes involved in various pathologic and physiologic processes. In cancer, MMPs contribute to processes from tumour initiation to establishment of distant metastases. Complex signalling and protein transport networks regulate MMP synthesis, cell surface presentation and release. Earlier attempts to disrupt MMP activity in patients have proven to be intolerable and with underwhelming clinical efficacy; thus targeting ancillary proteins that regulate MMP activity may be a useful therapeutic approach. Extracellular matrix metalloproteinase inducer (EMMPRIN) was originally characterized as a factor present on lung cancer cells, which stimulated collagenase (MMP-1) production in fibroblasts. Subsequent studies demonstrated that EMMPRIN was identical with several other protein factors, including basigin (Bsg), all of which are now commonly termed CD147. CD147 modulates the synthesis and activity of soluble and membrane-bound [membrane-type MMPs (MT-MMPs)] in various contexts via homophilic/heterophilic cell interactions, vesicular shedding or cell-autonomous processes. CD147 also participates in inflammation, nutrient and drug transporter activity, microbial pathology and developmental processes. Despite the hundreds of manuscripts demonstrating CD147-mediated MMP regulation, the molecular underpinnings governing this process have not been fully elucidated. The present review summarizes our present knowledge of the complex regulatory systems influencing CD147 biology and provides a framework to understand how CD147 may influence MMP activity. PMID:26604323

  15. How, with whom and when: an overview of CD147-mediated regulatory networks influencing matrix metalloproteinase activity.

    PubMed

    Grass, G Daniel; Toole, Bryan P

    2015-11-24

    Matrix metalloproteinases (MMPs) comprise a family of 23 zinc-dependent enzymes involved in various pathologic and physiologic processes. In cancer, MMPs contribute to processes from tumour initiation to establishment of distant metastases. Complex signalling and protein transport networks regulate MMP synthesis, cell surface presentation and release. Earlier attempts to disrupt MMP activity in patients have proven to be intolerable and with underwhelming clinical efficacy; thus targeting ancillary proteins that regulate MMP activity may be a useful therapeutic approach. Extracellular matrix metalloproteinase inducer (EMMPRIN) was originally characterized as a factor present on lung cancer cells, which stimulated collagenase (MMP-1) production in fibroblasts. Subsequent studies demonstrated that EMMPRIN was identical with several other protein factors, including basigin (Bsg), all of which are now commonly termed CD147. CD147 modulates the synthesis and activity of soluble and membrane-bound [membrane-type MMPs (MT-MMPs)] in various contexts via homophilic/heterophilic cell interactions, vesicular shedding or cell-autonomous processes. CD147 also participates in inflammation, nutrient and drug transporter activity, microbial pathology and developmental processes. Despite the hundreds of manuscripts demonstrating CD147-mediated MMP regulation, the molecular underpinnings governing this process have not been fully elucidated. The present review summarizes our present knowledge of the complex regulatory systems influencing CD147 biology and provides a framework to understand how CD147 may influence MMP activity.

  16. Structural basis for glutathione-mediated activation of the virulence regulatory protein PrfA in Listeria

    PubMed Central

    Hall, Michael; Grundström, Christin; Begum, Afshan; Lindberg, Mikael J.; Sauer, Uwe H.; Almqvist, Fredrik; Johansson, Jörgen; Sauer-Eriksson, A. Elisabeth

    2016-01-01

    Infection by the human bacterial pathogen Listeria monocytogenes is mainly controlled by the positive regulatory factor A (PrfA), a member of the Crp/Fnr family of transcriptional activators. Published data suggest that PrfA requires the binding of a cofactor for full activity, and it was recently proposed that glutathione (GSH) could fulfill this function. Here we report the crystal structures of PrfA in complex with GSH and in complex with GSH and its cognate DNA, the hly operator PrfA box motif. These structures reveal the structural basis for a GSH-mediated allosteric mode of activation of PrfA in the cytosol of the host cell. The crystal structure of PrfAWT in complex only with DNA confirms that PrfAWT can adopt a DNA binding-compatible structure without binding the GSH activator molecule. By binding to PrfA in the cytosol of the host cell, GSH induces the correct fold of the HTH motifs, thus priming the PrfA protein for DNA interaction. PMID:27930316

  17. Regulatory T cell-mediated resolution of lung injury: identification of potential target genes via expression profiling

    PubMed Central

    Aggarwal, Neil R.; D'Alessio, Franco R.; Tsushima, Kenji; Sidhaye, Venkataramana K.; Cheadle, Christopher; Grigoryev, Dmitry N.; Barnes, Kathleen C.

    2010-01-01

    In animal models of acute lung injury (ALI), gene expression studies have focused on the acute phase of illness, with little emphasis on resolution. In this study, the acute phase of intratracheal lipopolysaccharide (IT LPS)-induced lung injury was similar in wild-type (WT) and recombinase-activating gene-1-deficient (Rag-1−/−) lymphocyte-deficient mice, but resolution was impaired and resolution-phase lung gene expression remained different from baseline only in Rag-1−/− mice. By focusing on groups of genes involved in similar biological processes (gene ontologies) pertinent to inflammation and the immune response, we identified 102 genes at days 4 and 10 after IT LPS with significantly different expression between WT and Rag-1−/− mice. After adoptive transfer of isolated CD4+CD25+Foxp3+ regulatory T cells (Tregs) to Rag-1−/− mice at the time of IT LPS, resolution was similar to that in WT mice. Of the 102 genes distinctly changed in either WT or Rag-1−/− mice from our 7 gene ontologies, 19 genes reverted from the Rag-1−/− to the WT pattern of expression after adoptive transfer of Tregs, implicating those 19 genes in Treg-mediated resolution of ALI. PMID:20028937

  18. Transcriptome Profiling Reveals the Regulatory Mechanism Underlying Pollination Dependent and Parthenocarpic Fruit Set Mainly Mediated by Auxin and Gibberellin

    PubMed Central

    Tang, Ning; Deng, Wei; Hu, Guojian; Hu, Nan; Li, Zhengguo

    2015-01-01

    Background Fruit set is a key process for crop production in tomato which occurs after successful pollination and fertilization naturally. However, parthenocarpic fruit development can be uncoupled from fertilization triggered by exogenous auxin or gibberellins (GAs). Global transcriptome knowledge during fruit initiation would help to characterize the molecular mechanisms by which these two hormones regulate pollination-dependent and -independent fruit set. Principal Findings In this work, digital gene expression tag profiling (DGE) technology was applied to compare the transcriptomes from pollinated and 2, 4-D/GA3-treated ovaries. Activation of carbohydrate metabolism, cell division and expansion as well as the down-regulation of MADS-box is a comprehensive regulatory pathway during pollination-dependent and parthenocarpic fruit set. The signaling cascades of auxin and GA are significantly modulated. The feedback regulations of Aux/IAAs and DELLA genes which functioned to fine-tune auxin and GA response respectively play fundamental roles in triggering fruit initiation. In addition, auxin regulates GA synthesis via up-regulation of GA20ox1 and down-regulation of KNOX. Accordingly, the effect of auxin on fruit set is mediated by GA via ARF2 and IAA9 down-regulation, suggesting that both pollination-dependent and parthenocarpic fruit set depend on the crosstalk between auxin and GA. Significance This study characterizes the transcriptomic features of ovary development and more importantly unravels the integral roles of auxin and GA on pollination-dependent and parthenocarpic fruit set. PMID:25909657

  19. Endocrine mediators of seasonal growth in gilthead sea bream (Sparus aurata): the growth hormone and somatolactin paradigm.

    PubMed

    Mingarro, Mónica; Vega-Rubín de Celis, Silvia; Astola, Antonio; Pendón, Carlos; Valdivia, Manuel Martínez; Pérez-Sánchez, Jaume

    2002-09-01

    Regulation of somatolactin (SL) and the somatotropic axis was examined year-around at three different stocking times (spring, summer, and autumn) in a Mediterranean fish, the gilthead sea bream (Sparus aurata). The overall timing of plasma growth hormone (GH) increase was similar among trials (late spring-early summer), but the range of variation year-around was different and followed changes in food intake. Total plasma insulin-like growth factor-I primarily followed changes on growth rates, and a close positive correlation between IGF-I and thermal-unit growth coefficient (TGC) was found irrespective of fish stocking time. Thus, the activation of the somatotropic axis preceded always warm growth spurts, whereas the rise of SL in concurrence with low plasma cortisol levels was found at late autumn. This up-regulation of circulating SL titres preceded the winter inhibition of feeding, and it was more severe in big fish (spring and summer stocking times) than in small fish (autumn stocking time), growing with a relative high efficiency during the cold season despite of a severe hypertriglyceridemia and a high hepatosomatic index. These new insights provide good evidence for a different timing of GH and SL increases, and it is likely that the dominant role of SL in energy homeostasis is to be a mediator of the adaptation to fasting after replenishment of body fat stores, whereas GH and IGF-I are perceived as growth-promoting signals in times of food intake and increasing temperature and day-length.

  20. Hormone-Mediated Pattern Formation in Seedling of Plants: a Competitive Growth Dynamics Model

    NASA Astrophysics Data System (ADS)

    Kawaguchi, Satoshi; Mimura, Masayasu; Ohya, Tomoyuki; Oikawa, Noriko; Okabe, Hirotaka; Kai, Shoichi

    2001-10-01

    An ecologically relevant pattern formation process mediated by hormonal interactions among growing seedlings is modeled based on the experimental observations on the effects of indole acetic acid, which can act as an inhibitor and activator of root growth depending on its concentration. In the absence of any lateral root with constant hormone-sensitivity, the edge effect phenomenon is obtained depending on the secretion rate of hormone from the main root. Introduction of growth-stage-dependent hormone-sensitivity drastically amplifies the initial randomness, resulting in spatially irregular macroscopic patterns. When the lateral root growth is introduced, periodic patterns are obtained whose periodicity depends on the length of lateral roots. The growth-stage-dependent hormone-sensitivity and the lateral root growth are crucial for macroscopic periodic-pattern formation.

  1. Ultrasound-mediated interferon {beta} gene transfection inhibits growth of malignant melanoma

    SciTech Connect

    Yamaguchi, Kazuki; Feril, Loreto B.; Tachibana, Katsuro; Takahashi, Akira; Matsuo, Miki; Endo, Hitomi; Harada, Yoshimi; Nakayama, Juichiro

    2011-07-22

    Highlights: {yields} Successful ultrasound-mediated transfection of melanoma (C32) cells with IFN-{beta} genes both in vitro and in vivo. {yields} Ultrasound-mediated IFN-{beta} transfection inhibited proliferation of melanoma cells in vitro. {yields} Ultrasound-mediated IFN-{beta} transfection inhibited melanoma tumor growth in vivo. -- Abstract: We investigated the effects of ultrasound-mediated transfection (sonotransfection) of interferon {beta} (IFN-{beta}) gene on melanoma (C32) both in vitro and in vivo. C32 cells were sonotransfected with IFN-{beta} in vitro. Subcutaneous C32 tumors in mice were sonicated weekly immediately after intra-tumor injection with IFN-{beta} genes mixed with microbubbles. Successful sonotransfection with IFN-{beta} gene in vitro was confirmed by ELISA, which resulted in C32 growth inhibition. In vivo, the growth ratio of tumors transfected with IFN-{beta} gene was significantly lower than the other experimental groups. These results may lead to a new method of treatment against melanoma and other hard-to-treat cancers.

  2. A mitochondrial kinase complex is essential to mediate an ERK1/2-dependent phosphorylation of a key regulatory protein in steroid biosynthesis.

    PubMed

    Poderoso, Cecilia; Converso, Daniela P; Maloberti, Paula; Duarte, Alejandra; Neuman, Isabel; Galli, Soledad; Cornejo Maciel, Fabiana; Paz, Cristina; Carreras, María C; Poderoso, Juan J; Podestá, Ernesto J

    2008-01-16

    ERK1/2 is known to be involved in hormone-stimulated steroid synthesis, but its exact roles and the underlying mechanisms remain elusive. Both ERK1/2 phosphorylation and steroidogenesis may be triggered by cAMP/cAMP-dependent protein kinase (PKA)-dependent and-independent mechanisms; however, ERK1/2 activation by cAMP results in a maximal steroidogenic rate, whereas canonical activation by epidermal growth factor (EGF) does not. We demonstrate herein by Western blot analysis and confocal studies that temporal mitochondrial ERK1/2 activation is obligatory for PKA-mediated steroidogenesis in the Leydig-transformed MA-10 cell line. PKA activity leads to the phosphorylation of a constitutive mitochondrial MEK1/2 pool with a lower effect in cytosolic MEKs, while EGF allows predominant cytosolic MEK activation and nuclear pERK1/2 localization. These results would explain why PKA favors a more durable ERK1/2 activation in mitochondria than does EGF. By means of ex vivo experiments, we showed that mitochondrial maximal steroidogenesis occurred as a result of the mutual action of steroidogenic acute regulatory (StAR) protein -a key regulatory component in steroid biosynthesis-, active ERK1/2 and PKA. Our results indicate that there is an interaction between mitochondrial StAR and ERK1/2, involving a D domain with sequential basic-hydrophobic motifs similar to ERK substrates. As a result of this binding and only in the presence of cholesterol, ERK1/2 phosphorylates StAR at Ser(232). Directed mutagenesis of Ser(232) to a non-phosphorylable amino acid such as Ala (StAR S232A) inhibited in vitro StAR phosphorylation by active ERK1/2. Transient transfection of MA-10 cells with StAR S232A markedly reduced the yield of progesterone production. In summary, here we show that StAR is a novel substrate of ERK1/2, and that mitochondrial ERK1/2 is part of a multimeric protein kinase complex that regulates cholesterol transport. The role of MAPKs in mitochondrial function is underlined.

  3. Parasite-mediated growth patterns and nutritional constraints in a cavity-nesting bird.

    PubMed

    O'Brien, Erin L; Dawson, Russell D

    2008-01-01

    1. Trade-offs between growth and immunity of nestling birds can be influenced by parasites, but the magnitude of these effects may depend on availability of critical dietary nutrients. Owing to their importance for both immune system function and growth, dietary carotenoids have the potential to mediate parasite-induced developmental strategies of avian hosts. 2. The effects of ectoparasitic blow flies Protocalliphora spp. and dietary carotenoids (lutein and zeaxanthin) on immune function and patterns of growth in nestling mountain bluebirds Sialia currucoides were investigated by combining parasite removal and carotenoid supplementation treatments in a 2 x 2 design. 3. Supplemental carotenoids enhanced nestlings' T-cell-mediated immune response following intradermal injection of phytohaemagglutinin. 4. The effect of carotenoid supplementation on rate of mass gain depended on whether broods were exposed to parasites: among parasitized broods, those receiving supplemental carotenoids gained mass more rapidly than nonsupplemented broods, whereas there was no effect of supplemental carotenoids on growth of mass in broods that had parasites removed. This suggests that additional dietary carotenoids allowed nestlings to compensate for the otherwise detrimental effects of parasites on mass gain. For length of the eighth primary feather at fledging, early and late broods differed in their response to parasitism: early broods showed an increase in feather length when parasites were removed, while nestlings in late broods had shorter feathers in the absence of parasites. We suggest that this may reflect within-season variation in parasite-mediated growth strategies of nestlings. 5. Maternal condition was positively associated with mass, condition and rate of feather growth of offspring under all conditions, and also influenced nestling immunocompetence, but only in the absence of parasites. 6. We conclude that dietary carotenoids alleviate some of the detrimental effects of

  4. Control of liver size by RNAi-mediated multiplex knockdown and its application for discovery of regulatory mechanisms

    PubMed Central

    Yin, Hao; Bogorad, Roman L.; Barnes, Carmen; Walsha, Stephen; Zhuang, Iris; Nonaka, Hidenori; Ruda, Vera; Kuchimanchi, Satya; Nechev, Lubomir; Akinc, Akin; Xue, Wen; Zerial, Marino; Langer, Robert; Anderson, Daniel G.; Koteliansky, Victor

    2017-01-01

    Background and aims The Hippo pathway controls organ size through a negative regulation of the transcription co-activator Yap1. The overexpression of hyperactive mutant Yap1 or deletion of key components in the Hippo pathway leads to increased organ size in different species. Analysis of interactions of this pathway with other cellular signals corroborating organ size control is limited in part due to the difficulties associated with development of rodent models. Methods Here, we develop a new model of reversible induction of the liver size in mice using siRNA-nanoparticles targeting two kinases of Hippo pathway, namely, mammalian Ste20 family kinases 1 and 2 (Mst1 and Mst2), and an upstream regulator, neurofibromatosis type II (NF2). Results The triple siRNAs nanoparticle-induced hepatomegaly in mice phenocopies one observed with Mst1-/- Mst2-/- liver-specific depletion, as shown by extensive proliferation of hepatocytes and activation of Yap1. The simultaneous co-treatment with a fourth siRNA nanoparticle against Yap1 fully blocked the liver growth. Hippo pathway-induced liver enlargement is associated with p53 activation, evidenced by its accumulation in the nuclei and upregulation of its target genes. Moreover, injections of the triple siRNAs nanoparticle in p53LSL/LSL mice shows that livers lacking p53 expression grow faster and exceed the size of livers in p53 wild type animals, indicating a role of p53 in controlling Yap1-induced liver growth. Conclusion Our data show that siRNA-nanoparticulate manipulation of gene expression can provide the reversible control of organ size in adult animals, which presents a new avenue for the investigation of complex regulatory networks in liver. PMID:26658687

  5. Preservation of Gene Duplication Increases the Regulatory Spectrum of Ribosomal Protein Genes and Enhances Growth under Stress.

    PubMed

    Parenteau, Julie; Lavoie, Mathieu; Catala, Mathieu; Malik-Ghulam, Mustafa; Gagnon, Jules; Abou Elela, Sherif

    2015-12-22

    In baker's yeast, the majority of ribosomal protein genes (RPGs) are duplicated, and it was recently proposed that such duplications are preserved via the functional specialization of the duplicated genes. However, the origin and nature of duplicated RPGs' (dRPGs) functional specificity remain unclear. In this study, we show that differences in dRPG functions are generated by variations in the modality of gene expression and, to a lesser extent, by protein sequence. Analysis of the sequence and expression patterns of non-intron-containing RPGs indicates that each dRPG is controlled by specific regulatory sequences modulating its expression levels in response to changing growth conditions. Homogenization of dRPG sequences reduces cell tolerance to growth under stress without changing the number of expressed genes. Together, the data reveal a model where duplicated genes provide a means for modulating the expression of ribosomal proteins in response to stress.

  6. A Novel Pregnane X Receptor-mediated and Sterol Regulatory Element-binding Protein-independent Lipogenic Pathway*

    PubMed Central

    Zhou, Jie; Zhai, Yonggong; Mu, Ying; Gong, Haibiao; Uppal, Hirdesh; Toma, David; Ren, Songrong; Evans, Ronald M.; Xie, Wen

    2014-01-01

    The pregnane X receptor (PXR) was isolated as a xenosensor regulating xenobiotic responses. In this study, we show that PXR plays an endobiotic role by impacting lipid homeostasis. Expression of an activated PXR in the livers of transgenic mice resulted in an increased hepatic deposit of triglycerides. This PXR-mediated lipid accumulation was independent of the activation of the lipogenic transcriptional factor SREBP-1c (sterol regulatory element-binding protein 1c) and its primary lipogenic target enzymes, including fatty-acid synthase (FAS) and acetyl-CoA carboxylase 1 (ACC-1). Instead, the lipid accumulation in transgenic mice was associated with an increased expression of the free fatty acid transporter CD36 and several accessory lipogenic enzymes, such as stearoyl-CoA desaturase-1 (SCD-1) and long chain free fatty acid elongase. Studies using transgenic and knock-out mice showed that PXR is both necessary and sufficient for Cd36 activation. Promoter analyses revealed a DR-3-type of PXR-response element in the mouse Cd36 promoter, establishing Cd36 as a direct transcriptional target of PXR. The hepatic lipid accumulation and Cd36 induction were also seen in the hPXR “humanized” mice treated with the hPXR agonist rifampicin. The activation of PXR was also associated with an inhibition of pro-β-oxidative genes, such as peroxisome proliferator-activated receptor α (PPARα) and thiolase, and an up-regulation of PPARγ, a positive regulator of CD36. The cross-regulation of CD36 by PXR and PPARγ suggests that this fatty acid transporter may function as a common target of orphan nuclear receptors in their regulation of lipid homeostasis. PMID:16556603

  7. Regulatory T Cells Promote β-Catenin–Mediated Epithelium-to-Mesenchyme Transition During Radiation-Induced Pulmonary Fibrosis

    SciTech Connect

    Xiong, Shanshan; Pan, Xiujie; Xu, Long; Yang, Zhihua; Guo, Renfeng; Gu, Yongqing; Li, Ruoxi; Wang, Qianjun; Xiao, Fengjun; Du, Li; Zhou, Pingkun; Zhu, Maoxiang

    2015-10-01

    Purpose: Radiation-induced pulmonary fibrosis results from thoracic radiation therapy and severely limits radiation therapy approaches. CD4{sup +}CD25{sup +}FoxP3{sup +} regulatory T cells (Tregs) as well as epithelium-to-mesenchyme transition (EMT) cells are involved in pulmonary fibrosis induced by multiple factors. However, the mechanisms of Tregs and EMT cells in irradiation-induced pulmonary fibrosis remain unclear. In the present study, we investigated the influence of Tregs on EMT in radiation-induced pulmonary fibrosis. Methods and Materials: Mice thoraxes were irradiated (20 Gy), and Tregs were depleted by intraperitoneal injection of a monoclonal anti-CD25 antibody 2 hours after irradiation and every 7 days thereafter. Mice were treated on days 3, 7, and 14 and 1, 3, and 6 months post irradiation. The effectiveness of Treg depletion was assayed via flow cytometry. EMT and β-catenin in lung tissues were detected by immunohistochemistry. Tregs isolated from murine spleens were cultured with mouse lung epithelial (MLE) 12 cells, and short interfering RNA (siRNA) knockdown of β-catenin in MLE 12 cells was used to explore the effects of Tregs on EMT and β-catenin via flow cytometry and Western blotting. Results: Anti-CD25 antibody treatment depleted Tregs efficiently, attenuated the process of radiation-induced pulmonary fibrosis, hindered EMT, and reduced β-catenin accumulation in lung epithelial cells in vivo. The coculture of Tregs with irradiated MLE 12 cells showed that Tregs could promote EMT in MLE 12 cells and that the effect of Tregs on EMT was partially abrogated by β-catenin knockdown in vitro. Conclusions: Tregs can promote EMT in accelerating radiation-induced pulmonary fibrosis. This process is partially mediated through β-catenin. Our study suggests a new mechanism for EMT, promoted by Tregs, that accelerates radiation-induced pulmonary fibrosis.

  8. Simulating molecular mechanisms of the MDM2-mediated regulatory interactions: a conformational selection model of the MDM2 lid dynamics.

    PubMed

    Verkhivker, Gennady M

    2012-01-01

    Diversity and complexity of MDM2 mechanisms govern its principal function as the cellular antagonist of the p53 tumor suppressor. Structural and biophysical studies have demonstrated that MDM2 binding could be regulated by the dynamics of a pseudo-substrate lid motif. However, these experiments and subsequent computational studies have produced conflicting mechanistic models of MDM2 function and dynamics. We propose a unifying conformational selection model that can reconcile experimental findings and reveal a fundamental role of the lid as a dynamic regulator of MDM2-mediated binding. In this work, structure, dynamics and energetics of apo-MDM2 are studied as a function of posttranslational modifications and length of the lid. We found that the dynamic equilibrium between "closed" and "semi-closed" lid forms may be a fundamental characteristic of MDM2 regulatory interactions, which can be modulated by phosphorylation, phosphomimetic mutation as well as by the lid size. Our results revealed that these factors may regulate p53-MDM2 binding by fine-tuning the thermodynamic equilibrium between preexisting conformational states of apo-MDM2. In agreement with NMR studies, the effect of phosphorylation on MDM2 interactions was more pronounced with the truncated lid variant that favored the thermodynamically dominant closed form. The phosphomimetic mutation S17D may alter the lid dynamics by shifting the thermodynamic equilibrium towards the ensemble of "semi-closed" conformations. The dominant "semi-closed" lid form and weakened dependence on the phosphorylation seen in simulations with the complete lid can provide a rationale for binding of small p53-based mimetics and inhibitors without a direct competition with the lid dynamics. The results suggested that a conformational selection model of preexisting MDM2 states may provide a robust theoretical framework for understanding MDM2 dynamics. Probing biological functions and mechanisms of MDM2 regulation would require

  9. Effects of transforming growth factor beta-1 on growth-regulatory genes in tumour-derived human oral keratinocytes.

    PubMed Central

    Paterson, I. C.; Patel, V.; Sandy, J. R.; Prime, S. S.; Yeudall, W. A.

    1995-01-01

    This study examined the effect of transforming growth factor beta-1 (TGF-beta 1) on c-myc, RB1, junB and p53 expression together with pRb phosphorylation, in carcinoma-derived and normal human oral keratinocytes with a range of inhibitory responses to this ligand. Amplification of c-myc was observed in eight of eight tumour-derived cell lines and resulted in corresponding mRNA expression. The down-regulation of c-myc expression by TGF-beta 1 predominantly reflected growth inhibition by TGF-beta 1, but in two of eight tumour-derived cell lines which were partially responsive to TGF-beta 1 c-myc expression was unaltered by this ligand. While RB1 mRNA levels were unaltered by TGF-beta 1, the ligand caused the accumulation of the underphosphorylated form of the Rb protein in all cells irrespective of TGF-beta 1-induced growth arrest. junB expression was up-regulated by TGF-beta 1 in cells with a range of growth inhibitory responses. All cells contained mutant p53. TGF-beta 1 did not affect p53 mRNA expression in both tumour-derived and normal keratinocytes and there was no alteration in p53 protein levels in keratinocytes expressing stable p53 protein following TGF-beta 1 treatment. The data indicate that TGF-beta-induced growth control can exist independently of the presence of mutant p53 and the control of Rb phosphorylation and c-myc down-regulation. It may be that TGF-beta growth inhibition occurs via multiple mechanisms and that the loss of one pathway during tumour progression does not necessarily result in the abrogation of TGF-beta-induced growth control. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 PMID:7547241

  10. Scaling of submonolayer island growth with reversible adatom exchange in surfactant-mediated epitaxy

    NASA Astrophysics Data System (ADS)

    Wang, Daimu; Wang, Zhuping; Zhu, Hui

    2007-08-01

    Surfactant-mediated epitaxial growth is studied with a realistic model, which includes three main kinetic processes: diffusion of adatoms on the surfactant terrace, exchange of adatoms with their underneath surfactant atoms, and reexchange in which an exchanged adatom resurfaces to the top of the surfactant layer. The scaling behavior of nucleus density and island size distributions in the initial stage of growth is investigated by using kinetic Monte Carlo simulations. The results show that the temperature dependence of nucleus density and island size distributions governed by the reexchanging-controlled nucleation at high temperatures exhibits similar scaling behavior to that obtained by the standard diffusion-mediated nucleation at low temperatures. However, at intermediate temperatures, the exchanging-controlled nucleation leads to an increase of nucleus density with temperature, while the island size distribution scales to a monotonically decreasing function, showing nonstandard scaling behavior.

  11. Nucleation Behaviour in the Initial Stage of Surfactant-Mediated Epitaxial Growth

    NASA Astrophysics Data System (ADS)

    Wang, Dai-Mu; Sun, Xia; Ding, Ze-Jun; Wu, Zi-Qin

    2004-10-01

    The nucleation kinetics in the early stage of epitaxial growth mediated by a monolayer of surfactant is studied by using kinetic Monte Carlo simulations. Our simulation model includes three main kinetic parameters: a small barrier for adatom diffusion on the surfactant terrace, a higher barrier for the exchange of adatoms with their underneath surfactant atoms, and a highest barrier for the recovery exchange in which an exchanged adatom resurfaces to the top of the surfactant layer. The simulations reveal a distinct transition of nucleation behaviour as the different atomic processes are activated successively with increasing temperature. The total nucleus density as a function of temperature exhibits a complex N-shape with a minimum and a maximum, which define the transition temperatures. The characteristic behaviour of nucleation density is helpful to rationalize the experimental observations on the temperature dependence of growth mode in some surfactant-mediated epitaxial systems.

  12. Oncogene N-ras mediates selective inhibition of c-fos induction by nerve growth factor and basic fibroblast growth factor in a PC12 cell line.

    PubMed Central

    Thomson, T M; Green, S H; Trotta, R J; Burstein, D E; Pellicer, A

    1990-01-01

    A cell line was generated from U7 cells (a subline of PC12 rat pheochromocytoma cells) that contains a stably integrated transforming mouse N-ras (Lys-61) gene under the control of the long terminal repeat from mouse mammary tumor virus. Such cells, designated UR61, undergo neuronal differentiation upon exposure to nanomolar concentrations of dexamethasone, as a consequence of expression of the activated N-ras gene (I. Guerrero, A. Pellicer, and D.E. Burstein, Biochem, Biophys. Res. Commun. 150:1185-1192, 1988). Exposure of UR61 cells to either nerve growth factor (NGF) or basic fibroblast growth factor (bFGF) results in a marked induction of c-fos RNA, with kinetics paralleling those of NGF- or bFGF-induced expression of c-fos RNA in PC12 cells. Dexamethasone-induced expression of activated N-ras p21 results in blocking of c-fos RNA induction by NGF or bFGF in a time-dependent manner. Activated N-ras p21-mediated inhibition of c-fos RNA induction in UR61 cells is selective for NGF and bFGF and is not due to selective degradation of c-fos RNA. Normal and transforming N-ras can trans activate the chloramphenicol acetyltransferase gene linked to mouse c-fos regulatory sequences when transient expression assays are performed. Our observations suggest that N-ras p21 selectively interacts with pathways involved in induction of c-fos expression which initiate at the receptors for NGF and bFGF. Images PMID:2108319

  13. Targeting ASCT2-mediated glutamine uptake blocks prostate cancer growth and tumour development.

    PubMed

    Wang, Qian; Hardie, Rae-Anne; Hoy, Andrew J; van Geldermalsen, Michelle; Gao, Dadi; Fazli, Ladan; Sadowski, Martin C; Balaban, Seher; Schreuder, Mark; Nagarajah, Rajini; Wong, Justin J-L; Metierre, Cynthia; Pinello, Natalia; Otte, Nicholas J; Lehman, Melanie L; Gleave, Martin; Nelson, Colleen C; Bailey, Charles G; Ritchie, William; Rasko, John E J; Holst, Jeff

    2015-07-01

    Glutamine is conditionally essential in cancer cells, being utilized as a carbon and nitrogen source for macromolecule production, as well as for anaplerotic reactions fuelling the tricarboxylic acid (TCA) cycle. In this study, we demonstrated that the glutamine transporter ASCT2 (SLC1A5) is highly expressed in prostate cancer patient samples. Using LNCaP and PC-3 prostate cancer cell lines, we showed that chemical or shRNA-mediated inhibition of ASCT2 function in vitro decreases glutamine uptake, cell cycle progression through E2F transcription factors, mTORC1 pathway activation and cell growth. Chemical inhibition also reduces basal oxygen consumption and fatty acid synthesis, showing that downstream metabolic function is reliant on ASCT2-mediated glutamine uptake. Furthermore, shRNA knockdown of ASCT2 in PC-3 cell xenografts significantly inhibits tumour growth and metastasis in vivo, associated with the down-regulation of E2F cell cycle pathway proteins. In conclusion, ASCT2-mediated glutamine uptake is essential for multiple pathways regulating the cell cycle and cell growth, and is therefore a putative therapeutic target in prostate cancer.

  14. TOC1–PIF4 interaction mediates the circadian gating of thermoresponsive growth in Arabidopsis

    PubMed Central

    Zhu, Jia-Ying; Oh, Eunkyoo; Wang, Tina; Wang, Zhi-Yong

    2016-01-01

    Arabidopsis adapts to elevated temperature by promoting stem elongation and hyponastic growth through a temperature-responsive transcription factor PIF4. Here we show that the evening-expressed clock component TOC1 interacts with and inactivates PIF4, thereby suppressing thermoresponsive growth in the evening. We find that the expression of PIF4 target genes show circadian rhythms of thermosensitivity, with minimum responsiveness in the evening when TOC1 level is high. Loss of function of TOC1 and its close homologue PRR5 restores thermosensitivity in the evening, whereas TOC1 overexpression causes thermo insensitivity, demonstrating that TOC1 mediates the evening-specific inhibition of thermoresponses. We further show that PIF4 is required for thermoadaptation mediated by moderately elevated temperature. Our results demonstrate that the interaction between TOC1 and PIF4 mediates the circadian gating of thermoresponsive growth, which may serve to increase fitness by matching thermoresponsiveness with the day–night cycles of fluctuating temperature and light conditions. PMID:27966533

  15. Different receptors mediate the electrophysiological and growth cone responses of an identified neuron to applied dopamine.

    PubMed

    Dobson, K S; Dmetrichuk, J M; Spencer, G E

    2006-09-15

    Neurotransmitters are among the many cues that may guide developing axons toward appropriate targets in the developing nervous system. We have previously shown in the mollusk Lymnaea stagnalis that dopamine, released from an identified pre-synaptic cell, differentially affects growth cone behavior of its target and non-target cells in vitro. Here, we describe a group of non-target cells that also produce an inhibitory electrophysiological response to applied dopamine. We first determined, using pharmacological blockers, which receptors mediate this physiological response. We demonstrated that the dopaminergic electrophysiological responses of non-target cells were sensitive to a D2 receptor antagonist, as are known target cell responses. However, the non-target cell receptors were linked to different G-proteins and intracellular signaling pathways than the target cell receptors. Despite the presence of a D2-like receptor at the soma, the growth cone collapse of these non-target cells was mediated by D1-like receptors. This study shows that different dopamine receptor sub-types mediated the inhibitory physiological and growth cone responses of an identified cell type. We therefore not only provide further evidence that D2- and D1-like receptors can be present on the same neuron in invertebrates, but also show that these receptors are likely involved in very different cellular functions.

  16. New perspectives in cyclic AMP-mediated axon growth and guidance: The emerging epoch of Epac.

    PubMed

    Peace, Andrew G; Shewan, Derryck A

    2011-03-10

    In the search for a cure to brain and spinal cord injury much has been learned about the inhibitory environment of the central nervous system (CNS), and yet a clinical therapy remains elusive. In recent years great advances have been made in understanding intracellular molecular mechanisms that transduce cell surface receptor-mediated signals that neurons receive from their environment. Many of these signalling pathways share common mechanisms, which presents the possibility that manipulating activities of key cell signalling molecules such as those regulated by 3'-5'-cyclic adenosine monophosphate (cAMP) might allow axons to simultaneously overcome the inhibitory effects of a number of extracellular ligands. The identification of Epac, a novel direct intracellular target for cAMP, has opened up a new avenue of research that is beginning to explain how cAMP can mediate a range of neuronal functions including distinct axon growth and guidance decisions. With current research tools that allow more specific activation of proteins or knock-down of their expression, as well as quantitation of protein activities in live cells, it is already becoming clear that Epac plays highly important roles in the development and function of the nervous system. Here, we focus on emerging evidence that Epac mediates cAMP-regulated axon growth and chemoattraction, and thus represents a novel target for overcoming axon growth inhibition and promoting CNS regeneration.

  17. Unlabeled milk from cows treated with biosynthetic growth hormones: a case of regulatory abdication.

    PubMed

    Epstein, S S

    1996-01-01

    Levels of insulin-like growth factor-1 (IGF-1) are substantially elevated and more bioactive in the milk of cows hyperstimulated with the biosynthetic bovine growth hormones rBGH, and are further increased by pasteurization. IGF-1 is absorbed from the gastrointestinal tract, as evidenced by marked growth-promoting effects even in short-term tests in mature rats, and absorption is likely to be still higher in infants. Converging lines of evidence incriminate IGF-1 in rBGH milk as a potential risk factor for both breast and gastrointestinal cancers.

  18. Adiponectin regulate growth hormone secretion via adiponectin receptor mediated Ca(2+) signalling in rat somatotrophs in vitro.

    PubMed

    Steyn, F J; Boehme, F; Vargas, E; Wang, K; Parkington, H C; Rao, J R; Chen, C

    2009-08-01

    Obesity is associated with reduced levels of growth hormone (GH) and the disruption of pulsatile GH secretion. This results in relative GH deficiency. It is likely that a regulatory relationship between GH secretion and adipose tissue exists as the secretion of GH recovers to normal levels after a reduction in body weight. This report characterise the expression and interaction of adiponectin receptors 1 and 2 (AdipoR1 and AdipoR2) and adiponectin, respectively, in regulating the activity of GH secreting cells. Polymerase chain reaction analysis of the GH3 cell line, rat anterior pituitary gland and isolated somatotroph cells from transgenic GFP expressing mice confirmed the expression of both AdipoR1 and AdipoR2 in GH secretory cells. Because GH cells expressed both receptors, it is likely that the measured increase in GH secretion, observed in primary cultured rat pituitary cells after 30 min of incubation with full-length murine adiponectin, was mediated by a direct receptor regulated process. Adiponectin induced an increase in intracellular Ca(2+) through both the influx of extracellular Ca(2+) and the release of intracellular Ca(2+) stores resulting in the secretion of GH. Furthermore, results confirm that this increase in GH secretion depended mainly on an increase in Ca(2+) influx through L-type Ca(2+) channels. It is concluded that adiponectin directly regulates GH secretion from somatotrophs by binding to either adiponectin receptor, and that this is mediated via a similar process observed after the stimulation of GH secretion by GH-releasing hormone.

  19. Screening for feeding deterrent and insect growth regulatory activity of triterpenic saponins from Diploknema butyracea and Sapindus mukorossi.

    PubMed

    Saha, Supradip; Walia, Suresh; Kumar, Jitendra; Dhingra, Swaran; Parmar, Balraj S

    2010-01-13

    Antifeeding and insect growth regulatory effects of saponins and its hydrolyzed products from Diploknema butyracea and Sapindus mukorossi on the insect pest Spodoptera litura (F.) were investigated in the laboratory. D. butyracea saponins as well as their hydrolyzed prosapogenins were found to be better biologically active in controlling pests. A concentration of 1200 and 3400 mg L(-1) alkaline and acid hydrolyzed D. butyracea saponins exhibited significant antifeeding and toxic effects to third instar larvae when compared to the emulsified water as control. The n-BuOH extract after prep-HPLC separation provided two saponins from the D. butyracea saponin mixture: 3-O-[beta-D-glucopyarnosyl-beta-d-glucopyranosyl]-16-alpha-hydroxyprotobassic acid-28-O-[ara-glc-xyl]-ara (MI-I) and 3-O-beta-D-glucopyranosyl-glucopyranosyl-glucopyranosyl-16-alpha-hydroxyprotobassic acid-28-O-[ara-xyl-ara]-apiose (MI-III). The single saponin extracted from the S. mukorossi saponin mixture was 3-O-[beta-D-xyl(OAc).beta-D-arabinopyranosyl.beta-D-rhamnopyranosyl] hederagenin-28-O-[beta-D-glc.beta-D-glc.beta-D-rhamnopyranosyl] ester (SM-I). Five days after saponin treatment on larvae, the growth index (GI50) was reduced from 0.92% to 1520 ppm in alkaline hydrolyzed D. butyracea saponins. Upon hydrolysis, growth regulatory activity was improved in S. mukorossi saponin, whereas very little difference was found in antifeedant activity. Hydrophile-lipophile balance is important for the proper functioning of saponin/prosapogenin/sapogenin, which could be achieved by manipulating the sugar molecule in the triterpenic skeleton.

  20. Trauma or growth after a natural disaster? The mediating role of rumination processes

    PubMed Central

    García, Felipe E.; Cova, Félix; Rincón, Paulina; Vázquez, Carmelo

    2015-01-01

    The aim of this study was to test a cognitive model of posttraumatic symptoms (PTS) and posttraumatic growth (PTG) after exposure to a natural disaster. It was hypothesized that although subjective severity of trauma would be related to the severity of PTS, this relation would be mediated by brooding and cognitive strategies related to the presence of repetitive negative content in thoughts. Furthermore, the relation between severity and PTG would be fully mediated by deliberate rumination (DR), cognitive strategies related to conscious efforts focused on handling the event. To evaluate the cognitive model, adults (N=351) who lost their homes as a result of the earthquake and tsunami that occurred in Chile on February 27, 2010, were selected. Structural equation modeling was used to analyze the data. The resulting model had adequate indices of goodness adjustment and showed that brooding completely mediated the relation between subjective severity and PTS, and DR completely mediated the relation between subjective severity, brooding, and PTG. These results highlight the role of both the content and process of rumination in mediating the association between subjective severity of trauma, PTS, and PTG. The implications of these results for a more comprehensive model of symptom severity that occurs after trauma are discussed. PMID:26234365

  1. ANGUSTIFOLIA mediates one of the multiple SCRAMBLED signaling pathways regulating cell growth pattern in Arabidopsis thaliana.

    PubMed

    Kwak, Su-Hwan; Song, Sang-Kee; Lee, Myeong Min; Schiefelbein, John

    2015-09-25

    In Arabidopsis thaliana, an atypical leucine-rich repeat receptor-like kinase, SCRAMBLED (SCM), is required for multiple developmental processes including root epidermal cell fate determination, silique dehiscence, inflorescence growth, ovule morphogenesis, and tissue morphology. Previous work suggested that SCM regulates these multiple pathways using distinct mechanisms via interactions with specific downstream factors. ANGUSTIFOLIA (AN) is known to regulate cell and tissue morphogenesis by influencing cortical microtubule arrangement, and recently, the AN protein was reported to interact with the SCM protein. Therefore, we examined whether AN might be responsible for mediating some of the SCM-dependent phenotypes. We discovered that both scm and an mutant lines cause an abnormal spiral or twisting growth of roots, but only the scm mutant affected root epidermal patterning. The siliques of the an and scm mutants also exhibited spiral growth, as previously reported, but only the scm mutant altered silique dehiscence. Interestingly, we discovered that the spiral growth of roots and siliques of the scm mutant is rescued by a truncated SCM protein that lacks its kinase domain, and that a juxtamembrane domain of SCM was sufficient for AN binding in the yeast two-hybrid analysis. These results suggest that the AN protein is one of the critical downstream factors of SCM pathways specifically responsible for mediating its effects on cell/tissue morphogenesis through cortical microtubule arrangement.

  2. Redox signalling to nuclear regulatory proteins by reactive oxygen species contributes to oestrogen-induced growth of breast cancer cells

    PubMed Central

    Okoh, V O; Garba, N A; Penney, R B; Das, J; Deoraj, A; Singh, K P; Sarkar, S; Felty, Q; Yoo, C; Jackson, R M; Roy, D

    2015-01-01

    Background: 17β-Oestradiol (E2)-induced reactive oxygen species (ROS) have been implicated in regulating the growth of breast cancer cells. However, the underlying mechanism of this is not clear. Here we show how ROS through a novel redox signalling pathway involving nuclear respiratory factor-1 (NRF-1) and p27 contribute to E2-induced growth of MCF-7 breast cancer cells. Methods: Chromatin immunoprecipitation, qPCR, mass spectrometry, redox western blot, colony formation, cell proliferation, ROS assay, and immunofluorescence microscopy were used to study the role of NRF-1. Results: The major novel finding of this study is the demonstration of oxidative modification of phosphatases PTEN and CDC25A by E2-generated ROS along with the subsequent activation of AKT and ERK pathways that culminated in the activation of NRF-1 leading to the upregulation of cell cycle genes. 17β-Oestradiol-induced ROS by influencing nuclear proteins p27 and Jab1 also contributed to the growth of MCF-7 cells. Conclusions: Taken together, our results present evidence in the support of E2-induced ROS-mediated AKT signalling leading to the activation of NRF-1-regulated cell cycle genes as well as the impairment of p27 activity, which is presumably necessary for the growth of MCF-7 cells. These observations are important because they provide a new paradigm by which oestrogen may contribute to the growth of breast cancer. PMID:25965299

  3. MDP25, a novel calcium regulatory protein, mediates hypocotyl cell elongation by destabilizing cortical microtubules in Arabidopsis.

    PubMed

    Li, Jiejie; Wang, Xianling; Qin, Tao; Zhang, Yan; Liu, Xiaomin; Sun, Jingbo; Zhou, Yuan; Zhu, Lei; Zhang, Ziding; Yuan, Ming; Mao, Tonglin

    2011-12-01

    The regulation of hypocotyl elongation is important for plant growth. Microtubules play a crucial role during hypocotyl cell elongation. However, the molecular mechanism underlying this process is not well understood. In this study, we describe a novel Arabidopsis thaliana microtubule-destabilizing protein 25 (MDP25) as a negative regulator of hypocotyl cell elongation. We found that MDP25 directly bound to and destabilized microtubules to enhance microtubule depolymerization in vitro. The seedlings of mdp25 mutant Arabidopsis lines had longer etiolated hypocotyls. In addition, MDP25 overexpression resulted in significant overall shortening of hypocotyl cells, which exhibited destabilized cortical microtubules and abnormal cortical microtubule orientation, suggesting that MDP25 plays a crucial role in the negative regulation of hypocotyl cell elongation. Although MDP25 localized to the plasma membrane under normal conditions, increased calcium levels in cells caused MDP25 to partially dissociate from the plasma membrane and move into the cytosol. Cellular MDP25 bound to and destabilized cortical microtubules, resulting in their reorientation, and subsequently inhibited hypocotyl cell elongation. Our results suggest that MDP25 exerts its function on cortical microtubules by responding to cytoplasmic calcium levels to mediate hypocotyl cell elongation.

  4. Characterization of a two-component regulatory system that regulates succinate-mediated catabolite repression in Sinorhizobium meliloti.

    PubMed

    Garcia, Preston P; Bringhurst, Ryan M; Arango Pinedo, Catalina; Gage, Daniel J

    2010-11-01

    When they are available, Sinorhizobium meliloti utilizes C(4)-dicarboxylic acids as preferred carbon sources for growth while suppressing the utilization of some secondary carbon sources such as α- and β-galactosides. The phenomenon of using succinate as the sole carbon source in the presence of secondary carbon sources is termed succinate-mediated catabolite repression (SMCR). Genetic screening identified the gene sma0113 as needed for strong SMCR when S. meliloti was grown in succinate plus lactose, maltose, or raffinose. sma0113 and the gene immediately downstream, sma0114, encode the proteins Sma0113, an HWE histidine kinase with five PAS domains, and Sma0114, a CheY-like response regulator lacking a DNA-binding domain. sma0113 in-frame deletion mutants show a relief of catabolite repression compared to the wild type. sma0114 in-frame deletion mutants overproduce polyhydroxybutyrate (PHB), and this overproduction requires sma0113. Sma0113 may use its five PAS domains for redox level or energy state monitoring and use that information to regulate catabolite repression and related responses.

  5. Helicobacter pylori-Mediated Protection from Allergy Is Associated with IL-10-Secreting Peripheral Blood Regulatory T Cells

    PubMed Central

    Hussain, Khiyam; Letley, Darren P.; Greenaway, A. Borgel; Kenefeck, Rupert; Winter, Jody A.; Tomlinson, William; Rhead, Joanne; Staples, Emily; Kaneko, Kazuyo; Atherton, John C.; Robinson, Karen

    2016-01-01

    Helicobacter pylori infections are usually established in early childhood and continuously stimulate immunity, including T-helper 1 (Th1), Th17, and regulatory T-cell (Treg) responses, throughout life. Although known to be the major cause of peptic ulcer disease and gastric cancer, disease occurs in a minority of those who are infected. Recently, there has been much interest in beneficial effects arising from infection with this pathogen. Published data robustly show that the infection is protective against asthma in mouse models. Epidemiological studies show that H. pylori is inversely associated with human allergy and asthma, but there is a paucity of mechanistic data to explain this. Since Th1 and Treg responses are reported to protect against allergic responses, we investigated if there were links between the human systemic Th1 and Treg response to H. pylori and allergen-specific IgE levels. The human cytokine and T-cell responses were examined using peripheral blood mononuclear cells (PBMCs) from 49 infected and 58 uninfected adult patients. Concentrations of total and allergen-specific plasma IgE were determined by ELISA and ImmunoCAP assays. These responses were analyzed according to major virulence factor genotypes of the patients’ colonizing H. pylori strains. An in vitro assay was employed, using PBMCs from infected and uninfected donors, to determine the role of Treg cytokines in the suppression of IgE. Significantly higher frequencies of IL-10-secreting CD4+CD25hi Tregs, but not H. pylori-specific Th1 cells, were present in the peripheral blood of infected patients. Total and allergen-specific IgE concentrations were lower when there was a strong Treg response, and blocking IL-10 in vitro dramatically restored IgE responses. IgE concentrations were also significantly lower when patients were infected with CagA+ strains or those expressing the more active i1 form of VacA. The systemic IL-10+ Treg response is therefore likely to play a role in H. pylori-mediated

  6. Interleukin 10 and dendritic cells are the main suppression mediators of regulatory T cells in human neurocysticercosis.

    PubMed

    Arce-Sillas, A; Álvarez-Luquín, D D; Cárdenas, G; Casanova-Hernández, D; Fragoso, G; Hernández, M; Proaño Narváez, J V; García-Vázquez, F; Fleury, A; Sciutto, E; Adalid-Peralta, L

    2016-02-01

    Neurocysticercosis is caused by the establishment of Taenia solium cysticerci in the central nervous system. It is considered that, during co-evolution, the parasite developed strategies to modulate the host's immune response. The action mechanisms of regulatory T cells in controlling the immune response in neurocysticercosis are studied in this work. Higher blood levels of regulatory T cells with CD4(+) CD45RO(+) forkhead box protein 3 (FoxP3)(high) and CD4(+) CD25(high) FoxP3(+) CD95(high) phenotype and of non-regulatory CD4(+) CD45RO(+) FoxP3(med) T cells were found in neurocysticercosis patients with respect to controls. Interestingly, regulatory T cells express higher levels of cytotoxic T lymphocyte antigen 4 (CTLA-4), lymphocyte-activation gene 3 (LAG-3), programmed death 1 (PD-1) and glucocorticoid-induced tumour necrosis factor receptor (GITR), suggesting a cell-to-cell contact mechanism with dendritic cells. Furthermore, higher IL-10 and regulatory T cell type 1 (Tr1) levels were found in neurocysticercosis patients' peripheral blood, suggesting that the action mechanism of regulatory T cells involves the release of immunomodulatory cytokines. No evidence was found of the regulatory T cell role in inhibiting the proliferative response. Suppressive regulatory T cells from neurocysticercosis patients correlated negatively with late activated lymphocytes (CD4(+) CD38(+) ). Our results suggest that, during neurocysticercosis, regulatory T cells could control the immune response, probably by a cell-to-cell contact with dendritic cells and interleukin (IL)-10 release by Tr1, to create an immunomodulatory environment that may favour the development of T. solium cysticerci and their permanence in the central nervous system.

  7. An ancient transcriptional regulatory module for tip growth has been conserved throughout the vascular plant lineage.

    PubMed

    Cho, Hyung-Taeg

    2017-02-17

    The root hair development of vascular plants can be divided into two major processes, fate determination and hair morphogenesis, and the latter should be governed by the former so as to express the morphogenetic toolkits in a root hair-specific manner. Vascular plants, depending on taxa, show different fate-determining mechanisms for hair cell/non-hair cell fates, which leads to a question whether the downstream mophogenetic regulatory module is diverged accordingly to the upstream fate determiners or not. Our study demonstrates that the module of a transcription factor and a root hair-specific cis-element (RHE) for root hair-specific expression of morphogenetic toolkit genes is conserved in spite of different fate-determing mechanisms.

  8. Skeletal muscle hypertrophy and regeneration: interplay between the myogenic regulatory factors (MRFs) and insulin-like growth factors (IGFs) pathways.

    PubMed

    Zanou, Nadège; Gailly, Philippe

    2013-11-01

    Adult skeletal muscle can regenerate in response to muscle damage. This ability is conferred by the presence of myogenic stem cells called satellite cells. In response to stimuli such as injury or exercise, these cells become activated and express myogenic regulatory factors (MRFs), i.e., transcription factors of the myogenic lineage including Myf5, MyoD, myogenin, and Mrf4 to proliferate and differentiate into myofibers. The MRF family of proteins controls the transcription of important muscle-specific proteins such as myosin heavy chain and muscle creatine kinase. Different growth factors are secreted during muscle repair among which insulin-like growth factors (IGFs) are the only ones that promote both muscle cell proliferation and differentiation and that play a key role in muscle regeneration and hypertrophy. Different isoforms of IGFs are expressed during muscle repair: IGF-IEa, IGF-IEb, or IGF-IEc (also known as mechano growth factor, MGF) and IGF-II. MGF is expressed first and is observed in satellite cells and in proliferating myoblasts whereas IGF-Ia and IGF-II expression occurs at the state of muscle fiber formation. Interestingly, several studies report the induction of MRFs in response to IGFs stimulation. Inversely, IGFs expression may also be regulated by MRFs. Various mechanisms are proposed to support these interactions. In this review, we describe the general process of muscle hypertrophy and regeneration and decipher the interactions between the two groups of factors involved in the process.

  9. Targeting regulatory T cells.

    PubMed

    Ménétrier-Caux, Christine; Curiel, Tyler; Faget, Julien; Manuel, Manuarii; Caux, Christophe; Zou, Weiping

    2012-03-01

    Cancers express tumor-associated antigens that should elicit immune response to antagonize the tumor growth, but spontaneous immune rejection of established cancer is rare, suggesting an immunosuppressive environment hindering host antitumor immunity. Among the specific and active tumor-mediated mechanisms, CD4(+)CD25(high) T regulatory cells (Treg) are important mediators of active immune evasion in cancer. In this review, we will discuss Treg subpopulations and the mechanisms of their suppressive functions. Treg depletion improves endogenous antitumor immunity and the efficacy of active immunotherapy in animal models for cancer, suggesting that inhibiting Treg function could also improve the limited successes of human cancer immunotherapy. We will also discuss specific strategies for devising effective cancer immunotherapy targeting Treg.

