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Sample records for actin transforming growth

  1. Downregulation of the DNA repair enzyme apurinic/apyrimidinic endonuclease 1 stimulates transforming growth factor-β1 production and promotes actin rearrangement.

    PubMed

    Sakai, Yuri; Yamamori, Tohru; Yasui, Hironobu; Inanami, Osamu

    2015-05-22

    The DNA repair enzyme apurinic/apyrimidinic endonuclease 1 (APE1) plays a central role in base excision repair and functions as a reductive activator of various transcription factors. Multiple other functionalities have been ascribed to APE1 in addition to these major functions. A recent study showed that APE1 knockdown upregulated the expression of a set of genes related to extracellular matrix (ECM) production, indicating an additional novel biological role for this enzyme. Based on this finding, we have investigated the effect of APE1 downregulation on ECM-related gene expression and its biological consequences. Endogenous APE1 expression was downregulated in human cervical carcinoma HeLa cells and human lung carcinoma A549 cells using siRNA. When the expression of six ECM-related genes (TGFB1, LAMC1, FN1, COL1A1, COL3A1, and COL4A1) was evaluated, we found that APE1 knockdown upregulated the expression of TGFB1 in both cell lines. APE1 downregulation promoted actin rearrangement, inducing F-actin accumulation in HeLa cells and the dissipation of stress fibers in A549 cells. We also discovered that APE1 knockdown enhanced cellular motility in A549 cells, which was suppressed by the inhibition of transforming growth factor (TGF)-β1 signaling. These results suggested that APE1 controls the organization of actin cytoskeleton through the regulation of TGF-β1 expression, providing novel insights into the biological significance of APE1. PMID:25858321

  2. Transformation of actin-encapsulating liposomes induced by cytochalasin D.

    PubMed Central

    Miyata, H; Kinosita, K

    1994-01-01

    Liposomes encapsulating actin filaments were prepared by swelling at 0 degrees C lipid film consisting of a mixture of dimyristoyl phosphatidylcholine and cardiolipin (equal amounts by weight) in 100 microM rabbit skeletal muscle actin and 0.5 mM CaCl2 followed by polymerization of actin at 30 degrees C. Liposomes initially assumed either disk or dumbbell shape, but when cytochalasin D was added to the medium surrounding the liposomes, they were found to become spindle shaped. Liposomes containing bovine serum albumin that were given cytochalasin D and actin-containing liposomes that were given dimethylformamide, the solvent for cytochalasin D, did not transform. These results indicated actin-cytochalasin interaction is involved in the transformation process. Falling-ball viscometry and sedimentation analysis of actin solution indicated that cytochalasin cleaved actin filaments and caused depolymerization. The observation of polarized fluorescence of encapsulated actin labeled with acrylodan indicated that the actin filaments in the transformed liposomes aligned along the long axis of the liposomes. Because the actin filaments in the disk- or dumbbell-shaped liposomes formed bundles running along the liposome contour, the transformation was likely to be accompanied by the change in the actin filament arrangement in the liposomes, which was induced by actin-cytochalasin interaction. Images FIGURE 1 FIGURE 2 FIGURE 3 PMID:7948706

  3. Actin Dynamics in Growth Cone Motility and Navigation

    PubMed Central

    Gomez, Timothy M.; Letourneau, Paul C.

    2014-01-01

    Motile growth cones lead growing axons through developing tissues to synaptic targets. These behaviors depend on the organization and dynamics of actin filaments that fill the growth cone leading margin (peripheral (P-) domain). Actin filament organization in growth cones is regulated by actin-binding proteins that control all aspects of filament assembly, turnover, interactions with other filaments and cytoplasmic components, and participation in producing mechanical forces. Actin filament polymerization drives protrusion of sensory filopodia and lamellipodia, and actin filament connections to the plasma membrane link the filament network to adhesive contacts of filopodia and lamellipodia with other surfaces. These contacts stabilize protrusions and transduce mechanical forces generated by actomyosin activity into traction that pulls an elongating axon along the path towards its target. Adhesive ligands and extrinsic guidance cues bind growth cone receptors and trigger signaling activities involving Rho GTPases, kinases, phosphatases, cyclic nucleotides and [Ca++] fluxes. These signals regulate actin binding proteins to locally modulate actin polymerization, interactions and force transduction to steer the growth cone leading margin towards the sources of attractive cues and away from repellent guidance cues. PMID:24164353

  4. ROP Gtpase–Dependent Dynamics of Tip-Localized F-Actin Controls Tip Growth in Pollen Tubes

    PubMed Central

    Fu, Ying; Wu, Guang; Yang, Zhenbiao

    2001-01-01

    Tip-growing pollen tubes provide a useful model system to study polar growth. Although roles for tip-focused calcium gradient and tip-localized Rho-family GTPase in pollen tube growth is established, the existence and function of tip-localized F-actin have been controversial. Using the green fluorescent protein–tagged actin-binding domain of mouse talin, we found a dynamic form of tip-localized F-actin in tobacco pollen tubes, termed short actin bundles (SABs). The dynamics of SABs during polar growth in pollen tubes is regulated by Rop1At, a Rop GTPase belonging to the Rho family. When overexpressed, Rop1At transformed SAB into a network of fine filaments and induced a transverse actin band behind the tip, leading to depolarized growth. These changes were due to ectopic Rop1At localization to the apical region of the plasma membrane and were suppressed by guanine dissociation inhibitor overexpression, which removed ectopically localized Rop1At. Rop GTPase–activating protein (RopGAP1) overexpression, or Latrunculin B treatments, also recovered normal actin organization and tip growth in Rop1At-overexpressing tubes. Moreover, overexpression of RopGAP1 alone disrupted SABs and inhibited growth. Finally, SAB oscillates and appears at the tip before growth. Together, these results indicate that the dynamics of tip actin are essential for tip growth and provide the first direct evidence to link Rho GTPase to actin organization in controlling cell polarity and polar growth in plants. PMID:11238457

  5. Bacterial actin and tubulin homologs in cell growth and division.

    PubMed

    Busiek, Kimberly K; Margolin, William

    2015-03-16

    In contrast to the elaborate cytoskeletal machines harbored by eukaryotic cells, such as mitotic spindles, cytoskeletal structures detectable by typical negative stain electron microscopy are generally absent from bacterial cells. As a result, for decades it was thought that bacteria lacked cytoskeletal machines. Revolutions in genomics and fluorescence microscopy have confirmed the existence not only of smaller-scale cytoskeletal structures in bacteria, but also of widespread functional homologs of eukaryotic cytoskeletal proteins. The presence of actin, tubulin, and intermediate filament homologs in these relatively simple cells suggests that primitive cytoskeletons first arose in bacteria. In bacteria such as Escherichia coli, homologs of tubulin and actin directly interact with each other and are crucial for coordinating cell growth and division. The function and direct interactions between these proteins will be the focus of this review. PMID:25784047

  6. Dynamics of Actin Cables in Polarized Growth of the Filamentous Fungus Aspergillus nidulans

    PubMed Central

    Bergs, Anna; Ishitsuka, Yuji; Evangelinos, Minoas; Nienhaus, G. U.; Takeshita, Norio

    2016-01-01

    Highly polarized growth of filamentous fungi requires a continuous supply of proteins and lipids to the hyphal tip. This transport is managed by vesicle trafficking via the actin and microtubule cytoskeletons and their associated motor proteins. Particularly, actin cables originating from the hyphal tip are essential for hyphal growth. Although, specific marker proteins have been developed to visualize actin cables in filamentous fungi, the exact organization and dynamics of actin cables has remained elusive. Here, we observed actin cables using tropomyosin (TpmA) and Lifeact fused to fluorescent proteins in living Aspergillus nidulans hyphae and studied the dynamics and regulation. GFP tagged TpmA visualized dynamic actin cables formed from the hyphal tip with cycles of elongation and shrinkage. The elongation and shrinkage rates of actin cables were similar and approximately 0.6 μm/s. Comparison of actin markers revealed that high concentrations of Lifeact reduced actin dynamics. Simultaneous visualization of actin cables and microtubules suggests temporally and spatially coordinated polymerization and depolymerization between the two cytoskeletons. Our results provide new insights into the molecular mechanism of ordered polarized growth regulated by actin cables and microtubules. PMID:27242709

  7. Dynamics of Actin Cables in Polarized Growth of the Filamentous Fungus Aspergillus nidulans.

    PubMed

    Bergs, Anna; Ishitsuka, Yuji; Evangelinos, Minoas; Nienhaus, G U; Takeshita, Norio

    2016-01-01

    Highly polarized growth of filamentous fungi requires a continuous supply of proteins and lipids to the hyphal tip. This transport is managed by vesicle trafficking via the actin and microtubule cytoskeletons and their associated motor proteins. Particularly, actin cables originating from the hyphal tip are essential for hyphal growth. Although, specific marker proteins have been developed to visualize actin cables in filamentous fungi, the exact organization and dynamics of actin cables has remained elusive. Here, we observed actin cables using tropomyosin (TpmA) and Lifeact fused to fluorescent proteins in living Aspergillus nidulans hyphae and studied the dynamics and regulation. GFP tagged TpmA visualized dynamic actin cables formed from the hyphal tip with cycles of elongation and shrinkage. The elongation and shrinkage rates of actin cables were similar and approximately 0.6 μm/s. Comparison of actin markers revealed that high concentrations of Lifeact reduced actin dynamics. Simultaneous visualization of actin cables and microtubules suggests temporally and spatially coordinated polymerization and depolymerization between the two cytoskeletons. Our results provide new insights into the molecular mechanism of ordered polarized growth regulated by actin cables and microtubules. PMID:27242709

  8. Arabidopsis RIC1 Severs Actin Filaments at the Apex to Regulate Pollen Tube Growth

    PubMed Central

    Zhou, Zhenzhen; Shi, Haifan; Chen, Binqing; Zhang, Ruihui; Huang, Shanjin; Fu, Ying

    2015-01-01

    Pollen tubes deliver sperms to the ovule for fertilization via tip growth. The rapid turnover of F-actin in pollen tube tips plays an important role in this process. In this study, we demonstrate that Arabidopsis thaliana RIC1, a member of the ROP-interactive CRIB motif-containing protein family, regulates pollen tube growth via its F-actin severing activity. Knockout of RIC1 enhanced pollen tube elongation, while overexpression of RIC1 dramatically reduced tube growth. Pharmacological analysis indicated that RIC1 affected F-actin dynamics in pollen tubes. In vitro biochemical assays revealed that RIC1 directly bound and severed F-actin in the presence of Ca2+ in addition to interfering with F-actin turnover by capping F-actin at the barbed ends. In vivo, RIC1 localized primarily to the apical plasma membrane (PM) of pollen tubes. The level of RIC1 at the apical PM oscillated during pollen tube growth. The frequency of F-actin severing at the apex was notably decreased in ric1-1 pollen tubes but was increased in pollen tubes overexpressing RIC1. We propose that RIC1 regulates F-actin dynamics at the apical PM as well as the cytosol by severing F-actin and capping the barbed ends in the cytoplasm, establishing a novel mechanism that underlies the regulation of pollen tube growth. PMID:25804540

  9. Farnesyltransferase inhibition causes morphological reversion of ras-transformed cells by a complex mechanism that involves regulation of the actin cytoskeleton.

    PubMed Central

    Prendergast, G C; Davide, J P; deSolms, S J; Giuliani, E A; Graham, S L; Gibbs, J B; Oliff, A; Kohl, N E

    1994-01-01

    A potent and specific small molecule inhibitor of farnesyl-protein transferase, L-739,749, caused rapid morphological reversion and growth inhibition of ras-transformed fibroblasts (Rat1/ras cells). Morphological reversion occurred within 18 h of L-739,749 addition. The reverted phenotype was stable for several days in the absence of inhibitor before the transformed phenotype reappeared. Cell enlargement and actin stress fiber formation accompanied treatment of both Rat1/ras and normal Rat1 cells. Significantly, inhibition of Ras processing did not correlate with the initiation or maintenance of the reverted phenotype. While a single treatment with L-739,749 was sufficient to morphologically revert Rat1/ras cells, repetitive inhibitor treatment was required to significantly reduce cell growth rate. Thus, the effects of L-739,749 on transformed cell morphology and cytoskeletal actin organization could be separated from effects on cell growth, depending on whether exposure to a farnesyl-protein transferase inhibitor was transient or repetitive. In contrast, L-739,749 had no effect on the growth, morphology, or actin organization of v-raf-transformed cells. Taken together, the results suggest that the mechanism of morphological reversion is complex and may involve farnesylated proteins that control the organization of cytoskeletal actin. Images PMID:8196657

  10. Actinic keratosis

    MedlinePlus

    Solar keratosis; Sun-induced skin changes - keratosis; Keratosis - actinic (solar) ... Some actinic keratoses become squamous cell skin cancer . Have your health care provider look at all skin growths as soon as you find them. Your provider will ...

  11. Anillin Regulates Neuronal Migration and Neurite Growth by Linking RhoG to the Actin Cytoskeleton.

    PubMed

    Tian, Dong; Diao, Min; Jiang, Yuxiang; Sun, Lingfei; Zhang, Yan; Chen, Zhucheng; Huang, Shanjin; Ou, Guangshuo

    2015-05-01

    Neuronal migration and neurite growth are essential events in neural development, but it remains unclear how guidance cues are transduced through receptors to the actin cytoskeleton, which powers these processes. We report that a cytokinetic scaffold protein, Anillin, is redistributed to the leading edge of the C. elegans Q neuroblast during cell migration and neurite growth. To bypass the requirement for Anillin in cytokinesis, we used the somatic CRISPR-Cas9 technique to generate conditional mutations in Anillin. We demonstrate that Anillin regulates cell migration and growth cone extension by stabilizing the F-actin network at the leading edge. Our biochemical analysis shows that the actin-binding domain of Anillin is sufficient to stabilize F-actin by antagonizing the F-actin severing activity of Cofilin. We further uncover that the active form of RhoG/MIG-2 directly binds to Anillin and recruits it to the leading edge. Our results reveal a novel pathway in which Anillin transduces the RhoG signal to the actin cytoskeleton during neuronal migration and neurite growth. PMID:25843030

  12. Actin-dependent vacuolar occupancy of the cell determines auxin-induced growth repression

    PubMed Central

    Scheuring, David; Löfke, Christian; Krüger, Falco; Kittelmann, Maike; Eisa, Ahmed; Hughes, Louise; Smith, Richard S.; Hawes, Chris; Schumacher, Karin; Kleine-Vehn, Jürgen

    2016-01-01

    The cytoskeleton is an early attribute of cellular life, and its main components are composed of conserved proteins. The actin cytoskeleton has a direct impact on the control of cell size in animal cells, but its mechanistic contribution to cellular growth in plants remains largely elusive. Here, we reveal a role of actin in regulating cell size in plants. The actin cytoskeleton shows proximity to vacuoles, and the phytohormone auxin not only controls the organization of actin filaments but also impacts vacuolar morphogenesis in an actin-dependent manner. Pharmacological and genetic interference with the actin–myosin system abolishes the effect of auxin on vacuoles and thus disrupts its negative influence on cellular growth. SEM-based 3D nanometer-resolution imaging of the vacuoles revealed that auxin controls the constriction and luminal size of the vacuole. We show that this actin-dependent mechanism controls the relative vacuolar occupancy of the cell, thus suggesting an unanticipated mechanism for cytosol homeostasis during cellular growth. PMID:26715743

  13. Arabidopsis FIMBRIN5, an Actin Bundling Factor, Is Required for Pollen Germination and Pollen Tube Growth[W

    PubMed Central

    Wu, Youjun; Yan, Jin; Zhang, Ruihui; Qu, Xiaolu; Ren, Sulin; Chen, Naizhi; Huang, Shanjin

    2010-01-01

    Actin cables in pollen tubes serve as molecular tracks for cytoplasmic streaming and organelle movement and are formed by actin bundling factors like villins and fimbrins. However, the precise mechanisms by which actin cables are generated and maintained remain largely unknown. Fimbrins comprise a family of five members in Arabidopsis thaliana. Here, we characterized a fimbrin isoform, Arabidopsis FIMBRIN5 (FIM5). Our results show that FIM5 is required for the organization of actin cytoskeleton in pollen grains and pollen tubes, and FIM5 loss-of-function associates with a delay of pollen germination and inhibition of pollen tube growth. FIM5 decorates actin filaments throughout pollen grains and tubes. Actin filaments become redistributed in fim5 pollen grains and disorganized in fim5 pollen tubes. Specifically, actin cables protrude into the extreme tips, and their longitudinal arrangement is disrupted in the shank of fim5 pollen tubes. Consequently, the pattern and velocity of cytoplasmic streaming were altered in fim5 pollen tubes. Additionally, loss of FIM5 function rendered pollen germination and tube growth hypersensitive to the actin-depolymerizing drug latrunculin B. In vitro biochemical analyses indicated that FIM5 exhibits actin bundling activity and stabilizes actin filaments. Thus, we propose that FIM5 regulates actin dynamics and organization during pollen germination and tube growth via stabilizing actin filaments and organizing them into higher-order structures. PMID:21098731

  14. Spontaneous polarization in an interfacial growth model for actin filament networks with a rigorous mechanochemical coupling.

    PubMed

    John, Karin; Caillerie, Denis; Misbah, Chaouqi

    2014-11-01

    Many processes in eukaryotic cells, including cell motility, rely on the growth of branched actin networks from surfaces. Despite its central role the mechanochemical coupling mechanisms that guide the growth process are poorly understood, and a general continuum description combining growth and mechanics is lacking. We develop a theory that bridges the gap between mesoscale and continuum limit and propose a general framework providing the evolution law of actin networks growing under stress. This formulation opens an area for the systematic study of actin dynamics in arbitrary geometries. Our framework predicts a morphological instability of actin growth on a rigid sphere, leading to a spontaneous polarization of the network with a mode selection corresponding to a comet, as reported experimentally. We show that the mechanics of the contact between the network and the surface plays a crucial role, in that it determines directly the existence of the instability. We extract scaling laws relating growth dynamics and network properties offering basic perspectives for new experiments on growing actin networks. PMID:25493815

  15. Actin filament turnover drives leading edge growth during myelin sheath formation in the central nervous system

    PubMed Central

    Schmitt, Sebastian; Snaidero, Nicolas; Mitkovski, Mišo; Velte, Caroline; Brückner, Bastian R.; Alexopoulos, Ioannis; Czopka, Tim; Jung, Sang Y.; Rhee, Jeong S.; Janshoff, Andreas; Witke, Walter; Schaap, Iwan A.T.; Lyons, David A.; Simons, Mikael

    2016-01-01

    Summary During central nervous system development, oligodendrocytes wrap their plasma membrane around axons to generate multi-lamellar myelin sheaths. To drive growth at the leading edge of myelin at the interface with the axon, mechanical forces are necessary, but the underlying mechanisms are not known. Using an interdisciplinary approach that combines morphological, genetic and biophysical analyses, we identified a key role for actin filament network turnover in myelin growth. At the onset of myelin biogenesis, F-actin is redistributed to the leading edge, where its polymerization-based forces push out non-adhesive and motile protrusions. F-actin disassembly converts protrusions into sheets by reducing surface tension and in turn inducing membrane spreading and adhesion. We identified the actin depolymerizing factor ADF/Cofilin1, which mediates high F-actin turnover rates, as essential factor in this process. We propose that F-actin turnover is the driving force in myelin wrapping by regulating repetitive cycles of leading edge protrusion and spreading. PMID:26166299

  16. Interdependence of endomembrane trafficking and actin dynamics during polarized growth of Arabidopsis pollen tubes

    Technology Transfer Automated Retrieval System (TEKTRAN)

    During polarized growth of pollen tubes, endomembrane trafficking and actin polymerization are two critical processes that establish membrane/wall homeostasis and maintain growth polarity. Fine-tuned interactions between these two processes are therefore necessary but poorly understood. To better un...

  17. Nucleation and growth transformation kinetics

    NASA Astrophysics Data System (ADS)

    Erukhimovitch, V.; Baram, J.

    1995-03-01

    As a result of the reassessment of the Kolmogorov-Johnson-Mehl-Avrami (KJMA) theory for the kinetics of nucleation and growth transformations, an integral-equation formulation has been developed instead of the well-known and widely used Avrami equation. The presented formulation considers interfacial and diffusional growths, in one, two, and three dimensions, with both time-dependent and time-invariant nucleation and growth rates. The integral-equation model corrects reported inadequacies of the KJMA theory when applied in numerous experiments and various solid-state transformations. It is shown that in the example cases examined in this paper, crystallization from the amorphous state in melt-spun ribbons, isothermal aging of CuAlZn, pearlitic transition in an eutectoid steel, and crystallization in a PEKK polymer, the thermodynamic and kinetic interpretation and parameters extracted from best fits of the Avrami equations to the experimental data are erroneous. The KJMA formulation is a simplification of the real physical conditions. The main limitation of the new model is that almost all the integral equations representing the kinetics of solid-state transformations have no analytical solutions.

  18. Natural transformation occurs independently of the essential actin-like MreB cytoskeleton in Legionella pneumophila.

    PubMed

    Juan, Pierre-Alexandre; Attaiech, Laetitia; Charpentier, Xavier

    2015-01-01

    Natural transformation is the process by which bacteria can actively take up and integrate exogenous DNA thereby providing a source of genetic diversity. Under specific growth conditions the coordinated expression of several genes--a situation referred to as "competence"--allows bacteria to assemble a highly processive and dedicated system that can import high molecular weight DNA. Within the cell these large imported DNA molecules are protected from degradation and brought to the chromosome for recombination. Here, we report elevated expression of mreB during competence in the Gram-negative pathogen Legionella pneumophila. Interestingly a similar observation had previously been reported in the distantly-related Gram-positive organism Bacillus subtilis. MreB is often viewed as the bacterial actin homolog contributing to bacterial morphogenesis by coordinating peptidoglycan-synthesising complexes. In addition MreB is increasingly found to be involved in a growing number of processes including chromosome segregation and motor-driven motility. Using genetic and pharmacological approaches, we examined the possible role of MreB during natural transformation in L. pneumophila. Our data show that natural transformation does not require MreB dynamics and exclude a direct role of MreB filaments in the transport of foreign DNA and its recombination in the chromosome. PMID:26526572

  19. Natural transformation occurs independently of the essential actin-like MreB cytoskeleton in Legionella pneumophila

    PubMed Central

    Juan, Pierre-Alexandre; Attaiech, Laetitia; Charpentier, Xavier

    2015-01-01

    Natural transformation is the process by which bacteria can actively take up and integrate exogenous DNA thereby providing a source of genetic diversity. Under specific growth conditions the coordinated expression of several genes – a situation referred to as “competence” – allows bacteria to assemble a highly processive and dedicated system that can import high molecular weight DNA. Within the cell these large imported DNA molecules are protected from degradation and brought to the chromosome for recombination. Here, we report elevated expression of mreB during competence in the Gram-negative pathogen Legionella pneumophila. Interestingly a similar observation had previously been reported in the distantly-related Gram-positive organism Bacillus subtilis. MreB is often viewed as the bacterial actin homolog contributing to bacterial morphogenesis by coordinating peptidoglycan-synthesising complexes. In addition MreB is increasingly found to be involved in a growing number of processes including chromosome segregation and motor-driven motility. Using genetic and pharmacological approaches, we examined the possible role of MreB during natural transformation in L. pneumophila. Our data show that natural transformation does not require MreB dynamics and exclude a direct role of MreB filaments in the transport of foreign DNA and its recombination in the chromosome. PMID:26526572

  20. Arabidopsis Microtubule-Destabilizing Protein 25 Functions in Pollen Tube Growth by Severing Actin Filaments[W

    PubMed Central

    Qin, Tao; Liu, Xiaomin; Li, Jiejie; Sun, Jingbo; Song, Leina; Mao, Tonglin

    2014-01-01

    The formation of distinct actin filament arrays in the subapical region of pollen tubes is crucial for pollen tube growth. However, the molecular mechanisms underlying the organization and dynamics of the actin filaments in this region remain to be determined. This study shows that Arabidopsis thaliana MICROTUBULE-DESTABILIZING PROTEIN25 (MDP25) has the actin filament–severing activity of an actin binding protein. This protein negatively regulated pollen tube growth by modulating the organization and dynamics of actin filaments in the subapical region of pollen tubes. MDP25 loss of function resulted in enhanced pollen tube elongation and inefficient fertilization. MDP25 bound directly to actin filaments and severed individual actin filaments, in a manner that was dramatically enhanced by Ca2+, in vitro. Analysis of a mutant that bears a point mutation at the Ca2+ binding sites demonstrated that the subcellular localization of MDP25 was determined by cytosolic Ca2+ level in the subapical region of pollen tubes, where MDP25 was disassociated from the plasma membrane and moved into the cytosol. Time-lapse analysis showed that the F-actin-severing frequency significantly decreased and a high density of actin filaments was observed in the subapical region of mdp25-1 pollen tubes. This study reveals a mechanism whereby calcium enhances the actin filament–severing activity of MDP25 in the subapical region of pollen tubes to modulate pollen tube growth. PMID:24424096

  1. Cytoarchitecture of Kirsten sarcoma virus-transformed rat kidney fibroblasts: butyrate-induced reorganization within the actin microfilament network.

    PubMed

    Ryan, M P; Higgins, P J

    1988-10-01

    Murine sarcoma virus-transformed rat fibroblasts (KNRK cells) undergo marked cytoarchitectural reorganization during in vitro exposure to sodium-n-butyrate (NaB) resulting in restoration of (1) a more typical fibroblastoid morphology, (2) proper cell-to-cell orientation, and (3) substratum adherence. Augmented cell spreading, involving greater than 90% of the population, was a function of culture density and time of exposure to NaB (2 mM final concentration). Induced cell spreading reflected a 2.5- to 3.0-fold increase in both total cellular actin content and deposition of actin into the detergent-resistant cytoskeleton. Cytoskeletal actin deposition in response to NaB was accompanied by the formation of occasionally dense, parallel alignments of F-actin-containing microfilaments and by a dramatic increase in the size and incidence of actin-enriched membrane ruffles. Long-term NaB-treated cells exhibited parallel orientations of microfilaments similar to those found in untransformed fibroblasts. Increased cytoskeletal actin occurred within 24 hr of NaB exposure, correlating with the initial reorganization of actin-containing microfilaments detected microscopically, and reflected concomitant 3-fold increases in cellular alpha-actinin and fibronectin content. In contrast, the amount of vimentin, tropomyosin, and tubulin in NaB-treated cells was significantly decreased. NaB-induced morphologic restructuring of sarcoma virus-transformed fibroblasts, thus, impacts on all three basic cytoskeletal systems. Selective increases, however, were evident in particular cytoskeletal proteins (actin, alpha-actinin, fibronectin) implicated in microfilament networking and cell spreading. PMID:2844835

  2. Yeast Actin-Related Protein ARP6 Negatively Regulates Agrobacterium-Mediated Transformation of Yeast Cell.

    PubMed

    Luo, Yumei; Chen, Zikai; Zhu, Detu; Tu, Haitao; Pan, Shen Quan

    2015-01-01

    The yeasts, including Saccharomyces cerevisiae and Pichia pastoris, are single-cell eukaryotic organisms that can serve as models for human genetic diseases and hosts for large scale production of recombinant proteins in current biopharmaceutical industry. Thus, efficient genetic engineering tools for yeasts are of great research and economic values. Agrobacterium tumefaciens-mediated transformation (AMT) can transfer T-DNA into yeast cells as a method for genetic engineering. However, how the T-DNA is transferred into the yeast cells is not well established yet. Here our genetic screening of yeast knockout mutants identified a yeast actin-related protein ARP6 as a negative regulator of AMT. ARP6 is a critical member of the SWR1 chromatin remodeling complex (SWR-C); knocking out some other components of the complex also increased the transformation efficiency, suggesting that ARP6 might regulate AMT via SWR-C. Moreover, knockout of ARP6 led to disruption of microtubule integrity, higher uptake and degradation of virulence proteins, and increased DNA stability inside the cells, all of which resulted in enhanced transformation efficiency. Our findings have identified molecular and cellular mechanisms regulating AMT and a potential target for enhancing the transformation efficiency in yeast cells. PMID:26425545

  3. Yeast Actin-Related Protein ARP6 Negatively Regulates Agrobacterium-Mediated Transformation of Yeast Cell

    PubMed Central

    Luo, Yumei; Chen, Zikai; Zhu, Detu; Tu, Haitao; Pan, Shen Quan

    2015-01-01

    The yeasts, including Saccharomyces cerevisiae and Pichia pastoris, are single-cell eukaryotic organisms that can serve as models for human genetic diseases and hosts for large scale production of recombinant proteins in current biopharmaceutical industry. Thus, efficient genetic engineering tools for yeasts are of great research and economic values. Agrobacterium tumefaciens-mediated transformation (AMT) can transfer T-DNA into yeast cells as a method for genetic engineering. However, how the T-DNA is transferred into the yeast cells is not well established yet. Here our genetic screening of yeast knockout mutants identified a yeast actin-related protein ARP6 as a negative regulator of AMT. ARP6 is a critical member of the SWR1 chromatin remodeling complex (SWR-C); knocking out some other components of the complex also increased the transformation efficiency, suggesting that ARP6 might regulate AMT via SWR-C. Moreover, knockout of ARP6 led to disruption of microtubule integrity, higher uptake and degradation of virulence proteins, and increased DNA stability inside the cells, all of which resulted in enhanced transformation efficiency. Our findings have identified molecular and cellular mechanisms regulating AMT and a potential target for enhancing the transformation efficiency in yeast cells. PMID:26425545

  4. Coupling actin dynamics to phase-field in modeling neural growth.

    PubMed

    Najem, Sara; Grant, Martin

    2015-06-14

    In this paper we model the growth of a neural cell together with the actin dynamics taking place at its growing region by constructing a phase-field model. This is done by assigning auxiliary fields to different constituents of the cell in order to differentiate them. Specifically, the inner and outer regions of the neural cell are described by ϕ = 1 and ϕ = 0 respectively, whereas the inside and outside of its leading edge are portrayed by ψ = 1 and ψ = 0. This formulation inherently locates the boundary, which is required to determine the evolution of the underlying actin dynamics. Therefore, it provides an alternative to boundary tracking algorithms. Then the equations governing the molecular workings of the cell specifically those of actin are modified in order to satisfy their corresponding boundary conditions. PMID:25943025

  5. Initial stem cell adhesion on porous silicon surface: molecular architecture of actin cytoskeleton and filopodial growth

    NASA Astrophysics Data System (ADS)

    Collart-Dutilleul, Pierre-Yves; Panayotov, Ivan; Secret, Emilie; Cunin, Frédérique; Gergely, Csilla; Cuisinier, Frédéric; Martin, Marta

    2014-10-01

    The way cells explore their surrounding extracellular matrix (ECM) during development and migration is mediated by lamellipodia at their leading edge, acting as an actual motor pulling the cell forward. Lamellipodia are the primary area within the cell of actin microfilaments (filopodia) formation. In this work, we report on the use of porous silicon (pSi) scaffolds to mimic the ECM of mesenchymal stem cells from the dental pulp (DPSC) and breast cancer (MCF-7) cells. Our atomic force microscopy (AFM), fluorescence microscopy, and scanning electron microscopy (SEM) results show that pSi promoted the appearance of lateral filopodia protruding from the DPSC cell body and not only in the lamellipodia area. The formation of elongated lateral actin filaments suggests that pores provided the necessary anchorage points for protrusion growth. Although MCF-7 cells displayed a lower presence of organized actin network on both pSi and nonporous silicon, pSi stimulated the formation of extended cell protrusions.

  6. Arabidopsis VILLIN5, an Actin Filament Bundling and Severing Protein, Is Necessary for Normal Pollen Tube Growth[W

    PubMed Central

    Zhang, Hua; Qu, Xiaolu; Bao, Chanchan; Khurana, Parul; Wang, Qiannan; Xie, Yurong; Zheng, Yiyan; Chen, Naizhi; Blanchoin, Laurent; Staiger, Christopher J.; Huang, Shanjin

    2010-01-01

    A dynamic actin cytoskeleton is essential for pollen germination and tube growth. However, the molecular mechanisms underlying the organization and turnover of the actin cytoskeleton in pollen remain poorly understood. Villin plays a key role in the formation of higher-order structures from actin filaments and in the regulation of actin dynamics in eukaryotic cells. It belongs to the villin/gelsolin/fragmin superfamily of actin binding proteins and is composed of six gelsolin-homology domains at its core and a villin headpiece domain at its C terminus. Recently, several villin family members from plants have been shown to sever, cap, and bundle actin filaments in vitro. Here, we characterized a villin isovariant, Arabidopsis thaliana VILLIN5 (VLN5), that is highly and preferentially expressed in pollen. VLN5 loss-of-function retarded pollen tube growth and sensitized actin filaments in pollen grains and tubes to latrunculin B. In vitro biochemical analyses revealed that VLN5 is a typical member of the villin family and retains a full suite of activities, including barbed-end capping, filament bundling, and calcium-dependent severing. The severing activity was confirmed with time-lapse evanescent wave microscopy of individual actin filaments in vitro. We propose that VLN5 is a major regulator of actin filament stability and turnover that functions in concert with oscillatory calcium gradients in pollen and therefore plays an integral role in pollen germination and tube growth. PMID:20807879

  7. Bidirectional interactions between NOX2-type NADPH oxidase and the F-actin cytoskeleton in neuronal growth cones.

    PubMed

    Munnamalai, Vidhya; Weaver, Cory J; Weisheit, Corinne E; Venkatraman, Prahatha; Agim, Zeynep Sena; Quinn, Mark T; Suter, Daniel M

    2014-08-01

    NADPH oxidases are important for neuronal function but detailed subcellular localization studies have not been performed. Here, we provide the first evidence for the presence of functional NADPH oxidase 2 (NOX2)-type complex in neuronal growth cones and its bidirectional relationship with the actin cytoskeleton. NADPH oxidase inhibition resulted in reduced F-actin content, retrograde F-actin flow, and neurite outgrowth. Stimulation of NADPH oxidase via protein kinase C activation increased levels of hydrogen peroxide in the growth cone periphery. The main enzymatic NADPH oxidase subunit NOX2/gp91(phox) localized to the growth cone plasma membrane and showed little overlap with the regulatory subunit p40(phox) . p40(phox) itself exhibited colocalization with filopodial actin bundles. Differential subcellular fractionation revealed preferential association of NOX2/gp91(phox) and p40(phox) with the membrane and the cytoskeletal fraction, respectively. When neurite growth was evoked with beads coated with the cell adhesion molecule apCAM, we observed a significant increase in colocalization of p40(phox) with NOX2/gp91(phox) at apCAM adhesion sites. Together, these findings suggest a bidirectional functional relationship between NADPH oxidase activity and the actin cytoskeleton in neuronal growth cones, which contributes to the control of neurite outgrowth. We have previously shown that reactive oxygen species (ROS) are critical for actin organization and dynamics in neuronal growth cones as well as neurite outgrowth. Here, we report that the cytosolic subunit p40(phox) of the NOX2-type NADPH oxidase complex is partially associated with F-actin in neuronal growth cones, while ROS produced by this complex regulates F-actin dynamics and neurite growth. These findings provide evidence for a bidirectional relationship between NADPH oxidase activity and the actin cytoskeleton in neuronal growth cones. PMID:24702317

  8. Stabilization of actin bundles by a dynamin 1/cortactin ring complex is necessary for growth cone filopodia.

    PubMed

    Yamada, Hiroshi; Abe, Tadashi; Satoh, Ayano; Okazaki, Nana; Tago, Shota; Kobayashi, Kinue; Yoshida, Yumi; Oda, Yoshiya; Watanabe, Masami; Tomizawa, Kazuhito; Matsui, Hideki; Takei, Kohji

    2013-03-01

    Dynamin GTPase, a key molecule in endocytosis, mechanically severs the invaginated membrane upon GTP hydrolysis. Dynamin functions also in regulating actin cytoskeleton, but the mechanisms are yet to be defined. Here we show that dynamin 1, a neuronal isoform of dynamin, and cortactin form ring complexes, which twine around F-actin bundles and stabilize them. By negative-staining EM, dynamin 1-cortactin complexes appeared as "open" or "closed" rings depending on guanine nucleotide conditions. By pyrene actin assembly assay, dynamin 1 stimulated actin assembly in mouse brain cytosol. In vitro incubation of F-actin with both dynamin 1 and cortactin led to the formation of long and thick actin bundles, on which dynamin 1 and cortactin were periodically colocalized in puncta. A depolymerization assay revealed that dynamin 1 and cortactin increased the stability of actin bundles, most prominently in the presence of GTP. In rat cortical neurons and human neuroblastoma cell line, SH-SY5Y, both dynamin 1 and cortactin localized on actin filaments and the bundles at growth cone filopodia as revealed by immunoelectron microscopy. In SH-SY5Y cell, acute inhibition of dynamin 1 by application of dynamin inhibitor led to growth cone collapse. Cortactin knockdown also reduced growth cone filopodia. Together, our results strongly suggest that dynamin 1 and cortactin ring complex mechanically stabilizes F-actin bundles in growth cone filopodia. Thus, the GTPase-dependent mechanochemical enzyme property of dynamin is commonly used both in endocytosis and regulation of F-actin bundles by a dynamin 1-cortactin complex. PMID:23467367

  9. Emergence of Large-Scale Cell Morphology and Movement from Local Actin Filament Growth Dynamics

    PubMed Central

    Lacayo, Catherine I; Pincus, Zachary; VanDuijn, Martijn M; Wilson, Cyrus A; Fletcher, Daniel A; Gertler, Frank B; Mogilner, Alex; Theriot, Julie A

    2007-01-01

    phenotypes as large-scale consequences of kinetic contributions of VASP to actin filament growth and protection from capping at the leading edge. This work shows that the local effects of actin-remodeling proteins on cytoskeletal dynamics and organization can manifest as global modifications of the shape and behavior of migrating cells and that mathematical modeling can elucidate these large-scale cell behaviors from knowledge of detailed multiscale protein interactions. PMID:17760506

  10. Initial stem cell adhesion on porous silicon surface: molecular architecture of actin cytoskeleton and filopodial growth

    PubMed Central

    2014-01-01

    The way cells explore their surrounding extracellular matrix (ECM) during development and migration is mediated by lamellipodia at their leading edge, acting as an actual motor pulling the cell forward. Lamellipodia are the primary area within the cell of actin microfilaments (filopodia) formation. In this work, we report on the use of porous silicon (pSi) scaffolds to mimic the ECM of mesenchymal stem cells from the dental pulp (DPSC) and breast cancer (MCF-7) cells. Our atomic force microscopy (AFM), fluorescence microscopy, and scanning electron microscopy (SEM) results show that pSi promoted the appearance of lateral filopodia protruding from the DPSC cell body and not only in the lamellipodia area. The formation of elongated lateral actin filaments suggests that pores provided the necessary anchorage points for protrusion growth. Although MCF-7 cells displayed a lower presence of organized actin network on both pSi and nonporous silicon, pSi stimulated the formation of extended cell protrusions. PMID:25386101

  11. In β-actin knockouts, epigenetic reprogramming and rDNA transcription inactivation lead to growth and proliferation defects.

    PubMed

    Almuzzaini, Bader; Sarshad, Aishe A; Rahmanto, Aldwin S; Hansson, Magnus L; Von Euler, Anne; Sangfelt, Olle; Visa, Neus; Farrants, Ann-Kristin Östlund; Percipalle, Piergiorgio

    2016-08-01

    Actin and nuclear myosin 1 (NM1) are regulators of transcription and chromatin organization. Using a genome-wide approach, we report here that β-actin binds intergenic and genic regions across the mammalian genome, associated with both protein-coding and rRNA genes. Within the rDNA, the distribution of β-actin correlated with NM1 and the other subunits of the B-WICH complex, WSTF and SNF2h. In β-actin(-/-) mouse embryonic fibroblasts (MEFs), we found that rRNA synthesis levels decreased concomitantly with drops in RNA polymerase I (Pol I) and NM1 occupancies across the rRNA gene. Reintroduction of wild-type β-actin, in contrast to mutated forms with polymerization defects, efficiently rescued rRNA synthesis underscoring the direct role for a polymerization-competent form of β-actin in Pol I transcription. The rRNA synthesis defects in the β-actin(-/-) MEFs are a consequence of epigenetic reprogramming with up-regulation of the repressive mark H3K4me1 (monomethylation of lys4 on histone H3) and enhanced chromatin compaction at promoter-proximal enhancer (T0 sequence), which disturb binding of the transcription factor TTF1. We propose a novel genome-wide mechanism where the polymerase-associated β-actin synergizes with NM1 to coordinate permissive chromatin with Pol I transcription, cell growth, and proliferation.-Almuzzaini, B., Sarshad, A. A. , Rahmanto, A. S., Hansson, M. L., Von Euler, A., Sangfelt, O., Visa, N., Farrants, A.-K. Ö., Percipalle, P. In β-actin knockouts, epigenetic reprogramming and rDNA transcription inactivation lead to growth and proliferation defects. PMID:27127100

  12. Paclitaxel-loaded ethosomes®: potential treatment of squamous cell carcinoma, a malignant transformation of actinic keratoses.

    PubMed

    Paolino, Donatella; Celia, Christian; Trapasso, Elena; Cilurzo, Felisa; Fresta, Massimo

    2012-05-01

    Topical application of anticancer drugs for the treatment of malignancies represents a new challenge in dermatology, potentially being an alternative therapeutic approach for the efficacious treatment of non-melanoma skin cancer, that is, actinic keratoses, and malignant lesions of the skin caused by ultraviolet radiation. Anti-proliferative and antimitotic drugs, including many of the taxanes, are currently under investigation for the treatment of cutaneous malignant transformation of actinic keratoses, particularly the squamous cell carcinoma. Paclitaxel-loaded ethosomes® are proposed as topical drug delivery systems for the treatment of this pathology due to their suitable physicochemical characteristics and enhanced skin penetration ability for deep dermal delivery. Our in vitro data show that the skin application of paclitaxel-loaded ethosomes® improved the permeation of paclitaxel in a stratum corneum-epidermis membrane model and increased its anti-proliferative activity in a squamous cell carcinoma model as compared to the free drug. The results obtained encouraged the use of the paclitaxel-loaded ethosomes® as the formulation for the potential treatment of squamous cell carcinoma, a malignant transformation of actinic keratoses. PMID:22414731

  13. Bidirectional interactions between NOX2-type NADPH oxidase and the F-actin cytoskeleton in neuronal growth cones

    PubMed Central

    Munnamalai, Vidhya; Weaver, Cory J.; Weisheit, Corinne E.; Venkatraman, Prahatha; Agim, Zeynep Sena; Quinn, Mark T.; Suter, Daniel M.

    2014-01-01

    NADPH oxidases are important for neuronal function but detailed subcellular localization studies have not been performed. Here, we provide the first evidence for the presence of functional NOX2-type NADPH oxidase complex in neuronal growth cones and its bidirectional relationship with the actin cytoskeleton. NADPH oxidase inhibition resulted in reduced F-actin content, retrograde F-actin flow, and neurite outgrowth. Stimulation of NADPH oxidase via protein kinase C activation increased levels of hydrogen peroxide in the growth cone periphery. The main enzymatic NADPH oxidase subunit NOX2/gp91phox localized to the growth cone plasma membrane and showed little overlap with the regulatory subunit p40phox. p40phox itself exhibited co-localization with filopodial actin bundles. Differential subcellular fractionation revealed preferential association of NOX2/gp91phox and p40phox with the membrane and the cytoskeletal fraction, respectively. When neurite growth was evoked with beads coated with the cell adhesion molecule apCAM, we observed a significant increase in co-localization of p40phox with NOX2/gp91phox at apCAM adhesion sites. Together, these findings suggest a bidirectional functional relationship between NADPH oxidase activity and the actin cytoskeleton in neuronal growth cones, which contributes to the control of neurite outgrowth. PMID:24702317

  14. Regulation of the actin cytoskeleton in Helicobacter pylori-induced migration and invasive growth of gastric epithelial cells

    PubMed Central

    2011-01-01

    Dynamic rearrangement of the actin cytoskeleton is a significant hallmark of Helicobacter pylori (H. pylori) infected gastric epithelial cells leading to cell migration and invasive growth. Considering the cellular mechanisms, the type IV secretion system (T4SS) and the effector protein cytotoxin-associated gene A (CagA) of H. pylori are well-studied initiators of distinct signal transduction pathways in host cells targeting kinases, adaptor proteins, GTPases, actin binding and other proteins involved in the regulation of the actin lattice. In this review, we summarize recent findings of how H. pylori functionally interacts with the complex signaling network that controls the actin cytoskeleton of motile and invasive gastric epithelial cells. PMID:22044652

  15. Growth inhibition and changes in morphology and actin distribution in Acetabularia acetabulum by phalloidin and phalloidin derivatives.

    PubMed

    Sawitzky, H; Hanfstingl, U; Faulstich, H

    2003-03-01

    Effects on morphology and microfilament structure caused by phalloidin, phallacidin, and some semisynthetic phalloidin derivatives were studied in vegetative cells of the green alga Acetabularia acetabulum (L.) Silva. All phalloidin derivatives (except for phalloidin itself) caused growth stop of the alga after 1 day and (except for the fluorescein-labeled phalloidin) death of the cells after 4-7 days. Hair whorl tip growth and morphology as screened by light microscopy, as well as microfilament structure in tips, suggested that growth stop is correlated with a disorganization of actin filaments similar to that recently described for jasplakinolide (H. Sawitzky, S. Liebe, J. Willingale-Theune, D. Menzel, European Journal of Cell Biology 78: 424-433, 1999). Using rabbit muscle actin as a model target protein, we found that the toxic effects in vivo did not correlate with actin affinity values, suggesting that permeation through membranes must play a role. Indeed, the most lipophilic phalloidin derivatives benzoylphalloidin and dithiolanophalloidin were the most active in causing growth stop at ca. 100 microM. In comparison to the concentration of jasplakinolide required to cause similar effects (<3 microM), the two most active phalloidin derivatives exhibited an activity ca. 30 times lower. Nonetheless, lipophilic phalloidin derivatives can be used in algae, and probably also other cells, to modulate actin dynamics in vivo. In addition, we found that the fluorescent fluorescein isothiocyanate-phalloidin is able to enter living algal cells and stains actin structures brightly. Since it does not suppress actin dynamics, we suggest fluorescein isothiocyanate-phalloidin as a tool for studying rearrangements of actin structures in live cells, e.g., by confocal laser scanning microscopy. PMID:12664285

  16. How capping protein enhances actin filament growth and nucleation on biomimetic beads

    NASA Astrophysics Data System (ADS)

    Wang, Ruizhe; Carlsson, Anders E.

    2015-12-01

    Capping protein (CP), which caps the growing ends of actin filaments, accelerates actin-based motility. Recent experiments on biomimetic beads have shown that CP also enhances the rate of actin filament nucleation. Proposed explanations for these phenomena include (i) the actin funneling hypothesis (AFH), in which the presence of CP increases the free-actin concentration, and (ii) the monomer gating model, in which CP binding to actin filament barbed ends makes more monomers available for filament nucleation. To establish how CP increases the rates of filament elongation and nucleation on biomimetic beads, we perform a quantitative modeling analysis of actin polymerization, using rate equations that include actin filament nucleation, polymerization and capping, as modified by monomer depletion near the surface of the bead. With one adjustable parameter, our simulation results match previously measured time courses of polymerized actin and filament number. The results support a version of the AFH where CP increases the local actin monomer concentration at the bead surface, but leaves the global free-actin concentration nearly constant. Because the rate of filament nucleation increases with the monomer concentration, the increased local monomer concentration enhances actin filament nucleation. We derive a closed-form formula for the characteristic CP concentration where the local free-actin concentration reaches half the bulk value, and find it to be comparable to the global Arp2/3 complex concentration. We also propose an experimental protocol for distinguishing branching nucleation of filaments from spontaneous nucleation.

  17. A variational approach to the growth dynamics of pre-stressed actin filament networks

    NASA Astrophysics Data System (ADS)

    John, Karin; Stöter, Thomas; Misbah, Chaouqi

    2016-09-01

    In order to model the growth dynamics of elastic bodies with residual stresses a thermodynamically consistent approach is needed such that the cross-coupling between growth and mechanics can be correctly described. In the present work we apply a variational principle to the formulation of the interfacial growth dynamics of dendritic actin filament networks growing from biomimetic beads, an experimentally well studied system, where the buildup of residual stresses governs the network growth. We first introduce the material model for the network via a strain energy density for an isotropic weakly nonlinear elastic material and then derive consistently from this model the dynamic equations for the interfaces, i.e. for a polymerizing internal interface in contact with the bead and a depolymerizing external interface directed towards the solvent. We show that (i) this approach automatically preserves thermodynamic symmetry-properties, which is not the case for the often cited ‘rubber-band-model’ (Sekimoto et al 2004 Eur. Phys. J. E 13 247–59, Plastino et al 2004 Eur. Biophys. J. 33 310–20) and (ii) leads to a robust morphological instability of the treadmilling network interfaces. The nature of the instability depends on the interplay of the two dynamic interfaces. Depending on the biochemical conditions the network envelope evolves into a comet-like shape (i.e. the actin envelope thins out at one side and thickens on the opposite side of the bead) via a varicose instability or it breaks the symmetry via higher order zigzag modes. We conclude that morphological instabilities due to mechano-chemical coupling mechanisms and the presences of mechancial pre-stresses can play a major role in locally organizing the cytoskeleton of living cells.

  18. A variational approach to the growth dynamics of pre-stressed actin filament networks.

    PubMed

    John, Karin; Stöter, Thomas; Misbah, Chaouqi

    2016-09-21

    In order to model the growth dynamics of elastic bodies with residual stresses a thermodynamically consistent approach is needed such that the cross-coupling between growth and mechanics can be correctly described. In the present work we apply a variational principle to the formulation of the interfacial growth dynamics of dendritic actin filament networks growing from biomimetic beads, an experimentally well studied system, where the buildup of residual stresses governs the network growth. We first introduce the material model for the network via a strain energy density for an isotropic weakly nonlinear elastic material and then derive consistently from this model the dynamic equations for the interfaces, i.e. for a polymerizing internal interface in contact with the bead and a depolymerizing external interface directed towards the solvent. We show that (i) this approach automatically preserves thermodynamic symmetry-properties, which is not the case for the often cited 'rubber-band-model' (Sekimoto et al 2004 Eur. Phys. J. E 13 247-59, Plastino et al 2004 Eur. Biophys. J. 33 310-20) and (ii) leads to a robust morphological instability of the treadmilling network interfaces. The nature of the instability depends on the interplay of the two dynamic interfaces. Depending on the biochemical conditions the network envelope evolves into a comet-like shape (i.e. the actin envelope thins out at one side and thickens on the opposite side of the bead) via a varicose instability or it breaks the symmetry via higher order zigzag modes. We conclude that morphological instabilities due to mechano-chemical coupling mechanisms and the presences of mechancial pre-stresses can play a major role in locally organizing the cytoskeleton of living cells. PMID:27420637

  19. Grain nucleation and growth during phase transformations.

    PubMed

    Offerman, S E; van Dijk, N H; Sietsma, J; Grigull, S; Lauridsen, E M; Margulies, L; Poulsen, H F; Rekveldt, M Th; van der Zwaag, S

    2002-11-01

    The mechanical properties of polycrystalline materials are largely determined by the kinetics of the phase transformations during the production process. Progress in x-ray diffraction instrumentation at synchrotron sources has created an opportunity to study the transformation kinetics at the level of individual grains. Our measurements show that the activation energy for grain nucleation is at least two orders of magnitude smaller than that predicted by thermodynamic models. The observed growth curves of the newly formed grains confirm the parabolic growth model but also show three fundamentally different types of growth. Insight into the grain nucleation and growth mechanisms during phase transformations contributes to the development of materials with optimal mechanical properties. PMID:12411699

  20. The F-BAR protein Hof1 tunes formin activity to sculpt actin cables during polarized growth

    PubMed Central

    Graziano, Brian R.; Yu, Hoi-Ying E.; Alioto, Salvatore L.; Eskin, Julian A.; Ydenberg, Casey A.; Waterman, David P.; Garabedian, Mikael; Goode, Bruce L.

    2014-01-01

    Asymmetric cell growth and division rely on polarized actin cytoskeleton remodeling events, the regulation of which is poorly understood. In budding yeast, formins stimulate the assembly of an organized network of actin cables that direct polarized secretion. Here we show that the Fer/Cip4 homology–Bin amphiphysin Rvs protein Hof1, which has known roles in cytokinesis, also functions during polarized growth by directly controlling the activities of the formin Bnr1. A mutant lacking the C-terminal half of Hof1 displays misoriented and architecturally altered cables, along with impaired secretory vesicle traffic. In vitro, Hof1 inhibits the actin nucleation and elongation activities of Bnr1 without displacing the formin from filament ends. These effects depend on the Src homology 3 domain of Hof1, the formin homology 1 (FH1) domain of Bnr1, and Hof1 dimerization, suggesting a mechanism by which Hof1 “restrains” the otherwise flexible FH1-FH2 apparatus. In vivo, loss of inhibition does not alter actin levels in cables but, instead, cable shape and functionality. Thus Hof1 tunes formins to sculpt the actin cable network. PMID:24719456

  1. The Disruption of the Cytoskeleton during Semaphorin 3A induced Growth Cone Collapse Correlates with Differences in Actin Organization and Associated Binding Proteins

    PubMed Central

    Brown, Jacquelyn A; Bridgman, Paul C

    2010-01-01

    Repulsive guidance cues induce growth cone collapse or collapse and retraction. Collapse results from disruption and loss of the actin cytoskeleton. Actin rich regions of growth cones contain binding proteins that influence filament organization, such as Arp2/3, cortactin, and fascin, but little is known about the role that these proteins play in collapse. Here we show that Semaphorin 3A (Sema 3A), which is repulsive to mouse dorsal root ganglion neurons, has unequal effects on actin binding proteins and their associated filaments. The immunofluorescence staining intensity of Arp-2 and cortactin decreases relative to total protein, while in unextracted growth cones fascin increases. Fascin and myosin IIB staining redistribute and show increased overlap. The degree of actin filament loss during collapse correlates with filament superstructures detected by rotary shadow electron microscopy. Collapse results in the loss of branched f-actin meshworks, while actin bundles are partially retained to varying degrees. Taken together with the known affects of Sema 3A on actin, this suggests a model for collapse that follows a sequence; depolymerization of actin meshworks followed by partial depolymerization of fascin associated actin bundles and their movement to the neurite to complete collapse. The relocated fascin associated actin bundles may provide the substrate for actomyosin contractions that produce retraction. PMID:19513995

  2. Actin binding and proline rich motifs of CR16 play redundant role in growth of vrp1Delta cells.

    PubMed

    Meng, Lei; Rajmohan, Rajamuthiah; Yu, Shangjuan; Thanabalu, Thirumaran

    2007-05-25

    CR16, (Glucocorticoid-regulated) belongs to the verprolin family of proteins which are characterized by the presence of a V domain (verprolin) at the N-terminal. Expression of CR16 suppressed the growth and endocytosis defect of vrp1Delta strain without correcting the actin patch polarization defect. The V domain of CR16 is critical for suppression of the growth defect of vrp1Delta strain but not for localisation to cortical actin patches. Mutations in the actin binding motif alone did not abolish the activity of CR16 but the mutations in combination with deletion of N-terminal proline rich motif abolished the ability of CR16 to suppress the growth defect. This suggests that the V domain of CR16 has two functionally redundant motifs and either one of these motifs is sufficient for suppressing the growth defect of vrp1Delta strain. This is in contrast to the observation that both WIP and WIRE require the actin binding motif for their activity. PMID:17418095

  3. Tropomyosin-1 protects transformed alveolar epithelial cells against cigaret smoke extract through the stabilization of F-actin-dependent cell-cell junctions.

    PubMed

    Gagat, Maciej; Grzanka, Dariusz; Izdebska, Magdalena; Sroka, Wiktor Dariusz; Hałas-Wiśniewska, Marta; Grzanka, Alina

    2016-04-01

    The aim of the study was to estimate the effect of tropomyosin-1-based structural stabilization of F-actin in transformed human alveolar epithelial line H1299 cells subjected to high oxidative stress induced by cigaret smoke extract. We demonstrated here that cigaret smoke extract induces cell shrinking and detachment as a consequence of F-actin cytoskeleton degradation in H1299 cells not overexpressing tropomyosin-1. Furthermore, the treatment of these cells with cigaret smoke extract resulted in the loss of peripheral localization of ZO-1 and initiated apoptosis. In contrast, structural stabilization of F-actin, by overexpression of tropomyosin-1, preserved cell to cell interactions through the attenuation of cortical actin organization into thin fibers and thus protected these cells against oxidative stress-induced degradation of actin cytoskeleton and cell death. In conclusion, we suggest that structural stabilization of thin cortical F-actin fibers increases link between tight junctions proteins and actin cytoskeleton and thus protects H1299 cells against cigaret smoke extract. PMID:26805581

  4. A novel protein kinase gene ssp1+ is required for alteration of growth polarity and actin localization in fission yeast.

    PubMed Central

    Matsusaka, T; Hirata, D; Yanagida, M; Toda, T

    1995-01-01

    Temperature-sensitive suppressor mutants were isolated from two fission yeast mutants defective in cell shape control: ppe1, encoding a type 2A-like protein phosphatase, and sts5, one of 11 staurosporine-supersensitive mutants. Complementation tests showed that suppression was due to two chromosomal loci, ssp1 and ssp2. Cells of the ssp1 mutant grown at the restrictive temperature arrested uniformly with an elongated cell body and a 2C content of DNA. Interestingly, these mutant cells grew only in a monopolar manner. At a specific point in the G2 phase of the cell cycle, wild-type cells exhibit a drastic alteration in growth polarity, from mono- to bipolar. This change coincides with the distribution of cortical actin from one end of the cell to both ends. In the ssp1 mutant cells, cortical actin was localized only at one end, suggesting that the mutant fails to change growth polarity. Nucleotide sequence determination showed that ssp1+ encodes a novel protein kinase. Ectopic overexpression of ssp1+ resulted in an altered cell morphology and cortical actin was randomly dispersed within the cells. Immunocytological analysis revealed that the protein was primarily localized in the cytoplasm and that half of the protein existed in an insoluble fraction. These results show that the dynamics of actin-based growth polarity during the cell cycle are regulated, at least in part, by a novel set of protein kinases and phosphatases. Images PMID:7628434

  5. Formin-Dependent Synaptic Growth; Evidence that Dlar Signals via Diaphanous to Modulate Synaptic Actin and Dynamic Pioneer Microtubules

    PubMed Central

    Pawson, Catherine; Eaton, Benjamin A.; Davis, Graeme W.

    2008-01-01

    The diaphanous gene is the founding member of a family of Diaphanous Related Formin proteins (DRF). We identified diaphanous in a screen for genes that are necessary for the normal growth and stabilization of the Drosophila neuromuscular junction (NMJ). Here we demonstrate that diaphanous mutations perturb synaptic growth at the NMJ. Diaphanous protein is present both pre- and postsynaptically. However, genetic rescue experiments in combination with additional genetic interaction experiments support the conclusion that dia is necessary presynaptically for normal NMJ growth. We then document defects in both the actin and microtubule cytoskeletons in dia mutant nerve terminals. In so doing, we define and characterize a population of dynamic pioneer microtubules within the NMJ that are distinct from the bundled core of microtubules identified by the MAP1b-like protein Futsch. Defects in both synaptic actin and dynamic pioneer MTs are correlated with impaired synaptic growth in dia mutants. Finally, we present genetic evidence that Dia functions downstream of the presynaptic receptor tyrosine phosphatase Dlar and the Rho-type GEF trio to control NMJ growth. Based upon the established function of DRFs as Rho-GTPase dependent regulators of the cell cytoskeleton, we propose a model in which Diaphanous links receptor tyrosine phosphatase signaling at the plasma membrane to growth-dependent modulation of the synaptic actin and microtubule cytoskeletons. PMID:18971454

  6. Cdc42 and Actin Control Polarized Expression of TI-VAMP Vesicles to Neuronal Growth Cones and Their Fusion with the Plasma MembraneV⃞

    PubMed Central

    Alberts, Philipp; Rudge, Rachel; Irinopoulou, Theano; Danglot, Lydia; Gauthier-Rouvière, Cécile; Galli, Thierry

    2006-01-01

    Tetanus neurotoxin-insensitive vesicle-associated membrane protein (TI-VAMP)-mediated fusion of intracellular vesicles with the plasma membrane is crucial for neurite outgrowth, a pathway not requiring synaptobrevin-dependent exocytosis. Yet, it is not known how the TI-VAMP membrane trafficking pathway is regulated or how it is coordinated with cytoskeletal dynamics within the growth cone that guide neurite outgrowth. Here, we demonstrate that TI-VAMP, but not synaptobrevin 2, concentrates in the peripheral, F-actin-rich region of the growth cones of hippocampal neurons in primary culture. Its accumulation correlates with and depends upon the presence of F-actin. Moreover, acute stimulation of actin remodeling by homophilic activation of the adhesion molecule L1 induces a site-directed, actin-dependent recruitment of the TI-VAMP compartment. Expression of a dominant-positive mutant of Cdc42, a key regulator of cell polarity, stimulates formation of F-actin- and TI-VAMP-rich filopodia outside the growth cone. Furthermore, we report that Cdc42 activates exocytosis of pHLuorin tagged TI-VAMP in an actin-dependent manner. Collectively, our data suggest that Cdc42 and regulated assembly of the F-actin network control the accumulation and exocytosis of TI-VAMP-containing membrane vesicles in growth cones to coordinate membrane trafficking and actin remodeling during neurite outgrowth. PMID:16381811

  7. The Apical Actin Fringe Contributes to Localized Cell Wall Deposition and Polarized Growth in the Lily Pollen Tube1[W][OPEN

    PubMed Central

    Rounds, Caleb M.; Hepler, Peter K.; Winship, Lawrence J.

    2014-01-01

    In lily (Lilium formosanum) pollen tubes, pectin, a major component of the cell wall, is delivered through regulated exocytosis. The targeted transport and secretion of the pectin-containing vesicles may be controlled by the cortical actin fringe at the pollen tube apex. Here, we address the role of the actin fringe using three different inhibitors of growth: brefeldin A, latrunculin B, and potassium cyanide. Brefeldin A blocks membrane trafficking and inhibits exocytosis in pollen tubes; it also leads to the degradation of the actin fringe and the formation of an aggregate of filamentous actin at the base of the clear zone. Latrunculin B, which depolymerizes filamentous actin, markedly slows growth but allows focused pectin deposition to continue. Of note, the locus of deposition shifts frequently and correlates with changes in the direction of growth. Finally, potassium cyanide, an electron transport chain inhibitor, briefly stops growth while causing the actin fringe to completely disappear. Pectin deposition continues but lacks focus, instead being delivered in a wide arc across the pollen tube tip. These data support a model in which the actin fringe contributes to the focused secretion of pectin to the apical cell wall and, thus, to the polarized growth of the pollen tube. PMID:25037212

  8. Fibroblast growth factor (Fgf) 23 gene transcription depends on actin cytoskeleton reorganization.

    PubMed

    Fajol, Abul; Honisch, Sabina; Zhang, Bingbing; Schmidt, Sebastian; Alkahtani, Saad; Alarifi, Saud; Lang, Florian; Stournaras, Christos; Föller, Michael

    2016-03-01

    FGF23 regulates renal phosphate and vitamin D metabolism. Loss of FGF23 results in massive calcification and rapid aging. FGF23 production is stimulated by 1,25(OH)2 D3 and NFκB signaling. Here, we report that treatment of UMR106 osteoblast-like cells with 1,25(OH)2 D3 , inducing Fgf23 transcription, resulted in actin polymerization which was blocked by NFκB inhibitor wogonin. Interestingly, 1,25(OH)2 D3 -induced Fgf23 gene transcription was abolished by the actin microfilament-disrupting agent cytochalasin B, as well as by the inhibition of actin-regulating Rac1/PAK1 signaling. Our results provide strong evidence that actin redistribution regulated by the Rac1/PAK1 pathway participates in 1,25(OH)2 D3 -induced Fgf23 gene transcription. PMID:26878191

  9. The Arabidopsis Wave Complex: Mechanisms Of Localized Actin Polymerization And Growth

    SciTech Connect

    Daniel Szymanski

    2012-10-23

    The objective of this project was to discover the protein complexes and control mechanisms that determine the location of actin filament roadways in plant cells. Our work provided the first molecular description of protein complexes that are converted from inactive complexes to active actin filament nucleators in the cell. These discoveries provided a conceptual framework to control to roadways in plant cells that determine the location and delivery of plant metabolites and storage molecules that are relevant to the bioenergy economy.

  10. Actinic Keratosis

    MedlinePlus

    ... rashes clinical tools newsletter | contact Share | Actinic Keratosis (Solar Keratosis) Information for adults A A A Actinic ... the touch. Overview Actinic keratoses, also known as solar keratoses, are small rough or scaly areas of ...

  11. F-Actin Organization and Pollen Tube Tip Growth in Arabidopsis Are Dependent on the Gametophyte-Specific Armadillo Repeat Protein ARO1[W

    PubMed Central

    Gebert, Marina; Dresselhaus, Thomas; Sprunck, Stefanie

    2008-01-01

    The signal-mediated and spatially controlled assembly and dynamics of actin are crucial for maintaining shape, motility, and tip growth of eukaryotic cells. We report that a novel Armadillo repeat protein in Arabidopsis thaliana, ARMADILLO REPEAT ONLY1 (ARO1), is of fundamental importance for polar growth and F-actin organization in tip-growing pollen tubes. ARO1 is specifically expressed in the vegetative cell of pollen as well as in the egg cell. ARO1-GFP (for green fluorescent protein) fusion proteins accumulate most notably in pollen tube tips and partially colocalize with F-actin in the shank of pollen tubes. ARO1 knockout results in a highly disorganized actin cytoskeleton, growth depolarization, and ultimately tube growth arrest. Tip-localized ARO1-GFP is spatially shifted toward the future site of tip growth, indicating a role of ARO1 in the signaling network controlling tip growth and regulating actin organization. After the pollen tube discharges its contents into the receptive synergid, ARO1-GFP colocalizes with emerging F-actin structures near the site of sperm cell fusion, suggesting additional participation in the mechanism of sperm cell tracking toward the female gametes. The variable localization of ARO1 in the cytoplasm, the nucleus, and at the plasma membrane, however, indicates a multifunctional role like that of β-catenin/Armadillo and the p120 catenins. PMID:18931021

  12. The Spatial Distribution of the Exocyst and Actin Cortical Patches Is Sufficient To Organize Hyphal Tip Growth

    PubMed Central

    Caballero-Lima, David; Kaneva, Iliyana N.; Watton, Simon P.

    2013-01-01

    In the hyphal tip of Candida albicans we have made detailed quantitative measurements of (i) exocyst components, (ii) Rho1, the regulatory subunit of (1,3)-β-glucan synthase, (iii) Rom2, the specialized guanine-nucleotide exchange factor (GEF) of Rho1, and (iv) actin cortical patches, the sites of endocytosis. We use the resulting data to construct and test a quantitative 3-dimensional model of fungal hyphal growth based on the proposition that vesicles fuse with the hyphal tip at a rate determined by the local density of exocyst components. Enzymes such as (1,3)-β-glucan synthase thus embedded in the plasma membrane continue to synthesize the cell wall until they are removed by endocytosis. The model successfully predicts the shape and dimensions of the hyphae, provided that endocytosis acts to remove cell wall-synthesizing enzymes at the subapical bands of actin patches. Moreover, a key prediction of the model is that the distribution of the synthase is substantially broader than the area occupied by the exocyst. This prediction is borne out by our quantitative measurements. Thus, although the model highlights detailed issues that require further investigation, in general terms the pattern of tip growth of fungal hyphae can be satisfactorily explained by a simple but quantitative model rooted within the known molecular processes of polarized growth. Moreover, the methodology can be readily adapted to model other forms of polarized growth, such as that which occurs in plant pollen tubes. PMID:23666623

  13. Epidermal growth factor signaling in transformed cells

    PubMed Central

    Lindsey, Stephan; Langhans, Sigrid A.

    2016-01-01

    Members of the epidermal growth factor receptor (EGFR/ErbB) family play a critical role in normal cell growth and development. However, many ErbB family members, especially EGFR, are aberrantly expressed or deregulated in tumors and are thought to play crucial roles in cancer development and metastatic progression. In this chapter, we provide an overview of key mechanisms contributing to aberrant EGFR/ErbB signaling in transformed cells which results in many phenotypic changes associated with the earliest stages of tumor formation, including several hallmarks of epithelial-to-mesenchymal transition (EMT). These changes often occur through interaction with other major signaling pathways important to tumor progression resulting in a multitude of transcriptional changes that ultimately impact cell morphology, proliferation and adhesion, all of which are crucial for tumor progression. The resulting mesh of signaling networks will need to be taken into account as new regimens are designed for targeting EGFR for therapeutic intervention. As new insights into the molecular mechanisms of the cross-talk of EGFR signaling with other signaling pathways and their role in therapeutic resistance to anti-EGFR therapies are gained a continual reassessment of clinical therapeutic regimes and strategies will be required. Understanding the consequences and complexity of EGF signaling and how it relates to tumor progression is critical for the development of clinical compounds and establishing clinical protocols for the treatment of cancer. PMID:25619714

  14. Actin from Saccharomyces cerevisiae.

    PubMed Central

    Greer, C; Schekman, R

    1982-01-01

    Inhibition of DNase I activity has been used as an assay to purify actin from Saccharomyces cerevisiae (yeast actin). The final fraction, obtained after a 300-fold purification, is approximately 97% pure as judged by sodium dodecyl sulfate-gel electrophoresis. Like rabbit skeletal muscle actin, yeast actin has a molecular weight of about 43,000, forms 7-nm-diameter filaments when polymerization is induced by KCl or Mg2+, and can be decorated with a proteolytic fragment of muscle myosin (heavy meromyosin). Although heavy meromyosin ATPase activity is stimulated by rabbit muscle and yeast actins to approximately the same Vmax (2 mmol of Pi per min per mumol of heavy meromyosin), half-maximal activation (Kapp) is obtained with 14 micro M muscle actin, but requires approximately 135 micro M yeast actin. This difference suggests a low affinity of yeast actin for muscle myosin. Yeast and muscle filamentous actin respond similarly to cytochalasin and phalloidin, although the drugs have no effect on S. cerevisiae cell growth. Images PMID:6217414

  15. Myofibril growth during cardiac hypertrophy is regulated through dual phosphorylation and acetylation of the actin capping protein CapZ.

    PubMed

    Lin, Ying-Hsi; Warren, Chad M; Li, Jieli; McKinsey, Timothy A; Russell, Brenda

    2016-08-01

    The mechanotransduction signaling pathways initiated in heart muscle by increased mechanical loading are known to lead to long-term transcriptional changes and hypertrophy, but the rapid events for adaptation at the sarcomeric level are not fully understood. The goal of this study was to test the hypothesis that actin filament assembly during cardiomyocyte growth is regulated by post-translational modifications (PTMs) of CapZβ1. In rapidly hypertrophying neonatal rat ventricular myocytes (NRVMs) stimulated by phenylephrine (PE), two-dimensional gel electrophoresis (2DGE) of CapZβ1 revealed a shift toward more negative charge. Consistent with this, mass spectrometry identified CapZβ1 phosphorylation on serine-204 and acetylation on lysine-199, two residues which are near the actin binding surface of CapZβ1. Ectopic expression of dominant negative PKCɛ (dnPKCɛ) in NRVMs blunted the PE-induced increase in CapZ dynamics, as evidenced by the kinetic constant (Kfrap) of fluorescence recovery after photobleaching (FRAP), and concomitantly reduced phosphorylation and acetylation of CapZβ1. Furthermore, inhibition of class I histone deacetylases (HDACs) increased lysine-199 acetylation on CapZβ1, which increased Kfrap of CapZ and stimulated actin dynamics. Finally, we show that PE treatment of NRVMs results in decreased binding of HDAC3 to myofibrils, suggesting a signal-dependent mechanism for the regulation of sarcomere-associated CapZβ1 acetylation. Taken together, this dual regulation through phosphorylation and acetylation of CapZβ1 provides a novel model for the regulation of myofibril growth during cardiac hypertrophy. PMID:27185186

  16. Cortactin involvement in the keratinocyte growth factor and fibroblast growth factor 10 promotion of migration and cortical actin assembly in human keratinocytes

    SciTech Connect

    Ceccarelli, Simona; Cardinali, Giorgia; Aspite, Nicaela; Picardo, Mauro; Marchese, Cinzia; Torrisi, Maria Rosaria; Mancini, Patrizia . E-mail: patrizia.mancini@uniroma1.it

    2007-05-15

    Keratinocyte growth factor (KGF/FGF7) and fibroblast growth factor 10 (FGF10/KGF2) regulate keratinocyte proliferation and differentiation by binding to the tyrosine kinase KGF receptor (KGFR). KGF induces keratinocyte motility and cytoskeletal rearrangement, whereas a direct role of FGF10 on keratinocyte migration is not clearly established. Here we analyzed the motogenic activity of FGF10 and KGF on human keratinocytes. Migration assays and immunofluorescence of actin cytoskeleton revealed that FGF10 is less efficient than KGF in promoting migration and exerts a delayed effect in inducing lamellipodia and ruffles formation. Both growth factors promoted phosphorylation and subsequent membrane translocation of cortactin, an F-actin binding protein involved in cell migration; however, FGF10-induced cortactin phosphorylation was reduced, more transient and delayed with respect to that promoted by KGF. Cortactin phosphorylation induced by both growth factors was Src-dependent, while its membrane translocation and cell migration were blocked by either Src and PI3K inhibitors, suggesting that both pathways are involved in KGF- and FGF10-dependent motility. Furthermore, siRNA-mediated downregulation of cortactin inhibited KGF- and FGF10-induced migration. These results indicate that cortactin is involved in keratinocyte migration promoted by both KGF and FGF10.

  17. Transforming growth factor-beta 1 and fibroblast growth factors in rat growth plate.

    PubMed

    Jingushi, S; Scully, S P; Joyce, M E; Sugioka, Y; Bolander, M E

    1995-09-01

    Chondrocytes in the growth plate progress in an orderly fashion from resting through proliferating to hypertrophic cells. In the region of hypertrophic chondrocytes, the cartilage is invaded by capillary loops and endochondral ossification is initiated. It is currently believed that growth factors may regulate the proliferation and maturation of chondrocytes and the synthesis of extracellular matrix in the growth plate. The ordered sequence of proliferation and differentiation observed in the growth plate provides a unique opportunity to study the role of acidic fibroblast growth factor, basic fibroblast growth factor, and transforming growth factor-beta 1 in the regulation of these processes. In this study, expression of the mRNA of these growth factors was examined using total RNA extracted from the physis and epiphysis of rat tibias. Transforming growth factor-beta 1 mRNA was detected by Northern hybridization. Expression of the genes encoding acidic and basic fibroblast growth factors was demonstrated by polymerase chain reaction amplification. In addition, using polyclonal antibodies against these growth factors, we localized them by immunohistochemical analysis. Strong intracellular staining with a predominantly nuclear pattern was observed in chondrocytes from the proliferating and upper hypertrophic zones. In contrast, chondrocytes in the resting zone stained only faintly for the presence of these growth factors. Some chondrocytes in the resting zone adjacent to the proliferating zone stained with these antibodies, and the antibodies also stained cells in the zone of Ranvier, which regulates latitudinal bone growth. Lastly, the location of transforming growth factor-beta 1 was examined further with use of a polyclonal antipeptide antibody specific for its extracellular epitope.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:7472755

  18. ARF6 promotes the formation of Rac1 and WAVE-dependent ventral F-actin rosettes in breast cancer cells in response to epidermal growth factor.

    PubMed

    Marchesin, Valentina; Montagnac, Guillaume; Chavrier, Philippe

    2015-01-01

    Coordination between actin cytoskeleton assembly and localized polarization of intracellular trafficking routes is crucial for cancer cell migration. ARF6 has been implicated in the endocytic recycling of surface receptors and membrane components and in actin cytoskeleton remodeling. Here we show that overexpression of an ARF6 fast-cycling mutant in MDA-MB-231 breast cancer-derived cells to mimick ARF6 hyperactivation observed in invasive breast tumors induced a striking rearrangement of the actin cytoskeleton at the ventral cell surface. This phenotype consisted in the formation of dynamic actin-based podosome rosette-like structures expanding outward as wave positive for F-actin and actin cytoskeleton regulatory components including cortactin, Arp2/3 and SCAR/WAVE complexes and upstream Rac1 regulator. Ventral rosette-like structures were similarly induced in MDA-MB-231 cells in response to epidermal growth factor (EGF) stimulation and to Rac1 hyperactivation. In addition, interference with ARF6 expression attenuated activation and plasma membrane targeting of Rac1 in response to EGF treatment. Our data suggest a role for ARF6 in linking EGF-receptor signaling to Rac1 recruitment and activation at the plasma membrane to promote breast cancer cell directed migration. PMID:25799492

  19. Specific Transformation of Assembly with Actin Filaments and Molecular Motors in a Cell-Sized Self-Emerged Liposome

    NASA Astrophysics Data System (ADS)

    Takiguchi, Kingo; Negishi, Makiko; Tanaka-Takiguchi, Yohko; Hayashi, Masahito; Yoshikawa, Kenichi

    2014-12-01

    Eukaryotes, by the same combination of cytoskeleton and molecular motor, for example actin filament and myosin, can generate a variety of movements. For this diversity, the organization of biological machineries caused by the confinement and/or crowding effects of internal living cells, may play very important roles.

  20. Insulin-like growth factor binding protein-1 expression in baboon endometrial stromal cells: regulation by filamentous actin and requirement for de novo protein synthesis.

    PubMed

    Kim, J J; Jaffe, R C; Fazleabas, A T

    1999-02-01

    Stromal fibroblasts in the primate endometrium undergo dramatic morphological and biochemical changes in response to pregnancy. This transformation is characterized by the expression of insulin-like growth factor binding protein-1 (IGFBP-1). Stromal cells from the baboon endometrium of nonpregnant animals were cultured and subsequently treated with cytochalasin D to disrupt actin filaments. In response to cytochalasin D treatment, cells contracted and became rounded as early as 10 min after the initiation of treatment. When cytochalasin D was removed, cells reverted back to their original fibroblastic shape within 1 h. After cells were treated with cytochalasin D for 5 h, addition of (Bu)2cAMP and/or hormones (estradiol, medroxyprogesterone acetate, and relaxin) resulted in the expression of IGFBP-1 messenger RNA and protein within 24 h. Cells with an intact cytoskeleton did not express detectable levels of IGFBP-1 in response to hormones and/or (Bu)2cAMP. Furthermore, the addition of cycloheximide inhibited expression of IGFBP-1 in cytochalasin D-treated cells. Stromal cells were also isolated from early pregnant and simulated pregnant animals. Within 48 h, cells from both the pregnant and simulated pregnant animals produced IGFBP-1 in response to hormones and/or (Bu)2cAMP. In these studies, IGFBP-1 expression was also inhibited by cycloheximide. These studies suggest that induction of IGFBP-1 requires an intermediary protein and that alterations in the cytoskeleton may be involved. PMID:9927334

  1. Cardiac fibroblasts are predisposed to convert into myocyte phenotype: Specific effect of transforming growth factor. beta

    SciTech Connect

    Eghbali, M.; Tomek, R.; Woods, C.; Bhambi, B. )

    1991-02-01

    Cardiac fibroblasts are mainly responsible for the synthesis of major extracellular matrix proteins in the heart, including fibrillar collagen types I and III and fibronectin. In this report we show that these cells, when stimulated by transforming growth factor {beta}{sub 1} (TGF-{beta}{sub 1}), acquire certain myocyte-specific properties. Cultured cardiac fibroblasts from adult rabbit heart were treated with TGF-{beta}{sub 1}, (10-15 ng/ml) for different periods of time. Northern hybridization analysis of total RNA showed that cells treated with TGF-{beta}{sub 1} became stained with a monoclonal antibody to muscle-specific actin. After treatment of quiescent cells with TGF-{beta}{sub 1}, cell proliferation (as measured by ({sup 3}H)thymidine incorporation) was moderately increased. Cultured cardiac fibroblasts at the subconfluent stage, when exposed to TGF-{beta}{sub 1} in the presence of 10% fetal bovine serum, gave rise to a second generation of slowly growing cells that expressed muscle-specific actin filaments. The findings demonstrate that cardiac fibroblasts can be made to differentiate into cells that display many characteristics of cardiac myocytes. TGF-{beta}{sub 1} seems to be a specific inducer of such conversion.

  2. Actin Automata with Memory

    NASA Astrophysics Data System (ADS)

    Alonso-Sanz, Ramón; Adamatzky, Andy

    Actin is a globular protein which forms long polar filaments in eukaryotic. The actin filaments play the roles of cytoskeleton, motility units, information processing and learning. We model actin filament as a double chain of finite state machines, nodes, which take states “0” and “1”. The states are abstractions of absence and presence of a subthreshold charge on actin units corresponding to the nodes. All nodes update their state in parallel to discrete time. A node updates its current state depending on states of two closest neighbors in the node chain and two closest neighbors in the complementary chain. Previous models of actin automata consider momentary state transitions of nodes. We enrich the actin automata model by assuming that states of nodes depend not only on the current states of neighboring node but also on their past states. Thus, we assess the effect of memory of past states on the dynamics of acting automata. We demonstrate in computational experiments that memory slows down propagation of perturbations, decrease entropy of space-time patterns generated, transforms traveling localizations to stationary oscillators, and stationary oscillations to still patterns.

  3. A single-cell correlative nanoelectromechanosensing approach to detect cancerous transformation: monitoring the function of F-actin microfilaments in the modulation of the ion channel activity

    NASA Astrophysics Data System (ADS)

    AbdolahadThe Authors With Same Contributions., Mohammad; Saeidi, Ali; Janmaleki, Mohsen; Mashinchian, Omid; Taghinejad, Mohammad; Taghinejad, Hossein; Azimi, Soheil; Mahmoudi, Morteza; Mohajerzadeh, Shams

    2015-01-01

    Cancerous transformation may be dependent on correlation between electrical disruptions in the cell membrane and mechanical disruptions of cytoskeleton structures. Silicon nanotube (SiNT)-based electrical probes, as ultra-accurate signal recorders with subcellular resolution, may create many opportunities for fundamental biological research and biomedical applications. Here, we used this technology to electrically monitor cellular mechanosensing. The SiNT probe was combined with an electrically activated glass micropipette aspiration system to achieve a new cancer diagnostic technique that is based on real-time correlation between mechanical and electrical behaviour of single cells. Our studies demonstrated marked changes in the electrical response following increases in the mechanical aspiration force in healthy cells. In contrast, such responses were extremely weak for malignant cells. Confocal microscopy results showed the impact of actin microfilament remodelling on the reduction of the electrical response for aspirated cancer cells due to the significant role of actin in modulating the ion channel activity in the cell membrane.Cancerous transformation may be dependent on correlation between electrical disruptions in the cell membrane and mechanical disruptions of cytoskeleton structures. Silicon nanotube (SiNT)-based electrical probes, as ultra-accurate signal recorders with subcellular resolution, may create many opportunities for fundamental biological research and biomedical applications. Here, we used this technology to electrically monitor cellular mechanosensing. The SiNT probe was combined with an electrically activated glass micropipette aspiration system to achieve a new cancer diagnostic technique that is based on real-time correlation between mechanical and electrical behaviour of single cells. Our studies demonstrated marked changes in the electrical response following increases in the mechanical aspiration force in healthy cells. In contrast, such

  4. Actinic keratosis

    MedlinePlus

    ... example, if you work outdoors) Had many severe sunburns early in life Are older Symptoms Actinic keratosis ... and tanning salons. Other things to know about sun exposure: Sun exposure is stronger in or near surfaces ...

  5. Actinic Cheilitis

    MedlinePlus

    ... is a precancerous condition related to cumulative lifetime sun exposure. The lower lip is most often affected. Individuals ... Wearing barrier clothing (eg, wide-brimmed hats) and sunscreen-containing lip balms can aid in preventing actinic ...

  6. ROCK1 via LIM kinase regulates growth, maturation and actin based functions in mast cells

    PubMed Central

    Kapur, Reuben; Shi, Jianjian; Ghosh, Joydeep; Munugalavadla, Veerendra; Sims, Emily; Martin, Holly; Wei, Lei; Mali, Raghuveer Singh

    2016-01-01

    Understanding mast cell development is essential due to their critical role in regulating immunity and autoimmune diseases. Here, we show how Rho kinases (ROCK) regulate mast cell development and can function as therapeutic targets for treating allergic diseases. Rock1 deficiency results in delayed maturation of bone marrow derived mast cells (BMMCs) in response to IL-3 stimulation and reduced growth in response to stem cell factor (SCF) stimulation. Further, integrin-mediated adhesion and migration, and IgE-mediated degranulation are all impaired in Rock1-deficient BMMCs. To understand the mechanism behind altered mast cell development in Rock1−/− BMMCs, we analyzed the activation of ROCK and its downstream targets including LIM kinase (LIMK). We observed reduced activation of ROCK, LIMK, AKT and ERK1/2 in Rock1-deficient BMMCs in response to SCF stimulation. Further, loss of either Limk1 or Limk2 also demonstrated altered BMMC maturation and growth; combined deletion of both Limk1 and Limk2 resulted in further reduction in BMMC maturation and growth. In passive cutaneous anaphylaxis model, deficiency of Rock1 or treatment with ROCK inhibitor Fasudil protected mice against IgE-mediated challenge. Our results identify ROCK/LIMK pathway as a novel therapeutic target for treating allergic diseases involving mast cells. PMID:26943578

  7. Cells transformed by murine herpesvirus 68 (MHV-68) release compounds with transforming and transformed phenotype suppressing activity resembling growth factors.

    PubMed

    Šupolíková, M; Staňová, A Vojs; Kúdelová, M; Marák, J; Zelník, V; Golais, F

    2015-12-01

    In this study, we investigated the medium of three cell lines transformed with murine herpesvirus 68 (MHV-68) in vitro and in vivo, 68/HDF, 68/NIH3T3, and S11E, for the presence of compounds resembling growth factors of some herpesviruses which have displayed transforming and transformed phenotype suppressing activity in normal and tumor cells. When any of spent medium was added to cell culture we observed the onset of transformed phenotype in baby hamster kidney cells (BHK-21) cells and transformed phenotype suppressing activity in tumor human epithelial cells (HeLa). In media tested, we have identified the presence of putative growth factor related to MHV-68 (MHGF-68). Its bivalent properties have been blocked entirely by antisera against MHV-68 and two monoclonal antibodies against glycoprotein B (gB) of MHV-68 suggesting viral origin of MHGF-68. The results of initial efforts to separate MHGF-68 on FPLC Sephadex G15 column in the absence of salts revealed the loss of its transforming activity but transformed phenotype suppressing activity retained. On the other hand, the use of methanol-water mobile phase on RP-HPLC C18 column allowed separation of MHGF-68 to two compounds. Both separated fractions, had only the transforming activity to normal cells. Further experiments exploring the nature and the structure of hitherto unknown MHGF-68 are now in the progress to characterize its molecular and biological properties. PMID:26666191

  8. Steady-state nuclear actin levels are determined by export competent actin pool.

    PubMed

    Skarp, Kari-Pekka; Huet, Guillaume; Vartiainen, Maria K

    2013-10-01

    A number of studies in the last decade have irrevocably promoted actin into a fully fledged member of the nuclear compartment, where it, among other crucial tasks, facilitates transcription and chromatin remodeling. Changes in nuclear actin levels have been linked to different cellular processes: decreased nuclear actin to quiescence and increased nuclear actin to differentiation. Importin 9 and exportin 6 transport factors are responsible for the continuous nucleocytoplasmic shuttling of actin, but the mechanisms, which result in modulated actin levels, have not been characterized. We find that in cells growing under normal growth conditions, the levels of nuclear actin vary considerably from cell to cell. To understand the basis for this, we have extensively quantified several cellular parameters while at the same time recording the import and export rates of green fluorescent protein (GFP)-tagged actin. Surprisingly, our dataset shows that the ratio of nuclear to cytoplasmic fluorescence intensity, but not nuclear shape, size, cytoplasm size, or their ratio, correlates negatively with both import and export rate of actin. This suggests that high-nuclear actin content is maintained by both diminished import and export. The high nuclear actin containing cells still show high mobility of actin, but it is not export competent, suggesting increased binding of actin to nuclear complexes. Creation of such export incompetent actin pool would ensure enough actin is retained in the nucleus and make it available for the various nuclear functions described for actin. PMID:23749625

  9. Actinic reticuloid

    SciTech Connect

    Marx, J.L.; Vale, M.; Dermer, P.; Ragaz, A.; Michaelides, P.; Gladstein, A.H.

    1982-09-01

    A 58-year-old man has his condition diagnosed as actinic reticuloid on the basis of clinical and histologic findings and phototesting data. He had clinical features resembling mycosis fungoides in light-exposed areas. Histologic findings disclosed a bandlike infiltrate with atypical mononuclear cells in the dermis and scattered atypical cells in the epidermis. Electron microscopy disclosed mononuclear cells with bizarre, convoluted nuclei, resembling cerebriform cells of Lutzner. Phototesting disclosed a diminished minimal erythemal threshold to UV-B and UV-A. Microscopic changes resembling actinic reticuloid were reproduced in this patient 24 and 72 hours after exposure to 15 minimal erythemal doses of UV-B.

  10. Transforming growth factor-{beta}2 enhances differentiation of cardiac myocytes from embryonic stem cells

    SciTech Connect

    Kumar, Dinender . E-mail: Dinender.Kumar@uvm.edu; Sun, Baiming

    2005-06-24

    Stem cell therapy holds great promise for the treatment of injured myocardium, but is challenged by a limited supply of appropriate cells. Three different isoforms of transforming growth factor-{beta} (TGF-{beta}) -{beta}1, -{beta}2, and -{beta}3 exhibit distinct regulatory effects on cell growth, differentiation, and migration during embryonic development. We compared the effects of these three different isoforms on cardiomyocyte differentiation from embryonic stem (ES) cells. In contrast to TGF-{beta}1, or -{beta}3, treatment of mouse ES cells with TGF-{beta}2 isoform significantly increased embryoid body (EB) proliferation as well as the extent of the EB outgrowth that beat rhythmically. At 17 days, 49% of the EBs treated with TGF-{beta}2 exhibited spontaneous beating compared with 15% in controls. Cardiac myocyte specific protein markers sarcomeric myosin and {alpha}-actin were demonstrated in beating EBs and cells isolated from EBs. In conclusion, TGF-{beta}2 but not TGF-{beta}1, or -{beta}3 promotes cardiac myocyte differentiation from ES cells.

  11. Bidirectional actin transport is influenced by microtubule and actin stability.

    PubMed

    Chetta, Joshua; Love, James M; Bober, Brian G; Shah, Sameer B

    2015-11-01

    Local and long-distance transport of cytoskeletal proteins is vital to neuronal maintenance and growth. Though recent progress has provided insight into the movement of microtubules and neurofilaments, mechanisms underlying the movement of actin remain elusive, in large part due to rapid transitions between its filament states and its diverse cellular localization and function. In this work, we integrated live imaging of rat sensory neurons, image processing, multiple regression analysis, and mathematical modeling to perform the first quantitative, high-resolution investigation of GFP-actin identity and movement in individual axons. Our data revealed that filamentous actin densities arise along the length of the axon and move short but significant distances bidirectionally, with a net anterograde bias. We directly tested the role of actin and microtubules in this movement. We also confirmed a role for actin densities in extension of axonal filopodia, and demonstrated intermittent correlation of actin and mitochondrial movement. Our results support a novel mechanism underlying slow component axonal transport, in which the stability of both microtubule and actin cytoskeletal components influence the mobility of filamentous actin. PMID:26043972

  12. Waves of actin and microtubule polymerization drive microtubule-based transport and neurite growth before single axon formation

    PubMed Central

    Winans, Amy M; Collins, Sean R; Meyer, Tobias

    2016-01-01

    Many developing neurons transition through a multi-polar state with many competing neurites before assuming a unipolar state with one axon and multiple dendrites. Hallmarks of the multi-polar state are large fluctuations in microtubule-based transport into and outgrowth of different neurites, although what drives these fluctuations remains elusive. We show that actin waves, which stochastically migrate from the cell body towards neurite tips, direct microtubule-based transport during the multi-polar state. Our data argue for a mechanical control system whereby actin waves transiently widen the neurite shaft to allow increased microtubule polymerization to direct Kinesin-based transport and create bursts of neurite extension. Actin waves also require microtubule polymerization, arguing that positive feedback links these two components. We propose that actin waves create large stochastic fluctuations in microtubule-based transport and neurite outgrowth, promoting competition between neurites as they explore the environment until sufficient external cues can direct one to become the axon. DOI: http://dx.doi.org/10.7554/eLife.12387.001 PMID:26836307

  13. Plasma membrane-associated SCAR complex subunits promote cortical F-actin accumulation and normal growth characteristics in Arabidopsis roots

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The ARP2/3 complex, a highly conserved nucleator of F-actin polymerization, and its activator, the SCAR complex, have been shown to play important roles in leaf epidermal cell morphogenesis in Arabidopsis. However, the intracellular site(s) and function(s) of SCAR complex and ARP2/3 complex-depende...

  14. Vrp1p-Las17p interaction is critical for actin patch polarization but is not essential for growth or fluid phase endocytosis in S. cerevisiae.