  10. CBP-mediated FOXO-1 acetylation inhibits pancreatic tumor growth by targeting SirT.

    PubMed

    Pramanik, Kartick C; Fofaria, Neel M; Gupta, Parul; Srivastava, Sanjay K

    2014-03-01

    Here, we investigated the potential mechanism of capsaicin-mediated apoptosis in pancreatic cancer cells. Capsaicin treatment phosphorylated c-jun-NH2-kinase (JNK); forkhead box transcription factor, class O (FOXO1); and BIM in BxPC-3, AsPC-1, and L3.6PL cells. The expression of BIM increased in response to capsaicin treatment. Capsaicin treatment caused cleavage of caspase-3 and PARP, indicating apoptosis. Antioxidants tiron and PEG-catalase blocked capsaicin-mediated JNK/FOXO/BIM activation and protected the cells from apoptosis. Furthermore, capsaicin treatment caused a steady increase in the nuclear expression of FOXO-1, leading to increased DNA binding. Capsaicin-mediated expression of BIM was found to be directly dependent on the acetylation of FOXO-1. The expression of CREB-binding protein (CBP) was increased, whereas SirT-1 was reduced by capsaicin treatment. Using acetylation mimic or defective mutants, our result demonstrated that phosphorylation of FOXO-1 was mediated through acetylation by capsaicin treatment. JNK inhibitor attenuated the phosphorylation of FOXO-1, activation of BIM, and abrogated capsaicin-induced apoptosis. Moreover, silencing FOXO1 by siRNA blocked capsaicin-mediated activation of BIM and apoptosis, whereas overexpression of FOXO-1 augmented its effects. Silencing Bim drastically reduced capsaicin-mediated cleavage of caspase-3 and PARP, indicating the role of BIM in apoptosis. Oral administration of 5 mg/kg capsaicin substantially suppressed the growth of BxPC-3 tumor xenografts in athymic nude mice. Tumors from capsaicin-treated mice showed an increase in the phosphorylation of JNK, FOXO-1, BIM, and levels of CBP, cleavage of caspase-3, PARP, and decreased SirT-1 expression. Taken together, our results suggest that capsaicin activated JNK and FOXO-1, leading to the acetylation of FOXO-1 through CBP and SirT-1. Acetylated FOXO1 induced apoptosis in pancreatic cancer cells through BIM activation.

  11. miR-145 mediates the antiproliferative and gene regulatory effects of vitamin D3 by directly targeting E2F3 in gastric cancer cells

    PubMed Central

    Chang, Su'e; Gao, Ling; Yang, Yang; Tong, Dongdong; Guo, Bo; Liu, Liying; Li, Zongfang; Song, Tusheng; Huang, Chen

    2015-01-01

    VitaminD3 signaling is involved in inhibiting the development and progression of gastric cancer (GC), while the active vitamin D metabolite 1-alpha,25-dihydroxyvitamin D3 (1,25(OH)2D3)-mediated gene regulatory mechanisms in GC remain unclear. We found that miR-145 is induced by 1,25(OH)2D3 in a dose- and vitamin D receptor (VDR)-dependent manner in GC cells. Inhibition of miR-145 reverses the antiproliferative effect of 1,25(OH)2D3. Furthermore, miR-145 expression was lower in tumors compared with matched normal samples and correlated with increased the E2F3 transcription factor protein staining. Overexpression of miR-145 inhibited colony formation, cell viability and induced cell arrest in S-phase in GC cells by targeting E2F3 and CDK6. miR-145 inhibition consistently abrogates the 1,25(OH)2D3-mediated suppression of E2F3, CDK6, CDK2 and CCNA2 genes. Altogether, our results indicate that miR-145 mediates the antiproliferative and gene regulatory effects of vitamin D3 in GC cells and might hold promise for prognosis and therapeutic strategies for GC treatment. PMID:25762621

  12. Antisense oligonucleotide–mediated MDM4 exon 6 skipping impairs tumor growth

    PubMed Central

    Dewaele, Michael; Tabaglio, Tommaso; Willekens, Karen; Bezzi, Marco; Teo, Shun Xie; Low, Diana H.P.; Koh, Cheryl M.; Rambow, Florian; Fiers, Mark; Rogiers, Aljosja; Radaelli, Enrico; Al-Haddawi, Muthafar; Tan, Soo Yong; Hermans, Els; Amant, Frederic; Yan, Hualong; Lakshmanan, Manikandan; Koumar, Ratnacaram Chandrahas; Lim, Soon Thye; Derheimer, Frederick A.; Campbell, Robert M.; Bonday, Zahid; Tergaonkar, Vinay; Shackleton, Mark; Blattner, Christine; Marine, Jean-Christophe; Guccione, Ernesto

    2015-01-01

    MDM4 is a promising target for cancer therapy, as it is undetectable in most normal adult tissues but often upregulated in cancer cells to dampen p53 tumor-suppressor function. The mechanisms that underlie MDM4 upregulation in cancer cells are largely unknown. Here, we have shown that this key oncogenic event mainly depends on a specific alternative splicing switch. We determined that while a nonsense-mediated, decay-targeted isoform of MDM4 (MDM4-S) is produced in normal adult tissues as a result of exon 6 skipping, enhanced exon 6 inclusion leads to expression of full-length MDM4 in a large number of human cancers. Although this alternative splicing event is likely regulated by multiple splicing factors, we identified the SRSF3 oncoprotein as a key enhancer of exon 6 inclusion. In multiple human melanoma cell lines and in melanoma patient–derived xenograft (PDX) mouse models, antisense oligonucleotide–mediated (ASO-mediated) skipping of exon 6 decreased MDM4 abundance, inhibited melanoma growth, and enhanced sensitivity to MAPK-targeting therapeutics. Additionally, ASO-based MDM4 targeting reduced diffuse large B cell lymphoma PDX growth. As full-length MDM4 is enhanced in multiple human tumors, our data indicate that this strategy is applicable to a wide range of tumor types. We conclude that enhanced MDM4 exon 6 inclusion is a common oncogenic event and has potential as a clinically compatible therapeutic target. PMID:26595814

  13. Growth control in colon epithelial cells: gadolinium enhances calcium-mediated growth regulation.

    PubMed

    Attili, Durga; Jenkins, Brian; Aslam, Muhammad Nadeem; Dame, Michael K; Varani, James

    2012-12-01

    Gadolinium, a member of the lanthanoid family of transition metals, interacts with calcium-binding sites on proteins and other biological molecules. The overall goal of the present investigation was to determine if gadolinium could enhance calcium-induced epithelial cell growth inhibition in the colon. Gadolinium at concentrations as low as 1-5 μM combined with calcium inhibits proliferation of human colonic epithelial cells more effectively than calcium alone. Gadolinium had no detectable effect on calcium-induced differentiation in the same cells based on change in cell morphology, induction of E-cadherin synthesis, and translocation of E-cadherin from the cytosol to the cell surface. When the colon epithelial cells were treated with gadolinium and then exposed to increased calcium concentrations, movement of extracellular calcium into the cell was suppressed. In contrast, gadolinium treatment had no effect on ionomycin-induced release of stored intracellular calcium into the cytoplasm. Whether these in vitro observations can be translated into an approach for reducing abnormal proliferation in the colonic mucosa (including polyp formation) is not known. These results do, however, provide an explanation for our recent findings that a multi-mineral supplement containing all of the naturally occurring lanthanoid metals including gadolinium are more effective than calcium alone in preventing colon polyp formation in mice on a high-fat diet.

  14. The Pollen Receptor Kinase LePRK2 Mediates Growth-Promoting Signals and Positively Regulates Pollen Germination and Tube Growth

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In flowering plants, the process of pollen germination and tube growth is required for successful fertilization. A pollen receptor kinase from tomato, LePRK2, has been implicated in signaling during pollen germination and tube growth as well as in mediating pollen (tube)-pistil communication. Here w...

  15. The impact of stress on tumor growth: peripheral CRF mediates tumor-promoting effects of stress

    PubMed Central

    2010-01-01

    Introduction Stress has been shown to be a tumor promoting factor. Both clinical and laboratory studies have shown that chronic stress is associated with tumor growth in several types of cancer. Corticotropin Releasing Factor (CRF) is the major hypothalamic mediator of stress, but is also expressed in peripheral tissues. Earlier studies have shown that peripheral CRF affects breast cancer cell proliferation and motility. The aim of the present study was to assess the significance of peripheral CRF on tumor growth as a mediator of the response to stress in vivo. Methods For this purpose we used the 4T1 breast cancer cell line in cell culture and in vivo. Cells were treated with CRF in culture and gene specific arrays were performed to identify genes directly affected by CRF and involved in breast cancer cell growth. To assess the impact of peripheral CRF as a stress mediator in tumor growth, Balb/c mice were orthotopically injected with 4T1 cells in the mammary fat pad to induce breast tumors. Mice were subjected to repetitive immobilization stress as a model of chronic stress. To inhibit the action of CRF, the CRF antagonist antalarmin was injected intraperitoneally. Breast tissue samples were histologically analyzed and assessed for neoangiogenesis. Results Array analysis revealed among other genes that CRF induced the expression of SMAD2 and β-catenin, genes involved in breast cancer cell proliferation and cytoskeletal changes associated with metastasis. Cell transfection and luciferase assays confirmed the role of CRF in WNT- β-catenin signaling. CRF induced 4T1 cell proliferation and augmented the TGF-β action on proliferation confirming its impact on TGFβ/SMAD2 signaling. In addition, CRF promoted actin reorganization and cell migration, suggesting a direct tumor-promoting action. Chronic stress augmented tumor growth in 4T1 breast tumor bearing mice and peripheral administration of the CRF antagonist antalarmin suppressed this effect. Moreover, antalarmin

  16. Neurotropic arboviruses induce interferon regulatory factor 3-mediated neuronal responses that are cytoprotective, interferon independent, and inhibited by Western equine encephalitis virus capsid.

    PubMed

    Peltier, Daniel C; Lazear, Helen M; Farmer, Jocelyn R; Diamond, Michael S; Miller, David J

    2013-02-01

    Cell-intrinsic innate immune responses mediated by the transcription factor interferon regulatory factor 3 (IRF-3) are often vital for early pathogen control, and effective responses in neurons may be crucial to prevent the irreversible loss of these critical central nervous system cells after infection with neurotropic pathogens. To investigate this hypothesis, we used targeted molecular and genetic approaches with cultured neurons to study cell-intrinsic host defense pathways primarily using the neurotropic alphavirus western equine encephalitis virus (WEEV). We found that WEEV activated IRF-3-mediated neuronal innate immune pathways in a replication-dependent manner, and abrogation of IRF-3 function enhanced virus-mediated injury by WEEV and the unrelated flavivirus St. Louis encephalitis virus. Furthermore, IRF-3-dependent neuronal protection from virus-mediated cytopathology occurred independently of autocrine or paracrine type I interferon activity. Despite being partially controlled by IRF-3-dependent signals, WEEV also disrupted antiviral responses by inhibiting pattern recognition receptor pathways. This antagonist activity was mapped to the WEEV capsid gene, which disrupted signal transduction downstream of IRF-3 activation and was independent of capsid-mediated inhibition of host macromolecular synthesis. Overall, these results indicate that innate immune pathways have important cytoprotective activity in neurons and contribute to limiting injury associated with infection by neurotropic arboviruses.

  17. Regulatory cascade and biological activity of Beauveria bassiana oosporein that limits bacterial growth after host death.

    PubMed

    Fan, Yanhua; Liu, Xi; Keyhani, Nemat O; Tang, Guirong; Pei, Yan; Zhang, Wenwen; Tong, Sheng

    2017-02-28

    The regulatory network and biological functions of the fungal secondary metabolite oosporein have remained obscure. Beauveria bassiana has evolved the ability to parasitize insects and outcompete microbial challengers for assimilation of host nutrients. A novel zinc finger transcription factor, BbSmr1 (B. bassiana secondary metabolite regulator 1), was identified in a screen for oosporein overproduction. Deletion of Bbsmr1 resulted in up-regulation of the oosporein biosynthetic gene cluster (OpS genes) and constitutive oosporein production. Oosporein production was abolished in double mutants of Bbsmr1 and a second transcription factor, OpS3, within the oosporein gene cluster (ΔBbsmr1ΔOpS3), indicating that BbSmr1 acts as a negative regulator of OpS3 expression. Real-time quantitative PCR and a GFP promoter fusion construct of OpS1, the oosporein polyketide synthase, indicated that OpS1 is expressed mainly in insect cadavers at 24-48 h after death. Bacterial colony analysis in B. bassiana-infected insect hosts revealed increasing counts until host death, with a dramatic decrease (∼90%) after death that correlated with oosporein production. In vitro studies verified the inhibitory activity of oosporein against bacteria derived from insect cadavers. These results suggest that oosporein acts as an antimicrobial compound to limit microbial competition on B. bassiana-killed hosts, allowing the fungus to maximally use host nutrients to grow and sporulate on infected cadavers.

  18. Fetal production of growth factors and inflammatory mediators predicts pulmonary hypertension in congenital diaphragmatic hernia

    PubMed Central

    Fleck, Shannon; Bautista, Geoanna; Keating, Sheila M.; Lee, Tzong-Hae; Keller, Roberta L.; Moon-Grady, Anita J.; Gonzales, Kelly; Norris, Philip J.; Busch, Michael P.; Kim, CJ; Romero, Roberto; Lee, Hanmin; Miniati, Doug; MacKenzie, Tippi C.

    2014-01-01

    Background Congenital diaphragmatic hernia (CDH) represents a spectrum of lung hypoplasia and consequent pulmonary hypertension is an important cause of postnatal morbidity and mortality. We studied biomarkers at the maternal-fetal interface to understand factors associated with the persistence of pulmonary hypertension. Methods Maternal and cord blood samples from fetuses with CDH and unaffected controls were analyzed using a human 39plex immunoassay kit. Cellular trafficking between the mother and the fetu was quantified using quantitative real-time PCR for non-shared alleles. Biomarker profiles were then correlated with CDH severity based on the degree of pulmonary hypertension. Results Cord blood levels of epidermal growth factor, platelet-derived growth factor, and several inflammatory mediators increased significantly as the severity of CDH increased, while maternal levels growth factors and mediators decreased significantly with CDH severity. Maternal cells were increased in fetuses with severe CDH compared to controls, with elevated levels of the chemokine CXCL-10 in patients with the highest trafficking. Conclusion Patients with CDH demonstrate pro-inflammatory and chemotactic signals in fetal blood at the time of birth. Since some of these molecules have been implicated in the development of pulmonary hypertension, prenatal strategies targeting specific molecular pathways may be useful adjuncts to current fetal therapies. PMID:23770923

  19. Insect growth regulatory effects of linear diterpenoids and derivatives from Baccharis thymifolia.

    PubMed

    Juan Hikawczuk, Virginia E; Saad, J Roberto; Giordano, Oscar S; García, Celina; Martín, Tomás; Martín, Víctor S; Sosa, Marta E; Tonn, Carlos E

    2008-02-01

    Linear diterpenes isolated from aerial parts of Baccharis thymifolia ( 1- 3) were tested for insect growth inhibitory activity against Tenebrio molitor larvae. Compounds 4- 16 were prepared by various chemical transformations. The diterpenes 6,10-( E,E)-thymifodioic acid ( 1), the butenolide 3, and the derivatives 4, 11, and 14 produced developmental deficiencies in assays using topical application on fifth instar larvae of T. molitor. Compound 3 is a new natural product.

  20. A destructive cascade mediated by CCL2 facilitates prostate cancer growth in bone.

    PubMed

    Li, Xin; Loberg, Robert; Liao, Jinhui; Ying, Chi; Snyder, Linda A; Pienta, Kenneth J; McCauley, Laurie K

    2009-02-15

    Monocyte chemoattractant protein 1 (CCL2) is a recently identified prominent regulator of prostate cancer growth and metastasis. The purpose of this study was to investigate the mechanistic role of CCL2 in prostate cancer growth in bone. The present study found that CCL2 was up-regulated in osteoblasts (3-fold by PC-3 and 2-fold by VCaP conditioned medium) and endothelial cells (2-fold by PC-3 and VCaP conditioned medium). Parathyroid hormone-related protein (PTHrP) treatment of osteoblastic cells up-regulated CCL2 and was blocked by a PTHrP antagonist, suggesting that prostate cancer-derived PTHrP plays an important role in elevation of osteoblast-derived CCL2. CCL2 indirectly increased blood vessel formation in endothelial cells through vascular endothelial growth factor-A, which was up-regulated 2-fold with administration of CCL2 in prostate cancer cells. In vivo, anti-CCL2 treatment suppressed tumor growth in bone. The decreased tumor burden was associated with decreased bone resorption (serum TRAP5b levels were decreased by 50-60% in anti-CCL2-treated animals from VCaP or PC-3 cell osseous lesions) and microvessel density was decreased by 70% in anti-CCL2-treated animals with bone lesions from VCaP cells. These data suggest that a destructive cascade is driven by tumor cell-derived, PTHrP-mediated induction of CCL2, which facilitates tumor growth via enhanced osteoclastic and endothelial cell activity in bone marrow. Taken together, CCL2 mediates the interaction between tumor-derived factors and host-derived chemokines acting in cooperation to promote skeletal metastasis.

  1. Valproic acid overcomes transforming growth factor-β-mediated sorafenib resistance in hepatocellular carcinoma

    PubMed Central

    Matsuda, Yasunobu; Wakai, Toshifumi; Kubota, Masayuki; Osawa, Mami; Hirose, Yuki; Sakata, Jun; Kobayashi, Takashi; Fujimaki, Shun; Takamura, Masaaki; Yamagiwa, Satoshi; Aoyagi, Yutaka

    2014-01-01

    Sorafenib is a multi-kinase inhibitor approved for hepatocellular carcinoma, but rarely causes tumor regression in patients with chronic liver diseases. To investigate whether growth factor-mediated signaling is involved in sorafenib resistance, HepG2 and PLC/PRF/5 hepatoma cells were exposed to epidermal growth factor (EGF), hepatocyte growth factor (HGF) or transforming growth factor-β (TGF-β) prior to treatment with sorafenib. Furthermore, to identify an effective combination treatment with sorafenib, growth factor-sensitized cells were treated with sorafenib alone or in combination with celecoxib, lovastatin or valproic acid (VPA). Trypan blue staining and Annexin V assays showed that the cytotoxic effect of sorafenib was inhibited by 15-54% in cells sensitized to TGF-β (P<0.05). Western blotting analysis showed that TGF-β significantly activated extracellular signal-regulated kinase (ERK)-mediated AKT signaling, and sorafenib failed to suppress both ERK and AKT in TGF-β-sensitized cells. The decreased anti-tumor effect of sorafenib was rescued by chemical inhibition of ERK and AKT. When TGF-β-sensitized cells were treated with sorafenib plus VPA, the levels of phosphorylated ERK and AKT were considerably suppressed and the numbers of dead cells were increased by 3.7-5.7-fold compared with those exposed to sorafenib alone (P<0.05). Moreover, low dose sorafenib-induced cell migration was effectively suppressed by combination treatment with sorafenib and VPA. Collectively, TGF-β/ERK/AKT signaling might play a critical role in sorafenib resistance in hepatoma cells, and combination treatment with VPA may be effective against this drug resistance. PMID:24817927

  2. Transforming growth factor-beta1 mediates cellular response to DNA damage in situ

    NASA Technical Reports Server (NTRS)

    Ewan, Kenneth B.; Henshall-Powell, Rhonda L.; Ravani, Shraddha A.; Pajares, Maria Jose; Arteaga, Carlos; Warters, Ray; Akhurst, Rosemary J.; Barcellos-Hoff, Mary Helen

    2002-01-01

    Transforming growth factor (TGF)-beta1 is rapidly activated after ionizing radiation, but its specific role in cellular responses to DNA damage is not known. Here we use Tgfbeta1 knockout mice to show that radiation-induced apoptotic response is TGF-beta1 dependent in the mammary epithelium, and that both apoptosis and inhibition of proliferation in response to DNA damage decrease as a function of TGF-beta1 gene dose in embryonic epithelial tissues. Because apoptosis in these tissues has been shown previously to be p53 dependent, we then examined p53 protein activation. TGF-beta1 depletion, by either gene knockout or by using TGF-beta neutralizing antibodies, resulted in decreased p53 Ser-18 phosphorylation in irradiated mammary gland. These data indicate that TGF-beta1 is essential for rapid p53-mediated cellular responses that mediate cell fate decisions in situ.

  3. The expression of myogenic regulatory factors and muscle growth factors in the masticatory muscles of dystrophin-deficient (mdx) mice.

    PubMed

    Spassov, Alexander; Gredes, Tomasz; Gedrange, Tomasz; Lucke, Silke; Pavlovic, Dragan; Kunert-Keil, Christiane

    2011-06-01

    The activities of myogenic regulatory factors (MRF) and muscle growth factors increase in muscle that is undergoing regeneration, and may correspond to some specific changes. Little is known about the role of MRFs in masticatory muscles in mdx mice (the model of Duchenne muscular dystrophy) and particularly about their mRNA expression during the process of muscle regeneration. Using Taqman RT-PCR, we examined the mRNA expression of the MRFs myogenin and MyoD1 (myogenic differentiation 1), and of the muscle growth factors myostatin, IGF1 (insulin-like growth factor) and MGF (mechano-growth factor) in the masseter, temporal and tongue masticatory muscles of mdx mice (n = 6 to 10 per group). The myogenin mRNA expression in the mdx masseter and temporal muscle was found to have increased (P < 0.05), whereas the myostatin mRNA expressions in the mdx masseter (P < 0.005) and tongue (P < 0.05) were found to have diminished compared to those for the controls. The IGF and MGF mRNA amounts in the mdx mice remained unchanged. Inside the mdx animal group, gender-related differences in the mRNA expressions were also found. A higher mRNA expression of myogenin and MyoD1 in the mdx massterer and temporal muscles was found in females in comparison to males, and the level of myostatin was higher in the masseter and tongue muscle (P < 0.001 for all comparisons). Similar gender-related differences were also found within the control groups. This study reveals the intermuscular differences in the mRNA expression pattern of myogenin and myostatin in mdx mice. The existence of these differences implies that dystrophinopathy affects the skeletal muscles differentially. The finding of gender-related differences in the mRNA expression of the examined factors may indicate the importance of hormonal influences on muscle regeneration.

  4. 14-3-3 sigma and 14-3-3 zeta plays an opposite role in cell growth inhibition mediated by transforming growth factor-beta 1.

    PubMed

    Hong, Hye-Young; Jeon, Woo-Kwang; Bae, Eun-Jin; Kim, Shin-Tae; Lee, Ho-Jae; Kim, Seong-Jin; Kim, Byung-Chul

    2010-03-01

    The expression of 14-3-3 proteins is dysregulated in various types of cancer. This study was undertaken to investigate the effects of 14-3-3 zeta and 14-3-3 sigma on cell growth inhibition mediated by transforming growth factor-beta 1 (TGF-beta1). Mouse mammary epithelial cells (Eph4) that are transformed with oncogenic c-H-Ras (EpRas) and no longer sensitive to TGF-beta1-mediated growth inhibition displayed increased expression of 14-3-3 zeta and decreased expression of 14-3-3 sigma compared with parental Eph4 cells. Using small interfering RNA-mediated knockdown and overexpression of 14-3-3 sigma or 14-3-3 zeta, we showed that 14-3-3 sigma is required for TGF-beta1-mediated growth inhibition whereas 14-3-3 zeta negatively modulates this growth inhibitory response. Notably, overexpression of 14-3-3 zeta increased the level of Smad3 protein that is phosphorylated at linker regions and cannot mediate the TGF-beta1 growth inhibitory response. Consistent with this finding, mutation of the 14-3-3 zeta phosphorylation sites in Smad3 markedly reduced the 14-3-3 zeta-mediated inhibition of TGF-beta1-induced p15 promoter-reporter activity and cell cycle arrest, suggesting that these residues are critical targets of 14-3-3 zeta in the suppression of TGF-beta1-mediated growth. Taken together, our findings indicate that dysregulation of 14-3-3 sigma or 14-3-3 zeta contributes to TGF-beta1 resistance in cancer cells.

  5. Intronic Cis-Regulatory Modules Mediate Tissue-Specific and Microbial Control of angptl4/fiaf Transcription

    PubMed Central

    Camp, J. Gray; Jazwa, Amelia L.; Trent, Chad M.; Rawls, John F.

    2012-01-01

    The intestinal microbiota enhances dietary energy harvest leading to increased fat storage in adipose tissues. This effect is caused in part by the microbial suppression of intestinal epithelial expression of a circulating inhibitor of lipoprotein lipase called Angiopoietin-like 4 (Angptl4/Fiaf). To define the cis-regulatory mechanisms underlying intestine-specific and microbial control of Angptl4 transcription, we utilized the zebrafish system in which host regulatory DNA can be rapidly analyzed in a live, transparent, and gnotobiotic vertebrate. We found that zebrafish angptl4 is transcribed in multiple tissues including the liver, pancreatic islet, and intestinal epithelium, which is similar to its mammalian homologs. Zebrafish angptl4 is also specifically suppressed in the intestinal epithelium upon colonization with a microbiota. In vivo transgenic reporter assays identified discrete tissue-specific regulatory modules within angptl4 intron 3 sufficient to drive expression in the liver, pancreatic islet β-cells, or intestinal enterocytes. Comparative sequence analyses and heterologous functional assays of angptl4 intron 3 sequences from 12 teleost fish species revealed differential evolution of the islet and intestinal regulatory modules. High-resolution functional mapping and site-directed mutagenesis defined the minimal set of regulatory sequences required for intestinal activity. Strikingly, the microbiota suppressed the transcriptional activity of the intestine-specific regulatory module similar to the endogenous angptl4 gene. These results suggest that the microbiota might regulate host intestinal Angptl4 protein expression and peripheral fat storage by suppressing the activity of an intestine-specific transcriptional enhancer. This study provides a useful paradigm for understanding how microbial signals interact with tissue-specific regulatory networks to control the activity and evolution of host gene transcription. PMID:22479192

  6. Mediation of wound-related Rous sarcoma virus tumorigenesis by TFG (transforming growth factor)-. beta

    SciTech Connect

    Sieweke, M.H.; Bissell, M.J. ); Thompson, N.L.; Sporn, M.B. )

    1990-06-29

    In Rous sarcoma virus (RSV)-infected chickens, wounding leads to tumor formation with nearly 100% frequency in tissues that would otherwise remain tumor-free. Identifying molecular mediators of this phenomenon should yield important clues to the mechanisms involved in RSV tumorigenesis. Immunohistochemical staining showed that TGF-{beta} is present locally shortly after wounding, but not in unwounded controls. In addition, subcutaneous administration of recombinant transforming growth factor {beta}1 (TGF-{beta}1) could substitute completely for wounding in tumor induction. A treatment protocol of four doses of 800 nanograms of TGF-{beta} resulted in v-src-expressing tumors with 100% frequency; four doses of only 10 nanograms still led to tumor formation in 80% of the animals. This effect was specific, as other growth factors with suggested roles in would healing did not elicit the same response. Epidermal growth factor (EGF) or TGF-{alpha} had no effect, and platelet-derived growth factor (PDGF) or insulin-like growth factor-1 (IGF-1) yielded only occasional tumors after longer latency. TGF-{beta} release during the would-healing response may thus be a critical event that creates a conducive environment for RSV tumorigenesis and may act as a cofactor for transformation in this system. 31 refs., 3 figs., 2 tabs.

  7. Testosterone mediates seasonal growth of the song control nuclei in a tropical bird

    PubMed Central

    Small, Thomas W.; Brenowitz, Eliot A.; Wojtenek, Winfried; Moore, Ignacio T.

    2015-01-01

    In mid- to high-latitude songbirds seasonal reproduction is stimulated by increasing daylength accompanied by elevated plasma sex steroid levels, increased singing, and growth of the song control nuclei (SCN). Plasticity of the SCN and song behavior are primarily mediated by testosterone (T) and its metabolites in most species studied thus far. However, the majority of bird species are tropical and have less pronounced seasonal reproductive cycles. We have previously documented that equatorial rufous-collared sparrows (Zonotrichia capensis) exhibit seasonal neuroplasticity in the SCN. Manipulating T in these birds, however, did not alter singing behavior. In the current study we investigated whether T mediates plasticity of the SCN in a similar manner to temperate songbirds. In the first experiment we treated captive male birds with T or blank implants during the non-breeding season. In a second experiment we treated captive males with either blank implants, T-filled implants, T with Flutamide (FLU; an androgen receptor antagonist) or T with FLU and 1,4,6-androstatriene-3,17-dione (ATD; an estrogen synthesis inhibitor) during the breeding season. In both experiments, the volumes of brain areas HVC, Area X, and RA were measured along with singing behavior. In summary, T stimulated growth of HVC and RA and the combined effect of FLU and ATD reversed this effect in HVC. Area X was not affected by testosterone treatment in either experiment. Neither T-treated birds nor controls sang in captivity during either experiment. Together these data indicate that T mediates seasonal changes in the HVC and RA of both tropical and higher latitude bird species even if the environmental signals differ. However, unlike most higher latitude songbirds, we found no evidence that motivation to sing or growth of Area X are stimulated by T under captive conditions. PMID:26346733

  8. Egr2 and Egr3 in regulatory T cells cooperatively control systemic autoimmunity through Ltbp3-mediated TGF-β3 production.

    PubMed

    Morita, Kaoru; Okamura, Tomohisa; Inoue, Mariko; Komai, Toshihiko; Teruya, Shuzo; Iwasaki, Yukiko; Sumitomo, Shuji; Shoda, Hirofumi; Yamamoto, Kazuhiko; Fujio, Keishi

    2016-12-13

    Systemic lupus erythematosus (SLE) is a prototypical autoimmune disease characterized by multiorgan inflammation induced by autoantibodies. Early growth response gene 2 (Egr2), a transcription factor essential for T-cell anergy induction, controls systemic autoimmunity in mice and humans. We have previously identified a subpopulation of CD4(+) regulatory T cells, CD4(+)CD25(-)LAG3(+) cells, that characteristically express both Egr2 and LAG3 and control mice model of lupus via TGF-β3 production. However, due to the mild phenotype of lymphocyte-specific Egr2-deficient mice, the presence of an additional regulator has been speculated. Here, we show that Egr2 and Egr3 expressed in T cells cooperatively prevent humoral immune responses by supporting TGF-β3 secretion. T cell-specific Egr2/Egr3 double-deficient (Egr2/3DKO) mice spontaneously developed an early onset lupus-like disease that was more severe than in T cell-specific Egr2-deficient mice. In accordance with the observation that CD4(+)CD25(-)LAG3(+) cells from Egr2/3DKO mice completely lost the capacity to produce TGF-β3, the excessive germinal center reaction in Egr2/3DKO mice was suppressed by the adoptive transfer of WT CD4(+)CD25(-)LAG3(+) cells or treatment with a TGF-β3-expressing vector. Intriguingly, latent TGF-β binding protein (Ltbp)3 expression maintained by Egr2 and Egr3 was required for TGF-β3 production from CD4(+)CD25(-)LAG3(+) cells. Because Egr2 and Egr3 did not demonstrate cell intrinsic suppression of the development of follicular helper T cells, Egr2- and Egr3-dependent TGF-β3 production by CD4(+)CD25(-)LAG3(+) cells is critical for controlling excessive B-cell responses. The unique attributes of Egr2/Egr3 in T cells may provide an opportunity for developing novel therapeutics for autoantibody-mediated diseases including SLE.

  9. Egr2 and Egr3 in regulatory T cells cooperatively control systemic autoimmunity through Ltbp3-mediated TGF-β3 production

    PubMed Central

    Morita, Kaoru; Okamura, Tomohisa; Inoue, Mariko; Komai, Toshihiko; Teruya, Shuzo; Iwasaki, Yukiko; Sumitomo, Shuji; Shoda, Hirofumi; Yamamoto, Kazuhiko; Fujio, Keishi

    2016-01-01

    Systemic lupus erythematosus (SLE) is a prototypical autoimmune disease characterized by multiorgan inflammation induced by autoantibodies. Early growth response gene 2 (Egr2), a transcription factor essential for T-cell anergy induction, controls systemic autoimmunity in mice and humans. We have previously identified a subpopulation of CD4+ regulatory T cells, CD4+CD25−LAG3+ cells, that characteristically express both Egr2 and LAG3 and control mice model of lupus via TGF-β3 production. However, due to the mild phenotype of lymphocyte-specific Egr2-deficient mice, the presence of an additional regulator has been speculated. Here, we show that Egr2 and Egr3 expressed in T cells cooperatively prevent humoral immune responses by supporting TGF-β3 secretion. T cell-specific Egr2/Egr3 double-deficient (Egr2/3DKO) mice spontaneously developed an early onset lupus-like disease that was more severe than in T cell-specific Egr2-deficient mice. In accordance with the observation that CD4+CD25−LAG3+ cells from Egr2/3DKO mice completely lost the capacity to produce TGF-β3, the excessive germinal center reaction in Egr2/3DKO mice was suppressed by the adoptive transfer of WT CD4+CD25−LAG3+ cells or treatment with a TGF-β3–expressing vector. Intriguingly, latent TGF-β binding protein (Ltbp)3 expression maintained by Egr2 and Egr3 was required for TGF-β3 production from CD4+CD25−LAG3+ cells. Because Egr2 and Egr3 did not demonstrate cell intrinsic suppression of the development of follicular helper T cells, Egr2- and Egr3-dependent TGF-β3 production by CD4+CD25−LAG3+ cells is critical for controlling excessive B-cell responses. The unique attributes of Egr2/Egr3 in T cells may provide an opportunity for developing novel therapeutics for autoantibody-mediated diseases including SLE. PMID:27911796

  10. Regulatory functions of SnRK1 in stress-responsive gene expression and in plant growth and development.

    PubMed

    Cho, Young-Hee; Hong, Jung-Woo; Kim, Eun-Chul; Yoo, Sang-Dong

    2012-04-01

    Sucrose-nonfermentation1-related protein kinase1 (SnRK1) is an evolutionarily conserved energy sensor protein that regulates gene expression in response to energy depletion in plants. Efforts to elucidate the functions and mechanisms of this protein kinase are hampered, however, by inherent growth defects of snrk1-null mutant plants. To overcome these limitations and study SnRK1 functions in vivo, we applied a method combining transient expression in leaf mesophyll protoplasts and stable expression in transgenic plants. We found that both rice (Oryza sativa) and Arabidopsis (Arabidopsis thaliana) SnRK1 activities critically influence stress-inducible gene expression and the induction of stress tolerance. Genetic, molecular, and chromatin immunoprecipitation analyses further revealed that the nuclear SnRK1 modulated target gene transcription in a submergence-dependent manner. From early seedling development through late senescence, SnRK1 activities appeared to modulate developmental processes in the plants. Our findings offer insight into the regulatory functions of plant SnRK1 in stress-responsive gene regulation and in plant growth and development throughout the life cycle.

  11. Involvement of hexokinase1 in plant growth promotion as mediated by Burkholderia phytofirmans.

    PubMed

    Park, Jae Min; Lazarovits, George

    2014-06-01

    Potato plantlets inoculated with strain PsJN of the bacterium Burkholderia phytofirmans exhibit consistent and significant increases in plant growth under in vitro conditions, when compared with uninoculated plants. The greatest influence on the degree and type of growth enhancement that develops has been shown to be mediated by the sugar concentration in the agar media. Bacterial growth promotion has been suggested in other studies to be regulated by the sugar sensor enzyme hexokinase1, the role of which is activation of glucose phosphorylation. In this present study, we examined the co-relationship between root and stem development in potato plants treated with PsJN and the activity of hexokinase1. Plants grown in the presence of 1.5% and 3% sucrose showed increased levels of hexokinase1 activity only in the roots of inoculated plants, suggesting that the increased enzyme levels may be associated with root growth. Analysis for mRNA using reverse transcriptase did not reveal any significant differences in transcription levels of the gene between inoculated and uninoculated plants. When PsJN-inoculated plants were grown in 1.5% and 3% concentrations of glucose and fructose, stem height and mass, leaf number, root mass, and overall biomass increased. No growth promotion occurred when PsJN-inoculated plants were grown in 3% maltose. Subsequently, a hexokinase1 activity assay showed that PsJN-induced growth of potato plants was found to only occur when plants were grown in the presence of sugars that are recognized by the plant hexokinase1. The results suggest that PsJN may enhance sugar uptake in plants by direct or indirect stimulation of hexokinase1 activity in roots and this results in enhanced overall plant growth.

  12. The chemokine CXCL12 mediates the anti-amyloidogenic action of painless human nerve growth factor.

    PubMed

    Capsoni, Simona; Malerba, Francesca; Carucci, Nicola Maria; Rizzi, Caterina; Criscuolo, Chiara; Origlia, Nicola; Calvello, Mariantonietta; Viegi, Alessandro; Meli, Giovanni; Cattaneo, Antonino

    2017-01-01

    Nerve growth factor is a therapeutic candidate for Alzheimer's disease. Due to its pain-inducing activity, in current clinical trials nerve growth factor is delivered locally into the brain by neurosurgery, but data on the efficacy of local nerve growth factor delivery in decreasing amyloid-β deposition are not available. To reduce the nerve growth factor pain-inducing side effects, thus avoiding the need for local brain injection, we developed human painless nerve growth factor (hNGFp), inspired by the human genetic disease hereditary sensory and autonomic neuropathy type V. hNGFp has identical neurotrophic potency as wild-type human nerve growth factor, but a 10-fold lower pain sensitizing activity. In this study we first mimicked, in the 5xFAD mouse model, the intraparenchymal delivery of hNGFp used in clinical trials and found it to be ineffective in decreasing amyloid-β plaque load. On the contrary, the same dose of hNGFp delivered intranasally, which was widely biodistributed in the brain and did not induce pain, showed a potent anti-amyloidogenic action and rescued synaptic plasticity and memory deficits. We found that hNGFp acts on glial cells, modulating inflammatory proteins such as the soluble TNFα receptor II and the chemokine CXCL12. We further established that the rescuing effect by hNGFp is mediated by CXCL12, as pharmacological inhibition of CXCL12 receptor CXCR4 occludes most of hNGFp effects. These findings have significant therapeutic implications: (i) we established that a widespread exposure of the brain is required for nerve growth factor to fully exert its neuroprotective actions; and (ii) we have identified a new anti-neurodegenerative pathway as a broad target for new therapeutic opportunities for neurodegenerative diseases.

  13. The chemokine CXCL12 mediates the anti-amyloidogenic action of painless human nerve growth factor

    PubMed Central

    Capsoni, Simona; Malerba, Francesca; Carucci, Nicola Maria; Rizzi, Caterina; Criscuolo, Chiara; Origlia, Nicola; Calvello, Mariantonietta; Viegi, Alessandro; Meli, Giovanni

    2017-01-01

    Abstract Nerve growth factor is a therapeutic candidate for Alzheimer’s disease. Due to its pain-inducing activity, in current clinical trials nerve growth factor is delivered locally into the brain by neurosurgery, but data on the efficacy of local nerve growth factor delivery in decreasing amyloid-β deposition are not available. To reduce the nerve growth factor pain-inducing side effects, thus avoiding the need for local brain injection, we developed human painless nerve growth factor (hNGFp), inspired by the human genetic disease hereditary sensory and autonomic neuropathy type V. hNGFp has identical neurotrophic potency as wild-type human nerve growth factor, but a 10-fold lower pain sensitizing activity. In this study we first mimicked, in the 5xFAD mouse model, the intraparenchymal delivery of hNGFp used in clinical trials and found it to be ineffective in decreasing amyloid-β plaque load. On the contrary, the same dose of hNGFp delivered intranasally, which was widely biodistributed in the brain and did not induce pain, showed a potent anti-amyloidogenic action and rescued synaptic plasticity and memory deficits. We found that hNGFp acts on glial cells, modulating inflammatory proteins such as the soluble TNFα receptor II and the chemokine CXCL12. We further established that the rescuing effect by hNGFp is mediated by CXCL12, as pharmacological inhibition of CXCL12 receptor CXCR4 occludes most of hNGFp effects. These findings have significant therapeutic implications: (i) we established that a widespread exposure of the brain is required for nerve growth factor to fully exert its neuroprotective actions; and (ii) we have identified a new anti-neurodegenerative pathway as a broad target for new therapeutic opportunities for neurodegenerative diseases. PMID:28031222

  14. IL-17A and IL-2-expanded regulatory T cells cooperate to inhibit Th1-mediated rejection of MHC II disparate skin grafts.

    PubMed

    Vokaer, Benoît; Charbonnier, Louis-Marie; Lemaître, Philippe H; Spilleboudt, Chloé; Le Moine, Alain

    2013-01-01

    Several evidences suggest that regulatory T cells (Treg) promote Th17 differentiation. Based on this hypothesis, we tested the effect of IL-17A neutralization in a model of skin transplantation in which long-term graft survival depends on a strong in vivo Treg expansion induced by transient exogenous IL-2 administration. As expected, IL-2 supplementation prevented rejection of MHC class II disparate skin allografts but, surprisingly, not in IL-17A-deficient recipients. We attested that IL-17A was not required for IL-2-mediated Treg expansion, intragraft recruitment or suppressive capacities. Instead, IL-17A prevented allograft rejection by inhibiting Th1 alloreactivity independently of Tregs. Indeed, T-bet expression of naive alloreactive CD4+ T cells and the subsequent Th1 immune response was significantly enhanced in IL-17A deficient mice. Our results illustrate for the first time a protective role of IL-17A in CD4+-mediated allograft rejection process.

  15. Hammerhead Ribozyme-Mediated Knockdown of mRNA for Fibrotic Growth Factors: Transforming Growth Factor-Beta 1 and Connective Tissue Growth Factor

    PubMed Central

    Robinson, Paulette M.; Blalock, Timothy D.; Yuan, Rong; Lewin, Alfred S.; Schultz, Gregory S.

    2013-01-01

    Excessive scarring (fibrosis) is a major cause of pathologies in multiple tissues, including lung, liver, kidney, heart, cornea, and skin. The transforming growth factor- β (TGF- β) system has been shown to play a key role in regulating the formation of scar tissue throughout the body. Furthermore, connective tissue growth factor (CTGF) has been shown to mediate most of the fibrotic actions of TGF- β, including stimulation of synthesis of extracellular matrix and differentiation of fibroblasts into myofibroblasts. Currently, no approved drugs selectively and specifically regulate scar formation. Thus, there is a need for a drug that selectively targets the TGF- β cascade at the molecular level and has minimal off-target side effects. This chapter focuses on the design of hammerhead ribozymes, measurement of kinetic activity, and assessment of knockdown mRNAs of TGF- β and CTGF in cell cultures. PMID:22131029

  16. Dnmt2/Trdmt1 as Mediator of RNA Polymerase II Transcriptional Activity in Cardiac Growth

    PubMed Central

    Polo, Beatrice; Baudouy, Delphine; Kiani, Jafar; Michiels, Jean-François; Cuzin, François; Rassoulzadegan, Minoo

    2016-01-01

    Dnmt2/Trdmt1 is a methyltransferase, which has been shown to methylate tRNAs. Deficient mutants were reported to exhibit various, seemingly unrelated, defects in development and RNA-mediated epigenetic heredity. Here we report a role in a distinct developmental regulation effected by a noncoding RNA. We show that Dnmt2-deficiency in mice results in cardiac hypertrophy. Echocardiographic measurements revealed that cardiac function is preserved notwithstanding the increased dimensions of the organ due to cardiomyocyte enlargement. Mechanistically, activation of the P-TEFb complex, a critical step for cardiac growth, results from increased dissociation of the negatively regulating Rn7sk non-coding RNA component in Dnmt2-deficient cells. Our data suggest that Dnmt2 plays an unexpected role for regulation of cardiac growth by modulating activity of the P-TEFb complex. PMID:27270731

  17. Antimony mediated growth of high-density InAs quantum dots for photovoltaic cells

    SciTech Connect

    Tutu, F. K.; Wu, J.; Lam, P.; Tang, M.; Liu, H.; Miyashita, N.; Okada, Y.; Wilson, J.; Allison, R.

    2013-07-22

    We report enhanced solar cell performance using high-density InAs quantum dots. The high-density quantum dot was grown by antimony mediated molecular beam epitaxy. In-plane quantum dot density over 1 × 10{sup 11} cm{sup −2} was achieved by applying a few monolayers of antimony on the GaAs surface prior to quantum dot growth. The formation of defective large clusters was reduced by optimization of the growth temperature and InAs coverage. Comparing with a standard quantum dot solar cell without the incorporation of antimony, the high-density quantum dot solar cell demonstrates a distinct improvement in short-circuit current from 7.4 mA/cm{sup 2} to 8.3 mA/cm{sup 2}.

  18. Seed-mediated growth of ZnO nanorods on multiwalled carbon nanotubes.

    PubMed

    Li, Changqing; Jin, Zhong; Chu, Haibin; Li, Yan

    2008-09-01

    The heterostructures of ZnO nanorods on multiwalled carbon nanotubes (MWNTs) were fabricated by a seed-mediated growth method. First, the surfaces of the carbon nanotubes (CNTs) were coated in situ with mono-dispersed ZnO nanocrystals of about 7 nm by the reaction of zinc acetate and sodium hydroxide. These nanocrystals were then served as the seeds for further growth of ZnO nanorods. In the second step, ZnO nanorods were grown on MWNTs coated with ZnO nanocrystals in an aqueous solution of zinc nitrate and equimolar hexamethylenetetramine at 85 degrees C. Typically, the ZnO nanorods had the length of 300-600 nm and the diameter of 40-140 nm and took a random direction on the outside walls of MWNTs. The morphology of the ZnO nanorods was dependent on pH, reactant concentration, and growing time.

  19. Thyroid hormone-responsive genes mediate otolith growth and development during flatfish metamorphosis.

    PubMed

    Wang, X; Tan, Y; Sievers, Q; Sievers, B; Lee, M; Burrall, K; Schreiber, A M

    2011-01-01

    Flatfish begin life as up-right swimming, bilaterally symmetrical larvae that metamorphose into asymmetrically shaped juveniles that swim with a highly lateralized posture. We have previously shown that TH induces abrupt growth and mineralization of one component of the vestibular system, the otoliths, during early larval development and metamorphosis. Here we report that four of five vestibular-specific genes that we tested (alpha-tectorin, otogelin, otolith matrix protein, and otopetrins 1 and 2 that are known to be associated with otolith development in other vertebrates are up-regulated 1.5- to 7-fold in larval flatfish during spontaneous metamorphosis and/or following 72 h of TH treatment. These findings suggest that otolith growth and development are mediated by diverse TH-responsive genes during flatfish metamorphosis.

  20. Real-time imaging elucidates the role of H2O2 in regulating kinetics of epidermal growth factor-induced and Src-mediated tyrosine phosphorylation signaling

    NASA Astrophysics Data System (ADS)

    Su, Ting; Li, Xiangyong; Liu, Nisha; Pan, Shaotao; Lu, Jinling; Yang, Jie; Zhang, Zhihong

    2012-07-01

    Reversible oxidation is emerging as an important regulatory mechanism in protein tyrosine phosphorylation. Generation of hydrogen peroxide (H2O2), upon growth factor stimulation, is hypothesized to inhibit activity of protein tyrosine phosphatases (PTPs). This ensures that protein tyrosine kinases can elevate the steady-state level of protein tyrosine phosphorylation, which then allows propagation of the tyrosine phosphorylation signal. However, the effects of H2O2 on the kinetics of tyrosine phosphorylation signaling remain poorly understood, especially in living cells. Therefore, we used a genetically encoded Src kinase-specific biosensor based on fluorescence resonance energy transfer (FRET) to image the kinetics of the Src-mediated tyrosine phosphorylation signaling (Src signaling) induced by epidermal growth factor (EGF). We examined the kinetics under increased and decreased H2O2 levels. Through a straightforward, quantitative analysis method which characterized the signaling kinetics, we demonstrated that H2O2 modulated the amplitude and duration of the signal by inhibiting PTPs' activity. Our evidence also suggested the effect of H2O2 on Src activation is mediated by H2O2-dependent inhibition of PTPs. Furthermore, we provide evidence showing global elevation of intracellular H2O2 level attenuates EGF-induced Src signaling.

  1. The Involvement of Gibberellins in 1,8-Cineole-Mediated Inhibition of Sprout Growth in Russet Burbank Tubers

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The involvement of gibberellins in 1,8-cineole-mediated inhibition of tuber sprout growth was investigated in non-dormant field- and greenhouse-grown tubers of Russet Burbank. Continuous exposure of tubers to cineole in the vapor-phase resulted in a dose-dependent inhibition of sprout growth. Comp...

  2. Insect growth regulatory effects of some extracts and sterols from Myrtillocactus geometrizans (Cactaceae) against Spodoptera frugiperda and Tenebrio molitor.

    PubMed

    Céspedes, Carlos L; Salazar, J Rodrigo; Martínez, Mariano; Aranda, Eduardo

    2005-10-01

    A methanol extract from the roots and aerial parts of Myrtillocactus geometrizans (Cactaceae) yielded peniocerol 1, macdougallin 2, and chichipegenin 3. The natural products 1, 2 their mixtures, MeOH and CH(2)Cl(2) extracts showed insecticidal and insect growth regulatory activity against fall armyworm [Spodoptera frugiperda J. E. Smith (Lepidoptera: Noctuidae)], an important insect pest of corn, and [Tenebrio molitor (Coleoptera)], a pest of stored grains in Mexico. The most active compounds were 1, 2, and a mixture (M(2)) of 1 and 2 (6:4). All these extracts, compounds and the mixture had insect growth regulating (IGR) activity between 5.0 and 50.0 ppm and insecticidal effects between 50 and 300 ppm in diets. The extracts were insecticidal to larvae, with lethal doses between 100 and 200 ppm. These compounds appear to have selective effects on the pre-emergence metabolism of Coleoptera, because in all treatments of the larvae of T. molitor, pupation were shortened and this process show precociousness in relation to controls. In contrast to S. frugiperda larvae, onset of pupation was noticeably delayed. Emergence in both cases was drastically diminished. In both pupae and in the few adults that were able to emerge, many deformations were observed. The results of these assays indicated that the compounds were more active than other known natural insect growth inhibitors such as gedunin and methanol extracts of Cedrela salvadorensis and Yucca periculosa. Peniocerol, macdougallin and chichipegenin, as well as mixtures of these substances, may be useful as natural insecticidal agents.

  3. Caudatin Inhibits Human Glioma Cells Growth Through Triggering DNA Damage-Mediated Cell Cycle Arrest.

    PubMed

    Fu, Xiao-yan; Zhang, Shuai; Wang, Kun; Yang, Ming-feng; Fan, Cun-dong; Sun, Bao-liang

    2015-10-01

    Caudatin, one of the species of C-21 steroidal glycosides mainly isolated from the root of Cynanchum bungei Decne, exhibits potent anticancer activities. However, the mechanism remains poorly defined. In the present study, the growth inhibitory effect and mechanism of caudatin on human glioma cells were evaluated in vitro. The results revealed that caudatin time- and dose-dependently inhibited U251 and U87 cells growth. Flow cytometry analysis indicated that caudatin-induced growth inhibition against U251 and U87 cells was mainly achieved by the induction of G0/G1 and S-phase cell cycle arrest through triggering DNA damage, as convinced by the up-regulation of p53, p21, and histone phosphorylation, as well as the down-regulation of cyclin D1. Moreover, caudatin treatment also triggered the activation of ERK and inactivation of AKT pathway. LY294002 (an AKT inhibitor) addition enhanced caudation-induced AKT inhibition, indicating that caudatin inhibited U251 cells growth in an AKT-dependent manner. Taken together, these results indicate that caudatin may act as a novel cytostatic reagent against human glioma cells through the induction of DNA damage-mediated cell cycle arrest with the involvement of modulating MAPK and AKT pathways.

  4. Does social status within a dominance hierarchy mediate individual growth, residency and relocation?

    PubMed

    Akbaripasand, Abbas; Ramezani, J; Krkosek, Martin; Lokman, P Mark; Closs, Gerard P

    2014-11-01

    The availability of food, and hence energy, is known to influence the abundance, habitat choice and growth of individuals. In contrast, there is a paucity of knowledge on how the interaction of energy supply and social status determines patterns of residency and movement. This study tests whether the presence of conspecifics and an individual's social status in relation to food supply influence the fitness and movement of a drift-feeding fish (Galaxias fasciatus). Using an information-theoretic approach (AIC), our analysis indicated that the most parsimonious model of fish movement among pools was one that included food supply, social rank and fish relative growth rate. Our results indicated that subordinate fish relocated more frequently compared to dominant fish, most likely as a consequence of intra-specific competition that limited the access of these smaller fish to resources and constrained their growth. Our results suggest that energy constraints may force individuals to explore new habitats in an effort to find more energetically profitable patches. We conclude that intra-specific competition mediated through the social hierarchy amongst closely interacting individuals plays a key role in determining individual growth, residency and relocation.