    PubMed

    Wong, Ming Hwa; Meng, Lei; Rajmohan, Rajamuthiah; Yu, Shangjuan; Thanabalu, Thirumaran

    2010-12-01

    Vrp1p (yeast WIP) forms a protein complex with Las17p (yeast WASP), however the physiological significance of the interaction has not been fully characterized. Vrp1p residues, (788)MPKPR(792) are essential for Vrp1p-Las17p interaction. While C-Vrp1p(364-817) complements all the defects of the vrp1Δ strain, C-Vrp1p(364-817)(5A) ((788)AAAAA(792)) does not complement any of the defects, due to its inability to localize to cortical patches. Targeting C-Vrp1p(364-817)(5A) to membranes using CAAX motif (C-Vrp1p(364-817)(5A)-CAAX) rescued the growth and endocytosis defect but not the actin patch polarization defect of vrp1Δ. Vrp1p can localize to cortical patches, either by binding to Las17p through LBD (Las17 Binding Domain, Vrp1p(760-817)) or independent of Las17p through residues in N-Vrp1p(1-364). Unlike Vrp1p, Vrp1p(5A) localizes poorly to cortical patches and complements all the defects of vrp1Δ strain except actin patch polarization at elevated temperature. N-Vrp1p(1-364) complements all the defects of vrp1Δ strain except the actin patch polarization defect while N-Vrp1p(1-364)-LBD fusion protein complements all the defects. Thus our results show that while both Vrp1p and Las17p are essential for many cellular processes, the two proteins do not necessarily have to bind to each other to carry out these cellular functions. However, Las17p-Vrp1p interaction is essential for actin patch polarization at elevated temperature. PMID:20816901

  15. Transforming growth factor-beta and transforming growth factor beta-receptor expression in human meningioma cells.

    PubMed Central

    Johnson, M. D.; Federspiel, C. F.; Gold, L. I.; Moses, H. L.

    1992-01-01

    The transforming growth factor-beta (TGF beta) family in mammals includes three closely related peptides that influence proliferation and numerous physiologic processes in most mesenchymal cells. In this study, Northern blots, immunohistochemistry, TGF beta radioreceptor assays, TGF beta receptor affinity labeling and [3H] thymidine incorporation were used to evaluate whether primary cell cultures of human meningiomas synthesize the three TGF beta isoforms, bear TGF beta receptors, and respond to TGF beta. Transcripts for TGF beta 1 and 2 were detected in the three cases analyzed. Transforming growth factor-beta 1 immunoreactivity was detected in three of six cases, and TGF beta 2 and 3 immunoreactivity were detected in each case analyzed. Media conditioned by cells cultured from six meningiomas also contained latent TGF beta-like activity. Transforming growth factor-beta receptor cross-linking studies identified TGF beta binding sites corresponding to the type 1, type 2, and type 3 receptors on meningioma cells. Treatment with active TGF beta 1 produced a statistically significant reduction in [3H] thymidine incorporation after stimulation with 10% fetal calf serum and epidermal growth factor in all six cases studied. Images Figure 1 Figure 2 Figure 4 PMID:1325741

  16. Optogenetic control of PIP3: PIP3 is sufficient to induce the actin-based active part of growth cones and is regulated via endocytosis.

    PubMed

    Kakumoto, Toshiyuki; Nakata, Takao

    2013-01-01

    Phosphatidylinositol-3,4,5-trisphosphate (PIP3) is highly regulated in a spatiotemporal manner and plays multiple roles in individual cells. However, the local dynamics and primary functions of PIP3 in developing neurons remain unclear because of a lack of techniques for manipulating PIP3 spatiotemporally. We addressed this issue by combining optogenetic control and observation of endogenous PIP3 signaling. Endogenous PIP3 was abundant in actin-rich structures such as growth cones and "waves", and PIP3-rich plasma membranes moved actively within growth cones. To study the role of PIP3 in developing neurons, we developed a PI3K photoswitch that can induce production of PIP3 at specific locations upon blue light exposure. We succeeded in producing PIP3 locally in mouse hippocampal neurons. Local PIP3 elevation at neurite tips did not induce neurite elongation, but it was sufficient to induce the formation of filopodia and lamellipodia. Interestingly, ectopic PIP3 elevation alone activated membranes to form actin-based structures whose behavior was similar to that of growth-cone-like "waves". We also found that endocytosis regulates effective PIP3 concentration at plasma membranes. These results revealed the local dynamics and primary functions of PIP3, providing fundamental information about PIP3 signaling in neurons. PMID:23951027

  17. Connective tissue growth factor/CCN2-null mouse embryonic fibroblasts retain intact transforming growth factor-{beta} responsiveness

    SciTech Connect

    Mori, Yasuji; Hinchcliff, Monique; Wu, Minghua; Warner-Blankenship, Matthew; Lyons, Karen M.

    2008-03-10

    Background: The matricellular protein connective tissue growth factor (CCN2) has been implicated in pathological fibrosis, but its physiologic role remains elusive. In vitro, transforming growth factor-{beta} (TGF-{beta}) induces CCN2 expression in mesenchymal cells. Because CCN2 can enhance profibrotic responses elicited by TGF-{beta}, it has been proposed that CCN2 functions as an essential downstream signaling mediator for TGF-{beta}. To explore this notion, we characterized TGF-{beta}-induced activation of fibroblasts from CCN2-null (CCN2{sup -/-}) mouse embryos. Methods: The regulation of CCN2 expression was examined in vivo in a model of fibrosis induced by bleomycin. Cellular TGF-{beta} signal transduction and regulation of collagen gene expression were examined in CCN2{sup -/-} MEFs by immunohistochemistry, Northern, Western and RT-PCR analysis, immunocytochemistry and transient transfection assays. Results: Bleomycin-induced skin fibrosis in the mouse was associated with substantial CCN2 up-regulation in lesional fibroblasts. Whereas in vitro proliferation rate of CCN2{sup -/-} MEFs was markedly reduced compared to wild type MEFs, TGF-{beta}-induced activation of the Smad pathways, including Smad2 phosphorylation, Smad2/3 and Smad4 nuclear accumulation and Smad-dependent transcriptional responses, were unaffected by loss of CCN2. The stimulation of COL1A2 and fibronectin mRNA expression and promoter activity, and of corresponding protein levels, showed comparable time and dose-response in wild type and CCN2{sup -/-} MEFs, whereas stimulation of alpha smooth muscle actin and myofibroblast transdifferentiation showed subtle impairment in MEFs lacking CCN2. Conclusion: Whereas endogenous CCN2 plays a role in regulation of proliferation and TGF-{beta}-induced myofibroblast transdifferentiation, it appears to be dispensable for Smad-dependent stimulation of collagen and extracellular matrix synthesis in murine embryonic fibroblasts.

  18. Synthetic peptides that cause F-actin bundling and block actin depolymerization

    DOEpatents

    Sederoff, Heike; Huber, Steven C; Larabell, Carolyn A

    2011-10-18

    Synthetic peptides derived from sucrose synthase, and having homology to actin and actin-related proteins, sharing a common motif, useful for causing acting bundling and preventing actin depolymerization. Peptides exhibiting the common motif are described, as well as specific synthetic peptides which caused bundled actin and inhibit actin depolymerization. These peptides can be useful for treating a subject suffering from a disease characterized by cells having neoplastic growth, for anti-cancer therapeutics, delivered to subjects solely, or concomitantly or sequentially with other known cancer therapeutics. These peptides can also be used for stabilizing microfilaments in living cells and inhibiting growth of cells.

  19. Diosgenin attenuates hepatic stellate cell activation through transforming growth factor-β/Smad signaling pathway

    PubMed Central

    Xie, Wei-Lin; Jiang, Rong; Shen, Xiao-Lu; Chen, Zhi-Yu; Deng, Xiao-Ming

    2015-01-01

    Activation of hepatic stellate cells (HSC) plays a pivotal role in the development of hepatic fibrosis. Transforming growth factor-β1 (TGF-β1) is considered to be the main stimuli factor responsible for the activation of HSC. Diosgenin is a steroidal saponin found in several plants including Solanum and Dioscorea species, and it inhibited high glucose-induced renal tubular fibrosis. However, the effects of diosgenin against hepatic fibrosis remain elusive. Therefore, in this study, we investigated the effects of diosgenin on TGF-β1-induced HSCs and elucidate the possible mechanism of its anti-fibrotic effect. Our results demonstrated that diosgenin inhibited TGF-β1-induced HSC proliferation, reduced the expression of collagen I and α-smooth muscle actin (α-SMA), as well as the expression of TGF-β receptor I (TGF-β RI) and II. Moreover, diosgenin suppressed TGF-β1-induced phosphorylation of Smad3 in HSCs. In conclusion, our data demonstrate that diosgenin inhibited HSC-T6 cell proliferation and activation, at least in part, via the TGF-β1/Smad signaling pathway. These results provide that diosgenin may have potential to treat liver fibrosis. PMID:26884947

  20. Diosgenin attenuates hepatic stellate cell activation through transforming growth factor-β/Smad signaling pathway.

    PubMed

    Xie, Wei-Lin; Jiang, Rong; Shen, Xiao-Lu; Chen, Zhi-Yu; Deng, Xiao-Ming

    2015-01-01

    Activation of hepatic stellate cells (HSC) plays a pivotal role in the development of hepatic fibrosis. Transforming growth factor-β1 (TGF-β1) is considered to be the main stimuli factor responsible for the activation of HSC. Diosgenin is a steroidal saponin found in several plants including Solanum and Dioscorea species, and it inhibited high glucose-induced renal tubular fibrosis. However, the effects of diosgenin against hepatic fibrosis remain elusive. Therefore, in this study, we investigated the effects of diosgenin on TGF-β1-induced HSCs and elucidate the possible mechanism of its anti-fibrotic effect. Our results demonstrated that diosgenin inhibited TGF-β1-induced HSC proliferation, reduced the expression of collagen I and α-smooth muscle actin (α-SMA), as well as the expression of TGF-β receptor I (TGF-β RI) and II. Moreover, diosgenin suppressed TGF-β1-induced phosphorylation of Smad3 in HSCs. In conclusion, our data demonstrate that diosgenin inhibited HSC-T6 cell proliferation and activation, at least in part, via the TGF-β1/Smad signaling pathway. These results provide that diosgenin may have potential to treat liver fibrosis. PMID:26884947

  1. Osteogenic Responses in Fibroblasts Activated by Elastin Degradation Products and Transforming Growth Factor-β1

    PubMed Central

    Simionescu, Agneta; Simionescu, Dan T.; Vyavahare, Narendra R.

    2007-01-01

    Our objective was to establish the role of fibroblasts in medial vascular calcification, a pathological process known to be associated with elastin degradation and remodeling. Rat dermal fibroblasts were treated in vitro with elastin degradation products and transforming growth factor (TGF)-β1, factors usually present in deteriorated matrix environments. Cellular changes were monitored at the gene and protein level by reverse transcriptase-polymerase chain reaction, enzyme-linked immunosorbent assay, immunofluorescence, and von Kossa staining for calcium deposits. By 21 days, multicellular calcified nodules were formed in the presence of elastin degradation products and TGF-β1 separately and to a significantly greater extent when used together. Before mineralization, cells expressed α-smooth muscle actin and large amounts of collagen type I and matrix metalloproteinase-2, characteristic features of myofibroblasts, key elements in tissue remodeling and repair. Stimulated cells expressed increased levels of core-binding factor α1, osteocalcin, alkaline phosphatase, and osteoprotegerin, representative bone-regulating proteins. For most proteins analyzed, TGF-β1 synergistically amplified responses of fibroblasts to elastin degradation products. In conclusion, elastin degradation products and TGF-β1 promote myofibroblastic and osteogenic differentiation in fibroblasts. These results support the idea that elastin-related calcification involves dynamic remodeling events and suggest the possibility of a defective tissue repair process. PMID:17591959

  2. Transforming growth factor-beta stimulates epithelial-mesenchymal transformation in the proepicardium.

    PubMed

    Olivey, Harold E; Mundell, Nathan A; Austin, Anita F; Barnett, Joey V

    2006-01-01

    The proepicardium (PE) migrates over the heart and forms the epicardium. A subset of these PE-derived cells undergoes epithelial-mesenchymal transformation (EMT) and gives rise to cardiac fibroblasts and components of the coronary vasculature. We report that transforming growth factor-beta (TGFbeta) 1 and TGFbeta2 increase EMT in PE explants as measured by invasion into a collagen gel, loss of cytokeratin expression, and redistribution of ZO1. The type I TGFbeta receptors ALK2 and ALK5 are both expressed in the PE. However, only constitutively active (ca) ALK2 stimulates PE-derived epithelial cell activation, the first step in transformation, whereas caALK5 stimulates neither activation nor transformation in PE explants. Overexpression of Smad6, an inhibitor of ALK2 signaling, inhibits epithelial cell activation, whereas BMP7, a known ligand for ALK2, has no effect. These data demonstrate that TGFbeta stimulates transformation in the PE and suggest that ALK2 partially mediates this effect. PMID:16245329

  3. Nanowire growth by an electron beam induced massive phase transformation

    SciTech Connect

    Sood, Shantanu; Kisslinger, Kim; Gouma, Perena

    2014-11-15

    Tungsten trioxide nanowires of a high aspect ratio have been synthesized in-situ in a TEM under an electron beam of current density 14A/cm² due to a massive polymorphic reaction. Sol-gel processed pseudocubic phase nanocrystals of tungsten trioxide were seen to rapidly transform to one dimensional monoclinic phase configurations, and this reaction was independent of the substrate on which the material was deposited. The mechanism of the self-catalyzed polymorphic transition and accompanying radical shape change is a typical characteristic of metastable to stable phase transformations in nanostructured polymorphic metal oxides. A heuristic model is used to confirm the metastable to stable growth mechanism. The findings are important to the control electron beam deposition of nanowires for functional applications starting from colloidal precursors.

  4. Nanowire growth by an electron beam induced massive phase transformation

    DOE PAGESBeta

    Sood, Shantanu; Kisslinger, Kim; Gouma, Perena

    2014-11-15

    Tungsten trioxide nanowires of a high aspect ratio have been synthesized in-situ in a TEM under an electron beam of current density 14A/cm² due to a massive polymorphic reaction. Sol-gel processed pseudocubic phase nanocrystals of tungsten trioxide were seen to rapidly transform to one dimensional monoclinic phase configurations, and this reaction was independent of the substrate on which the material was deposited. The mechanism of the self-catalyzed polymorphic transition and accompanying radical shape change is a typical characteristic of metastable to stable phase transformations in nanostructured polymorphic metal oxides. A heuristic model is used to confirm the metastable to stablemore » growth mechanism. The findings are important to the control electron beam deposition of nanowires for functional applications starting from colloidal precursors.« less

  5. Tau co-organizes dynamic microtubule and actin networks

    PubMed Central

    Elie, Auréliane; Prezel, Elea; Guérin, Christophe; Denarier, Eric; Ramirez-Rios, Sacnicte; Serre, Laurence; Andrieux, Annie; Fourest-Lieuvin, Anne; Blanchoin, Laurent; Arnal, Isabelle

    2015-01-01

    The crosstalk between microtubules and actin is essential for cellular functions. However, mechanisms underlying the microtubule-actin organization by cross-linkers remain largely unexplored. Here, we report that tau, a neuronal microtubule-associated protein, binds to microtubules and actin simultaneously, promoting in vitro co-organization and coupled growth of both networks. By developing an original assay to visualize concomitant microtubule and actin assembly, we show that tau can induce guided polymerization of actin filaments along microtubule tracks and growth of single microtubules along actin filament bundles. Importantly, tau mediates microtubule-actin co-alignment without changing polymer growth properties. Mutagenesis studies further reveal that at least two of the four tau repeated motifs, primarily identified as tubulin-binding sites, are required to connect microtubules and actin. Tau thus represents a molecular linker between microtubule and actin networks, enabling a coordination of the two cytoskeletons that might be essential in various neuronal contexts. PMID:25944224

  6. Actin Interacting Protein1 and Actin Depolymerizing Factor Drive Rapid Actin Dynamics in Physcomitrella patens[W

    PubMed Central

    Augustine, Robert C.; Pattavina, Kelli A.; Tüzel, Erkan; Vidali, Luis; Bezanilla, Magdalena

    2011-01-01

    The remodeling of actin networks is required for a variety of cellular processes in eukaryotes. In plants, several actin binding proteins have been implicated in remodeling cortical actin filaments (F-actin). However, the extent to which these proteins support F-actin dynamics in planta has not been tested. Using reverse genetics, complementation analyses, and cell biological approaches, we assessed the in vivo function of two actin turnover proteins: actin interacting protein1 (AIP1) and actin depolymerizing factor (ADF). We report that AIP1 is a single-copy gene in the moss Physcomitrella patens. AIP1 knockout plants are viable but have reduced expansion of tip-growing cells. AIP1 is diffusely cytosolic and functions in a common genetic pathway with ADF to promote tip growth. Specifically, ADF can partially compensate for loss of AIP1, and AIP1 requires ADF for function. Consistent with a role in actin remodeling, AIP1 knockout lines accumulate F-actin bundles, have fewer dynamic ends, and have reduced severing frequency. Importantly, we demonstrate that AIP1 promotes and ADF is essential for cortical F-actin dynamics. PMID:22003077

  7. Rho, nuclear actin, and actin-binding proteins in the regulation of transcription and gene expression

    PubMed Central

    Rajakylä, Eeva Kaisa; Vartiainen, Maria K

    2014-01-01

    Actin cytoskeleton is one of the main targets of Rho GTPases, which act as molecular switches on many signaling pathways. During the past decade, actin has emerged as an important regulator of gene expression. Nuclear actin plays a key role in transcription, chromatin remodeling, and pre-mRNA processing. In addition, the “status” of the actin cytoskeleton is used as a signaling intermediate by at least the MKL1-SRF and Hippo-pathways, which culminate in the transcriptional regulation of cytoskeletal and growth-promoting genes, respectively. Rho GTPases may therefore regulate gene expression by controlling either cytoplasmic or nuclear actin dynamics. Although the regulation of nuclear actin polymerization is still poorly understood, many actin-binding proteins, which are downstream effectors of Rho, are found in the nuclear compartment. In this review, we discuss the possible mechanisms and key proteins that may mediate the transcriptional regulation by Rho GTPases through actin. PMID:24603113

  8. Rho, nuclear actin, and actin-binding proteins in the regulation of transcription and gene expression.

    PubMed

    Rajakylä, Eeva Kaisa; Vartiainen, Maria K

    2014-01-01

    Actin cytoskeleton is one of the main targets of Rho GTPases, which act as molecular switches on many signaling pathways. During the past decade, actin has emerged as an important regulator of gene expression. Nuclear actin plays a key role in transcription, chromatin remodeling, and pre-mRNA processing. In addition, the "status" of the actin cytoskeleton is used as a signaling intermediate by at least the MKL1-SRF and Hippo-pathways, which culminate in the transcriptional regulation of cytoskeletal and growth-promoting genes, respectively. Rho GTPases may therefore regulate gene expression by controlling either cytoplasmic or nuclear actin dynamics. Although the regulation of nuclear actin polymerization is still poorly understood, many actin-binding proteins, which are downstream effectors of Rho, are found in the nuclear compartment. In this review, we discuss the possible mechanisms and key proteins that may mediate the transcriptional regulation by Rho GTPases through actin. PMID:24603113

  9. Actinic Prurigo.

    PubMed

    Rodríguez-Carreón, Alma Angélica; Rodríguez-Lobato, Erika; Rodríguez-Gutiérrez, Georgina; Cuevas-González, Juan Carlos; Mancheno-Valencia, Alexandra; Solís-Arias, Martha Patricia; Vega-Memije, María Elisa; Hojyo-Tomoka, María Teresa; Domínguez-Soto, Luciano

    2015-01-01

    Actinic prurigo is an idiopathic photodermatosis that affects the skin, as well as the labial and conjunctival mucosa in indigenous and mestizo populations of Latin America. It starts predominantly in childhood, has a chronic course, and is exacerbated with solar exposure. Little is known of its pathophysiology, including the known mechanisms of the participation of HLA-DR4 and an abnormal immunologic response with increase of T CD4+ lymphocytes. The presence of IgE, eosinophils, and mast cells suggests that it is a hypersensitivity reaction (likely type IVa or b). The diagnosis is clinical, and the presence of lymphoid follicles in the mucosal histopathologic study of mucosa is pathognomonic. The best available treatment to date is thalidomide, despite its secondary effects. PMID:26861426

  10. Transforming growth factor (TGF)-. alpha. in human milk

    SciTech Connect

    Okada, Masaki; Wakai, Kae; Shizume, Kazuo ); Iwashita, Mitsutoshi ); Ohmura, Eiji; Kamiya, Yoshinobu; Murakami, Hitomi; Onoda, Noritaka; Tsushima, Toshio

    1991-01-01

    Transforming growth factor (TGF)-{alpha} and epidermal growth factor (EGF) were measured in human milk by means of homologous radioimmunoassay. As previously reported, EGF concentration in the colostrum was approximately 200 ng/ml and decreased to 50 ng/ml by day 7 postpartum. The value of immunoreactive (IR)-TGF-{alpha} was 2.2-7.2 ng/ml, much lower than that of EGF. In contrast to EGF, the concentration of IR-TGF-{alpha} was fairly stable during the 7 postpartum days. There was no relationship between the concentrations of IR-TGF-{alpha} and IR-EGF, suggesting that the regulatory mechanism in the release of the two growth factors is different. On gel-chromatography using a Sephadex G-50 column, IR-EGF appeared in the fraction corresponding to that of authentic human EGF, while 70%-80% of the IR-TGF-{alpha} was eluted as a species with a molecular weight greater than that of authentic human TGF-{alpha}. Although the physiological role of TGF-{alpha} in milk is not known, it is possible that it is involved in the development of the mammary gland and/or the growth of newborn infants.

  11. [Actinic Keratosis].

    PubMed

    Dejaco, D; Hauser, U; Zelger, B; Riechelmann, H

    2015-07-01

    Actinic keratosis is a cutaneous lesion characterized by proliferation of atypical epidermal keratinocytes due to prolonged exposure to exogenous factors such as ultraviolet radiation. AKs are in-situ-squamous cell carcinomas (PEC) of the skin. AK typically presents as erythematous, scaly patch or papule (classic AK), occasionally as thick, adherent scale on an erythematous base. Mostly fair-skinned adults are affected. AKs typically occur in areas of frequent sun exposure (balding scalp, face, "H-region", lateral neck, décolleté, dorsum of the hand and lower extremities). Actinic Cheilitis is the term used for AKs appearing on the lips. The diagnosis of AK is based on clinical examination including inspection and palpation. The typical palpable rough surface of AK often precedes a visible lesion. Dermoscopy may provide additional information. If diagnosis is uncertain and invasion suspected, biopsy and histopathologic evaluation should be performed. The potential for progression to invasive PECs mandates therapeutic intervention. Treatment options include topical and systemic therapies. Topical therapies are classified into physical, medical and combined physical-chemical approaches and a sequential combination of treatment modalities is possible. Topical-physical cryotherapy is the treatment of choice for isolated, non-hypertrophic AK. Topical-medical treatment, e. g. 5-fluoruracil (5FU) cream or Imiquomod or Ingenolmebutat application is used for multiple, non-hypertrophic AKs. For hypertrophic AKs, a dehorning pretreatment with salicinated vaseline is recommended. Isolated hypertrophic AKs often need cryotherapy with prolonged freezing time or several consecutive applications. Sequentially combined approaches are recommended for multiple, hypertrophic AKs. Photodynamic therapy (PDT) as example for a combined physical-chemical approach is an established treatment for multiple, non-hypertrophic and hypertrophic AKs. Prevention includes avoidance of sun and

  12. Basic science for the clinician 57: transforming growth factor β.

    PubMed

    Sigal, Leonard H

    2012-08-01

    As is so often the case, a molecule gets named for its first identified activity or apparent role and then that initial name sticks, even as new and perhaps fundamentally different activities emerge from later studies. It is the special power of evolution that takes a certain activity and then uses it over and over again in pursuit of apparently disparate goals in a maturing or mature organism. In general terms, transforming growth factor β (TGF-β) is intimately involved in a variety of differentiation and growth inhibition processes, in apoptosis, and in deposition of the extracellular matrix. Initially identified in its role in oncogenesis, TGF-β is now implicated in a number of vascular and rheumatologic disorders, perhaps most notably the scleroderma. TGF-β has been identified as a powerful influence in angiogenesis, wound healing, joint inflammation, tumor growth and metastasis, and, of course, immunoregulation. So "what is in a name?" A rose by any other name would smell as sweet and would still be immunologically active, even if the name is "misleading." PMID:22832292

  13. The latent transforming growth factor beta binding protein (LTBP) family.

    PubMed Central

    Oklü, R; Hesketh, R

    2000-01-01

    The transforming growth factor beta (TGFbeta) cytokines are a multi-functional family that exert a wide variety of effects on both normal and transformed mammalian cells. The secretion and activation of TGFbetas is regulated by their association with latency-associated proteins and latent TGFbeta binding proteins (LTBPs). Over the past few years, three members of the LTBP family have been identified, in addition to the protoype LTBP1 first sequenced in 1990. Three of the LTBP family are expressed in a variety of isoforms as a consequence of alternative splicing. This review summarizes the differences between the isoforms in terms of the effects on domain structure and hence possible function. The close identity between LTBPs and members of the fibrillin family, mutations in which have been linked directly to Marfan's syndrome, suggests that anomalous expression of LTBPs may be associated with disease. Recent data indicating that differential expression of LTBP1 isoforms occurs during the development of coronary heart disease is considered, together with evidence that modulation of LTBP function, and hence of TGFbeta activity, is associated with a variety of cancers. PMID:11104663

  14. Role of growth factors in the growth of normal and transformed cells

    SciTech Connect

    Lokeshwar, V.B.

    1989-01-01

    Growth factors play an important role in the growth of normal cells. However, their untimely and/or excess production leads to neoplastic transformation. The role of growth factors in the growth of normal cells was studied by investigating the mechanism of transmodulation of the cell surface EGF receptor number by protamine. Protamine increased the EGF stimulated mitogenic response in Swiss mouse 3T3 cells and A431 cells by increasing the number of functionally active EGF receptors. Protamine also increased EGF receptor number in plasma membranes and solubilized membranes. This was evidenced by an increase in both {sup 125}I-EGF-EGF-receptor complex and EGF stimulated phosphorylation of the EGF receptor. The solubilized EGF receptor was retained on a protamine-agarose gel indicating that protamine might increase EGF receptor number by directly activating cryptic EGF receptors in the plasma membranes. The role of growth factors in neoplastic transformation was studied by investigating the role of the oncogene v-sis in the growth of Simian sarcoma virus (SSV) transformed cells. The product of the oncogene v-sis is 94% homologous to the B chain of PDGF. This study found that (i) v-sis gene product is synthesized as a 32 kDa unglycosylated monomer which is glycosylated, dimerized and proteolytically processed into p36, p72, p68, p58, p44 and p27 mol. wt. species respectively. (ii) p36, p72, p68 and p58 are very likely formed in the endoplasmic reticulum and/or Golgi complex. A fraction of newly synthesized p72, p68 and p58 is degraded intracellularly at a fast rate. (iii) p44 is a secretory product which remains tightly associated with the cell surface. p44 is recaptured by the cells through interaction with cell surface PDGF receptors and degraded into p27. (iv) During long term cultures p44 is extracellularly cleaved into a 27 kDa product.

  15. Effects of transforming growth factor type beta on expression of cytoskeletal proteins in endosteal mouse osteoblastic cells

    SciTech Connect

    Lomri, A.; Marie, P.J. )

    1990-01-01

    Transforming growth factor beta (TGF beta) has been shown to influence the growth and differentiation of many cell types in vitro. We have examined the effects of TGF beta on cell morphology and cytoskeletal organization in relation to parameters of cell proliferation and differentiation in endosteal osteoblastic cells isolated from mouse caudal vertebrae. Treatment of mouse osteoblastic cells cultured in serum free medium for 24 hours with TGF beta (1.5-30 ng/mL) slightly (-23%) inhibited alkaline phosphatase activity. In parallel, TGF beta (0.5-30 ng/mL, 24 hours) greatly increased cell replication as evaluated by (3H)-thymidine incorporation into DNA (157% to 325% of controls). At a median dose (1.5 ng/mL) that affected both alkaline phosphatase and DNA synthesis (235% of controls) TGF beta induced rapid (six hours) cell respreading of quiescent mouse osteoblastic cells. This effect was associated with increased polymerization of actin, alpha actinin, and tubulins, as evaluated by both biochemical and immunofluorescence methods. In addition, TGF beta (1.5 ng/mL) increased the de novo biosynthesis of actin, alpha actinin, vimentin, and tubulins, as determined by {sup 35}S methionine labeling and fractionation of cytoskeletal proteins using two-dimensional gel electrophoresis. These effects were rapid and transient, as they occurred at six hours and were reversed after 24 hours of TGF beta exposure. The results indicate that the stimulatory effect of TGF beta on DNA synthesis in endosteal mouse osteoblastic cells is associated with a transient increase in cell spreading associated with enhanced polymerization and synthesis of cytoskeletal proteins.

  16. Zoledronic acid suppresses transforming growth factor-β-induced fibrogenesis by human gingival fibroblasts

    PubMed Central

    KOMATSU, YUKO; IBI, MIHO; CHOSA, NAOYUKI; KYAKUMOTO, SEIKO; KAMO, MASAHARU; SHIBATA, TOSHIYUKI; SUGIYAMA, YOSHIKI; ISHISAKI, AKIRA

    2016-01-01

    Bisphosphonates (BPs) are analogues of pyro-phosphate that are known to prevent bone resorption by inhibiting osteoclast activity. Nitrogen-containing BPs, such as zoledronic acid (ZA), are widely used in the treatment of osteoporosis and bone metastasis. However, despite having benefits, ZA has been reported to induce BP-related osteonecrosis of the jaw (BRONJ) in cancer patients. The molecular pathological mechanisms responsible for the development of BRONJ, including necrotic bone exposure after tooth extraction, remain to be elucidated. In this study, we examined the effects of ZA on the transforming growth factor-β (TGF-β)-induced myofibroblast (MF) differentiation of human gingival fibroblasts (hGFs) and the migratory activity of hGFs, which are important for wound closure by fibrous tissue formation. The ZA maximum concentration in serum (Cmax) was found to be approximately 1.47 µM, which clinically, is found after the intravenous administration of 4 mg ZA, and ZA at this dose is considered appropriate for the treatment of cancer bone metastasis or bone diseases, such as Erdheim-Chester disease. At Cmax, ZA significantly suppressed i) the TGF-β-induced promotion of cell viability, ii) the TGF-β-induced expression of MF markers such as α-smooth muscle actin (α-SMA) and type I collagen, iii) the TGF-β-induced migratory activity of hGFs and iv) the expression level of TGF-β type I receptor on the surfaces of hGFs, as well as the TGF-β-induced phosphorylation of Smad2/3. Thus, ZA suppresses TGF-β-induced fibrous tissue formation by hGFs, possibly through the inhibition of Smad-dependent signal transduction. Our findings partly elucidate the molecular mechanisms underlying BRONJ and may prove to be beneficial to the identification of drug targets for the treatment of this symptom at the molecular level. PMID:27176567

  17. Increased transforming growth factor beta 1 expression mediates ozone-induced airway fibrosis in mice

    PubMed Central

    Katre, Ashwini; Ballinger, Carol; Akhter, Hasina; Fanucchi, Michelle; Kim, Dae-Kee; Postlethwait, Edward; Liu, Rui-Ming

    2013-01-01

    Ozone (O3), a commonly encountered environmental pollutant, has been shown to induce pulmonary fibrosis in different animal models; the underlying mechanism, however, remains elusive. To investigate the molecular mechanism underlying O3-induced pulmonary fibrosis, 6- to 8-week-old C57BL/6 male mice were exposed to a cyclic O3 exposure protocol consisting of 2 days of filtered air and 5 days of O3 exposure (0.5 ppm, 8 h/day) for 5 and 10 cycles with or without intraperitoneal injection of IN-1233, a specific inhibitor of the type 1 receptor of transforming growth factor beta (TGF-β), the most potent profibrogenic cytokine. The results showed that O3 exposure for 5 or 10 cycles increased the TGF-β protein level in the epithelial lining fluid (ELF), associated with an increase in the expression of plasminogen activator inhibitor 1 (PAI-1), a TGF-β-responsive gene that plays a critical role in the development of fibrosis under various pathological conditions. Cyclic O3 exposure also increased the deposition of collagens and alpha smooth muscle actin (α-SMA) in airway walls. However, these fibrotic changes were not overt until after 10 cycles of O3 exposure. Importantly, blockage of the TGF-β signaling pathway with IN-1233 suppressed O3-induced Smad2/3 phosphorylation, PAI-1 expression, as well as collagens and α-SMA deposition in the lung. Our data demonstrate for the first time that O3 exposure increases TGF-β expression and activates TGF-β signaling pathways, which mediates O3-induced lung fibrotic responses in vivo. PMID:21689010

  18. Dedifferentiation of Immortalized Human Podocytes in Response to Transforming Growth Factor-β

    PubMed Central

    Herman-Edelstein, Michal; Thomas, Merlin C.; Thallas-Bonke, Vicki; Saleem, Moin; Cooper, Mark E.; Kantharidis, Phillip

    2011-01-01

    OBJECTIVE Diabetic nephropathy is associated with dedifferentiation of podocytes, losing the specialized features required for efficient glomerular function and acquiring a number of profibrotic, proinflammatory, and proliferative features. These result from tight junction and cytoskeletal rearrangement, augmented proliferation, and apoptosis. RESEARCH DESIGN AND METHODS Experiments were performed in conditionally immortalized human podocytes developed by transfection with the temperature-sensitive SV40-T gene. Cells were then cultured in the presence of transforming growth factor (TGF)-β1 or angiotensin II in the presence or absence of a selective inhibitor of the TGF-β type I receptor kinase, SB-431542. Gene and protein expression were then examined by real-time RT-PCR and immunofluorescence, and correlated with changes observed in vivo in experimental diabetes. RESULTS Treatment of cells with TGF-β1 resulted in dynamic changes in their morphology, starting with retraction and shortening of foot processes and finishing with the formation of broad and complex tight junctions between adjacent podocytes. This dedifferentiation was also associated with dose- and time-dependent reduction in the expression of glomerular epithelial markers (nephrin, p-cadherin, zonnula occludens-1) and increased expression of mesenchymal markers (α−smooth muscle actin, vimentin, nestin), matrix components (fibronectin, collagen I, and collagen IV α3), cellular proliferation, and apoptosis. The induction of diabetes in mice was also associated with similar changes in morphology, protein expression, and proliferation in glomerular podocytes. CONCLUSIONS In response to TGF-β and other TGF-dependent stimuli, mature podocytes undergo dedifferentiation that leads to effacement of foot processes, morphologic flattening, and increased formation of intercellular tight junctions. This simplification of their phenotype to a more embryonic form is also associated with reentry of mature

  19. New Aspects of Progesterone Interactions with the Actin Cytoskeleton and Neurosteroidogenesis in the Cerebellum and the Neuronal Growth Cone

    PubMed Central

    Wessel, Lisa; Olbrich, Laura; Brand-Saberi, Beate

    2014-01-01

    The impact of progesterone on neuronal tissues in the central (CNS) and peripheral (PNS) nervous system is of significant scientific and therapeutic interest. Glial and neuronal cells of vertebrates express steroidogenic enzymes, and are able to synthesize progesterone de novo from cholesterol. Progesterone is described to have neuroprotective, neuroreparative, anti-degenerative, and anti-apoptotic effects in the CNS and the PNS. Thus, the first clinical studies promise new therapeutic options using progesterone in the treatment of patients with traumatic brain injury. Additionally, experimental data from different animal models suggest further positive effects of progesterone on neurological diseases such as cerebral ischemia, peripheral nerve injury and amyothropic lateral sclerosis. In regard to this future clinical use of progesterone, we discuss in this review the underlying physiological principles of progesterone effects in neuronal tissues. Mechanisms leading to morphological reorganizations of neurons in the CNS and PNS affected by progesterone are addressed, with special focus on the actin cytoskeleton. Furthermore, new aspects of a progesterone-dependent regulation of neurosteroidogenesis mediated by the recently described progesterone binding protein PGRMC1 in the nervous system are discussed. PMID:25141866

  20. Actin-resistant DNAse I Expression From Oncolytic Adenovirus Enadenotucirev Enhances Its Intratumoral Spread and Reduces Tumor Growth.

    PubMed

    Tedcastle, Alison; Illingworth, Sam; Brown, Alice; Seymour, Leonard W; Fisher, Kerry D

    2016-04-01

    Spread of oncolytic viruses through tumor tissue is essential to effective virotherapy. Interstitial matrix is thought to be a significant barrier to virus particle convection between "islands" of tumor cells. One way to address this is to encode matrix-degrading enzymes within oncolytic viruses, for secretion from infected cells. To test the hypothesis that extracellular DNA provides an important barrier, we assessed the ability of DNase to promote virus spread. Nonreplicating Ad5 vectors expressing actin-resistant DNase (aDNAse I), proteinase K (PK), hyaluronidase (rhPH20), and chondroitinase ABC (CABC) were injected into established DLD human colorectal adenocarcinoma xenografts, transcomplemented with a replicating Ad5 virus. Each enzyme improved oncolysis by the replicating adenovirus, with no evidence of tumor cells being shed into the bloodstream. aDNAse I and rhPH20 hyaluronidase were then cloned into conditionally-replicating group B adenovirus, Enadenotucirev (EnAd). EnAd encoding each enzyme showed significantly better antitumor efficacy than the parental virus, with the aDNAse I-expressing virus showing improved spread. Both DNase and hyaluronidase activity was still measurable 32 days postinfection. This is the first time that extracellular DNA has been implicated as a barrier for interstitial virus spread, and suggests that oncolytic viruses expressing aDNAse I may be promising candidates for clinical translation. PMID:26708004

  1. Transforming growth factor-{beta}-inducible phosphorylation of Smad3.

    PubMed

    Wang, Guannan; Matsuura, Isao; He, Dongming; Liu, Fang

    2009-04-10

    Smad proteins transduce the transforming growth factor-beta (TGF-beta) signal at the cell surface into gene regulation in the nucleus. Upon TGF-beta treatment, the highly homologous Smad2 and Smad3 are phosphorylated by the TGF-beta receptor at the SSXS motif in the C-terminal tail. Here we show that in addition to the C-tail, three (S/T)-P sites in the Smad3 linker region, Ser(208), Ser(204), and Thr(179) are phosphorylated in response to TGF-beta. The linker phosphorylation peaks at 1 h after TGF-beta treatment, behind the peak of the C-tail phosphorylation. We provide evidence suggesting that the C-tail phosphorylation by the TGF-beta receptor is necessary for the TGF-beta-induced linker phosphorylation. Although the TGF-beta receptor is necessary for the linker phosphorylation, the receptor itself does not phosphorylate these sites. We further show that ERK is not responsible for TGF-beta-dependent phosphorylation of these three sites. We show that GSK3 accounts for TGF-beta-inducible Ser(204) phosphorylation. Flavopiridol, a pan-CDK inhibitor, abolishes TGF-beta-induced phosphorylation of Thr(179) and Ser(208), suggesting that the CDK family is responsible for phosphorylation of Thr(179) and Ser(208) in response to TGF-beta. Mutation of the linker phosphorylation sites to nonphosphorylatable residues increases the ability of Smad3 to activate a TGF-beta/Smad-target gene as well as the growth-inhibitory function of Smad3. Thus, these observations suggest that TGF-beta-induced phosphorylation of Smad3 linker sites inhibits its antiproliferative activity. PMID:19218245

  2. Dendritic Actin Filament Nucleation Causes Traveling Waves and Patches

    NASA Astrophysics Data System (ADS)

    Carlsson, Anders E.

    2010-06-01

    The polymerization of actin via branching at a cell membrane containing nucleation-promoting factors is simulated using a stochastic-growth methodology. The polymerized-actin distribution displays three types of behavior: (a) traveling waves, (b) moving patches, and (c) random fluctuations. Increasing actin concentration causes a transition from patches to waves. The waves and patches move by a treadmilling mechanism not involving myosin II. The effects of downregulation of key proteins on actin wave behavior are evaluated.

  3. Vitamin D inhibition of pro-fibrotic effects of transforming growth factor β1 in lung fibroblasts and epithelial cells

    PubMed Central

    Ramirez, Allan M.; Wongtrakool, Cherry; Welch, Teresa; Steinmeyer, Andreas; Zügel, Ulrich; Roman, Jesse

    2010-01-01

    The mechanisms that control fibroproliferation and matrix deposition in lung fibrosis remain unclear. We speculate that vitamin D deficiency may contribute to pulmonary fibrosis since vitamin D deficiency has been implicated in several diseases. First, we confirmed the presence of vitamin D receptors (VDR) in cultured NIH/3T3 and lung fibroblasts. Fibroblasts transfected with a vitamin D response element – reporter construct and exposed to the active vitamin D metabolite, 1,25(OH)2D3, showed increased promoter activity indicating VDR functionality in these cells. Testing the effects of 1,25(OH)2D3 on fibroblasts treated with transforming growth factor β1 (TGFβ1), considered a driver of many fibrotic disorders, we found that 1,25(OH)2D3 inhibited TGFβ1-induced fibroblast proliferation in a dose-dependent fashion. 1,25(OH)2D3 also inhibited TGFβ1 stimulation of α-smooth muscle actin expression and polymerization and prevented the upregulation of fibronectin and collagen in TGFβ1-treated fibroblasts. Finally, we examined how 1,25(OH)2D3 affects epithelial-mesenchymal transformation of lung epithelial cells upon exposure to TGFβ1. We showed that the TGFβ1-induced upregulation of mesenchymal cell markers and abnormal expression of epithelial cell markers were blunted by 1,25(OH)2D3. These observations suggest that under TGFβ1 stimulation, 1,25(OH)2D3 inhibits the profibrotic phenotype of lung fibroblasts and epithelial cells. PMID:19931390

  4. The Neuroprotective Functions of Transforming Growth Factor Beta Proteins

    PubMed Central

    Dobolyi, Arpád; Vincze, Csilla; Pál, Gabriella; Lovas, Gábor

    2012-01-01

    Transforming growth factor beta (TGF-β) proteins are multifunctional cytokines whose neural functions are increasingly recognized. The machinery of TGF-β signaling, including the serine kinase type transmembrane receptors, is present in the central nervous system. However, the 3 mammalian TGF-β subtypes have distinct distributions in the brain suggesting different neural functions. Evidence of their involvement in the development and plasticity of the nervous system as well as their functions in peripheral organs suggested that they also exhibit neuroprotective functions. Indeed, TGF-β expression is induced following a variety of types of brain tissue injury. The neuroprotective function of TGF-βs is most established following brain ischemia. Damage in experimental animal models of global and focal ischemia was shown to be attenuated by TGF-βs. In addition, support for their neuroprotective actions following trauma, sclerosis multiplex, neurodegenerative diseases, infections, and brain tumors is also accumulating. The review will also describe the potential mechanisms of neuroprotection exerted by TGF-βs including anti-inflammatory, -apoptotic, -excitotoxic actions as well as the promotion of scar formation, angiogenesis, and neuroregeneration. The participation of these mechanisms in the neuroprotective effects of TGF-βs during different brain lesions will also be discussed. PMID:22942700

  5. Arabidopsis VILLIN2 and VILLIN3 are required for the generation of thick actin filament bundles and for directional organ growth.

    PubMed

    van der Honing, Hannie S; Kieft, Henk; Emons, Anne Mie C; Ketelaar, Tijs

    2012-03-01

    In plant cells, actin filament bundles serve as tracks for myosin-dependent organelle movement and play a role in the organization of the cytoplasm. Although virtually all plant cells contain actin filament bundles, the role of the different actin-bundling proteins remains largely unknown. In this study, we investigated the role of the actin-bundling protein villin in Arabidopsis (Arabidopsis thaliana). We used Arabidopsis T-DNA insertion lines to generate a double mutant in which VILLIN2 (VLN2) and VLN3 transcripts are truncated. Leaves, stems, siliques, and roots of vln2 vln3 double mutant plants are twisted, which is caused by local differences in cell length. Microscopy analysis of the actin cytoskeleton showed that in these double mutant plants, thin actin filament bundles are more abundant while thick actin filament bundles are virtually absent. In contrast to full-length VLN3, truncated VLN3 lacking the headpiece region does not rescue the phenotype of the vln2 vln3 double mutant. Our results show that villin is involved in the generation of thick actin filament bundles in several cell types and suggest that these bundles are involved in the regulation of coordinated cell expansion. PMID:22209875

  6. Arabidopsis VILLIN2 and VILLIN3 Are Required for the Generation of Thick Actin Filament Bundles and for Directional Organ Growth[C][W

    PubMed Central

    van der Honing, Hannie S.; Kieft, Henk; Emons, Anne Mie C.; Ketelaar, Tijs

    2012-01-01

    In plant cells, actin filament bundles serve as tracks for myosin-dependent organelle movement and play a role in the organization of the cytoplasm. Although virtually all plant cells contain actin filament bundles, the role of the different actin-bundling proteins remains largely unknown. In this study, we investigated the role of the actin-bundling protein villin in Arabidopsis (Arabidopsis thaliana). We used Arabidopsis T-DNA insertion lines to generate a double mutant in which VILLIN2 (VLN2) and VLN3 transcripts are truncated. Leaves, stems, siliques, and roots of vln2 vln3 double mutant plants are twisted, which is caused by local differences in cell length. Microscopy analysis of the actin cytoskeleton showed that in these double mutant plants, thin actin filament bundles are more abundant while thick actin filament bundles are virtually absent. In contrast to full-length VLN3, truncated VLN3 lacking the headpiece region does not rescue the phenotype of the vln2 vln3 double mutant. Our results show that villin is involved in the generation of thick actin filament bundles in several cell types and suggest that these bundles are involved in the regulation of coordinated cell expansion. PMID:22209875

  7. Induction by transforming growth factor-β1 of epithelial to mesenchymal transition is a rare event in vitro

    PubMed Central

    Brown, Kimberly A; Aakre, Mary E; Gorska, Agnieska E; Price, James O; Eltom, Sakina E; Pietenpol, Jennifer A; Moses, Harold L

    2004-01-01

    Introduction Transforming growth factor (TGF)-β1 is proposed to inhibit the growth of epithelial cells in early tumorigenesis, and to promote tumor cell motility and invasion in the later stages of carcinogenesis through the induction of an epithelial to mesenchymal transition (EMT). EMT is a multistep process that is characterized by changes in cell morphology and dissociation of cell–cell contacts. Although there is growing interest in TGF-β1-mediated EMT, the phenotype is limited to only a few murine cell lines and mouse models. Methods To identify alternative cell systems in which to study TGF-β1-induced EMT, 18 human and mouse established cell lines and cultures of two human primary epithelial cell types were screened for TGF-β1-induced EMT by analysis of cell morphology, and localization of zonula occludens-1, E-cadherin, and F-actin. Sensitivity to TGF-β1 was also determined by [3H]thymidine incorporation, flow cytometry, phosphorylation of Smad2, and total levels of Smad2 and Smad3 in these cell lines and in six additional cancer cell lines. Results TGF-β1 inhibited the growth of most nontransformed cells screened, but many of the cancer cell lines were insensitive to the growth inhibitory effects of TGF-β1. In contrast, TGF-β1 induced Smad2 phosphorylation in the majority of cell lines, including cell lines resistant to TGF-β1-mediated cell cycle arrest. Of the cell lines screened only two underwent TGF-β1-induced EMT. Conclusion The results presented herein show that, although many cancer cell lines have lost sensitivity to the growth inhibitory effect of TGF-β1, most show evidence of TGF-β1 signal transduction, but only a few cell lines undergo TGF-β1-mediated EMT. PMID:15084245

  8. Computational model of polarized actin cables and cytokinetic actin ring formation in budding yeast

    PubMed Central

    Tang, Haosu; Bidone, Tamara C.

    2015-01-01

    The budding yeast actin cables and contractile ring are important for polarized growth and division, revealing basic aspects of cytoskeletal function. To study these formin-nucleated structures, we built a 3D computational model with actin filaments represented as beads connected by springs. Polymerization by formins at the bud tip and bud neck, crosslinking, severing, and myosin pulling, are included. Parameter values were estimated from prior experiments. The model generates actin cable structures and dynamics similar to those of wild type and formin deletion mutant cells. Simulations with increased polymerization rate result in long, wavy cables. Simulated pulling by type V myosin stretches actin cables. Increasing the affinity of actin filaments for the bud neck together with reduced myosin V pulling promotes the formation of a bundle of antiparallel filaments at the bud neck, which we suggest as a model for the assembly of actin filaments to the contractile ring. PMID:26538307

  9. Transforming growth factor-betas and vascular disorders.

    PubMed

    Bobik, Alex

    2006-08-01

    Transforming growth factor-beta (TGF-beta) superfamily members, TGF-beta and bone morphogenetic proteins (BMPs), are potent regulatory cytokines with diverse functions on vascular cells. They signal through heteromeric type I and II receptor complexes activating Smad-dependent and Smad-independent signals, which regulate proliferation, differentiation, and survival. They are potent regulators of vascular development and vessel remodeling and play key roles in atherosclerosis and restenosis, regulating endothelial, smooth muscle cell, macrophage, T cell, and probably vascular calcifying cell responses. In atherosclerosis, TGF-beta regulates lesion phenotype by controlling T-cell responses and stimulating smooth muscle cells to produce collagen. It contributes to restenosis by augmenting neointimal cell proliferation and collagen accumulation. Defective TGF-beta signaling in endothelial cells attributable to mutations in endoglin or the type I receptor ALK-1 leads to hereditary hemorrhagic telangiectasia, whereas defective BMP signaling attributable to mutations in the BMP receptor II has been associated with development of primary pulmonary hypertension. The development of mouse models with either cell type-specific or general inactivation of TGF-beta/BMP signaling has started to reveal the importance of the regulatory network of TGF-beta/BMP pathways in vivo and their significance for atherosclerosis, hereditary hemorrhagic telangiectasia, and primary pulmonary hypertension. This review highlights recent findings that have advanced our understanding of the roles of TGF-beta superfamily members in regulating vascular cell responses and provides likely avenues for future research that may lead to novel pharmacological therapies for the treatment or prevention of vascular disorders. PMID:16675726

  10. Modulation of growth and differentiation in normal human keratinocytes by transforming growth factor-beta

    SciTech Connect

    Matsumoto, K.; Hashimoto, K.; Hashiro, M.; Yoshimasa, H.; Yoshikawa, K. )

    1990-10-01

    The effect of transforming growth factor-type beta 1(TGF-beta) on the growth and differentiation of normal human skin keratinocytes cultured in serum-free medium was investigated. TGF-beta markedly inhibited the growth of keratinocytes at the concentrations greater than 2 ng/ml under low Ca2+ conditions (0.1 mM). Growth inhibition was accompanied by changes in cell functions related to proliferation. Remarkable inhibition of DNA synthesis was demonstrated by the decrease of (3H)thymidine incorporation. The decrease of (3H)thymidine incorporation was observed as early as 3 hr after addition of TGF-beta. TGF-beta also decreased c-myc messenger RNA (mRNA) expression 30 min after addition of TGF-beta. This rapid reduction of c-myc mRNA expression by TGF-beta treatment is possibly one of the main factors in the process of TGF-beta-induced growth inhibition of human keratinocytes. Since growth inhibition and induction of differentiation are closely related in human keratinocytes, the growth-inhibitory effect of TGF-beta under high Ca2+ conditions was examined. TGF-beta inhibited the growth of keratinocytes under high Ca2+ conditions in the same manner as under low Ca2+ conditions, suggesting that it is a strong growth inhibitor in both low and high Ca2+ environments. The induction of keratinocyte differentiation was evaluated by measuring involucrin expression and cornified envelope formation: TGF-beta at 20 ng/ml increased involucrin expression from 9.3% to 18.8% under high Ca2+ conditions, while it decreased involucrin expression from 7.0% to 3.3% under low Ca2+ conditions. Cornified envelope formation was modulated in a similar way by addition of TGF-beta: TGF-beta at 20 ng/ml decreased cornified envelope formation by 53% under low Ca2+ conditions, while it enhanced cornified envelope formation by 30.7% under high Ca2+ conditions.

  11. Profilin connects actin assembly with microtubule dynamics.

    PubMed

    Nejedla, Michaela; Sadi, Sara; Sulimenko, Vadym; de Almeida, Francisca Nunes; Blom, Hans; Draber, Pavel; Aspenström, Pontus; Karlsson, Roger

    2016-08-01

    Profilin controls actin nucleation and assembly processes in eukaryotic cells. Actin nucleation and elongation promoting factors (NEPFs) such as Ena/VASP, formins, and WASP-family proteins recruit profilin:actin for filament formation. Some of these are found to be microtubule associated, making actin polymerization from microtubule-associated platforms possible. Microtubules are implicated in focal adhesion turnover, cell polarity establishment, and migration, illustrating the coupling between actin and microtubule systems. Here we demonstrate that profilin is functionally linked to microtubules with formins and point to formins as major mediators of this association. To reach this conclusion, we combined different fluorescence microscopy techniques, including superresolution microscopy, with siRNA modulation of profilin expression and drug treatments to interfere with actin dynamics. Our studies show that profilin dynamically associates with microtubules and this fraction of profilin contributes to balance actin assembly during homeostatic cell growth and affects micro-tubule dynamics. Hence profilin functions as a regulator of microtubule (+)-end turnover in addition to being an actin control element. PMID:27307590

  12. Actin cytoskeleton demonstration in Trichomonas vaginalis and in other trichomonads.

    PubMed

    Brugerolle, G; Bricheux, G; Coffe, G

    1996-01-01

    The flagellate form of Trichomonas vaginalis (T v) transforms to amoeboid cells upon adherence to converslips. They grow and their nuclei divide without undergoing cytokinesis, yielding giant cells and a monolayer of T v F-actin was demonstrated in Trichomonas vaginalis by fluorescence microscopy using phalloidin and an anti-actin mAb which labelled the cytoplasm of both the flagellate and amoeboid forms. Comparative electrophoresis and immunoblotting established that the actin band has the same 42 kDa as muscle actin, but 2-D electrophoresis resolved the actin band into four spots; the two major spots observed were superimposable with major muscle actin isoforms. Electron microscopy demonstrated an ectoplasmic microfibrillar layer along the adhesion zone of amoeboid T v adhering to coverslips. Immunogold staining, using anti-actin monoclonal antibodies demonstrated that this layer was mainly composed of actin microfilaments. A comparative immunoblotting study comprising seven trichomonad species showed that all trichomonads studied expressed actin. The mAb Sigma A-4700 specific for an epitope on the actin C-terminal sequence labelled only actin of Trichomonas vaginalis, Tetratrichomonas gallinarum. Trichomitus batrachorum and Hypotrichomonas acosta, but not the actin of Tritrichomonas foetus, Tritrichomonas augusta and Monocercomonas sp. This discrimination between a 'trichomonas branch' and a 'tritrichomonas branch' is congruent with inferred sequence phylogeny from SSu rRNA and with classical phylogeny of trichomonads. PMID:9175265

  13. Zoledronic acid suppresses transforming growth factor-β-induced fibrogenesis by human gingival fibroblasts.

    PubMed

    Komatsu, Yuko; Ibi, Miho; Chosa, Naoyuki; Kyakumoto, Seiko; Kamo, Masaharu; Shibata, Toshiyuki; Sugiyama, Yoshiki; Ishisaki, Akira

    2016-07-01

    Bisphosphonates (BPs) are analogues of pyrophosphate that are known to prevent bone resorption by inhibiting osteoclast activity. Nitrogen-containing BPs, such as zoledronic acid (ZA), are widely used in the treatment of osteoporosis and bone metastasis. However, despite having benefits, ZA has been reported to induce BP-related osteonecrosis of the jaw (BRONJ) in cancer patients. The molecular pathological mechanisms responsible for the development of BRONJ, including necrotic bone exposure after tooth extraction, remain to be elucidated. In this study, we examined the effects of ZA on the transforming growth factor-β (TGF‑β)-induced myofibroblast (MF) differentiation of human gingival fibroblasts (hGFs) and the migratory activity of hGFs, which are important for wound closure by fibrous tissue formation. The ZA maximum concentration in serum (Cmax) was found to be approximately 1.47 µM, which clinically, is found after the intravenous administration of 4 mg ZA, and ZA at this dose is considered appropriate for the treatment of cancer bone metastasis or bone diseases, such as Erdheim-Chester disease. At Cmax, ZA significantly suppressed i) the TGF‑β-induced promotion of cell viability, ii) the TGF‑β-induced expression of MF markers such as α-smooth muscle actin (α-SMA) and type I collagen, iii) the TGF‑β-induced migratory activity of hGFs and iv) the expression level of TGF‑β type I receptor on the surfaces of hGFs, as well as the TGF‑β-induced phosphorylation of Smad2/3. Thus, ZA suppresses TGF‑β-induced fibrous tissue formation by hGFs, possibly through the inhibition of Smad‑dependent signal transduction. Our findings partly elucidate the molecular mechanisms underlying BRONJ and may prove to be beneficial to the identification of drug targets for the treatment of this symptom at the molecular level. PMID:27176567

  14. Epidemiology of actinic keratoses.

    PubMed

    Green, Adèle C

    2015-01-01

    The epidemiology of actinic keratoses (AKs) reflects their causation by cumulative sun exposure, with the highest prevalence seen in pale-skinned people living at low latitudes and on the most sun-exposed body sites, namely the hands, forearms and face. AKs are markers of increased risk of basal cell carcinoma, squamous cell carcinoma and melanoma, especially when they are numerous and have coalesced into an area of 'field cancerisation'. The major risk factors are male sex, advanced age, sun-sensitive complexion, high lifetime sun exposure and prolonged immunosuppression. Clinical counts of AKs enable the assessment and monitoring of AK burden, but accurate counting is notoriously difficult, especially when skin is severely sun damaged. AK counting has been repeatedly shown to be unreliable, even among expert dermatologists. Notwithstanding these challenges, qualitative assessment of the natural history of AKs shows a high turnover, with new lesions developing and with other lesions regressing. A very small proportion of AKs undergo malignant transformation, but the precise rate of transformation is unknown due to the inaccuracies in monitoring AK lesions over time. Primary prevention of AKs is achieved by limiting intense sun exposure through sun-protective behaviour, including seeking deep shade, wearing sun-protective clothing and applying sunscreen regularly to exposed skin, from an early age. PMID:25561199

  15. Transforming growth factor alpha and epidermal growth factor levels in bladder cancer and their relationship to epidermal growth factor receptor.

    PubMed Central

    Mellon, J. K.; Cook, S.; Chambers, P.; Neal, D. E.

    1996-01-01

    We have examined levels of epidermal growth factor (EGF) and transforming growth factor alpha (TGF-alpha) in neoplastic and non-neoplastic bladder tissue using a standard radioimmunoassay technique. Tumour samples had much higher TGF-alpha levels compared with EGF and TGF-alpha levels in malignant tissue were significantly higher than in benign bladder samples. There was, in addition, a difference in mean EGF levels from 'normal' bladder samples from non-tumour bearing areas of bladder in patients with bladder cancer compared with 'normal' bladder tissue obtained at the time of organ retrieval surgery. Levels of EGF and TGF-alpha did not correlate with levels of EGF receptor (EGFR) as determined by a radioligand binding method but levels of TGF-alpha > 10 ng gm-1 of tumour tissue did correlate with EGFR positivity defined using immunohistochemistry. These data suggest that TGF-alpha is the likely ligand for EGFR in bladder tumours. PMID:8605103

  16. Transforming your professional self: encouraging lifelong personal and professional growth.

    PubMed

    Rodts, Mary F; Lamb, Karen V

    2008-01-01

    Transforming from student nurse to registered nurse is often discussed in a capstone class or a hospital orientation program. Changes in professional plans later in the career continuum often occur, but are not always planned. This article discusses the challenges of change, the need for career planning, stages of role acquisition, role socialization, and role transformation. In addition, it outlines the importance of creating a career plan to meet future career goals. PMID:18385597

  17. A dynamic formin-dependent deep F-actin network in axons

    PubMed Central

    Ganguly, Archan; Tang, Yong; Wang, Lina; Ladt, Kelsey; Loi, Jonathan; Dargent, Bénédicte; Leterrier, Christophe

    2015-01-01

    Although actin at neuronal growth cones is well-studied, much less is known about actin organization and dynamics along axon shafts and presynaptic boutons. Using probes that selectively label filamentous-actin (F-actin), we found focal “actin hotspots” along axons—spaced ∼3–4 µm apart—where actin undergoes continuous assembly/disassembly. These foci are a nidus for vigorous actin polymerization, generating long filaments spurting bidirectionally along axons—a phenomenon we call “actin trails.” Super-resolution microscopy reveals intra-axonal deep actin filaments in addition to the subplasmalemmal “actin rings” described recently. F-actin hotspots colocalize with stationary axonal endosomes, and blocking vesicle transport diminishes the actin trails, suggesting mechanistic links between vesicles and F-actin kinetics. Actin trails are formin—but not Arp2/3—dependent and help enrich actin at presynaptic boutons. Finally, formin inhibition dramatically disrupts synaptic recycling. Collectively, available data suggest a two-tier F-actin organization in axons, with stable “actin rings” providing mechanical support to the plasma membrane and dynamic "actin trails" generating a flexible cytoskeletal network with putative physiological roles. PMID:26216902

  18. TERATOGENIC RESPONSES ARE MODULATED IN MICE LACKING EXPRESSION OF EPIDERMAL GROWTH FACTOR (EGF) AND TRANSFORMING GROWTH FACTOR-ALPHA (TGF)

    EPA Science Inventory

    TITLE:
    TERATOGENIC RESPONSES ARE MODULATED IN MICE LACKING EXPRESSION OF EPIDERMAL GROWTH FACTOR (EGF) AND TRANSFORMING GROWTH FACTOR-ALPHA (TGF). AUTHORS (ALL): Abbott, Barbara D.1; Best, Deborah S.1; Narotsky, Michael G.1. SPONSOR NAME: None INSTITUTIONS (ALL): 1. Repro Tox ...

  19. Actin Turnover-Mediated Gravity Response in Maize Root Apices

    PubMed Central

    Mancuso, Stefano; Barlow, Peter W; Volkmann, Dieter

    2006-01-01

    The dynamic actin cytoskeleton has been proposed to be linked to gravity sensing in plants but the mechanistic understanding of these processes remains unknown. We have performed detailed pharmacological analyses of the role of the dynamic actin cytoskeleton in gravibending of maize (Zea mays) root apices. Depolymerization of actin filaments with two drugs having different mode of their actions, cytochalasin D and latrunculin B, stimulated root gravibending. By contrast, drug-induced stimulation of actin polymerization and inhibition of actin turnover, using two different agents phalloidin and jasplakinolide, compromised the root gravibending. Importantly, all these actin drugs inhibited root growth to similar extents suggesting that high actin turnover is essential for the gravity-related growth responses rather than for the general growth process. Both latrunculin B and cytochalasin D treatments inhibited root growth but restored gravibending of the decapped root apices, indicating that there is a strong potential for effective actin-mediated gravity sensing outside the cap. This elusive gravity sensing outside the root cap is dependent not only on the high rate of actin turnover but also on weakening of myosin activities, as general inhibition of myosin ATPases induced stimulation of gravibending of the decapped root apices. Collectively, these data provide evidence for the actin turnover-mediated gravity sensing outside the root cap. PMID:19521476

  20. Transforming growth factor. beta. sub 1 is present at sites of extracellular matrix gene expression in human pulmonary fibrosis

    SciTech Connect

    Broekelmann, T.J.; Limper, A.H.; McDonald, J.A. ); Colby, T.V. )

    1991-08-01

    Idiopathic pulmonary fibrosis is an inexorably fatal disorder characterized by connective tissue deposition within the terminal air spaces resulting in loss of lung function and eventual respiratory failure. Previously, the authors demonstrated that foci of activated fibroblasts expressing high levels of fibronectin, procollagen, and smooth muscle actin and thus resembling those found in healing wounds are responsible for the connective tissue deposition and scarring in idiopathic pulmonary fibrosis. Using in situ hybridization and immunohistochemistry, they now demonstrate the presence of transforming growth factor {beta}{sub 1} (TGF-{beta}{sub 1}), a potent profibrotic cytokine, in the foci containing these activated fibroblasts. These results suggest that matrix-associated TGF-{beta}{sub 1} may serve as a stimulus for the persistent expression of connective tissue genes. One potential source of the TGF-{beta}{sub 1} is the alveolar macrophage, and they demonstrate the expression of abundant TGF-{beta}{sub 1} mRNA in alveolar macrophages in lung tissue from patients with idiopathic pulmonary fibrosis.

  1. Transforming Growth Factor β-1 Stimulates Profibrotic Epithelial Signaling to Activate Pericyte-Myofibroblast Transition in Obstructive Kidney Fibrosis

    PubMed Central

    Wu, Ching-Fang; Chiang, Wen-Chih; Lai, Chun-Fu; Chang, Fan-Chi; Chen, Yi-Ting; Chou, Yu-Hsiang; Wu, Ting-Hui; Linn, Geoffrey R.; Ling, Hong; Wu, Kwan-Dun; Tsai, Tun-Jun; Chen, Yung-Ming; Duffield, Jeremy S.; Lin, Shuei-Liong

    2014-01-01

    Pericytes have been identified as the major source of precursors of scar-producing myofibroblasts during kidney fibrosis. The underlying mechanisms triggering pericyte-myofibroblast transition are poorly understood. Transforming growth factor β-1 (TGF-β1) is well recognized as a pluripotent cytokine that drives organ fibrosis. We investigated the role of TGF-β1 in inducing profibrotic signaling from epithelial cells to activate pericyte-myofibroblast transition. Increased expression of TGF-β1 was detected predominantly in injured epithelium after unilateral ureteral obstruction, whereas downstream signaling from the TGF-β1 receptor increased in both injured epithelium and pericytes. In mice with ureteral obstruction that were treated with the pan anti–TGF-β antibody (1D11) or TGF-β receptor type I inhibitor (SB431542), kidney pericyte-myofibroblast transition was blunted. The consequence was marked attenuation of fibrosis. In addition, epithelial cell cycle G2/M arrest and production of profibrotic cytokines were both attenuated. Although TGF-β1 alone did not trigger pericyte proliferation in vitro, it robustly induced α smooth muscle actin (α-SMA). In cultured kidney epithelial cells, TGF-β1 stimulated G2/M arrest and production of profibrotic cytokines that had the capacity to stimulate proliferation and transition of pericytes to myofibroblasts. In conclusion, this study identified a novel link between injured epithelium and pericyte-myofibroblast transition through TGF-β1 during kidney fibrosis. PMID:23142380

  2. Dynamic Localization of G-actin During Membrane Protrusion in Neuronal Motility

    PubMed Central

    Lee, Chi Wai; Vitriol, Eric A.; Shim, Sangwoo; Wise, Ariel L.; Velayutham, Radhi P.; Zheng, James Q.

    2013-01-01

    Summary Background Actin-based cell motility is fundamental for the development, function, and malignant events of eukaryotic organisms. During neural development, axonal growth cones depend on rapid assembly and disassembly of actin filaments (F-actin) for their guided extension to specific targets for wiring. Monomeric globular actin (G-actin) is the building block for F-actin but is not considered to play a direct role in spatiotemporal control of actin dynamics in cell motility. Results Here we report that a pool of G-actin dynamically localizes to the leading edge of growth cones and neuroblastoma cells to spatially elevate the G-/F-actin ratio that drives membrane protrusion and cell movement. Loss of G-actin localization leads to the cessation and retraction of membrane protrusions. Moreover, G-actin localization occurs asymmetrically in growth cones during attractive turning. Finally, we identify the actin monomer binding proteins profilin and thymosin β4 as key molecules that localize actin monomers to the leading edge of lamellipodia for their motility. Conclusions Our results suggest that dynamic localization of G-actin provides a novel mechanism to regulate the spatiotemporal actin dynamics underlying membrane protrusion in cell locomotion and growth cone chemotaxis. PMID:23746641

  3. Exploring the Stability Limits of Actin and Its Suprastructures

    PubMed Central

    Rosin, Christopher; Erlkamp, Mirko; Ecken, Julian von der; Raunser, Stefan; Winter, Roland

    2014-01-01

    Actin is the main component of the microfilament system in eukaryotic cells and can be found in distinct morphological states. Global (G)-actin is able to assemble into highly organized, supramolecular cellular structures known as filamentous (F)-actin and bundled (B)-actin. To evaluate the structure and stability of G-, F-, and B-actin over a wide range of temperatures and pressures, we used Fourier transform infrared spectroscopy in combination with differential scanning and pressure perturbation calorimetry, small-angle x-ray scattering, laser confocal scanning microscopy, and transmission electron microscopy. Our analysis was designed to provide new (to our knowledge) insights into the stabilizing forces of actin self-assembly and to reveal the stability of the actin polymorphs, including in conditions encountered in extreme environments. In addition, we sought to explain the limited pressure stability of actin self-assembly observed in vivo. G-actin is not only the least temperature-stable but also the least pressure-stable actin species. Under abyssal conditions, where temperatures as low as 1–4°C and pressures up to 1 kbar are reached, G-actin is hardly stable. However, the supramolecular assemblies of actin are stable enough to withstand the extreme conditions usually encountered on Earth. Beyond ∼3–4 kbar, filamentous structures disassemble, and beyond ∼4 kbar, complete dissociation of F-actin structures is observed. Between ∼1 and 2 kbar, some disordering of actin assemblies commences, in agreement with in vivo observations. The limited pressure stability of the monomeric building block seems to be responsible for the suppression of actin assembly in the kbar pressure range. PMID:25517163

  4. Actin in Herpesvirus Infection

    PubMed Central

    Roberts, Kari L.; Baines, Joel D.

    2011-01-01

    Actin is important for a variety of cellular processes, including uptake of extracellular material and intracellular transport. Several emerging lines of evidence indicate that herpesviruses exploit actin and actin-associated myosin motors for viral entry, intranuclear transport of capsids, and virion egress. The goal of this review is to explore these processes and to highlight potential future directions for this area of research. PMID:21994736

  5. Increased fibroblast telomerase expression precedes myofibroblast α-smooth muscle actin expression in idiopathic pulmonary fibrosis

    PubMed Central

    Waisberg, Daniel Reis; Parra, Edwin Roger; Barbas-Filho, João Valente; Fernezlian, Sandra; Capelozzi, Vera Luiza

    2012-01-01

    OBJECTIVE: This study sought to identify the relationship between fibroblast telomerase expression, myofibroblasts, and telomerase-mediated regulatory signals in idiopathic pulmonary fibrosis. METHODS: Thirty-four surgical lung biopsies, which had been obtained from patients with idiopathic pulmonary fibrosis and histologically classified as usual interstitial pneumonia, were examined. Immunohistochemistry was used to evaluate fibroblast telomerase expression, myofibroblast α-smooth muscle actin expression and the tissue expression of interleukin-4, transforming growth factor-β, and basic fibroblast growth factor. The point-counting technique was used to quantify the expression of these markers in unaffected, collapsed, mural fibrosis, and honeycombing areas. The results were correlated to patient survival. RESULTS: Fibroblast telomerase expression and basic fibroblast growth factor tissue expression were higher in collapsed areas, whereas myofibroblast expression and interleukine-4 tissue expression were higher in areas of mural fibrosis. Transforming growth factor-β expression was higher in collapsed, mural fibrosis and honeycombing areas in comparison to unaffected areas. Positive correlations were found between basic fibroblast growth factor tissue expression and fibroblast telomerase expression and between interleukin-4 tissue expression and myofibroblast α-smooth muscle actin expression. Negative correlations were observed between interleukin-4 expression and basic fibroblast growth factor tissue expression in areas of mural fibrosis. Myofibroblast α-smooth muscle actin expression and interleukin-4 tissue expression in areas of mural fibrosis were negatively associated with patient survival. CONCLUSION: Fibroblast telomerase expression is higher in areas of early remodeling in lung tissues demonstrating typical interstitial pneumonia, whereas myofibroblast α-smooth muscle actin expression predominates in areas of late remodeling. These events seem to be

  6. Actin Rings of Power.

    PubMed

    Schwayer, Cornelia; Sikora, Mateusz; Slováková, Jana; Kardos, Roland; Heisenberg, Carl-Philipp

    2016-06-20

    Circular or ring-like actin structures play important roles in various developmental and physiological processes. Commonly, these rings are composed of actin filaments and myosin motors (actomyosin) that, upon activation, trigger ring constriction. Actomyosin ring constriction, in turn, has been implicated in key cellular processes ranging from cytokinesis to wound closure. Non-constricting actin ring-like structures also form at cell-cell contacts, where they exert a stabilizing function. Here, we review recent studies on the formation and function of actin ring-like structures in various morphogenetic processes, shedding light on how those different rings have been adapted to fulfill their specific roles. PMID:27326928

  7. Overexpression of granulocyte-macrophage colony-stimulating factor induces pulmonary granulation tissue formation and fibrosis by induction of transforming growth factor-beta 1 and myofibroblast accumulation.

    PubMed Central

    Xing, Z.; Tremblay, G. M.; Sime, P. J.; Gauldie, J.

    1997-01-01

    We have previously reported that transfer to rat lung of the granulocyte-macrophage colony-stimulating factor (GM-CSF) gene leads to high expression of GM-CSF between days 1 and 4 and granulation tissue formation followed by an irreversible fibrotic response starting from day 12 onward. In the current study, we investigated the underlying mechanisms. We found that GM-CSF overexpression did not enhance production of tumor necrosis factor-alpha in a significant manner at any time after GM-CSF gene transfer. However, the content of transforming growth factor-beta 1 in bronchoalveolar lavage fluid was markedly induced at day 4 and appeared to be maximal around day 7 and remained high at day 12. Macrophages purified from bronchoalveolar lavage fluid 7 days after GM-CSF gene transfer spontaneously released significant quantities of transforming growth factor-beta 1 protein in vitro. After peak transforming growth factor-beta 1 production was the emergence of alpha-smooth muscle actin-rich myofibroblasts. Accumulation of these cells was most prominent at day 12 within the granulation tissues and they were still present in fibrotic areas between days 12 and 24 and diminished markedly afterward. Thus, we provide the first in vivo evidence that tumor necrosis factor-alpha may be dissociated from participation in a fibrotic process in the lung and GM-CSF may play a more direct role in pulmonary fibrogenesis at least in part through its capability to induce transforming growth factor-beta 1 in macrophages and the subsequent emergence of myofibroblast phenotypes. This GM-CSF transgene lung model is useful for a stepwise dissection of both cellular and molecular events involved in pulmonary fibrosis. Images Figure 2 Figure 5 Figure 6 PMID:9006322

  8. Transforming Asian Pacific America: The Challenges of Growth and Diversity.

    ERIC Educational Resources Information Center

    Nakanishi, Don T.

    1994-01-01

    Examines the concept of Asian Pacific pan-ethnicity in light of the growing numbers and diversity of this population in the 1990s. While revised immigration laws, new immigrant groups, and continued hostility have brought new challenges, it is argued that Asian Pacific Americans can provide important leadership for the transformation of the United…

  9. Arabidopsis CROLIN1, a Novel Plant Actin-binding Protein, Functions in Cross-linking and Stabilizing Actin Filaments*

    PubMed Central

    Jia, Honglei; Li, Jisheng; Zhu, Jingen; Fan, Tingting; Qian, Dong; Zhou, Yuelong; Wang, Jiaojiao; Ren, Haiyun; Xiang, Yun; An, Lizhe

    2013-01-01

    Higher order actin filament structures are necessary for cytoplasmic streaming, organelle movement, and other physiological processes. However, the mechanism by which the higher order cytoskeleton is formed in plants remains unknown. In this study, we identified a novel actin-cross-linking protein family (named CROLIN) that is well conserved only in the plant kingdom. There are six isovariants of CROLIN in the Arabidopsis genome, with CROLIN1 specifically expressed in pollen. In vitro biochemical analyses showed that CROLIN1 is a novel actin-cross-linking protein with binding and stabilizing activities. Remarkably, CROLIN1 can cross-link actin bundles into actin networks. CROLIN1 loss of function induces pollen germination and pollen tube growth hypersensitive to latrunculin B. All of these results demonstrate that CROLIN1 may play an important role in stabilizing and remodeling actin filaments by binding to and cross-linking actin filaments. PMID:24072702

  10. [Actin in the wound healing process].

    PubMed

    Nowak, Dorota; Popow-Woźniak, Agnieszka; Raźnikiewicz, Linda; Malicka-Błaszkiewicz, Maria

    2009-01-01

    Wound healing is an important biological process of crucial value for organisms survival and retention of its proper functions. The recognition of molecular mechanisms of these phenomenon is still under investigation. The transition of mesenchymal fibroblasts to myofibroblasts is a key point in wound healing. The contraction ability of myofibroblast enables the shrinkage of a wound and closes its edges. Alpha smooth muscle actin (alpha-SMA), one of six actin isoforms, is a marker of compeletely differentiated myofibroblast. The regulation of differentiation process depends on many growth factors (especially TGF beta 1), the level of active thymosin beta 4, extracellular matrix proteins--including fibronectin, and also on specificity of microenvironment. Thymosin beta 4 is responsible for maintenance of pool of monomeric actin and actin filaments depolymerization. It can also act as a transcription factor, migration stimulator and immunomodulator, so this protein deserves for more attention in wound healing research field. PMID:19824469

  11. Stress-induced martensitic transformation in metastable austenitic stainless steels: Effect on fatigue crack growth rate

    NASA Astrophysics Data System (ADS)

    Khan, Z.; Ahmed, M.

    1996-04-01

    This paper addresses the influence of cyclic stress-induced martensitic transformation on fatigue crack growth rates in metastable austenitic stainless steels. At low applied stress and mean stress values in AISI type 301 stainless steel, fatigue crack growth rate is substantially retarded due to a cyclic stress-induced γ-α' and γ-ɛ martensitic transformation occurring at the crack-tip plastic zone. It is suggested that the transformation products produce a compressive residual stress at the tip of the fatigue crack, which essentially lowers the effective stress intensity and hence retards the fatigue crack growth rate. At high applied stress or mean stress values, fatigue crack growth rates in AISI type 301 steels become almost equal to those of stable AISI type 302 alloy. As the amount of transformed products increases (with an increase in applied or mean stress), the strain-hardening effect brought about by the transformed martensite phase appears to accelerate fatigue crack growth, offsetting the contribution from the compressive residual stress produced by the positive volume change of γ → α' or ɛ transformation.

  12. Direct dynamin–actin interactions regulate the actin cytoskeleton

    PubMed Central

    Gu, Changkyu; Yaddanapudi, Suma; Weins, Astrid; Osborn, Teresia; Reiser, Jochen; Pollak, Martin; Hartwig, John; Sever, Sanja

    2010-01-01

    The large GTPase dynamin assembles into higher order structures that are thought to promote endocytosis. Dynamin also regulates the actin cytoskeleton through an unknown, GTPase-dependent mechanism. Here, we identify a highly conserved site in dynamin that binds directly to actin filaments and aligns them into bundles. Point mutations in the actin-binding domain cause aberrant membrane ruffling and defective actin stress fibre formation in cells. Short actin filaments promote dynamin assembly into higher order structures, which in turn efficiently release the actin-capping protein (CP) gelsolin from barbed actin ends in vitro, allowing for elongation of actin filaments. Together, our results support a model in which assembled dynamin, generated through interactions with short actin filaments, promotes actin polymerization via displacement of actin-CPs. PMID:20935625

  13. Actin Mechanics and Fragmentation*

    PubMed Central

    De La Cruz, Enrique M.; Gardel, Margaret L.

    2015-01-01

    Cell physiological processes require the regulation and coordination of both mechanical and dynamical properties of the actin cytoskeleton. Here we review recent advances in understanding the mechanical properties and stability of actin filaments and how these properties are manifested at larger (network) length scales. We discuss how forces can influence local biochemical interactions, resulting in the formation of mechanically sensitive dynamic steady states. Understanding the regulation of such force-activated chemistries and dynamic steady states reflects an important challenge for future work that will provide valuable insights as to how the actin cytoskeleton engenders mechanoresponsiveness of living cells. PMID:25957404

  14. Substrate flexibility regulates growth and apoptosis of normal but not transformed cells

    NASA Technical Reports Server (NTRS)

    Wang, H. B.; Dembo, M.; Wang, Y. L.

    2000-01-01

    One of the hallmarks of oncogenic transformation is anchorage-independent growth (27). Here we demonstrate that responses to substrate rigidity play a major role in distinguishing the growth behavior of normal cells from that of transformed cells. We cultured normal or H-ras-transformed NIH 3T3 cells on flexible collagen-coated polyacrylamide substrates with similar chemical properties but different rigidity. Compared with cells cultured on stiff substrates, nontransformed cells on flexible substrates showed a decrease in the rate of DNA synthesis and an increase in the rate of apoptosis. These responses on flexible substrates are coupled to decreases in cell spreading area and traction forces. In contrast, transformed cells maintained their growth and apoptotic characteristics regardless of substrate flexibility. The responses in cell spreading area and traction forces to substrate flexibility were similarly diminished. Our results suggest that normal cells are capable of probing substrate rigidity and that proper mechanical feedback is required for regulating cell shape, cell growth, and survival. The loss of this response can explain the unregulated growth of transformed cells.

  15. Actin Polymerization is Stimulated by Actin Crosslinking Protein Palladin

    PubMed Central

    Gurung, Ritu; Yadav, Rahul; Brungardt, Joseph G.; Orlova, Albina; Egelman, Edward H.; Beck, Moriah R.

    2016-01-01

    The actin scaffold protein palladin regulates both normal cell migration and invasive cell motility, processes that require the coordinated regulation of actin dynamics. However, the potential effect of palladin on actin dynamics has remained elusive. Here we show that the actin binding immunoglobulin-like domain of palladin, which is directly responsible for both actin binding and bundling, also stimulates actin polymerization in vitro. Palladin eliminated the lag phase that is characteristic of the slow nucleation step of actin polymerization. Furthermore, palladin dramatically reduced depolymerization, slightly enhanced the elongation rate, and did not alter the critical concentration. Microscopy and in vitro crosslinking assays reveal differences in actin bundle architecture when palladin is incubated with actin before or after polymerization. These results suggest a model whereby palladin stimulates a polymerization-competent form of G-actin, akin to metal ions, either through charge neutralization or conformational changes. PMID:26607837

  16. Non-instantaneous growth characteristics of martensitic transformation in high Cr ferritic creep-resistant steel

    NASA Astrophysics Data System (ADS)

    Liu, Chenxi; Shao, Yi; Chen, Jianguo; Liu, Yongchang

    2016-08-01

    Microstructural observation and high-resolution dilatometry were employed to investigate kinetics of martensitic transformation in high Cr ferritic creep-resistant steel upon different quenching/cooling rates. By incorporating the classical athermal nucleation and impingement correction, a non-instantaneous growth model for martensitic transformation has been developed. The developed model describes austenite/martensite interface mobility during martensite growth. The growth rate of martensite is found to be varied from 1 × 10-6 to 3 × 10-6 m/s. The low interface mobility suggests that it is not appropriate to presume the instantaneous growth behavior of martensite. Moreover, based on the proposed model, nucleation rate of martensite under different cooling rates is found to be nearly the same, while the growth rate of martensite is promoted by increasing the cooling rate.

  17. The neuronal and actin commitment: Why do neurons need rings?

    PubMed

    Leite, Sérgio Carvalho; Sousa, Mónica Mendes

    2016-09-01

    The role of the actin cytoskeleton in neurons has been extensively studied in actin-enriched compartments such as the growth cone and dendritic spines. The recent discovery of actin rings in the axon shaft and in dendrites, together with the identification of axon actin trails, has advanced our understanding on actin organization and dynamics in neurons. However, specifically in the case of actin rings, the mechanisms regulating their nucleation and assembly, and the functions that they may exert in axons and dendrites remain largely unexplored. Here we discuss the possible structural, mechanistic and functional properties of the subcortical neuronal cytoskeleton putting the current knowledge in perspective with the information available on actin rings formed in other biological contexts, and with the organization of actin-spectrin lattices in other cell types. The detailed analysis of these novel neuronal actin ring structures, together with the elucidation of the function of actin-binding proteins in neuron biology, has a large potential to uncover new mechanisms of neuronal function under normal conditions that may have impact in our understanding of axon degeneration and regeneration. © 2016 Wiley Periodicals, Inc. PMID:26784007

  18. Environmental Particulate (PM2.5) Augments Stiffness-Induced Alveolar Epithelial Cell Mechanoactivation of Transforming Growth Factor Beta

    PubMed Central

    Dysart, Marilyn M.; Galvis, Boris R.; Russell, Armistead G.; Barker, Thomas H.

    2014-01-01

    Dysfunctional pulmonary homeostasis and repair, including diseases such as pulmonary fibrosis (PF), chronic obstructive pulmonary disease (COPD), and tumorigenesis have been increasing over the past decade, a fact that heavily implicates environmental influences. Several investigations have suggested that in response to increased transforming growth factor - beta (TGFβ) signaling, the alveolar type II (ATII) epithelial cell undergoes phenotypic changes that may contribute to the complex pathobiology of PF. We have previously demonstrated that increased tissue stiffness associated with PF is a potent extracellular matrix (ECM) signal for epithelial cell activation of TGFβ. The work reported here explores the relationship between tissue stiffness and exposure to environmental stimuli in the activation of TGFβ. We hypothesized that exposure of ATII cells to fine particulate matter (PM2.5) will result in enhanced cell contractility, TGFβ activation, and subsequent changes to ATII cell phenotype. ATII cells were cultured on increasingly stiff substrates with or without addition of PM2.5. Exposure to PM2.5 resulted in increased activation of TGFβ, increased cell contractility, and elongation of ATII cells. Most notably, on 8 kPa substrates, a stiffness greater than normal but less than established fibrotic lung, addition of PM2.5 resulted in increased cortical cell stiffness, enhanced actin staining and cell elongation; a result not seen in the absence of PM2.5. Our work suggests that PM2.5 exposure additionally enhances the existing interaction between ECM stiffness and TGFβ that has been previously reported. Furthermore, we show that this additional enhancement is likely a consequence of intracellular reactive oxygen species (ROS) leading to increased TGFβ signaling events. These results highlight the importance of both the micromechanical and biochemical environment in lung disease initiation and suggest that individuals in early stages of lung remodeling

  19. Normal Human Lung Epithelial Cells Inhibit Transforming Growth Factor-β Induced Myofibroblast Differentiation via Prostaglandin E2

    PubMed Central

    Epa, Amali P.; Thatcher, Thomas H.; Pollock, Stephen J.; Wahl, Lindsay A.; Lyda, Elizabeth; Kottmann, R. M.; Phipps, Richard P.; Sime, Patricia J.

    2015-01-01

    Introduction Idiopathic pulmonary fibrosis (IPF) is a chronic progressive disease with very few effective treatments. The key effector cells in fibrosis are believed to be fibroblasts, which differentiate to a contractile myofibroblast phenotype with enhanced capacity to proliferate and produce extracellular matrix. The role of the lung epithelium in fibrosis is unclear. While there is evidence that the epithelium is disrupted in IPF, it is not known whether this is a cause or a result of the fibroblast pathology. We hypothesized that healthy epithelial cells are required to maintain normal lung homeostasis and can inhibit the activation and differentiation of lung fibroblasts to the myofibroblast phenotype. To investigate this hypothesis, we employed a novel co-culture model with primary human lung epithelial cells and fibroblasts to investigate whether epithelial cells inhibit myofibroblast differentiation. Measurements and Main Results In the presence of transforming growth factor (TGF)-β, fibroblasts co-cultured with epithelial cells expressed significantly less α-smooth muscle actin and collagen and showed marked reduction in cell migration, collagen gel contraction, and cell proliferation compared to fibroblasts grown without epithelial cells. Epithelial cells from non-matching tissue origins were capable of inhibiting TGF-β induced myofibroblast differentiation in lung, keloid and Graves’ orbital fibroblasts. TGF-β promoted production of prostaglandin (PG) E2 in lung epithelial cells, and a PGE2 neutralizing antibody blocked the protective effect of epithelial cell co-culture. Conclusions We provide the first direct experimental evidence that lung epithelial cells inhibit TGF-β induced myofibroblast differentiation and pro-fibrotic phenotypes in fibroblasts. This effect is not restricted by tissue origin, and is mediated, at least in part, by PGE2. Our data support the hypothesis that the epithelium plays a crucial role in maintaining lung homeostasis

  20. Bereavement and transformation: a psycho-spiritual and post-traumatic growth perspective.

    PubMed

    Bray, Peter

    2013-09-01

    In the aftermath of loss, it is not uncommon for individuals to disclose psycho-spiritual experiences that lead to personal changes and psychological growth. This article argues for and outlines a broad conceptual framework that captures and acknowledges the significance of these subjective experiences. The model synthesises and integrates two approaches to healthy growth: Lawrence Calhoun and Richard Tedeschi's post-traumatic growth model and Stanislav and Christina Grof's model of psycho-spiritual transformation. A rationale for a broader model and underpinning theory is briefly discussed, and their shared understandings about the human potential for growth in the struggle through loss are explored. PMID:21948147

  1. Mutant Profilin Suppresses Mutant Actin-dependent Mitochondrial Phenotype in Saccharomyces cerevisiae*

    PubMed Central

    Wen, Kuo-Kuang; McKane, Melissa; Stokasimov, Ema; Rubenstein, Peter A.

    2011-01-01

    In the Saccharomyces cerevisiae actin-profilin interface, Ala167 of the actin barbed end W-loop and His372 near the C terminus form a clamp around a profilin segment containing residue Arg81 and Tyr79. Modeling suggests that altering steric packing in this interface regulates actin activity. An actin A167E mutation could increase interface crowding and alter actin regulation, and A167E does cause growth defects and mitochondrial dysfunction. We assessed whether a profilin Y79S mutation with its decreased mass could compensate for actin A167E crowding and rescue the mutant phenotype. Y79S profilin alone caused no growth defect in WT actin cells under standard conditions in rich medium and rescued the mitochondrial phenotype resulting from both the A167E and H372R actin mutations in vivo consistent with our model. Rescue did not result from effects of profilin on actin nucleotide exchange or direct effects of profilin on actin polymerization. Polymerization of A167E actin was less stimulated by formin Bni1 FH1-FH2 fragment than was WT actin. Addition of WT profilin to mixtures of A167E actin and formin fragment significantly altered polymerization kinetics from hyperbolic to a decidedly more sigmoidal behavior. Substitution of Y79S profilin in this system produced A167E behavior nearly identical to that of WT actin. A167E actin caused more dynamic actin cable behavior in vivo than observed with WT actin. Introduction of Y79S restored cable movement to a more normal phenotype. Our studies implicate the importance of the actin-profilin interface for formin-dependent actin and point to the involvement of formin and profilin in the maintenance of mitochondrial integrity and function. PMID:21956104

  2. Arabidopsis AtADF1 is functionally affected by mutations on actin binding sites.

    PubMed

    Dong, Chun-Hai; Tang, Wei-Ping; Liu, Jia-Yao

    2013-03-01

    The plant actin depolymerizing factor (ADF) binds to both monomeric and filamentous actin, and is directly involved in the depolymerization of actin filaments. To better understand the actin binding sites of the Arabidopsis thaliana L. AtADF1, we generated mutants of AtADF1 and investigated their functions in vitro and in vivo. Analysis of mutants harboring amino acid substitutions revealed that charged residues (Arg98 and Lys100) located at the α-helix 3 and forming an actin binding site together with the N-terminus are essential for both G- and F-actin binding. The basic residues on the β-strand 5 (K82/A) and the α-helix 4 (R135/A, R137/A) form another actin binding site that is important for F-actin binding. Using transient expression of CFP-tagged AtADF1 mutant proteins in onion (Allium cepa) peel epidermal cells and transgenic Arabidopsis thaliana L. plants overexpressing these mutants, we analyzed how these mutant proteins regulate actin organization and affect seedling growth. Our results show that the ADF mutants with a lower affinity for actin filament binding can still be functional, unless the affinity for actin monomers is also affected. The G-actin binding activity of the ADF plays an essential role in actin binding, depolymerization of actin polymers, and therefore in the control of actin organization. PMID:23190411

  3. The Molecular Evolution of Actin

    PubMed Central

    Hightower, Robin C.; Meagher, Richard B.