  5. Human Sarcoma Growth Is Sensitive to Small-Molecule Mediated AXIN Stabilization

    PubMed Central

    Rossi, Marco; Valensin, Silvia; Tunici, Patrizia; Mori, Elisa; Caradonna, Nicola; Varrone, Maurizio; Salerno, Massimiliano

    2014-01-01

    Sarcomas are mesenchymal tumors showing high molecular heterogeneity, reflected at the histological level by the existence of more than fifty different subtypes. Genetic and epigenetic evidences link aberrant activation of the Wnt signaling to growth and progression of human sarcomas. This phenomenon, mainly accomplished by autocrine loop activity, is sustained by gene amplification, over-expression of Wnt ligands and co-receptors or epigenetic silencing of endogenous Wnt antagonists. We previously showed that pharmacological inhibition of Wnt signaling mediated by Axin stabilization produced in vitro and in vivo antitumor activity in glioblastoma tumors. Here, we report that targeting different sarcoma cell lines with the Wnt inhibitor/Axin stabilizer SEN461 produces a less transformed phenotype, as supported by modulation of anchorage-independent growth in vitro. At the molecular level, SEN461 treatment enhanced the stability of the scaffold protein Axin1, a key negative regulator of the Wnt signaling with tumor suppressor function, resulting in downstream effects coherent with inhibition of canonical Wnt signaling. Genetic phenocopy of small molecule Axin stabilization, through Axin1 over-expression, coherently resulted in strong impairment of soft-agar growth. Importantly, sarcoma growth inhibition through pharmacological Axin stabilization was also observed in a xenograft model in vivo in female CD-1 nude mice. Our findings suggest the usefulness of Wnt inhibitors with Axin stabilization activity as a potentialyl clinical relevant strategy for certain types of sarcomas. PMID:24842792

  6. Yes is a central mediator of cell growth in malignant mesothelioma cells.

    PubMed

    Sato, Ayami; Sekine, Miki; Virgona, Nantiga; Ota, Masako; Yano, Tomohiro

    2012-11-01

    The constitutive activation of the Src family kinases (SFKs) has been established as a poor prognostic factor in malignant mesothelioma (MM), however, the family member(s) which contribute to the malignancy have not been defined. This study aimed to identify the SFK member(s) contributing to cell growth using RNA interference in various MM cell lines. Silencing of Yes but not of c-Src or Fyn in MM cells leads to cell growth suppression. This suppressive effect caused by Yes silencing mainly depends on G1 cell cycle arrest and partly the induction of apoptosis. Also, the knockout of Yes induces the inactivation of β-catenin signaling and subsequently decreases the levels of cyclin D necessary for G1-S transition in the cell cycle. In addition, Yes knockout has less effect on cell growth suppression in β-catenin-deficient H28 MM cells compared to other MM cells which express the catenin. Overall, we conclude that Yes is a central mediator for MM cell growth that is not shared with other SFKs such as c-Src.

  7. Brevundimonas diminuta mediated alleviation of arsenic toxicity and plant growth promotion in Oryza sativa L.

    PubMed

    Singh, Namrata; Marwa, Naina; Mishra, Shashank K; Mishra, Jyoti; Verma, Praveen C; Rathaur, Sushma; Singh, Nandita

    2016-03-01

    Arsenic (As), a toxic metalloid adversely affects plant growth in polluted areas. In the present study, we investigated the possibility of improving phytostablization of arsenic through application of new isolated strain Brevundimonas diminuta (NBRI012) in rice plant [Oryza sativa (L.) Var. Sarju 52] at two different concentrations [10ppm (low toxic) and 50ppm (high toxic)] of As. The plant growth promoting traits of bacterial strains revealed the inherent ability of siderophores, phosphate solubilisation, indole acetic acid (IAA), 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase production which may be associated with increased biomass, chlorophyll and MDA content of rice and thereby promoting plant growth. The study also revealed the As accumulation property of NBRI012 strain which could play an important role in As removal from contaminated soil. Furthermore, NBRI012 inoculation significantly restored the hampered root epidermal and cortical cell growth of rice plant and root hair elimination. Altogether our study highlights the multifarious role of B. diminuta in mediating stress tolerance and modulating translocation of As in edible part of rice plant.

  8. Wip1 inhibitor GSK2830371 inhibits neuroblastoma growth by inducing Chk2/p53-mediated apoptosis

    PubMed Central

    Chen, Zhenghu; Wang, Long; Yao, Dayong; Yang, Tianshu; Cao, Wen-Ming; Dou, Jun; Pang, Jonathan C.; Guan, Shan; Zhang, Huiyuan; Yu, Yang; Zhao, Yanling; Wang, Yongfeng; Xu, Xin; Shi, Yan; Patel, Roma; Zhang, Hong; Vasudevan, Sanjeev A.; Liu, Shangfeng; Yang, Jianhua; Nuchtern, Jed G.

    2016-01-01

    Neuroblastoma (NB) is the most common extracranial tumor in children. Unlike in most adult tumors, tumor suppressor protein 53 (p53) mutations occur with a relatively low frequency in NB and the downstream function of p53 is intact in NB cell lines. Wip1 is a negative regulator of p53 and hindrance of Wip1 activity by novel inhibitor GSK2830371 is a potential strategy to activate p53’s tumor suppressing function in NB. Yet, the in vivo efficacy and the possible mechanisms of GSK2830371 in NB have not yet been elucidated. Here we report that novel Wip1 inhibitor GSK2830371 induced Chk2/p53-mediated apoptosis in NB cells in a p53-dependent manner. In addition, GSK2830371 suppressed the colony-formation potential of p53 wild-type NB cell lines. Furthermore, GSK2830371 enhanced doxorubicin- (Dox) and etoposide- (VP-16) induced cytotoxicity in a subset of NB cell lines, including the chemoresistant LA-N-6 cell line. More importantly, GSK2830371 significantly inhibited tumor growth in an orthotopic xenograft NB mouse model by inducing Chk2/p53-mediated apoptosis in vivo. Taken together, this study suggests that GSK2830371 induces Chk2/p53-mediated apoptosis both in vitro and in vivo in a p53 dependent manner. PMID:27991505

  9. Platelet-derived growth factor-BB-mediated glycosaminoglycan synthesis is transduced through Akt.

    PubMed

    Cartel, Nicholas J; Wang, Jinxia; Post, Martin

    2002-04-01

    Previously we have demonstrated that the phosphoinositide 3-kinase (PI-3K) signal-transduction pathway mediates platelet-derived growth factor (PDGF)-BB-induced glycosaminoglycan (GAG) synthesis in fetal lung fibroblasts. In the present study we further investigated the signal-transduction pathway(s) that results in PDGF-BB-induced GAG synthesis. Over-expression of a soluble PDGF beta-receptor as well as a mutated form of the beta-receptor, unable to bind PI-3K, diminished GAG synthesis in fetal lung fibroblasts subsequent to PDGF-BB stimulation. The PI-3K inhibitor wortmannin blocked PDGF-BB-induced Akt activity as well as significantly diminishing PDGF-BB-mediated GAG synthesis. Expression of dominant-negative PI-3K also abrogated Akt activity and GAG synthesis. Furthermore, expression of dominant-negative Akt abrogated endogenous Akt activity, Rab3D phosphorylation and GAG synthesis, whereas expression of constitutively activated Akt stimulated Rab3D phosphorylation and GAG synthesis in the absence of PDGF-BB. Over-expression of wild-type PTEN (phosphatase and tensin homologue deleted in chromosome 10) inhibited Akt activity and concomitantly attenuated GAG synthesis in fibroblasts stimulated with PDGF-BB. These data suggest that Akt is an integral protein involved in PDGF-BB-mediated GAG regulation in fetal lung fibroblasts.

  10. Inhibition of tumor cell growth by Sigma1 ligand mediated translational repression

    SciTech Connect

    Kim, Felix J.; Schrock, Joel M.; Spino, Christina M.; Marino, Jacqueline C.; Pasternak, Gavril W.

    2012-09-21

    Highlights: Black-Right-Pointing-Pointer Sigma1 ligand treatment mediates decrease in tumor cell mass. Black-Right-Pointing-Pointer Identification of a Sigma1 ligand with reversible translational repressor actions. Black-Right-Pointing-Pointer Demonstration of a role for Sigma1 in cellular protein synthesis. -- Abstract: Treatment with sigma1 receptor (Sigma1) ligands can inhibit cell proliferation in vitro and tumor growth in vivo. However, the cellular pathways engaged in response to Sigma1 ligand treatment that contribute to these outcomes remain largely undefined. Here, we show that treatment with putative antagonists of Sigma1 decreases cell mass. This effect corresponds with repressed cap-dependent translation initiation in multiple breast and prostate cancer cell lines. Sigma1 antagonist treatment suppresses phosphorylation of translational regulator proteins p70S6K, S6, and 4E-BP1. RNAi-mediated knockdown of Sigma1 also results in translational repression, consistent with the effects of antagonist treatment. Sigma1 antagonist mediated translational repression and decreased cell size are both reversible. Together, these data reveal a role for Sigma1 in tumor cell protein synthesis, and demonstrate that small molecule Sigma1 ligands can be used as modulators of protein translation.

  11. The Wedelolactone Derivative Inhibits Estrogen Receptor-Mediated Breast, Endometrial, and Ovarian Cancer Cells Growth

    PubMed Central

    Xu, Defeng; Lin, Tzu-Hua; Cheng, Max A.; Chen, Lu-Min; Chang, Chawnshang; Yeh, Shuyuan

    2014-01-01

    Estrogen and estrogen receptor (ER)-mediated signaling pathways play important roles in the etiology and progression of human breast, endometrial, and ovarian cancers. Attenuating ER activities by natural products and their derivatives is a relatively practical strategy to control and reduce breast, endometrial, and ovarian cancer risk. Here, we found 3-butoxy-1,8,9-trihydroxy-6H-benzofuro[3,2-c]benzopyran-6-one (BTB), a new derivative of wedelolactone, could effectively inhibit the 17-estradiol (E2)-induced ER transactivation and suppress the growth of breast cancer as well as endometrial and ovarian cancer cells. Our results indicate that 2.5 μM BTB effectively suppresses ER-positive, but not ER-negative, breast, endometrial, and ovarian cancer cells. Furthermore, our data indicate that BTB can modulate ER transactivation and suppress the expression of E2-mediated ER target genes (Cyclin D1, E2F1, and TERT) in the ER-positive MCF-7, Ishikawa, and SKOV-3 cells. Importantly, this BTB mediated inhibition of ER activity is selective since BTB does not suppress the activities of other nuclear receptors, including glucocorticoid receptor and progesterone receptor, suggesting that BTB functions as a selective ER signaling inhibitor with the potential to treat breast, endometrial, and ovarian cancers. PMID:25221777

  12. Ethylene-Mediated Regulation of A2-Type CYCLINs Modulates Hyponastic Growth in Arabidopsis1[OPEN

    PubMed Central

    Polko, Joanna K.; van Rooij, Jop A.; Vanneste, Steffen; Pierik, Ronald; Ammerlaan, Ankie M.H.; Vergeer-van Eijk, Marleen H.; McLoughlin, Fionn; Gühl, Kerstin; Van Isterdael, Gert; Voesenek, Laurentius A.C.J.; Millenaar, Frank F.; Beeckman, Tom; Peeters, Anton J.M.; Marée, Athanasius F.M.; van Zanten, Martijn

    2015-01-01

    Upward leaf movement (hyponastic growth) is frequently observed in response to changing environmental conditions and can be induced by the phytohormone ethylene. Hyponasty results from differential growth (i.e. enhanced cell elongation at the proximal abaxial side of the petiole relative to the adaxial side). Here, we characterize Enhanced Hyponasty-d, an activation-tagged Arabidopsis (Arabidopsis thaliana) line with exaggerated hyponasty. This phenotype is associated with overexpression of the mitotic cyclin CYCLINA2;1 (CYCA2;1), which hints at a role for cell divisions in regulating hyponasty. Indeed, mathematical analysis suggested that the observed changes in abaxial cell elongation rates during ethylene treatment should result in a larger hyponastic amplitude than observed, unless a decrease in cell proliferation rate at the proximal abaxial side of the petiole relative to the adaxial side was implemented. Our model predicts that when this differential proliferation mechanism is disrupted by either ectopic overexpression or mutation of CYCA2;1, the hyponastic growth response becomes exaggerated. This is in accordance with experimental observations on CYCA2;1 overexpression lines and cyca2;1 knockouts. We therefore propose a bipartite mechanism controlling leaf movement: ethylene induces longitudinal cell expansion in the abaxial petiole epidermis to induce hyponasty and simultaneously affects its amplitude by controlling cell proliferation through CYCA2;1. Further corroborating the model, we found that ethylene treatment results in transcriptional down-regulation of A2-type CYCLINs and propose that this, and possibly other regulatory mechanisms affecting CYCA2;1, may contribute to this attenuation of hyponastic growth. PMID:26041787

  13. The SnRK2-APC/C(TE) regulatory module mediates the antagonistic action of gibberellic acid and abscisic acid pathways.

    PubMed

    Lin, Qibing; Wu, Fuqing; Sheng, Peike; Zhang, Zhe; Zhang, Xin; Guo, Xiuping; Wang, Jiulin; Cheng, Zhijun; Wang, Jie; Wang, Haiyang; Wan, Jianmin

    2015-08-14

    Abscisic acid (ABA) and gibberellic acid (GA) antagonistically regulate many developmental processes and responses to biotic or abiotic stresses in higher plants. However, the molecular mechanism underlying this antagonism is still poorly understood. Here, we show that loss-of-function mutation in rice Tiller Enhancer (TE), an activator of the APC/C(TE) complex, causes hypersensitivity and hyposensitivity to ABA and GA, respectively. We find that TE physically interacts with ABA receptor OsPYL/RCARs and promotes their degradation by the proteasome. Genetic analysis also shows OsPYL/RCARs act downstream of TE in mediating ABA responses. Conversely, ABA inhibits APC/C(TE) activity by phosphorylating TE through activating the SNF1-related protein kinases (SnRK2s), which may interrupt the interaction between TE and OsPYL/RCARs and subsequently stabilize OsPYL/RCARs. In contrast, GA can reduce the level of SnRK2s and may promote APC/C(TE)-mediated degradation of OsPYL/RCARs. Thus, we propose that the SnRK2-APC/C(TE) regulatory module represents a regulatory hub underlying the antagonistic action of GA and ABA in plants.

  14. Tyk2 mediates effects of urokinase on human vascular smooth muscle cell growth

    SciTech Connect

    Patecki, Margret; Schaewen, Markus von; Tkachuk, Sergey; Jerke, Uwe; Dietz, Rainer; Dumler, Inna; Kusch, Angelika . E-mail: angelika.kusch@charite.de

    2007-08-03

    The urokinase (uPA)/uPA receptor (uPAR) system plays a role in the response of the vessel wall to injury, presumably by modulating vascular smooth muscle cell (VSMC) functional behaviour. The Jak/Stat signaling pathway has been implicated to mediate the uPA/uPAR-directed cell migration and proliferation in VSMC. We have therefore investigated the underlying molecular mechanisms, which remained not completely understood. In particular, we aimed at identification of the kinase involved in the signaling cascade leading to Stat1 phosphorylation by uPA and its impact on VSMC growth. We performed expression in VSMC of kinase-deficient mutant forms of the Janus kinases Jak1 and Tyk2 and used different cell culture models imitating the response to vascular injury. We provide evidence that Tyk2, but not Jak1, mediates uPA-induced Stat1 phosphorylation and VSMC growth inhibition and suggest a novel function for Tyk2 as an important modulator of the uPA-directed VSMC functional behaviour at the place of injury.

  15. Tyk2 mediates effects of urokinase on human vascular smooth muscle cell growth.

    PubMed

    Patecki, Margret; von Schaewen, Markus; Tkachuk, Sergey; Jerke, Uwe; Dietz, Rainer; Dumler, Inna; Kusch, Angelika

    2007-08-03

    The urokinase (uPA)/uPA receptor (uPAR) system plays a role in the response of the vessel wall to injury, presumably by modulating vascular smooth muscle cell (VSMC) functional behaviour. The Jak/Stat signaling pathway has been implicated to mediate the uPA/uPAR-directed cell migration and proliferation in VSMC. We have therefore investigated the underlying molecular mechanisms, which remained not completely understood. In particular, we aimed at identification of the kinase involved in the signaling cascade leading to Stat1 phosphorylation by uPA and its impact on VSMC growth. We performed expression in VSMC of kinase-deficient mutant forms of the Janus kinases Jak1 and Tyk2 and used different cell culture models imitating the response to vascular injury. We provide evidence that Tyk2, but not Jak1, mediates uPA-induced Stat1 phosphorylation and VSMC growth inhibition and suggest a novel function for Tyk2 as an important modulator of the uPA-directed VSMC functional behaviour at the place of injury.

  16. Amplification of TLO Mediator Subunit Genes Facilitate Filamentous Growth in Candida Spp.

    PubMed Central

    Liu, Zhongle; Moran, Gary P.; Myers, Lawrence C.

    2016-01-01

    Filamentous growth is a hallmark of C. albicans pathogenicity compared to less-virulent ascomycetes. A multitude of transcription factors regulate filamentous growth in response to specific environmental cues. Our work, however, suggests the evolutionary history of C. albicans that resulted in its filamentous growth plasticity may be tied to a change in the general transcription machinery rather than transcription factors and their specific targets. A key genomic difference between C. albicans and its less-virulent relatives, including its closest relative C. dubliniensis, is the unique expansion of the TLO (TeLOmere-associated) gene family in C. albicans. Individual Tlo proteins are fungal-specific subunits of Mediator, a large multi-subunit eukaryotic transcriptional co-activator complex. This amplification results in a large pool of ‘free,’ non-Mediator associated, Tlo protein present in C. albicans, but not in C. dubliniensis or other ascomycetes with attenuated virulence. We show that engineering a large ‘free’ pool of the C. dubliniensis Tlo2 (CdTlo2) protein in C. dubliniensis, through overexpression, results in a number of filamentation phenotypes typically associated only with C. albicans. The amplitude of these phenotypes is proportional to the amount of overexpressed CdTlo2 protein. Overexpression of other C. dubliniensis and C. albicans Tlo proteins do result in these phenotypes. Tlo proteins and their orthologs contain a Mediator interaction domain, and a potent transcriptional activation domain. Nuclear localization of the CdTlo2 activation domain, facilitated naturally by the Tlo Mediator binding domain or artificially through an appended nuclear localization signal, is sufficient for the CdTlo2 overexpression phenotypes. A C. albicans med3 null mutant causes multiple defects including the inability to localize Tlo proteins to the nucleus and reduced virulence in a murine systemic infection model. Our data supports a model in which the

  17. Inhibitor of growth-4 mediates chromatin modification and has a suppressive effect on tumorigenesis and innate immunity.

    PubMed

    Mathema, Vivek Bhakta; Koh, Young-Sang

    2012-02-01

    Inhibitor of growth-4 (ING4) is a member of the ING family and acts as a tumor suppressor protein. ING4 is a promising candidate for cancer research due to its anti-angiogenic function and its role in the inhibition of cell migration, cell cycle, and induction of apoptosis. Interaction of this protein with the histone acetyl transferase complex plays a vital role in the regulation of multiple nuclear factor kappa light chain enhancer of activated B cells response elements and thus in the regulation of innate immunity. Splice variants of ING4 have different binding affinities to target sites, which results in the enhancement of its functional diversity. ING4 is among the few known regulatory proteins that can directly interact with chromatin as well as with transcription factors. The influence of ING4 on tumor necrosis factor-α, keratinocyte chemoattractant, interleukin (IL)-6, IL-8, matrix metalloproteinases, cyclooxygenase-2, and IκBα expression clearly demonstrates its critical role in the regulation of inflammatory mediators. Its interaction with liprin α1 and p53 contribute to mitigate cell spreading and induce apoptosis of cancer cells. Multiple factors including breast cancer melanoma suppressor-1 are upstream regulators of ING4 and are frequently deactivated in tumor cells. In the present review, the different properties of ING4 are discussed, and its activities are correlated with different aspects of cell physiology. Special emphasis is placed on our current understanding of ING4 with respect to its influence on chromatin modification, tumorigenesis, and innate immunity.

  18. Identification of a regulatory subunit of protein phosphatase 1 which mediates blue light signaling for stomatal opening.

    PubMed

    Takemiya, Atsushi; Yamauchi, Shota; Yano, Takayuki; Ariyoshi, Chie; Shimazaki, Ken-ichiro

    2013-01-01

    Protein phosphatase 1 (PP1) is a eukaryotic serine/threonine protein phosphatase comprised of a catalytic subunit (PP1c) and a regulatory subunit that modulates catalytic activity, subcellular localization and substrate specificity. PP1c positively regulates stomatal opening through blue light signaling between phototropins and the plasma membrane H(+)-ATPase in guard cells. However, the regulatory subunit functioning in this process is unknown. We identified Arabidopsis PRSL1 (PP1 regulatory subunit2-like protein1) as a regulatory subunit of PP1c. Tautomycin, a selective inhibitor of PP1c, inhibited blue light responses of stomata in the single mutants phot1 and phot2, supporting the idea that signals from phot1 and phot2 converge on PP1c. We obtained PRSL1 based on the sequence similarity to Vicia faba PRS2, a PP1c-binding protein isolated by a yeast two-hybrid screen. PRSL1 bound to Arabidopsis PP1c through its RVxF motif, a consensus PP1c-binding sequence. Arabidopsis prsl1 mutants were impaired in blue light-dependent stomatal opening, H(+) pumping and phosphorylation of the H(+)-ATPase, but showed normal phototropin activities. PRSL1 complemented the prsl1 phenotype, but not if the protein carried a mutation in the RVxF motif, suggesting that PRSL1 functions through binding PP1c via the RVxF motif. PRSL1 did not affect the catalytic activity of Arabidopsis PP1c but it stimulated the localization of PP1c in the cytoplasm. We conclude that PRSL1 functions as a regulatory subunit of PP1 and regulates blue light signaling in stomata.

  19. Hormone-mediated growth dynamics of the barley pericarp as revealed by magnetic resonance imaging and transcript profiling

    PubMed Central

    Pielot, Rainer; Kohl, Stefan; Manz, Bertram; Rutten, Twan; Weier, Diana; Tarkowská, Danuše; Rolčík, Jakub; Strnad, Miroslav; Volke, Frank; Weber, Hans

    2015-01-01

    The shape of the maternal pericarp affects cereal grain mass and yield. Pericarp growth was analysed by magnetic resonance imaging (MRI), revealing topological maps of mobile water in developing pericarp of barley (Hordeum vulgare) and displaying tissue regions actively elongating in specific temporal–spatial patterns. Correlation analysis of MRI signals and growth rates reveals that growth in length is mediated by dorsal and also lateral rather than ventral regions. Growth in thickness is related to ventral regions. Switching from dorsal to ventral growth is associated with differential expression of axial regulators of the HD-ZipIII and Kanadi/Ettin types, and NPH3 photoreceptors, suggesting light-mediated auxin re-distribution. Auxin increases with the highest levels in the basal pericarp at 6 days after fertilization (DAF), together with transcriptionally up-regulated auxin transport and signalling. Gibberellin biosynthesis is transcriptionally up-regulated only later, and levels of bioactive gibberellins increase from 7 to 13 DAF, with higher levels in ventral than dorsal regions. Differential gene expression related to cell expansion indicates genes related to apoplast acidification, wall relaxation, sugar cleavage, water transport, and cell wall biosynthesis. Candidate genes potentially involved in pericarp extension are distinguished by their temporal expression, representing potential isoforms responsible for dorsal-mediated early growth in length or ventral-mediated late growth in thickness. PMID:26276866

  20. Persistent Nociception Triggered by Nerve Growth Factor (NGF) Is Mediated by TRPV1 and Oxidative Mechanisms.

    PubMed

    Eskander, Michael A; Ruparel, Shivani; Green, Dustin P; Chen, Paul B; Por, Elaine D; Jeske, Nathaniel A; Gao, Xiaoli; Flores, Eric R; Hargreaves, Kenneth M

    2015-06-03

    Nerve growth factor (NGF) is elevated in certain chronic pain conditions and is a sufficient stimulus to cause lasting pain in humans, but the actual mechanisms underlying the persistent effects of NGF remain incompletely understood. We developed a rat model of NGF-induced persistent thermal hyperalgesia and mechanical allodynia to determine the role of transient receptor potential vanilloid 1 (TRPV1) and oxidative mechanisms in the persistent effects of NGF. Persistent thermal hypersensitivity and mechanical allodynia require de novo protein translation and are mediated by TRPV1 and oxidative mechanisms. By comparing effects after systemic (subcutaneous), spinal (intrathecal) or hindpaw (intraplantar) injections of test compounds, we determined that TRPV1 and oxidation mediate persistent thermal hypersensitivity via peripheral and spinal sites of action and mechanical allodynia via only a spinal site of action. Therefore, NGF-evoked thermal and mechanical allodynia are mediated by spatially distinct mechanisms. NGF treatment evoked sustained increases in peripheral and central TRPV1 activity, as demonstrated by increased capsaicin-evoked nocifensive responses, increased calcitonin gene-related peptide release from hindpaw skin biopsies, and increased capsaicin-evoked inward current and membrane expression of TRPV1 protein in dorsal root ganglia neurons. Finally, we showed that NGF treatment increased concentrations of linoleic and arachidonic-acid-derived oxidized TRPV1 agonists in spinal cord and skin biopsies. Furthermore, increases in oxidized TRPV1-active lipids were reduced by peripheral and spinal injections of compounds that completely blocked persistent nociception. Collectively, these data indicate that NGF evokes a persistent nociceptive state mediated by increased TRPV1 activity and oxidative mechanisms, including increased production of oxidized lipid TRPV1 agonists.

  1. Stimulatory effects of insulin-like growth factor-I on growth plate chondrogenesis are mediated by nuclear factor-kappaB p65.

    PubMed

    Wu, Shufang; Fadoju, Doris; Rezvani, Geoffrey; De Luca, Francesco

    2008-12-05

    Insulin-like growth factor-I (IGF-I) is an important regulator of endochondral ossification. However, little is known about the signaling pathways activated by IGF-I in growth plate chondrocytes. We have previously shown that NF-kappaB-p65 facilitates growth plate chondrogenesis. In this study, we first cultured rat metatarsal bones with IGF-I and/or pyrrolidine dithiocarbamate (PDTC), a known NF-kappaB inhibitor. The IGF-I-mediated stimulation of metatarsal growth and growth plate chondrogenesis was neutralized by PDTC. In rat growth plate chondrocytes, IGF-I induced NF-kappaB-p65 nuclear translocation. The inhibition of NF-kappaB-p65 expression and activity (by p65 short interfering RNA and PDTC, respectively) in chondrocytes reversed the IGF-I-mediated induction of cell proliferation and differentiation and the IGF-I-mediated prevention of cell apoptosis. Moreover, the inhibition of the phosphatidylinositol 3-kinase and Akt abolished the effects of IGF-I on NF-kappaB activation. In conclusion, our findings indicate that IGF-I stimulates growth plate chondrogenesis by activating NF-kappaB-p65 in chondrocytes.

  2. Eugenol-inhibited root growth in Avena fatua involves ROS-mediated oxidative damage.

    PubMed

    Ahuja, Nitina; Singh, Harminder Pal; Batish, Daizy Rani; Kohli, Ravinder Kumar

    2015-02-01

    Plant essential oils and their constituent monoterpenes are widely known plant growth retardants but their mechanism of action is not well understood. We explored the mechanism of phytotoxicity of eugenol, a monoterpenoid alcohol, proposed as a natural herbicide. Eugenol (100-1000 µM) retarded the germination of Avena fatua and strongly inhibited its root growth compared to the coleoptile growth. We further investigated the underlying physiological and biochemical alterations leading to the root growth inhibition. Eugenol induced the generation of reactive oxygen species (ROS) leading to oxidative stress and membrane damage in the root tissue. ROS generation measured in terms of hydrogen peroxide, superoxide anion and hydroxyl radical content increased significantly in the range of 24 to 144, 21 to 91, 46 to 173% over the control at 100 to 1000 µM eugenol, respectively. The disruption in membrane integrity was indicated by 25 to 125% increase in malondialdehyde (lipid peroxidation byproduct), and decreased conjugated diene content (~10 to 41%). The electrolyte leakage suggesting membrane damage increased both under light as well as dark conditions measured over a period from 0 to 30 h. In defense to the oxidative damage due to eugenol, a significant upregulation in the ROS-scavenging antioxidant enzyme machinery was observed. The activities of superoxide dismutases, catalases, ascorbate peroxidases, guaiacol peroxidases and glutathione reductases were elevated by ~1.5 to 2.8, 2 to 4.3, 1.9 to 5.0, 1.4 to 3.9, 2.5 to 5.5 times, respectively, in response to 100 to 1000 µM eugenol. The study concludes that eugenol inhibits early root growth through ROS-mediated oxidative damage, despite an activation of the antioxidant enzyme machinery.

  3. Activated carbon decreases invasive plant growth by mediating plant–microbe interactions

    PubMed Central

    Nolan, Nicole E.; Kulmatiski, Andrew; Beard, Karen H.; Norton, Jeanette M.

    2015-01-01

    There is growing appreciation for the idea that plant–soil interactions (e.g. allelopathy and plant–microbe feedbacks) may explain the success of some non-native plants. Where this is the case, native plant restoration may require management tools that change plant–soil interactions. Activated carbon (AC) is one such potential tool. Previous research has shown the potential for high concentrations of AC to restore native plant growth to areas dominated by non-natives on a small scale (1 m × 1 m plots). Here we (i) test the efficacy of different AC concentrations at a larger scale (15 m × 15 m plots), (ii) measure microbial responses to AC treatment and (iii) use a greenhouse experiment to identify the primary mechanism, allelopathy versus microbial changes, through which AC impacts native and non-native plant growth. Three years after large-scale applications, AC treatments decreased non-native plant cover and increased the ratio of native to non-native species cover, particularly at concentrations >400 g m−2. Activated carbon similarly decreased non-native plant growth in the greenhouse. This effect, however, was only observed in live soils, suggesting that AC effects were microbially mediated and not caused by direct allelopathy. Bacterial community analysis of field soils indicated that AC increased the relative abundance of an unidentified bacterium and an Actinomycetales and decreased the relative abundance of a Flavobacterium, suggesting that these organisms may play a role in AC effects on plant growth. Results support the idea that manipulations of plant–microbe interactions may provide novel and effective ways of directing plant growth and community development (e.g. native plant restoration). PMID:25387751

  4. F8-SIP mediated targeted photodynamic therapy leads to microvascular dysfunction and reduced glioma growth.

    PubMed

    Acker, G; Palumbo, A; Neri, D; Vajkoczy, P; Czabanka, M

    2016-08-01

    The extra domain A (ED A) of fibronectin has been identified as a tumor vessel specific neovascular marker in glioma. Antibody based vascular targeting against ED A of fibronectin allows precise accumulation of photosensitizer in glioma microvasculature and thereby promises to overcome drawbacks of current photodynamic therapy (PDT) for glioma treatment. Our aim was to characterize microcirculatory consequences of F8-small immunoprotein (SIP) mediated PDT by intravital microscopy (IVM) and to analyze the effects on glioma growth. For IVM SF126 glioma cells were implanted into dorsal skinfold-chamber of nude mice. PDT was performed after intravenous injection of photosensitizer (PS)-coupled F8-SIP or PBS (n = 4). IVM was performed before and after PDT for 4 days. Analysis included total and functional (TVD, FVD) vessel densities, perfusion index (PI), microvascular permeability and blood flow rate (Q). To assess tumor growth SF126 glioma cells were implanted subcutaneously. PDT was performed as a single and repetitive treatment after PS-F8-SIP injection (n = 5). Subcutaneous tumors were treated after uncoupled F8-SIP injection as control group (n = 5). PDT induced microvascular stasis and thrombosis with reduced FVD (24 h: 115.98 ± 0.7 vs. 200.8 ± 61.9 cm/cm(2)) and PI (39 ± 11 vs. 70 ± 10 %), whereas TVD was not altered (298 ± 39.2 vs. 278.2 ± 51 cm/cm(2)). Microvascular dysfunction recovered 4 days after treatment. Microvascular dysfunction led to a temporary reduction of glioma growth in the first 48 h after treatment with complete recovery 5 days after treatment. Repetitive PDT resulted in sustained reduction of tumor growth. F8-SIP mediated PDT leads to microvascular dysfunction and reduced glioma growth in a preclinical glioma model with recovery of microcirculation 4 days after treatment. Repetitive application of PDT overcomes microvascular recovery and leads to prolonged antiglioma effects.

  5. Protein Kinase A Regulatory Subunit Isoforms Regulate Growth and Differentiation in Mucor circinelloides: Essential Role of PKAR4

    PubMed Central

    Ocampo, J.; McCormack, B.; Navarro, E.; Moreno, S.; Garre, V.

    2012-01-01

    The protein kinase A (PKA) signaling pathway plays a role in regulating growth and differentiation in the dimorphic fungus Mucor circinelloides. PKA holoenzyme is comprised of two catalytic (C) and two regulatory (R) subunits. In M. circinelloides, four genes encode the PKAR1, PKAR2, PKAR3, and PKAR4 isoforms of R subunits. We have constructed null mutants and demonstrate that each isoform has a different role in growth and differentiation. The most striking finding is that pkaR4 is an essential gene, because only heterokaryons were obtained in knockout experiments. Heterokaryons with low levels of wild-type nuclei showed an impediment in the emission of the germ tube, suggesting a pivotal role of this gene in germ tube emergence. The remaining null strains showed different alterations in germ tube emergence, sporulation, and volume of the mother cell. The pkaR2 null mutant showed an accelerated germ tube emission and was the only mutant that germinated under anaerobic conditions when glycine was used as a nitrogen source, suggesting that pkaR2 participates in germ tube emergence by repressing it. From the measurement of the mRNA and protein levels of each isoform in the wild-type and knockout strains, it can be concluded that the expression of each subunit has its own mechanism of differential regulation. The PKAR1 and PKAR2 isoforms are posttranslationally modified by ubiquitylation, suggesting another regulation point in the specificity of the signal transduction. The results indicate that each R isoform has a different role in M. circinelloides physiology, controlling the dimorphism and contributing to the specificity of cyclic AMP (cAMP)-PKA pathway. PMID:22635921

  6. Binding studies of adhesion/growth-regulatory galectins with glycoconjugates monitored by surface plasmon resonance and NMR spectroscopy.

    PubMed

    Muñoz, F Javier; Santos, J Ignacio; Ardá, Ana; André, Sabine; Gabius, Hans-Joachim; Sinisterra, José V; Jiménez-Barbero, Jesús; Hernáiz, María J

    2010-06-28

    Functionalized fluorescent glycans have the potential to act as tools to detect and analyze protein-carbohydrate interactions. We present here a facile strategy for immobilization of functionalized lactose as a model disaccharide. Bioactivity was tested with three members of the adhesion/growth-regulatory galectins family in different types of assay, i.e. matrix in surface plasmon resonance (SPR), free ligand in solution by STD/trNOESY and docking measurements. In all cases, the activity of the disaccharide was maintained. The attachment of this new fluorescent glycoconjugate to the surface results in a well-defined interface, enabling desired orientational flexibility and enhanced access of binding partners. The results indicate that this new glycoconjugate exhibits binding affinity to galectin-1, 3 and CG-16. Kinetic analysis of the interaction between these galectins and immobilized glycoconjugate by SPR yielded a K(D) of 1.01 mM for galectin-1, 83.5 microM for galectin-3 and 0.28 mM for CG-16. No major contacts to the aglyconic part were detected, which might compromise the specificity of the binding process with other headgroups. Thus, testing these proteins offers the potential for medical applications to detect these endogenous effectors or further derivatives and characterize their carbohydrate specificity.

  7. Growth Hormone-Releasing Hormone and Its Analogues: Significance for MSCs-Mediated Angiogenesis

    PubMed Central

    Tao, Quanwei; Ma, Qunchao; Chen, Huiqiang; Wang, Jian'an

    2016-01-01

    Mesenchymal stromal cells (MSCs) are promising candidates for regenerative medicine because of their multipotency, immune-privilege, and paracrine properties including the potential to promote angiogenesis. Accumulating evidence suggests that the inherent properties of cytoprotection and tissue repair by native MSCs can be enhanced by various preconditioning stimuli implemented prior to cell transplantation. Growth hormone-releasing hormone (GHRH), a stimulator in extrahypothalamus systems including tumors, has attracted great attentions in recent years because GHRH and its agonists could promote angiogenesis in various tissues. GHRH and its agonists are proangiogenic in responsive tissues including tumors, and GHRH antagonists have been tested as antitumor agents through their ability to suppress angiogenesis and cell growth. GHRH-R is expressed by MSCs and evolving work from our laboratory indicates that treatment of MSCs with GHRH agonists prior to cell transplantation markedly enhanced the angiogenic potential and tissue reparative properties of MSCs through a STAT3 signaling pathway. In this review we summarized the possible effects of GHRH analogues on cell growth and development, as well as on the proangiogenic properties of MSCs. We also discussed the relationship between GHRH analogues and MSC-mediated angiogenesis. The analyses provide new insights into molecular pathways of MSCs-based therapies and their augmentation by GHRH analogues. PMID:27774107

  8. Lentivirus-mediated knockdown of NLK inhibits small-cell lung cancer growth and metastasis

    PubMed Central

    Lv, Mutian; Li, Yaming; Tian, Xin; Dai, Shundong; Sun, Jing; Jin, Guojiang; Jiang, Shenyi

    2016-01-01

    Nemo-like kinase (NLK), an evolutionarily conserved serine/threonine kinase, has been recognized as a critical regulator of various cancers. In this study, we investigated the role of NLK in human small-cell lung cancer (SCLC), which is the most aggressive form of lung cancer. NLK expression was evaluated by quantitative real-time polymerase chain reaction in 20 paired fresh SCLC tissue samples and found to be noticeably elevated in tumor tissues. Lentivirus-mediated RNAi efficiently suppressed NLK expression in NCI-H446 cells, resulting in a significant reduction in cell viability and proliferation in vitro. Moreover, knockdown of NLK led to cell cycle arrest at the S-phase via suppression of Cyclin A, CDK2, and CDC25A, which could contribute to cell growth inhibition. Furthermore, knockdown of NLK decreased the migration of NCI-H446 cells and downregulated matrix metalloproteinase 9. Treatment with NLK short hairpin RNA significantly reduced SCLC tumor growth in vivo. In conclusion, this study suggests that NLK plays an important role in the growth and metastasis of SCLC and may serve as a potential therapeutic target for the treatment of SCLC. PMID:27895463

  9. Cooperation of epidermis and inner tissues in auxin-mediated growth of maize coleoptiles.

    PubMed

    Kutschera, U; Bergfeld, R; Schopfer, P

    1987-02-01

    The function of the epidermis in auxinmediated elongation growth of maize (Zea mays L.) coleoptile segments was investigated. The following results were obtained: i) In the intact organ, there is a strong tissue tension produced by the expanding force of the inner tissues which is balanced by the contracting force of the outer epidermal wall. The compression imposed by the stretched outer epidermal wall upon the inner tissues gives rise to a wall-pressure difference which can be transformed into a water-potential difference between inner tissues and external medium (water) by removal of the outer epidermal wall. ii) Peeled segments fail to respond to auxin with normal growth. The plastic extensibility of the inner-tissue cell walls (measured with a constant-load extensiometer using living segments) is not influenced by auxin (or abscisic acid) in peeled or nonpeeled segments. It is concluded that auxin induces (and abscisic acid inhibits) elongation of the intact segment by increasing (decreasing) the extensibility specifically in the outer epidermal wall. In addition, tissue tension (and therewith the pressure acting on the outer epidermal wall) is maintained at a constant level over several hours of auxin-mediated growth, indicating that the inner cells also contribute actively to organ elongation. However, this contribution does not involve an increase of cell-wall extensibility, but a continuous shifting of the potential extension threshold (i.e., the length to which the inner tissues would extend by water uptake after peeling) ahead of the actual segment length. Thus, steady growth involves the coordinated action of wall loosening in the epidermis and regeneration of tissue tension by the inner tissues. iii) Electron micrographs show the accumulation of striking osmiophilic material (particles of approx. 0.3 μm diameter) specifically at the plasma membrane/cell-wall interface of the outer epidermal wall of auxin-treated segments. iv) Peeled segments fail to

  10. Regulatory T-Cell-Mediated Suppression of Conventional T-Cells and Dendritic Cells by Different cAMP Intracellular Pathways.

    PubMed

    Rueda, Cesar M; Jackson, Courtney M; Chougnet, Claire A

    2016-01-01

    Regulatory T-cells (Tregs) mediate their suppressive action by acting directly on conventional T-cells (Tcons) or dendritic cells (DCs). One mechanism of Treg suppression is the increase of cyclic adenosine 3',5'-monophosphate (cAMP) levels in target cells. Tregs utilize cAMP to control Tcon responses, such as proliferation and cytokine production. Tregs also exert their suppression on DCs, diminishing DC immunogenicity by downmodulating the expression of costimulatory molecules and actin polymerization at the immunological synapse. The Treg-mediated usage of cAMP occurs through two major mechanisms. The first involves the Treg-mediated influx of cAMP in target cells through gap junctions. The second is the conversion of adenosine triphosphate into adenosine by the ectonucleases CD39 and CD73 present on the surface of Tregs. Adenosine then binds to receptors on the surface of target cells, leading to increased intracellular cAMP levels in these targets. Downstream, cAMP can activate the canonical protein kinase A (PKA) pathway and the exchange protein activated by cyclic AMP (EPAC) non-canonical pathway. In this review, we discuss the most recent findings related to cAMP activation of PKA and EPAC, which are implicated in Treg homeostasis as well as the functional alterations induced by cAMP in cellular targets of Treg suppression.

  11. PGE{sub 2}-induced colon cancer growth is mediated by mTORC1

    SciTech Connect

    Dufour, Marc Faes, Seraina Dormond-Meuwly, Anne Demartines, Nicolas Dormond, Olivier

    2014-09-05

    Highlights: • PGE{sub 2} activates mTORC1 in colon cancer cells. • Inhibition of mTORC1 blocks PGE{sub 2} induced colon cancer cell growth. • mTORC1 is a signaling intermediary in PGE{sub 2} induced colon cancer cell responses. - Abstract: The inflammatory prostaglandin E{sub 2} (PGE{sub 2}) cytokine plays a key role in the development of colon cancer. Several studies have shown that PGE{sub 2} directly induces the growth of colon cancer cells and furthermore promotes tumor angiogenesis by increasing the production of the vascular endothelial growth factor (VEGF). The signaling intermediaries implicated in these processes have however not been fully characterized. In this report, we show that the mechanistic target of rapamycin complex 1 (mTORC1) plays an important role in PGE{sub 2}-induced colon cancer cell responses. Indeed, stimulation of LS174T cells with PGE{sub 2} increased mTORC1 activity as observed by the augmentation of S6 ribosomal protein phosphorylation, a downstream effector of mTORC1. The PGE{sub 2} EP{sub 4} receptor was responsible for transducing the signal to mTORC1. Moreover, PGE{sub 2} increased colon cancer cell proliferation as well as the growth of colon cancer cell colonies grown in matrigel and blocking mTORC1 by rapamycin or ATP-competitive inhibitors of mTOR abrogated these effects. Similarly, the inhibition of mTORC1 by downregulation of its component raptor using RNA interference blocked PGE{sub 2}-induced LS174T cell growth. Finally, stimulation of LS174T cells with PGE{sub 2} increased VEGF production which was also prevented by mTORC1 inhibition. Taken together, these results show that mTORC1 is an important signaling intermediary in PGE{sub 2} mediated colon cancer cell growth and VEGF production. They further support a role for mTORC1 in inflammation induced tumor growth.

  12. Akt- or MEK-mediated mTOR inhibition suppresses Nf1 optic glioma growth

    PubMed Central

    Kaul, Aparna; Toonen, Joseph A.; Cimino, Patrick J.; Gianino, Scott M.; Gutmann, David H.

    2015-01-01

    Background Children with neurofibromatosis type 1 (NF1) develop optic pathway gliomas, which result from impaired NF1 protein regulation of Ras activity. One obstacle to the implementation of biologically targeted therapies is an incomplete understanding of the individual contributions of the downstream Ras effectors (mitogen-activated protein kinase kinase [MEK], Akt) to optic glioma maintenance. This study was designed to address the importance of MEK and Akt signaling to Nf1 optic glioma growth. Methods Primary neonatal mouse astrocyte cultures were employed to determine the consequence of phosphatidylinositol-3 kinase (PI3K)/Akt and MEK inhibition on Nf1-deficient astrocyte growth. Nf1 optic glioma–bearing mice were used to assess the effect of Akt and MEK inhibition on tumor volume, proliferation, and retinal ganglion cell dysfunction. Results Both MEK and Akt were hyperactivated in Nf1-deficient astrocytes in vitro and in Nf1 murine optic gliomas in vivo. Pharmacologic PI3K or Akt inhibition reduced Nf1-deficient astrocyte proliferation to wild-type levels, while PI3K inhibition decreased Nf1 optic glioma volume and proliferation. Akt inhibition of Nf1-deficient astrocyte and optic glioma growth reflected Akt-dependent activation of mammalian target of rapamycin (mTOR). Sustained MEK pharmacologic blockade also attenuated Nf1-deficient astrocytes as well as Nf1 optic glioma volume and proliferation. Importantly, these MEK inhibitory effects resulted from p90RSK-mediated, Akt-independent mTOR activation. Finally, both PI3K and MEK inhibition reduced optic glioma–associated retinal ganglion cell loss and nerve fiber layer thinning. Conclusion These findings establish that the convergence of 2 distinct Ras effector pathways on mTOR signaling maintains Nf1 mouse optic glioma growth, supporting the evaluation of pharmacologic inhibitors that target mTOR function in future human NF1–optic pathway glioma clinical trials. PMID:25534823

  13. Synthesis and characterization of gold nanodogbones by the seeded mediated growth method.

    PubMed

    Huang, Chien-Jung; Chiu, Pin-Hsiang; Wang, Yeong-Her; Meen, Teen-Hang; Yang, Cheng-Fu

    2007-10-03

    Novel gold nanodogbones (GDBs) are successfully fabricated using a simple seeded mediated growth (SMG) method. The shapes of GDBs depend on the amount of added vitamin C solvent. The amount of vitamin C solvent was varied from 10 to 40 µl to investigate the influence of vitamin C solvent on the GDBs. It is found that the aspect ratios (R) of GDBs were in the range from 2.34 to 1.46, and the UV-vis absorption measurement revealed a pronounced blueshift on the longitudinal surface plasmon resonance (SPR) band from 713 to 676 nm. The GDBs were determined by x-ray diffraction (XRD) to be single-crystalline with a face-centered cubic (fcc) structure. The lattice constant calculated from this selected-area electron diffraction (SAED) pattern is 4.068 Å.

  14. Transforming growth factor-β: an important mediator in Helicobacter pylori-associated pathogenesis

    PubMed Central

    Li, Nianshuang; Xie, Chuan; Lu, Nong-Hua

    2015-01-01

    Helicobacter pylori (H.pylori) is a Gram-negative, microaerophilic, helical bacillus that specifically colonizes the gastric mucosa. The interaction of virulence factors, host genetic factors, and environmental factors contributes to the pathogenesis of H. pylori-associated conditions, such as atrophic gastritis and intestinal metaplasia. Infection with H. pylori has recently been recognized as the strongest risk factor for gastric cancer. As a pleiotropic cytokine, transforming growth factor (TGF)-β regulates various biological processes, including cell cycle, proliferation, apoptosis, and metastasis. Recent studies have shed new light on the involvement of TGF-β signaling in the pathogenesis of H. pylori infection. This review focuses on the potential etiological roles of TGF-β in H. pylori-mediated gastric pathogenesis. PMID:26583078

  15. Platelet Vascular Endothelial Growth Factor is a Potential Mediator of Transfusion-Related Acute Lung Injury

    PubMed Central

    Maloney, James P; Ambruso, Daniel R; Voelkel, Norbert F; Silliman, Christopher C

    2015-01-01

    Objective The occurrence of non-hemolytic transfusion reactions is highest with platelet and plasma administration. Some of these reactions are characterized by endothelial leak, especially transfusion related acute lung injury (TRALI). Elevated concentrations of inflammatory mediators secreted by contaminating leukocytes during blood product storage may contribute to such reactions, but platelet-secreted mediators may also contribute. We hypothesized that platelet storage leads to accumulation of the endothelial permeability mediator vascular endothelial growth factor (VEGF), and that intravascular administration of exogenous VEGF leads to extensive binding to its lung receptors. Methods Single donor, leukocyte-reduced apheresis platelet units were sampled over 5 days of storage. VEGF protein content of the centrifuged supernatant was determined by ELISA, and the potential contribution of VEGF from contaminating leukocytes was quantified. Isolated-perfused rat lungs were used to study the uptake of radiolabeled VEGF administered intravascularly, and the effect of unlabeled VEGF on lung leak. Results There was a time-dependent release of VEGF into the plasma fraction of the platelet concentrates (62 ± 9 pg/ml on day one, 149 ± 23 pg/ml on day 5; mean ± SEM, p<0.01, n=8) and a contribution by contaminating leukocytes was excluded. Exogenous 125I-VEGF bound avidly and specifically to the lung vasculature, and unlabeled VEGF in the lung perfusate caused vascular leak. Conclusion Rising concentrations of VEGF occur during storage of single donor platelet concentrates due to platelet secretion or disintegration, but not due to leukocyte contamination. Exogenous VEGF at these concentrations rapidly binds to its receptors in the lung vessels. At higher VEGF concentrations, VEGF causes vascular leak in uninjured lungs. These data provide further evidence that VEGF may contribute to the increased lung permeability seen in TRALI associated with platelet products. PMID

  16. Growth Hormone Mediates Its Protective Effect in Hepatic Apoptosis through Hnf6

    PubMed Central

    Wang, Kewei; Wang, Minhua; Gannon, Maureen

    2016-01-01

    Background and Aims Growth hormone (GH) not only supports hepatic metabolism but also protects against hepatocyte cell death. Hnf6 (or Oc1) belonging to the Onecut family of hepatocyte transcription factors known to regulate differentiated hepatic function, is a GH-responsive gene. We evaluate if GH mediates Hnf6 activity to attenuate hepatic apoptotic injury. Methods We used an animal model of hepatic apoptosis by bile duct ligation (BDL) with Hnf6 -/- (KO) mice in which hepatic Hnf6 was conditionally inactivated. GH was administered to adult wild type WT and KO mice for the 7 days of BDL to enhance Hnf6 expression. In vitro, primary hepatocytes derived from KO and WT liver were treated with LPS and hepatocyte apoptosis was assessed with and without GH treatment. Results In WT mice, GH treatment enhanced Hnf6 expression during BDL, inhibited Caspase -3, -8 and -9 responses and diminished hepatic apoptotic and fibrotic injury. GH-mediated upregulation of Hnf6 expression and parallel suppression of apoptosis and fibrosis in WT BDL liver were abrogated in KO mice. LPS activated apoptosis and suppressed Hnf6 expression in primary hepatocytes. GH/LPS co-treatment enhanced Hnf6 expression with corresponding attenuation of apoptosis in WT-derived hepatocytes, but not in KO hepatocytes. ChiP-on-ChiP and electromobility shift assays of KO and WT liver nuclear extracts identified Ciap1 (or Birc2) as an Hnf6-bound target gene. Ciap1 expression patterns closely follow Hnf6 expression in the liver and in hepatocytes. Conclusion GH broad protective actions on hepatocytes during liver injury are effected through Hnf6, with Hnf6 transcriptional activation of Ciap1 as an underlying molecular mediator. PMID:27936029

  17. H-Ras Mediates the Inhibitory Effect of Epidermal Growth Factor on the Epithelial Na+ Channel

    PubMed Central

    Lee, Il-Ha; Song, Sung-Hee; Cook, David I.; Dinudom, Anuwat

    2015-01-01

    The present study investigates the role of small G-proteins of the Ras family in the epidermal growth factor (EGF)-activated cellular signalling pathway that downregulates activity of the epithelial Na+ channel (ENaC). We found that H-Ras is a key component of this EGF-activated cellular signalling mechanism in M1 mouse collecting duct cells. Expression of a constitutively active H-Ras mutant inhibited the amiloride-sensitive current. The H-Ras-mediated signalling pathway that inhibits activity of ENaC involves c-Raf, and that the inhibitory effect of H-Ras on ENaC is abolished by the MEK1/2 inhibitor, PD98059. The inhibitory effect of H-Ras is not mediated by Nedd4-2, a ubiquitin protein ligase that regulates the abundance of ENaC at the cell surface membrane, or by a negative effect of H-Ras on proteolytic activation of the channel. The inhibitory effects of EGF and H-Ras on ENaC, however, were not observed in cells in which expression of caveolin-1 (Cav-1) had been knocked down by siRNA. These findings suggest that the inhibitory effect of EGF on ENaC-dependent Na+ absorption is mediated via the H-Ras/c-Raf, MEK/ERK signalling pathway, and that Cav-1 is an essential component of this EGF-activated signalling mechanism. Taken together with reports that mice expressing a constitutive mutant of H-Ras develop renal cysts, our findings suggest that H-Ras may play a key role in the regulation of renal ion transport and renal development. PMID:25774517

  18. Choroidal retinoic acid synthesis: a possible mediator between refractive error and compensatory eye growth.

    PubMed

    Mertz, J R; Wallman, J

    2000-04-01

    Research over the past two decades has shown that the growth of young eyes is guided by vision. If near- or far-sightedness is artificially imposed by spectacle lenses, eyes of primates and chicks compensate by changing their rate of elongation, thereby growing back to the pre-lens optical condition. Little is known about what chemical signals might mediate between visual effects on the retina and alterations of eye growth. We present five findings that point to choroidal retinoic acid possibly being such a mediator. First, the chick choroid can convert retinol into all-trans-retinoic acid at the rate of 11 +/- 3 pmoles mg protein(-1) hr(-1), compared to 1.3 +/- 0.3 for retina/RPE and no conversion for sclera. Second, those visual conditions that cause increased rates of ocular elongation (diffusers or negative lens wear) produce a sharp decrease in all-trans-retinoic acid synthesis to levels barely detectable with our assay. In contrast, visual conditions which result in decreased rates of ocular elongation (recovery from diffusers or positive lens wear) produce a four- to five-fold increase in the formation of all-trans-retinoic acid. Third, the choroidal retinoic acid is found bound to a 28-32 kD protein. Fourth, a large fraction of the choroidal retinoic acid synthesized in culture is found in a nucleus-enriched fraction of sclera. Finally, application of retinoic acid to cultured sclera at physiological concentrations produced an inhibition of proteoglycan production (as assessed by measuring sulfate incorporation) with a EC50 of 8 x 10(-7) M. These results show that the synthesis of choroidal retinoic acid is modulated by those visual manipulations that influence ocular elongation and that this retinoic acid may reach the sclera in concentrations adequate to modulate scleral proteoglycan formation.

  19. Piwi-like 2 mediates fibroblast growth factor signaling during gastrulation of zebrafish embryo.

    PubMed

    Zhao, Jun; Sun, Huaqin; Deng, Wenqian; Li, Dan; Liu, Yanyan; Lu, Yilu; Liu, Yunqiang; Tao, Dachang; Zhang, Sizhong; Ma, Yongxin

    2010-09-01

    Piwi (P-element-induced wimpy testis) proteins have been shown to play important roles in maintenance of germ line stem cells, germ cell proliferation and differentiation, and control of Piwi-interacting RNAs (PiRNAs). PiRNAs comprise a broad class of small noncoding RNAs that function as an endogenous defense system against transposable elements. Fibroblast growth factor (Fgf) signals, mediated partly by no tail gene (ntl), are responsible for patterning embryo and mesoderm formation. To understand the function of Piwi proteins, we used zebrafish as a model system. In zebrafish, piwi-like 2 gene (piwil2) is also required for germ cell differentiation and meiosis. Here we report that piwil2 knockdown is able to inhibit the expression of fibroblast growth factor 8a (fgf8a). In contrast, injection with piwil2 mRNA enhances fgf8a expression. Knockdown of piwil2 reduces the inductive effect of fgf8a on dorsalized phenotype, in which embryos extend to an oval shape at the end of epiboly stage. Coinjection with fgf8a and piwil2 mRNAs led to more seriously dorsalized phenotype than coinjection with fgf8a mRNA and piwil2-cMO. In addition, knockdown of piwil2 inhibits the inductive effect of fgf8a on ntl, whereas overexpression of piwil2 enhances the inductive effect of fgf8a on ntl. We also demonstrate that piwil2 positively regulates ntl expression at bud stage, while piwil2 negatively regulates ntl expression at 24 hours post-fertilization. Thus, the functional consequences of piwil2 expression vary during early development of zebrafish embryo. Taken together, we suggest that zebrafish piwil2 is a mediator of Fgf signals in gastrula period.

  20. Sphingosine-1-phosphate mediates epidermal growth factor-induced muscle satellite cell activation.

    PubMed

    Nagata, Yosuke; Ohashi, Kazuya; Wada, Eiji; Yuasa, Yuki; Shiozuka, Masataka; Nonomura, Yoshiaki; Matsuda, Ryoichi

    2014-08-01

    Skeletal muscle can regenerate repeatedly due to the presence of resident stem cells, called satellite cells. Because satellite cells are usually quiescent, they must be activated before participating in muscle regeneration in response to stimuli such as injury, overloading, and stretch. Although satellite cell activation is a crucial step in muscle regeneration, little is known of the molecular mechanisms controlling this process. Recent work showed that the bioactive lipid sphingosine-1-phosphate (S1P) plays crucial roles in the activation, proliferation, and differentiation of muscle satellite cells. We investigated the role of growth factors in S1P-mediated satellite cell activation. We found that epidermal growth factor (EGF) in combination with insulin induced proliferation of quiescent undifferentiated mouse myoblast C2C12 cells, which are also known as reserve cells, in serum-free conditions. Sphingosine kinase activity increased when reserve cells were stimulated with EGF. Treatment of reserve cells with the D-erythro-N,N-dimethylsphingosine, Sphingosine Kinase Inhibitor, or siRNA duplexes specific for sphingosine kinase 1, suppressed EGF-induced C2C12 activation. We also present the evidence showing the S1P receptor S1P2 is involved in EGF-induced reserve cell activation. Moreover, we demonstrated a combination of insulin and EGF promoted activation of satellite cells on single myofibers in a manner dependent on SPHK and S1P2. Taken together, our observations show that EGF-induced satellite cell activation is mediated by S1P and its receptor.

  1. The growth and reproduction performance of TALEN-mediated β-lactoglobulin-knockout bucks.

    PubMed

    Ge, Hengtao; Cui, Chenchen; Liu, Jun; Luo, Yan; Quan, Fusheng; Jin, Yaping; Zhang, Yong

    2016-10-01

    With the technological development of several engineered endonucleases (EENs), such as zinc-finger nucleases, transcription activator-like effector nucleases (TALENs) and CRISPR/Cas9, gene targeting by homologous recombination has been efficiently improved to generate site-specifically genetically modified livestock. However, few studies have been done to investigate the health and fertility of these animals. The purpose of the present study is to investigate if gene targeting events and a recloning procedure would affect the production traits of EEN-mediated gene targeted bucks. TALEN-mediated β-lactoglobulin (BLG) gene mono-allelic knockout (BLG (+/-)) goats and bi-allelic knockout (BLG (-/-)) buck produced by using sequential gene targeting combined with recloning in fibroblasts from BLG (+/-) buck were used to evaluate their health and fertility. Birth weight and postnatal growth of BLG (+/-) bucks were similar to the wild-type goats. None of the parameters for both fresh and frozen-thawed semen quality were significantly different in BLG (+/-) or BLG (-/-) bucks compared to their corresponding comparators. In vitro fertilization (IVF) test revealed that the proportion of IVF oocytes developing to the blastocyst stage was identical among BLG (+/-), BLG (-/-) and wild-type bucks. Conception rates of artificial insemination were respectively 42.3, 38.0 and 42.6 % for frozen-thawed semen from the BLG (+/-), BLG (-/-) and wild-type bucks. In addition, germline transmission of the targeted BLG modification was in accordance with Mendelian rules. These results demonstrated that the analyzed growth and reproductive traits were not impacted by targeting BLG gene and recloning, implicating the potential for dairy goat breeding of BLG (+/-) and BLG (-/-) bucks.

  2. Berberine inhibits cell growth and mediates caspase-independent cell death in human pancreatic cancer cells.

    PubMed

    Pinto-Garcia, Lina; Efferth, Thomas; Torres, Amada; Hoheisel, Jörg D; Youns, Mahmoud

    2010-08-01

    Pancreatic cancer is one of the most aggressive human malignancies with an increasing incidence worldwide. In addition to the poor survival rates, combinations using gemcitabine as a backbone have failed to show any benefit beyond monotherapy. These facts underscore an urgent need for novel therapeutic options and motivated us to study the effect of berberine on pancreatic cancer cells. Here, we undertook an mRNA-based gene expression profiling study in order to get deeper insight into the molecular targets mediating the growth inhibitory effects of berberine on pancreatic cancer cells compared to normal ones. Twenty-four hours after treatment, berberine showed preferential selectivity toward pancreatic cancer cells compared to normal ones. Moreover, expression profiling and Ingenuity pathway analysis results showed that the cytotoxicity of berberine was accompanied with an activation of BRCA1-mediated DNA damage response, G1/S and G2/M cell cycle checkpoint regulation, and P53 signalling pathways. The activation of these signalling pathways might be explained by the fact that berberine intercalates DNA and induces DNA strand break through inhibition of topoisomerases and induction of DNA lesions.

  3. EHD1 mediates vesicle trafficking required for normal muscle growth and transverse tubule development.

    PubMed

    Posey, Avery D; Swanson, Kaitlin E; Alvarez, Manuel G; Krishnan, Swathi; Earley, Judy U; Band, Hamid; Pytel, Peter; McNally, Elizabeth M; Demonbreun, Alexis R

    2014-03-15

    EHD proteins have been implicated in intracellular trafficking, especially endocytic recycling, where they mediate receptor and lipid recycling back to the plasma membrane. Additionally, EHDs help regulate cytoskeletal reorganization and induce tubule formation. It was previously shown that EHD proteins bind directly to the C2 domains in myoferlin, a protein that regulates myoblast fusion. Loss of myoferlin impairs normal myoblast fusion leading to smaller muscles in vivo but the intracellular pathways perturbed by loss of myoferlin function are not well known. We now characterized muscle development in EHD1-null mice. EHD1-null myoblasts display defective receptor recycling and mislocalization of key muscle proteins, including caveolin-3 and Fer1L5, a related ferlin protein homologous to myoferlin. Additionally, EHD1-null myoblast fusion is reduced. We found that loss of EHD1 leads to smaller muscles and myofibers in vivo. In wildtype skeletal muscle EHD1 localizes to the transverse tubule (T-tubule), and loss of EHD1 results in overgrowth of T-tubules with excess vesicle accumulation in skeletal muscle. We provide evidence that tubule formation in myoblasts relies on a functional EHD1 ATPase domain. Moreover, we extended our studies to show EHD1 regulates BIN1 induced tubule formation. These data, taken together and with the known interaction between EHD and ferlin proteins, suggests that the EHD proteins coordinate growth and development likely through mediating vesicle recycling and the ability to reorganize the cytoskeleton.

  4. EHD1 mediates vesicle trafficking required for normal muscle growth and tubule development

    PubMed Central

    Posey, Avery D.; Swanson, Kaitlin E.; Alvarez, Manuel G.; Krishnan, Swathi; Earley, Judy E.; Band, Hamid; Pytel, Peter; McNally, Elizabeth M.; Demonbreun, Alexis R.

    2014-01-01

    EHD proteins have been implicated in intracellular trafficking, especially endocytic recycling, where they mediate receptor and lipid recycling back to the plasma membrane. Additionally, EHDs help regulate cytoskeletal reorganization and induce tubule formation. It was previously shown that EHD proteins bind directly to the C2 domains in myoferlin, a protein that regulates myoblast fusion. Loss of myoferlin impairs normal myoblast fusion leading to smaller muscles in vivo but the intracellular pathways perturbed by loss of myoferlin function are not well known. We now characterized muscle development in EHD1-null mice. EHD1-null myoblasts display defective receptor recycling and mislocalization of key muscle proteins, including caveolin-3 and Fer1L5, a related ferlin protein homologous to myoferlin. Additionally, EHD1-null myoblast fusion is reduced. We found that loss of EHD1 leads to smaller muscles and myofibers in vivo. In wildtype skeletal muscle EHD1 localizes to the transverse tubule (T-tubule), and loss of EHD1 results in overgrowth of T-tubules with excess vesicle accumulation in skeletal muscle. We provide evidence that tubule formation in myoblasts relies on a functional EHD1 ATPase domain. Moreover, we extended our studies to show EHD1 regulates BIN1 induced tubule formation. These data, taken together and with the known interaction between EHD and ferlin proteins, suggests that the EHD proteins coordinate growth and development likely through mediating vesicle recycling and the ability to reorganize the cytoskeleton. PMID:24440153

  5. Distal regulatory element of the STAT1 gene potentially mediates positive feedback control of STAT1 expression.

    PubMed

    Yuasa, Katsutoshi; Hijikata, Takao

    2016-01-01

    We previously identified a distal regulatory element located approximately 5.5-kb upstream of the signal transducer and activator of transcription 1 (STAT1) gene, thereafter designating it as 5.5-kb upstream regulatory region (5.5URR). In this study, we investigated the functional roles of 5.5URR in the transcriptional regulation of STAT1 gene. A chromosome conformation capture assay indicated physical interaction of 5.5URR with the STAT1 core promoter. In luciferase reporter assays, 5.5URR-combined STAT1 core promoter exhibited significant increase in reporter activity enhanced by forced STAT1 expression or interferon (IFN) treatment, but STAT1 core promoter alone did not. The 5.5URR contained IFN-stimulated response element and GAS sites, which bound STAT1 complexes in electrophoretic mobility shift assays. Consistently, chromatin immunoprecipitation (ChIP) assays of HEK293 cells with Halo-tagged STAT1 expression indicated the association of Halo-tagged STAT1 with 5.5URR. ChIP assays with IFN treatment demonstrated that IFNs promoted the recruitment of Halo-tagged STAT1 to 5.5URR. Forced STAT1 expression or IFN treatment increased the expression of endogenous STAT1 and other IFN signaling pathway components, such as STAT2, IRF9 and IRF1, besides IFN-responsive genes. Collectively, the results suggest that 5.5URR may provide a regulatory platform for positive feedback control of STAT1 expression possibly to amplify or sustain the intracellular IFN signals.

  6. Pim-2 Kinase Influences Regulatory T Cell Function and Stability by Mediating Foxp3 Protein N-terminal Phosphorylation*

    PubMed Central

    Deng, Guoping; Nagai, Yasuhiro; Xiao, Yan; Li, Zhiyuan; Dai, Shujia; Ohtani, Takuya; Banham, Alison; Li, Bin; Wu, Shiaw-Lin; Hancock, Wayne; Samanta, Arabinda; Zhang, Hongtao; Greene, Mark I.

    2015-01-01

    Regulation of the extent of immune responses is a requirement to maintain self-tolerance and limit inflammatory processes. CD4+CD25+Foxp3+ regulatory T (Treg) cells play a role in regulation. The Foxp3 transcription factor is considered a dominant regulator for Treg cell development and function. Foxp3 function itself is directly regulated by multiple posttranslational modifications that occur in response to various external stimuli. The Foxp3 protein is a component of several dynamic macromolecular regulatory complexes. The complexes change constituents over time and through different signals to regulate the development and function of regulatory T cells. Here we identified a mechanism regulating Foxp3 level and activity that operates through discrete phosphorylation. The Pim-2 kinase can phosphorylate Foxp3, leading to decreased suppressive functions of Treg cells. The amino-terminal domain of Foxp3 is modified at several sites by Pim-2 kinase. This modification leads to altered expression of proteins related to Treg cell functions and increased Treg cell lineage stability. Treg cell suppressive function can be up-regulated by either pharmacologically inhibiting Pim-2 kinase activity or by genetically knocking out Pim-2 in rodent Treg cells. Deficiency of Pim-2 activity increases murine host resistance to dextran sodium sulfate-induced colitis in vivo, and a Pim-2 small molecule kinase inhibitor also modified Treg cell functions. Our studies define a pathway for limiting the regulation of Foxp3 function because the Pim-2 kinase represents a potential therapeutic target for modulating the Treg cell suppressive activities in controlling immune responses. PMID:25987564

  7. Pim-2 Kinase Influences Regulatory T Cell Function and Stability by Mediating Foxp3 Protein N-terminal Phosphorylation.

    PubMed

    Deng, Guoping; Nagai, Yasuhiro; Xiao, Yan; Li, Zhiyuan; Dai, Shujia; Ohtani, Takuya; Banham, Alison; Li, Bin; Wu, Shiaw-Lin; Hancock, Wayne; Samanta, Arabinda; Zhang, Hongtao; Greene, Mark I

    2015-08-14

    Regulation of the extent of immune responses is a requirement to maintain self-tolerance and limit inflammatory processes. CD4(+)CD25(+)Foxp3(+) regulatory T (Treg) cells play a role in regulation. The Foxp3 transcription factor is considered a dominant regulator for Treg cell development and function. Foxp3 function itself is directly regulated by multiple posttranslational modifications that occur in response to various external stimuli. The Foxp3 protein is a component of several dynamic macromolecular regulatory complexes. The complexes change constituents over time and through different signals to regulate the development and function of regulatory T cells. Here we identified a mechanism regulating Foxp3 level and activity that operates through discrete phosphorylation. The Pim-2 kinase can phosphorylate Foxp3, leading to decreased suppressive functions of Treg cells. The amino-terminal domain of Foxp3 is modified at several sites by Pim-2 kinase. This modification leads to altered expression of proteins related to Treg cell functions and increased Treg cell lineage stability. Treg cell suppressive function can be up-regulated by either pharmacologically inhibiting Pim-2 kinase activity or by genetically knocking out Pim-2 in rodent Treg cells. Deficiency of Pim-2 activity increases murine host resistance to dextran sodium sulfate-induced colitis in vivo, and a Pim-2 small molecule kinase inhibitor also modified Treg cell functions. Our studies define a pathway for limiting the regulation of Foxp3 function because the Pim-2 kinase represents a potential therapeutic target for modulating the Treg cell suppressive activities in controlling immune responses.

  8. Inactivation of PI(3)K p110δ breaks regulatory T-cell-mediated immune tolerance to cancer.

    PubMed

    Ali, Khaled; Soond, Dalya R; Piñeiro, Roberto; Hagemann, Thorsten; Pearce, Wayne; Lim, Ee Lyn; Bouabe, Hicham; Scudamore, Cheryl L; Hancox, Timothy; Maecker, Heather; Friedman, Lori; Turner, Martin; Okkenhaug, Klaus; Vanhaesebroeck, Bart

    2014-06-19

    Inhibitors against the p110δ isoform of phosphoinositide-3-OH kinase (PI(3)K) have shown remarkable therapeutic efficacy in some human leukaemias. As p110δ is primarily expressed in leukocytes, drugs against p110δ have not been considered for the treatment of solid tumours. Here we report that p110δ inactivation in mice protects against a broad range of cancers, including non-haematological solid tumours. We demonstrate that p110δ inactivation in regulatory T cells unleashes CD8(+) cytotoxic T cells and induces tumour regression. Thus, p110δ inhibitors can break tumour-induced immune tolerance and should be considered for wider use in oncology.

  9. Surface proteome analysis identifies platelet derived growth factor receptor-alpha as a critical mediator of transforming growth factor-beta-induced collagen secretion.

    PubMed

    Heinzelmann, Katharina; Noskovičová, Nina; Merl-Pham, Juliane; Preissler, Gerhard; Winter, Hauke; Lindner, Michael; Hatz, Rudolf; Hauck, Stefanie M; Behr, Jürgen; Eickelberg, Oliver

    2016-05-01

    Fibroblasts are extracellular matrix-producing cells in the lung. Fibroblast activation by transforming growth factor-beta leads to myofibroblast-differentiation and increased extracellular matrix deposition, a hallmark of pulmonary fibrosis. While fibroblast function with respect to migration, invasion, and extracellular matrix deposition has been well-explored, little is known about the surface proteome of lung fibroblasts in general and its specific response to fibrogenic growth factors, in particular transforming growth factor-beta. We thus performed a cell-surface proteome analysis of primary human lung fibroblasts in presence/absence of transforming growth factor-beta, followed by characterization of our findings using FACS analysis, Western blot, and siRNA-mediated knockdown experiments. We identified 213 surface proteins significantly regulated by transforming growth factor-beta, platelet derived growth factor receptor-alpha being one of the top down-regulated proteins. Transforming growth factor beta-induced downregulation of platelet derived growth factor receptor-alpha induced upregulation of platelet derived growth factor receptor-beta expression and phosphorylation of Akt, a downstream target of platelet derived growth factor signaling. Importantly, collagen type V expression and secretion was strongly increased after forced knockdown of platelet derived growth factor receptor-alpha, an effect that was potentiated by transforming growth factor-beta. We therefore show previously underappreciated cross-talk of transforming growth factor-beta and platelet derived growth factor signaling in human lung fibroblasts, resulting in increased extracellular matrix deposition in a platelet derived growth factor receptor-alpha dependent manner. These findings are of particular importance for the treatment of lung fibrosis patients with high pulmonary transforming growth factor-beta activity.

  10. Sphingosine-1-phosphate mediates epidermal growth factor-induced muscle satellite cell activation

    SciTech Connect

    Nagata, Yosuke Ohashi, Kazuya; Wada, Eiji; Yuasa, Yuki; Shiozuka, Masataka; Nonomura, Yoshiaki; Matsuda, Ryoichi

    2014-08-01

    Skeletal muscle can regenerate repeatedly due to the presence of resident stem cells, called satellite cells. Because satellite cells are usually quiescent, they must be activated before participating in muscle regeneration in response to stimuli such as injury, overloading, and stretch. Although satellite cell activation is a crucial step in muscle regeneration, little is known of the molecular mechanisms controlling this process. Recent work showed that the bioactive lipid sphingosine-1-phosphate (S1P) plays crucial roles in the activation, proliferation, and differentiation of muscle satellite cells. We investigated the role of growth factors in S1P-mediated satellite cell activation. We found that epidermal growth factor (EGF) in combination with insulin induced proliferation of quiescent undifferentiated mouse myoblast C2C12 cells, which are also known as reserve cells, in serum-free conditions. Sphingosine kinase activity increased when reserve cells were stimulated with EGF. Treatment of reserve cells with the D-erythro-N,N-dimethylsphingosine, Sphingosine Kinase Inhibitor, or siRNA duplexes specific for sphingosine kinase 1, suppressed EGF-induced C2C12 activation. We also present the evidence showing the S1P receptor S1P2 is involved in EGF-induced reserve cell activation. Moreover, we demonstrated a combination of insulin and EGF promoted activation of satellite cells on single myofibers in a manner dependent on SPHK and S1P2. Taken together, our observations show that EGF-induced satellite cell activation is mediated by S1P and its receptor. - Highlights: • EGF in combination with insulin induces proliferation of quiescent C2C12 cells. • Sphingosine kinase activity increases when reserve cells are stimulated with EGF. • EGF-induced activation of reserve cells is dependent on sphingosine kinase and ERK. • The S1P receptor S1P2 is involved in EGF-induced reserve cell activation. • EGF-induced reserve cell activation is mediated by S1P and its

  11. The effect of a brief sprint interval exercise on growth factors and inflammatory mediators.

    PubMed

    Meckel, Yoav; Eliakim, Alon; Seraev, Mariana; Zaldivar, Frank; Cooper, Dan M; Sagiv, Michael; Nemet, Dan

    2009-01-01

    Exercise training efficiency depends on the intensity, volume, duration, and frequency of training, as well as on the athlete's ability to tolerate it. Recent efforts to quantify the effects of aerobic exercise training on hormonal response have suggested that exercise leads to simultaneous changes of antagonistic mediators. The effects of anaerobic exercise on these mediators are not known. Therefore, the aim of the present study was to evaluate the effects of a brief sprint interval session on the balance between anabolic (growth hormone [GH]--> insulin-like growth factor [IGF]-I axis) and catabolic hormones (cortisol), and circulating inflammatory cytokines such as interleukin (IL)-6. Twelve healthy elite junior handball players (17-20 years) participated in the study. Exercise consisted of a 4 x 250-m run on a treadmill, at a constant intensity of 80% of the personal maximal speed. Each run was separated by 3 minutes of rest. Blood samples were collected before, immediately after each 250-m run, and 1 hour after the last run. Exercise led to significant increases in GH (0.3 +/- 0.2 to 5.1 +/- 2.2 ngxml, p < 0.05), IGF binding protein (IGFBP)-3 (4191 +/- 2.48 to 4875 +/- 301 ngxml, p < 0.05), IL-6 (1.3 +/- 0.2 to 2.1 +/- 0.3 pgxml, p < 0.002), testosterone, and testosterone/cortisol ratio, and to a significant decrease in IGFBP-1 levels. Levels of IL-6 remained elevated 1 hour after the end of exercise. Exercise had no significant effects on IGF-I and cortisol levels. Changes in the GH-IGF-I axis and testosterone/cortisol ratio after the brief sprint interval exercise suggested exercise-related anabolic adaptations. The increase in IL-6 may indicate its important role in muscle tissue repair after anaerobic exercise. Changes in the anabolic-catabolic hormonal balance and in inflammatory mediators can be used as an objective tool to gauge the training intensity of different types of anaerobic exercises and training periods.

  12. Connective Tissue Growth Factor in Regulation of RhoA Mediated Cytoskeletal Tension Associated Osteogenesis of Mouse Adipose-Derived Stromal Cells

    PubMed Central

    Xu, Yue; Wagner, Diane R.; Bekerman, Elena; Chiou, Michael; James, Aaron W.; Carter, Dennis; Longaker, Michael T.

    2010-01-01

    Background Cytoskeletal tension is an intracellular mechanism through which cells convert a mechanical signal into a biochemical response, including production of cytokines and activation of various signaling pathways. Methods/Principal Findings Adipose-derived stromal cells (ASCs) were allowed to spread into large cells by seeding them at a low-density (1,250 cells/cm2), which was observed to induce osteogenesis. Conversely, ASCs seeded at a high-density (25,000 cells/cm2) featured small cells that promoted adipogenesis. RhoA and actin filaments were altered by changes in cell size. Blocking actin polymerization by Cytochalasin D influenced cytoskeletal tension and differentiation of ASCs. To understand the potential regulatory mechanisms leading to actin cytoskeletal tension, cDNA microarray was performed on large and small ASCs. Connective tissue growth factor (CTGF) was identified as a major regulator of osteogenesis associated with RhoA mediated cytoskeletal tension. Subsequently, knock-down of CTGF by siRNA in ASCs inhibited this osteogenesis. Conclusions/Significance We conclude that CTGF is important in the regulation of cytoskeletal tension mediated ASC osteogenic differentiation. PMID:20585662

  13. Gene regulatory networks mediating canonical Wnt signal-directed control of pluripotency and differentiation in embryo stem cells.

    PubMed

    Zhang, Xiaoxiao; Peterson, Kevin A; Liu, X Shirley; McMahon, Andrew P; Ohba, Shinsuke

    2013-12-01

    Canonical Wnt signaling supports the pluripotency of embryonic stem cells (ESCs) but also promotes differentiation of early mammalian cell lineages. To explain these paradoxical observations, we explored the gene regulatory networks at play. Canonical Wnt signaling is intertwined with the pluripotency network comprising Nanog, Oct4, and Sox2 in mouse ESCs. In defined media supporting the derivation and propagation of ESCs, Tcf3 and β-catenin interact with Oct4; Tcf3 binds to Sox motif within Oct-Sox composite motifs that are also bound by Oct4-Sox2 complexes. Furthermore, canonical Wnt signaling upregulates the activity of the Pou5f1 distal enhancer via the Sox motif in ESCs. When viewed in the context of published studies on Tcf3 and β-catenin mutants, our findings suggest Tcf3 counters pluripotency by competition with Sox2 at these sites, and Tcf3 inhibition is blocked by β-catenin entry into this complex. Wnt pathway stimulation also triggers β-catenin association at regulatory elements with classic Lef/Tcf motifs associated with differentiation programs. The failure to activate these targets in the presence of a mitogen-activated protein kinase kinase (MEK)/extracellular signal-regulated kinase (ERK) inhibitor essential for ESC culture suggests MEK/ERK signaling and canonical Wnt signaling combine to promote ESC differentiation.

  14. Transient low-dose methotrexate generates B regulatory cells that mediate antigen-specific tolerance to alglucosidase alfa.

    PubMed

    Joly, Marguerite S; Martin, Roderick P; Mitra-Kaushik, Shibani; Phillips, Lucy; D'Angona, Alida; Richards, Susan M; Joseph, Alexandra M

    2014-10-15

    Biologic drugs, including enzyme-replacement therapies, can elicit anti-drug Abs (ADA) that may interfere with drug efficacy and impact patient safety. In an effort to control ADA, we focused on identifying regimens of immune tolerance induction that may be readily available for clinical use. Data generated in both wild-type mice and a Pompe disease mouse model demonstrate that single-cycle, low-dose methotrexate can be as effective as three cycles of methotrexate in providing a long-lived reduction in alglucosidase alfa-specific ADA. In addition, we show that methotrexate induces Ag-specific tolerance as mice generate similar Ab responses to an irrelevant Ag regardless of prior methotrexate treatment. Methotrexate-induced immune tolerance does not seem to involve cell depletion, but rather a specific expansion of IL-10- and TGF-β-secreting B cells that express Foxp3, suggesting an induction of regulatory B cells. The mechanism of immune tolerance induction appears to be IL-10 dependent, as methotrexate does not induce immune tolerance in IL-10 knockout mice. Splenic B cells from animals that have been tolerized to alglucosidase alfa with methotrexate can transfer tolerance to naive hosts. We hypothesize that methotrexate induction treatment concomitant with initial exposure to the biotherapeutic can induce Ag-specific immune tolerance in mice through a mechanism that appears to involve the induction of regulatory B cells.

  15. Activin A-Smad Signaling Mediates Connective Tissue Growth Factor Synthesis in Liver Progenitor Cells

    PubMed Central

    Ding, Ze-Yang; Jin, Guan-Nan; Wang, Wei; Sun, Yi-Min; Chen, Wei-Xun; Chen, Lin; Liang, Hui-Fang; Datta, Pran K.; Zhang, Ming-Zhi; Zhang, Bixiang; Chen, Xiao-Ping

    2016-01-01

    Liver progenitor cells (LPCs) are activated in chronic liver damage and may contribute to liver fibrosis. Our previous investigation reported that LPCs produced connective tissue growth factor (CTGF/CCN2), an inducer of liver fibrosis, yet the regulatory mechanism of the production of CTGF/CCN2 in LPCs remains elusive. In this study, we report that Activin A is an inducer of CTGF/CCN2 in LPCs. Here we show that expression of both Activin A and CTGF/CCN2 were upregulated in the cirrhotic liver, and the expression of Activin A positively correlates with that of CTGF/CCN2 in liver tissues. We go on to show that Activin A induced de novo synthesis of CTGF/CCN2 in LPC cell lines LE/6 and WB-F344. Furthermore, Activin A contributed to autonomous production of CTGF/CCN2 in liver progenitor cells (LPCs) via activation of the Smad signaling pathway. Smad2, 3 and 4 were all required for this induction. Collectively, these results provide evidence for the fibrotic role of LPCs in the liver and suggest that the Activin A-Smad-CTGF/CCN2 signaling in LPCs may be a therapeutic target of liver fibrosis. PMID:27011166

  16. Activin A-Smad Signaling Mediates Connective Tissue Growth Factor Synthesis in Liver Progenitor Cells.

    PubMed

    Ding, Ze-Yang; Jin, Guan-Nan; Wang, Wei; Sun, Yi-Min; Chen, Wei-Xun; Chen, Lin; Liang, Hui-Fang; Datta, Pran K; Zhang, Ming-Zhi; Zhang, Bixiang; Chen, Xiao-Ping

    2016-03-22

    Liver progenitor cells (LPCs) are activated in chronic liver damage and may contribute to liver fibrosis. Our previous investigation reported that LPCs produced connective tissue growth factor (CTGF/CCN2), an inducer of liver fibrosis, yet the regulatory mechanism of the production of CTGF/CCN2 in LPCs remains elusive. In this study, we report that Activin A is an inducer of CTGF/CCN2 in LPCs. Here we show that expression of both Activin A and CTGF/CCN2 were upregulated in the cirrhotic liver, and the expression of Activin A positively correlates with that of CTGF/CCN2 in liver tissues. We go on to show that Activin A induced de novo synthesis of CTGF/CCN2 in LPC cell lines LE/6 and WB-F344. Furthermore, Activin A contributed to autonomous production of CTGF/CCN2 in liver progenitor cells (LPCs) via activation of the Smad signaling pathway. Smad2, 3 and 4 were all required for this induction. Collectively, these results provide evidence for the fibrotic role of LPCs in the liver and suggest that the Activin A-Smad-CTGF/CCN2 signaling in LPCs may be a therapeutic target of liver fibrosis.

  17. The regulatory gene areA mediating nitrogen metabolite repression in Aspergillus nidulans. Mutations affecting specificity of gene activation alter a loop residue of a putative zinc finger.

    PubMed Central

    Kudla, B; Caddick, M X; Langdon, T; Martinez-Rossi, N M; Bennett, C F; Sibley, S; Davies, R W; Arst, H N

    1990-01-01

    The regulatory gene areA mediating nitrogen metabolite repression in Aspergillus nidulans has been sequenced and its transcript mapped and orientated. A single ORF can encode a protein of 719 amino acids. A 52 amino acid region including a putative 'zinc finger' strongly resembles putative DNA binding regions of the major regulatory protein of erythroid cells. The derived protein sequence also contains a highly acidic region possibly involved in gene activation and 22 copies of the motif S(T)PXX, abundant in DNA binding proteins. Analysis of chromosomal rearrangements and transformation with deletion clones identified 342 N-terminal and 124 C-terminal residues as inessential and localized a C-terminal region required for nitrogen metabolite repressibility. A -1 frameshift eliminating the inessential 122 C-terminal amino acids is a surprising loss-of-function mutation. Extraordinary basicity of the replacement C terminus might explain its phenotype. Mutant sequencing also identified a polypeptide chain termination and several missense mutations, but most interesting are sequence changes associated with specificity mutations. A mutation elevating expression of some structural genes under areA control whilst reducing or not affecting expression of others is a leucine to valine change in the zinc finger loop. It reverts to a partly reciprocal phenotype by replacing the mutant valine by methionine. Images Fig.2 Fig.4 Fig.5 Fig. 8. Fig. 9. PMID:1970293

  18. Growth suppression of Leydig TM3 cells mediated by aryl hydrocarbon receptor

    SciTech Connect

    Iseki, Minoru; Ikuta, Togo; Kobayashi, Tetsuya; Kawajiri, Kaname . E-mail: kawajiri@cancer-c.pref.saitama.jp

    2005-06-17

    Exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin induces developmental toxicity in reproductive organs. To elucidate the function of AhR, we generated stable transformants of TM3 cells overexpressing wild-type aryl hydrocarbon receptor (AhR) or its mutants which carried mutations in nuclear localization signal or nuclear export signal. In the presence of 3-methylcholanthrene (MC), proliferation of the cells transfected with wild-type AhR was completely suppressed, whereas cells expressing AhR mutants proliferated in a manner equivalent to control TM3 cells, suggesting AhR-dependent growth inhibition. The suppression was associated with up-regulation of cyclin-dependent kinase inhibitor p21{sup Cip1}, which was abolished by pretreatment with actinomycin D. A p38 MAPK specific inhibitor, SB203580, blocked the increase of p21{sup Cip1} mRNA in response to MC. Treatment with indigo, another AhR ligand, failed to increase of p21{sup Cip1} mRNA, although up-regulation of mRNA for CYP1A1 was observed. These data suggest AhR in Leydig cells mediates growth inhibition by inducing p21{sup Cip1}.

  19. Mitochondrial dysfunction mediated by cytoplasmic acidification results in pollen tube growth cessation in Pyrus pyrifolia.

    PubMed

    Gao, Yongbin; Zhou, Hongsheng; Chen, Jianqing; Jiang, Xueting; Tao, Shutian; Wu, Juyou; Zhang, Shaoling

    2015-04-01

    The length of pollen tubes grown in synthetic media is normally shorter than those grown in vivo. However, the mechanism(s) underlying the cessation of pollen tube growth under culture conditions remain(s) largely unknown. Here, we report a previously unknown correlation between vacuolar function and the cell's ability to sustain mitochondrial functions in pear pollen tubes. The pear pollen tubes in vitro grew slowly after 15 hours post-cultured (HPC) and nearly ceased growth at 18 HPC. There was increased malondialdehyde content and membrane ion leakage at 15 HPC compared with 12 HPC. Furthermore, cytoplasmic acidification mainly mediated by decreased vacuolar H(+)-ATPase [V-ATPase, Enzyme Commission (EC) 3.6.1.3] activity was observed in pollen tubes after 15 HPC, and this further resulted in mitochondrial dysfunction, including mitochondrial structure disruption, mitochondrial membrane potential collapse and decreases in both oxygen consumption and ATP production. Our findings suggest that vacuoles and mitochondria intimately linked in regulating pollen tube elongation.

  20. Knockdown of Mediator Complex Subunit 19 Suppresses the Growth and Invasion of Prostate Cancer Cells

    PubMed Central

    Zhao, Hongwei; Lv, Wei; Chen, Jian; Wan, Fengchun; Liu, Dongfu; Gao, Zhenli; Wu, Jitao

    2017-01-01

    Prostate cancer (PCa) is one of the most common cancers in elderly men. Mediator Complex Subunit 19 (Med19) is overexpressed and plays promotional roles in many cancers. However, the roles of Med19 in PCa are still obscure. In this study, by using immunohistochemical staining, we found higher expression level of Med19 in PCa tissues than in adjacent benign prostate tissues. We then knocked down the Med19 expression in PCa cell lines LNCaP and PC3 by using lentivirus siRNA. Cell proliferation, anchor-independent growth, migration, and invasion were suppressed in Med19 knockdown PCa cells. In nude mice xenograft model, we found that Med19 knockdown PCa cells formed smaller tumors with lower proliferation index than did control cells. In the mechanism study, we found that Med19 could regulate genes involved in cell proliferation, cell cycle, and epithelial-mesenchymal transition, including P27, pAKT, pPI3K, IGF1R, E-Cadherin, N-Cadherin, Vimentin, ZEB2, Snail-1 and Snail-2. Targeting Med19 in PCa cells could inhibit the PCa growth and metastasis, and might be a therapeutic option for PCa in the future. PMID:28125713

  1. Medial gastrocnemius muscle growth during adolescence is mediated by increased fascicle diameter rather than by longitudinal fascicle growth.

    PubMed

    Weide, Guido; Huijing, Peter A; Maas, Josina C; Becher, Jules G; Harlaar, Jaap; Jaspers, Richard T

    2015-06-01

    Using a cross-sectional design, the purpose of this study was to determine how pennate gastrocnemius medialis (GM) muscle geometry changes as a function of adolescent age. Sixteen healthy adolescent males (aged 10-19 years) participated in this study. GM muscle geometry was measured within the mid-longitudinal plane obtained from a 3D voxel-array composed of transverse ultrasound images. Images were taken at footplate angles corresponding to standardised externally applied footplate moments (between 4 Nm plantar flexion and 6 Nm dorsal flexion). Muscle activity was recorded using surface electromyography (EMG), expressed as a percentage of maximal voluntary contraction (%MVC). To minimise the effects of muscle excitation, EMG inclusion criteria were set at <10% of MVC. In practice, however, normalised EMG levels were much lower. For adolescent subjects with increasing ages, GM muscle (belly) length increased due to an increase in the length component of the physiological cross-sectional area measured within the mid-longitudinal plane. No difference was found between fascicles at different ages, but the aponeurosis length and pennation angle increased by 0.5 cm year(-1) and 0.5° per year, respectively. Footplate angles corresponding to externally applied 0 and 4 Nm plantarflexion moments were not associated with different adolescent ages. In contrast, footplate angles corresponding to externally applied 4 and 6 Nm dorsal flexion moments decreased by 10° between 10 and 19 years. In conclusion, we found that in adolescents' pennate GM muscles, longitudinal muscle growth is mediated predominantly by increased muscle fascicle diameter.

  2. Transforming growth factor-beta1 inhibits tissue engineering cartilage absorption via inducing the generation of regulatory T cells.

    PubMed

    Li, Chichi; Bi, Wei; Gong, Yiming; Ding, Xiaojun; Guo, Xuehua; Sun, Jian; Cui, Lei; Yu, Youcheng

    2016-02-01

    The objective of the present study was to explore the mechanisms of transforming growth factor (TGF)-β1 inhibiting the absorption of tissue engineering cartilage. We transfected TGF-β1 gene into bone marrow mesenchymal stem cells (BMMSCs) and co-cultured with interferon (IFN)-γ and tumour necrosis factor (TNF)-α and CD4(+) CD25(-) T lymphocytes. We then characterized the morphological changes, apoptosis and characterization of chondrogenic-committed cells from TGF-β1(+) BMMSCs and explored their mechanisms. Results showed that BMMSCs apoptosis and tissue engineering cartilage absorption in the group with added IFN-γ and TNF-α were greater than in the control group. In contrast, there was little BMMSC apoptosis and absorption by tissue engineering cartilage in the group with added CD4(+) CD25(-) T lymphocytes; Foxp3(+) T cells and CD25(+) CD39(+) T cells were found. In contrast, no type II collagen or Foxp3(+) T cells or CD25(+) CD39(+) T cells was found in the TGF-β1(-) BMMSC group. The data suggest that IFN-γ and TNF-α induced BMMSCs apoptosis and absorption of tissue engineering cartilage, but the newborn regulatory T (Treg) cells inhibited the function of IFN-γ and TNF-α and protected BMMSCs and tissue engineering cartilage. TGF-β1not only played a cartilage inductive role, but also inhibited the absorption of tissue engineering cartilage. The pathway proposed in our study may simulate the actual reaction procedure after implantation of BMMSCs and tissue engineering cartilage in vivo.

  3. Combined Large-Scale Phenotyping and Transcriptomics in Maize Reveals a Robust Growth Regulatory Network1[OPEN

    PubMed Central

    Herman, Dorota; Slabbinck, Bram; Pè, Mario Enrico

    2016-01-01

    Leaves are vital organs for biomass and seed production because of their role in the generation of metabolic energy and organic compounds. A better understanding of the molecular networks underlying leaf development is crucial to sustain global requirements for food and renewable energy. Here, we combined transcriptome profiling of proliferative leaf tissue with in-depth phenotyping of the fourth leaf at later stages of development in 197 recombinant inbred lines of two different maize (Zea mays) populations. Previously, correlation analysis in a classical biparental mapping population identified 1,740 genes correlated with at least one of 14 traits. Here, we extended these results with data from a multiparent advanced generation intercross population. As expected, the phenotypic variability was found to be larger in the latter population than in the biparental population, although general conclusions on the correlations among the traits are comparable. Data integration from the two diverse populations allowed us to identify a set of 226 genes that are robustly associated with diverse leaf traits. This set of genes is enriched for transcriptional regulators and genes involved in protein synthesis and cell wall metabolism. In order to investigate the molecular network context of the candidate gene set, we integrated our data with publicly available functional genomics data and identified a growth regulatory network of 185 genes. Our results illustrate the power of combining in-depth phenotyping with transcriptomics in mapping populations to dissect the genetic control of complex traits and present a set of candidate genes for use in biomass improvement. PMID:26754667

  4. FoxO proteins mediate hypoxic induction of connective tissue growth factor in endothelial cells.

    PubMed

    Samarin, Jana; Wessel, Julia; Cicha, Iwona; Kroening, Sven; Warnecke, Christina; Goppelt-Struebe, Margarete

    2010-02-12

    Hypoxia, a driving force in neovascularization, promotes alterations in gene expression mediated by hypoxia-inducible factor (HIF)-1alpha. Connective tissue growth factor (CTGF, CCN2) is a modulator of endothelial cell growth and migration, but its regulation by hypoxia is poorly understood. Therefore, we analyzed signaling pathways involved in the regulation of CTGF by hypoxia in endothelial cells. Exposure to low oxygen tension or treatment with the hypoxia-mimetic dimethyloxalyl glycine (DMOG) stabilized HIF-1alpha and up-regulated CTGF in human umbilical vein endothelial cells and in a murine microvascular endothelial cell line. Induction of CTGF correlated with a HIF-dependent increase in protein and mRNA levels, and nuclear accumulation of the transcription factor FoxO3a. By contrast, gene expression and cellular localization of FoxO1 were not significantly altered by hypoxia. Expression of CTGF was strongly reduced by siRNA silencing of FoxO1 or FoxO3a. Furthermore, nuclear exclusion of FoxO1/3a transcription factors by inhibition of serine/threonine protein phosphatases by okadaic acid inhibited CTGF expression, providing evidence for both FoxO proteins as regulators of CTGF expression. The DMOG-stimulated induction of CTGF was further increased when endothelial cells were co-incubated with transforming growth factor-beta, an activator of Smad signaling. Activation of RhoA-Rho kinase signaling by the microtubule-disrupting drug combretastatin A4 also enhanced the DMOG-induced CTGF expression, thus placing CTGF induction by hypoxia in a network of interacting signaling pathways. Our findings provide evidence that FoxO1, hypoxia-stimulated expression of FoxO3a and its nuclear accumulation are required for the induction of CTGF by hypoxia in endothelial cells.

  5. Endostatin inhibits androgen-independent prostate cancer growth by suppressing nuclear receptor-mediated oxidative stress.

    PubMed

    Lee, Joo Hyoung; Kang, Minsung; Wang, Hong; Naik, Gurudatta; Mobley, James A; Sonpavde, Guru; Garvey, W Timothy; Darley-Usmar, Victor M; Ponnazhagan, Selvarangan

    2017-04-01

    Androgen-deprivation therapy has been identified to induce oxidative stress in prostate cancer (PCa), leading to reactivation of androgen receptor (AR) signaling in a hormone-refractory manner. Thus, antioxidant therapies have gained attention as adjuvants for castration-resistant PCa. Here, we report for the first time that human endostatin (ES) prevents androgen-independent growth phenotype in PCa cells through its molecular targeting of AR and glucocorticoid receptor (GR) and downstream pro-oxidant signaling. This reversal after ES treatment significantly decreased PCa cell proliferation through down-regulation of GR and up-regulation of manganese superoxide dismutase and reduced glutathione levels. Proteome and biochemical analyses of ES-treated PCa cells further indicated a significant up-regulation of enzymes in the major reactive oxygen species (ROS) scavenging machinery, including catalase, glutathione synthetase, glutathione reductase, NADPH-cytochrome P450 reductase, biliverdin reductase, and thioredoxin reductase, resulting in a concomitant reduction of intracellular ROS. ES further augmented the antioxidant system through up-regulation of glucose influx, the pentose phosphate pathway, and NAD salvaging pathways. This shift in cancer cell redox homeostasis by ES significantly decreased the effect of protumorigenic oxidative machinery on androgen-independent PCa growth, suggesting that ES can suppress GR-induced resistant phenotype upon AR antagonism and that the dual targeting action of ES on AR and GR can be further translated to PCa therapy.-Lee, J. H., Kang, M., Wang, H., Naik, G., Mobley, J. A., Sonpavde, G., Garvey, W. T., Darley-Usmar, V. M., Ponnazhagan, S. Endostatin inhibits androgen-independent prostate cancer growth by suppressing nuclear receptor-mediated oxidative stress.

  6. Previtellogenic oocyte growth in salmon: relationships among body growth, plasma insulin-like growth factor-1, estradiol-17beta, follicle-stimulating hormone and expression of ovarian genes for insulin-like growth factors, steroidogenic-acute regulatory protein and receptors for gonadotropins, growth hormone, and somatolactin.

    PubMed

    Campbell, B; Dickey, J; Beckman, B; Young, G; Pierce, A; Fukada, H; Swanson, P

    2006-07-01

    Body growth during critical periods is known to be an important factor in determining the age of maturity and fecundity in fish. However, the endocrine mechanisms controlling oogenesis in fish and the effects of growth on this process are poorly understood. In this study interactions between the growth and reproductive systems were examined by monitoring changes in various components of the FSH-ovary axis, plasma insulin-like growth factor 1 (Igf1), and ovarian gene expression in relation to body and previtellogenic oocyte growth in coho salmon. Samples were collected from females during two hypothesized critical periods when growth influences maturation in this species. Body growth during the fall-spring months was strongly related to the degree of oocyte development, with larger fish possessing more advanced oocytes than smaller, slower growing fish. The accumulation of cortical alveoli in the oocytes was associated with increases in plasma and pituitary FSH, plasma estradiol-17beta, and ovarian steroidogenic acute regulatory protein (star) gene expression, whereas ovarian transcripts for growth hormone receptor and somatolactin receptor decreased. As oocytes accumulated lipid droplets, a general increase occurred in plasma Igf1 and components of the FSH-ovary axis, including plasma FSH, estradiol-17beta, and ovarian mRNAs for gonadotropin receptors, star, igf1, and igf2. A consistent positive relationship between plasma Igf1, estradiol-17beta, and pituitary FSH during growth in the spring suggests that these factors are important links in the mechanism by which body growth influences the rate of oocyte development.

  7. Extracellular signal-regulated kinase 2 (ERK-2) mediated phosphorylation regulates nucleo-cytoplasmic shuttling and cell growth control of Ras-associated tumor suppressor protein, RASSF2

    SciTech Connect

    Kumari, Gita; Mahalingam, S.

    2009-10-01

    Ras GTPase controls the normal cell growth through binding with an array of effector molecules, such as Raf and PI3-kinase in a GTP-dependent manner. RASSF2, a member of the Ras association domain family, is known to be involved in the suppression of cell growth and is frequently down-regulated in various tumor tissues by promoter hypermethylation. In the present study, we demonstrate that RASSF2 shuttles between nucleus and cytoplasm by a signal-mediated process and its export from the nucleus is sensitive to leptomycin B. Amino acids between 240 to 260 in the C-terminus of RASSF2 harbor a functional nuclear export signal (NES), which is necessary and sufficient for efficient export of RASSF2 from the nucleus. Substitution of conserved Ile254, Val257 and Leu259 within the minimal NES impaired RASSF2 export from the nucleus. In addition, wild type but not the nuclear export defective RASSF2 mutant interacts with export receptor, CRM-1 and exported from the nucleus. Surprisingly, we observed nucleolar localization for the nuclear export defective mutant suggesting the possibility that RASSF2 may localize in different cellular compartments transiently in a cell cycle dependent manner and the observed nuclear localization for wild type protein may be due to faster export kinetics from the nucleolus. Furthermore, our data suggest that RASSF2 is specifically phosphorylated by MAPK/ERK-2 and the inhibitors of MAPK pathway impair the phosphorylation and subsequently block the export of RASSF2 from the nucleus. These data clearly suggest that ERK-2 mediated phosphorylation plays an important role in regulating the nucleo-cytoplasmic shuttling of RASSF2. Interestingly, nuclear import defective mutant of RASSF2 failed to induce cell cycle arrest at G1/S phase and apoptosis suggesting that RASSF2 regulates cell growth in a nuclear localization dependent manner. Collectively, these data provided evidence for the first time that MAPK/ERK-2 mediated phosphorylation regulates

  8. Classical dendritic cells are required for dietary antigen-mediated peripheral regulatory T cell and tolerance induction

    PubMed Central

    Esterházy, Daria; Loschko, Jakob; London, Mariya; Jove, Veronica; Oliveira, Thiago Y.; Mucida, Daniel

    2016-01-01

    Oral tolerance prevents pathological inflammatory responses towards innocuous foreign antigens via peripheral regulatory T cells (pTreg cells). However, whether a particular subset of antigen-presenting cells (APCs) is required during dietary antigen exposure to instruct naïve CD4+ T cells to differentiate into pTreg cells has not been defined. Using myeloid lineage-specific APC depletion in mice, we found that monocyte-derived APCs are dispensable, while classical dendritic cells (cDCs) are critical for pTreg cell induction and oral tolerance. CD11b− cDCs from the gut-draining lymph nodes efficiently induced pTreg cells, and conversely, loss of IRF8-dependent CD11b− cDCs impaired their polarization, although oral tolerance remained intact. These data reveal the hierarchy of cDC subsets in pTreg cell induction and their redundancy during oral tolerance development. PMID:27019226

  9. p38 MAPK-inhibited dendritic cells induce superior antitumor immune responses and overcome regulatory T cell-mediated immunosuppression

    PubMed Central

    Lu, Yong; Zhang, Mingjun; Wang, Siqing; Hong, Bangxing; Wang, Zhiqiang; Li, Haiyan; Zheng, Yuhuan; Yang, Jing; Davis, Richard E.; Qian, Jianfei; Hou, Jian; Yi, Qing

    2014-01-01

    Dendritic cell (DC)-based cancer immunotherapy is a promising method but so far has demonstrated limited clinical benefits. Regulatory T cells (Treg) represent a major obstacle to cancer immunotherapy approaches. Here we show that inhibiting p38 MAPK during DC differentiation enables DCs to activate tumor-specific effector T cells (Teff), inhibiting the conversion of Treg and compromising Treg inhibitory effects on Teff. Inhibition of p38 MAPK in DCs lowers expression of PPARγ, activating p50 and upregulation of OX40L expression in DCs. OX40L/OX40 interactions between DCs and Teff and/or Treg are critical for priming effective and therapeutic antitumor responses. Similarly, p38 MAPK inhibition also augments the T cell-stimulatory capacity of human monocyte-derived DCs in the presence of Treg. These findings contribute to ongoing efforts to improve DC-based immunotherapy in human cancers. PMID:24957461

  10. Sugar signals and the control of plant growth and development.

    PubMed

    Lastdrager, Jeroen; Hanson, Johannes; Smeekens, Sjef

    2014-03-01

    Sugars have a central regulatory function in steering plant growth. This review focuses on information presented in the past 2 years on key players in sugar-mediated plant growth regulation, with emphasis on trehalose 6-phosphate, target of rapamycin kinase, and Snf1-related kinase 1 regulatory systems. The regulation of protein synthesis by sugars is fundamental to plant growth control, and recent advances in our understanding of the regulation of translation by sugars will be discussed.

  11. Targeting extracellular domains D4 and D7 of vascular endothelial growth factor receptor 2 reveals allosteric receptor regulatory sites.

    PubMed

    Hyde, Caroline A C; Giese, Alexandra; Stuttfeld, Edward; Abram Saliba, Johan; Villemagne, Denis; Schleier, Thomas; Binz, H Kaspar; Ballmer-Hofer, Kurt

    2012-10-01

    Vascular endothelial growth factors (VEGFs) activate three receptor tyrosine kinases, VEGFR-1, -2, and -3, which regulate angiogenic and lymphangiogenic signaling. VEGFR-2 is the most prominent receptor in angiogenic signaling by VEGF ligands. The extracellular part of VEGF receptors consists of seven immunoglobulin homology domains (Ig domains). Earlier studies showed that domains 2 and 3 (D23) mediate ligand binding, while structural analysis of dimeric ligand/receptor complexes by electron microscopy and small-angle solution scattering revealed additional homotypic contacts in membrane-proximal Ig domains D4 and D7. Here we show that D4 and D7 are indispensable for receptor signaling. To confirm the essential role of these domains in signaling, we isolated VEGFR-2-inhibitory "designed ankyrin repeat proteins" (DARPins) that interact with D23, D4, or D7. DARPins that interact with D23 inhibited ligand binding, receptor dimerization, and receptor kinase activation, while DARPins specific for D4 or D7 did not prevent ligand binding or receptor dimerization but effectively blocked receptor signaling and functional output. These data show that D4 and D7 allosterically regulate VEGFR-2 activity. We propose that these extracellular-domain-specific DARPins represent a novel generation of receptor-inhibitory drugs for in vivo applications such as targeting of VEGFRs in medical diagnostics and for treating vascular pathologies.