    1986-01-01

    We have investigated the molecular evolution of plant and nonplant actin genes comparing nucleotide and amino acid sequences of 20 actin genes. Nucleotide changes resulting in amino acid substitutions (replacement substitutions) ranged from 3–7% for all pairwise comparisons of animal actin genes with the following exceptions. Comparisons between higher animal muscle actin gene sequences and comparisons between higher animal cytoplasmic actin gene sequences indicated <3% divergence. Comparisons between plant and nonplant actin genes revealed, with two exceptions, 11–15% replacement substitution. In the analysis of plant actins, replacement substitution between soybean actin genes SAc1, SAc3, SAc4 and maize actin gene MAc1 ranged from 8–10%, whereas these members within the soybean actin gene family ranged from 6–9% replacement substitution. The rate of sequence divergence of plant actin sequences appears to be similar to that observed for animal actins. Furthermore, these and other data suggest that the plant actin gene family is ancient and that the families of soybean and maize actin genes have diverged from a single common ancestral plant actin gene that originated long before the divergence of monocots and dicots. The soybean actin multigene family encodes at least three classes of actin. These classes each contain a pair of actin genes that have been designated kappa (SAc1, SAc6), lambda (SAc2, SAc4) and mu (SAc3, SAc7). The three classes of soybean actin are more divergent in nucleotide sequence from one another than higher animal cytoplasmic actin is divergent from muscle actin. The location and distribution of amino acid changes were compared between actin proteins from all sources. A comparison of the hydropathy of all actin sequences, except from Oxytricha, indicated a strong similarity in hydropathic character between all plant and nonplant actins despite the greater number of replacement substitutions in plant actins. These protein sequence

  4. TEM Study of the Growth Mechanism, Phase Transformation, and Core/shell Structure of Semiconductor Nanowires

    NASA Astrophysics Data System (ADS)

    Wong, Tai Lun

    In this thesis, the fabrication and characterization of one-dimensional nanostructures have been studied systematically to understand the growth mechanism and structure transformation of one-dimensional nanostructures. The growth behavior of the ultrathin ZnSe nanowires with diameter less than 60 nm was found to be different from classical vapor-liquid-solid (VLS) process. The growth rate increases when the diameter of nanowires decreases, in contrast to the classical VLS process in which the growth rate increases with the diameter. The nucleation, initial growth, growth rates, defects, interface structures and growth direction of the nanowires were investigated by high resolution transmission electron microscopy (HRTEM). We found the structure and growth direction of ultra-thin nanowires are highly sensitive to growth temperatures and diameters of nanowires. At a low growth temperature (380°C), the growth direction for most nanowires is along <111>. Planar defects were found throughout the nanowires. At a high growth temperature (530°C), uniform nanowires with diameters around 10nm were grown along <110> and <112> directions, and the nanowires with diameters larger than 20nm were mainly grown along <111> direction. The possible growth mechanism of ultrathin nanowires was proposed by combining the solid catalytic growth with the interface diffusion theory, in order to explain how the growth temperature and the size of the catalysts influent the morphology, growth direction and growth rate of ultrathin nanowires. Structural and phase transformation of a nickel coated Si nanowire to NiSi2/SiC core-shell nanowire heterostructures has been investigated by the in-situ Transmission Electron Microscope (TEM). The phase transformation is a single-site nucleation process and therefore a single crystalline NiSi2 core resulted in the core-shell nanowire heterostructures. The transformation of the Si nanowire to NiSi2/SiC core-shell nanowire heterostructures was extremely

  5. Competition of two distinct actin networks for actin defines a bistable switch for cell polarization

    PubMed Central

    Lomakin, Alexis J.; Lee, Kun-Chun; Han, Sangyoon J.; Bui, D A.; Davidson, Michael; Mogilner, Alex; Danuser, Gaudenz

    2015-01-01

    Symmetry-breaking polarization enables functional plasticity of cells and tissues and is yet not well understood. Here we show that epithelial cells, hard-wired to maintain a static morphology and to preserve tissue organization, can spontaneously switch to a migratory polarized phenotype upon relaxation of the actomyosin cytoskeleton. We find that myosin-II engages actin in the formation of cortical actomyosin bundles and thus makes it unavailable for deployment in the process of dendritic growth normally driving cell motility. At low contractility regimes epithelial cells polarize in a front-back manner due to emergence of actin retrograde flows powered by dendritic polymerization of actin. Coupled to cell movement, the flows transport myosin-II from the front to the back of the cell, where the motor locally “locks” actin in contractile bundles. This polarization mechanism could be employed by embryonic and cancer epithelial cells in microenvironments where high contractility-driven cell motion is inefficient. PMID:26414403

  6. Wnt Signalling Promotes Actin Dynamics during Axon Remodelling through the Actin-Binding Protein Eps8

    PubMed Central

    Salinas, Patricia C.

    2015-01-01

    Upon arrival at their synaptic targets, axons slow down their growth and extensively remodel before the assembly of presynaptic boutons. Wnt proteins are target-derived secreted factors that promote axonal remodelling and synaptic assembly. In the developing spinal cord, Wnts secreted by motor neurons promote axonal remodelling of NT-3 responsive dorsal root ganglia neurons. Axon remodelling induced by Wnts is characterised by growth cone pausing and enlargement, processes that depend on the re-organisation of microtubules. However, the contribution of the actin cytoskeleton has remained unexplored. Here, we demonstrate that Wnt3a regulates the actin cytoskeleton by rapidly inducing F-actin accumulation in growth cones from rodent DRG neurons through the scaffold protein Dishevelled-1 (Dvl1) and the serine-threonine kinase Gsk3β. Importantly, these changes in actin cytoskeleton occurs before enlargement of the growth cones is evident. Time-lapse imaging shows that Wnt3a increases lamellar protrusion and filopodia velocity. In addition, pharmacological inhibition of actin assembly demonstrates that Wnt3a increases actin dynamics. Through a yeast-two hybrid screen, we identified the actin-binding protein Eps8 as a direct interactor of Dvl1, a scaffold protein crucial for the Wnt signalling pathway. Gain of function of Eps8 mimics Wnt-mediated axon remodelling, whereas Eps8 silencing blocks the axon remodelling activity of Wnt3a. Importantly, blockade of the Dvl1-Eps8 interaction completely abolishes Wnt3a-mediated axonal remodelling. These findings demonstrate a novel role for Wnt-Dvl1 signalling through Eps8 in the regulation of axonal remodeling. PMID:26252776

  7. Intranuclear Actin Regulates Osteogenesis

    PubMed Central

    Sen, Buer; Xie, Zhihui; Uzer, Gunes; Thompson, William R.; Styner, Maya; Wu, Xin; Rubin, Janet

    2016-01-01

    Depolymerization of the actin cytoskeleton induces nuclear trafficking of regulatory proteins and global effects on gene transcription. We here show that in mesenchymal stem cells (MSCs), cytochalasin D treatment causes rapid cofilin-/importin-9-dependent transfer of G-actin into the nucleus. The continued presence of intranuclear actin, which forms rod-like structures that stain with phalloidin, is associated with induction of robust expression of the osteogenic genes osterix and osteocalcin in a Runx2-dependent manner, and leads to acquisition of osteogenic phenotype. Adipogenic differentiation also occurs, but to a lesser degree. Intranuclear actin leads to nuclear export of Yes-associated protein (YAP); maintenance of nuclear YAP inhibits Runx2 initiation of osteogenesis. Injection of cytochalasin into the tibial marrow space of live mice results in abundant bone formation within the space of 1 week. In sum, increased intranuclear actin forces MSC into osteogenic lineage through controlling Runx2 activity; this process may be useful for clinical objectives of forming bone. PMID:26140478

  8. Effects of transformed ferrite growth on the tensile fracture characteristics of a dual-phase steel

    NASA Astrophysics Data System (ADS)

    Jeong, W. C.; Kim, C. H.

    1988-02-01

    The effects of transformed ferrite growth on the tensile fracture characteristics of a dual-phase steel were investigated by observing crack initiation, propagation, and fracture behaviors. Crack initiation occurred by decohesion between martensite and ferrite. However, cracks propagated along the ferrite-martensite interface in a high temperature quenched specimen, whereas in specimens quenched from lower temperature cracks propagated into the martensite particle. Tensile fracture behaviors were not strongly influenced by the cooling rate. At both cooling rates of 5.6 and 0.1 °C/sec, specimens quenched from high temperature fractured by partially brittle fracture mode, but fracture mode changed to ductile mode as the quenching temperature decreased. The effect of transformed ferrite on the fracture mode was not substantially different from that of retained ferrite. However, the crack initiation and propagation was influenced by the variation in martensite distribution caused by different growth behavior of transformed ferrite.

  9. Stable Glass Transformation to Supercooled Liquid via Surface-Initiated Growth Front

    SciTech Connect

    Swallen, Stephen F.; Traynor, Katherine; McMahon, Robert J.; Ediger, M. D.; Mates, Thomas E.

    2009-02-13

    Highly stable glasses of tris-naphthylbenzene transform into a liquid when annealed above the glass transition temperature T{sub g}. In contrast to the predictions of standard models, the observed transformation is spatially inhomogeneous. Secondary ion mass spectrometry experiments on isotopically labeled multilayer films show that the liquid grows into the stable glass with sharp growth fronts initiated at the free surface and at the interface with the substrate. For the free surface, the growth velocity is constant in time and has the same temperature dependence as self-diffusion in the equilibrium supercooled liquid. These stable glasses are packed so efficiently that surfaces and interfaces are required to initiate the transformation to the liquid even well above T{sub g}.

  10. He-Ne laser influenced actin filaments alleviate the damage of UV-B in wheat

    NASA Astrophysics Data System (ADS)

    Chen, Huize; Han, Rong

    2015-01-01

    This work investigated the use of a He-Ne laser in alleviating the damaging effects of ultraviolet-B (UV-B) radiation on wheat seedlings by influenced actin filaments. Triticum aestivum seedlings were irradiated with either enhanced UV-B (10.08 KJ m-2 d-1) or a combination of UV-B light and the He-Ne laser. Plants were also exposed to the He-Ne laser alone. In order to compare the effect of the He-Ne laser, red light (same power and wavelength as the He-Ne laser) treatment and the combined UV-B and red light treatment were added. Moreover, wheat seedlings were treated with actin special drugs, including cytochalasin B (CB) and jasplakinolide (JAS). We analyzed the growth of the seedlings, the distribution of actin filaments (AFs), DNA laddering and ACTIN expression in the different groups. The results showed that enhanced UV-B produced negative effects on the growth of wheat seedlings while implementing the He-Ne laser partially alleviated the injury. With the red light treatment, there are no positive effects. The ACTIN expression stayed the same in the different treatments, while the distribution and the protein content are different. The Fourier transform infrared (FTIR) microspectroscopic results further established significant changes in the chemical composition of the wall material. These results suggested that the He-Ne laser alleviated the damaging effects of UV-B radiation in wheat seedlings by changing the characteristics of the AFs.

  11. Myosins, Actin and Autophagy.

    PubMed

    Kruppa, Antonina J; Kendrick-Jones, John; Buss, Folma

    2016-08-01

    Myosin motor proteins working together with the actin cytoskeleton drive a wide range of cellular processes. In this review, we focus on their roles in autophagy - the pathway the cell uses to ensure homeostasis by targeting pathogens, misfolded proteins and damaged organelles for degradation. The actin cytoskeleton regulated by a host of nucleating, anchoring and stabilizing proteins provides the filament network for the delivery of essential membrane vesicles from different cellular compartments to the autophagosome. Actin networks have also been implicated in structurally supporting the expanding phagophore, moving autophagosomes and enabling efficient fusion with the lysosome. Only a few myosins have so far been shown to play a role in autophagy. Non-muscle myosin IIA functions in the early stages delivering membrane for the initial formation of the autophagosome, whereas myosin IC and myosin VI are involved in the final stages providing specific membranes for autophagosome maturation and its fusion with the lysosome. PMID:27146966

  12. Divergent effects of epidermal growth factor and transforming growth factors on a human endometrial carcinoma cell line.

    PubMed

    Korc, M; Haussler, C A; Trookman, N S

    1987-09-15

    Epidermal growth factor (EGF), at concentrations ranging from 0.83 to 4.98 nM, markedly inhibited the proliferation of RL95-2 cells that were seeded at low plating densities (4.7 X 10(3) cells/cm2). Under the same incubation conditions, 16.6 pM EGF enhanced cell proliferation. At high plating densities (2.5 X 10(4) cells/cm2) 0.83 nM EGF also stimulated cell proliferation. Both the inhibitory and stimulatory effects of EGF were mimicked by transforming growth factor-alpha (TGF-alpha). However, the inhibitory action of TGF-alpha was always greater that of EGF. Binding studies with 125I-labeled TGF-alpha indicated that maximal cell surface binding of TGF-alpha occurred at 15 min, whereas maximal internalization occurred at 45 min. Both cell surface and internalized radioactivity declined sharply thereafter. Analysis of radioactivity released into the incubation medium during pulse-chase experiments indicated that RL95-2 cells extensively degraded both TGF-alpha and EGF. The lysosomotropic compound methylamine arrested the generation of low-molecular-weight degradation products of EGF, but not of TGF-alpha. In contrast to EGF and TGF-alpha, transforming growth factor-beta (TGF-beta) inhibited the proliferation of RL95-2 cells that were seeded at either low or high plating densities. Further, transforming growth factor-beta induced the appearance of large cuboidal cells that were readily distinguished from cells treated with either EGF or TGF-alpha. These findings point to complex regulatory actions of growth factors on the proliferation of RL95-2 cells and suggest that the processing of TGF-alpha following EGF receptor activation is distinct from the processing of EGF. PMID:3497713

  13. Force Generation by Endocytic Actin Patches in Budding Yeast

    PubMed Central

    Carlsson, Anders E.; Bayly, Philip V.

    2014-01-01

    Membrane deformation during endocytosis in yeast is driven by local, templated assembly of a sequence of proteins including polymerized actin and curvature-generating coat proteins such as clathrin. Actin polymerization is required for successful endocytosis, but it is not known by what mechanisms actin polymerization generates the required pulling forces. To address this issue, we develop a simulation method in which the actin network at the protein patch is modeled as an active gel. The deformation of the gel is treated using a finite-element approach. We explore the effects and interplay of three different types of force driving invagination: 1), forces perpendicular to the membrane, generated by differences between actin polymerization rates at the edge of the patch and those at the center; 2), the inherent curvature of the coat-protein layer; and 3), forces parallel to the membrane that buckle the coat protein layer, generated by an actomyosin contractile ring. We find that with optimistic estimates for the stall stress of actin gel growth and the shear modulus of the actin gel, actin polymerization can generate almost enough force to overcome the turgor pressure. In combination with the other mechanisms, actin polymerization can the force over the critical value. PMID:24739159

  14. Atomic mechanisms of diffusional nucleation and growth and comparisons with their counterparts in shear transformations

    NASA Astrophysics Data System (ADS)

    Aaronson, Hubert I.

    1993-02-01

    An integrated overview is presented of a viewpoint on the present understanding of nucleation and growth mechanisms in both diffusional and shear (martensitic) transformations. Special emphasis is placed on the roles played by the anisotropy of interphase boundary structure and energy and also upon elastic shear strain energy in both types of transformation. Even though diffusional nucleation is based on random statistical fluctuations, use of the time reversal principle shows that interfacial energy anisotropy leads to accurately reproducible orientation relationships and hence to partially or fully coherent boundaries, even when nucleation at a grain boundary requires an irrational orientation relationship to obtain. Since the fully coherent boundary areas separating most linear misfit compensating defects are wholly immobile during diffusional growth because of the improbability of moving substitutional atoms even temporarily into interstitial sites under conditions normally encountered, partially and fully coherent interphase boundaries should be immovable without the intervention of growth ledges. These ledges, however, must be heavily kinked and usually irregular in both spacing and path if they, too, are not to be similarly trapped. On the other hand, the large shear strain energy usually associated with martensite requires that its formation be initiated through a process which avoids the activation barrier associated with nucleation, perhaps by the Olson-Cohen matrix dislocation rearrangement mechanism. During growth, certain ledges on martensite plates serve as transformation dislocations and perform the crystal structure change (Bain strain). However, the terraces between these ledges in martensite (unlike those present during diffusional growth) are also mobile during non-fcc/hcp transformations; glissile dislocations on these terraces perform the lattice invariant deformation. Growth ledges operative during both diffusional and shear growth probably

  15. Growth-arrest-specific 7C protein inhibits tumor metastasis via the N-WASP/FAK/F-actin and hnRNP U/β-TrCP/β-catenin pathways in lung cancer

    PubMed Central

    Chang, Jer-Wei; Mao, Jiou-Shan; Tsai, Charng-Dar; Wu, Pei-Chen; Lin, Cuei-Jyuan; Lu, Yi-Lin; Liao, Sheng-You; Cheng, Hung-Chi; Hsu, Han-Shui

    2015-01-01

    Growth-arrest-specific 7 (GAS7) belongs to a group of adaptor proteins that coordinate the actin cytoskeleton. Among human GAS7 isoforms, only GAS7C possesses a Src homology 3 domain. We report here that GAS7C acts as a migration suppressor and can serve as a prognostic biomarker in lung cancer. GAS7C overexpression reduces lung cancer migration, whereas GAS7C knockdown enhances cancer cell migration. Importantly, ectopically overexpressed GAS7C binds tightly with N-WASP thus inactivates the fibronectin/integrin/FAK pathway, which in turn leads to the suppression of F-actin dynamics. In addition, overexpression of GAS7C sequesters hnRNP U and thus decreases the level of β-catenin protein via the β-TrCP ubiquitin-degradation pathway. The anti-metastatic effect of GAS7C overexpression was also confirmed using lung cancer xenografts. Our clinical data indicated that 23.6% (25/106) of lung cancer patients showed low expression of GAS7C mRNA which correlated with a poorer overall survival. In addition, low GAS7C mRNA expression was detected in 60.0% of metastatic lung cancer patients, indicating an association between low GAS7C expression and cancer progression. A significant inverse correlation between mRNA expression and promoter hypermethylation was also found, which suggests that the low level of GAS7C expression was partly due to promoter hypermethylation. Our results provide novel evidence that low GAS7C correlates with poor prognosis and promotes metastasis in lung cancer. Low GAS7C increases cancer cell motility by promoting N-WASP/FAK/F-actin cytoskeleton dynamics. It also enhances β-catenin stability via hnRNP U/β-TrCP complex formation. Therefore, GAS7C acts as a metastasis suppressor in lung cancer. PMID:26506240

  16. Actin Filament Segmentation Using Dynamic Programming

    PubMed Central

    Li, Hongsheng; Shen, Tian; Huang, Xiaolei

    2011-01-01

    We introduce a novel algorithm for actin filament segmentation in 2D TIRFM image sequences. This problem is difficult because actin filaments dynamically change shapes during their growth, and the TIRFM images are usually noisy. We ask a user to specify the two tips of a filament of interest in the first frame. We then model the segmentation problem in an image sequence as a temporal chain, where its states are tip locations; given candidate tip locations, actin filaments' body points are inferred by a dynamic programming method, which adaptively generates candidate solutions. Combining candidate tip locations and their inferred body points, the temporal chain model is efficiently optimized using another dynamic programming method. Evaluation on noisy TIRFM image sequences demonstrates the accuracy and robustness of this approach. PMID:21761674

  17. Combined effects of interleukin-1α and transforming growth factor-β1 on modulation of human cardiac fibroblast function.

    PubMed

    van Nieuwenhoven, Frans A; Hemmings, Karen E; Porter, Karen E; Turner, Neil A

    2013-01-01

    During cardiac remodeling, cardiac fibroblasts (CF) are influenced by increased levels of interleukin-1α (IL-1α) and transforming growth factor-β1 (TGFβ1). The present study investigated the interaction between these two important cytokines on function of human CF and their differentiation to myofibroblasts (CMF). CF were isolated from human atrial appendage and exposed to IL-1α and/or TGFβ1 (both 0.1 ng/ml). mRNA expression levels of selected genes were determined after 6-24h by real-time RT-PCR, while protein levels were analyzed at 24-48 h by ELISA or western blot. Activation of canonical signaling pathways (NFκB, Smad3, p38 MAPK) was determined by western blotting. Differentiation to CMF was examined by collagen gel contraction assays. Exposure of CF to IL-1α alone enhanced levels of IL-6, IL-8, matrix metalloproteinase-3 (MMP3) and collagen III (COL3A1), but reduced the CMF markers α-smooth muscle actin (αSMA) and connective tissue growth factor (CTGF/CCN2). By contrast, TGFβ1 alone had minor effects on IL-6, IL-8 and MMP3 levels, but significantly increased levels of the CMF markers αSMA, CTGF, COL1A1 and COL3A1. Co-stimulation with both IL-1α and TGFβ1 increased MMP3 expression synergistically. Furthermore, while TGFβ1 had no effect on IL-1α-induced IL-6 or IL-8 levels, co-stimulation inhibited the TGFβ1-induced increase in αSMA and blocked the gel contraction caused by TGFβ1. Combining IL-1α and TGFβ1 had no apparent effect on their canonical signaling pathways. In conclusion, IL-1α and TGFβ1 act synergistically to stimulate MMP3 expression in CF. Moreover, IL-1α has a dominant inhibitory effect on the phenotypic switch of CF to CMF induced by TGFβ1. PMID:23583823

  18. A synthetic mechano-growth factor E peptide promotes rat tenocyte migration by lessening cell stiffness and increasing F-actin formation via the FAK-ERK1/2 signaling pathway

    SciTech Connect

    Zhang, Bingyu; Luo, Qing; Mao, Xinjian; Xu, Baiyao; Yang, Li; Ju, Yang; Song, Guanbin

    2014-03-10

    Tendon injuries are common in sports and are frequent reasons for orthopedic consultations. The management of damaged tendons is one of the most challenging problems in orthopedics. Mechano-growth factor (MGF), a recently discovered growth repair factor, plays positive roles in tissue repair through the improvement of cell proliferation and migration and the protection of cells against injury-induced apoptosis. However, it remains unclear whether MGF has the potential to accelerate tendon repair. We used a scratch wound assay in this study to demonstrate that MGF-C25E (a synthetic mechano-growth factor E peptide) promotes the migration of rat tenocytes and that this promotion is accompanied by an elevation in the expression of the following signaling molecules: focal adhesion kinase (FAK) and extracellular signal regulated kinase1/2 (ERK1/2). Inhibitors of the FAK and ERK1/2 pathways inhibited the MGF-C25E-induced tenocyte migration, indicating that MGF-C25E promotes tenocyte migration through the FAK-ERK1/2 signaling pathway. The analysis of the mechanical properties showed that the Young's modulus of tenocytes was decreased through treatment of MGF-C25E, and an obvious formation of pseudopodia and F-actin was observed in MGF-C25E-treated tenocytes. The inhibition of the FAK or ERK1/2 signals restored the decrease in Young's modulus and inhibited the formation of pseudopodia and F-actin. Overall, our study demonstrated that MGF-C25E promotes rat tenocyte migration by lessening cell stiffness and increasing pseudopodia formation via the FAK-ERK1/2 signaling pathway. - Highlights: • Mechano-growth factor E peptide (MGF-C25E) promotes migration of rat tenocytes. • MGF-C25E activates the FAK-ERK1/2 pathway in rat tenocytes. • MGF-C25E induces the actin remodeling and the formation of pseudopodia, and decreases the stiffness in rat tenocytes. • MGF-C25E promotes tenocyte migration via altering stiffness and forming pseudopodia by the activation of the FAK-ERK1

  19. A small molecule inhibitor of tropomyosin dissociates actin binding from tropomyosin-directed regulation of actin dynamics

    PubMed Central

    Bonello, Teresa T.; Janco, Miro; Hook, Jeff; Byun, Alex; Appaduray, Mark; Dedova, Irina; Hitchcock-DeGregori, Sarah; Hardeman, Edna C.; Stehn, Justine R.; Böcking, Till; Gunning, Peter W.

    2016-01-01

    The tropomyosin family of proteins form end-to-end polymers along the actin filament. Tumour cells rely on specific tropomyosin-containing actin filament populations for growth and survival. To dissect out the role of tropomyosin in actin filament regulation we use the small molecule TR100 directed against the C terminus of the tropomyosin isoform Tpm3.1. TR100 nullifies the effect of Tpm3.1 on actin depolymerisation but surprisingly Tpm3.1 retains the capacity to bind F-actin in a cooperative manner. In vivo analysis also confirms that, in the presence of TR100, fluorescently tagged Tpm3.1 recovers normally into stress fibers. Assembling end-to-end along the actin filament is thereby not sufficient for tropomyosin to fulfil its function. Rather, regulation of F-actin stability by tropomyosin requires fidelity of information communicated at the barbed end of the actin filament. This distinction has significant implications for perturbing tropomyosin-dependent actin filament function in the context of anti-cancer drug development. PMID:26804624

  20. TRANSFORMATION

    SciTech Connect

    LACKS,S.A.

    2003-10-09

    Transformation, which alters the genetic makeup of an individual, is a concept that intrigues the human imagination. In Streptococcus pneumoniae such transformation was first demonstrated. Perhaps our fascination with genetics derived from our ancestors observing their own progeny, with its retention and assortment of parental traits, but such interest must have been accelerated after the dawn of agriculture. It was in pea plants that Gregor Mendel in the late 1800s examined inherited traits and found them to be determined by physical elements, or genes, passed from parents to progeny. In our day, the material basis of these genetic determinants was revealed to be DNA by the lowly bacteria, in particular, the pneumococcus. For this species, transformation by free DNA is a sexual process that enables cells to sport new combinations of genes and traits. Genetic transformation of the type found in S. pneumoniae occurs naturally in many species of bacteria (70), but, initially only a few other transformable species were found, namely, Haemophilus influenzae, Neisseria meningitides, Neisseria gonorrheae, and Bacillus subtilis (96). Natural transformation, which requires a set of genes evolved for the purpose, contrasts with artificial transformation, which is accomplished by shocking cells either electrically, as in electroporation, or by ionic and temperature shifts. Although such artificial treatments can introduce very small amounts of DNA into virtually any type of cell, the amounts introduced by natural transformation are a million-fold greater, and S. pneumoniae can take up as much as 10% of its cellular DNA content (40).

  1. Organization and regulation of the actin cytoskeleton in the pollen tube

    PubMed Central

    Qu, Xiaolu; Jiang, Yuxiang; Chang, Ming; Liu, Xiaonan; Zhang, Ruihui; Huang, Shanjin

    2015-01-01

    Proper organization of the actin cytoskeleton is crucial for pollen tube growth. However, the precise mechanisms by which the actin cytoskeleton regulates pollen tube growth remain to be further elucidated. The functions of the actin cytoskeleton are dictated by its spatial organization and dynamics. However, early observations of the distribution of actin filaments at the pollen tube apex were quite perplexing, resulting in decades of controversial debate. Fortunately, due to improvements in fixation regimens for staining actin filaments in fixed pollen tubes, as well as the adoption of appropriate markers for visualizing actin filaments in living pollen tubes, this issue has been resolved and has given rise to the consensus view of the spatial distribution of actin filaments throughout the entire pollen tube. Importantly, recent descriptions of the dynamics of individual actin filaments in the apical region have expanded our understanding of the function of actin in regulation of pollen tube growth. Furthermore, careful documentation of the function and mode of action of several actin-binding proteins expressed in pollen have provided novel insights into the regulation of actin spatial distribution and dynamics. In the current review, we summarize our understanding of the organization, dynamics, and regulation of the actin cytoskeleton in the pollen tube. PMID:25620974

  2. Handling Arabidopsis plants: growth, preservation of seeds, transformation, and genetic crosses.

    PubMed

    Rivero, Luz; Scholl, Randy; Holomuzki, Nicholas; Crist, Deborah; Grotewold, Erich; Brkljacic, Jelena

    2014-01-01

    Growing healthy plants is essential for the advancement of Arabidopsis thaliana (Arabidopsis) research. Over the last 20 years, the Arabidopsis Biological Resource Center (ABRC) has collected and developed a series of best-practice protocols, some of which are presented in this chapter. Arabidopsis can be grown in a variety of locations, growth media, and environmental conditions. Most laboratory accessions and their mutant or transgenic derivatives flower after 4-5 weeks and set seeds after 7-8 weeks, under standard growth conditions (soil, long day, 23 ºC). Some mutant genotypes, natural accessions, and Arabidopsis relatives require strict control of growth conditions best provided by growth rooms, chambers, or incubators. Other lines can be grown in less-controlled greenhouse settings. Although the majority of lines can be grown in soil, certain experimental purposes require utilization of sterile solid or liquid growth media. These include the selection of primary transformants, identification of homozygous lethal individuals in a segregating population, or bulking of a large amount of plant material. The importance of controlling, observing, and recording growth conditions is emphasized and appropriate equipment required to perform monitoring of these conditions is listed. Proper conditions for seed harvesting and preservation, as well as seed quality control, are also described. Plant transformation and genetic crosses, two of the methods that revolutionized Arabidopsis genetics, are introduced as well. PMID:24057358

  3. Arp2/3 complex-dependent actin networks constrain myosin II function in driving retrograde actin flow.

    PubMed

    Yang, Qing; Zhang, Xiao-Feng; Pollard, Thomas D; Forscher, Paul

    2012-06-25

    The Arp2/3 complex nucleates actin filaments to generate networks at the leading edge of motile cells. Nonmuscle myosin II produces contractile forces involved in driving actin network translocation. We inhibited the Arp2/3 complex and/or myosin II with small molecules to investigate their respective functions in neuronal growth cone actin dynamics. Inhibition of the Arp2/3 complex with CK666 reduced barbed end actin assembly site density at the leading edge, disrupted actin veils, and resulted in veil retraction. Strikingly, retrograde actin flow rates increased with Arp2/3 complex inhibition; however, when myosin II activity was blocked, Arp2/3 complex inhibition now resulted in slowing of retrograde actin flow and veils no longer retracted. Retrograde flow rate increases induced by Arp2/3 complex inhibition were independent of Rho kinase activity. These results provide evidence that, although the Arp2/3 complex and myosin II are spatially segregated, actin networks assembled by the Arp2/3 complex can restrict myosin II-dependent contractility with consequent effects on growth cone motility. PMID:22711700

  4. Sustained activation of fibroblast transforming growth factor-beta/Smad signaling in a murine model of scleroderma.

    PubMed

    Takagawa, Shinsuke; Lakos, Gabriella; Mori, Yasuji; Yamamoto, Toshiyuki; Nishioka, Kiyoshi; Varga, John

    2003-07-01

    Transforming growth factor-beta is responsible for triggering a cascade of events leading to fibrosis in scleroderma. The Smads are intracellular signal transducers recently shown to mediate fibroblast activation and other profibrotic responses elicited by transforming growth factor-betain vitro. To understand better the involvement of Smads in the pathogenesis of fibrosis, we examined Smad expression and activation in situ in a murine model of scleroderma. Bleomycin injections induced striking dermal infiltration with macrophages by 3 d, and progressive fibrosis by 2 wk. Infiltrating macrophages and resident fibroblasts expressed Smad3, the positive mediator for transforming growth factor-beta responses. Importantly, in bleomycin-injected skin, fibroblasts showed predominantly nuclear localization of Smad3 and intense staining for phospho-Smad2/3. Furthermore, phosphorylated Smad2/3 in fibroblasts was detected even after the resolution of inflammation. Expression of Smad7, the endogenous inhibitor of transforming growth factor-beta/Smad signaling, was strongly induced in dermal cells by transforming growth factor-beta, but not by bleomycin injections. Collectively, these results indicate that bleomycin-induced murine scleroderma is associated with rapid and sustained induction of transforming growth factor-beta/Smad signaling in resident dermal fibroblasts. Despite apparent activation of the intracellular transforming growth factor-beta signaling pathway in the lesional dermis, the expression of transforming growth factor-beta-inducible Smad7 was not upregulated. In light of the critical function of Smad7 as an endogenous inhibitor of Smad signaling that restricts the duration and magnitude of transforming growth factor-beta responses, and as a mediator of apoptosis, relative Smad7 deficiency observed in the present studies may account for sustained activation of transforming growth factor-beta/Smad signaling in lesional tissues. These findings raise the

  5. TRANSFORMER

    DOEpatents

    Baker, W.R.

    1959-08-25

    Transformers of a type adapted for use with extreme high power vacuum tubes where current requirements may be of the order of 2,000 to 200,000 amperes are described. The transformer casing has the form of a re-entrant section being extended through an opening in one end of the cylinder to form a coaxial terminal arrangement. A toroidal multi-turn primary winding is disposed within the casing in coaxial relationship therein. In a second embodiment, means are provided for forming the casing as a multi-turn secondary. The transformer is characterized by minimized resistance heating, minimized external magnetic flux, and an economical construction.

  6. Transforming Growth Factor β/activin signalling induces epithelial cell flattening during Drosophila oogenesis

    PubMed Central

    Brigaud, Isabelle; Duteyrat, Jean-Luc; Chlasta, Julien; Le Bail, Sandrine; Couderc, Jean-Louis; Grammont, Muriel

    2015-01-01

    ABSTRACT Although the regulation of epithelial morphogenesis is essential for the formation of tissues and organs in multicellular organisms, little is known about how signalling pathways control cell shape changes in space and time. In the Drosophila ovarian epithelium, the transition from a cuboidal to a squamous shape is accompanied by a wave of cell flattening and by the ordered remodelling of E-cadherin-based adherens junctions. We show that activation of the TGFβ pathway is crucial to determine the timing, the degree and the dynamic of cell flattening. Within these cells, TGFβ signalling controls cell-autonomously the formation of Actin filament and the localisation of activated Myosin II, indicating that internal forces are generated and used to remodel AJ and to promote cytoskeleton rearrangement. Our results also reveal that TGFβ signalling controls Notch activity and that its functions are partly executed through Notch. Thus, we demonstrate that the cells that undergo the cuboidal-to-squamous transition produce active cell-shaping mechanisms, rather than passively flattening in response to a global force generated by the growth of the underlying cells. Thus, our work on TGFβ signalling provides new insights into the mechanisms through which signal transduction cascades orchestrate cell shape changes to generate proper organ structure. PMID:25681395

  7. [THE ROLE OF TRANSFORMING GROWTH FACTOR-B IN IMMUNOPATHOGENESIS OF DISEASES OF CONNECTIVE TISSUE].

    PubMed

    Rudoi, A S; Moskalev, A V; Sboitchakov, V B

    2016-02-01

    The recent studies of molecular physiology of fibrillin and pathophysiology of inherent disorders of structure and function of connective tissue such as dissection and aneurysm of aorta, myxomatously altered cusps and prolapses of mitral valve, syndrome of hyper-mobility of joints, demonstrated that important role in development of these malformations play alterations of transfer of signals by growth factors and matrix cellular interaction. These conditions under manifesting Marfan's syndrome can be a consequence of anomalies of fibrillin-1 which deficiency unbrakes process of activation of transforming growth factor-β (TGFβ). The involvement of TGFβ in pathogenesis of Marfan's syndrome permits consider antagonists of angiotensin-transforming enzymes as potential pharmaceuticals in therapy of this disease. The article presents analysis of publications' data related to this problem. PMID:27455564

  8. Transformation of the Herbicide Sulcotrione into a Root Growth Enhancer Compound by Sequential Photolysis and Hydrolysis.

    PubMed

    Goujon, Eric; Maruel, Sandra; Richard, Claire; Goupil, Pascale; Ledoigt, Gérard

    2016-01-27

    Xanthene-1,9-dione-3,4-dihydro-6-methylsulfonyl (1), the main product of sulcotrione phototransformation on plant leaves, was slowly hydrolyzed into 2-hydroxy-4-methylsulfonylbenzoic acid (2) and 1,3-cyclohexanedione (3) in aqueous solution. Interestingly, the rate of hydrolysis was significantly enhanced in the presence of roots of monocotyledonous plants, while the same treatment showed adverse effects on broadleaf weeds. Root growth enhancement varied according to the plant species and concentrations of compound 2, as shown with Zea mays roots. Compound 2 is a derivative of salicylic acid that is known to be a plant signaling messenger. Compound 2 was, therefore, able to mimic some known effects of this phytohormone. This work showed that a pesticide like sulcotrione was transformed into a compound exhibiting a positive impact on plant growth. This study exemplified a rarely reported situation where chemical and biological chain reactions transformed a xenobiotic into a compound exhibiting potential beneficial effects. PMID:26654319

  9. 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.

  10. [Effects of nitrogen regulators on fertilizer nitrogen transformation in meadow cinnamon soil and on pakchoi growth].

    PubMed

    Sun, Zhi-Mei; Zhang, Kuo; Liu, Jian-Tao; Si, Huan-Sen; Wang, Yan-Qun

    2012-09-01

    Soil incubation test and pot experiment were conducted to investigate the effects of dicyandiamide (DCD) and its combination with nano-carbon on the transformation of fertilizers (urea and ammonium bicarbonate) nitrogen (N) in meadow cinnamon soil, a typical soil type in North China Plain, and on the growth of pakchoi (Brassica chinensis). In the first two weeks after applying urea and ammonium bicarbonate, the soil NH4+-N and NO3(-)-N contents varied greatly, but little variation was observed since then. The effects of the applied fertilizer N on the pakchoi growth and its N use efficiency differed significantly at early growth stages, but had little difference at harvesting stage. The DCD inhibited the transformation of the fertilizer N (especially ammonium bicarbonate N) into nitrate markedly, and this effect increased with increasing DCD dose. Under the conditions of our experiment, the optimal application rate of DCD was 1.0-1.5% of applied fertilize N, which could increase the pakchoi yield significantly, improve the leaf color, decrease the plant nitrate contents, and increase the fertilizer N use efficiency. The combination of DCD and nano-carbon exerted a synergistic effect on inhibiting soil ammonium oxidation, and also, promoted the pakchoi growth and N utilization at early growth stages significantly and decreased the plant nitrate level at harvesting stage. PMID:23286007

  11. Synthesis of a 35-Member Stereoisomer Library of Bistramide A: Evaluation of Effects on Actin State, Cell Cycle and Tumor Cell Growth

    PubMed Central

    Wrona, Iwona E.; Lowe, Jason T.; Turbyville, Thomas J.; Johnson, Tanya R.; Beignet, Julien; Beutler, John A.; Panek, James S.

    2011-01-01

    Synthesis and preliminary biological evaluation of a 35-member library of bistramide A stereoisomers are reported. All eight stereoisomers of the C1-C13 tetrahydropyran fragment of the molecule were prepared utilizing crotylsilane reagents 9 and 10 in our [4+2]-annulation methodology. In addition, the four isomers of the C14-C18 γ-amino acid unit were accessed via a Lewis acid mediated crotylation reaction using both enantiomers of organosilane 11. The spiroketal subunit of bistramide A was modified at the C39-alcohol to give another point of stereochemical diversification. The fragments were coupled using standard peptide coupling protocol to provide 35 stereoisomers of the natural product. These stereochemical analogs were screened for their effects on cellular actin and cytotoxicity against cancer cell lines (UO-31 renal and SF-295 CNS). The results of these assays identified one analog, 1.21, with enhanced potency relative to the natural product, bistramide A. PMID:19191575

  12. The Actin-Binding Protein α-Adducin Is Required for Maintaining Axon Diameter.

    PubMed

    Leite, Sérgio Carvalho; Sampaio, Paula; Sousa, Vera Filipe; Nogueira-Rodrigues, Joana; Pinto-Costa, Rita; Peters, Luanne Laurel; Brites, Pedro; Sousa, Mónica Mendes

    2016-04-19

    The actin-binding protein adducin was recently identified as a component of the neuronal subcortical cytoskeleton. Here, we analyzed mice lacking adducin to uncover the function of this protein in actin rings. α-adducin knockout mice presented progressive axon enlargement in the spinal cord and optic and sciatic nerves, followed by axon degeneration and loss. Using stimulated emission depletion super-resolution microscopy, we show that a periodic subcortical actin cytoskeleton is assembled in every neuron type inspected including retinal ganglion cells and dorsal root ganglia neurons. In neurons devoid of adducin, the actin ring diameter increased, although the inter-ring periodicity was maintained. In vitro, the actin ring diameter adjusted as axons grew, suggesting the lattice is dynamic. Our data support a model in which adducin activity is not essential for actin ring assembly and periodicity but is necessary to control the diameter of both actin rings and axons and actin filament growth within rings. PMID:27068466

  13. Severe inflammatory bowel disease associated with congenital alteration of transforming growth factor beta signaling.

    PubMed

    Naviglio, Samuele; Arrigo, Serena; Martelossi, Stefano; Villanacci, Vincenzo; Tommasini, Alberto; Loganes, Claudia; Fabretto, Antonella; Vignola, Silvia; Lonardi, Silvia; Ventura, Alessandro

    2014-08-01

    Transforming growth factor beta is a pleiotropic cytokine which plays a central role in the homeostasis of the immune system. A complex dysregulation of its signaling occurs in Loeys-Dietz syndrome, a monogenic disorder caused by mutations of transforming growth factor beta receptors type 1 or type 2, characterized by skeletal involvement, craniofacial abnormalities, and arterial tortuosity with a strong predisposition for aneurysm and dissection. In addition, several immunologic abnormalities have been described in these patients, including an increased risk of allergic disorders as well as eosinophilic gastrointestinal disorders. The occurrence of inflammatory bowel disorders has been also reported, but it is poorly documented. We describe two unrelated children with Loeys-Dietz syndrome affected by severe chronic inflammatory colitis appearing at an early age. The intestinal disease presented similar features in both patients, including a histopathological picture of non-eosinophilic chronic ulcerative colitis, striking elevation of inflammatory markers, and a distinctly severe clinical course leading to failure to thrive, with resistance to multiple immunosuppressive treatments. One of the patients also presented autoimmune thyroiditis. Our report confirms that chronic ulcerative colitis may be associated with Loeys-Dietz syndrome. This finding suggests that an alteration of transforming growth factor beta signaling may by itself predispose to inflammatory colitis in humans, and represent an invaluable model to understand inflammatory bowel diseases. PMID:24486179

  14. Deletion of ocular transforming growth factor β signaling mimics essential characteristics of diabetic retinopathy.

    PubMed

    Braunger, Barbara M; Leimbeck, Sarah V; Schlecht, Anja; Volz, Cornelia; Jägle, Herbert; Tamm, Ernst R

    2015-06-01

    Diabetic retinopathy, a major cause of blindness, is characterized by a distinct phenotype. The molecular causes of the phenotype are not sufficiently clear. Here, we report that deletion of transforming growth factor β signaling in the retinal microenvironment of newborn mice induces changes that largely mimic the phenotype of nonproliferative and proliferative diabetic retinopathy in humans. Lack of transforming growth factor β signaling leads to the formation of abundant microaneurysms, leaky capillaries, and retinal hemorrhages. Retinal capillaries are not covered by differentiated pericytes, but by a coat of vascular smooth muscle-like cells and a thickened basal lamina. Reactive microglia is found in close association with retinal capillaries. In older animals, loss of endothelial cells and the formation of ghost vessels are observed, findings that correlate with the induction of angiogenic molecules and the accumulation of retinal hypoxia-inducible factor 1α, indicating hypoxia. Consequently, retinal and vitreal neovascularization occurs, a scenario that leads to retinal detachment, vitreal hemorrhages, neuronal apoptosis, and impairment of sensory function. We conclude that transforming growth factor β signaling is required for the differentiation of retinal pericytes during vascular development of the retina. Lack of differentiated pericytes initiates a scenario of structural and functional changes in the retina that mimics those of diabetic retinopathy strongly indicating a common mechanism. PMID:25857227

  15. Role of LncRNA-activated by transforming growth factor beta in the progression of hepatitis C virus-related liver fibrosis.

    PubMed

    Fu, Na; Niu, Xuemin; Wang, Yang; Du, Huijuan; Wang, Baoyu; Du, Jinghua; Li, Ya; Wang, Rongqi; Zhang, Yuguo; Zhao, Suxian; Sun, Dianxing; Qiao, Liang; Nan, Yuemin

    2016-08-01

    Long non-coding RNA (LncRNA)-activated by transforming growth factor-beta (LncRNA-ATB) is a key regulator of transforming growth factor-beta (TGF-β) signaling pathway, and is positively correlated with the development of liver cirrhosis and vascular invasion of hepatocellular carcinoma (HCC). However, the role of LncRNA-ATB in hepatitis C virus (HCV)-related liver fibrosis remains largely unknown. In the present study, we confirmed a high expression level of LncRNA-ATB in the liver tissues and plasma samples of patients with HCV-related hepatic fibrosis, and the plasma level of LncRNA-ATB was significantly correlated with liver fibrosis stages. Furthermore, increased expression level of LncRNA-ATB was also present in activated hepatic stellate cells (HSCs), and knockdown of LncRNA-ATB inhibited the expression of alpha-smooth muscle actin (α-SMA) and alpha-1 type I collagen (Col1A1). LncRNA-ATB was found to share the common miRNA responsive element of miR-425-5p with TGF-β type II receptor (TGF-βRII) and SMAD2. Ectopic expression of LncRNA-ATB in HSCs could upregulate the protein expression of TGF-βRII and SMAD2 by inhibiting the endogenous miR-425-5p. Moreover, overexpression of miR-425-5p could partly abrogate the expression of TGF-βRII and SMAD2 induced by LncRNA-ATB. Hence, we conclude that LncRNA-ATB promotes HCV-induced liver fibrogenesis by activating HSCs and increasing collagen I production through competitively binding to miR-425-5p. LncRNA-ATB may be a novel diagnostic biomarker and a potential therapeutic target for HCV-related hepatic fibrosis. PMID:27585228

  16. Notch induces myofibroblast differentiation of alveolar epithelial cells via transforming growth factor-{beta}-Smad3 pathway.

    PubMed

    Aoyagi-Ikeda, Kana; Maeno, Toshitaka; Matsui, Hiroki; Ueno, Manabu; Hara, Kenichiro; Aoki, Yasuhiro; Aoki, Fumiaki; Shimizu, Takehisa; Doi, Hiroshi; Kawai-Kowase, Keiko; Iso, Tatsuya; Suga, Tatsuo; Arai, Masashi; Kurabayashi, Masahiko

    2011-07-01

    Notch is an ancient cell-signaling system that regulates the specification of cell fate. This study examined the role of Notch in the epithelial-mesenchymal transition (EMT) and myofibroblast differentiation of cultured RLE-6TN cells (i.e., rat alveolar epithelial cells). The activation of Notch, either by ectopic expression of the Notch intracellular domain or by the co-culture of RLE-6TN cells with L-Jagged1 cells, induces the expression of smooth muscle α-actin (SMA) and other mesenchymal marker genes (collagen I and vimentin), and reduces the expression of epithelial marker genes (E-cadherin, occludin, and zonula occludens-1). The pharmacologic inhibition of the endogenous Notch signal significantly inhibited the transforming growth factor-β (TGF-β)-induced expression of SMA. Cell migratory capacity was increased by Notch. Luciferase assays revealed that the CC(A/T)(6)GG (CArG) box and the TGF-β control element (TCE) are required for Notch-induced SMA gene transcription. DNA microarray analysis revealed that members of the TGF-β family as well as Jagged1 were induced in RLE-6TN cells by Notch. Western blot analysis showed that Notch induced the phosphorylation of Smad3, and the TGF-β receptor type I/activin receptor-like kinase 5 (ALK5) kinase inhibitor SB431542 markedly reduced the Notch-induced expression of SMA. Enzyme-linked immunosorbent assays confirmed the production of TGF-β1 from RLE-6TN cells by Notch. Immunohistochemistry of a bleomycin-induced model of pulmonary fibrosis and lung specimens from patients with idiopathic interstitial pneumonias showed that Notch was strongly expressed in myofibroblasts, identified as SMA-positive cells. These data indicate that Notch induces myofibroblast differentiation through a TGF-β-Smad3 pathway that activates SMA gene transcription in a CArG-dependent and TCE-dependent manner in alveolar epithelial cells. Our data also imply that Notch induces the EMT phenotype, with increased migratory behavior in

  17. Transforming growth factor beta mRNA increases during liver regeneration: a possible paracrine mechanism of growth regulation.

    PubMed Central

    Braun, L; Mead, J E; Panzica, M; Mikumo, R; Bell, G I; Fausto, N

    1988-01-01

    Transforming growth factor beta (TGF-beta) is a growth factor with multiple biological properties including stimulation and inhibition of cell proliferation. To determine whether TGF-beta is involved in hepatocyte growth responses in vivo, we measured the levels of TGF-beta mRNA in normal liver and during liver regeneration after partial hepatectomy in rats. TGF-beta mRNA increases in the regenerating liver and reaches a peak (about 8 times higher than basal levels) after the major wave of hepatocyte cell division and mitosis have taken place and after the peak expression of the ras protooncogenes. Although hepatocytes from normal and regenerating liver respond to TGF-beta, they do not synthesize TGF-beta mRNA. Instead, the message is present in liver nonparenchymal cells and is particularly abundant in cell fractions enriched for endothelial cells. TGF-beta inhibits epidermal growth factor-induced DNA synthesis in vitro in hepatocytes from normal or regenerating liver, although the dose-response curves vary according to the culture medium used. We conclude that TGF-beta may function as the effector of an inhibitory paracrine loop that is activated during liver regeneration, perhaps to prevent uncontrolled hepatocyte proliferation. Images PMID:3422749

  18. Tumor-host interactions in the gallbladder suppress distal angiogenesis and tumor growth: involvement of transforming growth factor beta1.