  12. Targeting Extracellular Domains D4 and D7 of Vascular Endothelial Growth Factor Receptor 2 Reveals Allosteric Receptor Regulatory Sites

    PubMed Central

    Hyde, Caroline A. C.; Giese, Alexandra; Stuttfeld, Edward; Abram Saliba, Johan; Villemagne, Denis; Schleier, Thomas; Binz, H. Kaspar

    2012-01-01

    Vascular endothelial growth factors (VEGFs) activate three receptor tyrosine kinases, VEGFR-1, -2, and -3, which regulate angiogenic and lymphangiogenic signaling. VEGFR-2 is the most prominent receptor in angiogenic signaling by VEGF ligands. The extracellular part of VEGF receptors consists of seven immunoglobulin homology domains (Ig domains). Earlier studies showed that domains 2 and 3 (D23) mediate ligand binding, while structural analysis of dimeric ligand/receptor complexes by electron microscopy and small-angle solution scattering revealed additional homotypic contacts in membrane-proximal Ig domains D4 and D7. Here we show that D4 and D7 are indispensable for receptor signaling. To confirm the essential role of these domains in signaling, we isolated VEGFR-2-inhibitory “designed ankyrin repeat proteins” (DARPins) that interact with D23, D4, or D7. DARPins that interact with D23 inhibited ligand binding, receptor dimerization, and receptor kinase activation, while DARPins specific for D4 or D7 did not prevent ligand binding or receptor dimerization but effectively blocked receptor signaling and functional output. These data show that D4 and D7 allosterically regulate VEGFR-2 activity. We propose that these extracellular-domain-specific DARPins represent a novel generation of receptor-inhibitory drugs for in vivo applications such as targeting of VEGFRs in medical diagnostics and for treating vascular pathologies. PMID:22801374

  13. Cognitive and Emotion-Regulatory Mediators of the Relationship between Behavioral Approach System Sensitivity and Non-Suicidal Self-Injury Frequency

    PubMed Central

    Burke, Taylor A.; Stange, Jonathan P.; Hamilton, Jessica L.; Cohen, Jonah N.; O’Garro-Moore, Jared; Daryanani, Issar; Abramson, Lyn Y.; Alloy, Lauren B.

    2014-01-01

    Non-suicidal self-injury (NSSI) is highly prevalent among late adolescents and predicts the onset of suicidal ideation and behavior. Although research has established an association between the behavioral approach system (BAS) and NSSI, less research has explored mechanisms underlying this relationship. The authors examined negative and positive emotion regulation patterns, as well as the BAS-relevant cognitive style of self-criticism, as potential mechanisms through which a hypersensitive BAS might be related to NSSI frequency. Late adolescents (N=177) with high and moderate BAS levels completed measures of self-criticism, positive emotion regulation, brooding, and both lifetime and last year frequency of NSSI. Results indicated that self-criticism and positive emotion dampening independently mediated the relationship between BAS and last year frequency of NSSI. Self-criticism also mediated the relationship between BAS and lifetime frequency of NSSI. Results suggest that cognitive and emotion-regulatory styles may help to explain why high BAS individuals are likely to engage in NSSI. PMID:25443691

  14. Amphiregulin activates regulatory T lymphocytes and suppresses CD8+ T cell-mediated anti-tumor response in hepatocellular carcinoma cells

    PubMed Central

    Yuan, Chun-Hui; Sun, Xiao-Ming; Zhu, Cheng-Liang; Liu, Shao-Ping; Wu, Long; Chen, Hao; Feng, Mao-Hui; Wu, Ke; Wang, Fu-Bing

    2015-01-01

    CD8+ T cell-mediated immune response plays an important role in inhibiting progression of hepatocellular carcinoma (HCC). For strategic immunotherapy, it is critical to understand why some of the tumor cells escape from this immune attack. In this study, we investigated how HCC cells alter endogenous anti-tumor immunity and their related signaling pathways. We found that HCC cells, both in vitro and in vivo, substantially secret and express amphiregulin (AR). AR in turn activates immunosuppressive function of intratumoral CD4+Foxp3+ regulatory T cells (Tregs), a major inhibitor of CD8+ T cells. Using either lentiviral siRNA, or AR neutralizing antibody, we blocked the expression and function of AR to test the specificity of AR mediated activation of Tregs, Biochemical and cell biology studies were followed and confirmed that blocking of AR inhibited Tregs activation. In addition, we found that AR can trigger the activation of rapamycin complex 1(mTORC1) signaling in Tregs. The mTORC1 inhibitor rapamycin treatment led to compromise Treg function and resulted in enhancing anti-tumor function of CD8+ T cells. Blocking AR/EGFR signaling in Tregs with Gefitinib also enhanced anti-tumor immunity and decreased tumor size in a mouse xenograft tumor model. Taken together, our study suggested a novel mechanism of functional interaction between HCC and Tregs for regulating anti-tumor function of CD8+ T cells. PMID:26451607

  15. Exposure of FVIII in the Presence of Phosphatidyl Serine Reduces Generation of Memory B-Cells and Induces Regulatory T-Cell-Mediated Hyporesponsiveness in Hemophilia A Mice.

    PubMed

    Ramakrishnan, Radha; Davidowitz, Andrew; Balu-Iyer, Sathy V

    2015-08-01

    A major complication of replacement therapy with Factor VIII (FVIII) for hemophilia A (HA) is the development of unwanted immune responses. Previous studies showed that administration of FVIII in the presence of phosphatidyl serine (PS) reduced the development of anti-FVIII antibodies in HA mice. However, the impact of PS-mediated effects on immunological memory, such as generation of memory B-cells, is not clear. The effect of PS on memory B-cells was therefore investigated using adoptive transfer approach in FVIII(-/-) HA mice. Adoptive transfer of memory B-cells from a PS-FVIII-treated group to naïve mice followed by challenge of the recipient mice with FVIII showed a significantly reduced anti-FVIII antibody response in the recipient mice, compared with animals that received memory B-cells from free FVIII and FVIII-charge matched phosphatidyl glycerol (PG) group. The decrease in memory B-cell response is accompanied by an increase in FoxP3 expressing regulatory T-cells (Tregs). Flow cytometry studies showed that the generation of Tregs is higher in PS-treated animals as compared with FVIII and FVIII-PG treated animals. The PS-mediated hyporesponsiveness was found to be antigen-specific. The PS-FVIII immunization showed hyporesponsiveness toward FVIII rechallenge but not against ovalbumin (OVA) rechallenge, an unrelated antigen. This demonstrates that PS reduces immunologic memory of FVIII and induces antigen-specific peripheral tolerance in HA mice.

  16. Reappraising contemporary theories of subcortical participation in language: proposing an interhemispheric regulatory function for the subthalamic nucleus (STN) in the mediation of high-level linguistic processes.

    PubMed

    Whelan, Brooke-Mai; Murdoch, Bruce E; Theodoros, Deborah G; Silburn, Peter; Hall, Bruce

    2004-02-01

    Apropos the basal ganglia, the dominant striatum and globus pallidus internus (GPi) have been hypothesised to represent integral components of subcortical language circuitry. Working subcortical language theories, however, have failed thus far to consider a role for the STN in the mediation of linguistic processes, a structure recently defined as the driving force of basal ganglia output. The aim of this research was to investigate the impact of surgically induced functional inhibition of the STN upon linguistic abilities, within the context of established models of basal ganglia participation in language. Two males with surgically induced 'lesions' of the dominant and non-dominant dorsolateral STN, aimed at relieving Parkinsonian motor symptoms, served as experimental subjects. General and high-level language profiles were compiled for each subject up to 1 month prior to and 3 months following neurosurgery, within the drug-on state (i.e., when optimally medicated). Comparable post-operative alterations in linguistic performance were observed subsequent to surgically induced functional inhibition of the left and right STN. More specifically, higher proportions of reliable decline as opposed to improvement in post-operative performance were demonstrated by both subjects on complex language tasks, hypothesised to entail the interplay of cognitive-linguistic processes. The outcomes of the current research challenge unilateralised models of functional basal ganglia organisation with the proposal of a potential interhemispheric regulatory function for the STN in the mediation of high-level linguistic processes.

  17. Exacerbation of spontaneous autoimmune nephritis following regulatory T cell depletion in B cell lymphoma 2-interacting mediator knock-out mice.

    PubMed

    Wang, Y M; Zhang, G Y; Wang, Y; Hu, M; Zhou, J J; Sawyer, A; Cao, Q; Wang, Y; Zheng, G; Lee, V W S; Harris, D C H; Alexander, S I

    2017-02-02

    Regulatory T cells (Tregs ) have been recognized as central mediators for maintaining peripheral tolerance and limiting autoimmune diseases. The loss of Tregs or their function has been associated with exacerbation of autoimmune disease. However, the temporary loss of Tregs in the chronic spontaneous disease model has not been investigated. In this study, we evaluated the role of Tregs in a novel chronic spontaneous glomerulonephritis model of B cell lymphoma 2-interacting mediator (Bim) knock-out mice by transient depleting Tregs . Bim is a pro-apoptotic member of the B cell lymphoma 2 (Bcl-2) family. Bim knock-out (Bim(-/-) ) mice fail to delete autoreactive T cells in thymus, leading to chronic spontaneous autoimmune kidney disease. We found that Treg depletion in Bim(-/-) mice exacerbated the kidney injury with increased proteinuria, impaired kidney function, weight loss and greater histological injury compared with wild-type mice. There was a significant increase in interstitial infiltrate of inflammatory cells, antibody deposition and tubular damage. Furthermore, the serum levels of cytokines interleukin (IL)-2, IL-4, IL-6, IL-10, IL-17α, interferon (IFN)-γ and tumour necrosis factor (TNF)-α were increased significantly after Treg depletion in Bim(-/-) mice. This study demonstrates that transient depletion of Tregs leads to enhanced self-reactive T effector cell function followed by exacerbation of kidney disease in the chronic spontaneous kidney disease model of Bim-deficient mice.

  18. A Structural Model of Parental Alcoholism, Family Functioning, and Psychological Health: The Mediating Effects of Hardiness and Personal Growth Orientation.

    ERIC Educational Resources Information Center

    Robitschek, Christine; Kashubeck, Susan

    1999-01-01

    This study sought to: (a) determine whether personal-growth orientation and hardiness mediated the relations of parental alcoholism and family functioning to psychological well-being and distress; (b) determine whether this model was invariant across men and women; and (c) examine the role of parental alcoholism in a model that included family…

  19. Redox-mediated activation of latent transforming growth factor-beta 1

    NASA Technical Reports Server (NTRS)

    Barcellos-Hoff, M. H.; Dix, T. A.; Chatterjee, A. (Principal Investigator)

    1996-01-01

    Transforming growth factor beta 1 (TGF beta) is a multifunctional cytokine that orchestrates response to injury via ubiquitous cell surface receptors. The biological activity of TGF beta is restrained by its secretion as a latent complex (LTGF beta) such that activation determines the extent of TGF beta activity during physiological and pathological events. TGF beta action has been implicated in a variety of reactive oxygen-mediated tissue processes, particularly inflammation, and in pathologies such as reperfusion injury, rheumatoid arthritis, and atherosclerosis. It was recently shown to be rapidly activated after in vivo radiation exposure, which also generates reactive oxygen species (ROS). In the present studies, the potential for redox-mediated LTGF beta activation was investigated using a cell-free system in which ROS were generated in solution by ionizing radiation or metal ion-catalyzed ascorbate reaction. Irradiation (100 Gray) of recombinant human LTGF beta in solution induced 26% activation compared with that elicited by standard thermal activation. Metal-catalyzed ascorbate oxidation elicited extremely efficient recombinant LTGF beta activation that matched or exceeded thermal activation. The efficiency of ascorbate activation depended on ascorbate concentrations and the presence of transition metal ions. We postulate that oxidation of specific amino acids in the latency-conferring peptide leads to a conformation change in the latent complex that allows release of TGF beta. Oxidative activation offers a novel route for the involvement of TGF beta in tissue processes in which ROS are implicated and endows LTGF beta with the ability to act as a sensor of oxidative stress and, by releasing TGF beta, to function as a signal for orchestrating the response of multiple cell types. LTGF beta redox sensitivity is presumably directed toward recovery of homeostasis; however, oxidation may also be a mechanism of LTGF beta activation that can be deleterious during

  20. UBPY-mediated epidermal growth factor receptor (EGFR) de-ubiquitination promotes EGFR degradation.

    PubMed

    Alwan, Husam A J; van Leeuwen, Jeroen E M

    2007-01-19

    Whereas poly-ubiquitination targets protein substrates for proteasomal degradation, mono-ubiquitination is known to regulate protein trafficking in the endosomal system and to target cargo proteins for lysosomal degradation. The role of the de-ubiquitinating enzymes AMSH and UBPY in endosomal trafficking of cargo proteins such as the epidermal growth factor receptor (EGFR) has only very recently been the subject of study and is already a matter of debate. Although one report (Mizuno, E., Iura, T., Mukai, A., Yoshimori, T., Kitamura, N., and Komada, M. (2005) Mol. Biol. Cell 16, 5163-5174) concludes that UBPY negatively regulates EGFR degradation by de-ubiquitinating the EGFR on endosomes, another report (Row, P. E., Prior, I. A., McCullough, J., Clague, M. J., and Urbe, S. (2006) J. Biol. Chem. 281, 12618-12624) concludes that UBPY-mediated EGFR de-ubiquitination is essential for EGFR degradation. Here, we demonstrate that Usp8/UBPY, the mammalian ortholog of budding yeast Ubp4/Doa4, constitutively co-precipitates in a bivalent manner with the EGFR. Moreover, UBPY is a substrate for Src-family tyrosine kinases that are activated after ligand-induced EGFR activation. Using overexpression of three different recombinant dominant negative UBPY mutants (UBPY C748A mutant, UBPY 1-505, and UBPY 640-1080) in NIH3T3 and HEK293 cells, we demonstrate that UBPY affects both constitutive and ligand-induced (i) EGFR ubiquitination, (ii) EGFR expression levels, and (iii) the appearance of intermediate EGFR degradation products as well as (iv) downstream mitogen-activated protein kinase signal transduction. Our findings provide further evidence in favor of the model that UBPY-mediated EGFR de-ubiquitination promotes EGFR degradation.

  1. Hypoxia simultaneously alters satellite cell-mediated angiogenesis and hepatocyte growth factor expression.

    PubMed

    Flann, K L; Rathbone, C R; Cole, L C; Liu, X; Allen, R E; Rhoads, R P

    2014-05-01

    Skeletal muscle regeneration is a multifaceted process requiring the spatial and temporal coordination of myogenesis as well as angiogenesis. Hepatocyte growth factor (HGF) plays a pivotal role in myogenesis by activating satellite cells (SC) in regenerating muscle and likely plays a role as a contributor to revascularization. Moreover, repair of a functional blood supply is critical to ameliorate tissue ischemia and restore skeletal muscle function, however effects of hypoxia on satellite cell-mediated angiogenesis remain unclear. The objective of this study was to examine the role of HGF and effect of hypoxia on the capacity of satellite cells to promote angiogenesis. To characterize the role of HGF, a microvascular fragment (MVF) culture model coupled with satellite cell conditioned media (CM) was employed. The activity of HGF was specifically blocked in SC CM reducing sprout length compared to control CM. In contrast, MVF sprout number did not differ between control or HGF-deficient SC CM media. Next, we cultured MVF in the presence of CM from satellite cells exposed to normoxic (20% O2 ) or hypoxic (1% O2 ) conditions. Hypoxic CM recapitulated a MVF angiogenic response identical to HGF deficient satellite cell CM. Hypoxic conditions increased satellite cell HIF-1α protein abundance and VEGF mRNA abundance but decreased HGF mRNA abundance compared to normoxic satellite cells. Consistent with reduced HGF gene expression, HGF promoter activity decreased during hypoxia. Taken together, this data indicates that hypoxic modulation of satellite cell-mediated angiogenesis involves a reduction in satellite cell HGF expression.

  2. Vascular endothelial growth factor mediates reactive angiogenesis in the postnatal developing brain.

    PubMed

    Ment, L R; Stewart, W B; Fronc, R; Seashore, C; Mahooti, S; Scaramuzzino, D; Madri, J A

    1997-05-20

    Although chronic sublethal hypoxia has been shown to promote angiogenesis in the developing brain, the pathogenesis of this response is unknown. We hypothesized that this response may be mediated in part by vascular endothelial growth factor (VEGF). We reared newborn rats (P3) in a chamber with FIO2 of 9.5 +/- 1% (exposed, E). At P33, the animals were removed from the chamber and the brains prepared for immunohistochemistry, mRNA extraction, or horseradish peroxidase (HRP) permeability studies. We also isolated beagle brain germinal matrix endothelial cells from PND 1 beagle pups and placed them in three-dimensional (3-D) coculture with PND 1 rat forebrain astrocytes. Cultures were grown for 6 days in 11% O2 and compared to control 3-D cocultures. When compared to age-matched controls, the experimental rats had significantly increased cortical vascular density (vessels/mm2: 518 +/- 18 vs. 400 +/- 15, P = 0.025). HRP studies demonstrated significantly increased permeability in all cortical vessels examined in experimental rats compared to controls. Compared to controls, VEGF mRNA from hypoxic pups was increased 2.4 times, and immunohistochemical studies of VEGF protein confirmed this finding. Similarly, when compared to controls, hypoxic cocultures of brain microvascular endothelial cells and astrocytes demonstrated significant increase in tubelike structures representing in vitro angiogenesis. Additionally, astrocyte VEGF protein levels increased 4.4-fold in hypoxic compared to control astrocyte cultures and VEGF protein levels increased 1.7-fold in hypoxic compared to control cocultures. Finally, addition of VEGF (10 ng/ml culture medium) to BBMEC alone in 3-D culture elicited not only significant proliferation (P = 0.001) but also increased tube formation. These data demonstrate that the developing brain responds to chronic sublethal hypoxia with increases in permeability and angiogenesis and suggest that VEGF mediates this response.

  3. Ca2+/calmodulin-dependent transcriptional pathways: potential mediators of skeletal muscle growth and development.

    PubMed

    Al-Shanti, Nasser; Stewart, Claire E

    2009-11-01

    The loss of muscle mass with age and disuse has a significant impact on the physiological and social well-being of the aged; this is an increasingly important problem as the population becomes skewed towards older age. Exercise has psychological benefits but it also impacts on muscle protein synthesis and degradation, increasing muscle tissue volume in both young and older individuals. Skeletal muscle hypertrophy involves an increase in muscle mass and cross-sectional area and associated increased myofibrillar protein content. Attempts to understand the molecular mechanisms that underlie muscle growth, development and maintenance, have focused on characterising the molecular pathways that initiate, maintain and regenerate skeletal muscle. Such understanding may aid in improving targeted interventional therapies for age-related muscle loss and muscle wasting associated with diseases. Two major routes through which skeletal muscle development and growth are regulated are insulin-like growth factor I (IGF-I) and Ca(2+)/calmodulin-dependent transcriptional pathways. Many reviews have focused on understanding the signalling pathways of IGF-I and its receptor, which govern skeletal muscle hypertrophy. However, alternative molecular signalling pathways such as the Ca(2+)/calmodulin-dependent transcriptional pathways should also be considered as potential mediators of muscle growth. These latter pathways have received relatively little attention and the purpose herein is to highlight the progress being made in the understanding of these pathways and associated molecules: calmodulin, calmodulin kinases (CaMKs), calcineurin and nuclear factor of activated T-cell (NFAT), which are involved in skeletal muscle regulation. We describe: (1) how conformational changes in the Ca(2+) sensor calmodulin result in the exposure of binding pockets for the target proteins (CaMKs and calcineurin). (2) How Calmodulin consequently activates either the Ca(2+)/calmodulin-dependent kinases

  4. Early growth response 1 mediates the systemic and hepatic inflammatory response initiated by hemorrhagic shock.

    PubMed

    Prince, Jose M; Ming, Mei Jian; Levy, Ryan M; Liu, Shubing; Pinsky, David J; Vodovotz, Yoram; Billiar, Timothy R

    2007-02-01

    Hemorrhagic shock (HS) is a major cause of morbidity and mortality in trauma patients. The early growth response 1 (Egr-1) transcription factor is induced by a variety of cellular stresses, including hypoxia, and may function as a master switch to trigger the expression of numerous key inflammatory mediators. We hypothesized that HS would induce hepatic expression of Egr-1 and that Egr-1 upregulates the inflammatory response after HS. The Egr-1 mice and wild-type (WT) controls (n>or=5 for all groups) were subjected to HS alone or HS followed by resuscitation (HS/R). Other mice were subjected to a sham procedure which included general anesthesia and vessel cannulation but no shock (sham). After the HS, HS/R, or sham procedures, mice were euthanized for determination of serum concentrations of interleukin (IL) 6, IL-10, and alanine aminotransferase. Northern blot analysis was performed to evaluate Egr-1 messenger RNA (mRNA) expression. Liver whole cell lysates were evaluated for Egr-1 protein expression by Western blot analysis. Hepatic expression of IL-6, granulocyte colony-stimulating factor, and intracellular adhesion molecule 1 mRNA was determined by semiquantitative reverse transcriptase-polymerase chain reaction. The Egr-1 DNA binding was assessed using the electrophoretic mobility shift assay. Hemorrhagic shock results in a rapid and transient hepatic expression of Egr-1 mRNA in WT mice by 1 h, whereas protein and DNA binding activity was evident by 2.5 h. The Egr-1 mRNA expression diminished after 4 h of resuscitation, whereas Egr-1 protein expression and DNA binding activity persisted through resuscitation. The Egr-1 mice exhibited decreased levels of hepatic inflammatory mediators compared with WT controls with a decrease in hepatic mRNA levels of IL-6 by 42%, granulocyte colony-stimulating factor by 39%, and intracellular adhesion molecule 1 by 43%. Similarly, Egr-1 mice demonstrated a decreased systemic inflammatory response and hepatic injury after HS

  5. Growth kinetics of Ge islands during Ga-surfactant-mediated ultrahigh vacuum chemical vapor deposition on Si(001)

    NASA Astrophysics Data System (ADS)

    Portavoce, A.; Kammler, M.; Hull, R.; Reuter, M. C.; Copel, M.; Ross, F. M.

    2004-11-01

    The surfactant effect of Ga pre-deposited prior to ultrahigh vacuum (UHV) chemical vapor deposition (CVD) of Ge on Si(001) is investigated using in situ transmission electron microscopy. Island shape, nucleation and growth kinetics, and the influence of growth temperature and pressure are studied. When 1 monolayer of Ga covers the surface, Ge island nucleation occurs after a 5-10× longer growth time than without Ga for the same growth temperature and pressure, and the island density then varies with time in a more complex way than for Ga-free growth. The islands formed have a different shape and aspect ratio compared to conventional hut and dome clusters, and the critical lateral size for dislocation nucleation is reduced. Ex situ medium energy ion scattering and x-ray photoelectron scattering measurements of the total amount of Ge deposited before island nucleation demonstrate that the Ga layer decreases the surface reactivity and inhibits Ge growth. We discuss a simple model in which the surfactant-mediated growth kinetics are principally driven by the balance between Ge deposition and Ga desorption. We show that Ga surfactant-mediated UHV CVD can be used to produce Ge quantum dots with a higher surface density and a thinner wetting layer than is possible using conventional CVD.

  6. Quiescence of Memory CD8(+) T Cells Is Mediated by Regulatory T Cells through Inhibitory Receptor CTLA-4.

    PubMed

    Kalia, Vandana; Penny, Laura Anne; Yuzefpolskiy, Yevgeniy; Baumann, Florian Martin; Sarkar, Surojit

    2015-06-16

    Immune memory cells are poised to rapidly expand and elaborate effector functions upon reinfection yet exist in a functionally quiescent state. The paradigm is that memory T cells remain inactive due to lack of T cell receptor (TCR) stimuli. Here, we report that regulatory T (Treg) cells orchestrate memory T cell quiescence by suppressing effector and proliferation programs through inhibitory receptor, cytotoxic-T-lymphocyte-associated protein-4 (CTLA-4). Loss of Treg cells resulted in activation of genome-wide transcriptional programs characteristic of effector T cells and drove transitioning as well as established memory CD8(+) T cells toward terminally differentiated KLRG-1(hi)IL-7Rα(lo)GzmB(hi) phenotype, with compromised metabolic fitness, longevity, polyfunctionality, and protective efficacy. CTLA-4 functionally replaced Treg cells in trans to rescue memory T cell defects and restore homeostasis. These studies present the CTLA-4-CD28-CD80/CD86 axis as a potential target to accelerate vaccine-induced immunity and improve T cell memory quality in current cancer immunotherapies proposing transient Treg cell ablation.

  7. A specific CD4 epitope bound by tregalizumab mediates activation of regulatory T cells by a unique signaling pathway

    PubMed Central

    Helling, Bianca; König, Martin; Dälken, Benjamin; Engling, Andre; Krömer, Wolfgang; Heim, Katharina; Wallmeier, Holger; Haas, Jürgen; Wildemann, Brigitte; Fritz, Brigitte; Jonuleit, Helmut; Kubach, Jan; Dingermann, Theodor; Radeke, Heinfried H; Osterroth, Frank; Uherek, Christoph; Czeloth, Niklas; Schüttrumpf, Jörg

    2015-01-01

    CD4+CD25+ regulatory T cells (Tregs) represent a specialized subpopulation of T cells, which are essential for maintaining peripheral tolerance and preventing autoimmunity. The immunomodulatory effects of Tregs depend on their activation status. Here we show that, in contrast to conventional anti-CD4 monoclonal antibodies (mAbs), the humanized CD4-specific monoclonal antibody tregalizumab (BT-061) is able to selectively activate the suppressive properties of Tregs in vitro. BT-061 activates Tregs by binding to CD4 and activation of signaling downstream pathways. The specific functionality of BT-061 may be explained by the recognition of a unique, conformational epitope on domain 2 of the CD4 molecule that is not recognized by other anti-CD4 mAbs. We found that, due to this special epitope binding, BT-061 induces a unique phosphorylation of T-cell receptor complex-associated signaling molecules. This is sufficient to activate the function of Tregs without activating effector T cells. Furthermore, BT-061 does not induce the release of pro-inflammatory cytokines. These results demonstrate that BT-061 stimulation via the CD4 receptor is able to induce T-cell receptor-independent activation of Tregs. Selective activation of Tregs via CD4 is a promising approach for the treatment of autoimmune diseases where insufficient Treg activity has been described. Clinical investigation of this new approach is currently ongoing. PMID:25512343

  8. Chaperone-rich tumor cell lysate-mediated activation of antigen-presenting cells resists regulatory T cell suppression.

    PubMed

    Larmonier, Nicolas; Cantrell, Jessica; Lacasse, Collin; Li, Gang; Janikashvili, Nona; Situ, Elaine; Sepassi, Marjan; Andreansky, Samita; Katsanis, Emmanuel

    2008-04-01

    CD4(+)CD25(+) regulatory T lymphocytes (Tregs) critically contribute to the mechanisms of cancer-induced tolerance. These cells suppress anti-tumoral CD8(+) and CD4(+) T lymphocytes and can also restrain the function of APCs. We have previously documented the immunostimulatory effects of a chaperone-rich cell lysate (CRCL) anti-cancer vaccine. Tumor-derived CRCL induces tumor immunity in vivo, partly by promoting dendritic cell (DC) and macrophage activation. In the current study, we evaluated the effects of CD4(+)CD25(+)forkhead box P3(+) Tregs isolated from mice bearing 12B1 bcr-abl(+) leukemia on DC and macrophages that had been activated by 12B1-derived CRCL. CRCL-activated DC and macrophages resisted Treg suppression, as the production of proinflammatory cytokines, the activation of transcription factor NF-kappaB, and their immunostimulatory potential was unaffected by Tregs. Our results thus highlight CRCL as a powerful adjuvant endowed with the capacity to overcome tumor-induced Treg-inhibitory effects on APCs.

  9. Growth inhibition effect of HMME-mediated PDT on hepatocellular carcinoma HepG2 cells.

    PubMed

    Liu, Lifeng; Song, Yuanjian; Ma, Limin; Zang, Lixin; Tao, Lili; Zhang, Zhiguo; Han, Jiwu

    2014-09-01

    Photodynamic therapy (PDT) is considered a promising new strategy for liver cancer treatment. Three elements of PDT--optical output power, irradiation time, and photosensitizer concentration--play important roles in promoting cell death. This research aimed to characterize the effects of hematoporphyrin monomethyl ether (HMME)-based PDT on hepatocellular carcinoma cells HepG2 and thus elucidate the relationship between cell death and the three elements mentioned earlier. Furthermore, in this study, we present a parameter that represents the cumulative effects of these elements. The accumulation of HMME in HepG2 cells was observed by fluorescence microscopy. The absorption spectrum of HMME was detected using fluorescence spectral analysis. The viability of the treated cells was determined using the MTT assay, and cell apoptosis was evaluated using flow cytometry. We found that the fluorescence intensity was positively correlated with the incubation time for up to 2 h. The cell growth inhibition rate was significantly high and gradually increased with increasing concentrations of HMME or increasing light intensity, which was calculated as optical output power × irradiation time. Further analysis revealed an e-exponential decay of the cell survival rate to the product of the HMME concentration and the light intensity. We defined the product as parameter B (B = optical output power × irradiation time × HMME concentration). Similarly, the rate of cell apoptosis showed roughly e-exponential growth to parameter B. In conclusion, HMME-mediated PDT can significantly kill HepG2 cells, and the killing effect was related to the cumulative effects of the optical output power, the irradiation time, and the HMME concentration. Therefore, the newly defined parameter B, as a comprehensive physical quantity, may be of great significance for the regulation of light and photosensitizer according to patient-specific conditions in clinical practice.

  10. Necdin, a p53-Target Gene, Is an Inhibitor of p53-Mediated Growth Arrest

    PubMed Central

    Lafontaine, Julie; Rodier, Francis; Ouellet, Véronique; Mes-Masson, Anne-Marie

    2012-01-01

    In vitro, cellular immortalization and transformation define a model for multistep carcinogenesis and current ongoing challenges include the identification of specific molecular events associated with steps along this oncogenic pathway. Here, using NIH3T3 cells, we identified transcriptionally related events associated with the expression of Polyomavirus Large-T antigen (PyLT), a potent viral oncogene. We propose that a subset of these alterations in gene expression may be related to the early events that contribute to carcinogenesis. The proposed tumor suppressor Necdin, known to be regulated by p53, was within a group of genes that was consistently upregulated in the presence of PyLT. While Necdin is induced following p53 activation with different genotoxic stresses, Necdin induction by PyLT did not involve p53 activation or the Rb-binding site of PyLT. Necdin depletion by shRNA conferred a proliferative advantage to NIH3T3 and PyLT-expressing NIH3T3 (NIHLT) cells. In contrast, our results demonstrate that although overexpression of Necdin induced a growth arrest in NIH3T3 and NIHLT cells, a growing population rapidly emerged from these arrested cells. This population no longer showed significant proliferation defects despite high Necdin expression. Moreover, we established that Necdin is a negative regulator of p53-mediated growth arrest induced by nutlin-3, suggesting that Necdin upregulation could contribute to the bypass of a p53-response in p53 wild type tumors. To support this, we characterized Necdin expression in low malignant potential ovarian cancer (LMP) where p53 mutations rarely occur. Elevated levels of Necdin expression were observed in LMP when compared to aggressive serous ovarian cancers. We propose that in some contexts, the constitutive expression of Necdin could contribute to cancer promotion by delaying appropriate p53 responses and potentially promote genomic instability. PMID:22355404

  11. Using hierarchical linear growth models to evaluate protective mechanisms that mediate science achievement

    NASA Astrophysics Data System (ADS)

    von Secker, Clare Elaine

    The study of students at risk is a major topic of science education policy and discussion. Much research has focused on describing conditions and problems associated with the statistical risk of low science achievement among individuals who are members of groups characterized by problems such as poverty and social disadvantage. But outcomes attributed to these factors do not explain the nature and extent of mechanisms that account for differences in performance among individuals at risk. There is ample theoretical and empirical evidence that demographic differences should be conceptualized as social contexts, or collections of variables, that alter the psychological significance and social demands of life events, and affect subsequent relationships between risk and resilience. The hierarchical linear growth models used in this dissertation provide greater specification of the role of social context and the protective effects of attitude, expectations, parenting practices, peer influences, and learning opportunities on science achievement. While the individual influences of these protective factors on science achievement were small, their cumulative effect was substantial. Meta-analysis conducted on the effects associated with psychological and environmental processes that mediate risk mechanisms in sixteen social contexts revealed twenty-two significant differences between groups of students. Positive attitudes, high expectations, and more intense science course-taking had positive effects on achievement of all students, although these factors were not equally protective in all social contexts. In general, effects associated with authoritative parenting and peer influences were negative, regardless of social context. An evaluation comparing the performance and stability of hierarchical linear growth models with traditional repeated measures models is included as well.

  12. Vasopressin regulation of epithelial colonic proliferation and permeability is mediated by pericryptal platelet-derived growth factor A.

    PubMed

    Miró, Lluïsa; Pérez-Bosque, Anna; Maijó, Mònica; Naftalin, Richard J; Moretó, Miquel

    2014-10-01

    Arginine vasopressin (AVP) has trophic effects on the rat distal colon, increasing the growth of pericryptal myofibroblasts and reducing the colonic crypt wall permeability. This study aimed to reproduce in vitro the effects of AVP observed in vivo using cultures of human CCD-18Co myofibroblasts and T84 colonic epithelial cells. Proliferation of myofibroblasts was quantified by bromodeoxyuridine incorporation; the expression of platelet-derived growth factor A (PDGFA), platelet-derived growth factor B, epidermal growth factor, transforming growth factor-β and vascular endothelial growth factor was measured by PCR and the expression of epithelial junction proteins by Western blot. Arginine vasopressin stimulated myofibroblast proliferation and the expression of PDGFA without affecting the expression of platelet-derived growth factor B, epidermal growth factor, transforming growth factor-β or vascular endothelial growth factor. These effects were prevented when AVP receptor inhibitors were present in the medium. Pre-incubation of CCD-18Co cells with anti-PDGF antibody or with an inhibitor of the PDGF receptor abolished the effects of AVP. When colonocytes were incubated with medium obtained from myofibroblasts incubated with AVP, both cell proliferation and the expression of epithelial junction proteins increased; however, direct incubation of colonocytes with AVP did not modify these variables. These results demonstrate that AVP stimulates myofibroblast proliferation and induces PDGFA secretion, implying that PDGFA mediates local myofibroblast proliferation by an autocrine feedback loop and regulates epithelial proliferation and permeability by a paracrine mechanism.

  13. Platelet-derived growth factor CC-mediated neuroprotection against HIV Tat involves TRPC-mediated inactivation of GSK 3beta.

    PubMed

    Peng, Fuwang; Yao, Honghong; Akturk, Halis Kaan; Buch, Shilpa

    2012-01-01

    Platelet-derived growth factor-CC (PDGF-CC) is the third member of the PDGF family, and has been implicated both in embryogenesis and development of the CNS. The biological function of this isoform however, remains largely unexplored in the context of HIV-associated dementia (HAD). In the present study, we demonstrate that exposure of human neuroblastoma cells SH-SY5Y to HIV transactivator protein Tat resulted in decreased intrinsic expression of PDGF-CC as evidenced by RT-PCR and western blot assays. Reciprocally, pretreatment of SH-SY5Y cells with PDGF-CC abrogated Tat-mediated neurotoxicity by mitigating apoptosis and neurite & MAP-2 loss. Using pharmacological and loss of function approaches we identified the role of phosphatidylinositol 3-kinase (PI3K)/Akt signaling in PDGF-CC-mediated neuroprotection. We report herein a novel role about the involvement of transient receptor potential canonical (TRPC) channel 1 in modulation of calcium transients in PDGF-CC-mediated neuroprotection. Furthermore we also demonstrated PDGF-CC-mediated inactivation of the downstream mediator--glycogen synthase kinase 3β (GSK3β) evidenced by its phosphorylation at Ser-9. This was further validated by gain and loss of function studies using cells transfected with either the wild type or mutant GSK3β constructs. Intriguingly, pretreatment of cells with either the PI3K inhibitor or TRPC blocker resulted in failure of PDGF-CC to inactivate GSK3β, thereby suggesting the intersection of PI3K and TRPC signaling at GSK3β. Taken together our findings lead to the suggestion that PDGF-CC could be developed as a therapeutic target to reverse Tat-mediated neurotoxicity with implications for HAD.

  14. Development of tolerogenic dendritic cells and regulatory T cells favors exponential bacterial growth and survival during early respiratory tularemia.

    PubMed

    Periasamy, Sivakumar; Singh, Anju; Sahay, Bikash; Rahman, Tabassum; Feustel, Paul J; Pham, Giang H; Gosselin, Edmund J; Sellati, Timothy J

    2011-09-01

    Tularemia is a vector-borne zoonosis caused by Ft, a Gram-negative, facultative intracellular bacterium. Ft exists in two clinically relevant forms, the European biovar B (holarctica), which produces acute, although mild, self-limiting infections, and the more virulent United States biovar A (tularensis), which is often associated with pneumonic tularemia and more severe disease. In a mouse model of tularemia, respiratory infection with the virulence-attenuated Type B (LVS) or highly virulent Type A (SchuS4) strain engenders peribronchiolar and perivascular inflammation. Paradoxically, despite an intense neutrophilic infiltrate and high bacterial burden, T(h)1-type proinflammatory cytokines (e.g., TNF, IL-1β, IL-6, and IL-12) are absent within the first ∼72 h of pulmonary infection. It has been suggested that the bacterium has the capacity to actively suppress or block NF-κB signaling, thus causing an initial delay in up-regulation of inflammatory mediators. However, our previously published findings and those presented herein contradict this paradigm and instead, strongly support an alternative hypothesis. Rather than blocking NF-κB, Ft actually triggers TLR2-dependent NF-κB signaling, resulting in the development and activation of tDCs and the release of anti-inflammatory cytokines (e.g., IL-10 and TGF-β). In turn, these cytokines stimulate development and proliferation of T(regs) that may restrain T(h)1-type proinflammatory cytokine release early during tularemic infection. The highly regulated and overall anti-inflammatory milieu established in the lung is permissive for unfettered growth and survival of Ft. The capacity of Ft to evoke such a response represents an important immune-evasive strategy.

  15. Development of tolerogenic dendritic cells and regulatory T cells favors exponential bacterial growth and survival during early respiratory tularemia

    PubMed Central

    Periasamy, Sivakumar; Singh, Anju; Sahay, Bikash; Rahman, Tabassum; Feustel, Paul J.; Pham, Giang H.; Gosselin, Edmund J.; Sellati, Timothy J.

    2011-01-01

    Tularemia is a vector-borne zoonosis caused by Ft, a Gram-negative, facultative intracellular bacterium. Ft exists in two clinically relevant forms, the European biovar B (holarctica), which produces acute, although mild, self-limiting infections, and the more virulent United States biovar A (tularensis), which is often associated with pneumonic tularemia and more severe disease. In a mouse model of tularemia, respiratory infection with the virulence-attenuated Type B (LVS) or highly virulent Type A (SchuS4) strain engenders peribronchiolar and perivascular inflammation. Paradoxically, despite an intense neutrophilic infiltrate and high bacterial burden, Th1-type proinflammatory cytokines (e.g., TNF, IL-1β, IL-6, and IL-12) are absent within the first ∼72 h of pulmonary infection. It has been suggested that the bacterium has the capacity to actively suppress or block NF-κB signaling, thus causing an initial delay in up-regulation of inflammatory mediators. However, our previously published findings and those presented herein contradict this paradigm and instead, strongly support an alternative hypothesis. Rather than blocking NF-κB, Ft actually triggers TLR2-dependent NF-κB signaling, resulting in the development and activation of tDCs and the release of anti-inflammatory cytokines (e.g., IL-10 and TGF-β). In turn, these cytokines stimulate development and proliferation of Tregs that may restrain Th1-type proinflammatory cytokine release early during tularemic infection. The highly regulated and overall anti-inflammatory milieu established in the lung is permissive for unfettered growth and survival of Ft. The capacity of Ft to evoke such a response represents an important immune-evasive strategy. PMID:21724804

  16. Effects of Myoga on Memory and Synaptic Plasticity by Regulating Nerve Growth Factor-Mediated Signaling.

    PubMed

    Kim, Hyo Geun; Lim, Soonmin; Hong, Jongki; Kim, Ae-Jung; Oh, Myung Sook

    2016-02-01

    The flower bud of Zingiber mioga Roscoe, known as 'myoga' or Japanese ginger, has a pungent aroma and is commonly consumed as a spice, with pickles, or as a health supplement in Eastern Asia. Here, we evaluated the activity of myoga in the brain, focusing especially on nerve growth factor (NGF), which is believed to mediate synaptic plasticity, supporting learning and memory. In a rat primary hippocampal astrocyte culture system, treatment with myoga extract for 24 h significantly stimulated the production of NGF. In mice administered myoga extract for 14 days, 200 and 400 mg/kg/day treatment resulted in increased NGF levels in the hippocampus. Myoga extract treatment also regulated the phosphorylation of extracellular signal-regulated kinases and cAMP response element-binding protein in the mouse hippocampus, leading to increased synaptic plasticity. In addition, it significantly increased novel object recognition time and spontaneous alternation, indicating improvement in learning and memory. These results suggest that myoga helps regulate NGF and synaptic plasticity, increasing memory ability.

  17. Parathyroid hormone mediates bone growth through the regulation of osteoblast proliferation and differentiation.

    PubMed

    Pettway, Glenda J; Meganck, Jeffrey A; Koh, Amy J; Keller, Evan T; Goldstein, Steven A; McCauley, Laurie K

    2008-04-01

    PTH (1-34) is the only FDA-approved anabolic agent for osteoporosis treatment in the U.S., but its mechanisms are not completely understood. This study investigated PTH effects on osteogenic cells at various stages of differentiation and proliferation using an engineered bone growth model in vivo. Ossicles were generated from bone marrow stromal cells (BMSCs) implanted in immunocompromised mice. Three weeks of PTH (40 microg/kg/day) or vehicle treatment initiated 1 day, 1, 2, or 3 weeks after BMSC implantation resulted in an anabolic response in PTH-treated implants (via histomorphometry and muCT) in all treatment groups. A novel in vivo tracking strategy with luciferase tagged BMSCs and weekly bioluminescent imaging of ossicles revealed increased donor cell proliferation in PTH-treated ossicles. The greatest increase occurred during the first week, and the activity remained elevated in PTH-treated implants over time. Zoledronic acid (ZA) was combined with PTH to delineate interactive mechanisms of these bone active agents. Combining ZA with PTH treatment reduced the PTH-mediated increase in luciferase BMSC activity, serum osteocalcin, and serum tartrate resistant acid phosphotase-5b (TRAP-5b) but ZA did not reduce the PTH-induced increase in total bone. Since zoledronic acid reduced PTH-induced proliferation without reducing bone volume, these data suggest that combining PTH and bisphosphonate therapy warrants further investigation in the treatment of bone disorders.

  18. Class 3 semaphorin mediates dendrite growth in adult newborn neurons through Cdk5/FAK pathway.

    PubMed

    Ng, Teclise; Ryu, Jae Ryun; Sohn, Jae Ho; Tan, Terence; Song, Hongjun; Ming, Guo-Li; Goh, Eyleen L K

    2013-01-01

    Class 3 semaphorins are well-known axonal guidance cues during the embryonic development of mammalian nervous system. However, their activity on postnatally differentiated neurons in neurogenic regions of adult brains has not been characterized. We found that silencing of semaphorin receptors neuropilins (NRP) 1 or 2 in neural progenitors at the adult mouse dentate gyrus resulted in newly differentiated neurons with shorter dendrites and simpler branching in vivo. Tyrosine phosphorylation (Tyr 397) and serine phosphorylation (Ser 732) of FAK were essential for these effects. Semaphorin 3A and 3F mediate serine phosphorylation of FAK through the activation of Cdk5. Silencing of either Cdk5 or FAK in newborn neurons phenocopied the defects in dendritic development seen upon silencing of NRP1 or NRP2. Furthermore, in vivo overexpression of Cdk5 or FAK rescued the dendritic phenotypes seen in NRP1 and NRP2 deficient neurons. These results point to a novel role for class 3 semaphorins in promoting dendritic growth and branching during adult hippocampal neurogenesis through the activation of Cdk5-FAK signaling pathway.

  19. Adipocyte JAK2 mediates growth hormone–induced hepatic insulin resistance

    PubMed Central

    Corbit, Kevin C.; Camporez, João Paulo G.; Tran, Jennifer L.; Wilson, Camella G.; Lowe, Dylan A.; Nordstrom, Sarah M.; Ganeshan, Kirthana; Perry, Rachel J.; Weiss, Ethan J.

    2017-01-01

    For nearly 100 years, growth hormone (GH) has been known to affect insulin sensitivity and risk of diabetes. However, the tissue governing the effects of GH signaling on insulin and glucose homeostasis remains unknown. Excess GH reduces fat mass and insulin sensitivity. Conversely, GH insensitivity (GHI) is associated with increased adiposity, augmented insulin sensitivity, and protection from diabetes. Here, we induce adipocyte-specific GHI through conditional deletion of Jak2 (JAK2A), an obligate transducer of GH signaling. Similar to whole-body GHI, JAK2A mice had increased adiposity and extreme insulin sensitivity. Loss of adipocyte Jak2 augmented hepatic insulin sensitivity and conferred resistance to diet-induced metabolic stress without overt changes in circulating fatty acids. While GH injections induced hepatic insulin resistance in control mice, the diabetogenic action was absent in JAK2A mice. Adipocyte GH signaling directly impinged on both adipose and hepatic insulin signal transduction. Collectively, our results show that adipose tissue governs the effects of GH on insulin and glucose homeostasis. Further, we show that JAK2 mediates liver insulin sensitivity via an extrahepatic, adipose tissue–dependent mechanism. PMID:28194444

  20. Bladder cancer cell growth and motility implicate cannabinoid 2 receptor-mediated modifications of sphingolipids metabolism

    PubMed Central

    Bettiga, Arianna; Aureli, Massimo; Colciago, Giorgia; Murdica, Valentina; Moschini, Marco; Lucianò, Roberta; Canals, Daniel; Hannun, Yusuf; Hedlund, Petter; Lavorgna, Giovanni; Colombo, Renzo; Bassi, Rosaria; Samarani, Maura; Montorsi, Francesco; Salonia, Andrea; Benigni, Fabio

    2017-01-01

    The inhibitory effects demonstrated by activation of cannabinoid receptors (CB) on cancer proliferation and migration may also play critical roles in controlling bladder cancer (BC). CB expression on human normal and BC specimens was tested by immunohistochemistry. Human BC cells RT4 and RT112 were challenged with CB agonists and assessed for proliferation, apoptosis, and motility. Cellular sphingolipids (SL) constitution and metabolism were evaluated after metabolic labelling. CB1-2 were detected in BC specimens, but only CB2 was more expressed in the tumour. Both cell lines expressed similar CB2. Exposure to CB2 agonists inhibited BC growth, down-modulated Akt, induced caspase 3-activation and modified SL metabolism. Baseline SL analysis in cell lines showed differences linked to unique migratory behaviours and cytoskeletal re-arrangements. CB2 activation changed the SL composition of more aggressive RT112 cells by reducing (p < 0.01) Gb3 ganglioside (−50 ± 3%) and sphingosine 1-phosphate (S1P, −40 ± 4%), which ended up to reduction in cell motility (−46 ± 5%) with inhibition of p-SRC. CB2-selective antagonists, gene silencing and an inhibitor of SL biosynthesis partially prevented CB2 agonist-induced effects on cell viability and motility. CB2 activation led to ceramide-mediated BC cell apoptosis independently of SL constitutive composition, which instead was modulated by CB2 agonists to reduce cell motility. PMID:28191815

  1. Placental growth factor mediates aldosterone-dependent vascular injury in mice.

    PubMed

    Jaffe, Iris Z; Newfell, Brenna G; Aronovitz, Mark; Mohammad, Najwa N; McGraw, Adam P; Perreault, Roger E; Carmeliet, Peter; Ehsan, Afshin; Mendelsohn, Michael E

    2010-11-01

    In clinical trials, aldosterone antagonists reduce cardiovascular ischemia and mortality by unknown mechanisms. Aldosterone is a steroid hormone that signals through renal mineralocorticoid receptors (MRs) to regulate blood pressure. MRs are expressed and regulate gene transcription in human vascular cells, suggesting that aldosterone might have direct vascular effects. Using gene expression profiling, we identify the pro-proliferative VEGF family member placental growth factor (PGF) as an aldosterone-regulated vascular MR target gene in mice and humans. Aldosterone-activated vascular MR stimulated Pgf gene transcription and increased PGF protein expression and secretion in the mouse vasculature. In mouse vessels with endothelial damage and human vessels from patients with atherosclerosis, aldosterone enhanced expression of PGF and its receptor, FMS-like tyrosine kinase 1 (Flt1). In atherosclerotic human vessels, MR antagonists inhibited PGF expression. In vivo, aldosterone infusion augmented vascular remodeling in mouse carotids following wire injury, an effect that was lost in Pgf-/- mice. In summary, we have identified PGF as what we believe to be a novel downstream target of vascular MR that mediates aldosterone augmentation of vascular injury. These findings suggest a non-renal mechanism for the vascular protective effects of aldosterone antagonists in humans and support targeting the vascular aldosterone/MR/PGF/Flt1 pathway as a therapeutic strategy for ischemic cardiovascular disease.

  2. Sulfatase modifying factor 1-mediated fibroblast growth factor signaling primes hematopoietic multilineage development.

    PubMed

    Buono, Mario; Visigalli, Ilaria; Bergamasco, Roberta; Biffi, Alessandra; Cosma, Maria Pia

    2010-08-02

    Self-renewal and differentiation of hematopoietic stem cells (HSCs) are balanced by the concerted activities of the fibroblast growth factor (FGF), Wnt, and Notch pathways, which are tuned by enzyme-mediated remodeling of heparan sulfate proteoglycans (HSPGs). Sulfatase modifying factor 1 (SUMF1) activates the Sulf1 and Sulf2 sulfatases that remodel the HSPGs, and is mutated in patients with multiple sulfatase deficiency. Here, we show that the FGF signaling pathway is constitutively activated in Sumf1(-/-) HSCs and hematopoietic stem progenitor cells (HSPCs). These cells show increased p-extracellular signal-regulated kinase levels, which in turn promote beta-catenin accumulation. Constitutive activation of FGF signaling results in a block in erythroid differentiation at the chromatophilic erythroblast stage, and of B lymphocyte differentiation at the pro-B cell stage. A reduction in mature myeloid cells and an aberrant development of T lymphocytes are also seen. These defects are rescued in vivo by blocking the FGF pathway in Sumf1(-/-) mice. Transplantation of Sumf1(-/-) HSPCs into wild-type mice reconstituted the phenotype of the donors, suggesting a cell autonomous defect. These data indicate that Sumf1 controls HSPC differentiation and hematopoietic lineage development through FGF and Wnt signaling.

  3. Understanding the Seed-Mediated Growth of Gold Nanorods through a Fractional Factorial Design of Experiments.

    PubMed

    Burrows, Nathan D; Harvey, Samantha; Idesis, Fred A; Murphy, Catherine J

    2017-02-28

    Since the development of simple, aqueous protocols for the synthesis of anisotropic metal nanoparticles, research into many promising, valuable applications of gold nanorods has grown considerably, but a number of challenges remain, including gold-particle yield, robustness to minor impurities, and precise control of gold nanorod surface chemistry. Herein we present the results of a composite fractional factorial series of experiments designed to screen seven additional potential avenues of control and to understand the seed-mediated silver-assisted synthesis of gold nanorods. These synthesis variables are the amount of sodium borohydride used and the rate of stirring when producing seed nanoparticles, the age of and the amount of seeds added, the reaction temperature, the amounts of silver nitrate and ascorbic acid added, and the age of the reduced growth solution before seed nanoparticles are added to initiate rod formation. This statistical experimental design and analysis method, besides determining which experimental variables are important and which are not when synthesizing gold nanorods (and quantifying their effects), gives further insight into the mechanism of growth by measuring the degree to which variables interact with each other by mapping out their mechanistic connections. This work demonstrates that when forming gold nanorods by the reduction of auric ions by ascorbic acid onto seed nanoparticles, ascorbic acid determines how much gold is reduced, and the amount of seeds determine how it is divided, yet both influence the intrinsic growth rates, in both width and length, of the forming nanorods. Furthermore, this work shows that the reduction of gold proceeds via direct reduction on the surface of seeds and not through a disproportionation reaction. Further control over the length of gold nanorods can be achieved by tuning the amount of silver nitrate or the reaction temperature. This work shows that silver does not directly influence rod length or

  4. Inhibition of NF-κB by a PXR-dependent pathway mediates counter-regulatory activities of rifaximin on innate immunity in intestinal epithelial cells.