    PubMed

    Gohongi, T; Fukumura, D; Boucher, Y; Yun, C O; Soff, G A; Compton, C; Todoroki, T; Jain, R K

    1999-10-01

    Angiogenesis inhibitors produced by a primary tumor can create a systemic anti-angiogenic environment and maintain metastatic tumor cells in a state of dormancy. We show here that the gallbladder microenvironment modulates the production of transforming growth factor (TGF)-beta1, a multifunctional cytokine that functions as an endogenous anti-angiogenic and anti-tumor factor in a cranial window preparation. We found that a wide variety of human gallbladder tumors express TGF-beta1 irrespective of histologic type. We implanted a gel impregnated with basic fibroblast growth factor or Mz-ChA-2 tumor in the cranial windows of mice without tumors or mice with subcutaneous or gallbladder tumors to study angiogenesis and tumor growth at a secondary site. Angiogenesis, leukocyte-endothelial interaction in vessels and tumor growth in the cranial window were substantially inhibited in mice with gallbladder tumors. The concentration of TGF-beta1 in the plasma of mice with gallbladder tumors was 300% higher than that in the plasma of mice without tumors or with subcutaneous tumors. In contrast, there was no difference in the plasma levels of other anti- and pro-angiogenic factors. Treatment with neutralizing antibody against TGF-beta1 reversed both angiogenesis suppression and inhibition of leukocyte rolling induced by gallbladder tumors. TGF-beta1 also inhibited Mz-ChA-2 tumor cell proliferation. Our results indicate that the production of anti-angiogenesis/proliferation factors is regulated by tumor-host interactions. PMID:10502827

  19. Epidermal growth factor (EGF) antagonizes transforming growth factor (TGF)-beta1-induced collagen lattice contraction by human skin fibroblasts.

    PubMed

    Park, J S; Kim, J Y; Cho, J Y; Kang, J S; Yu, Y H

    2000-12-01

    Wound contraction plays an important role in healing, but in extreme conditions, it may lead to excessive scar formation and pathological wound contracture. To date, the key regulator of excessive contracture is known to be transforming growth factor-beta (TGF-beta1). In this study, we have evaluated epidermal growth factor (EGF) antagonism in fibroblast-populated collagen lattice (FPCL) gel contraction, which has been generally used as an in vitro model thought to mimic wound contraction in vivo. As expected, TGF-beta1 treatment enhanced normal fibroblast-induced collagen gel contraction in a dose-dependent manner. In contrast, EGF did not affect normal gel formation, but significantly antagonized TGF-beta1-induced gel formation (p<0.05 at 100 ng/ml), whereas the other growth factor, platelet-derived growth factor (PDGF), did not altered either normal or TGF-beta1-induced gel contractions. Similarly, EGF treatment, but not PDGF, also significantly suppressed TGF-beta1 release that was autologously elicited by TGF-beta1 treatment (p<0.01 at 100 ng/ml). Therefore, the results suggest that EGF may negatively regulate the role of TGF-beta1 through attenuating autologous release of TGF-beta1. PMID:11145189

  20. Arabidopsis FIM5 decorates apical actin filaments and regulates their organization in the pollen tube

    PubMed Central

    Zhang, Meng; Zhang, Ruihui; Qu, Xiaolu; Huang, Shanjin

    2016-01-01

    The actin cytoskeleton is increasingly recognized as a major regulator of pollen tube growth. Actin filaments have distinct distribution patterns and dynamic properties within different regions of the pollen tube. Apical actin filaments are highly dynamic and crucial for pollen tube growth. However, how apical actin filaments are generated and properly constructed remains an open question. Here we showed that Arabidopsis fimbrin5 (FIM5) decorates filamentous structures throughout the entire tube but is apically concentrated. Apical actin structures are disorganized to different degrees in the pollen tubes of fim5 loss-of-function mutants. Further observations suggest that apical actin structures are not constructed properly because apical actin filaments cannot be maintained at the cortex of fim5 pollen tubes. Actin filaments appeared to be more curved in fim5 pollen tubes and this was confirmed by measurements showing that the convolutedness and the rate of change of convolutedness of actin filaments was significantly increased in fim5 pollen tubes. This suggests that the rigidity of the actin filaments may be compromised in fim5 pollen tubes. Further, the apical cell wall composition is altered, implying that tip-directed vesicle trafficking events are impaired in fim5 pollen tubes. Thus, we found that FIM5 decorates apical actin filaments and regulates their organization in order to drive polarized pollen tube growth. PMID:27117336

  1. Symmetry breaking in actin gels - Implications for cellular motility

    NASA Astrophysics Data System (ADS)

    John, Karin; Peyla, Philippe; Misbah, Chaouqi

    2007-03-01

    The physical origin of cell motility is not fully understood. Recently minimal model systems have shown, that polymerizing actin itself can produce a motile force, without the help of motor proteins. Pathogens like Shigella or Listeria use actin to propel themselves forward in their host cell. The same process can be mimicked with polystyrene beads covered with the activating protein ActA, which reside in a solution containing actin monomers. ActA induces the growth of an actin gel at the bead surface. Initially the gel grows symmetrically around the bead until a critical size is reached. Subsequently one observes a symmetry breaking and the gel starts to grow asymmetrically around the bead developing a tail of actin at one side. This symmetry breaking is accompanied by a directed movement of the bead, with the actin tail trailing behind the bead. Force generation relies on the combination of two properties: growth and elasticity of the actin gel. We study this phenomenon theoretically within the framework of a linear elasticity theory and linear flux-force relationships for the evolution of an elastic gel around a hard sphere. Conditions for a parity symmetry breaking are identified analytically and illustrated numerically with the help of a phasefield model.

  2. Expression of transforming growth factor-β1 in neonatal rats with hyperoxia-induced bronchopulmonary dysplasia and its relationship with lung development.

    PubMed

    Yan, B; Zhong, W; He, Q M; Zhang, S Y; Yu, J K; Pan, Y L

    2016-01-01

    The aim of this study was to detect the expression of transforming growth factor-ß1 (TGF-ß1) in neonatal rats with hyperoxia-induced bronchopulmonary dysplasia (BPD) and to explore its relationship with lung development. Forty-eight rats (2-3 days old) were randomly divided into a hyperoxia group and a control group (N = 24) which were then fed in ≥95% oxygen atmosphere and air, respectively. On the 1st, 3rd and 7th days of hyperoxia exposure, morphological changes of lung tissues were observed under an optical microscope. TGF-ß1 mRNA and protein levels in lung tissues were detected by real-time quantitative polymerase chain reaction and western blot, respectively. With increasing time of hyperoxia exposure, the hyperoxia group gradually suffered from pathological changes such as poor development of lung tissues, alveolar simplification, decrease in the number of alveoli, and hindered pulmonary microvascular development. On the 7th day of hyperoxia exposure, TGF-ß1 mRNA and protein levels (relative to b-actin) of the hyperoxia group (0.34 ± 0.19 and 0.21 ± 0.09, respectively) were significantly lower than those of the control group (0.83 ± 0.45 and 0.57 ± 0.45, respectively; P < 0.05). TGF-ß1 participates in the pathogenesis of BPD as an important regulatory factor during pulmonary vascular development. PMID:27173325

  3. Epigallocatechin-3-gallate attenuates transforming growth factor-β1 induced epithelial-mesenchymal transition via Nrf2 regulation in renal tubular epithelial cells.

    PubMed

    Wang, Yanqiu; Liu, Na; Su, Xuesong; Zhou, Guangyu; Sun, Guangping; Du, Feng; Bian, Xiaohui; Wang, Bowen

    2015-03-01

    Transforming growth factor-β1 (TGF-β1) induced epithelial-mesenchymal transition (EMT) plays an important role in renal fibrotic process regulation. Epigallocatechin-3-gallate (EGCG) exerts a protective effect against acute renal damage through its anti-oxidative effect by activating the Nrf2 signaling pathway. This study aims to investigate whether EGCG prevents TGF-β1 induced EMT and whether this effect acts via the Nrf2-mediated suppression of TGF-β1 signaling. MTT was used for cytotoxicity of EGCG examination and Western blotting and immunofluorescence were used for protein expression analysis. Results showed that EGCG prevented TGF-β1 mediated EMT and Smad 2 and Smad 3 phosphorylation in a dose dependent manner in NRK-52E cells. In addition, EGCG increased Nrf2 nuclear accumulation. Overexpression of Nrf2 blocked the phosphorylation of Smad 2 and Smad 3 mediated by TGF-β1 and decreased protein expression of plasminogen activator inhibitor 1 (PAI-1) and α-smooth muscle actin (α-SMA). Furthermore, siRNA-mediated knockdown of Nrf2 gene completely blocked the effects of EGCG, indicated by the reduced expressions of type I collagen (Col-I) and α-SMA were restored. In summary, EGCG inhibits TGF-β1 induced EMT and fibrotic proteins expression by Nrf2 activation. This study reveals a possible underlying mechanism of the renal protective effects of EGCG, and may provide a potential candidate to renal fibrosis therapy. PMID:25776510

  4. Involvement of epithelial-to-mesenchymal transition and associated transforming growth factor-β/Smad signaling in paraquat-induced pulmonary fibrosis.

    PubMed

    Han, Ying-Ying; Shen, Peng; Chang, Wen-Xiu

    2015-12-01

    Paraquat (PQ) is a highly toxic herbicide which is able to induce pulmonary fibrosis in humans and animals. The epithelial‑to‑mesenchymal transition (EMT) was demonstrated to be an important factor in pulmonary fibrosis. However, it has remained elusive whether PQ induces pulmonary fibrosis via EMT, which was therefore investigated in the present study. In addition, the underlying mechanisms of PQ‑induced EMT were examined in vitro. Hematoxylin and eosin staining of rat lung tissues demonstrated that PQ induced pulmonary fibrosis in vivo. Western blot analysis then revealed that the expression of epithelial cell marker E‑cadherin was significantly decreased, while the expression of mesenchymal markers α‑smooth‑muscle actin and vimentin was significantly increased in rat lung tissues and A549 cells following PQ treatment. Transforming growth factor (TGF)‑β/Smad signaling was also induced by PQ as evidenced by increased expression of TGF‑β1 and Smad2. However, PQ‑induced EMT in A549 cells was abolished by transfection with TGF‑β1‑specific small hairpin RNA. In conclusion, the present study demonstrated that PQ induced EMT in vivo and in vitro, which may be an important process in the development of PQ‑induced pulmonary fibrosis. In addition, TGF-β/Smad signaling was involved in PQ-induced EMT. PMID:26499763

  5. Knockdown of elF3a inhibits collagen synthesis in renal fibroblasts via Inhibition of transforming growth factor-β1/Smad signaling pathway.

    PubMed

    Zhang, Yun-Fang; Wang, Qi; Luo, Jie; Yang, Shen; Wang, Jie-Lin; Li, Hong-Yan

    2015-01-01

    Renal fibrosis is characterized by an exacerbated accumulation of deposition of the extracellular matrix (ECM). The eukaryotic translation initiation factor (eIF) 3a is the largest subunit of the eIF3 complex and has been involved in pulmonary fibrosis. However, the role of eIF3a in rental fibrosis is still unclear. Therefore, in this study, we investigated the role of eIF3a in rental fibrosis and explored the underlying mechanism. Our study found that eIF3a was up-regulated in renal fibrotic tissues and transforming growth factor (TGF)-β1-treated HK-2 cells. In addition, knockdown of eIF3a significantly inhibited TGF-β1-induced expression levels of α-smooth muscle actin (α-SMA) and collagen I. Furthermore, knockdown of eIF3a attenuated TGF-β1-induced Smad3 activation in HK-2 cells. Taken together, these results suggest that knockdown of eIF3a inhibits collagen synthesis in renal fibroblasts via inhibition of TGF-β1/Smad signaling pathway, and eIF3a may be a potential molecular target for the treatment of renal fibrosis. PMID:26464640

  6. Localization of type I procollagen gene expression in silica-induced granulomatous lung disease and implication of transforming growth factor-beta as a mediator of fibrosis.

    PubMed Central

    Mariani, T. J.; Roby, J. D.; Mecham, R. P.; Parks, W. C.; Crouch, E.; Pierce, R. A.

    1996-01-01

    We have used the silica-induced model of pulmonary injury in the rat to study the pattern of collagen expression in granulomatous lung inflammation. A single intratracheal instillation of silica into adult rats resulted in granulomatous inflammation leading to fibrosis and alveolar proteinosis. The development of disease in these animals was characterized over a 27-day period after treatment by means of histological, biochemical, and molecular analyses. Biochemical analyses indicated that significant increases in the weights of silicotic lungs were due to elevated amounts of DNA and total protein. Analysis of hydroxyproline content showed a 15-fold increase in this amino acid in silicotic lungs, confirming the development of a fibrotic reaction. In situ hybridization for type I procollagen mRNA displayed increased gene expression in the parenchyma, conducting airways, and vasculature of silicotic rats. Within the parenchyma, type I procollagen was expressed uniquely within granulomatous lesions. Immunohistochemistry indicated type I procollagen was being expressed by an alpha-smooth muscle actin-negative population of cells. Immunolocalization of extra-cellular transforming growth factor-beta showed coincident temporal and spatial overlap with type I procollagen expression, implicating this cytokine as a mediator of collagen gene expression in this model. Images Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 PMID:8546202

  7. Reconstitution of Actin-based Motility by Vasodilator-stimulated Phosphoprotein (VASP) Depends on the Recruitment of F-actin Seeds from the Solution Produced by Cofilin*

    PubMed Central

    Siton, Orit; Bernheim-Groswasser, Anne

    2014-01-01

    Vasodilator-stimulated phosphoprotein (VASP) is active in many filopodium-based and cytoskeleton reorganization processes. It is not fully understood how VASP directly functions in actin-based motility and how regulatory proteins affect its function. Here, we combine bead motility assay and single filament experiments. In the presence of a bundling component, actin bundles that grow from the surface of WT-VASP-coated beads induced movement of the beads. VASP promotes actin-based movement alone, in the absence of other actin nucleators. We propose that at physiological salt conditions VASP nucleation activity is too weak to promote motility and bundle formation. Rather, VASP recruits F-actin seeds from the solution and promotes their elongation. Cofilin has a crucial role in the nucleation of these F-actin seeds, notably under conditions of unfavorable spontaneous actin nucleation. We explored the role of multiple VASP variants. We found that the VASP-F-actin binding domain is required for the recruitment of F-actin seeds from the solution. We also found that the interaction of profilin-actin complexes with the VASP-proline-rich domain and the binding of the VASP-F-actin binding domain to the side of growing filaments is critical for transforming actin polymerization into motion. At the single filament level, profilin mediates both filament elongation rate and VASP anti-capping activity. Binding of profilin-actin complexes increases the polymerization efficiency by VASP but decreases its efficiency as an anti-capper; binding of free profilin creates the opposite effect. Finally, we found that an additional component such as methylcellulose or fascin is required for actin bundle formation and motility mediated by VASP. PMID:25246528

  8. Staining Fission Yeast Filamentous Actin with Fluorescent Phalloidin Conjugates.

    PubMed

    Hagan, Iain M

    2016-01-01

    The Schizosaccharomyces pombe filamentous (F)-actin cytoskeleton drives cell growth, morphogenesis, endocytosis, and cytokinesis. The protocol described here reveals the distribution of F-actin in fixed cells through the use of fluorescently conjugated phalloidin. Simultaneous staining of cell wall landmarks (with calcofluor) and chromatin (with 4',6-diamidino-2-phenylindole, or DAPI) makes this rapid staining procedure highly effective for staging cell cycle progression, monitoring morphogenetic abnormalities, and assessing the impact of environmental and genetic changes on the integrity of the F-actin cytoskeleton. PMID:27250943

  9. Transcriptional pathways associated with the slow growth phenotype of transformed Anaplasma marginale

    PubMed Central

    2013-01-01

    Background The ability to genetically manipulate bacteria has been fundamentally important for both basic biological discovery and translational research to develop new vaccines and antibiotics. Experimental alteration of the genetic content of prokaryotic pathogens has revealed both expected functional relationships and unexpected phenotypic consequences. Slow growth phenotypes have been reported for multiple transformed bacterial species, including extracellular and intracellular pathogens. Understanding the genes and pathways responsible for the slow growth phenotype provides the opportunity to develop attenuated vaccines as well as bacteriostatic antibiotics. Transformed Anaplasma marginale, a rickettsial pathogen, exhibits slow growth in vitro and in vivo as compared to the parent wild type strain, providing the opportunity to identify the underlying genes and pathways associated with this phenotype. Results Whole genome transcriptional profiling allowed for identification of specific genes and pathways altered in transformed A. marginale. Genes found immediately upstream and downstream of the insertion site, including a four gene operon encoding key outer membrane proteins, were not differentially transcribed between wild type and transformed A. marginale. This lack of significant difference in transcription of flanking genes and the large size of the insert relative to the genome were consistent with a trans rather than a cis effect. Transcriptional profiling across the complete genome identified the most differentially transcribed genes, including an iron transporter, an RNA cleaving enzyme and several genes involved in translation. In order to confirm the trend seen in translation-related genes, K-means clustering and Gene Set Enrichment Analysis (GSEA) were applied. These algorithms allowed evaluation of the behavior of genes as groups that share transcriptional status or biological function. Clustering and GSEA confirmed the initial observations and

  10. Shape and size transformation of gold nanorods (GNRs) via oxidation process: A reverse growth mechanism

    NASA Astrophysics Data System (ADS)

    Chandrasekar, Govindasamy; Mougin, Karine; Haidara, Hamidou; Vidal, Loïc; Gnecco, Enrico

    2011-02-01

    The anisotropic shape transformation of gold nanorods (GNRs) with H2O2 was observed in the presence of "cethyl trimethylammonium bromide" (CTAB). The adequate oxidative dissolution of GNR is provided by the following autocatalytic scheme with H2O2: Au0 → Au+, Au0 + Aun+ → 2Au3+, n = 1 and 3. The shape transformation of the GNRs was investigated by UV-vis spectroscopy and transmission electron microscopy (TEM). As-synthesised GNRs exhibit transverse plasmon band (TPB) at 523 nm and longitudinal plasmon band (LPB) at 731 nm. Upon H2O2 oxidation, the LPB showed a systematic hypsochromic (blue) shift, while TPB stays at ca. 523 nm. In addition, a new emerging peak observed at ca. 390 nm due to Au(III)-CTAB complex formation during the oxidation. TEM analysis of as-synthesised GNRs with H2O2 confirmed the shape transformation to spherical particles with 10 nm size in 2 h, whereas centrifuged nanorod solution showed no changes in the aspect ratio under the same condition. Au3+ ions produced from oxidation, complex with excess free CTAB and approach the nanorods preferentially at the end, leading to spatially directed oxidation. This work provides some information to the crystal stability and the growth mechanism of GNRs, as both growth and shortening reactions occur preferentially at the edge of single-crystalline GNRs, all directed by Br- ions.

  11. Hic-5 Regulates Actin Cytoskeletal Reorganization and Expression of Fibrogenic Markers and Myocilin in Trabecular Meshwork Cells

    PubMed Central

    Pattabiraman, Padmanabhan Paranji; Rao, Ponugoti Vasantha

    2015-01-01

    Purpose To explore the role of inducible focal adhesion (FA) protein Hic-5 in actin cytoskeletal reorganization, FA formation, fibrogenic activity, and expression of myocilin in trabecular meshwork (TM) cells. Methods Using primary cultures of human TM (HTM) cells, the effects of various external factors on Hic-5 protein levels, as well as the effects of recombinant Hic-5 and Hic-5 small interfering RNA (siRNA) on actin cytoskeleton, FAs, myocilin, α-smooth muscle actin (αSMA), and collagen-1 were determined by immunofluorescence and immunoblot analyses. Results Hic-5 distributes discretely to the FAs in HTM cells and throughout the TM and Schlemm's canal of the human aqueous humor (AH) outflow pathway. Transforming growth factor-β2 (TGF-β2), endothelin-1, lysophosphatidic acid, hydrogen peroxide, and RhoA significantly increased Hic-5 protein levels in HTM cells in association with reorganization of actin cytoskeleton and FAs. While recombinant Hic-5 induced actin stress fibers, FAs, αv integrin redistribution to the FAs, increased levels of αSMA, collagen-1, and myocilin, Hic-5 siRNA suppressed most of these responses in HTM cells. Hic-5 siRNA also suppressed TGF-β2-induced fibrogenic activity and dexamethasone-induced myocilin expression in HTM cells. Conclusions Taken together, these results reveal that Hic-5, whose levels were increased by various external factors implicated in elevated intraocular pressure, induces actin cytoskeletal reorganization, FAs, expression of fibrogenic markers, and myocilin in HTM cells. These characteristics of Hic-5 in TM cells indicate its importance in regulation of AH outflow through the TM in both normal and glaucomatous eyes. PMID:26313302

  12. Evidence That an Unconventional Actin Can Provide Essential F-Actin Function and That a Surveillance System Monitors F-Actin Integrity in Chlamydomonas.

    PubMed

    Onishi, Masayuki; Pringle, John R; Cross, Frederick R

    2016-03-01

    Actin is one of the most conserved eukaryotic proteins. It is thought to have multiple essential cellular roles and to function primarily or exclusively as filaments ("F-actin"). Chlamydomonas has been an enigma, because a null mutation (ida5-1) in its single gene for conventional actin does not affect growth. A highly divergent actin gene, NAP1, is upregulated in ida5-1 cells, but it has been unclear whether NAP1 can form filaments or provide actin function. Here, we used the actin-depolymerizing drug latrunculin B (LatB), the F-actin-specific probe Lifeact-Venus, and genetic and molecular methods to resolve these issues. LatB-treated wild-type cells continue to proliferate; they initially lose Lifeact-stained structures but recover them concomitant with upregulation of NAP1. Thirty-nine LatB-sensitive mutants fell into four genes (NAP1 and LAT1-LAT3) in which we identified the causative mutations using a novel combinatorial pool-sequencing strategy. LAT1-LAT3 are required for NAP1 upregulation upon LatB treatment, and ectopic expression of NAP1 largely rescues the LatB sensitivity of the lat1-lat3 mutants, suggesting that the LAT gene products comprise a regulatory hierarchy with NAP1 expression as the major functional output. Selection of LatB-resistant revertants of a nap1 mutant yielded dominant IDA5 mutations that presumably render F-IDA5 resistant to LatB, and nap1 and lat mutations are synthetically lethal with ida5-1 in the absence of LatB. We conclude that both IDA5 and the divergent NAP1 can form filaments and redundantly provide essential F-actin functions and that a novel surveillance system, probably responding to a loss of F-actin, triggers NAP1 expression and perhaps other compensatory responses. PMID:26715672

  13. Force Generation, Polymerization Dynamics and Nucleation of Actin Filaments

    NASA Astrophysics Data System (ADS)

    Wang, Ruizhe

    We study force generation and actin filament dynamics using stochastic and deterministic methods. First, we treat force generation of bundled actin filaments by polymerization via molecular-level stochastic simulations. In the widely-used Brownian Ratchet model, actin filaments grow freely whenever the tip-obstacle gap created by thermal fluctuation exceeds the monomer size. We name this model the Perfect Brownian Ratchet (PBR) model. In the PBR model, actin monomer diffusion is treated implicitly. We perform a series of simulations based on the PBR, in which obstacle motion is treated explicitly; in most previous studies, obstacle motion has been treated implicitly. We find that the cooperativity of filaments is generally weak in the PBR model, meaning that more filaments would grow more slowly given the same force per filament. Closed-form formulas are also developed, which match the simulation results. These portable and accurate formulas provide guidance for experiments and upper and lower bounds for theoretical analyses. We also studied a variation of the PBR, called the Diffusing Brownian Ratchet (DBR) model, in which both actin monomer and obstacle diffusion are treated explicitly. We find that the growth rate of multiple filaments is even lower, compared with that in PBR. This finding challenges the widely-accepted PBR assumption and suggests that pushing the study of actin dynamics down to the sub-nanometer level yields new insights. We subsequently used a rate equation approach to model the effect of local depletion of actin monomers on the nucleation of actin filaments on biomimetic beads, and how the effect is regulated by capping protein (CP). We find that near the bead surface, a higher CP concentration increases local actin concentration, which leads to an enhanced activities of actin filaments' nucleation. Our model analysis matches the experimental results and lends support to an important but undervalued hypothesis proposed by Carlier and

  14. Mammalian dwarfins are phosphorylated in response to transforming growth factor beta and are implicated in control of cell growth.

    PubMed Central

    Yingling, J M; Das, P; Savage, C; Zhang, M; Padgett, R W; Wang, X F

    1996-01-01

    The dwarfin protein family has been genetically implicated in transforming growth factor beta (TGF-beta)-like signaling pathways in Drosophila and Caenorhabditis elegans. To investigate the role of these proteins in mammalian signaling pathways, we have isolated and studied two murine dwarfins, dwarfin-A and dwarfin-C. Using antibodies against dwarfin-A and dwarfin-C, we show that these two dwarfins and an immunogenically related protein, presumably also a dwarfin, are phosphorylated in a time- and dose-dependent manner in response to TGF-beta. Bone morphogenetic protein 2, a TGF-beta superfamily ligand, induces phosphorylation of only the related dwarfin protein. Thus, TGF-beta superfamily members may use overlapping yet distinct dwarfins to mediate their intracellular signals. Furthermore, transient overexpression of either dwarfin-A or dwarfin-C causes growth arrest, implicating the dwarfins in growth regulation. This work provides strong biochemical and preliminary functional evidence that dwarfin-A and dwarfin-C represent prototypic members of a family of mammalian proteins that may serve as mediators of signaling pathways for TGF-beta superfamily members. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 PMID:8799132

  15. Expression of transforming growth factor alpha and epidermal growth factor receptor in rat lung neoplasms induced by plutonium-239

    SciTech Connect

    Stegelmeier, B.L.; Gillett, N.A.; Hahn, F.F.; Kelly, G.; Rebar, A.H.

    1994-11-01

    Ninety-two rat lung proliferative lesions and neoplasms induced by inhaled {sup 239}PuO{sub 2} were evaluated for aberrant expression of transforming growth factor alpha (TGF-{alpha}) and epidermal growth factor receptor (EGFR). Expression of TGF-{alpha} protein, measured by immunohistochemistry, was higher in 94% of the squamous cell carcinomas and 87% of the foci of alveolar epithelial squamous metaplasia than that exhibited by the normal-appearing, adjacent lung parenchyma. In contrast, only 20% of adenocarcinomas and foci of epithelial hyperplasia expressed elevated levels of TGF-{alpha}. Many neoplasms expressing TGF-{alpha} also expressed excessive levels of EGFR mRNA. Southern and DNA slot blot analyses showed that the elevated EGFR expression was not due to amplification of the EGFR gene. These data suggest that increased amounts of TGF-{alpha} were early alterations in the progression of plutonium-induced squamous cell carcinoma, and these increases may occur in parallel with overexpression of the receptor for this growth factor. Together, these alterations create a potential autocrine loop for sustaining clonal expansion of cells initiated by high-LET radiation. 44 refs., 4 figs., 1 tab.

  16. Mechanism of Actin Network Attachment to Moving Membranes

    PubMed Central

    Co, Carl; Wong, Derek T.; Gierke, Sarah; Chang, Vicky; Taunton, Jack

    2007-01-01

    Summary Actin filament networks exert protrusive and attachment forces on membranes and thereby drive membrane deformation and movement. Here, we show that N-WASP WH2 domains play a previously unanticipated role in vesicle movement by transiently attaching actin filament barbed ends to the membrane. To dissect the attachment mechanism, we reconstituted the propulsive motility of lipid-coated glass beads using purified soluble proteins. N-WASP WH2 mutants assembled actin comet tails and initiated movement, but the comet tails catastrophically detached from the membrane. When presented on the surface of a lipid-coated bead, WH2 domains were sufficient to maintain comet tail attachment. In v-Src-transformed fibroblasts, N-WASP WH2 mutants were severely defective in the formation of circular podosome arrays. In addition to creating an attachment force, interactions between WH2 domains and barbed ends may locally amplify signals for dendritic actin nucleation. PMID:17350575

  17. Role of actin in auxin transport and transduction of gravity

    NASA Astrophysics Data System (ADS)

    Hu, S.; Basu, S.; Brady, S.; Muday, G.

    Transport of the plant hormone auxin is polar and the direction of the hormone movement appears to be controlled by asymmetric distribution of auxin transport protein complexes. Changes in the direction of auxin transport are believed to drive asymmetric growth in response to changes in the gravity vector. To test the possibility that asymmetric distribution of the auxin transport protein complex is mediated by attachment to the actin cytoskeleton, a variety of experimental approaches have been used. The most direct demonstration of the role of the actin cytoskeleton in localization of the protein complex is the ability of one protein in this complex to bind to affinity columns containing actin filaments. Additionally, treatments of plant tissues with drugs that fragment the actin c toskeleton reducey polar transport. In order to explore this actin interaction and the affect of gravity on auxin transport and developmental polarity, embryos of the brown alga, Fucus have been examined. Fucus zygotes are initially symmetrical, but develop asymmetry in response to environmental gradients, with light gradients being the best- characterized signal. Gravity will polarize these embryos and gravity-induced polarity is randomized by clinorotation. Auxin transport also appears necessary for environmental controls of polarity, since auxin efflux inhibitors perturb both photo- and gravity-polarization at a very discrete temporal window within six hours after fertilization. The actin cytoskeleton has previously been shown to reorganize after fertilization of Fucus embryos leading to formation of an actin patch at the site of polar outgrowth. These actin patches still form in Fucus embryos treated with auxin efflux inhibitors, yet the position of these patches is randomized. Together, these results suggest that there are connections between the actin cytoskeleton, auxin transport, and gravity oriented growth and development. (Supported by NASA Grant: NAG2-1203)

  18. Differentiation of Bone Marrow Mesenchymal Stem Cells to Cardiomyocyte-Like Cells Is Regulated by the Combined Low Dose Treatment of Transforming Growth Factor-β1 and 5-Azacytidine.

    PubMed

    Shi, Shutian; Wu, Xingxin; Wang, Xiao; Hao, Wen; Miao, Huangtai; Zhen, Lei; Nie, Shaoping

    2016-01-01

    Bone marrow mesenchymal stem cells (BMMSCs) are used in cardiac tissue engineering for the regeneration of diseased hearts. We examined the differentiation of rat BMMSCs into cardiomyocyte-like cells when induced with a combined low dose treatment of transforming growth factor-β1 (TGF-β1) and 5-azacytidine (5-AZA). Results showed that cell proliferation in the combined low dose treatment group of TGF-β1 and 5-AZA was increased compared with the TGF-β1 group or the 5-AZA group. The cell apoptosis was relieved by combined TGF-β1 and 5-AZA treatment compared to 5-AZA treatment alone. The number of cells positive for myosin heavy chain, connexin-43, α-actin, and troponin I in the combined treatment group was higher than those observed in the TGF-β1 group or the 5-AZA group. Moreover, the combined low dose treatment group of TGF-β1 and 5-AZA reveals the strongest expression of troponin I, α-actin, and phosphorylated extracellular signal-regulated protein kinases 1 and 2 (p-ErK1/2) among the treatment groups. These results suggest that the combined low dose treatment of TGF-β1 and 5-AZA can improve the differentiation potential of rat BMMSCs into cardiomyocyte-like cells and alleviate cell damage effects in vitro. The mechanism that is involved in influencing differentiation may be associated with p-ErK1/2. PMID:26697074

  19. Differentiation of Bone Marrow Mesenchymal Stem Cells to Cardiomyocyte-Like Cells Is Regulated by the Combined Low Dose Treatment of Transforming Growth Factor-β1 and 5-Azacytidine

    PubMed Central

    Shi, Shutian; Wu, Xingxin; Wang, Xiao; Hao, Wen; Miao, Huangtai; Zhen, Lei; Nie, Shaoping

    2016-01-01

    Bone marrow mesenchymal stem cells (BMMSCs) are used in cardiac tissue engineering for the regeneration of diseased hearts. We examined the differentiation of rat BMMSCs into cardiomyocyte-like cells when induced with a combined low dose treatment of transforming growth factor-β1 (TGF-β1) and 5-azacytidine (5-AZA). Results showed that cell proliferation in the combined low dose treatment group of TGF-β1 and 5-AZA was increased compared with the TGF-β1 group or the 5-AZA group. The cell apoptosis was relieved by combined TGF-β1 and 5-AZA treatment compared to 5-AZA treatment alone. The number of cells positive for myosin heavy chain, connexin-43, α-actin, and troponin I in the combined treatment group was higher than those observed in the TGF-β1 group or the 5-AZA group. Moreover, the combined low dose treatment group of TGF-β1 and 5-AZA reveals the strongest expression of troponin I, α-actin, and phosphorylated extracellular signal-regulated protein kinases 1 and 2 (p-ErK1/2) among the treatment groups. These results suggest that the combined low dose treatment of TGF-β1 and 5-AZA can improve the differentiation potential of rat BMMSCs into cardiomyocyte-like cells and alleviate cell damage effects in vitro. The mechanism that is involved in influencing differentiation may be associated with p-ErK1/2. PMID:26697074

  20. Capping of the barbed ends of actin filaments by a high-affinity profilin-actin complex.

    PubMed

    DiNubile, M J; Huang, S

    1997-01-01

    . Dissociation of the tight profilactin complex may serve as a unique mechanism by which profilin helps regulate actin filament growth. PMID:9227851

  1. Transforming growth factor-β in breast cancer: too much, too late

    PubMed Central

    Barcellos-Hoff, Mary Helen; Akhurst, Rosemary J

    2009-01-01

    The contribution of transforming growth factor (TGF)β to breast cancer has been studied from a myriad perspectives since seminal studies more than two decades ago. Although the action of TGFβ as a canonical tumor suppressor in breast is without a doubt, there is compelling evidence that TGFβ is frequently subverted in a malignant plexus that drives breast cancer. New knowledge that TGFβ regulates the DNA damage response, which underlies cancer therapy, reveals another facet of TGFβ biology that impedes cancer control. Too much TGFβ, too late in cancer progression is the fundamental motivation for pharmaceutical inhibition. PMID:19291273

  2. Influence of phase transformation on stress evolution during growth of metal thin films on silicon.

    PubMed

    Fillon, A; Abadias, G; Michel, A; Jaouen, C; Villechaise, P

    2010-03-01

    In situ stress measurements during two-dimensional growth of low mobility metal films on amorphous Si were used to demonstrate the impact of interface reactivity and phase transformation on stress evolution. Using Mo1-xSix films as examples, the results show that the tensile stress rise, which develops after the film has become crystalline, is correlated with an increase in lateral grain size. The origin of the tensile stress is attributed to the volume change resulting from the alloy crystallization, which occurs at a concentration-dependent critical thickness. PMID:20366996

  3. A novel transforming growth factor beta2 antisense transcript in mammalian lung.

    PubMed Central

    Coker, R K; Laurent, G J; Dabbagh, K; Dawson, J; McAnulty, R J

    1998-01-01

    Transforming growth factor (TGF) beta2 gene expression was examined in murine, rat and human lung by in situ hybridization with riboprobes. Hybridization signal was observed in a variety of cells with the sense probe, and Northern-blot analysis with this probe demonstrated the presence of a novel 3.5 kb transcript. This first report suggesting the existance of a natural TGFbeta2 antisense transcript raises the possibility that such a transcript may play a role in regulating TGFbeta2 production. PMID:9601055

  4. Phase transformations during the growth of paracetamol crystals from the vapor phase

    NASA Astrophysics Data System (ADS)

    Belyaev, A. P.; Rubets, V. P.; Antipov, V. V.; Bordei, N. S.

    2014-07-01

    Phase transformations during the growth of paracetamol crystals from the vapor phase are studied by differential scanning calorimetry. It is found that the vapor-crystal phase transition is actually a superposition of two phase transitions: a first-order phase transition with variable density and a second-order phase transition with variable ordering. The latter, being a diffuse phase transition, results in the formation of a new, "pretransition," phase irreversibly spent in the course of the transition, which ends in the appearance of orthorhombic crystals. X-ray diffraction data and micrograph are presented.

  5. Phase and structural transformations in annealed copper coatings in relation to oxide whisker growth

    NASA Astrophysics Data System (ADS)

    Dorogov, M. V.; Priezzheva, A. N.; Vlassov, S.; Kink, I.; Shulga, E.; Dorogin, L. M.; Lõhmus, R.; Tyurkov, M. N.; Vikarchuk, A. A.; Romanov, A. E.

    2015-08-01

    We describe structural and phase transformation in copper coatings made of microparticles during heating and annealing in air in the temperature range up to 400 °C. Such thermal treatment is accompanied by intensive CuO nanowhisker growth on the coating surface and the formation of the layered oxide phases (Cu2O and CuO) in the coating interior. X-ray diffraction and focused ion beam (FIB) are employed to characterize the multilayer structure of annealed copper coatings. Formation of volumetric defects such as voids and cracks in the coating is demonstrated.

  6. Rho/Rock cross-talks with transforming growth factor-β/Smad pathway participates in lung fibroblast-myofibroblast differentiation.

    PubMed

    Ji, Hong; Tang, Haiying; Lin, Hongli; Mao, Jingwei; Gao, Lili; Liu, Jia; Wu, Taihua

    2014-11-01

    The differentiation of fibroblasts, which are promoted by transforming growth factor-β (TGF-β)/Smad, is involved in the process of pulmonary fibrosis. The Rho/Rho-associated coiled-coil-forming protein kinase (Rock) pathway may regulate the fibroblast differentiation and myofibroblast expression of α-smooth muscle actin (α-SMA), however, the mechanism is not clear. The aim of the present study was to evaluate the role of Rho/Rock and TGF-β/Smad in TGF-β1-induced lung fibroblasts differentiation. Human embryonic lung fibroblasts were stimulated by TGF-β1, Y-27632 (inhibitor of Rho/Rock signaling) and staurosporine (inhibitor of TGF-β/Smad signaling). The α-SMA expression, cell cycle progression, content of the extracellular matrix (ECM) in cell culture supernatants and the expression of RhoA, RhoC, Rock1 and Smad2 were detected. The results demonstrated that α-SMA-positive cells significantly increased following TGF-β1 stimulation. Rho/Rock and TGF-β/Smad inhibitors suppressed TGF-β1-induced lung fibroblast differentiation. The inhibitors increased G0/G1 and decreased S and G2/M percentages. The concentrations of the ECM proteins in the supernatant were significantly increased by TGF-β1 stimulation, whereas they were decreased by inhibitor stimulation. RhoA, RhoC, Rock1, Smad2 and tissue inhibitor of metalloproteinase-1 were upregulated by TGF-β1 stimulation. The Rho/Rock inhibitor downregulated Smad2 expression and the TGF-β/Smad inhibitor downregulated RhoA, RhoC and Rock1 expression. Therefore, the Rho/Rock pathway and Smad signaling were involved in the process of lung fibroblasts transformation, induced by TGF-β1, to myofibroblasts. The two pathways may undergo cross-talk in the lung fibroblasts differentiation in vitro. PMID:25279146

  7. Transforming Growth Factor-β3 Therapy Delays Postoperative Reossification and Improves Craniofacial Growth in Craniosynostotic Rabbits.

    PubMed

    Gilbert, J; Karski, M; Smith, T D; Burrows, A M; Norbutt, C; Siegel, M I; Costello, B J; Cray, J J; Losee, J E; Moursi, A M; Cooper, G M; Mooney, M P

    2016-03-01

    Postoperative reossification is a common clinical correlate following surgery. It has been suggested that an underexpression of transforming growth factor-β3 (TGF-β3) may be related to craniosynostosis and postoperative reossification. Adding TGF-β3 may delay reossification and improve postoperative growth. The present study was designed to test this hypothesis. Thirty 10-day-old New Zealand white rabbits with hereditary coronal suture synostosis were divided into three groups: (1) suturectomy controls (n = 14), (2) suturectomy treated with bovine serum albumin (n = 8), and (3) suturectomy treated with TGF-β3 protein (n = 8). At 10 days of age, a 3-mm × 15-mm coronal suturectomy was performed, and serial three-dimensional (3D) computed tomography (CT) scans and cephalographs were taken at 10, 25, 42, and 84 days of age. Calvaria were harvested at 84 days of age for histomorphometric analysis. Mean differences were analyzed using a group by age analysis of variance. Analysis of the 3D CT scan data revealed that sites treated with TGF-β3 had significantly (P < .05) greater defect areas and significantly (P < .05) greater intracranial volumes through 84 days of age compared with controls. Histomorphometry showed that sites treated with TGF-β3 had patent suturectomy sites and significantly (P < .001) less new bone in the suturectomy site compared with controls. Serial radiograph data revealed significant (P < .05) differences in craniofacial growth from 25 to 84 days in TGF-β3-treated rabbits compared with controls. Data show that TGF-β3 administration delayed reossification and improved craniofacial growth in this rabbit model. These findings also suggest that this molecular-based therapy may have potential clinical use. PMID:26090789

  8. Harnessing high density lipoproteins to block transforming growth factor beta and to inhibit the growth of liver tumor metastases.

    PubMed

    Medina-Echeverz, José; Fioravanti, Jessica; Díaz-Valdés, Nancy; Frank, Kathrin; Aranda, Fernando; Gomar, Celia; Ardaiz, Nuria; Dotor, Javier; Umansky, Viktor; Prieto, Jesús; Berraondo, Pedro

    2014-01-01

    Transforming growth factor β (TGF-β) is a powerful promoter of cancer progression and a key target for antitumor therapy. As cancer cells exhibit active cholesterol metabolism, high density lipoproteins (HDLs) appear as an attractive delivery system for anticancer TGFβ-inhibitory molecules. We constructed a plasmid encoding a potent TGF-β-blocking peptide (P144) linked to apolipoprotein A-I (ApoA-I) through a flexible linker (pApoLinkerP144). The ApoLinkerP144 sequence was then incorporated into a hepatotropic adeno-associated vector (AAVApoLinkerP144). The aim was to induce hepatocytes to produce HDLs containing a modified ApoA-I capable of blocking TGF-β. We observed that transduction of the murine liver with pApoLinkerP144 led to the appearance of a fraction of circulating HDL containing the fusion protein. These HDLs were able to attenuate TGF-β signaling in the liver and to enhance IL-12 -mediated IFN-γ production. Treatment of liver metastasis of MC38 colorectal cancer with AAVApoLinkerP144 resulted in a significant reduction of tumor growth and enhanced expression of IFN-γ and GM-CSF in cancerous tissue. ApoLinkerP144 also delayed MC38 liver metastasis in Rag2-/-IL2rγ-/- immunodeficient mice. This effect was associated with downregulation of TGF-β target genes essential for metastatic niche conditioning. Finally, in a subset of ret transgenic mice, a model of aggressive spontaneous metastatic melanoma, AAVApoLinkerP144 delayed tumor growth in association with increased CD8+ T cell numbers in regional lymph nodes. In conclusion, modification of HDLs to transport TGF-β-blocking molecules is a novel and promising approach to inhibit the growth of liver metastases by immunological and non-immunological mechanisms. PMID:24797128

  9. Harnessing High Density Lipoproteins to Block Transforming Growth Factor Beta and to Inhibit the Growth of Liver Tumor Metastases

    PubMed Central

    Medina-Echeverz, José; Fioravanti, Jessica; Díaz-Valdés, Nancy; Frank, Kathrin; Aranda, Fernando; Gomar, Celia; Ardaiz, Nuria; Dotor, Javier; Umansky, Viktor; Prieto, Jesús; Berraondo, Pedro

    2014-01-01

    Transforming growth factor β (TGF-β) is a powerful promoter of cancer progression and a key target for antitumor therapy. As cancer cells exhibit active cholesterol metabolism, high density lipoproteins (HDLs) appear as an attractive delivery system for anticancer TGFβ-inhibitory molecules. We constructed a plasmid encoding a potent TGF-β-blocking peptide (P144) linked to apolipoprotein A-I (ApoA-I) through a flexible linker (pApoLinkerP144). The ApoLinkerP144 sequence was then incorporated into a hepatotropic adeno-associated vector (AAVApoLinkerP144). The aim was to induce hepatocytes to produce HDLs containing a modified ApoA-I capable of blocking TGF-β. We observed that transduction of the murine liver with pApoLinkerP144 led to the appearance of a fraction of circulating HDL containing the fusion protein. These HDLs were able to attenuate TGF-β signaling in the liver and to enhance IL-12 -mediated IFN-γ production. Treatment of liver metastasis of MC38 colorectal cancer with AAVApoLinkerP144 resulted in a significant reduction of tumor growth and enhanced expression of IFN-γ and GM-CSF in cancerous tissue. ApoLinkerP144 also delayed MC38 liver metastasis in Rag2−/−IL2rγ−/− immunodeficient mice. This effect was associated with downregulation of TGF-β target genes essential for metastatic niche conditioning. Finally, in a subset of ret transgenic mice, a model of aggressive spontaneous metastatic melanoma, AAVApoLinkerP144 delayed tumor growth in association with increased CD8+ T cell numbers in regional lymph nodes. In conclusion, modification of HDLs to transport TGF-β-blocking molecules is a novel and promising approach to inhibit the growth of liver metastases by immunological and non-immunological mechanisms. PMID:24797128

  10. The actin cytoskeleton is a suppressor of the endogenous skewing behaviour of Arabidopsis primary roots in microgravity.

    PubMed

    Nakashima, J; Liao, F; Sparks, J A; Tang, Y; Blancaflor, E B

    2014-01-01

    Before plants can be effectively utilised as a component of enclosed life-support systems for space exploration, it is important to understand the molecular mechanisms by which they develop in microgravity. Using the Biological Research in Canisters (BRIC) hardware on board the second to the last flight of the Space Shuttle Discovery (STS-131 mission), we studied how microgravity impacts root growth in Arabidopsis thaliana. Ground-based studies showed that the actin cytoskeleton negatively regulates root gravity responses on Earth, leading us to hypothesise that actin might also be an important modulator of root growth behaviour in space. We investigated how microgravity impacted root growth of wild type (ecotype Columbia) and a mutant (act2-3) disrupted in a root-expressed vegetative actin isoform (ACTIN2). Roots of etiolated wild-type and act2-3 seedlings grown in space skewed vigorously toward the left, which was unexpected given the reduced directional cue provided by gravity. The left-handed directional root growth in space was more pronounced in act2-3 mutants than wild type. To quantify differences in root orientation of these two genotypes in space, we developed an algorithm where single root images were converted into binary images using computational edge detection methods. Binary images were processed with Fast Fourier Transformation (FFT), and histogram and entropy were used to determine spectral distribution, such that high entropy values corresponded to roots that deviated more strongly from linear orientation whereas low entropy values represented straight roots. We found that act2-3 roots had a statistically stronger skewing/coiling response than wild-type roots, but such differences were not apparent on Earth. Ultrastructural studies revealed that newly developed cell walls of space-grown act2-3 roots were more severely disrupted compared to space-grown wild type, and ground control wild-type and act2-3 roots. Collectively, our results provide

  11. [Transforming growth factor beta-1: structure, function, and regulation mechanisms in cancer].