    PubMed

    Mencarelli, Andrea; Renga, Barbara; Palladino, Giuseppe; Claudio, D'Amore; Ricci, Patrizia; Distrutti, Eleonora; Barbanti, Miriam; Baldelli, Franco; Fiorucci, Stefano

    2011-10-01

    A dysregulated interaction between intestinal epithelial cells (IEC) and components of innate immunity is a hallmark of inflammatory bowel diseases. Rifaximin is a poorly absorbed oral antimicrobial agent increasingly used in the treatment of inflammatory bowel diseases that has been demonstrated to act as a gut-specific ligand for the human nuclear receptor pregnane-X receptor (PXR). In the present study we investigated, whether activation of PXR in IEC by rifaximin, emanates counter-regulatory signals and modulates the expression of cytokines or chemokines mechanistically involved in dysregulated intestinal immune homeostasis documented in inflammatory bowel diseases. Our results demonstrate that primary IEC express PXR that regulate the pattern of cytokines and chemokines expressed. PXR silencing decreases TGF-β and IP-10 while increases the expression of TNF-α, IL-8, Rantes and increase the production of PGE2. This pattern is further exacerbated by treating anti-PXR siRNA cells with bacterial endotoxin (LPS). Exposure to rifaximin caused a robust attenuation of generation of inflammatory mediators caused by LPS and increased the generation of TGF-β. PXR silencing completely abrogated these anti-inflammatory effects of rifaximin. By Western blot analysis we found that rifaximin abrogates the binding of NF-κB caused by LPS. Finally, exposure of human colon biopsies from inflammatory bowel diseases patients to rifaximin reduced mRNA levels of IL-8, Rantes, MIP-3α and TNFα induced by LPS. Collectively, these data establish that rifaximin exerts counter-regulatory activities at the interface between enteric bacteria and intestinal epithelial cells. The ability of rifaximin to activate PXR contributes to the maintenance of the intestinal immune homeostasis.

  5. The identification and characterization of specific ARF-Aux/IAA regulatory modules in plant growth and development.

    PubMed

    Krogan, Naden T; Berleth, Thomas

    2015-01-01

    The current model of auxin-inducible transcription describes numerous regulatory interactions between AUXIN RESPONSE FACTORs (ARFs) and Aux/IAAs. However, specific relationships between individual members of these families in planta remain largely uncharacterized. Using a systems biology approach, the entire suite of Aux/IAA genes directly regulated by the developmentally pivotal ARF MONOPTEROS (MP) was recently determined for multiple Arabidopsis tissue types. This study showed that MP directly targets distinct subclades of Aux/IAAs, revealing potential regulatory modules of redundantly acting Aux/IAAs involved in MP-dependent processes. Further, functional analyses indicated that the protein products of these targeted Aux/IAAs negatively feedback on MP. Thus, comprehensive identification of Aux/IAAs targeted by individual ARFs will generate biologically meaningful networks of ARF-Aux/IAA regulatory modules controlling distinct plant pathways.

  6. The molecular mechanism of mediation of adsorbed serum proteins to endothelial cells adhesion and growth on biomaterials.

    PubMed

    Yang, Dayun; Lü, Xiaoying; Hong, Ying; Xi, Tingfei; Zhang, Deyuan

    2013-07-01

    To explore molecular mechanism of mediation of adsorbed proteins to cell adhesion and growth on biomaterials, this study examined endothelial cell adhesion, morphology and viability on bare and titanium nitride (TiN) coated nickel titanium (NiTi) alloys and chitosan film firstly, and then identified the type and amount of serum proteins adsorbed on the three surfaces by proteomic technology. Subsequently, the mediation role of the identified proteins to cell adhesion and growth was investigated with bioinformatics analyses, and further confirmed by a series of cellular and molecular biological experiments. Results showed that the type and amount of adsorbed serum proteins associated with cell adhesion and growth was obviously higher on the alloys than on the chitosan film, and these proteins mediated endothelial cell adhesion and growth on the alloys via four ways. First, proteins such as adiponectin in the adsorbed protein layer bound with cell surface receptors to generate signal transduction, which activated cell surface integrins through increasing intracellular calcium level. Another way, thrombospondin 1 in the adsorbed protein layer promoted TGF-β signaling pathway activation and enhanced integrins expression. The third, RGD sequence containing proteins such as fibronectin 1, vitronectin and thrombospondin 1 in the adsorbed protein layer bound with activated integrins to activate focal adhesion pathway, increased focal adhesion formation and actin cytoskeleton organization and mediated cell adhesion and spreading. In addition, the activated focal adhesion pathway promoted the expression of cell growth related genes and resulted in cell proliferation. The fourth route, coagulation factor II (F2) and fibronectin 1 in the adsorbed protein layer bound with cell surface F2 receptor and integrin, activated regulation of actin cytoskeleton pathway and regulated actin cytoskeleton organization.

  7. Volatile-Mediated Effects Predominate in Paraburkholderia phytofirmans Growth Promotion and Salt Stress Tolerance of Arabidopsis thaliana.

    PubMed

    Ledger, Thomas; Rojas, Sandy; Timmermann, Tania; Pinedo, Ignacio; Poupin, María J; Garrido, Tatiana; Richter, Pablo; Tamayo, Javier; Donoso, Raúl

    2016-01-01

    Abiotic stress has a growing impact on plant growth and agricultural activity worldwide. Specific plant growth promoting rhizobacteria have been reported to stimulate growth and tolerance to abiotic stress in plants, and molecular mechanisms like phytohormone synthesis and 1-aminocyclopropane-1-carboxylate deamination are usual candidates proposed to mediate these bacterial effects. Paraburkholderia phytofirmans PsJN is able to promote growth of several plant hosts, and improve their tolerance to chilling, drought and salinity. This work investigated bacterial determinants involved in PsJN stimulation of growth and salinity tolerance in Arabidopsis thaliana, showing bacteria enable plants to survive long-term salinity treatment, accumulating less sodium within leaf tissues relative to non-inoculated controls. Inactivation of specific bacterial genes encoding ACC deaminase, auxin catabolism, N-acyl-homoserine-lactone production, and flagellin synthesis showed these functions have little influence on bacterial induction of salinity tolerance. Volatile organic compound emission from strain PsJN was shown to reproduce the effects of direct bacterial inoculation of roots, increasing plant growth rate and tolerance to salinity evaluated both in vitro and in soil. Furthermore, early exposure to VOCs from P. phytofirmans was sufficient to stimulate long-term effects observed in Arabidopsis growth in the presence and absence of salinity. Organic compounds were analyzed in the headspace of PsJN cultures, showing production of 2-undecanone, 7-hexanol, 3-methylbutanol and dimethyl disulfide. Exposure of A. thaliana to different quantities of these molecules showed that they are able to influence growth in a wide range of added amounts. Exposure to a blend of the first three compounds was found to mimic the effects of PsJN on both general growth promotion and salinity tolerance. To our knowledge, this is the first report on volatile compound-mediated induction of plant abiotic

  8. Posttraumatic growth in patients who survived cardiac surgery: the predictive and mediating roles of faith-based factors.

    PubMed

    Ai, Amy L; Hall, Daniel; Pargament, Kenneth; Tice, Terrence N

    2013-04-01

    Despite the growing knowledge of posttraumatic growth, only a few studies have examined personal growth in the context of cardiac health. Similarly, longitudinal research is lacking on the implications of religion/spirituality for patients with advanced cardiac diseases. This paper aims to explore the effect of preoperative religious coping on long-term postoperative personal growth and potential mediation in this effect. Analyses capitalized on a preoperative survey and medical indices from the Society of Thoracic Surgeons' National Database of patients undergoing cardiac surgery. Participants in the current follow-up study completed a mailed survey 30 months after surgery. Hierarchical regression analysis was performed to evaluate the extent to which preoperative use of religious coping predicted growth at follow-up, after controlling for key demographics, medical indices, mental health, and protective factors. Predictors of posttraumatic growth at follow-up were positive religious coping and a living status without a partner. Medical indices, optimistic expectations, social support, and other religious factors were unrelated to posttraumatic growth. Including religious factors diminished effects of gender, age, and race. Including perceived spiritual support completely eliminated the role of positive religious coping, indicating mediation. Preoperative positive religious coping may have a long-term effect on postoperative personal growth, explainable by higher spiritual connections as a part of significance-making. These results suggest that spirituality may play a favorable role in cardiac patients' posttraumatic growth after surviving a life-altering operation. The elimination of demographic effects may help explain previously mixed findings concerning the association between these factors and personal growth.

  9. Volatile-Mediated Effects Predominate in Paraburkholderia phytofirmans Growth Promotion and Salt Stress Tolerance of Arabidopsis thaliana

    PubMed Central

    Ledger, Thomas; Rojas, Sandy; Timmermann, Tania; Pinedo, Ignacio; Poupin, María J.; Garrido, Tatiana; Richter, Pablo; Tamayo, Javier

    2016-01-01

    Abiotic stress has a growing impact on plant growth and agricultural activity worldwide. Specific plant growth promoting rhizobacteria have been reported to stimulate growth and tolerance to abiotic stress in plants, and molecular mechanisms like phytohormone synthesis and 1-aminocyclopropane-1-carboxylate deamination are usual candidates proposed to mediate these bacterial effects. Paraburkholderia phytofirmans PsJN is able to promote growth of several plant hosts, and improve their tolerance to chilling, drought and salinity. This work investigated bacterial determinants involved in PsJN stimulation of growth and salinity tolerance in Arabidopsis thaliana, showing bacteria enable plants to survive long-term salinity treatment, accumulating less sodium within leaf tissues relative to non-inoculated controls. Inactivation of specific bacterial genes encoding ACC deaminase, auxin catabolism, N-acyl-homoserine-lactone production, and flagellin synthesis showed these functions have little influence on bacterial induction of salinity tolerance. Volatile organic compound emission from strain PsJN was shown to reproduce the effects of direct bacterial inoculation of roots, increasing plant growth rate and tolerance to salinity evaluated both in vitro and in soil. Furthermore, early exposure to VOCs from P. phytofirmans was sufficient to stimulate long-term effects observed in Arabidopsis growth in the presence and absence of salinity. Organic compounds were analyzed in the headspace of PsJN cultures, showing production of 2-undecanone, 7-hexanol, 3-methylbutanol and dimethyl disulfide. Exposure of A. thaliana to different quantities of these molecules showed that they are able to influence growth in a wide range of added amounts. Exposure to a blend of the first three compounds was found to mimic the effects of PsJN on both general growth promotion and salinity tolerance. To our knowledge, this is the first report on volatile compound-mediated induction of plant abiotic

  10. Regulatory effects of introduction of an exogenous FGF2 gene on other growth factor genes in a healing tendon.

    PubMed

    Tang, Jin Bo; Chen, Chuan Hao; Zhou, You Lang; McKeever, Clarie; Liu, Paul Y

    2014-01-01

    In this study of a tendon injury model, we investigated how injection of a vector incorporating one growth factor gene changes expression levels of multiple growth factor genes in the healing process. The flexor tendon of chicken toes was completely cut and repaired surgically. The tendons in the experimental arm were injected with an adeno-associated virus-2 vector incorporating basic fibroblast growth-factor gene, whereas the tendons in the control arm were not injected or injected with sham vectors. Using real-time polymerase chain reaction, we found that, within the tendon healing period, a set of growth factor genes-transforming growth factor-β1, vascular endothelial growth factor, and connective tissue growth factor-were significantly up-regulated. Expression of the platelet-derived growth factor-B gene was not changed, and the insulin-like growth factor was down-regulated. A tendon marker gene, scleraxis, was significantly up-regulated in the period. Our study revealed an intriguing finding that introduction of one growth factor gene in the healing tendon modulated expression of multiple growth factor genes. We believe this study may have significant implications in determining the approach of gene therapy, and the findings substantiate that gene therapy using a single growth factor could affect multiple growth factors.

  11. The isolation and characterization of growth regulatory factors produced by a herpes simplex virus Type 2 transformed mouse tumor cell line, H238

    SciTech Connect

    Stagg, R.B.

    1988-01-01

    This study was performed in an attempt to associate HSV-2-transformation with specific growth factors in order to develop a testable model for HSV-2-transformation. We report here the isolation and characterization of four growth regulatory factors produced by H238, an HSV-2-transformed mouse tumor cell line. These factors were separated from the H238-CM by heparin-sepharose affinity chromatography into three peaks of mitogenic activity and a fourth containing inhibitory activity for splenocytes. The three peaks of mitogenic activity have been identified based on physiochemical characteristics: the first supported the anchorage-independent growth of EGF treated NRK-c-49 cells and resembles transforming growth factor-{beta} (TGF-{beta}); the second bound to lectin-coated sepharose beads and was sensitive to trypsin, neuroaminidase, and the reducing agent dithiothreitol (DTT) and, resembled a platelet-derived growth factor (PDGF)-like factor; and the third displaced ({sup 125}I)-labeled basic fibroblast growth factor (bFGF) in a dose-dependent fashion when tested with a radioimmune assay. The fourth peak was inhibitory for a variety of splenocyte function assays. A model for the interaction of these factors in vivo is presented with an emphasis on testability.

  12. Attachment Styles and Personal Growth following Romantic Breakups: The Mediating Roles of Distress, Rumination, and Tendency to Rebound

    PubMed Central

    Marshall, Tara C.; Bejanyan, Kathrine; Ferenczi, Nelli

    2013-01-01

    The purpose of this research was to examine the associations of attachment anxiety and avoidance with personal growth following relationship dissolution, and to test breakup distress, rumination, and tendency to rebound with new partners as mediators of these associations. Study 1 (N = 411) and Study 2 (N = 465) measured attachment style, breakup distress, and personal growth; Study 2 additionally measured ruminative reflection, brooding, and proclivity to rebound with new partners. Structural equation modelling revealed in both studies that anxiety was indirectly associated with greater personal growth through heightened breakup distress, whereas avoidance was indirectly associated with lower personal growth through inhibited breakup distress. Study 2 further showed that the positive association of breakup distress with personal growth was accounted for by enhanced reflection and brooding, and that anxious individuals’ greater personal growth was also explained by their proclivity to rebound. These findings suggest that anxious individuals’ hyperactivated breakup distress may act as a catalyst for personal growth by promoting the cognitive processing of breakup-related thoughts and emotions, whereas avoidant individuals’ deactivated distress may inhibit personal growth by suppressing this cognitive work. PMID:24066169

  13. Extracellular calmodulin regulates growth and cAMP-mediated chemotaxis in Dictyostelium discoideum

    SciTech Connect

    O'Day, Danton H.; Huber, Robert J.; Suarez, Andres

    2012-09-07

    Highlights: Black-Right-Pointing-Pointer Extracellular calmodulin is present throughout growth and development in Dictyostelium. Black-Right-Pointing-Pointer Extracellular calmodulin localizes within the ECM during development. Black-Right-Pointing-Pointer Extracellular calmodulin inhibits cell proliferation and increases chemotaxis. Black-Right-Pointing-Pointer Extracellular calmodulin exists in eukaryotic microbes. Black-Right-Pointing-Pointer Extracellular calmodulin may be functionally as important as intracellular calmodulin. -- Abstract: The existence of extracellular calmodulin (CaM) has had a long and controversial history. CaM is a ubiquitous calcium-binding protein that has been found in every eukaryotic cell system. Calcium-free apo-CaM and Ca{sup 2+}/CaM exert their effects by binding to and regulating the activity of CaM-binding proteins (CaMBPs). Most of the research done to date on CaM and its CaMBPs has focused on their intracellular functions. The presence of extracellular CaM is well established in a number of plants where it functions in proliferation, cell wall regeneration, gene regulation and germination. While CaM has been detected extracellularly in several animal species, including frog, rat, rabbit and human, its extracellular localization and functions are less well established. In contrast the study of extracellular CaM in eukaryotic microbes remains to be done. Here we show that CaM is constitutively expressed and secreted throughout asexual development in Dictyostelium where the presence of extracellular CaM dose-dependently inhibits cell proliferation but increases cAMP mediated chemotaxis. During development, extracellular CaM localizes within the slime sheath where it coexists with at least one CaMBP, the matricellular CaM-binding protein CyrA. Coupled with previous research, this work provides direct evidence for the existence of extracellular CaM in the Dictyostelium and provides insight into its functions in this model amoebozoan.

  14. Methylglyoxal, a glycolysis side-product, induces Hsp90 glycation and YAP-mediated tumor growth and metastasis.

    PubMed

    Nokin, Marie-Julie; Durieux, Florence; Peixoto, Paul; Chiavarina, Barbara; Peulen, Olivier; Blomme, Arnaud; Turtoi, Andrei; Costanza, Brunella; Smargiasso, Nicolas; Baiwir, Dominique; Scheijen, Jean L; Schalkwijk, Casper G; Leenders, Justine; De Tullio, Pascal; Bianchi, Elettra; Thiry, Marc; Uchida, Koji; Spiegel, David A; Cochrane, James R; Hutton, Craig A; De Pauw, Edwin; Delvenne, Philippe; Belpomme, Dominique; Castronovo, Vincent; Bellahcène, Akeila

    2016-10-19

    Metabolic reprogramming toward aerobic glycolysis unavoidably induces methylglyoxal (MG) formation in cancer cells. MG mediates the glycation of proteins to form advanced glycation end products (AGEs). We have recently demonstrated that MG-induced AGEs are a common feature of breast cancer. Little is known regarding the impact of MG-mediated carbonyl stress on tumor progression. Breast tumors with MG stress presented with high nuclear YAP, a key transcriptional co-activator regulating tumor growth and invasion. Elevated MG levels resulted in sustained YAP nuclear localization/activity that could be reverted using Carnosine, a scavenger for MG. MG treatment affected Hsp90 chaperone activity and decreased its binding to LATS1, a key kinase of the Hippo pathway. Cancer cells with high MG stress showed enhanced growth and metastatic potential in vivo. These findings reinforce the cumulative evidence pointing to hyperglycemia as a risk factor for cancer incidence and bring renewed interest in MG scavengers for cancer treatment.

  15. TIR1/AFB-Aux/IAA auxin perception mediates rapid cell wall acidification and growth of Arabidopsis hypocotyls

    PubMed Central

    Fendrych, Matyáš; Leung, Jeffrey; Friml, Jiří

    2016-01-01

    Despite being composed of immobile cells, plants reorient along directional stimuli. The hormone auxin is redistributed in stimulated organs leading to differential growth and bending. Auxin application triggers rapid cell wall acidification and elongation of aerial organs of plants, but the molecular players mediating these effects are still controversial. Here we use genetically-encoded pH and auxin signaling sensors, pharmacological and genetic manipulations available for Arabidopsis etiolated hypocotyls to clarify how auxin is perceived and the downstream growth executed. We show that auxin-induced acidification occurs by local activation of H+-ATPases, which in the context of gravity response is restricted to the lower organ side. This auxin-stimulated acidification and growth require TIR1/AFB-Aux/IAA nuclear auxin perception. In addition, auxin-induced gene transcription and specifically SAUR proteins are crucial downstream mediators of this growth. Our study provides strong experimental support for the acid growth theory and clarified the contribution of the upstream auxin perception mechanisms. DOI: http://dx.doi.org/10.7554/eLife.19048.001 PMID:27627746

  16. TIR1/AFB-Aux/IAA auxin perception mediates rapid cell wall acidification and growth of Arabidopsis hypocotyls.

    PubMed

    Fendrych, Matyáš; Leung, Jeffrey; Friml, Jiří

    2016-09-14

    Despite being composed of immobile cells, plants reorient along directional stimuli. The hormone auxin is redistributed in stimulated organs leading to differential growth and bending. Auxin application triggers rapid cell wall acidification and elongation of aerial organs of plants, but the molecular players mediating these effects are still controversial. Here we use genetically-encoded pH and auxin signaling sensors, pharmacological and genetic manipulations available for Arabidopsis etiolated hypocotyls to clarify how auxin is perceived and the downstream growth executed. We show that auxin-induced acidification occurs by local activation of H(+)-ATPases, which in the context of gravity response is restricted to the lower organ side. This auxin-stimulated acidification and growth require TIR1/AFB-Aux/IAA nuclear auxin perception. In addition, auxin-induced gene transcription and specifically SAUR proteins are crucial downstream mediators of this growth. Our study provides strong experimental support for the acid growth theory and clarified the contribution of the upstream auxin perception mechanisms.

  17. Inhibition of miR301 enhances Akt-mediated cell proliferation by accumulation of PTEN in nucleus and its effects on cell-cycle regulatory proteins

    PubMed Central

    Jain, Mayur V.; Shareef, Ahmad; Likus, Wirginia; Cieślar-Pobuda, Artur; Ghavami, Saeid; Łos, Marek J.

    2016-01-01

    Micro-RNAs (miRs) represent an innovative class of genes that act as regulators of gene expression. Recently, the aberrant expression of several miRs has been associated with different types of cancers. In this study, we show that miR301 inhibition influences PI3K-Akt pathway activity. Akt overexpression in MCF7 and MDAMB468 cells caused downregulation of miR301 expression. This effect was confirmed by co-transfection of miR301-modulators in the presence of Akt. Cells overexpressing miR301-inhibitor and Akt, exhibited increased migration and proliferation. Experimental results also confirmed PI3K, PTEN and FoxF2 as regulatory targets for miR301. Furthermore, Akt expression in conjunction with miR301-inhibitor increased nuclear accumulation of PTEN, thus preventing it from downregulating the PI3K-signalling. In summary, our data emphasize the importance of miR301 inhibition on PI3K-Akt pathway-mediated cellular functions. Hence, it opens new avenues for the development of new anti-cancer agents preferentially targeting PI3K-Akt pathway. PMID:26967567

  18. PR55α, a Regulatory Subunit of PP2A, Specifically Regulates PP2A-mediated β-Catenin Dephosphorylation

    PubMed Central

    Zhang, Wen; Yang, Jun; Liu, Yajuan; Chen, Xi; Yu, Tianxin; Jia, Jianhang; Liu, Chunming

    2009-01-01

    A central question in Wnt signaling is the regulation of β-catenin phosphorylation and degradation. Multiple kinases, including CKIα and GSK3, are involved in β-catenin phosphorylation. Protein phosphatases such as PP2A and PP1 have been implicated in the regulation of β-catenin. However, which phosphatase dephosphorylates β-catenin in vivo and how the specificity of β-catenin dephosphorylation is regulated are not clear. In this study, we show that PP2A regulates β-catenin phosphorylation and degradation in vivo. We demonstrate that PP2A is required for Wnt/β-catenin signaling in Drosophila. Moreover, we have identified PR55α as the regulatory subunit of PP2A that controls β-catenin phosphorylation and degradation. PR55α, but not the catalytic subunit, PP2Ac, directly interacts with β-catenin. RNA interference knockdown of PR55α elevates β-catenin phosphorylation and decreases Wnt signaling, whereas overexpressing PR55α enhances Wnt signaling. Taken together, our results suggest that PR55α specifically regulates PP2A-mediated β-catenin dephosphorylation and plays an essential role in Wnt signaling. PMID:19556239

  19. Combined QSAR-based virtual screening and fluorescence binding assay to identify natural product mediators of Interferon Regulatory Factor 7 (IRF-7) in pulmonary infection.

    PubMed

    Liu, Y; Huang, L; Ye, H; Lv, X

    2016-11-01

    Interferon regulatory factor-7 (IRF-7) is involved in pulmonary infection and pneumonia. Here, a synthetic strategy that combined quantitative structure-activity relationship (QSAR)-based virtual screening and in vitro binding assay was described to identify new and potent mediator ligands of IRF-7 from natural products. In the procedure, a QSAR scoring function was developed and validated using Gaussian process (GP) regression and a structure-based set of protein-ligand affinity data. By integrating hotspot pocket prediction, pharmacokinetics profile analysis and molecular docking calculations, the scoring function was successfully applied to virtual screening against a large library of structurally diverse, drug-like natural products. With the method we were able to identify a number of potential hits, from which several compounds were found to have moderate or high affinity to IRF-7 using fluorescence binding assays, with dissociation constants Kd at micromolar level. We have also examined the structural basis and noncovalent interactions of computationally modelled IRF-7 complex with its potent ligands. It is revealed that hydrophobic forces and van der Waals contacts play a central role in stabilization of the complex architecture, while few hydrogen bonds confer additional specificity for the protein-ligand recognition.

  20. Vancomycin susceptibility in methicillin-resistant Staphylococcus aureus is mediated by YycHI activation of the WalRK essential two-component regulatory system

    PubMed Central

    Cameron, David R.; Jiang, Jhih-Hang; Kostoulias, Xenia; Foxwell, Daniel J.; Peleg, Anton Y.

    2016-01-01

    The treatment of infections caused by methicillin-resistant Staphylococcus aureus is complicated by the emergence of strains with intermediate-level resistance to vancomycin (termed VISA). We have characterised a molecular pathway involved in the in vivo evolution of VISA mediated by the regulatory proteins YycH and YycI. In contrast to their function in other bacterial species, we report a positive role for these auxiliary proteins in regulation of the two-component regulator WalRK. Transcriptional profiling of yycH and yycI deletion mutants revealed downregulation of the ‘WalRK regulon’ including cell wall hydrolase genes atlA and sle1, with functional autolysis assays supporting these data by showing an impaired autolytic phenotype for each deletion strain. Using bacterial-two hybrid assays, we showed that YycH and YycI interact, and that YycHI also interacts with the sensor kinase WalK, forming a ternary protein complex. Mutation to YycH or YycI associated with clinical VISA strains had a deleterious impact on the YycHI/WalK complex, suggesting that the interaction is important for the regulation of WalRK. Taken together, we have described a novel antibiotic resistance strategy for the human pathogen S. aureus, whereby YycHI mutations are selected for in vivo leading to reduced WalRK activation, impaired cell wall turnover and ultimately reduced vancomycin efficacy. PMID:27600558

  1. TOP1 and 2, polysaccharides from Taraxacum officinale, attenuate CCl(4)-induced hepatic damage through the modulation of NF-kappaB and its regulatory mediators.

    PubMed

    Park, Chung Mu; Youn, Hyun Joo; Chang, Hee Kyung; Song, Young Sun

    2010-05-01

    In this work, we estimate the inhibitory effect of two polysaccharides from Taraxacum officinale (TOP) on CCl(4)-induced oxidative stress and inflammation in Sprague-Dawley rats. TOP1 and 2 (304, 92 mg/kg bw) were administered for 7 days via a stomach sonde, and hepatitis was induced by a single dose of CCl(4) (50% CCl(4)/olive oil; 0.5 mL/kg bw) administration. CCl(4) significantly elevated serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) activities. Histopathological observation further revealed that CCl(4)-induced moderate levels of inflammatory cell infiltration, centrilobular fatty change, apoptosis, and necrosis. However, TOPs pretreatment markedly decreased AST and ALT activities as well as hepatic lesions. TOPs also increased free radical scavenging activity, as exhibited by a lowered TBARS concentration. TOPs pretreatment also reversed other hepatitis-associated symptoms, including GSH depletion, inhibited anti-oxidative enzyme activities, up-regulation of NF-kappaB and increased expression of its regulatory inflammatory mediators, such as inducible nitric oxide synthase (iNOS), cyclooxygenase (COX)-2, tumor necrosis factor (TNF)-alpha, and interleukin (IL)-1beta. These results suggest that TOPs have a hepatoprotective effect by modulating inflammatory responses and ameliorating oxidative stress.

  2. Immune stimulatory receptor CD40 is required for T-cell suppression and T regulatory cell activation mediated by myeloid-derived suppressor cells in cancer.

    PubMed

    Pan, Ping-Ying; Ma, Ge; Weber, Kaare J; Ozao-Choy, Junko; Wang, George; Yin, Bingjiao; Divino, Celia M; Chen, Shu-Hsia

    2010-01-01

    Immune tolerance to tumors is often associated with accumulation of myeloid-derived suppressor cells (MDSC) and an increase in the number of T-regulatory cells (Treg). In tumor-bearing mice, MDSCs can themselves facilitate the generation of tumor-specific Tregs. In this study, we demonstrate that expression of the immune stimulatory receptor CD40 on MDSCs is required to induce T-cell tolerance and Treg accumulation. In an immune reconstitution model, adoptive transfer of Gr-1+CD115+ monocytic MDSCs derived from CD40-deficient mice failed to recapitulate the ability of wild-type MDSCs to induce tolerance and Treg development in vivo. Agonistic anti-CD40 antibodies phenocopied the effect of CD40 deficiency and also improved the therapeutic efficacy of IL-12 and 4-1BB immunotherapy in the treatment of advanced tumors. Our findings suggest that CD40 is essential not only for MDSC-mediated immune suppression but also for tumor-specific Treg expansion. Blockade of CD40-CD40L interaction between MDSC and Treg may provide a new strategy to ablate tumoral immune suppression and thereby heighten responses to immunotherapy.

  3. The Regulatory Role of Rolipram on Inflammatory Mediators and Cholinergic/Adrenergic Stimulation-Induced Signals in Isolated Primary Mouse Submandibular Gland Cells

    PubMed Central

    Lee, Dong Un; Shin, Dong Min; Hong, Jeong Hee

    2016-01-01

    Exposure to bacterial lipopolysaccharides (LPS) induces inflammatory signals in salivary glands. We investigated the regulatory role of phosphodiesterase 4 (PDE4) inhibitor rolipram on inflammatory mediators and cholinergic/adrenergic stimulation-induced intracellular Ca2+ signaling in salivary acinar and ductal cells. Submandibular gland (SMG) expressed PDE4A through 4D mRNA and PDE4 was localized in the luminal membrane of SMG. LPS induced Ca2+ signaling and ROS production in SMG. Treatment with rolipram blocked LPS-induced Ca2+ increase and ROS production. The application of histamine evoked Ca2+ signals and ROS production, which were attenuated by rolipram in SMG cells. Moreover, LPS-induced NLRP3 inflammasome and cleaved caspase-1 were inhibited by rolipram. The inhibitory role of rolipram in ROS-induced Ca2+ signaling was mainly observed in acinar cells and not in ductal cells. Rolipram also protected SMG acinar but not ductal cells from LPS-induced cell membrane damage. In the case of cholinergic/adrenergic stimulation, carbachol/isoproterenol-induced Ca2+ signals were upregulated by the treatment of rolipram in SMG. In the case of cAMP-dependent ductal bicarbonate secretion by rolipram, no effect was observed on the modulation of ductal chloride/bicarbonate exchange activity. Rolipram could suppress the inflammatory signals and could be a potential therapeutic strategy against LPS-induced inflammation to protect the salivary gland cells. PMID:27143817

  4. Fusogenic-oligoarginine peptide-mediated silencing of the CIP2A oncogene suppresses oral cancer tumor growth in vivo.

    PubMed

    Alexander-Bryant, Angela A; Dumitriu, Anca; Attaway, Christopher C; Yu, Hong; Jakymiw, Andrew

    2015-11-28

    Intracellular delivery and endosomal escape of functional small interfering RNAs (siRNAs) remain major barriers limiting the clinical translation of RNA interference (RNAi)-based therapeutics. Recently, we demonstrated that a cell-penetrating endosome-disruptive peptide we synthesized, termed 599, enhanced the intracellular delivery and bioavailability of siRNAs designed to target the CIP2A oncoprotein (siCIP2A) into oral cancer cells and consequently inhibited oral cancer cell invasiveness and anchorage-independent growth in vitro. Thus, to further assess the therapeutic potential of the 599 peptide in mediating RNAi-based therapeutics for oral cancer and its prospective applicability in clinical settings, the objective of the current study was to determine whether intratumoral dosing of the 599 peptide-siCIP2A complex could induce silencing of CIP2A and consequently impair tumor growth using a xenograft oral cancer mouse model. Our results demonstrate that the 599 peptide is able to protect siRNAs from degradation by serum and ribonucleases in vitro and upon intratumoral injection in vivo, confirming the stability of the 599 peptide-siRNA complex and its potential for therapeutic utility. Moreover, 599 peptide-mediated delivery of siCIP2A to tumor tissue induces CIP2A silencing without any associated toxicity, consequently resulting in reduction of the mitotic index and significant inhibition of tumor growth. Together, these data suggest that the 599 peptide carrier is a clinically effective mediator of RNAi-based cancer therapeutics.

  5. Mitogen-Inducible Gene-6 Mediates Feedback Inhibition from Mutated BRAF towards the Epidermal Growth Factor Receptor and Thereby Limits Malignant Transformation.

    PubMed

    Milewska, Malgorzata; Romano, David; Herrero, Ana; Guerriero, Maria Luisa; Birtwistle, Marc; Quehenberger, Franz; Hatzl, Stefan; Kholodenko, Boris N; Segatto, Oreste; Kolch, Walter; Zebisch, Armin

    2015-01-01

    BRAF functions in the RAS-extracellular signal-regulated kinase (ERK) signaling cascade. Activation of this pathway is necessary to mediate the transforming potential of oncogenic BRAF, however, it may also cause a negative feedback that inhibits the epidermal growth factor receptor (EGFR). Mitogen-inducible gene-6 (MIG-6) is a potent inhibitor of the EGFR and has been demonstrated to function as a tumor suppressor. As MIG-6 can be induced via RAS-ERK signaling, we investigated its potential involvement in this negative regulatory loop. Focus formation assays were performed and demonstrated that MIG-6 significantly reduces malignant transformation induced by oncogenic BRAF. Although this genetic interaction was mirrored by a physical interaction between MIG-6 and BRAF, we did not observe a direct regulation of BRAF kinase activity by MIG-6. Interestingly, a selective chemical EGFR inhibitor suppressed transformation to a similar degree as MIG-6, whereas combining these approaches had no synergistic effect. By analyzing a range of BRAF mutated and wildtype cell line models, we could show that BRAF V600E causes a strong upregulation of MIG-6, which was mediated at the transcriptional level via the RAS-ERK pathway and resulted in downregulation of EGFR activation. This feedback loop is operational in tumors, as shown by the analysis of almost 400 patients with papillary thyroid cancer (PTC). Presence of BRAF V600E correlated with increased MIG-6 expression on the one hand, and with inactivation of the EGFR and of PI3K/AKT signaling on the other hand. Importantly, we also observed a more aggressive disease phenotype when BRAF V600E coexisted with low MIG-6 expression. Finally, analysis of methylation data was performed and revealed that higher methylation of MIG-6 correlated to its decreased expression. Taken together, we demonstrate that MIG-6 efficiently reduces cellular transformation driven by oncogenic BRAF by orchestrating a negative feedback circuit directed

  6. Mitogen-Inducible Gene-6 Mediates Feedback Inhibition from Mutated BRAF towards the Epidermal Growth Factor Receptor and Thereby Limits Malignant Transformation

    PubMed Central

    Milewska, Malgorzata; Romano, David; Herrero, Ana; Guerriero, Maria Luisa; Birtwistle, Marc; Quehenberger, Franz; Hatzl, Stefan; Kholodenko, Boris N.; Segatto, Oreste; Kolch, Walter; Zebisch, Armin

    2015-01-01

    BRAF functions in the RAS-extracellular signal-regulated kinase (ERK) signaling cascade. Activation of this pathway is necessary to mediate the transforming potential of oncogenic BRAF, however, it may also cause a negative feedback that inhibits the epidermal growth factor receptor (EGFR). Mitogen-inducible gene-6 (MIG-6) is a potent inhibitor of the EGFR and has been demonstrated to function as a tumor suppressor. As MIG-6 can be induced via RAS-ERK signaling, we investigated its potential involvement in this negative regulatory loop. Focus formation assays were performed and demonstrated that MIG-6 significantly reduces malignant transformation induced by oncogenic BRAF. Although this genetic interaction was mirrored by a physical interaction between MIG-6 and BRAF, we did not observe a direct regulation of BRAF kinase activity by MIG-6. Interestingly, a selective chemical EGFR inhibitor suppressed transformation to a similar degree as MIG-6, whereas combining these approaches had no synergistic effect. By analyzing a range of BRAF mutated and wildtype cell line models, we could show that BRAF V600E causes a strong upregulation of MIG-6, which was mediated at the transcriptional level via the RAS-ERK pathway and resulted in downregulation of EGFR activation. This feedback loop is operational in tumors, as shown by the analysis of almost 400 patients with papillary thyroid cancer (PTC). Presence of BRAF V600E correlated with increased MIG-6 expression on the one hand, and with inactivation of the EGFR and of PI3K/AKT signaling on the other hand. Importantly, we also observed a more aggressive disease phenotype when BRAF V600E coexisted with low MIG-6 expression. Finally, analysis of methylation data was performed and revealed that higher methylation of MIG-6 correlated to its decreased expression. Taken together, we demonstrate that MIG-6 efficiently reduces cellular transformation driven by oncogenic BRAF by orchestrating a negative feedback circuit directed

  7. Functional cooperation between Smad proteins and activator protein-1 regulates transforming growth factor-beta-mediated induction of endothelin-1 expression.

    PubMed

    Rodríguez-Pascual, Fernando; Redondo-Horcajo, Mariano; Lamas, Santiago

    2003-06-27

    Endothelin-1 (ET-1) is a 21-amino-acid potent vasoconstrictor peptide that is mainly produced by vascular endothelial cells. Expression of the ET-1 gene is subject to complex regulation by numerous factors, among which transforming growth factor-beta (TGF-beta) is one of the most important. It has been widely documented that TGF-beta increases ET-1 mRNA and peptide levels. We have explored the mechanism by which TGF-beta upregulates ET-1 expression in endothelial cells. Transcriptional activation of the ET-1 promoter accounted for the TGF-beta-induced increase in ET-1 mRNA levels. We have identified within the ET-1 promoter two DNA elements indispensable for TGF-beta-mediated induction of ET-1: an activator protein-1 (AP-1) site at -108/-102, known to be important for constitutive and induced expression, and a novel regulatory sequence located at -193/-171, which constitutes a specific binding site for Smad transcription factors. Mutation of both elements abolished TGF-beta responsiveness. Binding of Smad3/Smad4 and c-Jun to their corresponding DNA elements was evidenced by electrophoretic mobility shift assays. Furthermore, the coactivator CREB-binding protein (CBP)/p300 was found to play an essential role in the induction of the gene. The simultaneous requirement for two distinct and independent DNA elements suggests that Smads and activator protein-1 functionally cooperate through CBP/p300 to mediate TGF-beta-induced transcriptional activation of the ET-1 gene.

  8. Inhibition of CD1d-mediated antigen presentation by the transforming growth factor-β/Smad signalling pathway.

    PubMed

    Bailey, Jennifer C; Iyer, Abhirami K; Renukaradhya, Gourapura J; Lin, Yinling; Nguyen, Hoa; Brutkiewicz, Randy R

    2014-12-01

    CD1d-mediated lipid antigen presentation activates a subset of innate immune lymphocytes called invariant natural killer T (NKT) cells that, by virtue of their potent cytokine production, bridge the innate and adaptive immune systems. Transforming growth factor (TGF-β) is a known immune modulator that can activate the mitogen-activated protein kinase p38; we have previously shown that p38 is a negative regulator of CD1d-mediated antigen presentation. Several studies implicate a role for TGF-β in the activation of p38. Therefore, we hypothesized that TGF-β would impair antigen presentation by CD1d. Indeed, a dose-dependent decrease in CD1d-mediated antigen presentation and impairment of lipid antigen processing was observed in response to TGF-β treatment. However, it was found that this inhibition was not through p38 activation. Instead, Smads 2, 3 and 4, downstream elements of the TGF-β canonical signalling pathway, contributed to the observed effects. In marked contrast to that observed with CD1d, TGF-β was found to enhance MHC class II-mediated antigen presentation. Overall, these results suggest that the canonical TGF-β/Smad pathway negatively regulates an important arm of the host's innate immune responses - CD1d-mediated lipid antigen presentation to NKT cells.

  9. Investigating the use of in situ liquid cell scanning transmission electron microscopy to explore DNA-mediated gold nanoparticle growth

    NASA Astrophysics Data System (ADS)

    Nguy, Amanda

    Engineering nanoparticles with desired shape-dependent properties is the key to many applications in nanotechnology. Although many synthetic procedures exist to produce anisotropic gold nanoparticles, the dynamics of growth are typically unknown or hypothetical. In the case of seed-mediated growth in the presence of DNA into anisotropic nanoparticles, it is not known exactly how DNA directs growth into specific morphologies. A series of preliminary experiments were carried out to contribute to the investigation of the possible mechanism of DNA-mediated growth of gold nanoprisms into gold nanostars using liquid cell scanning transmission electron microscopy (STEM). Imaging in the liquid phase was achieved through the use of a liquid cell platform and liquid cell holder that allow the sample to be contained within a “chip sandwich” between two electron transparent windows. Ex situ growth experiments were performed using Au-T30 NPrisms (30-base thymine oligonucleotide-coated gold nanoprisms) that are expected to grow into gold nanostars. Growth to form these nanostars were imaged using TEM (transmission electron microscopy) and liquid cell STEM (scanning transmission electron microscopy). An attempt to perform in situ growth experiments with the same Au-T30 nanoprisms revealed challenges in obtaining desired morphology results due to the environmental differences within the liquid cell compared to the ex situ environment. Different parameters in the experimental method were explored including fluid line set up, simultaneous and alternating reagent addition, and the effect of different liquid cell volumes to ensure adequate flow of reagents into the liquid cell. Lastly, the binding affinities were compared for T30 and A30 DNA incubated with gold nanoparticles using zeta potential measurements, absorption spectroscopy, and isothermal titration calorimetry (ITC). It was previously reported thymine bases have a lower binding affinity to gold surfaces than

  10. RegulonDB (version 5.0): Escherichia coli K-12 transcriptional regulatory network, operon organization, and growth conditions

    PubMed Central

    Salgado, Heladia; Gama-Castro, Socorro; Peralta-Gil, Martín; Díaz-Peredo, Edgar; Sánchez-Solano, Fabiola; Santos-Zavaleta, Alberto; Martínez-Flores, Irma; Jiménez-Jacinto, Verónica; Bonavides-Martínez, César; Segura-Salazar, Juan; Martínez-Antonio, Agustino; Collado-Vides, Julio

    2006-01-01

    RegulonDB is the internationally recognized reference database of Escherichia coli K-12 offering curated knowledge of the regulatory network and operon organization. It is currently the largest electronically-encoded database of the regulatory network of any free-living organism. We present here the recently launched RegulonDB version 5.0 radically different in content, interface design and capabilities. Continuous curation of original scientific literature provides the evidence behind every single object and feature. This knowledge is complemented with comprehensive computational predictions across the complete genome. Literature-based and predicted data are clearly distinguished in the database. Starting with this version, RegulonDB public releases are synchronized with those of EcoCyc since our curation supports both databases. The complex biology of regulation is simplified in a navigation scheme based on three major streams: genes, operons and regulons. Regulatory knowledge is directly available in every navigation step. Displays combine graphic and textual information and are organized allowing different levels of detail and biological context. This knowledge is the backbone of an integrated system for the graphic display of the network, graphic and tabular microarray comparisons with curated and predicted objects, as well as predictions across bacterial genomes, and predicted networks of functionally related gene products. Access RegulonDB at . PMID:16381895

  11. Different regulatory and cytotoxic CD4+ T lymphocyte profiles in renal transplants with antibody-mediated chronic rejection or long-term good graft function.

    PubMed

    Giaretta, Fulvia; Bussolino, Stefania; Beltramo, Silvia; Fop, Fabrizio; Rossetti, Maura; Messina, Maria; Cantaluppi, Vincenzo; Ranghino, Andrea; Basso, Elisa; Camussi, Giovanni; Segoloni, Giuseppe Paolo; Biancone, Luigi

    2013-01-01

    Comparative analysis of the different subsets of CD4(+) T-lymphocytes may provide hints on the immunologic mechanisms operating in the long-term fate of a kidney transplant. We analyzed peripheral regulatory CD4(+) T cells (Tregs) and CD4(+) cytotoxic T lymphocytes (CTLs) in antibody-mediated chronic rejection (AMCR), in middle-term kidney transplants (2-4 years, MTKT) with good graft function and rejection-free history, in long-term kidney transplants (>15 years, LTKT) and in normal healthy subjects (NHS). Transplant groups with good prognosis (MTKT and LTKT) displayed a significant lower amount of CD4(+)CD25(high) T lymphocytes than NHS, with a trend of a higher percentage in AMCR than in MTKT and LTKT. However, CD4(+)CD25(high) Foxp3(+) cells were significantly higher in LTKT and MTKT than AMCR. Characterization of CD4(+)CD25(high) T cells showed a marked increase of intracellular CTLA-4 in the AMCR group in respect to the other transplant groups, while the expression of the surface molecule seemed to follow a reverse trend. In addition, CD27, a costimulatory receptor involved in long-term T cell survival and prevention of immune tolerance, is significantly reduced in CD4(+)CD25(high) and CD4(+)Foxp3(+) T cells in the LTKT in respect to the other transplant groups. CD4(+)CD25(high)CD45RO(+) and CD4(+)Foxp3(+)CD45RO(+) regulatory T cells with memory function were increased in LTKT compared to NHS and for the latter also in AMCR group. Finally, CD4(+)CTLs that were quantified on the basis of granzyme A expression, were more represented in AMCR patients in comparison to the other groups. Strikingly, CD27 in the CD4(+)CTLs was suppressed in LTKT and MTKT and markedly expressed in AMCR group. No significant differences in the expression of CD28 were observed among different groups. In conclusion, different profiles of Tregs and CD4(+)CTL populations correlate with different long-term conditions of kidney-transplanted patients, suggesting their role in the development

  12. S6K1 and E2FB are in mutually antagonistic regulatory links controlling cell growth and proliferation in Arabidopsis.

    PubMed

    Henriques, Rossana; Magyar, Zoltán; Bögre, László

    2013-06-01

    Plant development is dependent on the coordination between growth and cell proliferation. The nutrient sensing TOR kinase and its downstream target, the 40S ribosomal S6 Kinase, are central controllers of cell growth that were also shown to determine cell size by inhibiting the onset of mitosis in yeast and animal cells. We have shown that the Arabidopsis S6 Kinase1 inhibits cell proliferation through the RBR-E2FB complex. S6K1 interacts with RBR via its N-terminal RBR binding motif, promotes its nuclear localization and consequent RBR-dependent repression of cell cycle genes through E2FB. Here we show that S6K1 and E2FB are in a mutually antagonistic relationship both in their protein abundance and in their activity. We propose that this double inhibitory regulatory connection between S6K1 and E2FB forms a regulatory switch that might be important to determine whether cells divide or grow.

  13. RhoC Mediates Epidermal Growth Factor-Stimulated Migration and Invasion in Head and Neck Squamous Cell Carcinoma

    PubMed Central

    Tumur, Zohra; Katebzadeh, Shahbaz; Guerra, Carlos; Bhushan, Lokesh; Alkam, Tursun; Henson, Bradley S.

    2015-01-01

    Epidermal growth factor receptor (EGFR) is overexpressed in head and neck squamous cell carcinoma (HNSCC) where it has been shown to promote tumor cell invasion upon phosphorylation. One mechanism by which EGFR promotes tumor progression is by activating signal cascades that lead to loss of E-cadherin, a transmembrane glycoprotein of the cell-cell adherence junctions; however mediators of these signaling cascades are not fully understood. One such mediator, RhoC, is activated upon a number of external stimuli, such as epidermal growth factor (EGF), but its role as a mediator of EGF-stimulated migration and invasion has not been elucidated in HNSCC. In the present study, we investigate the role of RhoC as a mediator of EGF-stimulated migration and invasion in HNSCC. We show that upon EGF stimulation, EGFR and RhoC were strongly activated in HNSCC. This resulted in activation of the phosphatidylinositol 3-Kinase Akt pathway (PI3K-Akt), phosphorylation of GSK-3β at the Ser9 residue, and subsequent down regulation of E-cadherin cell surface expression resulting in increased tumor cell invasion. Knockdown of RhoC restored E-cadherin expression and inhibited EGF-stimulated migration and invasion. This is the first report in HNSCC demonstrating the role RhoC plays in mediating EGF-stimulated migration and invasion by down-regulating the PI3K-Akt pathway and E-cadherin expression. RhoC may serve as a treatment target for HNSCC. PMID:25622907

  14. Hsa-miRNA-765 as a key mediator for inhibiting growth, migration and invasion in fulvestrant-treated prostate cancer.

    PubMed

    Leung, Yuet-Kin; Chan, Queeny Kwan-Yi; Ng, Chi-Fai; Ma, Fanny Man-Ting; Tse, Ho-Man; To, Ka-Fai; Maranchie, Jodi; Ho, Shuk-Mei; Lau, Kin-Mang

    2014-01-01

    Fulvestrant (ICI-182,780) has recently been shown to effectively suppress prostate cancer cell growth in vitro and in vivo. But it is unclear whether microRNAs play a role in regulating oncogene expression in fulvestrant-treated prostate cancer. Here, this study reports hsa-miR-765 as the first fulvestrant-driven, ERβ-regulated miRNA exhibiting significant tumor suppressor activities like fulvestrant, against prostate cancer cell growth via blockage of cell-cycle progression at the G2/M transition, and cell migration and invasion possibly via reduction of filopodia/intense stress-fiber formation. Fulvestrant was shown to upregulate hsa-miR-765 expression through recruitment of ERβ to the 5'-regulatory-region of hsa-miR-765. HMGA1, an oncogenic protein in prostate cancer, was identified as a downstream target of hsa-miR-765 and fulvestrant in cell-based experiments and a clinical study. Both the antiestrogen and the hsa-miR-765 mimic suppressed HMGA1 protein expression. In a neo-adjuvant study, levels of hsa-miR-765 were increased and HMGA1 expression was almost completely lost in prostate cancer specimens from patients treated with a single dose (250 mg) of fulvestrant 28 days before prostatectomy. These findings reveal a novel fulvestrant signaling cascade involving ERβ-mediated transcriptional upregulation of hsa-miR-765 that suppresses HMGA1 protein expression as part of the mechanism underlying the tumor suppressor action of fulvestrant in prostate cancer.

  15. Co-Targeting Prostate Cancer Epithelium and Bone Stroma by Human Osteonectin-Promoter–Mediated Suicide Gene Therapy Effectively Inhibits Androgen-Independent Prostate Cancer Growth

    PubMed Central

    Sung, Shian-Ying; Chang, Junn-Liang; Chen, Kuan-Chou; Yeh, Shauh-Der; Liu, Yun-Ru; Su, Yen-Hao; Hsueh, Chia-Yen; Chung, Leland W. K.; Hsieh, Chia-Ling

    2016-01-01

    Stromal-epithelial interaction has been shown to promote local tumor growth and distant metastasis. We sought to create a promising gene therapy approach that co-targets cancer and its supporting stromal cells for combating castration-resistant prostate tumors. Herein, we demonstrated that human osteonectin is overexpressed in the prostate cancer epithelium and tumor stroma in comparison with their normal counterpart. We designed a novel human osteonectin promoter (hON-522E) containing positive transcriptional regulatory elements identified in both the promoter and exon 1 region of the human osteonectin gene. In vitro reporter assays revealed that the hON-522E promoter is highly active in androgen receptor negative and metastatic prostate cancer and bone stromal cells compared to androgen receptor-positive prostate cancer cells. Moreover, in vivo prostate-tumor–promoting activity of the hON-522E promoter was confirmed by intravenous administration of an adenoviral vector containing the hON-522E promoter-driven luciferase gene (Ad-522E-Luc) into mice bearing orthotopic human prostate tumor xenografts. In addition, an adenoviral vector with the hON-522E-promoter–driven herpes simplex virus thymidine kinase gene (Ad-522E-TK) was highly effective against the growth of androgen-independent human prostate cancer PC3M and bone stromal cell line in vitro and in pre-established PC3M tumors in vivo upon addition of the prodrug ganciclovir. Because of the heterogeneity of human prostate tumors, hON-522E promoter-mediated gene therapy has the potential for the treatment of hormone refractory and bone metastatic prostate cancers. PMID:27054343

  16. Growth Differentiation Factor-8 Decreases StAR Expression Through ALK5-Mediated Smad3 and ERK1/2 Signaling Pathways in Luteinized Human Granulosa Cells.

    PubMed

    Fang, Lanlan; Chang, Hsun-Ming; Cheng, Jung-Chien; Yu, Yiping; Leung, Peter C K; Sun, Ying-Pu

    2015-12-01

    Growth differentiation factor-8 (GDF-8) has been recently shown to be expressed in human granulosa cells, and the mature form of GDF-8 protein can be detected in the follicular fluid. However, the biological function and significance of this growth factor in the human ovary remains to be determined. Here, we investigated the effects of GDF-8 on steroidogenic enzyme expression and the potential mechanisms of action in luteinized human granulosa cells. We demonstrated that treatment with GDF-8 did not affect the mRNA levels of P450 side-chain cleavage enzyme and 3β-hydroxysteroid dehydrogenase, whereas it significantly down-regulated steroidogenic acute regulatory protein (StAR) expression and decreased progesterone production. The suppressive effect of GDF-8 on StAR expression was abolished by the inhibition of the TGF-β type I receptor. In addition, treatment with GDF-8 activated both Smad2/3 and ERK1/2 signaling pathways. Furthermore, knockdown of activin receptor-like kinase 5 reversed the effects of GDF-8 on Smad2/3 phosphorylation and StAR expression. The inhibition of Smad3 or ERK1/2 signaling pathways attenuated the GDF-8-induced down-regulation of StAR and production of progesterone. Interestingly, the concentrations of GDF-8 were negatively correlated with those of progesterone in human follicular fluid. These results indicate a novel autocrine function of GDF-8 to down-regulate StAR expression and decrease progesterone production in luteinized human granulosa cells, most likely through activin receptor-like kinase 5-mediated Smad3 and ERK1/2 signaling pathways. Our findings suggest that granulosa cells might play a critical role in the regulation of progesterone production to prevent premature luteinization during the final stage of folliculogenesis.

  17. Co-Targeting Prostate Cancer Epithelium and Bone Stroma by Human Osteonectin-Promoter-Mediated Suicide Gene Therapy Effectively Inhibits Androgen-Independent Prostate Cancer Growth.