    PubMed

    Peralta-Zaragoza, O; Lagunas-Martínez, A; Madrid-Marina, V

    2001-01-01

    Transforming growth factor beta-1 (TGF-beta 1) is produced by several cell lineages such as lymphocytes, macrophages, and dendritic cells, and its expression serves in both autocrine and paracrine modes to control the differentiation, proliferation, and state of activation of these and other cells. In general, TGF-beta 1 has pleiotropic properties on the immune response during the development of infection diseases and cancer; however, the mechanisms of action and regulation of gene expression of this cytokine are poorly understood, in this review, the biological properties and the molecular mechanisms that regulate TGF-beta 1 gene expression are described, to understand the role of this cytokine in growth and cell differentiation. The knowledge of molecular mechanisms of gene expression of TGF-beta 1 may serve to develop new cancer therapies. The English version of this paper is available at: http://www.insp.mx/salud/index.html PMID:11547595

  12. Enhanced jun gene expression is an early genomic response to transforming growth factor beta stimulation.

    PubMed Central

    Pertovaara, L; Sistonen, L; Bos, T J; Vogt, P K; Keski-Oja, J; Alitalo, K

    1989-01-01

    Transforming growth factor beta (TGF beta) is a multifunctional polypeptide that regulates proliferation, differentiation, and other functions of many cell types. The pathway of TGF beta signal transduction in cells is unknown. We report here that an early effect of TGF beta is an enhancement of the expression of two genes encoding serum- and phorbol ester tumor promoter-regulated transcription factors: the junB gene and the c-jun proto-oncogene, respectively. This stimulation was observed in human lung adenocarcinoma A549 cells which were growth inhibited by TGF beta, AKR-2B mouse embryo fibroblasts which were growth stimulated by TGF beta, and K562 human erythroleukemia cells, which were not appreciably affected in their growth by TGF beta. The increase in jun mRNA occurred with picomolar TGF beta concentrations within 1 h of TGF beta stimulation, reached a peak between 1 and 5 h in different cells, and declined gradually to base-line levels. This mRNA response was followed by a large increase in the biosynthesis of the c-jun protein (AP-1), as shown by metabolic labeling and immunoprecipitation analysis. However, differential and cell type-specific regulation appeared to determine the timing and magnitude of the response of each jun gene in a given cell. In AKR-2B and NIH 3T3 cells, only junB was induced by TGF beta, evidently in a protein synthesis-independent fashion. The junB response to TGF beta was maintained in c-Ha-ras and neu oncogene-transformed cells. Thus, one of the earliest genomic responses to TGF beta may involve nuclear signal transduction and amplification by the junB and c-jun transcription factors in concert with c-fos, which is also induced. The differential activation of the jun genes may explain some of the pleiotropic effects of TGF beta. Images PMID:2725496

  13. Steric Effects Induce Geometric Remodeling of Actin Bundles in Filopodia.

    PubMed

    Dobramysl, Ulrich; Papoian, Garegin A; Erban, Radek

    2016-05-10

    Filopodia are ubiquitous fingerlike protrusions, spawned by many eukaryotic cells, to probe and interact with their environments. Polymerization dynamics of actin filaments, comprising the structural core of filopodia, largely determine their instantaneous lengths and overall lifetimes. The polymerization reactions at the filopodial tip require transport of G-actin, which enter the filopodial tube from the filopodial base and diffuse toward the filament barbed ends near the tip. Actin filaments are mechanically coupled into a tight bundle by cross-linker proteins. Interestingly, many of these proteins are relatively short, restricting the free diffusion of cytosolic G-actin throughout the bundle and, in particular, its penetration into the bundle core. To investigate the effect of steric restrictions on G-actin diffusion by the porous structure of filopodial actin filament bundle, we used a particle-based stochastic simulation approach. We discovered that excluded volume interactions result in partial and then full collapse of central filaments in the bundle, leading to a hollowed-out structure. The latter may further collapse radially due to the activity of cross-linking proteins, hence producing conical-shaped filament bundles. Interestingly, electron microscopy experiments on mature filopodia indeed frequently reveal actin bundles that are narrow at the tip and wider at the base. Overall, our work demonstrates that excluded volume effects in the context of reaction-diffusion processes in porous networks may lead to unexpected geometric growth patterns and complicated, history-dependent dynamics of intermediate metastable configurations. PMID:27166814

  14. The yin-yang of dendrite morphology: unity of actin and microtubules.

    PubMed

    Georges, Penelope C; Hadzimichalis, Norell M; Sweet, Eric S; Firestein, Bonnie L

    2008-12-01

    Actin and microtubules (MT) are targets of numerous molecular pathways that control neurite outgrowth. To generate a neuronal protrusion, coordinated structural changes of the actin and MT cytoskeletons must occur. Neurite formation occurs when actin filaments (F-actin) are destabilized, filopodia are extended, and MTs invade filopodia. This process results in either axon or dendrite formation. Axonal branching involves interplay between F-actin and MTs, with F-actin and MTs influencing polymerization, stabilization, and maintenance of each other. Our knowledge of the mechanisms regulating development of the axon, however, far eclipses our understanding of dendritic development and branching. The two classes of neurites, while fundamentally similar in their ability to elongate and branch, dramatically differ in growth rate, orientation of polarized MT bundles, and mechanisms that initiate branching. In this review, we focus on how F-actin, MTs, and proteins that link the two cytoskeletons coordinate to specifically initiate dendritic events. PMID:18987787

  15. Estimating canopy leaf area index in the late stages of wheat growth using continuous wavelet transform

    NASA Astrophysics Data System (ADS)

    Huang, Yan; Tian, Qingjiu; Wang, Lei; Geng, Jun; Lyu, Chunguang

    2014-01-01

    The existing hyperspectral vegetation indices used for estimating the canopy leaf area index (LAI) of winter wheat (Triticum aestivum L.) performed well, but the use of such indices at late growth stages can lead to inaccurate results. To improve the performance of LAI models for wheat in late growth stages, the continuous wavelet transform (CWT) method was applied in this study and used to decompose the canopy reflectance and its first derivative into wavelet coefficients. The correlation scalograms of wavelet coefficients and the LAI were then constructed and used to extract the top 1% correlated region as the wavelet feature. The canopy LAI estimation model for late growth wheat was established at last and compared with models based on 12 different types of hyperspectral vegetation indices. The results showed that, compared with the estimation models using the hyperspectral vegetation indices (for which the R2 values were all less than 0.15 and the root-mean-square errors (RMSEs) were greater than 1), the CWT-based canopy LAI estimation model for late growth wheat had obvious improvements in accuracy (maximum R2 of 0.53 and minimum of RMSE of 0.78). Hence, this new method shows promise for use in agricultural and ecological applications.

  16. Biochemical characterization of the Drosophila dpp protein, a member of the transforming growth factor beta family of growth factors.

    PubMed Central

    Panganiban, G E; Rashka, K E; Neitzel, M D; Hoffmann, F M

    1990-01-01

    The decapentaplegic (dpp) gene of Drosophila melanogaster is required for pattern formation in the embryo and for viability of the epithelial cells in the imaginal disks. The dpp protein product predicted from the DNA sequence is similar to members of a family of growth factors that includes transforming growth factor beta (TGF-beta). We have produced polyclonal antibodies to a recombinant dpp protein made in bacteria and used a metallothionein promoter to express a dpp cDNA in Drosophila S2 cells. Similar to other proteins in the TGF-beta family, the dpp protein produced by the Drosophila cells was proteolytically cleaved, and both portions of the protein were secreted from the cells. The amino-terminal 47-kilodalton (kDa) peptide was found in the medium and in the proteins adhering to the plastic petri dish. The carboxy-terminal peptide, the region with sequence similarity to the active ligand portion of TGF-beta, was found extracellularly as a 30-kDa homodimer. Most of the 30-kDa homodimer was in the S2 cell protein adsorbed onto the surface of the plastic dish. The dpp protein could be released into solution by increased salt concentration and nonionic detergent. Under these conditions, the amino-terminal and carboxy-terminal portions of dpp were not associated in a stable complex. Images PMID:1692958

  17. 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. PMID:21477078

  18. Growth Hormone Induces Transforming Growth Factor-Beta-Induced Protein in Podocytes: Implications for Podocyte Depletion and Proteinuria.

    PubMed

    Chitra, P Swathi; Swathi, T; Sahay, Rakesh; Reddy, G Bhanuprakash; Menon, Ram K; Kumar, P Anil

    2015-09-01

    The glomerular podocytes form a major size selective barrier for the filtration of serum proteins and reduced podocyte number is a critical event in the pathogenesis of proteinuria during diabetic nephropathy (DN). An elevated level of growth hormone (GH) is implicated as a causative factor in the development of nephropathy in patients with type 1 diabetes mellitus. We have previously shown that podocytes express GH receptor and are a target for GH action. To elucidate the molecular basis for the effects of GH on podocyte depletion, we conducted PCR-array analyses for extracellular matrix and adhesion molecules in podocytes. Our studies reveal that GH increases expression of a gene that encodes transforming growth factor-beta-induced protein (TGFBIp) expression. Similarly, microarray data retrieved from the Nephromine database revealed elevation of TGFBIp in patients with DN. Treatment with GH results in increased secretion of extracellular TGFBIp by podocytes. Both GH and TGFBIp induced apoptosis and epithelial mesenchymal transition (EMT) of podocytes. Exposure of podocytes to GH and TGFBIp resulted in increased migration of cells and altered podocyte permeability to albumin across podocyte monolayer. Administration of GH to rats induced EMT and apoptosis in the glomerular fraction of the kidney. Therefore, we conclude that the GH-dependent increase in TGFBIp in the podocyte is one of the mechanisms responsible for podocyte depletion in DN. PMID:25740786

  19. The Stationary-Phase Cells of Saccharomyces cerevisiae Display Dynamic Actin Filaments Required for Processes Extending Chronological Life Span

    PubMed Central

    Lejskova, Renata; Malcova, Ivana

    2015-01-01

    Stationary-growth-phase Saccharomyces cerevisiae yeast cultures consist of nondividing cells that undergo chronological aging. For their successful survival, the turnover of proteins and organelles, ensured by autophagy and the activation of mitochondria, is performed. Some of these processes are engaged in by the actin cytoskeleton. In S. cerevisiae stationary-phase cells, F actin has been shown to form static aggregates named actin bodies, subsequently cited to be markers of quiescence. Our in vivo analyses revealed that stationary-phase cultures contain cells with dynamic actin filaments, besides the cells with static actin bodies. The cells with dynamic actin displayed active endocytosis and autophagy and well-developed mitochondrial networks. Even more, stationary-phase cell cultures grown under calorie restriction predominantly contained cells with actin cables, confirming that the presence of actin cables is linked to successful adaptation to stationary phase. Cells with actin bodies were inactive in endocytosis and autophagy and displayed aberrations in mitochondrial networks. Notably, cells of the respiratory activity-deficient cox4Δ strain displayed the same mitochondrial aberrations and actin bodies only. Additionally, our results indicate that mitochondrial dysfunction precedes the formation of actin bodies and the appearance of actin bodies corresponds to decreased cell fitness. We conclude that the F-actin status reflects the extent of damage that arises from exponential growth. PMID:26351139

  20. Dental pulp stem cells suppress the proliferation of lymphocytes via transforming growth factor-β1.

    PubMed

    Ding, Gang; Niu, Jianyi; Liu, Yi

    2015-04-01

    Dental pulp stem cells (DPSCs) possess self-renewal capability, multi-lineage differentiation potential, and can generate a dentin-pulp-like tissue in vivo, which is promising for tooth regeneration. To enlarge the cells resource of DPSCs and explore the feasibility of DPSCs-mediated immune therapy, it is prerequisite to investigate the immunological properties of DPSCs and the underlying mechanisms. Human DPSCs and peripheral blood mononuclear cells were isolated and cultured. Then we used lymphocytes proliferation assays, cytokines detection, Transwell cultures, neutralization experiments, and flow cytometry to examine the in vitro immune characteristics of DPSCs. We found that DPSCs failed to stimulate allogeneic T cells proliferation and suppressed T cells proliferation, B cells proliferation, and mixed lymphocyte reaction. In addition, DPSCs could up-regulate IL-10, down-regulate the production of IL-2, IL-17, and IFN-γ, and did not affect the production of IL-6. Monoclonal antibody against transforming growth factor-β1 restored the T cells proliferation inhibited by DPSCs. Moreover, the population of regulatory T cells increased significantly and T-helper 17 cells decreased significantly in peripheral blood mononuclear cells co-cultured with DPSCs. These data confirmed that DPSCs are low immunogenic, could inhibit the proliferation of lymphocytes, regulate the production of cytokines in vitro, and the secretion of transforming growth factor-β1 may be involved in this event. PMID:25605036

  1. Retinoic acid modulates rat Ito cell proliferation, collagen, and transforming growth factor beta production.

    PubMed Central

    Davis, B H; Kramer, R T; Davidson, N O

    1990-01-01

    Recent studies suggest that vitamin A plays an inhibitory role with respect to "activation" of the hepatic Ito cell, a likely effector of hepatic fibrogenesis. Ito cell "activation" during fibrogenesis is characterized by a decrease in intracellular vitamin A and an increase in cellular proliferation and collagen production. To explore the hypothesis that retinoids have the capacity to diminish Ito cell activation, cultured Ito cells were exposed to retinoic acid and its effects assessed on three key features: cell proliferation, collagen protein production and mRNA abundance, and transforming growth factor beta protein production. Retinoic acid was 100-1,000X more potent than retinol with respect to inhibition of Ito cell proliferation. Interstitial collagen and transforming growth factor beta production were also reduced by 10(-6) M retinoic acid. The relative abundance of type I collagen mRNA however, was not significantly altered. By contrast, retinoic acid administration to rats caused a marked reduction in the abundance of type I collagen mRNA in both total hepatic and purified Ito cell RNA. The relative abundance of rat hepatic fibronectin or apolipoprotein E mRNA was not significantly altered. These studies demonstrate that retinoic acid can differentially modulate several key features of hepatic fibrogenesis in vitro and in vivo. Images PMID:2254460

  2. Toll-Like Receptor 4 Signaling Augments Transforming Growth Factor-β Responses

    PubMed Central

    Bhattacharyya, Swati; Kelley, Kathleen; Melichian, Denisa S.; Tamaki, Zenshiro; Fang, Feng; Su, Yunyun; Feng, Gilbert; Pope, Richard M.; Budinger, G.R. Scott; Mutlu, Gökhan M.; Lafyatis, Robert; Radstake, Timothy; Feghali-Bostwick, Carol; Varga, John

    2014-01-01

    Because recent studies implicate Toll-like receptors (TLRs) in the pathogenesis of fibrosis, we sought to investigate the in vitro and in vivo role and mechanism of TLR4-mediated fibroblast responses in fibrogenesis. We found that TLR4 was constitutively expressed, and accumulation of endogenous TLR4 ligands significantly elevated, in lesional skin and lung tissues from patients with scleroderma. Activation of TLR4 signaling in explanted fibroblasts resulted in enhanced collagen synthesis and increased expression of multiple genes involved in tissue remodeling and extracellular matrix homeostasis. Moreover, TLR4 dramatically enhanced the sensitivity of fibroblasts to the stimulatory effect of transforming growth factor-β1. These profibrotic responses were abrogated by both genetic and pharmacological disruption of TLR4 signaling in vitro, and skin fibrosis induced by bleomycin in vivo was attenuated in mice harboring a mutated TLR4. Activation of TLR4 in fibroblasts augmented the intensity of canonical Smad signaling, and was accompanied by suppression of anti-fibrotic microRNA expression. Together, these results suggest a novel model to account for persistent fibrogenesis in scleroderma, in which activation of fibroblast TLR4 signaling, triggered by damage-associated endogenous TLR4 ligands, results in augmented transforming growth factor-β1 sensitivity with increased matrix production and progressive connective tissue remodeling. Under these conditions, fibroblast TLR4 serves as the switch for converting self-limited tissue repair into intractable fibrosis. PMID:23141927

  3. Strong magnetic field-assisted growth of carbon nanofibers and its microstructural transformation mechanism

    PubMed Central

    Luo, Chengzhi; Fu, Qiang; Pan, Chunxu

    2015-01-01

    It is well-known that electric and magnetic fields can control the growth direction, morphology and microstructure of one-dimensional carbon nanomaterials (1-DCNMs), which plays a key role for its potential applications in micro-nano-electrics and devices. In this paper, we introduce a novel process for controlling growth of carbon nanofibers (CNFs) with assistance of a strong magnetic field (up to 0.5 T in the center) in a chemical vapor deposition (CVD) system. The results reveal that: 1) The CNFs get bundled when grown in the presence of a strong magnetic field and slightly get aligned parallel to the direction of the magnetic field; 2) The CNFs diameter become narrowed and homogenized with increase of the magnetic field; 3) With the increase of the magnetic field, the microstructure of CNFs is gradually changed, i.e., the strong magnetic field makes the disordered “solid-cored” CNFs transform into a kind of bamboo-liked carbon nanotubes; 4) We propose a mechanism that the reason for these variations and transformation is due to diamagnetic property of carbon atoms, so that it has direction selectivity in the precipitation process. PMID:25761381

  4. Strong magnetic field-assisted growth of carbon nanofibers and its microstructural transformation mechanism.

    PubMed

    Luo, Chengzhi; Fu, Qiang; Pan, Chunxu

    2015-01-01

    It is well-known that electric and magnetic fields can control the growth direction, morphology and microstructure of one-dimensional carbon nanomaterials (1-DCNMs), which plays a key role for its potential applications in micro-nano-electrics and devices. In this paper, we introduce a novel process for controlling growth of carbon nanofibers (CNFs) with assistance of a strong magnetic field (up to 0.5 T in the center) in a chemical vapor deposition (CVD) system. The results reveal that: 1) The CNFs get bundled when grown in the presence of a strong magnetic field and slightly get aligned parallel to the direction of the magnetic field; 2) The CNFs diameter become narrowed and homogenized with increase of the magnetic field; 3) With the increase of the magnetic field, the microstructure of CNFs is gradually changed, i.e., the strong magnetic field makes the disordered "solid-cored" CNFs transform into a kind of bamboo-liked carbon nanotubes; 4) We propose a mechanism that the reason for these variations and transformation is due to diamagnetic property of carbon atoms, so that it has direction selectivity in the precipitation process. PMID:25761381

  5. Strong magnetic field-assisted growth of carbon nanofibers and its microstructural transformation mechanism

    NASA Astrophysics Data System (ADS)

    Luo, Chengzhi; Fu, Qiang; Pan, Chunxu

    2015-03-01

    It is well-known that electric and magnetic fields can control the growth direction, morphology and microstructure of one-dimensional carbon nanomaterials (1-DCNMs), which plays a key role for its potential applications in micro-nano-electrics and devices. In this paper, we introduce a novel process for controlling growth of carbon nanofibers (CNFs) with assistance of a strong magnetic field (up to 0.5 T in the center) in a chemical vapor deposition (CVD) system. The results reveal that: 1) The CNFs get bundled when grown in the presence of a strong magnetic field and slightly get aligned parallel to the direction of the magnetic field; 2) The CNFs diameter become narrowed and homogenized with increase of the magnetic field; 3) With the increase of the magnetic field, the microstructure of CNFs is gradually changed, i.e., the strong magnetic field makes the disordered ``solid-cored'' CNFs transform into a kind of bamboo-liked carbon nanotubes; 4) We propose a mechanism that the reason for these variations and transformation is due to diamagnetic property of carbon atoms, so that it has direction selectivity in the precipitation process.

  6. Alterations in cellular differentiation, mitogenesis, cytoskeleton and growth characteristics during Syrian hamster embryo cell multistep in vitro transformation.

    PubMed

    Isfort, R J; Cody, D B; Doersen, C J; Kerckaert, G A; Leboeuf, R A

    1994-10-01

    In vitro Syrian hamster embryo (SHE) cell transformation is a neoplastic process that proceeds through several identifiable consecutive stages including in vitro morphological transformation (mt), acquisition of immortality (I+), acquisition of tumorigenicity (T+) and tumor-derived cells (I'TD). Eight transformed lineages consisting of cells at the mt, I+, T+ and I'TD stages were assayed for alterations in general markers of cell differentiation, mitogenic signaling pathways, cytoskeleton and cellular growth in 3D matrix. Alterations in cellular differentiation markers included a decrease in H19 gene expression and placental alkaline phosphatase enzymatic activity at the mt stage in all lineages examined with a complete absence of H19 gene expression and placental alkaline phosphatase enzymatic activity by the I'TD stage in a majority of transformed lineages. Changes in mitogenic signaling pathways included the production of autocrine growth factors and alterations in growth factor-induced immediate early gene expression by the I'TD stage of transformation in the majority of transformed lineages investigated. By the I'TD stage of transformation in most lineages, changes in both the cytoskeleton (including a decrease in tropomyosin-I gene expression) and the Matrigel growth characteristics of SHE cells were observed. PMID:7927892

  7. Electrostatic Interactions Between the Bni1p Formin FH2 Domain and Actin Influence Actin Filament Nucleation

    PubMed Central

    Baker, Joseph L.; Courtemanche, Naomi; Parton, Daniel L.; McCullagh, Martin; Pollard, Thomas D.; Voth, Gregory A.

    2014-01-01

    SUMMARY Formins catalyze nucleation and growth of actin filaments. Here we study the structure and interactions of actin with the FH2 domain of budding yeast formin Bni1p. We built an all-atom model of the formin dimer on an Oda actin filament 7-mer and studied structural relaxation and inter-protein interactions by molecular dynamics simulations. These simulations produced a refined model for the FH2 dimer associated with the barbed end of the filament and revealed electrostatic interactions between the formin knob and actin target-binding cleft. Mutations of two formin residues contributing to these interactions (R1423N, K1467L or both) reduced the interaction energies between the proteins, and in coarse-grained simulations the formin lost more inter-protein contacts with an actin dimer than with an actin 7-mer. Biochemical experiments confirmed a strong influence of these mutations on Bni1p-mediated actin filament nucleation, but not elongation, suggesting that different interactions contribute to these two functions of formins. PMID:25482541

  8. Arabidopsis Actin Depolymerizing Factor4 Modulates the Stochastic Dynamic Behavior of Actin Filaments in the Cortical Array of Epidermal Cells[C][W

    PubMed Central

    Henty, Jessica L.; Bledsoe, Samuel W.; Khurana, Parul; Meagher, Richard B.; Day, Brad; Blanchoin, Laurent; Staiger, Christopher J.

    2011-01-01

    Actin filament arrays are constantly remodeled as the needs of cells change as well as during responses to biotic and abiotic stimuli. Previous studies demonstrate that many single actin filaments in the cortical array of living Arabidopsis thaliana epidermal cells undergo stochastic dynamics, a combination of rapid growth balanced by disassembly from prolific severing activity. Filament turnover and dynamics are well understood from in vitro biochemical analyses and simple reconstituted systems. However, the identification in living cells of the molecular players involved in controlling actin dynamics awaits the use of model systems, especially ones where the power of genetics can be combined with imaging of individual actin filaments at high spatial and temporal resolution. Here, we test the hypothesis that actin depolymerizing factor (ADF)/cofilin contributes to stochastic filament severing and facilitates actin turnover. A knockout mutant for Arabidopsis ADF4 has longer hypocotyls and epidermal cells when compared with wild-type seedlings. This correlates with a change in actin filament architecture; cytoskeletal arrays in adf4 cells are significantly more bundled and less dense than in wild-type cells. Several parameters of single actin filament turnover are also altered. Notably, adf4 mutant cells have a 2.5-fold reduced severing frequency as well as significantly increased actin filament lengths and lifetimes. Thus, we provide evidence that ADF4 contributes to the stochastic dynamic turnover of actin filaments in plant cells. PMID:22010035

  9. ATP differentially upregulates fibroblast growth factor 2 and transforming growth factor α in neonatal and adult mice: effect on neuroproliferation.

    PubMed

    Jia, C; Cussen, A R; Hegg, C C

    2011-03-17

    Multiple neurotrophic factors play a role in proliferation, differentiation and survival in the olfactory epithelium (OE); however, the signaling cascade has not been fully elucidated. We tested the hypotheses that ATP induces the synthesis and secretion of two neurotrophic factors, fibroblast growth factor 2 (FGF2) and transforming growth factor alpha (TGFα), and that these neurotrophic factors have a role in inducing proliferation. Protein levels of FGF2 and TGFα were increased 20 h post-intranasal instillation of ATP compared to vehicle control in adult Swiss Webster mice. Pre-intranasal treatment with purinergic receptor antagonist pyridoxal-phosphate-6-azophenyl-20,40-disulfonic acid (PPADS) significantly blocked this ATP-induced increase, indicating that upregulation of FGF2 and TGFα expression is mediated by purinergic receptor activation. However, in neonatal mouse, intranasal instillation of ATP significantly increased the protein levels of FGF2, but not TGFα. Likewise, ATP evoked the secretion of FGF2, but not TGFα, from neonatal mouse olfactory epithelial slices and PPADS significantly blocked ATP-evoked FGF2 release. To determine the role of FGF2 and TGFα in inducing proliferation, 5-bromo-2-deoxyuridine (BrdU) incorporation was examined in adult olfactory epithelium. Intranasal treatment with FGF receptor inhibitor PD173074 or epidermal growth factor receptor inhibitor AG1478 following ATP instillation significantly blocked ATP-induced BrdU incorporation. Collectively, these data demonstrate that ATP induces proliferation in adult mouse olfactory epithelium by promoting FGF2 and TGFα synthesis and activation of their receptors. These data suggest that different mechanisms regulate neurogenesis in neonatal and adult OE, and FGF2 and TGFα may have different roles throughout development. PMID:21187124

  10. Recruitment of Matrix Metalloproteinase-9 (MMP-9) to the Fibroblast Cell Surface by Lysyl Hydroxylase 3 (LH3) Triggers Transforming Growth Factor-β (TGF-β) Activation and Fibroblast Differentiation*

    PubMed Central

    Dayer, Cynthia; Stamenkovic, Ivan

    2015-01-01

    Solid tumor growth triggers a wound healing response. Similar to wound healing, fibroblasts in the tumor stroma differentiate into myofibroblasts (also referred to as cancer-associated fibroblasts) primarily, but not exclusively, in response to transforming growth factor-β (TGF-β). Myofibroblasts in turn enhance tumor progression by remodeling the stroma. Among proteases implicated in stroma remodeling, matrix metalloproteinases (MMPs), including MMP-9, play a prominent role. Recent evidence indicates that MMP-9 recruitment to the tumor cell surface enhances tumor growth and invasion. In the present work, we addressed the potential relevance of MMP-9 recruitment to and activity at the surface of fibroblasts. We show that recruitment of MMP-9 to the fibroblast cell surface occurs through its fibronectin-like (FN) domain and that the molecule responsible for the recruitment is lysyl hydroxylase 3 (LH3). Functional assays suggest that both pro- and active MMP-9 trigger α-smooth muscle actin expression in cultured fibroblasts, reflecting myofibroblast differentiation, possibly as a result of TGF-β activation. Moreover, the recombinant FN domain inhibited both MMP-9-induced TGF-β activation and α-smooth muscle actin expression by displacing MMP-9 from the fibroblast cell surface. Together our results uncover LH3 as a new docking receptor of MMP-9 on the fibroblast cell surface and demonstrate that the MMP-9 FN domain is essential for the interaction. They also show that the recombinant FN domain inhibits MMP-9-induced TGF-β activation and fibroblast differentiation, providing a potentially attractive therapeutic reagent toward attenuating tumor progression where MMP-9 activity is strongly implicated. PMID:25825495

  11. 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

  12. Reduced susceptibility of mice overexpressing transforming growth factor α to dextran sodium sulphate induced colitis

    PubMed Central

    Egger, B; Carey, H; Procaccino, F; Chai, N; Sandgren, E; Lakshmanan, J; Buslon, V; French, S; Buchler, M; Eysselein, V

    1998-01-01

    Background—Transforming growth factor α (TGF-α) knockout mice have increased susceptibility to dextran sodium sulphate (DSS) induced colitis. 
Aim—To substantiate the findings that TGF-α is a key mediator of colonic mucosal protection and/or repair mechanisms by evaluating the susceptibility of mice overexpressing TGF-α to DSS induced colitis. 
Methods—TGF-α overexpression was induced in transgenic mice by ZnSO4 administration in drinking water (TG+). Three groups were used as controls: one transgenic group without ZnSO4 administration (TG−), and two non-transgenic littermate groups receiving ZnSO4 (Non-TG+) or only water (Non-TG−). Acute colitis was induced in all groups by administration of DSS (5%, w/v) in drinking water for six days ad libitum. 
Results—About 35-39% of the entire colonic mucosa was destroyed in Non-TG−, Non-TG+, and TG− animals compared with 9% in TG+ mice. The crypt damage score was 18.7 (0.9), 18.2 (1.0), 18.9(0.8), and 6.8 (1.5) (means (SEM)) in Non-TG−, Non-TG+, TG−, and TG+ mice respectively. Mucin and bromodeoxyuridine staining were markedly enhanced in colons of TG+ mice compared with controls, indicating increased mucosal protection and regeneration. 
Conclusions—The significantly reduced susceptibility of mice overexpressing TGF-α to DSS further substantiates that endogenous TGF-α is a pivotal mediator of protection and/or healing mechanisms in the colon. 

 Keywords: transforming growth factor α; epidermal growth factor; dextran sodium sulphate; colitis; inflammatory bowel disease; transgenic mice PMID:9771407

  13. Transforming growth factor-beta as a differentiating factor for cultured smooth muscle cells.

    PubMed

    Gawaziuk, J P; X; Sheikh, F; Cheng, Z-Q; Cattini, P A; Stephens, N L

    2007-10-01

    The aim of the present study was to determine whether the development of supercontractile smooth muscle cells, contributing to the nonspecific hyperreactivity of airways in asthmatic patients, is due to transforming growth factor (TGF)-beta. In cultured smooth muscle cells starved by removal of 10% foetal bovine serum for 7 days, growth arrest was seen; 30% became elongated and demonstrated super contractility. Study of conditioned medium suggested that the differentiating factor was TGF-beta. Sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) was carried out on conditioned medium from the arrested cells. Two protein bands were identified as matrix metalloproteinase (MMP)-2 and TGF-beta1. To determine second messenger signalling by SMAD2, Western blotting and confocal microscopy were employed. Conditioned medium from arrested cultures showed the presence of MMP-2 and TGF-beta1, as revealed by SDS-PAGE; 68- and 25-kDa bands were seen. Differentiation was confirmed by upregulation of marker proteins, smooth muscle type myosin heavy chain and myosin light chain kinase. Confirmation was obtained by downregulating these proteins with decorin treatment, which reduces the levels of active TGF-beta and an adenoviral dominant-negative vector coding for a mutated type II TGF-beta-receptor. Activation of second messenger signalling was demonstrated immunocytochemically by the presence of phosphorylated SMAD2 and SMAD4. Transforming growth factor-beta is likely to be the differentiating factor responsible for the development of these supercontractile smooth muscle cells. The development of such cells in vivo after cessation of an asthmatic attack could contribute to the nonspecific hyperreactivity of airways seen in patients. PMID:17596270

  14. Nerve growth factor stimulates axon outgrowth through negative regulation of growth cone actomyosin restraint of microtubule advance

    PubMed Central

    Turney, Stephen G.; Ahmed, Mostafa; Chandrasekar, Indra; Wysolmerski, Robert B.; Goeckeler, Zoe M.; Rioux, Robert M.; Whitesides, George M.; Bridgman, Paul C.

    2016-01-01

    Nerve growth factor (NGF) promotes growth, differentiation, and survival of sensory neurons in the mammalian nervous system. Little is known about how NGF elicits faster axon outgrowth or how growth cones integrate and transform signal input to motor output. Using cultured mouse dorsal root ganglion neurons, we found that myosin II (MII) is required for NGF to stimulate faster axon outgrowth. From experiments inducing loss or gain of function of MII, specific MII isoforms, and vinculin-dependent adhesion-cytoskeletal coupling, we determined that NGF causes decreased vinculin-dependent actomyosin restraint of microtubule advance. Inhibition of MII blocked NGF stimulation, indicating the central role of restraint in directed outgrowth. The restraint consists of myosin IIB- and IIA-dependent processes: retrograde actin network flow and transverse actin bundling, respectively. The processes differentially contribute on laminin-1 and fibronectin due to selective actin tethering to adhesions. On laminin-1, NGF induced greater vinculin-dependent adhesion–cytoskeletal coupling, which slowed retrograde actin network flow (i.e., it regulated the molecular clutch). On fibronectin, NGF caused inactivation of myosin IIA, which negatively regulated actin bundling. On both substrates, the result was the same: NGF-induced weakening of MII-dependent restraint led to dynamic microtubules entering the actin-rich periphery more frequently, giving rise to faster elongation. PMID:26631553

  15. Reconstitution of actin-based motility of Listeria and Shigella using pure proteins

    NASA Astrophysics Data System (ADS)

    Loisel, Thomas P.; Boujemaa, Rajaa; Pantaloni, Dominique; Carlier, Marie-France

    1999-10-01

    Actin polymerization is essential for cell locomotion and is thought to generate the force responsible for cellular protrusions. The Arp2/3 complex is required to stimulate actin assembly at the leading edge in response to signalling. The bacteria Listeria and Shigella bypass the signalling pathway and harness the Arp2/3 complex to induce actin assembly and to propel themselves in living cells. However, the Arp2/3 complex alone is insufficient to promote movement. Here we have used pure components of the actin cytoskeleton to reconstitute sustained movement in Listeria and Shigella in vitro. Actin-based propulsion is driven by the free energy released by ATP hydrolysis linked to actin polymerization, and does not require myosin. In addition to actin and activated Arp2/3 complex, actin depolymerizing factor (ADF, or cofilin) and capping protein are also required for motility as they maintain a high steady-state level of G-actin, which controls the rate of unidirectional growth of actin filaments at the surface of the bacterium. The movement is more effective when profilin, α-actinin and VASP (for Listeria) are also included. These results have implications for our understanding of the mechanism of actin-based motility in cells.

  16. Pin1 promotes transforming growth factor-beta-induced migration and invasion.

    PubMed

    Matsuura, Isao; Chiang, Keng-Nan; Lai, Chen-Yu; He, Dongming; Wang, Guannan; Ramkumar, Romila; Uchida, Takafumi; Ryo, Akihide; Lu, Kunping; Liu, Fang

    2010-01-15

    Transforming growth factor-beta (TGF-beta) regulates a wide variety of biological activities. It induces potent growth-inhibitory responses in normal cells but promotes migration and invasion of cancer cells. Smads mediate the TGF-beta responses. TGF-beta binding to the cell surface receptors leads to the phosphorylation of Smad2/3 in their C terminus as well as in the proline-rich linker region. The serine/threonine phosphorylation sites in the linker region are followed by the proline residue. Pin1, a peptidyl-prolyl cis/trans isomerase, recognizes phosphorylated serine/threonine-proline motifs. Here we show that Smad2/3 interacts with Pin1 in a TGF-beta-dependent manner. We further show that the phosphorylated threonine 179-proline motif in the Smad3 linker region is the major binding site for Pin1. Although epidermal growth factor also induces phosphorylation of threonine 179 and other residues in the Smad3 linker region the same as TGF-beta, Pin1 is unable to bind to the epidermal growth factor-stimulated Smad3. Further analysis suggests that phosphorylation of Smad3 in the C terminus is necessary for the interaction with Pin1. Depletion of Pin1 by small hairpin RNA does not significantly affect TGF-beta-induced growth-inhibitory responses and a number of TGF-beta/Smad target genes analyzed. In contrast, knockdown of Pin1 in human PC3 prostate cancer cells strongly inhibited TGF-beta-mediated migration and invasion. Accordingly, TGF-beta induction of N-cadherin, which plays an important role in migration and invasion, is markedly reduced when Pin1 is depleted in PC3 cells. Because Pin1 is overexpressed in many cancers, our findings highlight the importance of Pin1 in TGF-beta-induced migration and invasion of cancer cells. PMID:19920136

  17. 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

  18. Regulators of Actin Dynamics in Gastrointestinal Tract Tumors

    PubMed Central

    Steinestel, Konrad; Wardelmann, Eva; Hartmann, Wolfgang; Grünewald, Inga

    2015-01-01

    Reorganization of the actin cytoskeleton underlies cell migration in a wide variety of physiological and pathological processes, such as embryonic development, wound healing, and tumor cell invasion. It has been shown that actin assembly and disassembly are precisely regulated by intracellular signaling cascades that respond to changes in the cell microenvironment, ligand binding to surface receptors, or oncogenic transformation of the cell. Actin-nucleating and actin-depolymerizing (ANFs/ADFs) and nucleation-promoting factors (NPFs) regulate cytoskeletal dynamics at the leading edge of migrating cells, thereby modulating cell shape; these proteins facilitate cellular movement and mediate degradation of the surrounding extracellular matrix by secretion of lytic proteases, thus eliminating barriers for tumor cell invasion. Accordingly, expression and activity of these actin-binding proteins have been linked to enhanced metastasis and poor prognosis in a variety of malignancies. In this review, we will summarize what is known about expression patterns and the functional role of actin regulators in gastrointestinal tumors and evaluate first pharmacological approaches to prevent invasion and metastatic dissemination of malignant cells. PMID:26345720

  19. Regulators of Actin Dynamics in Gastrointestinal Tract Tumors.

    PubMed

    Steinestel, Konrad; Wardelmann, Eva; Hartmann, Wolfgang; Grünewald, Inga

    2015-01-01

    Reorganization of the actin cytoskeleton underlies cell migration in a wide variety of physiological and pathological processes, such as embryonic development, wound healing, and tumor cell invasion. It has been shown that actin assembly and disassembly are precisely regulated by intracellular signaling cascades that respond to changes in the cell microenvironment, ligand binding to surface receptors, or oncogenic transformation of the cell. Actin-nucleating and actin-depolymerizing (ANFs/ADFs) and nucleation-promoting factors (NPFs) regulate cytoskeletal dynamics at the leading edge of migrating cells, thereby modulating cell shape; these proteins facilitate cellular movement and mediate degradation of the surrounding extracellular matrix by secretion of lytic proteases, thus eliminating barriers for tumor cell invasion. Accordingly, expression and activity of these actin-binding proteins have been linked to enhanced metastasis and poor prognosis in a variety of malignancies. In this review, we will summarize what is known about expression patterns and the functional role of actin regulators in gastrointestinal tumors and evaluate first pharmacological approaches to prevent invasion and metastatic dissemination of malignant cells. PMID:26345720

  20. Structural Polymorphism of the Actin-Espin System: A Prototypical System of Filaments and Linkers in Stereocilia

    SciTech Connect

    Purdy, Kirstin R.; Wong, Gerard C. L.; Bartles, James R.

    2007-02-02

    We examine the interaction between cytoskeletal F-actin and espin 3A, a prototypical actin bundling protein found in sensory cell microvilli, including ear cell stereocilia. Espin induces twist distortions in F-actin as well as facilitates bundle formation. Mutations in one of the two F-actin binding sites of espin, which have been implicated in deafness, can tune espin-actin interactions and radically transform the system's phase behavior. These results are compared to recent theoretical work on the general phase behavior linker-rod systems.

  1. Structural Polymorphism of the Actin-Espin System: A Prototypical System of Filaments and Linkers in Stereocilia

    NASA Astrophysics Data System (ADS)

    Purdy, Kirstin R.; Bartles, James R.; Wong, Gerard C. L.

    2007-02-01

    We examine the interaction between cytoskeletal F-actin and espin 3A, a prototypical actin bundling protein found in sensory cell microvilli, including ear cell stereocilia. Espin induces twist distortions in F-actin as well as facilitates bundle formation. Mutations in one of the two F-actin binding sites of espin, which have been implicated in deafness, can tune espin-actin interactions and radically transform the system’s phase behavior. These results are compared to recent theoretical work on the general phase behavior linker-rod systems.

  2. Regulation of Transforming Growth Factor β1, Platelet-Derived Growth Factor, and Basic Fibroblast Growth Factor by Silicone Gel Sheeting in Early-Stage Scarring

    PubMed Central

    Choi, Jaehoon; Lee, Eun Hee; Park, Sang Woo

    2015-01-01

    Background Hypertrophic scars and keloids are associated with abnormal levels of growth factors. Silicone gel sheets are effective in treating and preventing hypertrophic scars and keloids. There has been no report on the change in growth factors in the scar tissue following the use of silicone gel sheeting for scar prevention. A prospective controlled trial was performed to evaluate whether growth factors are altered by the application of a silicone gel sheet on a fresh surgical scar. Methods Four of seven enrolled patients completed the study. Transforming growth factor (TGF)-β1, platelet-derived growth factor (PDGF), and basic fibroblast growth factor (bFGF) were investigated immunohistochemically in biopsies taken from five scars at 4 months following surgery. Results In both the epidermis and the dermis, the expression of TGF-β1 (P=0.042 and P=0.042) and PDGF (P=0.043 and P=0.042) was significantly lower in the case of silicone gel sheet-treated scars than in the case of untreated scars. The expression of bFGF in the dermis was significantly higher in the case of silicone gel sheet-treated scars than in the case of untreated scars (P=0.042), but in the epidermis, the expression of bFGF showed no significant difference between the groups (P=0.655). Conclusions The levels of TGF-β1, PDGF, and bFGF are altered by the silicone gel sheet treatment, which might be one of the mechanisms of action in scar prevention. PMID:25606485

  3. Tracheal Smooth Muscle Cells Stimulated by Stem Cell Factor-c-Kit Coordinate the Production of Transforming Growth Factor-β1 and Fibroblast Growth Factor-2 Mediated by Chemokine (C-C Motif) Ligand 3.

    PubMed

    Oliveira, Luis Cezar Farias de; Danilucci, Taís Marolato; Chaves-Neto, Antonio Hernandes; Campanelli, Ana Paula; Silva, Tereza Cristina Cardoso da; Oliveira, Sandra Helena Penha

    2016-06-01

    The aim of this study was to evaluate the mechanism involved in the stem cell factor (SCF)-induced production of fibroblast growth factor-2 (FGF-2), transforming growth factor-β1 (TGF-β1), and chemokine (C-C motif) ligand 3 (CCL3) in tracheal smooth muscle cells (tSMCs) and the signaling pathway involved in the process. tSMC primary cultures were stimulated with SCF and evaluated at 24 h. Cells treated with specific antibodies did not show any immunolabeling for cytokeratin or fibroblast activation protein, but were positive for α-smooth muscle actin, indicating the purity of the primary cell line. Western blot analysis showed constitutive phosphorylation of c-Kit, as well as increased total protein and phosphorylated c-Kit levels in tSMCs after SCF stimulation. Flow cytometry analysis also showed an increase in cell-surface c-Kit expression in the presence of SCF. SCF induced TGF-β mRNA expression in tSMCs, as well as the production of TGF-β1, CCL3, and FGF-2. Pretreatment with anti-CCL3 antibody blocked TGF-β1 expression and partially inhibited FGF-2 production. On the other hand, anti-c-Kit antibody blocked TGF-β1 expression and FGF-2 production. Thus, TGF-β1 and FGF-2 production were mediated by CCL3 production through c-Kit. Pretreatment with mitogen-activated protein kinase kinase 1, p38, and Jun N-terminal kinase inhibitors showed that the effects mediated by SCF were involved with the modulation of mitogen-activated protein kinase (MAPK) pathways. Development of inhibitors targeting CCL3 through MAPK activation could thus be an attractive strategy to inhibit tSMC activation during asthma. PMID:27123814

  4. Transforming growth factor-beta and platelet-derived growth factor signal via c-Jun N-terminal kinase-dependent Smad2/3 phosphorylation in rat hepatic stellate cells after acute liver injury.