    PubMed

    Sung, Shian-Ying; Chang, Junn-Liang; Chen, Kuan-Chou; Yeh, Shauh-Der; Liu, Yun-Ru; Su, Yen-Hao; Hsueh, Chia-Yen; Chung, Leland W K; Hsieh, Chia-Ling

    2016-01-01

    Stromal-epithelial interaction has been shown to promote local tumor growth and distant metastasis. We sought to create a promising gene therapy approach that co-targets cancer and its supporting stromal cells for combating castration-resistant prostate tumors. Herein, we demonstrated that human osteonectin is overexpressed in the prostate cancer epithelium and tumor stroma in comparison with their normal counterpart. We designed a novel human osteonectin promoter (hON-522E) containing positive transcriptional regulatory elements identified in both the promoter and exon 1 region of the human osteonectin gene. In vitro reporter assays revealed that the hON-522E promoter is highly active in androgen receptor negative and metastatic prostate cancer and bone stromal cells compared to androgen receptor-positive prostate cancer cells. Moreover, in vivo prostate-tumor-promoting activity of the hON-522E promoter was confirmed by intravenous administration of an adenoviral vector containing the hON-522E promoter-driven luciferase gene (Ad-522E-Luc) into mice bearing orthotopic human prostate tumor xenografts. In addition, an adenoviral vector with the hON-522E-promoter-driven herpes simplex virus thymidine kinase gene (Ad-522E-TK) was highly effective against the growth of androgen-independent human prostate cancer PC3M and bone stromal cell line in vitro and in pre-established PC3M tumors in vivo upon addition of the prodrug ganciclovir. Because of the heterogeneity of human prostate tumors, hON-522E promoter-mediated gene therapy has the potential for the treatment of hormone refractory and bone metastatic prostate cancers.

  18. AAV-Mediated angiotensin 1-7 overexpression inhibits tumor growth of lung cancer in vitro and in vivo

    PubMed Central

    Mao, Yingying; Wang, Shengyao; Yan, Renhe; Bai, Na; Li, Andrew; Zhang, Yanling; Du, Hongyan; Chen, Baihong; Sumners, Colin; Li, Jinlong; Li, Hongwei

    2017-01-01

    Ang-(1-7) inhibits lung cancer cell growth both in vitro and in vivo. However, the molecular mechanism of action is unclear and also the rapid degradation of Ang-(1-7) in vivo limits its clinical application. Here, we have demonstrated that Ang- (1-7) inhibits lung cancer cell growth by interrupting pre-replicative complex assembly and restrains epithelial-mesenchymal transition via Cdc6 inhibition. Furthermore, we constructed a mutant adeno-associated viral vector AAV8 (Y733F) that produced stable and high efficient Ang-(1-7) expression in a xenograft tumor model. The results show that AAV8-mediated Ang-(1-7) over-expression can remarkably suppress tumor growth in vivo by down-regulating Cdc6 and anti-angiogenesis. Ang-(1-7) over-expression via the AAV8 method may be a promising strategy for lung cancer treatment. PMID:27861149

  19. Physiological state, growth mode, and oxidative stress play a role in Cd(II)-mediated inhibition of Nitrosomonas europaea 19718.

    PubMed

    Chandran, Kartik; Love, Nancy G

    2008-04-01

    The goal of this study was to determine the impact of physiological growth states (batch exponential and batch stationary growth) and growth modes (substrate-limited chemostat, substrate-sufficient exponential batch, and substrate-depleted stationary batch growth) on several measures of growth and responses to Cd(II)-mediated inhibition of Nitrosomonas europaea strain 19718. The specific oxygen uptake rate (sOUR) was the most sensitive indicator of inhibition among the different responses analyzed, including total cell abundance, membrane integrity, intracellular 16S rRNA/DNA ratio, and amoA expression. This observation remained true irrespective of the physiological state, the growth mode, or the mode of Cd(II) exposure. Based on the sOUR, a strong time-dependent exacerbation of inhibition (in terms of an inhibition coefficient [K(i)]) in exponential batch cultures was observed. Long-term inhibition levels (based on K(i) estimates) in metabolically active chemostat and exponential batch cultures were also especially severe and comparable. In contrast, the inhibition level in stationary-phase cultures was 10-fold lower and invariable with exposure time. Different strategies for surviving substrate limitation (a 10-fold increase in amoA expression) and starvation (the retention of 16S rRNA levels) in N. europaea cultures were observed. amoA expression was most negatively impacted by Cd(II) exposure in the chemostat cultures, was less impacted in exponential batch cultures, and was least impacted in stationary batch cultures. Although the amoA response was consistent with that of the sOUR, the amoA response was not as strong. The intracellular 16S rRNA/DNA ratio, as determined by fluorescence in situ hybridization, also did not uniformly correlate with the sOUR under conditions of inhibition or no inhibition. Finally, Cd(II)-mediated inhibition of N. europaea was attributed partially to oxidative stress.

  20. The putative guanine nucleotide exchange factor RicA mediates upstream signaling for growth and development in Aspergillus.

    PubMed

    Kwon, Nak-Jung; Park, Hee-Soo; Jung, Seunho; Kim, Sun Chang; Yu, Jae-Hyuk

    2012-11-01

    Heterotrimeric G proteins (G proteins) govern growth, development, and secondary metabolism in various fungi. Here, we characterized ricA, which encodes a putative GDP/GTP exchange factor for G proteins in the model fungus Aspergillus nidulans and the opportunistic human pathogen Aspergillus fumigatus. In both species, ricA mRNA accumulates during vegetative growth and early developmental phases, but it is not present in spores. The deletion of ricA results in severely impaired colony growth and the total (for A. nidulans) or near (for A. fumigatus) absence of asexual sporulation (conidiation). The overexpression (OE) of the A. fumigatus ricA gene (AfricA) restores growth and conidiation in the ΔAnricA mutant to some extent, indicating partial conservation of RicA function in Aspergillus. A series of double mutant analyses revealed that the removal of RgsA (an RGS protein of the GanB Gα subunit), but not sfgA, flbA, rgsB, or rgsC, restored vegetative growth and conidiation in ΔAnricA. Furthermore, we found that RicA can physically interact with GanB in yeast and in vitro. Moreover, the presence of two copies or OE of pkaA suppresses the profound defects caused by ΔAnricA, indicating that RicA-mediated growth and developmental signaling is primarily through GanB and PkaA in A. nidulans. Despite the lack of conidiation, brlA and vosA mRNAs accumulated to normal levels in the ΔricA mutant. In addition, mutants overexpressing fluG or brlA (OEfluG or OEbrlA) failed to restore development in the ΔAnricA mutant. These findings suggest that the commencement of asexual development requires unknown RicA-mediated signaling input in A. nidulans.

  1. Effect of cadmium exposure on primary tumor growth and cell-mediated cytotoxicity in mice bearing MSB sarcomas.

    PubMed

    Kerkvliet, N I; Koller, L D; Baecher, L G; Brauner, J A

    1979-08-01

    In vivo MSB tumor growth and cell-mediated cytotoxicity (CMC) to MSB tumor cells in vitro were studied in male C57BL/6 mice exposed to 0, 3, 30, or 300 ppm Cd as CdCl2 in their drinking water for 21 weeks prior to and during tumor growth. CMC was assessed on days 5, 12, and 19 post injection with the use of both a 51Cr release assay and a 51Cr post-label assay. Cd exposure significantly inhibited the growth of MSB tumors in vivo and enhanced the levels of CMC in the tumor-bearing hosts. Peak levels of CMC on day 12 post tumor injection were significantly increased in Cd-exposed animals. However, whereas the inhibition of tumor growth was directly dependent on the dose of Cd, the enhancement of CMC was inversely related to dosage. These data suggested that other mechanisms in addition to increased CMC were involved in tumor growth inhibition. Possible factors such as direct inhibition of tumor growth by Cd and decreased serum blocking levels in Cd-exposed animals are discussed.

  2. Contributions of two-component regulatory systems, alternative sigma factors, and negative regulators to Listeria monocytogenes cold adaptation and cold growth.

    PubMed

    Chan, Yvonne C; Hu, Yuewei; Chaturongakul, Soraya; Files, Kali D; Bowen, Barbara M; Boor, Kathryn J; Wiedmann, Martin

    2008-02-01

    The ability of Listeria monocytogenes to grow at refrigeration temperatures is critical for transmission of this foodborne pathogen. We evaluated the contributions of different transcriptional regulators and two-component regulatory systems to L. monocytogenes cold adaptation and cold growth. L. monocytogenes parent strain 10403S and selected isogenic null mutants in genes encoding four alternative sigma factors (sigB, sigH, sigC, and sigL), two regulators of sigmaB (rsbT and rsbV), two negative regulators (ctsR and hrcA), and 15 two-component response regulators were grown in brain heart infusion broth at 4 degrees C with (i) a high-concentration starting inoculum (10(8) CFU/ml), (ii) a low-concentration starting inoculum (102 CFU/ml), and (iii) a high-concentration starting inoculum of cold-adapted cells. With a starting inoculum of 10(8) CFU/ml, null mutants in genes encoding selected alternative sigma factors (DeltasigH, DeltasigC, and DeltasigL), a negative regulator (DeltactsR), regulators of sigmaB (DeltarsbT and DeltarsbV), and selected two-component response regulators (DeltalisR, Deltalmo1172, and Deltalmo1060) had significantly reduced growth (P < 0.05) compared with the parent strain after 12 days at 4 degrees C. The growth defect for DeltasigL was limited and was not confirmed by optical density (OD600) measurement data. With a starting inoculum of 102 CFU/ml and after monitoring growth at 4 degrees C over 84 days, only the DeltactsR strain had a consistent but limited growth defect; the other mutant strains had either no growth defects or limited growth defects apparent at only one or two of the nine sampling points evaluated during the 84-day growth period (DeltasigB, DeltasigC, and Deltalmo1172). With a 10(8) CFU/ml starting inoculum of cold-adapted cells, none of the mutant strains that had a growth defect when inoculation was performed with cells pregrown at 37 degrees C had reduced growth as compared with the parent strain after 12 days at 4

  3. Characterization of intracellular growth regulator icgR by utilizing transcriptomics to identify mediators of pathogenesis in Shigella flexneri.

    PubMed

    Morris, Carolyn R; Grassel, Christen L; Redman, Julia C; Sahl, Jason W; Barry, Eileen M; Rasko, David A

    2013-09-01

    Shigella species Gram-negative bacteria which cause a diarrheal disease, known as shigellosis, by invading and destroying the colonic mucosa and inducing a robust inflammatory response. With no vaccine available, shigellosis annually kills over 600,000 children in developing countries. This study demonstrates the utility of combining high-throughput bioinformatic methods with in vitro and in vivo assays to provide new insights into pathogenesis. Comparisons of in vivo and in vitro gene expression identified genes associated with intracellular growth. Additional bioinformatics analyses identified genes that are present in S. flexneri isolates but not in the three other Shigella species. Comparison of these two analyses revealed nine genes that are differentially expressed during invasion and that are specific to S. flexneri. One gene, a DeoR family transcriptional regulator with decreased expression during invasion, was further characterized and is now designated icgR, for intracellular growth regulator. Deletion of icgR caused no difference in growth in vitro but resulted in increased intracellular replication in HCT-8 cells. Further in vitro and in vivo studies using high-throughput sequencing of RNA transcripts (RNA-seq) of an isogenic ΔicgR mutant identified 34 genes that were upregulated under both growth conditions. This combined informatics and functional approach has allowed the characterization of a gene and pathway previously unknown in Shigella pathogenesis and provides a framework for further identification of novel virulence factors and regulatory pathways.

  4. The Mediator Kinase Module Restrains Epidermal Growth Factor Receptor Signaling and Represses Vulval Cell Fate Specification in Caenorhabditis elegans.

    PubMed

    Grants, Jennifer M; Ying, Lisa T L; Yoda, Akinori; You, Charlotte C; Okano, Hideyuki; Sawa, Hitoshi; Taubert, Stefan

    2016-02-01

    Cell signaling pathways that control proliferation and determine cell fates are tightly regulated to prevent developmental anomalies and cancer. Transcription factors and coregulators are important effectors of signaling pathway output, as they regulate downstream gene programs. In Caenorhabditis elegans, several subunits of the Mediator transcriptional coregulator complex promote or inhibit vulva development, but pertinent mechanisms are poorly defined. Here, we show that Mediator's dissociable cyclin dependent kinase 8 (CDK8) module (CKM), consisting of cdk-8, cic-1/Cyclin C, mdt-12/dpy-22, and mdt-13/let-19, is required to inhibit ectopic vulval cell fates downstream of the epidermal growth factor receptor (EGFR)-Ras-extracellular signal-regulated kinase (ERK) pathway. cdk-8 inhibits ectopic vulva formation by acting downstream of mpk-1/ERK, cell autonomously in vulval cells, and in a kinase-dependent manner. We also provide evidence that the CKM acts as a corepressor for the Ets-family transcription factor LIN-1, as cdk-8 promotes transcriptional repression by LIN-1. In addition, we find that CKM mutation alters Mediator subunit requirements in vulva development: the mdt-23/sur-2 subunit, which is required for vulva development in wild-type worms, is dispensable for ectopic vulva formation in CKM mutants, which instead display hallmarks of unrestrained Mediator tail module activity. We propose a model whereby the CKM controls EGFR-Ras-ERK transcriptional output by corepressing LIN-1 and by fine tuning Mediator specificity, thus balancing transcriptional repression vs. activation in a critical developmental signaling pathway. Collectively, these data offer an explanation for CKM repression of EGFR signaling output and ectopic vulva formation and provide the first evidence of Mediator CKM-tail module subunit crosstalk in animals.

  5. Sodium Benzoate, a Food Additive and a Metabolite of Cinnamon, Enriches Regulatory T Cells via STAT6-Mediated Upregulation of TGF-β.

    PubMed

    Kundu, Madhuchhanda; Mondal, Susanta; Roy, Avik; Martinson, Jeffrey L; Pahan, Kalipada

    2016-10-15

    Upregulation and/or maintenance of regulatory T cells (Tregs) during autoimmune insults may have therapeutic efficacy in autoimmune diseases. Earlier we have reported that sodium benzoate (NaB), a metabolite of cinnamon and a Food and Drug Administration-approved drug against urea cycle disorders, upregulates Tregs and protects mice from experimental allergic encephalomyelitis, an animal model of multiple sclerosis. However, mechanisms by which NaB increases Tregs are poorly understood. Because TGF-β is an important inducer of Tregs, we examined the effect of NaB on the status of TGF-β. In this study, we demonstrated that NaB induced the expression of TGF-β mRNA and protein in normal as well as proteolipid protein-primed splenocytes. The presence of a consensus STAT6 binding site in the promoter of the TGF-β gene, activation of STAT6 in splenocytes by NaB, recruitment of STAT6 to the TGF-β promoter by NaB, and abrogation of NaB-induced expression of TGF-β in splenocytes by small interfering RNA knockdown of STAT6 suggest that NaB induces the expression of TGF-β via activation of STAT6. Furthermore, we demonstrated that blocking of TGF-β by neutralizing Abs abrogated NaB-mediated protection of Tregs and experimental allergic encephalomyelitis. These studies identify a new function of NaB in upregulating TGF-β via activation of STAT6, which may be beneficial in MS patients.

  6. Mediator subunits MED1 and MED24 cooperatively contribute to pubertal mammary gland development and growth of breast carcinoma cells.

    PubMed

    Hasegawa, Natsumi; Sumitomo, Akiko; Fujita, Azusa; Aritome, Nami; Mizuta, Shumpei; Matsui, Keiji; Ishino, Ruri; Inoue, Kana; Urahama, Norinaga; Nose, Junko; Mukohara, Toru; Kamoshida, Shingo; Roeder, Robert G; Ito, Mitsuhiro

    2012-04-01

    The Mediator subunit MED1 is essential for mammary gland development and lactation, whose contribution through direct interaction with estrogen receptors (ERs) is restricted to involvement in pubertal mammary gland development and luminal cell differentiation. Here, we provide evidence that the MED24-containing submodule of Mediator functionally communicates specifically with MED1 in pubertal mammary gland development. Mammary glands from MED1/MED24 double heterozygous knockout mice showed profound retardation in ductal branching during puberty, while single haploinsufficient glands developed normally. DNA synthesis of both luminal and basal cells were impaired in double mutant mice, and the expression of ER-targeted genes encoding E2F1 and cyclin D1, which promote progression through the G(1)/S phase of the cell cycle, was attenuated. Luciferase reporter assays employing double mutant mouse embryonic fibroblasts showed selective impairment in ER functions. Various breast carcinoma cell lines expressed abundant amounts of MED1, MED24, and MED30, and attenuated expression of MED1 and MED24 in breast carcinoma cells led to attenuated DNA synthesis and growth. These results indicate functional communications between the MED1 subunit and the MED24-containing submodule that mediate estrogen receptor functions and growth of both normal mammary epithelial cells and breast carcinoma cells.

  7. Prolonged induction of p21Cip1/WAF1/CDK2/PCNA complex by epidermal growth factor receptor activation mediates ligand-induced A431 cell growth inhibition

    PubMed Central

    1995-01-01

    Proliferation of some cultured human tumor cell lines bearing high numbers of epidermal growth factor (EGF) receptors is paradoxically inhibited by EGF in nanomolar concentrations. In the present study, we have investigated the biochemical mechanism of growth inhibition in A431 human squamous carcinoma cells exposed to exogenous EGF. In parallel, we studied a selected subpopulation, A431-F, which is resistant to EGF-mediated growth inhibition. We observed a marked reduction in cyclin-dependent kinase-2 (CDK2) activity when A431 and A431-F cells were cultured with 20 nM EGF for 4 h. After further continuous exposure of A431 cells to EGF, the CDK2 activity remained at a low level and was accompanied by persistent G1 arrest. In contrast, the early reduced CDK2 activity and G1 accumulation in A431-F cells was only transient. We found that, at early time points (4-8 h), EGF induces p21Cip1/WAF1 mRNA and protein expression in both EGF-sensitive A431 cells and EGF-resistant A431-F cells. But only in A431 cells, was p21Cip1/WAF1 expression sustained at a significantly increased level for up to 5 d after addition of EGF. Induction of p21Cip1/WAF1 by EGF could be inhibited by a specific EGF receptor tyrosine kinase inhibitor, tyrphostin AG1478, suggesting that p21Cip1/WAF1 induction was a consequence of receptor tyrosine kinase activation by EGF. We also demonstrated that the increased p21Cip1/WAF1 was associated with both CDK2 and proliferating cell nuclear antigen (PCNA). Taken together, our results demonstrate that p21Cip1/WAF1 is an important mediator of EGF-induced G1 arrest and growth inhibition in A431 cells. PMID:7559780

  8. The Orphan Nuclear Receptor ERRγ Regulates Hepatic CB1 Receptor-Mediated Fibroblast Growth Factor 21 Gene Expression

    PubMed Central

    Jung, Yoon Seok; Lee, Ji-Min; Kim, Don-Kyu; Lee, Yong-Soo; Kim, Ki-Sun; Kim, Yong-Hoon; Kim, Jina; Lee, Myung-Shik; Lee, In-Kyu; Kim, Seong Heon; Cho, Sung Jin; Jeong, Won-Il; Lee, Chul-Ho; Harris, Robert A.; Choi, Hueng-Sik

    2016-01-01

    Background Fibroblast growth factor 21 (FGF21), a stress inducible hepatokine, is synthesized in the liver and plays important roles in glucose and lipid metabolism. However, the mechanism of hepatic cannabinoid type 1 (CB1) receptor-mediated induction of FGF21 gene expression is largely unknown. Results Activation of the hepatic CB1 receptor by arachidonyl-2’-chloroethylamide (ACEA), a CB1 receptor selective agonist, significantly increased FGF21 gene expression. Overexpression of estrogen-related receptor (ERR) γ increased FGF21 gene expression and secretion both in hepatocytes and mice, whereas knockdown of ERRγ decreased ACEA-mediated FGF21 gene expression and secretion. Moreover, ERRγ, but not ERRα and ERRβ, induced FGF21 gene promoter activity. In addition, deletion and mutation analysis of the FGF21 promoter identified a putative ERRγ-binding motif (AGGTGC, a near-consensus response element). A chromatin immunoprecipitation assay revealed direct binding of ERRγ to the FGF21 gene promoter. Finally, GSK5182, an ERRγ inverse agonist, significantly inhibited hepatic CB1 receptor-mediated FGF21 gene expression and secretion. Conclusion Based on our data, we conclude that ERRγ plays a key role in hepatic CB1 receptor-mediated induction of FGF21 gene expression and secretion. PMID:27455076

  9. Docking protein SNT1 is a critical mediator of fibroblast growth factor signaling during Xenopus embryonic development.

    PubMed

    Akagi, Keiko; Kyun Park, Eui; Mood, Kathleen; Daar, Ira O

    2002-03-01

    The docking protein SNT1/FRS2 (fibroblast growth factor receptor substrate 2) is implicated in the transmission of extracellular signals from several growth factor receptors to the mitogen-activated protein (MAP) kinase signaling cascade, but its biological function during development is not well characterized. Here, we show that the Xenopus homolog of mammalian SNT1/FRS-2 (XSNT1) plays a critical role in the appropriate formation of mesoderm-derived tissue during embryogenesis. XSNT1 has an expression pattern that is quite similar to the fibroblast growth factor receptor-1 (FGFR1) during Xenopus development. Ectopic expression of XSNT1 markedly enhanced the embryonic defects induced by an activated FGF receptor, and increased the MAP kinase activity as well as the expression of a mesodermal marker in response to FGF receptor signaling. A loss-of-function study using antisense XSNT1 morpholino oligonucleotides (XSNT-AS) shows severe malformation of trunk and posterior structures. Moreover, XSNT-AS disrupts muscle and notochord formation, and inhibits FGFR-induced MAP kinase activation. In ectodermal explants, XSNT-AS blocks FGFR-mediated induction of mesoderm and the accompanying elongation movements. Our results indicate that XSNT1 is a critical mediator of FGF signaling and is required for early Xenopus development.

  10. Personal growth initiative and identity formation in adolescence through young adulthood: mediating processes on the pathway to well-being.

    PubMed

    Luyckx, Koen; Robitschek, Christine

    2014-10-01

    Developing a personal identity is a challenging task throughout adolescence and the transition to adulthood. The present study sampling 551 14-35 year olds (54.1% female) examined personal growth initiative (PGI) as a potential predictor of core identity processes and explored whether identity functioned as a mediator on the pathway from PGI to self-esteem and depressive symptoms. Path analyses from a structural equation modeling approach indicated that all four components of PGI (i.e., planfulness, readiness for change, intentional behavior, and using resources) predicted different commitment and exploration processes, with planfulness being the most consistent predictor. Important age differences linking PGI-components to identity exploration were found. Finally, especially the degree to which individuals identified themselves with their identity commitments and the degree to which they relied on ruminative or maladaptive forms of identity exploration mediated pathways from PGI to self-esteem and depressive symptoms. Implications and suggestions for future research are discussed.

  11. Regulatory effects of Spirulina complex polysaccharides on growth of murine RSV-M glioma cells through Toll-like receptor 4.

    PubMed

    Kawanishi, Yu; Tominaga, Akira; Okuyama, Hiromi; Fukuoka, Satoshi; Taguchi, Takahiro; Kusumoto, Yutaka; Yawata, Toshio; Fujimoto, Yasunori; Ono, Shiro; Shimizu, Keiji

    2013-01-01

    This study is the first to report that Spirulina complex polysaccharides (CPS) suppress glioma growth by down-regulating angiogenesis via a Toll-like receptor 4 signal. Murine RSV-M glioma cells were implanted s.c. into C3H/HeN mice and TLR4 mutant C3H/HeJ mice. Treatment with either Spirulina CPS or Escherichia coli (E. coli) lipopolysaccharides (LPS) strongly suppressed RSV-M glioma cell growth in C3H/HeN, but not C3H/HeJ, mice. Glioma cells stimulated production of interleukin (IL)-17 in both C3H/HeN and C3H/HeJ tumor-bearing mice. Treatment with E. coli LPS induced much greater IL-17 production in tumor-bearing C3H/HeN mice than in tumor-bearing C3H/HeJ mice. In C3H/HeN mice, treatment with Spirulina CPS suppressed growth of re-transplanted glioma; however, treatment with E. coli LPS did not, suggesting that Spirulina CPS enhance the immune response. Administration of anti-cluster of differentiation (CD)8, anti-CD4, anti-CD8 antibodies, and anti-asialo GM1 antibodies enhanced tumor growth, suggesting that T cells and natural killer cells or macrophages are involved in suppression of tumor growth by Spirulina CPS. Although anti-interferon-γ antibodies had no effect on glioma cell growth, anti-IL-17 antibodies administered four days after tumor transplantation suppressed growth similarly to treatment with Spirulina CPS. Less angiogenesis was observed in gliomas from Spirulina CPS-treated mice than in those from saline- or E. coli LPS-treated mice. These findings suggest that, in C3H/HeN mice, Spirulina CPS antagonize glioma cell growth by down-regulating angiogenesis, and that this down-regulation is mediated in part by regulating IL-17 production.

  12. Suppression of polymorphonuclear (PMN) and monocyte-mediated inhibition of Candida albicans growth by delta-9-tetrahydrocannabinol

    SciTech Connect

    Djeu, J.Y.; Parapanios, A.; Halkias, D.; Friedman, H.

    1986-03-05

    This study was an in vitro attempt to identify the effector cells responsible for growth inhibition of the opportunistic fungus, candida albicans, and to determine if THC or another marijuana derivatives, 11-hydroxyTHC, would adversely affect their function. Using a 24h radiolabel assay, the authors found that growth inhibition of C. albicans was primarily mediated by PMN and monocytes that could be isolated normal human peripheral blood. Both effector cell types caused almost complete inhibition of Candida growth at effector/target ratio of 300/1 and inhibition was often still seen at 30/1-. Incubation of PMN, PBL, or monocytes for 1 hr at 37C with THC or 11-hydroxyTHC caused a marked suppression of function in all 3 cell populations. Maximal suppression was obtained with 7.5-10..mu..g/ml of the drugs in medium containing 10% fetal bovine serum (FBS) or with 2-4..mu..g/ml in 1% FBS. These drug concentrations did not affect lymphoid cell viability or candida growth in the absence of lymphoid effector cells. Marijuana derivatives, therefore, are doubly dangerous in that opportunistic fungi such as C. albicans can grow in their presence while the effector cells that control fungal growth are readily inactivated.

  13. Relationship between nutritionally-mediated placental growth restriction and fetal growth, body composition and endocrine status during late gestation in adolescent sheep.

    PubMed

    Wallace, J M; Bourke, D A; Aitken, R P; Palmer, R M; Da Silva, P; Cruickshank, M A

    2000-01-01

    The aim was to investigate the consequences of nutritionally-mediated placental growth restriction on fetal organ growth, conformation, body composition and endocrine status during late gestation. Embryos recovered from superovulated adult ewes inseminated by a single sire were transferred in singleton to the uterus of peripubertal adolescent recipients. Post-transfer, adolescent dams were offered a high (H) or moderate (M) level of a complete diet to promote rapid or moderate maternal growth rates, respectively (n=7 per group). After day 100 of gestation the feed intake of the M dams was adjusted weekly to maintain body condition score. Liveweight gain during the first 100 days of gestation was 301+/-24 and 90+/-4.6 g/day for the H and M groups, respectively. Maternal plasma concentrations of insulin, IGF-I and urea were significantly higher and non-esterified fatty acid concentrations significantly lower in H compared with M dams prior to slaughter on day 128 of gestation. At this stage of gestation, total placentome weight was 50 per cent lower in H compared with M groups (P< 0.001) and was associated with a 37 per cent reduction in fetal weight (P< 0.01). All variables of fetal conformation and absolute fetal organ weights, with the exception of the adrenal glands, were lower (P< 0. 05) in the fetuses from H intake dams. However, relative fetal organ weights expressed as g/kg fetal body weight, with the exception of the gut, were not influenced by maternal dietary intake. Furthermore, fetal weight but not maternal nutritional group were predictive of individual organ weight for all organs dissected. Together these results imply that growth restriction in the fetuses derived from H intake dams was largely symmetrical. Fetal plasma concentrations of insulin, IGF-I and glucose were attenuated (P< 0.05) in fetuses from H compared with M groups. The lower fetal body weight in the former group was associated with a reduction in absolute but not relative crude protein

  14. HB-EGF release mediates glucose-induced activation of the epidermal growth factor receptor in mesangial cells.

    PubMed

    Uttarwar, L; Peng, F; Wu, D; Kumar, S; Gao, B; Ingram, A J; Krepinsky, J C

    2011-04-01

    Glomerular matrix accumulation is a hallmark of diabetic nephropathy. We showed that transactivation of the epidermal growth factor receptor (EGFR) is an important mediator of matrix upregulation in mesangial cells (MC) in response to high glucose (HG). Here, we study the mechanism of EGFR transactivation. In primary MC, EGFR transactivation by 1 h of HG (30 mM) was unaffected by inhibitors of protein kinase C, reactive oxygen species, or the angiotensin II AT1 receptor. However, general metalloprotease inhibition, as well as specific inhibitors of heparin-binding EGF-like growth factor (HB-EGF), prevented both EGFR and downstream Akt activation. HB-EGF was released into the medium by 30 min of HG, and this depended on metalloprotease activity. One of the metalloproteases shown to cleave proHB-EGF is ADAM17 (TACE). HG, but not an osmotic control, activated ADAM17, and its inhibition prevented EGFR and Akt activation and HB-EGF release into the medium. siRNA to either ADAM17 or HB-EGF prevented HG-induced EGFR transactivation. We previously showed that EGFR/Akt signaling increases transforming growth factor (TGF)-β1 transcription through the transcription factor activator protein (AP)-1. HG-induced AP-1 activation, as assessed by EMSA, was abrogated by inhibitors of metalloproteases, HB-EGF and ADAM17. HB-EGF and ADAM17 siRNA also prevented AP-1 activation. Finally, these inhibitors and siRNA prevented TGF-β1 upregulation by HG. Thus, HG-induced EGFR transactivation in MC is mediated by the release of HB-EGF, which requires activity of the metalloprotease ADAM17. The mechanism of ADAM17 activation awaits identification. Targeting upstream mediators of EGFR transactivation including HB-EGF or ADAM17 provides novel therapeutic targets for the treatment of diabetic nephropathy.

  15. Diet and energy-sensing inputs affect TorC1-mediated axon misrouting but not TorC2-directed synapse growth in a Drosophila model of tuberous sclerosis.

    PubMed

    Dimitroff, Brian; Howe, Katie; Watson, Adrienne; Campion, Bridget; Lee, Hyun-Gwan; Zhao, Na; O'Connor, Michael B; Neufeld, Thomas P; Selleck, Scott B

    2012-01-01

    The Target of Rapamycin (TOR) growth regulatory system is influenced by a number of different inputs, including growth factor signaling, nutrient availability, and cellular energy levels. While the effects of TOR on cell and organismal growth have been well characterized, this pathway also has profound effects on neural development and behavior. Hyperactivation of the TOR pathway by mutations in the upstream TOR inhibitors TSC1 (tuberous sclerosis complex 1) or TSC2 promotes benign tumors and neurological and behavioral deficits, a syndrome known as tuberous sclerosis (TS). In Drosophila, neuron-specific overexpression of Rheb, the direct downstream target inhibited by Tsc1/Tsc2, produced significant synapse overgrowth, axon misrouting, and phototaxis deficits. To understand how misregulation of Tor signaling affects neural and behavioral development, we examined the influence of growth factor, nutrient, and energy sensing inputs on these neurodevelopmental phenotypes. Neural expression of Pi3K, a principal mediator of growth factor inputs to Tor, caused synapse overgrowth similar to Rheb, but did not disrupt axon guidance or phototaxis. Dietary restriction rescued Rheb-mediated behavioral and axon guidance deficits, as did overexpression of AMPK, a component of the cellular energy sensing pathway, but neither was able to rescue synapse overgrowth. While axon guidance and behavioral phenotypes were affected by altering the function of a Tor complex 1 (TorC1) component, Raptor, or a TORC1 downstream element (S6k), synapse overgrowth was only suppressed by reducing the function of Tor complex 2 (TorC2) components (Rictor, Sin1). These findings demonstrate that different inputs to Tor signaling have distinct activities in nervous system development, and that Tor provides an important connection between nutrient-energy sensing systems and patterning of the nervous system.

  16. Androgen Receptor-Mediated Growth Suppression of HPr-1AR and PC3-Lenti-AR Prostate Epithelial Cells

    PubMed Central

    Bolton, Eric C.

    2015-01-01

    The androgen receptor (AR) mediates the developmental, physiologic, and pathologic effects of androgens including 5α-dihydrotestosterone (DHT). However, the mechanisms whereby AR regulates growth suppression and differentiation of luminal epithelial cells in the prostate gland and proliferation of malignant versions of these cells are not well understood, though they are central to prostate development, homeostasis, and neoplasia. Here, we identify androgen-responsive genes that restrain cell cycle progression and proliferation of human prostate epithelial cell lines (HPr-1AR and PC3-Lenti-AR), and we investigate the mechanisms through which AR regulates their expression. DHT inhibited proliferation of HPr-1AR and PC3-Lenti-AR, and cell cycle analysis revealed a prolonged G1 interval. In the cell cycle, the G1/S-phase transition is initiated by the activity of cyclin D and cyclin-dependent kinase (CDK) complexes, which relieve growth suppression. In HPr-1AR, cyclin D1/2 and CDK4/6 mRNAs were androgen-repressed, whereas CDK inhibitor, CDKN1A, mRNA was androgen-induced. The regulation of these transcripts was AR-dependent, and involved multiple mechanisms. Similar AR-mediated down-regulation of CDK4/6 mRNAs and up-regulation of CDKN1A mRNA occurred in PC3-Lenti-AR. Further, CDK4/6 overexpression suppressed DHT-inhibited cell cycle progression and proliferation of HPr-1AR and PC3-Lenti-AR, whereas CDKN1A overexpression induced cell cycle arrest. We therefore propose that AR-mediated growth suppression of HPr-1AR involves cyclin D1 mRNA decay, transcriptional repression of cyclin D2 and CDK4/6, and transcriptional activation of CDKN1A, which serve to decrease CDK4/6 activity. AR-mediated inhibition of PC3-Lenti-AR proliferation occurs through a similar mechanism, albeit without down-regulation of cyclin D. Our findings provide insight into AR-mediated regulation of prostate epithelial cell proliferation. PMID:26372468

  17. An auxin-mediated shift toward growth isotropy promotes organ formation at the shoot meristem in Arabidopsis.

    PubMed

    Sassi, Massimiliano; Ali, Olivier; Boudon, Frédéric; Cloarec, Gladys; Abad, Ursula; Cellier, Coralie; Chen, Xu; Gilles, Benjamin; Milani, Pascale; Friml, Jiří; Vernoux, Teva; Godin, Christophe; Hamant, Olivier; Traas, Jan

    2014-10-06

    To control morphogenesis, molecular regulatory networks have to interfere with the mechanical properties of the individual cells of developing organs and tissues, but how this is achieved is not well known. We study this issue here in the shoot meristem of higher plants, a group of undifferentiated cells where complex changes in growth rates and directions lead to the continuous formation of new organs. Here, we show that the plant hormone auxin plays an important role in this process via a dual, local effect on the extracellular matrix, the cell wall, which determines cell shape. Our study reveals that auxin not only causes a limited reduction in wall stiffness but also directly interferes with wall anisotropy via the regulation of cortical microtubule dynamics. We further show that to induce growth isotropy and organ outgrowth, auxin somehow interferes with the cortical microtubule-ordering activity of a network of proteins, including AUXIN BINDING PROTEIN 1 and KATANIN 1. Numerical simulations further indicate that the induced isotropy is sufficient to amplify the effects of the relatively minor changes in wall stiffness to promote organogenesis and the establishment of new growth axes in a robust manner.

  18. The Mediator Kinase Module Restrains Epidermal Growth Factor Receptor Signaling and Represses Vulval Cell Fate Specification in Caenorhabditis elegans

    PubMed Central

    Grants, Jennifer M.; Ying, Lisa T. L.; Yoda, Akinori; You, Charlotte C.; Okano, Hideyuki; Sawa, Hitoshi; Taubert, Stefan

    2016-01-01

    Cell signaling pathways that control proliferation and determine cell fates are tightly regulated to prevent developmental anomalies and cancer. Transcription factors and coregulators are important effectors of signaling pathway output, as they regulate downstream gene programs. In Caenorhabditis elegans, several subunits of the Mediator transcriptional coregulator complex promote or inhibit vulva development, but pertinent mechanisms are poorly defined. Here, we show that Mediator’s dissociable cyclin dependent kinase 8 (CDK8) module (CKM), consisting of cdk-8, cic-1/Cyclin C, mdt-12/dpy-22, and mdt-13/let-19, is required to inhibit ectopic vulval cell fates downstream of the epidermal growth factor receptor (EGFR)-Ras-extracellular signal-regulated kinase (ERK) pathway. cdk-8 inhibits ectopic vulva formation by acting downstream of mpk-1/ERK, cell autonomously in vulval cells, and in a kinase-dependent manner. We also provide evidence that the CKM acts as a corepressor for the Ets-family transcription factor LIN-1, as cdk-8 promotes transcriptional repression by LIN-1. In addition, we find that CKM mutation alters Mediator subunit requirements in vulva development: the mdt-23/sur-2 subunit, which is required for vulva development in wild-type worms, is dispensable for ectopic vulva formation in CKM mutants, which instead display hallmarks of unrestrained Mediator tail module activity. We propose a model whereby the CKM controls EGFR-Ras-ERK transcriptional output by corepressing LIN-1 and by fine tuning Mediator specificity, thus balancing transcriptional repression vs. activation in a critical developmental signaling pathway. Collectively, these data offer an explanation for CKM repression of EGFR signaling output and ectopic vulva formation and provide the first evidence of Mediator CKM-tail module subunit crosstalk in animals. PMID:26715664

  19. Modulation of HCV reinfection after orthotopic liver transplantation by fibroblast growth factor-2 and other non-interferon mediators

    PubMed Central

    Van, Nguyen Dinh; Falk, Christine S; Sandmann, Lisa; Vondran, Florian W R; Helfritz, Fabian; Wedemeyer, Heiner; Manns, Michael P; Ciesek, Sandra; von Hahn, Thomas

    2016-01-01

    Objective In HCV infected individuals graft infection occurs shortly after orthotopic liver transplantation (OLT). We aimed to describe the composition of the inflammatory response at this time, how it affects the HCV replication cycle and identify novel proviral and antiviral factors. Design We used a Luminex assay to quantify 50 inflammatory mediators in sera before and shortly after OLT. In vitro grown HCV based on the JFH-1 isolate were used to characterise the effects of patient sera and individual mediators on HCV. Results Although the mediator composition is highly variable between individuals, sera drawn immediately post-OLT significantly enhance HCV infectivity compared with control sera from before OLT in about half of the cases. Among 27 non-interferon inflammatory mediators fibroblast growth factor (FGF)-2 stood out as it enhanced HCV RNA replication and release of infectious particles. The effect was concentration-dependent and detectable in dividing and non-dividing cells. Moreover, pharmacological inhibition of FGF-2 receptor signalling abrogated the enhancing effect of FGF-2 and inhibited HCV replication in the absence of serum FGF-2 suggesting that HCV replication is dependent on basal activation of the FGF-2 triggered signalling pathway. Finally, in individuals with chronic HCV infection with high viral load, serum FGF-2 was significantly higher compared with those with low viral load. Conclusions Although no single mediator may account for this effect, serum shortly post-OLT enhances HCV infection. FGF-2 is a novel endogenous driver of HCV replication and a potential therapeutic target. PMID:25800783

  20. Regulatory function of organic carbon supplementation on biodiesel production during growth and nutrient stress phases of mixotrophic microalgae cultivation.

    PubMed

    Chandra, Rashmi; Rohit, M V; Swamy, Y V; Venkata Mohan, S

    2014-08-01

    Critical role of organic carbon supplementation on the lipid synthesis during growth and nutrient deprived stress phase was investigated in present study. Mixotrophic cultivation showed relatively higher biomass productivity at lower carbon loading condition (500mgCOD/l). Nutrient deprivation induced physiological stress and glucose supplementation with 2000mgCOD/l supported higher lipid accumulation (26%). Glucose supplementation in mixotrophic growth phase showed distinct influence on biomass growth whereas glucose supplementation in nutrient starvation resulted in higher lipid storage. Compositional variation in FAME profile was observed with respect to saturated fatty acids when operated with increasing glucose concentrations. Mixotrophic mode of cultivation showed remarkable benefits of nutrient removal and organic carbon supplementation influenced greatly on biodiesel production which can be easily scaled up to pilot plant and large scale production facilities.

  1. Overexpression of the PP2A regulatory subunit Tap46 leads to enhanced plant growth through stimulation of the TOR signalling pathway.

    PubMed

    Ahn, Chang Sook; Ahn, Hee-Kyung; Pai, Hyun-Sook

    2015-02-01

    Tap46, a regulatory subunit of protein phosphatase 2A (PP2A), plays an essential role in plant growth and development through a functional link with the Target of Rapamycin (TOR) signalling pathway. Here, we have characterized the molecular mechanisms behind a gain-of-function phenotype of Tap46 and its relationship with TOR to gain further insights into Tap46 function in plants. Constitutive overexpression of Tap46 in Arabidopsis resulted in overall growth stimulation with enlarged organs, such as leaves and siliques. Kinematic analysis of leaf growth revealed that increased cell size was mainly responsible for the leaf enlargement. Tap46 overexpression also enhanced seed size and viability under accelerated ageing conditions. Enhanced plant growth was also observed in dexamethasone (DEX)-inducible Tap46 overexpression Arabidopsis lines, accompanied by increased cellular activities of nitrate-assimilating enzymes. DEX-induced Tap46 overexpression and Tap46 RNAi resulted in increased and decreased phosphorylation of S6 kinase (S6K), respectively, which is a sensitive indicator of endogenous TOR activity, and Tap46 interacted with S6K in planta based on bimolecular fluorescence complementation and co-immunoprecipitation. Furthermore, inactivation of TOR by estradiol-inducible RNAi or rapamycin treatment decreased Tap46 protein levels, but increased PP2A catalytic subunit levels. Real-time quantitative PCR analysis revealed that Tap46 overexpression induced transcriptional modulation of genes involved in nitrogen metabolism, ribosome biogenesis, and lignin biosynthesis. These findings suggest that Tap46 modulates plant growth as a positive effector of the TOR signalling pathway and Tap46/PP2Ac protein abundance is regulated by TOR activity.

  2. A phenomenological model for mechanically mediated growth, remodeling, damage, and plasticity of gel-derived tissue engineered blood vessels.

    PubMed

    Raykin, Julia; Rachev, Alexander I; Gleason, Rudolph L

    2009-10-01

    Mechanical stimulation has been shown to dramatically improve mechanical and functional properties of gel-derived tissue engineered blood vessels (TEBVs). Adjusting factors such as cell source, type of extracellular matrix, cross-linking, magnitude, frequency, and time course of mechanical stimuli (among many other factors) make interpretation of experimental results challenging. Interpretation of data from such multifactor experiments requires modeling. We present a modeling framework and simulations for mechanically mediated growth, remodeling, plasticity, and damage of gel-derived TEBVs that merge ideas from classical plasticity, volumetric growth, and continuum damage mechanics. Our results are compared with published data and suggest that this model framework can predict the evolution of geometry and material behavior under common experimental loading scenarios.

  3. Development of mechanisms associated with neurogenic-mediated skin inflammation during the growth of rats.

    PubMed

    Ohshima, Mihoko; Miyake, Mio; Takeda, Masanori; Muto, Taichiro; Ueda, Norishi; Ito, Komei; Sakamoto, Tatsuo

    2010-04-01

    Neurogenic-mediated inflammation may be associated with several inflammatory skin diseases including atopic dermatitis. However, age-dependent differences in neurogenic-mediated skin responses are not fully understood. We compared skin plasma leakage in rats aged 2 and 8 wk, which was induced by topical capsaicin, topical formalin, and intracutaneous substance P, whose effects are mediated via tachykinin NK1 receptors. Evans blue dye extravasation served as an index of the increase in skin vascular permeability. Capsaicin, formalin, and substance P caused a skin response in a dose-dependent manner in both age groups. However, the skin response was much greater in adults than in pups. In addition, the localization of sensory C-fibers and tachykinin NK1 receptors in the skin was investigated by immunofluorescent staining with antisubstance P and antitachykinin NK1 receptor antibodies, respectively. Substance P-immunoreactive nerves were detected throughout the dermis and tachykinin NK1 receptors were mainly detected in blood vessel walls in the dermis in both age groups. However, they were more sparsely distributed in pups. In conclusion, the weak neurogenic-mediated skin inflammation in pups is probably because of immature neural mechanisms associated with skin inflammation such as reduced innervation of sensory C-fibers and low expression of tachykinin NK1 receptors.

  4. Fibroblast growth factor 10-fibroblast growth factor receptor 2b mediated signaling is not required for adult glandular stomach homeostasis.

    PubMed

    Speer, Allison L; Al Alam, Denise; Sala, Frederic G; Ford, Henri R; Bellusci, Saverio; Grikscheit, Tracy C

    2012-01-01

    The signaling pathways that are essential for gastric organogenesis have been studied in some detail; however, those that regulate the maintenance of the gastric epithelium during adult homeostasis remain unclear. In this study, we investigated the role of Fibroblast growth factor 10 (FGF10) and its main receptor, Fibroblast growth factor receptor 2b (FGFR2b), in adult glandular stomach homeostasis. We first showed that mouse adult glandular stomach expressed Fgf10, its receptors, Fgfr1b and Fgfr2b, and most of the other FGFR2b ligands (Fgf1, Fgf7, Fgf22) except for Fgf3 and Fgf20. Fgf10 expression was mesenchymal whereas FGFR1 and FGFR2 expression were mostly epithelial. Studying double transgenic mice that allow inducible overexpression of Fgf10 in adult mice, we showed that Fgf10 overexpression in normal adult glandular stomach increased epithelial proliferation, drove mucous neck cell differentiation, and reduced parietal and chief cell differentiation. Although a similar phenotype can be associated with the development of metaplasia, we found that Fgf10 overexpression for a short duration does not cause metaplasia. Finally, investigating double transgenic mice that allow the expression of a soluble form of Fgfr2b, FGF10's main receptor, which acts as a dominant negative, we found no significant changes in gastric epithelial proliferation or differentiation in the mutants. Our work provides evidence, for the first time, that the FGF10-FGFR2b signaling pathway is not required for epithelial proliferation and differentiation during adult glandular stomach homeostasis.

  5. Pαx6 Expression in Postmitotic Neurons Mediates the Growth of Axons in Response to SFRP1

    PubMed Central

    Sebastián-Serrano, Alvaro; Sandonis, Africa; Cardozo, Marcos; Rodríguez-Tornos, Fernanda M.; Bovolenta, Paola; Nieto, Marta

    2012-01-01

    During development, the mechanisms that specify neuronal subclasses are coupled to those that determine their axonal response to guidance cues. Pax6 is a homedomain transcription factor required for the specification of a variety of neural precursors. After cell cycle exit, Pax6 expression is often shut down in the precursor progeny and most postmitotic neurons no longer express detectable levels of the protein. There are however exceptions and high Pax6 protein levels are found, for example, in postmitotic retinal ganglion cells (RGCs), dopaminergic neurons of the olfactory bulb and the limbic system in the telencephalon. The function of Pax6 in these differentiating neurons remains mostly elusive. Here, we demonstrate that Pax6 mediates the response of growing axons to SFRP1, a secreted molecule expressed in several Pax6-positive forebrain territories. Forced expression of Pax6 in cultured postmitotic cortical neurons, which do not normally express Pax6, was sufficient to increment axonal length. Growth was blocked by the addition of anti-SFRP1 antibodies, whereas exogenously added SFRP1 increased axonal growth of Pax6-transfected neurons but not that of control or untransfected cortical neurons. In the reverse scenario, shRNA-mediated knock-down of Pax6 in mouse retinal explants specifically abolished RGCs axonal growth induced by SFRP1, but had no effect on RGCs differentiation and it did not modify the effect of Shh or Netrin on axon growth. Taken together these results demonstrate that expression of Pax6 is necessary and sufficient to render postmitotic neurons competent to respond to SFRP1. These results reveal a novel and unexpected function of Pax6 in postmitotic neurons and situate Pax6 and SFRP1 as pair regulators of axonal connectivity. PMID:22359602

  6. Plant growth regulatory effect and insecticidal activity of the extracts of the Tree of Heaven (Ailanthus altissima L.)

    PubMed Central

    Tsao, Rong; Romanchuk, Frieda E; Peterson, Chris J; Coats, Joel R

    2002-01-01

    Background There is an urgent need to explore and utilize naturally occurring products for combating harmful agricultural and public health pests. Secondary metabolites in the leaves of the Tree of Heaven, Ailanthus altissima L. have been reported to be herbicidal and insecticidal. The mode of action, however, of the active compounds in A. altissima are not understood. In this paper, we report the chemical characteristics of the herbicidal and insecticidal components in this tree, and will discuss the effect of light on the bioactivity of the active components. Results Extracts from the fresh leaves of A. altissima showed a strong plant germination/growth inhibitory effect in laboratory bioassays against alfalfa (Medicago sativa). The effect was dose-dependent. The growth inhibitory components were in the methylene chloride soluble fraction of the extract. The effect was greater in the light than in the dark. Other fractions had plant growth enhancing effect at lower concentrations. The extract was slightly insecticidal against yellow fever mosquito larvae (Aedes aegypti). Conclusions The extract or its semi-purified fractions of A. altissima were strong plant growth inhibitors, therefore good candidates as potential environmentally safe and effective agricultural pest management agents. The finding that light affects the activity will be useful in the application of such natural products. PMID:11860616

  7. Lentivirus-mediated RNAi knockdown of insulin-like growth factor-1 receptor inhibits the growth and invasion of hepatocellular carcinoma via down-regulating midkine expression

    PubMed Central

    Huang, Qiu Yan; Tang, Hui Jun; Wang, Min; Cao, Guo Li; Yi, Ting Zhuang; Wu, Sheng Lan; Xu, Wei Jie; Tang, Shao Hui

    2016-01-01

    The insulin-like growth factor-1 receptor (IGF-1R) overexpression contributes to the development of a variety of cancers. The present study explored the role of IGF-1R in the development and progression of hepatocellular carcinoma (HCC) and the possibility of IGF-1R silencing by lentivirus-mediated RNA interference (RNAi) as a therapeutic target for HCC. We showed that IGF-1R mRNA was up-regulated in Huh7 and Hep3B cells and human HCC tissues, and that IGF-1R knockdown by RNAi led to decreased proliferation, apoptosis induction, and decreased migration and invasion of Huh7 and Hep3B cells. Further, the in vivo study indicated that IGF-1R knockdown markedly diminished the tumorigenesis and metastasis of Huh7 xenograft. Moreover, the intratumoral administration of lentivirus-IGF-1R siRNA led to significant tumor growth inhibition in an established Huh7 xenograft model. Mechanistic investigations showed that midkine was found to be the most significantly down-regulated protein in Huh7 cells with IGF-1R knockdown, and ectopic overexpression of midkine significantly rescued inhibition of Huh7 cell proliferation, migration, and invasion caused by IGF-1R suppression. Collectively, these data suggest that IGF-1R inhibition by RNAi can significantly suppress HCC growth and invasion at least partially through down-regulating midkine expression, and IGF-1R is a potential target for HCC gene therapy. PMID:27813495

  8. Spectroscopic studies of nucleic acid additions during seed-mediated growth of gold nanoparticles

    PubMed Central

    Tapp, Maeling; Sullivan, Rick; Dennis, Patrick; Naik, Rajesh R.

    2015-01-01

    The effect of adding nucleic acids to gold seeds during the growth stage of either nanospheres or nanorods was investigated using UV-Vis spectroscopy to reveal any oligonucleotide base or structure-specific effects on nanoparticle growth kinetics or plasmonic signatures. Spectral data indicate that the presence of DNA duplexes during seed ageing drastically accelerated nanosphere growth while the addition of single-stranded polyadenine at any point during seed ageing induces nanosphere aggregation. For seeds added to a gold nanorod growth solution, single-stranded polythymine induces a modest blue-shift in the longitudinal peak wavelength. Moreover, a particular sequence comprised of 50% thymine bases was found to induce a faster, more dramatic blue-shift in the longitudinal peak wavelength compared to any of the homopolymer incubation cases. Monomeric forms of the nucleic acids, however, do not yield discernable spectral differences in any of the gold suspensions studied. PMID:25960601

  9. Teacher Prejudice as a Mediating Factor in Student Growth and Development.

    ERIC Educational Resources Information Center

    Gay, Geneva

    1979-01-01

    Teacher prejudices exert powerful influences on student growth and development and are naturally restrictive whether the prejudice comes from biases about intelligence, sexuality, ethnicity, physical appearance, or socioeconomic status. (JMF)

  10. Expression of the rat sterol regulatory element-binding protein-1c gene in response to insulin is mediated by increased transactivating capacity of specificity protein 1 (Sp1).

    PubMed

    Deng, Xiong; Yellaturu, Chandrahasa; Cagen, Lauren; Wilcox, Henry G; Park, Edwards A; Raghow, Rajendra; Elam, Marshall B

    2007-06-15

    The induction of genes involved in lipid biosynthesis by insulin is mediated in part by the sterol regulatory element-binding protein-1c (SREBP-1c). SREBP-1c is directly regulated by insulin by transcriptional and post-transcriptional mechanisms. Previously, we have demonstrated that the insulin-responsive cis-acting unit of the rat SREBP-1c promoter is composed of several elements that include a sterol regulatory element, two liver X receptor elements, and a number of conserved GC boxes. Here we systematically dissected the role of these GC boxes and report that five bona fide Sp1-binding elements of the SREBP-1c promoter determine its basal and insulin-induced activation. Luciferase expression driven by the rat SREBP-1c promoter was accelerated by ectopic expression of Sp1, and insulin further enhanced the transactivation potential of Sp1. Introduction of a small interfering RNA against Sp1 reduced both basal and insulin-induced activation of the SREBP-1c promoter. We also found that Sp1 interacted with both SREBP-1c and LXRalpha proteins and that insulin promoted these interactions. Chromatin immunoprecipitation studies revealed that insulin facilitated the recruitment of the steroid receptor coactivator-1 to the SREBP-1c promoter. These studies identify a novel mechanism by which maximal activation of the rat SREBP-1c gene expression by insulin is mediated by Sp1 and its enhanced ability to interact with other transcriptional regulatory proteins.

  11. MDP25, A Novel Calcium Regulatory Protein, Mediates Hypocotyl Cell Elongation by Destabilizing Cortical Microtubules in Arabidopsis[C][W][OA

    PubMed Central

    Li, Jiejie; Wang, Xianling; Qin, Tao; Zhang, Yan; Liu, Xiaomin; Sun, Jingbo; Zhou, Yuan; Zhu, Lei; Zhang, Ziding; Yuan, Ming; Mao, Tonglin

    2011-01-01

    The regulation of hypocotyl elongation is important for plant growth. Microtubules play a crucial role during hypocotyl cell elongation. However, the molecular mechanism underlying this process is not well understood. In this study, we describe a novel Arabidopsis thaliana microtubule-destabilizing protein 25 (MDP25) as a negative regulator of hypocotyl cell elongation. We found that MDP25 directly bound to and destabilized microtubules to enhance microtubule depolymerization in vitro. The seedlings of mdp25 mutant Arabidopsis lines had longer etiolated hypocotyls. In addition, MDP25 overexpression resulted in significant overall shortening of hypocotyl cells, which exhibited destabilized cortical microtubules and abnormal cortical microtubule orientation, suggesting that MDP25 plays a crucial role in the negative regulation of hypocotyl cell elongation. Although MDP25 localized to the plasma membrane under normal conditions, increased calcium levels in cells caused MDP25 to partially dissociate from the plasma membrane and move into the cytosol. Cellular MDP25 bound to and destabilized cortical microtubules, resulting in their reorientation, and subsequently inhibited hypocotyl cell elongation. Our results suggest that MDP25 exerts its function on cortical microtubules by responding to cytoplasmic calcium levels to mediate hypocotyl cell elongation. PMID:22209764

  12. Contrasting growth phenology of native and invasive forest shrubs mediated by genome size.

    PubMed

    Fridley, Jason D; Craddock, Alaä

    2015-08-01

    Examination of the significance of genome size to plant invasions has been largely restricted to its association with growth rate. We investigated the novel hypothesis that genome size is related to forest invasions through its association with growth phenology, as a result of the ability of large-genome species to grow more effectively through cell expansion at cool temperatures. We monitored the spring leaf phenology of 54 species of eastern USA deciduous forests, including native and invasive shrubs of six common genera. We used new measurements of genome size to evaluate its association with spring budbreak, cell size, summer leaf production rate, and photosynthetic capacity. In a phylogenetic hierarchical model that differentiated native and invasive species as a function of summer growth rate and spring budbreak timing, species with smaller genomes exhibited both faster growth and delayed budbreak compared with those with larger nuclear DNA content. Growth rate, but not budbreak timing, was associated with whether a species was native or invasive. Our results support genome size as a broad indicator of the growth behavior of woody species. Surprisingly, invaders of deciduous forests show the same small-genome tendencies of invaders of more open habitats, supporting genome size as a robust indicator of invasiveness.

  13. The Staphylococcus aureus SrrAB Regulatory System Modulates Hydrogen Peroxide Resistance Factors, Which Imparts Protection to Aconitase during Aerobic Growth

    PubMed Central

    Mashruwala, Ameya A.; Boyd, Jeffrey M.

    2017-01-01

    The SrrAB two-component regulatory system (TCRS) positively influences the transcription of genes involved in aerobic respiration in response to changes in respiratory flux. Hydrogen peroxide (H2O2) can arise as a byproduct of spontaneous interactions between dioxygen and components of respiratory pathways. H2O2 damages cellular factors including protein associated iron-sulfur cluster prosthetic groups. We found that a Staphylococcus aureus strain lacking the SrrAB two-component regulatory system (TCRS) is sensitive to H2O2 intoxication. We tested the hypothesis that SrrAB manages the mutually inclusive expression of genes required for aerobic respiration and H2O2 resistance. Consistent with our hypothesis, a ΔsrrAB strain had decreased transcription of genes encoding for H2O2 resistance factors (kat, ahpC, dps). SrrAB was not required for the inducing the transcription of these genes in cells challenged with H2O2. Purified SrrA bound to the promoter region for dps suggesting that SrrA directly influences dps transcription. The H2O2 sensitivity of the ΔsrrAB strain was alleviated by iron chelation or deletion of the gene encoding for the peroxide regulon repressor (PerR). The positive influence of SrrAB upon H2O2 metabolism bestowed protection upon the solvent accessible iron-sulfur (FeS) cluster of aconitase from H2O2 poisoning. SrrAB also positively influenced transcription of scdA (ytfE), which encodes for a FeS cluster repair protein. Finally, we found that SrrAB positively influences H2O2 resistance only during periods of high dioxygen-dependent respiratory activity. SrrAB did not influence H2O2 resistance when cellular respiration was diminished as a result of decreased dioxygen availability, and negatively influenced it in the absence of respiration (fermentative growth). We propose a model whereby SrrAB-dependent regulatory patterns facilitate the adaptation of cells to changes in dioxygen concentrations, and thereby aids in the prevention of H2O2

  14. Calcium-Mediated Induction of Paradoxical Growth following Caspofungin Treatment Is Associated with Calcineurin Activation and Phosphorylation in Aspergillus fumigatus.

    PubMed

    Juvvadi, Praveen R; Muñoz, Alberto; Lamoth, Frédéric; Soderblom, Erik J; Moseley, M Arthur; Read, Nick D; Steinbach, William J

    2015-08-01

    The echinocandin antifungal drug caspofungin at high concentrations reverses the growth inhibition of Aspergillus fumigatus, a phenomenon known as the "paradoxical effect," which is not consistently observed with other echinocandins (micafungin and anidulafungin). Previous studies of A. fumigatus revealed the loss of the paradoxical effect following pharmacological or genetic inhibition of calcineurin, yet the underlying mechanism is poorly understood. Here, we utilized a codon-optimized bioluminescent Ca(2+) reporter aequorin expression system in A. fumigatus and showed that caspofungin elicits a transient increase in cytosolic free Ca(2+) ([Ca(2+)]c) in the fungus that acts as the initial trigger of the paradoxical effect by activating calmodulin-calcineurin signaling. While the increase in [Ca(2+)]c was also observed upon treatment with micafungin, another echinocandin without the paradoxical effect, a higher [Ca(2+)]c increase was noted with the paradoxical-growth concentration of caspofungin. Treatments with a Ca(2+)-selective chelator, BAPTA [1,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid], or the L-type Ca(2+) channel blocker verapamil abolished caspofungin-mediated paradoxical growth in both the wild-type and the echinocandin-resistant (EMFR-S678P) strains. Concomitant with increased [Ca(2+)]c levels at higher concentrations of caspofungin, calmodulin and calcineurin gene expression was enhanced. Phosphoproteomic analysis revealed that calcineurin is activated through phosphorylation at its serine-proline-rich region (SPRR), a domain previously shown to be essential for regulation of hyphal growth, only at a paradoxical-growth concentration of caspofungin. Our results indicate that as opposed to micafungin, the increased [Ca(2+)]c at high concentrations of caspofungin activates calmodulin-calcineurin signaling at both a transcriptional and a posttranslational level and ultimately leads to paradoxical fungal growth.

  15. Salt stress differentially affects growth-mediating β-expansins in resistant and sensitive maize (Zea mays L.).

    PubMed

    Geilfus, Christoph-Martin; Zörb, Christian; Mühling, Karl H

    2010-12-01

    Salinity mainly reduces shoot growth by the inhibition of cell division and elongation. Expansins loosen plant cell walls. Moreover, the expression of some isoforms is clearly correlated with growth. Effects of salinity on β-expansin transcripts protein abundance were recently reported for different crop species. This study provides a broad analysis of the impact of an 8-day 100mM NaCl stress treatment on the mRNA expression of different maize (Zea mays L.) β-Expansin isoforms using real-time quantitative RT-PCR. The composite β-expansin protein expression was analyzed by western blotting using an anti-peptide antibody raised against a conserved 15-amino-acid region shared by vegetatively expressed β-expansin isoforms. For the first time, changes in β-expansin transcript and protein abundance have been analyzed together with the salinity-induced inhibition of shoot growth. A salt-resistant and a salt-sensitive cultivar were compared in order to elucidate physiological changes. Genotypic differences in the relative concentration of six β-expansin transcripts together with differences in the abundance β-expansin protein are shown in response NaCl stress. In salt-sensitive Lector, reduced β-expansin protein expression was found to correlate positively with reduced shoot growth under stress. A down-regulation of ZmExpB2, ZmExpB6, and ZmExpB8 transcripts possibly contribute to this decrease in protein abundance. In contrast, the maintenance of shoot growth in salt-resistant SR03 might be related to an unaffected abundance of growth-mediating β-expansin proteins in the shoot. Our data suggest that the up-regulation of ZmExpB2, ZmExpB6, and ZmExpB8 may sustain the stable expression of β-expansin protein under conditions of salt stress.

  16. Atropine-enhanced, antigen challenge-induced airway hyperreactivity in guinea pigs is mediated by eosinophils and nerve growth factor.

    PubMed

    Verbout, Norah G; Jacoby, David B; Gleich, Gerald J; Fryer, Allison D

    2009-08-01

    Although anticholinergic therapy inhibits bronchoconstriction in asthmatic patients and antigen-challenged animals, administration of atropine 1 h before antigen challenge significantly potentiates airway hyperreactivity and eosinophil activation measured 24 h later. This potentiation in airway hyperreactivity is related to increased eosinophil activation and is mediated at the level of the airway nerves. Since eosinophils produce nerve growth factor (NGF), which is known to play a role in antigen-induced airway hyperreactivity, we tested whether NGF mediates atropine-enhanced, antigen challenge-induced hyperreactivity. Antibody to NGF (Ab NGF) was administered to sensitized guinea pigs with and without atropine pretreatment (1 mg/kg iv) 1 h before challenge. At 24 h after challenge, animals were anesthetized, vagotomized, paralyzed, and ventilated. Electrical stimulation of both vagus nerves caused bronchoconstriction that was increased in challenged animals. Atropine pretreatment potentiated antigen challenge-induced hyperreactivity. Ab NGF did not affect eosinophils or inflammatory cells in any group, nor did it prevent hyperreactivity in challenged animals that were not pretreated with atropine. However, Ab NGF did prevent atropine-enhanced, antigen challenge-induced hyperreactivity and eosinophil activation (assessed by immunohistochemistry). This effect was specific to NGF, since animals given control IgG remained hyperreactive. These data suggest that anticholinergic therapy amplifies eosinophil interactions with airway nerves via NGF. Therefore, therapeutic strategies that target both eosinophil activation and NGF-mediated inflammatory processes in allergic asthma are likely to be beneficial.

  17. Local Epidermal Growth Factor Receptor Signaling Mediates the Systemic Pathogenic Effects of Staphylococcus aureus Toxic Shock Syndrome

    PubMed Central

    Gillman, Aaron N.; Stach, Christopher S.; Schlievert, Patrick M.; Peterson, Marnie L.

    2016-01-01

    Secreted factors of Staphylococcus aureus can activate host signaling from the epidermal growth factor receptor (EGFR). The superantigen toxic shock syndrome toxin-1 (TSST-1) contributes to mucosal cytokine production through a disintegrin and metalloproteinase (ADAM)-mediated shedding of EGFR ligands and subsequent EGFR activation. The secreted hemolysin, α-toxin, can also induce EGFR signaling and directly interacts with ADAM10, a sheddase of EGFR ligands. The current work explores the role of EGFR signaling in menstrual toxic shock syndrome (mTSS), a disease mediated by TSST-1. The data presented show that TSST-1 and α-toxin induce ADAM- and EGFR-dependent cytokine production from human vaginal epithelial cells. TSST-1 and α-toxin also induce cytokine production from an ex vivo porcine vaginal mucosa (PVM) model. EGFR signaling is responsible for the majority of IL-8 production from PVM in response to secreted toxins and live S. aureus. Finally, data are presented demonstrating that inhibition of EGFR signaling with the EGFR-specific tyrosine kinase inhibitor AG1478 significantly increases survival in a rabbit model of mTSS. These data indicate that EGFR signaling is critical for progression of an S. aureus exotoxin-mediated disease and may represent an attractive host target for therapeutics. PMID:27414801

  18. Methylglyoxal, a glycolysis side-product, induces Hsp90 glycation and YAP-mediated tumor growth and metastasis

    PubMed Central

    Nokin, Marie-Julie; Durieux, Florence; Peixoto, Paul; Chiavarina, Barbara; Peulen, Olivier; Blomme, Arnaud; Turtoi, Andrei; Costanza, Brunella; Smargiasso, Nicolas; Baiwir, Dominique; Scheijen, Jean L; Schalkwijk, Casper G; Leenders, Justine; De Tullio, Pascal; Bianchi, Elettra; Thiry, Marc; Uchida, Koji; Spiegel, David A; Cochrane, James R; Hutton, Craig A; De Pauw, Edwin; Delvenne, Philippe; Belpomme, Dominique; Castronovo, Vincent; Bellahcène, Akeila

    2016-01-01

    Metabolic reprogramming toward aerobic glycolysis unavoidably induces methylglyoxal (MG) formation in cancer cells. MG mediates the glycation of proteins to form advanced glycation end products (AGEs). We have recently demonstrated that MG-induced AGEs are a common feature of breast cancer. Little is known regarding the impact of MG-mediated carbonyl stress on tumor progression. Breast tumors with MG stress presented with high nuclear YAP, a key transcriptional co-activator regulating tumor growth and invasion. Elevated MG levels resulted in sustained YAP nuclear localization/activity that could be reverted using Carnosine, a scavenger for MG. MG treatment affected Hsp90 chaperone activity and decreased its binding to LATS1, a key kinase of the Hippo pathway. Cancer cells with high MG stress showed enhanced growth and metastatic potential in vivo. These findings reinforce the cumulative evidence pointing to hyperglycemia as a risk factor for cancer incidence and bring renewed interest in MG scavengers for cancer treatment. DOI: http://dx.doi.org/10.7554/eLife.19375.001 PMID:27759563

  19. Focal Adhesion Kinase-mediated Phosphorylation of Beclin1 Protein Suppresses Cardiomyocyte Autophagy and Initiates Hypertrophic Growth*♦

    PubMed Central

    Cheng, Zhaokang; Zhu, Qiang; Dee, Rachel; Opheim, Zachary; Mack, Christopher P.; Cyr, Douglas M.; Taylor, Joan M.

    2017-01-01

    Autophagy is an evolutionarily conserved intracellular degradation/recycling system that is essential for cellular homeostasis but is dysregulated in a number of diseases, including myocardial hypertrophy. Although it is clear that limiting or accelerating autophagic flux can result in pathological cardiac remodeling, the physiological signaling pathways that fine-tune cardiac autophagy are poorly understood. Herein, we demonstrated that stimulation of cardiomyocytes with phenylephrine (PE), a well known hypertrophic agonist, suppresses autophagy and that activation of focal adhesion kinase (FAK) is necessary for PE-stimulated autophagy suppression and subsequent initiation of hypertrophic growth. Mechanistically, we showed that FAK phosphorylates Beclin1, a core autophagy protein, on Tyr-233 and that this post-translational modification limits Beclin1 association with Atg14L and reduces Beclin1-dependent autophagosome formation. Remarkably, although ectopic expression of wild-type Beclin1 promoted cardiomyocyte atrophy, expression of a Y233E phosphomimetic variant of Beclin1 failed to affect cardiomyocyte size. Moreover, genetic depletion of Beclin1 attenuated PE-mediated/FAK-dependent initiation of myocyte hypertrophy in vivo. Collectively, these findings identify FAK as a novel negative regulator of Beclin1-mediated autophagy and indicate that this pathway can facilitate the promotion of compensatory hypertrophic growth. This novel mechanism to limit Beclin1 activity has important implications for treating a variety of pathologies associated with altered autophagic flux. PMID:27994061

  20. Activation of microtubule dynamics increases neuronal growth via the nerve growth factor (NGF)- and Gαs-mediated signaling pathways.

    PubMed

    Sarma, Tulika; Koutsouris, Athanasia; Yu, Jiang Zhu; Krbanjevic, Aleksandar; Hope, Thomas J; Rasenick, Mark M

    2015-04-17

    Signals that activate the G protein Gαs and promote neuronal differentiation evoke Gαs internalization in rat pheochromocytoma (PC12) cells. These agents also significantly increase Gαs association with microtubules, resulting in an increase in microtubule dynamics because of the activation of tubulin GTPase by Gαs. To determine the function of Gαs/microtubule association in neuronal development, we used real-time trafficking of a GFP-Gαs fusion protein. GFP-Gαs concentrates at the distal end of the neurites in differentiated living PC12 cells as well as in cultured hippocampal neurons. Gαs translocates to specialized membrane compartments at tips of growing neurites. A dominant-negative Gα chimera that interferes with Gαs binding to tubulin and activation of tubulin GTPase attenuates neurite elongation and neurite number both in PC12 cells and primary hippocampal neurons. This effect is greatest on differentiation induced by activated Gαs. Together, these data suggest that activated Gαs translocates from the plasma membrane and, through interaction with tubulin/microtubules in the cytosol, is important for neurite formation, development, and outgrowth. Characterization of neuronal G protein dynamics and their contribution to microtubule dynamics is important for understanding the molecular mechanisms by which G protein-coupled receptor signaling orchestrates neuronal growth and differentiation.

  1. Restoration of DNA-Binding and Growth-Suppressive Activity of Mutant Forms of p53 Via a PCAF-Mediated Acetylation Pathway

    PubMed Central

    PEREZ, RICARDO E.; KNIGHTS, CHAD D.; SAHU, GEETARAM; CATANIA, JASON; KOLUKULA, VAMSI K.; STOLER, DANIEL; GRAESSMANN, ADOLF; OGRYZKO, VASILY; PISHVAIAN, MICHAEL; ALBANESE, CHRISTOPHER; AVANTAGGIATI, MARIA LAURA

    2013-01-01

    Tumor-derived mutant forms of p53 compromise its DNA binding, transcriptional, and growth regulatory activity in a manner that is dependent upon the cell-type and the type of mutation. Given the high frequency of p53 mutations in human tumors, reactivation of the p53 pathway has been widely proposed as beneficial for cancer therapy. In support of this possibility p53 mutants possess a certain degree of conformational flexibility that allows for re-induction of function by a number of structurally different artificial compounds or by short peptides. This raises the question of whether physiological pathways for p53 mutant reactivation also exist and can be exploited therapeutically. The activity of wild-type p53 is modulated by various acetyl-transferases and deacetylases, but whether acetylation influences signaling by p53 mutant is still unknown. Here, we show that the PCAF acetyl-transferase is down-regulated in tumors harboring p53 mutants, where its re-expression leads to p53 acetylation and to cell death. Furthermore, acetylation restores the DNA-binding ability of p53 mutants in vitro and expression of PCAF, or treatment with deacetylase inhibitors, promotes their binding to p53-regulated promoters and transcriptional activity in vivo. These data suggest that PCAF-mediated acetylation rescues activity of at least a set of p53 mutations. Therefore, we propose that dis-regulation of PCAF activity is a pre-requisite for p53 mutant loss of function and for the oncogenic potential acquired by neoplastic cells expressing these proteins. Our findings offer a new rationale for therapeutic targeting of PCAF activity in tumors harboring oncogenic versions of p53. PMID:20589832

  2. ROLES OF EPIDERMAL GROWTH FACTOR (EGF) AND TRANSFORMING GROWTH FACTOR-ALPHA (TGF-A) IN MEDIATION OF DIOXIN (TCDD)-INDUCED DELAYS IN DEVELOPMENT OF THE MOUSE MAMMARY GLAND

    EPA Science Inventory

    Roles of Epidermal Growth Factor (EGF) and Transforming Growth Factor-alpha (TGF-a) in Mediation of Dioxin (TCDD)-Induced Delays in Development of the Mouse Mammary Gland.
    Suzanne E. Fenton, Barbara Abbott, Lamont Bryant, and Angela Buckalew. U.S. EPA, NHEERL, Reproductive Tox...

  3. Core cell cycle regulatory genes in rice and their expression profiles across the growth zone of the leaf.

    PubMed

    Pettkó-Szandtner, A; Cserháti, M; Barrôco, R M; Hariharan, S; Dudits, D; Beemster, G T S

    2015-11-01

    Rice (Oryza sativa L.) as a model and crop plant with a sequenced genome offers an outstanding experimental system for discovering and functionally analyzing the major cell cycle control elements in a cereal species. In this study, we identified the core cell cycle genes in the rice genome through a hidden Markov model search and multiple alignments supported with the use of short protein sequence probes. In total we present 55 rice putative cell cycle genes with locus identity, chromosomal location, approximate chromosome position and EST accession number. These cell cycle genes include nine cyclin dependent-kinase (CDK) genes, 27 cyclin genes, one CKS gene, two RBR genes, nine E2F/DP/DEL genes, six KRP genes, and one WEE gene. We also provide characteristic protein sequence signatures encoded by CDK and cyclin gene variants. Promoter analysis by the FootPrinter program discovered several motifs in the regulatory region of the core cell cycle genes. As a first step towards functional characterization we performed transcript analysis by RT-PCR to determine gene specific variation in transcript levels along the rice leaves. The meristematic zone of the leaves where cells are actively dividing was identified based on kinematic analysis and flow cytometry. As expected, expression of the majority of cell cycle genes was exclusively associated with the meristematic region. However genes such as different D-type cyclins, DEL1, KRP1/3, and RBR2 were also expressed in leaf segments representing the transition zone in which cells start differentiation.

  4. Model for surfactant-mediated growth of Ge on Pb-covered Si(111) surfaces

    NASA Astrophysics Data System (ADS)

    Bȩben, Janusz; Hwang, Ing-Shouh; Chang, Tien-Chih; Tsong, Tien T.

    2001-01-01

    A model of surfactant-mediated epitaxy is investigated using the kinetic Monte Carlo method. This model assumes that (1) adatom-adatom interaction on the surfactant layer is weakly repulsive for dimers, and (2) a concerted atomic exchange of adatoms with surfactant atoms occurs when adatom clusters above the surfactant layer reach a threshold size. All essential features observed in a recent study of Ge/Pb/Si(111) reported by Hwang, Chang, and Tsong can be satisfactorily explained with this model.

  5. Increased expression of platelet-derived growth factor associated protein-1 is associated with PDGF-B mediated glioma progression.

    PubMed

    Sharma, Vinay Kumar; Singh, Anand; Srivastava, Sandeep Kumar; Kumar, Vignesh; Gardi, Nilesh Laxman; Nalwa, Aasma; Dinda, Amit Kumar; Chattopadhyay, Parthaprasad; Yadav, Savita

    2016-09-01

    The current treatment therapies available for malignant gliomas are inadequate. There is an urgent need to develop more effective therapies by characterizing the molecular pathogenesis of the disease. Over expression of platelet-derived growth factor (PDGF) ligands and receptors have been reported in malignant gliomas. Platelet-derived growth factor associated protein-1 (PDAP-1) is reported to modulate the mitogenic activity of PDGF ligands, but to date, there is no information concerning its role in PDGF-mediated glioma cell proliferation. This study aimed to characterize the role of PDAP-1 in PDGF-mediated glioma proliferation. The expression of PDAP-1 was observed to be significantly increased (p<0.05) in grade IV glioma tissue and cell lines compared to grade III. siRNA-mediated knockdown of PDAP-1 reduced the expression of PDGF-B and its downstream genes (Akt1/Protein kinase B (PKB) and phosphoinositide-dependent kinase-1 (PDK1) by up to 50%. In PDAP-1 knockdown glioma cells, more than a twofold reduction was also observed in the level of phosphorylated Akt. Interestingly, knockdown of PDAP-1 in combination with PDGF-B antibody inhibited glioma cell proliferation through activation of Caspase 3/7 and 9. We also demonstrate that PDAP-1 co-localizes with PDGF-B in the cytoplasm of glioma cells, and an interaction between both of the proteins was established. Collectively, these findings suggest that the expression of PDAP-1 is associated with disease malignancy, and its inhibition reduced the proliferation of malignant glioma cells through down-regulation of PDGF-B/Akt/PDK1 signaling. Thus, this study establishes PDAP-1 as an effecter of PDGF signaling in glioma cells and suggests that it could also be a promising therapeutic target.

  6. A Cyclin Dependent Kinase Regulatory Subunit (CKS) Gene of Pigeonpea Imparts Abiotic Stress Tolerance and Regulates Plant Growth and Development in Arabidopsis.

    PubMed

    Tamirisa, Srinath; Vudem, Dashavantha R; Khareedu, Venkateswara R

    2017-01-01

    Frequent climatic changes in conjunction with other extreme environmental factors are known to affect growth, development and productivity of diverse crop plants. Pigeonpea, a major grain legume of the semiarid tropics, endowed with an excellent deep-root system, is known as one of the important drought tolerant crop plants. Cyclin dependent kinases (CDKs) are core cell cycle regulators and play important role in different aspects of plant growth and development. The cyclin-dependent kinase regulatory subunit gene (CKS) was isolated from the cDNA library of pigeonpea plants subjected to drought stress. Pigeonpea CKS (CcCKS) gene expression was detected in both the root and leaf tissues of pigeonpea and was upregulated by polyethylene glycol (PEG), mannitol, NaCl and abscisic acid (ABA) treatments. The overexpression of CcCKS gene in Arabidopsis significantly enhanced tolerance of transgenics to drought and salt stresses as evidenced by different physiological parameters. Under stress conditions, transgenics showed higher biomass, decreased rate of water loss, decreased MDA levels, higher free proline contents, and glutathione levels. Moreover, under stress conditions transgenics exhibited lower stomatal conductance, lower transpiration, and higher photosynthetic rates. However, under normal conditions, CcCKS-transgenics displayed decreased plant growth rate, increased cell size and decreased stomatal number compared to those of wild-type plants. Real-time polymerase chain reaction revealed that CcCKS could regulate the expression of both ABA-dependent and ABA-independent genes associated with abiotic stress tolerance as well as plant growth and development. As such, the CcCKS seems promising and might serve as a potential candidate gene for enhancing the abiotic stress tolerance of crop plants.

  7. A Cyclin Dependent Kinase Regulatory Subunit (CKS) Gene of Pigeonpea Imparts Abiotic Stress Tolerance and Regulates Plant Growth and Development in Arabidopsis

    PubMed Central

    Tamirisa, Srinath; Vudem, Dashavantha R.; Khareedu, Venkateswara R.

    2017-01-01

    Frequent climatic changes in conjunction with other extreme environmental factors are known to affect growth, development and productivity of diverse crop plants. Pigeonpea, a major grain legume of the semiarid tropics, endowed with an excellent deep-root system, is known as one of the important drought tolerant crop plants. Cyclin dependent kinases (CDKs) are core cell cycle regulators and play important role in different aspects of plant growth and development. The cyclin-dependent kinase regulatory subunit gene (CKS) was isolated from the cDNA library of pigeonpea plants subjected to drought stress. Pigeonpea CKS (CcCKS) gene expression was detected in both the root and leaf tissues of pigeonpea and was upregulated by polyethylene glycol (PEG), mannitol, NaCl and abscisic acid (ABA) treatments. The overexpression of CcCKS gene in Arabidopsis significantly enhanced tolerance of transgenics to drought and salt stresses as evidenced by different physiological parameters. Under stress conditions, transgenics showed higher biomass, decreased rate of water loss, decreased MDA levels, higher free proline contents, and glutathione levels. Moreover, under stress conditions transgenics exhibited lower stomatal conductance, lower transpiration, and higher photosynthetic rates. However, under normal conditions, CcCKS-transgenics displayed decreased plant growth rate, increased cell size and decreased stomatal number compared to those of wild-type plants. Real-time polymerase chain reaction revealed that CcCKS could regulate the expression of both ABA-dependent and ABA-independent genes associated with abiotic stress tolerance as well as plant growth and development. As such, the CcCKS seems promising and might serve as a potential candidate gene for enhancing the abiotic stress tolerance of crop plants. PMID:28239388

  8. Antheraea pernyi silk sericin mediating biomimetic nucleation and growth of hydroxylapatite crystals promoting bone matrix formation.

    PubMed

    Jiayao, Zhuang; Guanshan, Zhou; Jinchi, Zhang; Yuyin, Chen; Yongqiang, Zhu

    2017-03-01

    Bone biomineralization is well-regulated processes mediated by extracellular matrix proteins. The materials that can direct nucleation of hydroxylapatite (HAp) crystals and assembly of well-structured material-minerals complex are the key to mimicking the natural mineralization. This study used sericin from Antheraea pernyi (A.pernyi), non-mulberry silkworm cocoon as template to mediate nucleation of HAp crystals. Here we find out that AS (Antheraea pernyi sericin) can nucleate the formation HAp crystals in simulated body fluid verified by XRD and FTIR observations. The HAp crystals are organized into nano-rods oriented with c-axis preferentially parallel to the long axis of AS due to hydrogen bonds and electrostatic interaction and finally aggregated into HAp globule. The cell culture of human bone marrow-derived mesenchymal stem cells (BMSCs) showed that the HAp crystals mediated by AS not only stimulate cell adhesion and proliferation but also promote 0f osteogenic differentiation, suggesting that the resultant mineralized AS biomaterial has potential in promoting bone formation. Thus our work will provide significant implication on biomineralization of A. pernyi silk sericin as a potential scaffold for tissue engineering.

  9. S100A11, an Dual Mediator for Growth Regulation of Human Keratinocytes

    PubMed Central

    Sakaguchi, Masakiyo; Sonegawa, Hiroyuki; Murata, Hitoshi; Kitazoe, Midori; Futami, Jun-ichiro; Kataoka, Ken; Yamada, Hidenori

    2008-01-01

    We previously revealed a novel signal pathway involving S100A11 for inhibition of the growth of normal human keratinocytes (NHK) caused by high Ca++ or transforming growth factor β. Exposure to either agent resulted in transfer of S100A11 to nuclei, where it induced p21WAF1. In contrast, S100A11 has been shown to be overexpressed in many human cancers. To address this apparent discrepancy, we analyzed possible new functions of S100A11, and we provide herein evidence that 1) S100A11 is actively secreted by NHK; 2) extracellular S100A11 acts on NHK to enhance the production of epidermal growth factor family proteins, resulting in growth stimulation; 3) receptor for advanced glycation end products, nuclear factor-κB, Akt, and cAMP response element-binding protein are involved in the S100A11-triggered signal transduction; and 4) production and secretion of S100A11 are markedly enhanced in human squamous cancer cells. These findings indicate that S100A11 plays a dual role in growth regulation of epithelial cells. PMID:17978094

  10. The metalloproteinase stromelysin-1 (transin) mediates PC12 cell growth cone invasiveness through basal laminae.

    PubMed

    Nordstrom, L A; Lochner, J; Yeung, W; Ciment, G

    1995-02-01

    Matrix metalloproteinases have been implicated in various extracellular matrix remodeling events that occur during normal development and in a number of pathologies. In previous work with PC12 rat pheochromocytoma cells, we found that the matrix metalloproteinase stromelysin-1 (ST1) was highly induced by nerve growth factor (NGF), but not by epidermal growth factor (EGF). Here, we show that ST1 immunoreactivity is present in growth cones of NGF-treated PC12 cells, but not EGF-treated or untreated cells. To determine whether ST1 expression confers neurite invasiveness, three lines of PC12 cells were produced that constitutively express ST1 antisense mRNA. These lines expressed and secreted significantly reduced levels of ST1 protein, as determined by immunoblot and immunocytochemical methods, but otherwise responded normally to NGF-treatment by elaborating neurites. We found, however, that the neurites of these ST1 antisense cells showed a significantly reduced ability to penetrate a Matrigel reconstituted basal lamina, as compared to the parental cells, suggesting that ST1 confers neurite invasiveness. Finally, we show that ST1 is also expressed in vivo in sections through Embryonic Day 15 rat embryos, including neurons of both the peripheral and central nervous systems. These data indicate that ST1 may play a role in axonal growth in vivo, including a role in growth cone invasiveness.

  11. Nerve growth factor delivery by ultrasound-mediated nanobubble destruction as a treatment for acute spinal cord injury in rats

    PubMed Central

    Song, Zhaojun; Wang, Zhigang; Shen, Jieliang; Xu, Shengxi; Hu, Zhenming

    2017-01-01

    Background Spinal cord injuries (SCIs) can cause severe disability or death. Treatment options include surgical intervention, drug therapy, and stem cell transplantation. However, the efficacy of these methods for functional recovery remains unsatisfactory. Purpose This study was conducted to explore the effect of ultrasound (US)-mediated destruction of poly(lactic-co-glycolic acid) (PLGA) nanobubbles (NBs) expressing nerve growth factor (NGF) (NGF/PLGA NBs) on nerve regeneration in rats following SCI. Materials and methods Adult male Sprague Dawley rats were randomly divided into four treatment groups after Allen hit models of SCI were established. The groups were normal saline (NS) group, NGF and NBs group, NGF and US group, and NGF/PLGA NBs and US group. Histological changes after SCI were observed by hematoxylin and eosin staining. Neuron viability was determined by Nissl staining. Terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling staining was used to examine cell apoptosis. NGF gene and protein expressions were detected by quantitative reverse transcription polymerase chain reaction and Western blotting. Green fluorescent protein expression in the spinal cord was examined using an inverted fluorescence microscope. The recovery of neural function was determined using the Basso, Beattie, and Bresnahan test. Results NGF therapy using US-mediated NGF/PLGA NBs destruction significantly increased NGF expression, attenuated histological injury, decreased neuron loss, inhibited neuronal apoptosis in injured spinal cords, and increased BBB scores in rats with SCI. Conclusion US-mediated NGF/PLGA NBs destruction effectively transfects the NGF gene into target tissues and has a significant effect on the injured spinal cord. The combination of US irradiation and gene therapy through NGF/PLGA NBs holds great promise for the future of nanomedicine and the development of noninvasive treatment options for SCI and other diseases. PMID:28280337

  12. Constitutive NF-κB activation and tumor-growth promotion by Romo1-mediated reactive oxygen species production

    SciTech Connect

    Chung, Jin Sil; Lee, Sora; Yoo, Young Do

    2014-08-08

    Highlights: • Romo1 expression is required for constitutive nuclear DNA-binding activity of NF-κB. • Romo1 depletion suppresses tumor growth in vivo. • Romo1 presents a potential therapeutic target for diseases. - Abstract: Deregulation of nuclear factor-κB (NF-κB) and related pathways contribute to tumor cell proliferation and invasion. Mechanisms for constitutive NF-κB activation are not fully explained; however, the underlying defects appear to generate and maintain pro-oxidative conditions. In hepatocellular carcinoma (HCC) tissues, up-regulation of reactive oxygen species modulator 1 (Romo1) correlates positively with tumor size. In the present study, we showed that Romo1 expression is required to maintain constitutive nuclear DNA-binding activity of NF-κB and transcriptional activity through constitutive IκBα phosphorylation. Overexpression of Romo1 promoted p65 nuclear translocation and DNA-binding activity. We also show that Romo1 depletion suppressed anchorage-independent colony formation by HCC cells and suppressed tumor growth in vivo. Based on these findings, Romo1 may be a principal regulatory factor in the maintenance of constitutive NF-κB activation in tumor cells. In the interest of anti-proliferative treatments for cancer, Romo1 may also present a productive target for drug development.

  13. Monolayer-Mediated Growth of Organic Semiconductor Films with Improved Device Performance.

    PubMed

    Huang, Lizhen; Hu, Xiaorong; Chi, Lifeng

    2015-09-15

    Increased interest in wearable and smart electronics is driving numerous research works on organic electronics. The control of film growth and patterning is of great importance when targeting high-performance organic semiconductor devices. In this Feature Article, we summarize our recent work focusing on the growth, crystallization, and device operation of organic semiconductors intermediated by ultrathin organic films (in most cases, only a monolayer). The site-selective growth, modified crystallization and morphology, and improved device performance of organic semiconductor films are demonstrated with the help of the inducing layers, including patterned and uniform Langmuir-Blodgett monolayers, crystalline ultrathin organic films, and self-assembled polymer brush films. The introduction of the inducing layers could dramatically change the diffusion of the organic semiconductors on the surface and the interactions between the active layer with the inducing layer, leading to improved aggregation/crystallization behavior and device performance.

  14. Nitrogen-mediated Carbon Nanotube Growth: Diameter reduction, metallicity, bundle dispersability, and bamboo formation

    SciTech Connect

    Sumpter, Bobby G; Meunier, Vincent; Terrones Maldonado, Mauricio; Terrones Maldonado, Humberto; Romo Herrera, Jose M; Cruz Silva, Eduardo; Cullen, David A; Smith, David J

    2007-01-01

    Carbon nanotube growth in the presence of nitrogen has been the subject of much experimental scrutiny, sparking intense debate about the role of nitrogen in the formation of diverse structural features, including shortened length, reduced diameters, and bamboo-like multilayered nanotubules. In this letter, the origin of these features is elucidated using a combination of experimental and theoretical techniques, showing that N acts as a surfactant during growth. N doping enhances the formation of smaller diameter tubes and it can also promote tube closure which includes a relatively large amount of N atoms into the tube lattice, leading to bamboo-like structures. Our findings demonstrate that the mechanism is independent of the tube chirality and suggest a simple procedure for controlling the growth of bamboo-like nanotube morphologies.

  15. Photo-regeneration of severed gel with iniferter-mediated photo-growth.

    PubMed

    Singh, Awaneesh; Kuksenok, Olga; Johnson, Jeremiah A; Balazs, Anna C

    2017-03-08

    Using dissipative particle dynamics (DPD), we developed a computational approach to model the light-induced regeneration of a gel matrix when a significant portion of the material is severed. We considered photo-controlled radical polymerization (photo-CRP) within polymer networks with embedded iniferter groups: the "photo-growth" strategy. Absorption of light by the iniferter groups turns on the polymerization process, which inserts monomers and cross-linkers into the network strands. Photo-growth allows us to effectively regenerate a severed gel under the application of light even when the severed parts are not in direct contact. The growth process can be turned off once the polymerization is near completion to yield a new cross-linked gel that resembles the uncut material.

  16. CD43 promotes cells transformation by preventing merlin-mediated contact inhibition of growth.

    PubMed

    Camacho-Concha, Nohemi; Olivos-Ortiz, Amiel; Nuñez-Rivera, Alfredo; Pedroza-Saavedra, Adolfo; Gutierrez-Xicotencatl, Lourdes; Rosenstein, Yvonne; Pedraza-Alva, Gustavo

    2013-01-01

    In normal tissues, strict control of tissue size is achieved by regulating cell numbers. The mechanism that controls total cell number is known as contact inhibition of growth and it depends on the NF2/Merlin pathway. Negative regulation of this pathway by deleterious mutations or by oncogenes results in cell transformation and tumor progression. Here we provide evidence that the CD43 sialomucin cooperates with oncogenic signals to promote cell transformation by abrogating the contact inhibition of growth through a molecular mechanism that involves AKT-dependent Merlin phosphorylation and degradation. Accordingly, inhibition of endogenous CD43 expression by RNA interference in lung, cervix and colon human cancer cells impaired tumor growth in vivo. These data underscore a previously unidentified role for CD43 in non-hematopoietic tumor progression.

  17. CD43 Promotes Cells Transformation by Preventing Merlin-Mediated Contact Inhibition of Growth

    PubMed Central

    Camacho-Concha, Nohemi; Olivos-Ortiz, Amiel; Nuñez-Rivera, Alfredo; Pedroza-Saavedra, Adolfo; Gutierrez-Xicotencatl, Lourdes; Rosenstein, Yvonne; Pedraza-Alva, Gustavo

    2013-01-01

    In normal tissues, strict control of tissue size is achieved by regulating cell numbers. The mechanism that controls total cell number is known as contact inhibition of growth and it depends on the NF2/Merlin pathway. Negative regulation of this pathway by deleterious mutations or by oncogenes results in cell transformation and tumor progression. Here we provide evidence that the CD43 sialomucin cooperates with oncogenic signals to promote cell transformation by abrogating the contact inhibition of growth through a molecular mechanism that involves AKT-dependent Merlin phosphorylation and degradation. Accordingly, inhibition of endogenous CD43 expression by RNA interference in lung, cervix and colon human cancer cells impaired tumor growth in vivo. These data underscore a previously unidentified role for CD43 in non-hematopoietic tumor progression. PMID:24260485

  18. A novel histone deacetylase inhibitor augments tamoxifen-mediated attenuation of breast carcinoma growth.

    PubMed

    Restall, Christina; Doherty, Judy; Liu, Hong Bin; Genovese, Rosemary; Paiman, Lisa; Byron, Keith A; Anderson, Robin L; Dear, Anthony E

    2009-07-15

    Earlier we generated novel derivatives of the hydroxamate-based histone deacetylase inhibitor (HDACi), Oxamflatin (Ox), which demonstrate considerable HDACi activity. Here the effects of one such derivative, Metacept-1 (MCT-1), alone or in combination with tamoxifen on mammary tumour growth have been assessed in a syngeneic orthotopic model. MCT-1 alone resulted in a trend towards inhibition of growth of 4T1.2 mammary tumours. Since the combination of MCT-1 and tamoxifen up-regulates estrogen receptor expression in 4T1.2 cells in vitro, we tested this combination and found a significant reduction in primary tumour growth over tamoxifen treatment alone. Taken together, these observations suggest that the novel HDACi MCT-1 may warrant further exploration in the treatment of estrogen receptor positive breast carcinoma, particularly when used in combination with conventional agents such as tamoxifen.

  19. Phosphoinositide 3-kinase p110δ mediates estrogen- and FSH-stimulated ovarian follicle growth.

    PubMed

    Li, Qian; He, Hui; Zhang, Yin-Li; Li, Xiao-Meng; Guo, Xuejiang; Huo, Ran; Bi, Ye; Li, Jing; Fan, Heng-Yu; Sha, Jiahao

    2013-09-01

    In the mammalian ovary, primordial follicles are generated early in life and remain dormant for prolonged periods. Their growth resumes via primordial follicle activation, and they continue to grow until the preovulatory stage under the regulation of hormones and growth factors, such as estrogen, FSH, and IGF-1. Both FSH and IGF-1 activate the phosphatidylinositol-3 kinase (PI3K)/Akt (acute transforming retrovirus thymoma protein kinase) signaling pathway in granulosa cells (GCs), yet it remains inconclusive whether the PI3K pathway is crucial for follicle growth. In this study, we investigated the p110δ isoform (encoded by the Pik3cd gene) of PI3K catalytic subunit expression in the mouse ovary and its function in fertility. Pik3cd-null females were subfertile, exhibited fewer growing follicles and more atretic antral follicles in the ovary, and responded poorly to exogenous gonadotropins compared with controls. Ovary transplantation showed that Pik3cd-null ovaries responded poorly to FSH stimulation in vitro; this confirmed that the follicle growth defect was intrinsically ovarian. In addition, estradiol (E2)-stimulated follicle growth and GC proliferation in preantral follicles was impaired in Pik3cd-null ovaries. FSH and E2 substantially activated the PI3K/Akt pathway in GCs of control mice but not in those of Pik3cd-null mice. However, primordial follicle activation and oocyte meiotic maturation were not affected by Pik3cd knockout. Taken together, our findings indicate that the p110δ isoform of the PI3K catalytic subunit is a key component of the PI3K pathway for both FSH and E2-stimulated follicle growth in ovarian GCs; however, it is not required for primordial follicle activation and oocyte development.

  20. Defect-mediated snaking: A new growth mechanism for localized structures

    NASA Astrophysics Data System (ADS)

    Ma, Y.-P.; Burke, J.; Knobloch, E.

    2010-10-01

    Stationary spatially localized patterns in parametrically driven systems are studied, focusing on the 2:1 and 1:1 resonance tongues as described by the forced complex Ginzburg-Landau equation. Homoclinic snaking is identified in both cases and the nature of the growth of the localized structures along the snaking branches is described. The structures grow from a central defect that inserts new rolls on either side, while pushing existing rolls outwards. This growth mechanism differs fundamentally from that found in other systems exhibiting homoclinic snaking in which new rolls are added at the fronts that connect the structure to the background homogeneous state.

  1. Identification of a novel BET bromodomain inhibitor-sensitive, gene regulatory circuit that controls Rituximab response and tumour growth in aggressive lymphoid cancers

    PubMed Central

    Emadali, Anouk; Rousseaux, Sophie; Bruder-Costa, Juliana; Rome, Claire; Duley, Samuel; Hamaidia, Sieme; Betton, Patricia; Debernardi, Alexandra; Leroux, Dominique; Bernay, Benoit; Kieffer-Jaquinod, Sylvie; Combes, Florence; Ferri, Elena; McKenna, Charles E; Petosa, Carlo; Bruley, Christophe; Garin, Jérôme; Ferro, Myriam; Gressin, Rémy; Callanan, Mary B; Khochbin, Saadi

    2013-01-01

    Immuno-chemotherapy elicit high response rates in B-cell non-Hodgkin lymphoma but heterogeneity in response duration is observed, with some patients achieving cure and others showing refractory disease or relapse. Using a transcriptome-powered targeted proteomics screen, we discovered a gene regulatory circuit involving the nuclear factor CYCLON which characterizes aggressive disease and resistance to the anti-CD20 monoclonal antibody, Rituximab, in high-risk B-cell lymphoma. CYCLON knockdown was found to inhibit the aggressivity of MYC-overexpressing tumours in mice and to modulate gene expression programs of biological relevance to lymphoma. Furthermore, CYCLON knockdown increased the sensitivity of human lymphoma B cells to Rituximab in vitro and in vivo. Strikingly, this effect could be mimicked by in vitro treatment of lymphoma B cells with a small molecule inhibitor for BET bromodomain proteins (JQ1). In summary, this work has identified CYCLON as a new MYC cooperating factor that autonomously drives aggressive tumour growth and Rituximab resistance in lymphoma. This resistance mechanism is amenable to next-generation epigenetic therapy by BET bromodomain inhibition, thereby providing a new combination therapy rationale for high-risk lymphoma. The nuclear factor CYCLON is a new MYC cooperating factor that drives tumor growth and Rituximab resistance in lymphoma. This resistance mechanism can be targeted by next-generation epigenetic therapy by BET bromodomain inhibition downstream of MYC. PMID:23828858

  2. Role of Flightless-I (Drosophila) homolog in the transcription activation of type I collagen gene mediated by transforming growth factor beta

    SciTech Connect

    Lim, Mi-Sun; Jeong, Kwang Won

    2014-11-21

    Highlights: • FLII activates TGFβ-mediated expression of COL1A2 gene. • TGFβ induces the association of FLII with SMAD3 and BRG1 in A549 cells. • FLII is required for the recruitment of SWI/SNF complex and chromatin accessibility to COL1A2 promoter. - Abstract: Flightless-I (Drosophila) homolog (FLII) is a nuclear receptor coactivator that is known to interact with other transcriptional regulators such as the SWI/SNF complex, an ATP-dependent chromatin-remodeling complex, at the promoter or enhancer region of estrogen receptor (ER)-α target genes. However, little is known about the role of FLII during transcription initiation in the transforming growth factor beta (TGFβ)/SMAD-dependent signaling pathway. Here, we demonstrate that FLII functions as a coactivator in the expression of type I collagen gene induced by TGFβ in A549 cells. FLII activates the reporter gene driven by COL1A2 promoter in a dose-dependent manner. Co-expression of GRIP1, CARM1, or p300 did not show any synergistic activation of transcription. Furthermore, the level of COL1A2 expression correlated with the endogenous level of FLII mRNA level. Depletion of FLII resulted in a reduction of TGFβ-induced expression of COL1A2 gene. In contrast, over-expression of FLII caused an increase in the endogenous expression of COL1A2. We also showed that FLII is associated with Brahma-related gene 1 (BRG1) as well as SMAD in A549 cells. Notably, the recruitment of BRG1 to the COL1A2 promoter region was decreased in FLII-depleted A549 cells, suggesting that FLII is required for TGFβ-induced chromatin remodeling, which is carried out by the SWI/SNF complex. Furthermore, formaldehyde-assisted isolation of regulatory elements (FAIRE)-quantitative polymerase chain reaction (qPCR) experiments revealed that depletion of FLII caused a reduction in chromatin accessibility at the COL1A2 promoter. These results suggest that FLII plays a critical role in TGFβ/SMAD-mediated transcription of the COL1A2 gene

  3. Plasmodium falciparum infection and age influence parasite growth inhibition mediated by IgG in Beninese infants.

    PubMed

    Adamou, Rafiou; Chénou, Francine; Sadissou, Ibrahim; Sonon, Paulin; Dechavanne, Célia; Djilali-Saïah, Abdelkader; Cottrell, Gilles; Le Port, Agnès; Massougbodji, Achille; Remarque, Edmond J; Luty, Adrian J F; Sanni, Ambaliou; Garcia, André; Migot-Nabias, Florence; Milet, Jacqueline; Courtin, David

    2016-07-01

    Antibodies that impede the invasion of Plasmodium falciparum (P. falciparum) merozoites into erythrocytes play a critical role in anti-malarial immunity. The Growth Inhibition Assay (GIA) is an in vitro measure of the functional capacity of such antibodies to limit erythrocyte invasion and/or parasite growth. Up to now, it is unclear whether growth-inhibitory activity correlates with protection from clinical disease and there are inconsistent results from studies performed with GIA. Studies that have focused on the relationship between IgGs and their in vitro parasite Growth Inhibition Activity (GIAc) in infants aged less than two years old are rare. Here, we used clinical and parasitological data to precisely define symptomatic or asymptomatic infection with P. falciparum in groups of infants followed-up actively for 18 months post-natally. We quantified the levels of IgG1 and IgG3 directed to a panel of candidate P. falciparum vaccine antigens (AMA-1, MSP1, 2, 3 and GLURP) using ELISA and the functional activity of IgG was quantified using GIA. Data were then correlated with individuals' infection status. At 18 months of age, infants harbouring infections at the time of blood sampling had an average 19% less GIAc than those not infected (p=0.004, multivariate linear regression). GIAc decreased from 12 to 18 months of age (p=0.003, Wilcoxon matched pairs test). Antibody levels quantified at 18 months in infants were strongly correlated with their exposure to malarial infection, however GIAc was not correlated with malaria infectious status (asymptomatic and symptomatic groups). In conclusion, both infection status at blood draw and age influence parasite growth inhibition mediated by IgG in the GIA. Both factors must be taken into account when correlations between GIAc and anti-malarial protection or vaccine efficacy have to be made.

  4. Brain-derived neurotrophic factor mediates activity-dependent dendritic growth in nonpyramidal neocortical interneurons in developing organotypic cultures.

    PubMed

    Jin, Xiaoming; Hu, Hang; Mathers, Peter H; Agmon, Ariel

    2003-07-02

    Brain-derived neurotrophic factor (BDNF) promotes postnatal maturation of GABAergic inhibition in the cerebral and cerebellar cortices, and its expression and release are enhanced by neuronal activity, suggesting that it acts in a feedback manner to maintain a balance between excitation and inhibition during development. BDNF promotes differentiation of cerebellar, hippocampal, and neostriatal inhibitory neurons, but its effects on the dendritic development of neocortical inhibitory interneurons remain unknown. Here, we show that BDNF mediates depolarization-induced dendritic growth and branching in neocortical interneurons. To visualize inhibitory interneurons, we biolistically transfected organotypic cortical slice cultures from neonatal mice with green fluorescent protein (GFP) driven by the glutamic acid decarboxylase (GAD)67 promoter. Nearly all GAD67-GFP-expressing neurons were nonpyramidal, many contained GABA, and some expressed markers of neurochemically defined GABAergic subtypes, indicating that GAD67-GFP-expressing neurons were GABAergic. We traced dendritic trees from confocal images of the same GAD67-GFP-expressing neurons before and after a 5 d growth period, and quantified the change in total dendritic length (TDL) and total dendritic branch points (TDBPs) for each neuron. GAD67-GFP-expressing neurons growing in control medium exhibited a 20% increase in TDL, but in 200 ng/ml BDNF or 10 mm KCl, this increase nearly doubled and was accompanied by a significant increase in TDBPs. Blocking action potentials with TTX did not prevent the BDNF-induced growth, but antibodies against BDNF blocked the growth-promoting effect of KCl. We conclude that BDNF, released by neocortical pyramidal neurons in response to depolarization, enhances dendritic growth and branching in nearby inhibitory interneurons.

  5. Wnt/RANKL-mediated bone growth promoting effects of blueberries in weanling rats

    Technology Transfer Automated Retrieval System (TEKTRAN)

    We studied the effects of dietary blueberry supplementation on bone growth in weanling rats. Weanling male and female rats were fed AIN-93G semi-purified diets supplemented with 10% whole blueberry powder for 14 and 30 days beginning on PND 21. In both sexes tibial bone mineral density and content a...

  6. Aridopsis COBRA-LIKE 10, a GPI-anchored protien, mediates directional growth of pollen tubes

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Successful reproduction of flowering plants requires constant communication between female tissues and growing pollen tubes. Female cells secrete molecules and peptides as nutrients or guidance cues for fast and directional tube growth, which is executed by dynamic changes of intracellular activitie...

  7. Detection of neurotransmitters by a light scattering technique based on seed-mediated growth of gold nanoparticles

    NASA Astrophysics Data System (ADS)

    Shang, Li; Dong, Shaojun

    2008-03-01

    A simple light scattering detection method for neurotransmitters has been developed, based on the growth of gold nanoparticles. Neurotransmitters (dopamine, L-dopa, noradrenaline and adrenaline) can effectively function as active reducing agents for generating gold nanoparticles, w