    PubMed

    Yoshida, Katsunori; Matsuzaki, Koichi; Mori, Shigeo; Tahashi, Yoshiya; Yamagata, Hideo; Furukawa, Fukiko; Seki, Toshihito; Nishizawa, Mikio; Fujisawa, Junichi; Okazaki, Kazuichi

    2005-04-01

    After liver injury, transforming growth factor-beta (TGF-beta) and platelet-derived growth factor (PDGF) regulate the activation of hepatic stellate cells (HSCs) and tissue remodeling. Mechanisms of PDGF signaling in the TGF-beta-triggered cascade are not completely understood. TGF-beta signaling involves phosphorylation of Smad2 and Smad3 at linker and C-terminal regions. Using antibodies to distinguish Smad2/3 phosphorylated at linker regions from those phosphorylated at C-terminal regions, we investigated Smad2/3-mediated signaling in rat liver injured by CCl(4) administration and in cultured HSCs. In acute liver injury, Smad2/3 were transiently phosphorylated at both regions. Although linker-phosphorylated Smad2 remained in the cytoplasm of alpha-smooth muscle actin-immunoreactive mesenchymal cells adjacent to necrotic hepatocytes in centrilobular areas, linker-phosphorylated Smad3 accumulated in the nuclei. c-Jun N-terminal kinase (JNK) in the activated HSCs directly phosphorylated Smad2/3 at linker regions. Co-treatment of primary cultured HSCs with TGF-beta and PDGF activated the JNK pathway, subsequently inducing endogenous linker phosphorylation of Smad2/3. The JNK pathway may be involved in migration of resident HSCs within the space of Disse to the sites of tissue damage because the JNK inhibitor SP600125 inhibited HSC migration induced by TGF-beta and PDGF signals. Moreover, treatment of HSCs with both TGF-beta and PDGF increased transcriptional activity of plasminogen activator inhibitor-1 through linker phosphorylation of Smad3. In conclusion, TGF-beta and PDGF activate HSCs by transmitting their signals through JNK-mediated Smad2/3 phosphorylation at linker regions, both in vivo and in vitro. PMID:15793284

  5. Transforming Growth Factor-β1 (TGF-β1)-stimulated Fibroblast to Myofibroblast Differentiation Is Mediated by Hyaluronan (HA)-facilitated Epidermal Growth Factor Receptor (EGFR) and CD44 Co-localization in Lipid Rafts*

    PubMed Central

    Midgley, Adam C.; Rogers, Mathew; Hallett, Maurice B.; Clayton, Aled; Bowen, Timothy; Phillips, Aled O.; Steadman, Robert

    2013-01-01

    Fibroblast to myofibroblast differentiation drives effective wound healing and is largely regulated by the cytokine transforming growth factor-β1 (TGF-β1). Myofibroblasts express α-smooth muscle actin and are present in granulation tissue, where they are responsible for wound contraction. Our previous studies show that fibroblast differentiation in response to TGF-β1 is dependent on and mediated by the linear polysaccharide hyaluronan (HA). Both the HA receptor, CD44, and the epidermal growth factor receptor (EGFR) are involved in this differentiation response. The aim of this study was to understand the mechanisms linking HA-, CD44-, and EGFR-regulated TGF-β1-dependent differentiation. CD44 and EGFR co-localization within membrane-bound lipid rafts was necessary for differentiation, and this triggered downstream mitogen-activated protein kinase (MAPK/ERK) and Ca2+/calmodulin kinase II (CaMKII) activation. We also found that ERK phosphorylation was upstream of CaMKII phosphorylation, that ERK activation was necessary for CaMKII signaling, and that both kinases were essential for differentiation. In addition, HA synthase-2 (HAS2) siRNA attenuated both ERK and CaMKII signaling and sequestration of CD44 into lipid rafts, preventing differentiation. In summary, the data suggest that HAS2-dependent production of HA facilitates TGF-β1-dependent fibroblast differentiation through promoting CD44 interaction with EGFR held within membrane-bound lipid rafts. This induces MAPK/ERK, followed by CaMKII activation, leading to differentiation. This pathway is synergistic with the classical TGF-β1-dependent SMAD-signaling pathway and may provide a novel opportunity for intervention in wound healing. PMID:23589287

  6. Mammary tumor suppression by transforming growth factor beta 1 transgene expression.

    PubMed Central

    Pierce, D F; Gorska, A E; Chytil, A; Meise, K S; Page, D L; Coffey, R J; Moses, H L

    1995-01-01

    In cell culture, type alpha transforming growth factor (TGF-alpha) stimulates epithelial cell growth, whereas TGF-beta 1 overrides this stimulatory effect and is growth inhibitory. Transgenic mice that overexpress TGF-alpha under control of the mouse mammary tumor virus (MMTV) promoter/enhancer exhibit mammary ductal hyperplasia and stochastic development of mammary carcinomas, a process that can be accelerated by administration of the chemical carcinogen 7,12-dimethylbenz[a]anthracene. MMTV-TGF-beta 1 transgenic mice display mammary ductal hypoplasia and do not develop mammary tumors. We report that in crossbreeding experiments involving the production of mice carrying both the MMTV-TGF-beta 1 and MMTV-TGF-alpha transgenes, there is marked suppression of mammary tumor formation and that MMTV-TGF-beta 1 transgenic mice are resistant to 7,12-dimethylbenz[a]anthracene-induced mammary tumor formation. These data demonstrate that overexpression of TGF-beta 1 in vivo can markedly suppress mammary tumor development. Images Fig. 1 Fig. 3 PMID:7753792

  7. Direct transfer of transforming growth factor beta 1 gene into arteries stimulates fibrocellular hyperplasia.

    PubMed Central

    Nabel, E G; Shum, L; Pompili, V J; Yang, Z Y; San, H; Shu, H B; Liptay, S; Gold, L; Gordon, D; Derynck, R

    1993-01-01

    The arterial wall responds to thrombosis or mechanical injury through the induction of specific gene products that increase cellular proliferation and connective tissue formation. These changes result in intimal hyperplasia that is observed in restenosis and the early phases of atherosclerosis. Transforming growth factor beta 1 (TGF-beta 1) is a secreted multi-functional protein that plays an important role in embryonal development and in repair following tissue injury. However, the function of TGF-beta 1 in vascular cell growth in vivo has not been defined. In this report, we have evaluated the role of TGF-beta 1 in the pathophysiology of intimal and medial hyperplasia by gene transfer of an expression plasmid encoding active TGF-beta 1 into porcine arteries. Expression of TGF-beta 1 in normal arteries resulted in substantial extracellular matrix production accompanied by intimal and medial hyperplasia. Increased procollagen, collagen, and proteoglycan synthesis in the neointima was demonstrated by immunohistochemistry relative to control transfected arteries. Expression of TGF-beta 1 induced a distinctly different program of gene expression and biologic response from the platelet-derived growth factor B (PDGF B) gene: procollagen synthesis induced by TGF-beta 1 was greater, and cellular proliferation was less prominent. These findings show that TGF-beta 1 differentially modulates extracellular matrix production and cellular proliferation in the arterial wall in vivo and could play a reparative role in the response to arterial injury. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 PMID:8248168

  8. Capsaicin Inhibits Preferentially the NADH Oxidase and Growth of Transformed Cells in Culture

    NASA Astrophysics Data System (ADS)

    Morre, D. James; Chueh, Pin-Ju; Morre, Dorothy M.

    1995-03-01

    A hormone- and growth factor-stimulated NADH oxidase of the mammalian plasma membrane, constitutively activated in transformed cells, was inhibited preferentially in HeLa, ovarian carcinoma, mammary adenocarcinoma, and HL-60 cells, all of human origin, by the naturally occurring quinone analog capsaicin (8-methyl-N-vanillyl-6-noneamide), compared with plasma membranes from human mammary epithelial, rat liver, normal rat kidney cells, or HL-60 cells induced to differentiate with dimethyl sulfoxide. With cells in culture, capsaicin preferentially inhibited growth of HeLa, ovarian carcinoma, mammary adenocarcinoma, and HL-60 cells but was largely without effect on the mammary epithelial cells, rat kidney cells, or HL-60 cells induced to differentiate with dimethyl sulfoxide. Inhibited cells became smaller and cell death was accompanied by a condensed and fragmented appearance of the nuclear DNA, as revealed by fluorescence microscopy with 4',6-diamidino-2-phenylindole, suggestive of apoptosis. The findings correlate capsaicin inhibition of cell surface NADH oxidase activity and inhibition of growth that correlate with capsaicin-induced apoptosis.

  9. Regulation of the ovarian reserve by members of the transforming growth factor beta family

    PubMed Central

    Pangas, Stephanie A.

    2012-01-01

    Genetic or environmental factors that affect the endowment of oocytes, their assembly nto primordial follicles, or their subsequent entry into the growing follicle pool can disrupt reproductive function and may underlie disorders such as primary ovarian insufficiency. Mouse models have been instrumental in identifying genes important in ovarian development, and a number of genes now associated with ovarian dysfunction in women were first identified as causing reproductive defects in knockout mice. The transforming growth factor beta (TGFB) family consists of developmentally important growth factors that include the TGFBs, anti-Müllerian hormone (AMH), activins, bone morphogenetic proteins (BMPs), and growth and differentiation factor 9 (GDF9). The ovarian primordial follicle pool is the source of oocytes in adults. Development of this pool can be grossly divided into three key processes: (1) establishment of oocytes during embryogenesis followed by (2) assembly and (3) activation of the primordial follicle. Disruptions in any of these processes may cause reproductive dysfunction. Most members of the TGFB family show pivotal roles in each of these areas. Understanding the phenotypes of various mouse models for this protein family will be directly relevant to understanding how disruptions in TGFB family signaling result in reproductive diseases in women and will present new areas for development of tailored diagnostics and interventions for infertility. PMID:22847922

  10. Microsatellite mutation of type II transforming growth factor-beta receptor is rare in atherosclerotic plaques.

    PubMed

    Clark, K J; Cary, N R; Grace, A A; Metcalfe, J C

    2001-04-01

    A somatic mutation within a microsatellite polyA tract in the coding region of the type II transforming growth factor (TGF)-beta receptor gene was reported to occur in human atherosclerotic and restenotic lesions. This mutation occurs frequently in colorectal cancer with the replication error repair phenotype and results in loss of sensitivity to the growth inhibitory effects of TGF-beta in cells from the tumors. The mutation was proposed to account for the clonal expansion of vascular smooth muscle cells observed in atherosclerotic plaques, through loss of the growth inhibitory effect of TGF-beta. The frequency of the mutation and the extent of clonal expansion of the mutated cells have major implications for the mechanism of atherogenesis and therapeutic strategies. We analyzed a set of 22 coronary arterial and 9 aortic samples containing early to advanced atherosclerotic lesions for the mutation in the type II TGF-beta receptor polyA tract. Only 1 coronary arterial sample from an advanced lesion showed detectable amounts of the mutation, present at a low level (8% of the DNA sample). The data imply that the mutation occurs only at low frequency and is not a major mechanistic contributor to the development of atherosclerosis. PMID:11304472

  11. Immunohistochemical localization of transforming growth factor beta isoforms in asbestos-related diseases.

    PubMed Central

    Jagirdar, J; Lee, T C; Reibman, J; Gold, L I; Aston, C; Bégin, R; Rom, W N

    1997-01-01

    Transforming growth factor beta (TGF-beta), a multifunctional cytokine and growth factor, plays a key role in scarring and fibrotic processes because of its ability to induce extracellular matrix proteins and modulate the growth and immune function of many cell types. These effects are important in inflammatory disorders with fibrosis and cancer. The asbestos-related diseases are characterized by fibrosis in the lower respiratory tract and pleura and increased occurrence of lung cancer and mesothelioma. We performed immunohistochemistry with isoform-specific antibodies to the three TGF-beta isoforms on 16 autopsy lungs from Quebec, Canada, asbestos miners and millers. There was increased immunolocalization of all three TGF-beta isoforms in the fibrotic lesions of asbestosis and pleural fibrosis. The hyperplastic type II pneumocytes contained all three isoforms. By contrast, there was differential spatial immunostaining for the TGF-beta isoforms in malignant mesothelioma, with TGF-beta 1 in the stroma but TGF-beta 2 in the tumor cells. These data are consistent with an important role for TGF-beta in accumulation of extracellular matrix and cell proliferation in asbestos-related diseases. Images Figure 1. Figure 2. Figure 3. Figure 4. Figure 5. Figure 6. Figure 7. Figure 8. Figure 9. PMID:9400723

  12. Lysyl oxidase binds transforming growth factor-beta and regulates its signaling via amine oxidase activity.

    PubMed

    Atsawasuwan, Phimon; Mochida, Yoshiyuki; Katafuchi, Michitsuna; Kaku, Masaru; Fong, Keith S K; Csiszar, Katalin; Yamauchi, Mitsuo

    2008-12-01

    Lysyl oxidase (LOX), an amine oxidase critical for the initiation of collagen and elastin cross-linking, has recently been shown to regulate cellular activities possibly by modulating the functions of growth factors. In this study, we investigated the interaction between LOX and transforming growth factor-beta1 (TGF-beta1), a potent growth factor abundant in bone, the effect of LOX on TGF-beta1 signaling, and its potential mechanism. The specific binding between mature LOX and mature TGF-beta1 was demonstrated by immunoprecipitation and glutathione S-transferase pulldown assay in vitro. Both proteins were colocalized in the extracellular matrix in an osteoblastic cell culture system, and the binding complex was identified in the mineral-associated fraction of bone matrix. Furthermore, LOX suppressed TGF-beta1-induced Smad3 phosphorylation likely through its amine oxidase activity. The data indicate that LOX binds to mature TGF-beta1 and enzymatically regulates its signaling in bone and thus may play an important role in bone maintenance and remodeling. PMID:18835815

  13. Competition for actin between two distinct F-actin networks defines a bistable switch for cell polarization.

    PubMed

    Lomakin, Alexis J; Lee, Kun-Chun; Han, Sangyoon J; Bui, Duyen A; Davidson, Michael; Mogilner, Alex; Danuser, Gaudenz

    2015-11-01

    Symmetry-breaking polarization enables functional plasticity of cells and tissues and is yet not well understood. Here we show that epithelial cells, hard-wired to maintain a static morphology and to preserve tissue organization, can spontaneously switch to a migratory polarized phenotype after relaxation of the actomyosin cytoskeleton. We find that myosin II engages actin in the formation of cortical actomyosin bundles and thus makes it unavailable for deployment in the process of dendritic growth normally driving cell motility. Under low-contractility regimes, epithelial cells polarize in a front-back manner owing to the emergence of actin retrograde flows powered by dendritic polymerization of actin. Coupled to cell movement, the flows transport myosin II from the front to the back of the cell, where the motor locally 'locks' actin in contractile bundles. This polarization mechanism could be employed by embryonic and cancer epithelial cells in microenvironments where high-contractility-driven cell motion is inefficient. PMID:26414403

  14. Platelet-derived growth factor and transforming growth factor-beta enhance tissue repair activities by unique mechanisms.

    PubMed

    Pierce, G F; Mustoe, T A; Lingelbach, J; Masakowski, V R; Griffin, G L; Senior, R M; Deuel, T F

    1989-07-01

    Platelet-derived growth factor (PDGF) and transforming growth factor-beta (TGF-beta) markedly potentiate tissue repair in vivo. In the present experiments, both in vitro and in vivo responses to PDGF and TGF-beta were tested to identify mechanisms whereby these growth factors might each enhance the wound-healing response. Recombinant human PDGF B-chain homodimers (PDGF-BB) and TGF-beta 1 had identical dose-response curves in chemotactic assays with monocytes and fibroblasts as the natural proteins from platelets. Single applications of PDGF-BB (2 micrograms, 80 pmol) and TGF-beta 1 (20 micrograms, 600 pmol) were next applied to linear incisions in rats and each enhanced the strength required to disrupt the wounds at 5 d up to 212% of paired control wounds. Histological analysis of treated wounds demonstrated an in vivo chemotactic response of macrophages and fibroblasts to both PDGF-BB and to TGF-beta 1 but the response to TGF-beta 1 was significantly less than that observed with PDGF-BB. Marked increases of procollagen type I were observed by immunohistochemical staining in fibroblasts in treated wounds during the first week. The augmented breaking strength of TGF-beta 1 was not observed 2 and 3 wk after wounding. However, the positive influence of PDGF-BB on wound breaking strength persisted through the 7 wk of testing. Furthermore, PDGF-BB-treated wounds had persistently increased numbers of fibroblasts and granulation tissue through day 21, whereas the enhanced cellular influx in TGF-beta 1-treated wounds was not detectable beyond day 7. Wound macrophages and fibroblasts from PDGF-BB-treated wounds contained sharply increased levels of immunohistochemically detectable intracellular TGF-beta. Furthermore, PDGF-BB in vitro induced a marked, time-dependent stimulation of TGF-beta mRNA levels in cultured normal rat kidney fibroblasts. The results suggest that TGF-beta transiently attracts fibroblasts into the wound and may stimulate collagen synthesis directly. In

  15. Smad-Independent Transforming Growth Factor-β Regulation of Early Growth Response-1 and Sustained Expression in Fibrosis

    PubMed Central

    Bhattacharyya, Swati; Chen, Shu-Jen; Wu, Minghua; Warner-Blankenship, Matthew; Ning, Hongyan; Lakos, Gabriella; Mori, Yasuji; Chang, Eric; Nihijima, Chihiro; Takehara, Kazuhiro; Feghali-Bostwick, Carol; Varga, John

    2008-01-01

    Transforming growth factor-β (TGF-β) plays a key role in scleroderma pathogenesis. The transcription factor early growth response-1 (Egr-1) mediates the stimulation of collagen transcription elicited by TGF-β and is necessary for the development of pulmonary fibrosis in mice. Here, we report that TGF-β causes a time- and dose-dependent increase in Egr-1 protein and mRNA levels and enhanced transcription of the Egr-1 gene via serum response elements in normal fibroblasts. The ability of TGF-β to stimulate Egr-1 was preserved in Smad3-null mice and in explanted Smad3-null fibroblasts. The response was blocked by a specific mitogen-activated protein kinase kinase 1 (MEK1) inhibitor but not by an ALK5 kinase inhibitor. Furthermore, MEK1 was phosphorylated by TGF-β, which was sufficient to drive Egr-1 transactivation. Stimulation by TGF-β enhanced the transcriptional activity of Elk-1 via the MEK-extracellular signal-regulated kinase 1/2 pathway. Bleomycin-induced scleroderma in the mouse was accompanied by increased Egr-1 accumulation in lesional fibroblasts. Furthermore, biopsies of lesional skin and lung from patients with scleroderma showed increased Egr-1 levels, which were highest in early diffuse disease. Moreover, both Egr-1 mRNA and protein were elevated in explanted scleroderma skin fibroblasts in vitro. Together, these findings define a Smad-independent TGF-β signal transduction mechanism that underlies the stimulation of Egr-1, demonstrate for the first time sustained Egr-1 up-regulation in fibrotic lesions and suggests that Egr-1 has a role in the induction and progression of fibrosis. PMID:18772333

  16. Actin-based phagosome motility.

    PubMed

    Zhang, Fangliang; Southwick, Frederick S; Purich, Daniel L

    2002-10-01

    Despite abundant evidence of actin's involvement at the particle internalization stage of phagocytosis, little is known about whether phagosomes undergo the same type of actin-based motility as observed with endocytic vesicles or such intracellular pathogens as Listeria and Shigella. By employing video microscopy to follow the fate of latex bead-containing phagosomes within the cytoplasm of bone marrow macrophages, we have made the novel observation of actin-based phagosome motility. Immunofluorescence microscopy confirmed that phagosomes containing IgG-opsonized, bovine serum albumin (or BSA) -coated or uncoated latex beads all formed actin-rich rocket tails that persisted only during a brief, 1-2 min period of actin-based motility. Average speeds of actin-based phagosome motility were 0.13 +/- 0.06 microm/s for IgG-coated beads, 0.14 +/- 0.04 microm/s for BSA-coated beads, and 0.11+/- 0.03 microm/s for uncoated beads. Moreover, the speeds and motile-phase duration of each type of phagosome were comparable to the behavior of pinosomes [Merrifield et al., 1999: Nat. Cell Biol. 1:72-74.]. Determination of optimal conditions for observing and analyzing actin-based phagosome motility should facilitate future investigations of phagocytosis and phagosome maturation. PMID:12211106

  17. Autocrine and exogenous transforming growth factor beta control cell cycle inhibition through pathways with different sensitivity.

    PubMed

    Wang, Jing; Sergina, Natalia; Ko, Tien C; Gong, Jiangeng; Brattain, Michael G

    2004-09-17

    Human colon carcinoma cells HCT116 that lack transforming growth factor beta (TGF-beta) type II receptor (RII) demonstrated restoration of autocrine TGF-beta activity upon reexpression of RII without restoring inhibitory responses to exogenous TGF-beta treatment. RII transfectants (designated RII Cl 37) had a longer lag phase relative to NEO-transfected control cells (designated NEO pool) before entering exponential growth in tissue culture. The prolonged growth arrest of RII Cl 37 cells was associated with markedly reduced cyclin-dependent kinase (CDK)2 activity. Our results demonstrate that p21 induction by autocrine TGF-beta is responsible for reduced CDK2 activity, which at least partially contributes to prolonged growth arrest and reduced cell proliferation in RII Cl 37 cells. In contrast to RII transfectants, HCT116 cells transfected with chromosome 3 (designated HCT116Ch3), which bears the RII gene, restored the response to exogenous TGF-beta as well as autocrine TGF-beta activity. Autocrine TGF-beta activity in HCT116Ch3 cells induced p21 expression as seen in RII Cl 37 cells; however, in addition to autocrine activity, HCT116Ch3 cells responded to exogenous TGF-beta as decreased CDK4 expression and reduced pRb phosphorylation mediated a TGF-beta inhibitory response in these cells. These results indicate that autocrine TGF-beta regulates the cell cycle through a pathway different from exogenous TGF-beta in the sense that p21 is a more sensitive effector of the TGF-beta signaling pathway, which can be induced and saturated by autocrine TGF-beta, whereas CDK4 inhibition is a less sensitive effector, which can only be activated by high levels of exogenous TGF-beta PMID:15271980

  18. Stromal inhibition of prostatic epithelial cell proliferation not mediated by transforming growth factor beta.

    PubMed Central

    Kooistra, A.; van den Eijnden-van Raaij, A. J.; Klaij, I. A.; Romijn, J. C.; Schröder, F. H.

    1995-01-01

    The paracrine influence of prostatic stroma on the proliferation of prostatic epithelial cells was investigated. Stromal cells from the human prostate have previously been shown to inhibit anchorage-dependent as well as anchorage-independent growth of the prostatic tumour epithelial cell lines PC-3 and LNCaP. Antiproliferative activity, mediated by a diffusible factor in the stromal cell conditioned medium, was found to be produced specifically by prostatic stromal cells. In the present study the characteristics of this factor were examined. It is demonstrated that prostate stroma-derived inhibiting factor is an acid- and heat-labile, dithiothreitol-sensitive protein. Although some similarities with type beta transforming growth factor (TGF-beta)-like inhibitors are apparent, evidence is presented that the factor is not identical to TGF-beta or to the TGF-beta-like factors activin and inhibin. Absence of TGF-beta activity was shown by the lack of inhibitory response of the TGF-beta-sensitive mink lung cell line CCL-64 to prostate stromal cell conditioned medium and to concentrated, partially purified preparations of the inhibitor. Furthermore, neutralising antibodies against TGF-beta 1 or TGF-beta 2 did not cause a decline in the level of PC-3 growth inhibition caused by partially purified inhibitor. Using Northern blot analyses, we excluded the involvement of inhibin or activin. It is concluded that the prostate stroma-derived factor may be a novel growth inhibitor different from any of the currently described inhibiting factors. Images Figure 5 PMID:7543773

  19. Interferon-alpha inhibits murine macrophage transforming growth factor-beta mRNA expression.

    PubMed

    Dhanani, S; Huang, M; Wang, J; Dubinett, S M

    1994-06-01

    Transforming growth factor-beta (TGF-beta), a multifunctional polypeptide is produced by a wide variety of cells and regulates a broad array of physiological and pathological functions. TGF-beta appears to play a central role in pulmonary fibrosis and may contribute to tumor-associated immunosuppression. Alveolar macrophages are a rich source of TGF-beta and are intimately involved in lung inflammation. We therefore chose to study TGF-beta regulation in murine alveolar macrophages as well as an immortalized peritoneal macrophage cell line (IC-21). Murine macrophages were incubated with cytokines to evaluate their role in regulating TGF-beta mRNA expression. We conclude that IFN-alpha downregulates TGF-beta mRNA expression in murine macrophages. PMID:8088926

  20. The pleiotropic roles of transforming growth factor beta inhomeostasis and carcinogenesis of endocrine organs.

    SciTech Connect

    Fleisch, Markus C.; Maxwell, Christopher A.; Barcellos-Hoff,Mary-Helen

    2006-01-13

    Transforming growth factor beta (TGF-beta) is a ubiquitous cytokine that plays a critical role in numerous pathways regulating cellular and tissue homeostasis. TGF-beta is regulated by hormones and is a primary mediator of hormone response in uterus, prostate and mammary gland. This review will address the role of TGF-beta in regulating hormone dependent proliferation and morphogenesis. The subversion of TGF-beta regulation during the processes of carcinogenesis, with particular emphasis on its effects on genetic stability and epithelial to mesenchymal transition (EMT), will also be examined. An understanding of the multiple and complex mechanisms of TGF-beta regulation of epithelial function, and the ultimate loss of TGF-beta function during carcinogenesis, will be critical in the design of novel therapeutic interventions for endocrine-related cancers.

  1. Role of transforming growth factor-beta (TGF) beta in the physiopathology of rheumatoid arthritis.

    PubMed

    Gonzalo-Gil, Elena; Galindo-Izquierdo, María

    2014-01-01

    Transforming growth factor-beta (TGF-β) is a cytokine with pleiotropic functions in hematopoiesis, angiogenesis, cell proliferation, differentiation, migration and apoptosis. Although its role in rheumatoid arthritis is not well defined, TGF-β activation leads to functional immunomodulatory effects according to environmental conditions. The function of TGF-β in the development of arthritis in murine models has been extensively studied with controversial results. Recent findings point to a non-relevant role for TGF-β in a mice model of collagen-induced arthritis. The study of TGF-β on T-cell responses has shown controversial results as an inhibitor or promoter of the inflammatory response. This paper presents a review of the role of TGF-β in animal models of arthritis. PMID:24685296

  2. 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

  3. Mechanistic basis and clinical relevance of the role of transforming growth factor-β in cancer

    PubMed Central

    Lin, Run-Long; Zhao, Lu-Jun

    2015-01-01

    Transforming growth factor-β (TGF-β) is a key factor in cancer development and progression. TGF-β can suppress tumorigenesis by inhibiting cell cycle progression and stimulating apoptosis in early stages of cancer progression. However, TGF-β can modulate cancer-related processes, such as cell invasion, distant metastasis, and microenvironment modification that may be used by cancer cells to their advantage in late stages. Corresponding mechanisms include angiogenesis promotion, anti-tumor immunity suppression, and epithelial-to-mesenchymal transition (EMT) induction. The correlation between TGF-β expression and cancer prognosis has also been extensively investigated. Results suggest that TGF-β pathway can be targeted to treat cancer; as such, the feasibility of this treatment is investigated in clinical trials. PMID:26779375

  4. Serum deprivation response inhibits breast cancer progression by blocking transforming growth factor-β signaling.

    PubMed

    Tian, Yao; Yu, Yue; Hou, Li-Kun; Chi, Jiang-Rui; Mao, Jie-Fei; Xia, Li; Wang, Xin; Wang, Ping; Cao, Xu-Chen

    2016-03-01

    Serum deprivation response (SDPR), a key substrate for protein kinase C, play a critical role in inducing membrane curvature and participate in the formation of caveolae. However, the function of SDPR in cancer development and progression is still not clear. Here, we found that SDPR is downregulated in human breast cancer. Overexpression of SDPR suppresses cell proliferation and invasion in MDA-MB-231 cells, while depletion of SDPR promotes cell proliferation and invasion in MCF10A cells. Subsequently, SDPR depletion induces epithelial-mesenchymal transition (EMT)-like phenotype. Finally, knockdown of SDPR activates transforming growth factor-β (TGF-β) signaling by upregulation of TGF-β1 expression. In conclusion, our results showed that SDPR inhibits breast cancer progression by blocking TGF-β signaling. Serum deprivation response suppresses cell proliferation and invasion in breast cancer cells. SDPR depletion induces epithelial-mesenchymal transition by activation of TGF-β signaling. PMID:26749136

  5. The Role of Transforming Growth Factor β1 in the Regulation of Blood Pressure

    PubMed Central

    Matsuki, Kota; Hathaway, Catherine K.; Lawrence, Marlon G.; Smithies, Oliver; Kakoki, Masao

    2016-01-01

    Although human association studies suggest a link between polymorphisms in the gene encoding transforming growth factor (TGF) β1 and differing blood pressure levels, a causative mechanism for this correlation remains elusive. Recently we have generated a series of mice with graded expression of TGFβ1, ranging from approximately 10% to 300% compared to normal. We have found that blood pressure and plasma volume are negatively regulated by TGFβ1. Of note, the 10% hypomorph exhibits primary aldosteronism and markedly impaired urinary excretion of water and electrolytes. We here review previous literature highlighting the importance of TGFβ signaling as a natriuretic system, which we postulate is a causative mechanism explaining how polymorphisms in TGFβ1 could influence blood pressure levels. PMID:25801626

  6. CYLD negatively regulates transforming growth factor-β-signalling via deubiquitinating Akt

    PubMed Central

    Lim, Jae Hyang; Jono, Hirofumi; Komatsu, Kensei; Woo, Chang-Hoon; Lee, Jiyun; Miyata, Masanori; Matsuno, Takashi; Xu, Xiangbin; Huang, Yuxian; Zhang, Wenhong; Park, Soo Hyun; Kim, Yu-Il; Choi, Yoo-Duk; Shen, Huahao; Heo, Kyung-Sun; Xu, Haodong; Bourne, Patricia; Koga, Tomoaki; Xu, Haidong; Yan, Chen; Wang, Binghe; Chen, Lin-Feng; Feng, Xin-Hua; Li, Jian-Dong

    2012-01-01

    Lung injury, whether induced by infection or caustic chemicals, initiates a series of complex wound-healing responses. If uncontrolled, these responses may lead to fibrotic lung diseases and loss of function. Thus, resolution of lung injury must be tightly regulated. The key regulatory proteins required for tightly controlling the resolution of lung injury have yet to be identified. Here we show that loss of deubiquitinase CYLD led to the development of lung fibrosis in mice after infection with Streptococcus pneumoniae. CYLD inhibited transforming growth factor-β-signalling and prevented lung fibrosis by decreasing the stability of Smad3 in an E3 ligase carboxy terminus of Hsc70-interacting protein-dependent manner. Moreover, CYLD decreases Smad3 stability by deubiquitinating K63-polyubiquitinated Akt. Together, our results unveil a role for CYLD in tightly regulating the resolution of lung injury and preventing fibrosis by deubiquitinating Akt. These studies may help develop new therapeutic strategies for preventing lung fibrosis. PMID:22491319

  7. 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.

  8. Enhanced jun gene expression is an early genomic response to transforming growth factor. beta. stimulation

    SciTech Connect

    Pertovaara, L.; Sistonen, L.; Keski-Oja, J.; Alitalo, K. ); Bos, T.J.; Vogt, P.K. . Dept. of Microbiology)

    1989-03-01

    Transforming growth factor {beta} (TGF{beta}) is a multifunctional polypeptide4 that regulates proliferation, differentiation, and other functions of many cell types. The pathway of TGF{beta} signal transduction in cells is unknown. The authors report here that an early effect of TGF{beta} is an enhancement of the expression of two genes encoding serum- and phorbol ester tumor promoter-regulated transcription factors: the junB gene and the c-jun proto-oncogene, respectively. This stimulation was observed in human lung adenocarcinoma A549 cells which were growth inhibited by TGF{beta}, AKR-2B mouse embryo fibroblasts which were growth stimulated by TGF{beta}, and K562 human erythroleukemia cells, which were not appreciably affected in their growth by TFG{beta}. The increase in jun mRNA occurred with picomolar TGF{beta} concentrations within 1 h of TGF{beta} stimulation, reached a peak between 1 and 5 h in different cells, and declined gradually to base-fine levels. This mRNA response was followed by a large increase in the biosynthesis of the c-jun protein (AP-1), as shown by metabolic labeling and immunoprecipitation analysis. However, differential and cell type-specific regulation appeared to determine the timing and magnitude of the response of each jun gene in a given cell. In AKR-2B and NIH 3T3 cells, only junB was induced by TGF{beta}, evidently in a protein synthesis-independent fashion. The junB response to TGF{beta} was maintained in c-Ha-ras and neu oncogene-transformed cells. Thus, one of the earliest genomic responses to TGF{beta} may involve nuclear signal transduction and amplification by the junB and c-jun transcription factors in concert with c-fos, which is also induced. The differential activation of the jun genes may explain some of the pleiotropic effects of TGF{beta}.

  9. Fullerenol Nanoparticles with Structural Activity Induce Variable Intracellular Actin Filament Morphologies.

    PubMed

    Jin, Junjiang; Dong, Ying; Wang, Ying; Xia, Lin; Gu, Weihong; Bai, Xue; Chang, Yanan; Zhang, Mingyi; Chen, Kui; Li, Juan; Zhao, Lina; Xing, Gengmei

    2016-06-01

    Fullerenol nanoparticles are promising for various biological applications; many studies have shown that they induce variable and diverse biological effects including side effects. Separation and purification of two fractions of fullerenols has demonstrated that they have varied chemical structures on the surfaces of their carbon cages. Actin is an important structural protein that is able to transform functional structures under varied physiological conditions. We assessed the abilities of the two fractions of fullerenols to attach to actin and induce variable morphological features in actin filament structures. Specifically the fullerenol fraction with a surface electric charge of -1.913 ± 0.008q (x10(-6) C) has percentages of C-OH and C=O on the carbon cage of 16.14 ± 0.60 and 17.55 ± 0.69. These features allow it to form intermolecular hydrogen bonds with actin at a stoichiometric ratio of four fullerenols per actin subunit. Molecular simulations revealed these specific binding sites and binding modes in atomic details in the interaction between the active fullerenol and actin filament. Conversely, these interactions were not possible for the other fraction of fullerenol with that percentages of C-OH and C=O on the carbon cage were 15.59 ± 0.01 and 1.94 ± 0.11. Neither sample induced appreciable cytotoxicity or acute cell death. After entering cells, active fullerenol binding to actin induces variable morphological features and may transform ATP-actin to ADP-actin. These changes facilitate the binding of ADF/cofilin, allowing cofilin to sever actin filaments to form cofilin/actin/fullerenol rods. Our findings suggest that fullerenol with structural activity binding disturbs actin filament structure, which may inhibit locomotion of cell or induce chronic side effects in to cells. PMID:27319217

  10. Roles of Asp179 and Glu270 in ADP-Ribosylation of Actin by Clostridium perfringens Iota Toxin

    PubMed Central

    Belyy, Alexander; Tabakova, Irina; Lang, Alexander E.; Jank, Thomas; Belyi, Yury; Aktories, Klaus

    2015-01-01

    Clostridium perfringens iota toxin is a binary toxin composed of the enzymatically active component Ia and receptor binding component Ib. Ia is an ADP-ribosyltransferase, which modifies Arg177 of actin. The previously determined crystal structure of the actin-Ia complex suggested involvement of Asp179 of actin in the ADP-ribosylation reaction. To gain more insights into the structural requirements of actin to serve as a substrate for toxin-catalyzed ADP-ribosylation, we engineered Saccharomyces cerevisiae strains, in which wild type actin was replaced by actin variants with substitutions in residues located on the Ia-actin interface. Expression of the actin mutant Arg177Lys resulted in complete resistance towards Ia. Actin mutation of Asp179 did not change Ia-induced ADP-ribosylation and growth inhibition of S. cerevisiae. By contrast, substitution of Glu270 of actin inhibited the toxic action of Ia and the ADP-ribosylation of actin. In vitro transcribed/translated human β-actin confirmed the crucial role of Glu270 in ADP-ribosylation of actin by Ia. PMID:26713879

  11. Transforming Growth Factor-Beta and Oxidative Stress Interplay: Implications in Tumorigenesis and Cancer Progression

    PubMed Central

    Krstić, Jelena; Trivanović, Drenka; Mojsilović, Slavko; Santibanez, Juan F.

    2015-01-01

    Transforming growth factor-beta (TGF-β) and oxidative stress/Reactive Oxygen Species (ROS) both have pivotal roles in health and disease. In this review we are analyzing the interplay between TGF-β and ROS in tumorigenesis and cancer progression. They have contradictory roles in cancer progression since both can have antitumor effects, through the induction of cell death, senescence and cell cycle arrest, and protumor effects by contributing to cancer cell spreading, proliferation, survival, and metastasis. TGF-β can control ROS production directly or by downregulating antioxidative systems. Meanwhile, ROS can influence TGF-β signaling and increase its expression as well as its activation from the latent complex. This way, both are building a strong interplay which can be taken as an advantage by cancer cells in order to increment their malignancy. In addition, both TGF-β and ROS are able to induce cell senescence, which in one way protects damaged cells from neoplastic transformation but also may collaborate in cancer progression. The mutual collaboration of TGF-β and ROS in tumorigenesis is highly complex, and, due to their differential roles in tumor progression, careful consideration should be taken when thinking of combinatorial targeting in cancer therapies. PMID:26078812

  12. Transforming growth factor-alpha promotes mammary tumorigenesis through selective survival and growth of secretory epithelial cells.

    PubMed Central

    Smith, G. H.; Sharp, R.; Kordon, E. C.; Jhappan, C.; Merlino, G.

    1995-01-01

    Transforming growth factor (TGF)-alpha stimulates the growth and development of mammary epithelial cells and is implicated in the pathogenesis of human breast cancer. In this report we evaluate the consequences of overexpressing TGF-alpha in the mammary gland of transgenic mice and examine associated cellular mechanisms. When operating on a FVB/N genetic background (line MT100), TGF-alpha induced the stochastic development of mammary adenomas and adenocarcinomas f secretory epithelial origin in 64% of multiparous females. In contrast, tumors were exceedingly rare in virgin MT100 females, MT100 males, and multiparous FVB/N females. In MT100 females multiple foci of hyperplastic secretory lesions preceded the development of frank tumors; these initial lesions appeared during the involution period after the first lactation. Serial transplantation of these hyperplasias indicated an absence of proliferative immortality. Nevertheless, they gave rise to tumors at a low frequency and after a prolonged latency in virgin hosts; in multiparous hosts, tumors developed earlier and at a high incidence. The TGF-alpha transgene was highly expressed in hyperplasias and tumors but not in virgin and nonlesion-bearing tissue, suggesting that TGF-alpha overexpression provides a selective growth advantage. TGF-alpha also induced at lactation a 6.4-fold increase in DNA synthesis in MT100 epithelial cells, many of which were binucleated. MT100 mammary tissue experienced an obvious delay in involution, resulting in the postlactational survival of a significant population of unregressed secretory epithelial cells. In contrast, another line of transgenic mice on a CD-1 genetic background (MT42), in which TGF-alpha overexpression induced liver but not mammary tumors, failed to demonstrate postlactational epithelial cell survival. These data show that TGF-alpha promotes mammary tumorigenesis in multiparous MT100 mice by stimulating secretory epithelial cell proliferation during lactation and

  13. Formin' actin in the nucleus.

    PubMed

    Baarlink, Christian; Grosse, Robert

    2014-01-01

    Many if not most proteins can, under certain conditions, change cellular compartments, such as, for example, shuttling from the cytoplasm to the nucleus. Thus, many proteins may exert functions in various and very different subcellular locations, depending on the signaling context. A large amount of actin regulatory proteins has been detected in the mammalian cell nucleus, although their potential roles are much debated and are just beginning to emerge. Recently, members of the formin family of actin nucleators were also reported to dynamically localize to the nuclear environment. Here we discuss our findings that specific diaphanous-related formins can promote nuclear actin assembly in a signal-dependent manner. PMID:24637338

  14. Bacterial actins and their diversity

    PubMed Central

    Ozyamak, Ertan; Kollman, Justin M.; Komeili, Arash

    2015-01-01

    For many years bacteria were considered rather simple organisms, but the dogmatic notion that subcellular organization is a eukaryotic trait has been overthrown for more than a decade. The discovery of homologs of the eukaryotic cytoskeletal proteins actin, tubulin, and intermediate filaments in bacteria has been instrumental in changing this view. Over the recent years we gained an incredible level of insight into the diverse family of bacterial actins and their molecular workings. Here we review the functional, biochemical and structural features of the most well-studied bacterial actins. PMID:24015924

  15. Expression and localization of epidermal growth factor, transforming growth factor-α and epidermal growth factor receptor in the canine testis

    PubMed Central

    TAMADA, Hiromichi; TAKEMOTO, Kohei; TOMINAGA, Masato; KAWATE, Noritoshi; TAKAHASHI, Masahiro; HATOYA, Shingo; MATSUYAMA, Satoshi; INABA, Toshio; SAWADA, Tsutomu

    2015-01-01

    Gene expression of epidermal growth factor (EGF), transforming growth factor-α (TGF-α) and EGF receptor (EGF-R) and the localization of the corresponding proteins in the canine testis were studied. Levels of mRNA expressions were determined by semiquantitative reverse transcription polymerase chain reaction in the testes of the peripubertal (4–6 months), young adult (3–4 years), advanced adult (7–8 years) and senescent (11–16 years) groups. The EGF-R mRNA level in the testes of the peripubertal group was significantly higher than those in the other groups, whereas there was no difference in EGF and TGF-α mRNA levels among groups. Immunohistochemical stainings for EGF, TGF-α and EGF-R in the testis revealed that immunoreactivity in the seminiferous epithelium and Sertoli cell was weak and nonspecific for the stage of spermatogenesis, and distinct staining was found in Leydig cells. These results suggest that the EGF family of growth factors may be involved in testicular maturation and function in the dog. PMID:26498203

  16. Interrelationship of in situ growth stress evolution and phase transformations in Ti/W multilayered thin films

    NASA Astrophysics Data System (ADS)

    Wan, Li; Yu, Xiao-xiang; Zhou, Xuyang; Thompson, Gregory

    2016-06-01

    This paper addresses the in situ growth stress evolution and phase transformation of bcc to hcp Ti in Ti/W multilayered thin films. A series of equal layer thicknesses from 20 nm to 1 nm were deposited. As the bilayer thickness reduced, the overall film stress became less compressive until the Ti transformed from hcp (at the larger layer thicknesses) to bcc in the 1 nm/1 nm multilayer. The pseudomorphic bcc stabilization resulted in a recovery of the compressive stress to values near that for the bulk phase stabilized for the 5 nm/5 nm multilayer. A discernable change in stress slope was noted for the bcc to hcp Ti transition as a function of Ti layer thickness. The stress states for each film, during film growth, are rationalized by the lattice matching of the phase with the growth surface. These results are coupled to a molecular dynamics deposition simulation which revealed good agreement with the experimentally observed transformation thickness.

  17. A Histologically Distinctive Interstitial Pneumonia Induced by Overexpression of the Interleukin 6, Transforming Growth Factor β1, or Platelet-Derived Growth Factor B Gene

    NASA Astrophysics Data System (ADS)

    Yoshida, Mitsuhiro; Sakuma, Junko; Hayashi, Seiji; Abe, Kin'ya; Saito, Izumu; Harada, Shizuko; Sakatani, Mitsunoir; Yamamoto, Satoru; Matsumoto, Norinao; Kaneda, Yasufumi; Kishmoto, Tadamitsu

    1995-10-01

    Interstitial pneumonia is characterized by alveolitis with resulting fibrosis of the interstitium. To determine the relevance of humoral factors in the pathogenesis of interstitial pneumonia, we introduced expression vectors into Wistar rats via the trachea to locally overexpress humoral factors in the lungs. Human interleukin (IL) 6 and IL-6 receptor genes induced lymphocytic alveolitis without marked fibroblast proliferation. In contrast, overexpression of human transforming growth factor β1 or human platelet-derived growth factor B gene induced only mild or apparent cellular infiltration in the alveoli, respectively. However, both factors induced significant proliferation of fibroblasts and deposition of collagen fibrils. These histopathologic changes induced by the transforming growth factor β1 and platelet-derived growth factor B gene are partly akin to those changes seen in lung tissues from patients with pulmonary fibrosis and markedly contrast with the changes induced by overexpression of the IL-6 and IL-6 receptor genes that mimics lymphocytic interstitial pneumonia.

  18. Transforming Growth Factor-β1 Antagonizes the Transcription, Expression, and Vascular Signaling of Guanylyl Cyclase/Natriuretic Peptide Receptor A: Role of δEF1

    PubMed Central

    Sen, Anagha; Kumar, Prerna; Garg, Renu; Lindsey, Sarah H.; Katakam, Prasad V.G.; Bloodworth, Meaghan; Pandey, Kailash N.

    2016-01-01

    The objective of this study was to determine the role of transforming growth factor-beta 1 (TGF-β1) in transcriptional regulation and function of guanylyl cyclase-A/natriuretic peptide receptor-A (GC-A/NPRA) gene (Npr1) and whether a cross-talk exists between these two hormonal systems in target cells. After treatments of primary cultured rat thoracic aortic vascular smooth muscle cells (RTASMCs) and mouse mesangial cells (MMCs) with TGF-β1, the Npr1 promoter construct embodying delta-crystallin enhancer binding factor 1 (δEF1) site showed 85% reduction in luciferase activity in a time- and dose-dependent manner. TGF-β1 also significantly attenuated luciferase activity of Npr1 promoter by 62% and decreased the ANP-mediated relaxation of mouse denuded aortic rings ex vivo. Treatment of cells with TGF-β1, stimulated the protein levels of δEF1 by 2.4- to 2.8-fold and also significantly enhanced the phosphorylation of Smad 2/3; however, markedly reduced Npr1 mRNA and receptor protein levels. Overexpression of δEF1 showed a reduction in Npr1 promoter activity by 75% while the deletion or site-directed mutagenesis of δEF1 sites in Npr1 promoter, eliminated the TGF-β1-mediated repression of Npr1 transcription. TGF-β1 significantly increased the expression of α-smooth muscle actin and collagen type 1 alpha 2 in RTASMCs, which were markedly attenuated by ANP in NPRA overexpressing cells. Together, the present results suggest that an antagonistic cascade exists between TGF-β1/Smad/δEF1 pathways and Npr1 expression and receptor signaling relevant to renal and vascular remodeling, which might be critical in the regulation of blood pressure and cardiovascular homeostasis. PMID:26934489

  19. Lactic Acid is Elevated in Idiopathic Pulmonary Fibrosis and Induces Myofibroblast Differentiation Via pH-Dependent Activation of Transforming Growth Factor-β

    SciTech Connect

    Kottman, R. M.; Kulkarni, Ajit A.; Smolnycki, Katie A.; Lyda, Elizabeth; Dahanayake, Thinesh; Salibi, Rami; Honnons, Sylvie; Jones, Carolyn; Isern, Nancy G.; Hu, Jian Z.; Nathan, Steven D.; Grant, Geraldine; Phipps, Richard P.; Sime, Patricia J.

    2012-10-15

    Rationale: Idiopathic pulmonary fibrosis (IPF) is a complex disease for which the pathogenesis is poorly understood. In this study, we identified lactic acid as a metabolite that is elevated in the lung tissue of patients with IPF. Objectives: This study examines the effect of lactic acid on myofibroblast differentiation and pulmonary fibrosis. Methods:We used metabolomic analysis to examine cellular metabolism in lung tissuefrom patients with IPFanddeterminedthe effects of lactic acid and lactate dehydrogenase-5 (LDH5) overexpression on myofibroblast differentiation and transforming growth factor (TGF)-b activation in vitro. Measurements and Main Results: Lactic acid concentrations from healthy and IPF lung tissue were determined by nuclear magnetic resonance spectroscopy; a-smooth muscle actin, calponin, and LDH5 expression were assessed by Western blot of cell culture lysates. Lactic acid and LDH5 were significantly elevated in IPF lung tissue compared with controls. Physiologic concentrations of lactic acid induced myofibroblast differentiation via activation of TGF-b. TGF-b induced expression of LDH5 via hypoxia-inducible factor 1a (HIF1a). Importantly, overexpression of both HIF1a and LDH5 in human lung fibroblasts induced myofibroblast differentiation and synergized with low dose TGF-b to induce differentiation. Furthermore, inhibition of both HIF1a and LDH5 inhibited TGF-b–induced myofibroblast differentiation. Conclusions: We have identified the metabolite lactic acid as an important mediator of myofibroblast differentiation via a pHdependent activation of TGF-b. We propose that the metabolic milieu of the lung, and potentially other tissues, is an important driving force behind myofibroblast differentiation and potentially the initiation and progression of fibrotic disorders.

  20. Thalidomide Accelerates the Degradation of Extracellular Matrix in Rat Hepatic Cirrhosis via Down-Regulation of Transforming Growth Factor-β1

    PubMed Central

    Meng, Qingshun; Liu, Jie; Wang, Chuanfang

    2015-01-01

    Purpose The degradation of the extracellular matrix has been shown to play an important role in the treatment of hepatic cirrhosis. In this study, the effect of thalidomide on the degradation of extracellular matrix was evaluated in a rat model of hepatic cirrhosis. Materials and Methods Cirrhosis was induced in Wistar rats by intraperitoneal injection of carbon tetrachloride (CCl4) three times weekly for 8 weeks. Then CCl4 was discontinued and thalidomide (100 mg/kg) or its vehicle was administered daily by gavage for 6 weeks. Serum hyaluronic acid, laminin, procollagen type III, and collagen type IV were examined by using a radioimmunoassay. Matrix metalloproteinase-13 (MMP-13), tissue inhibitor of metalloproteinase-1 (TIMP-1), and α-smooth muscle actin (α-SMA) protein in the liver, transforming growth factor β1 (TGF-β1) protein in cytoplasm by using immunohistochemistry and Western blot analysis, and MMP-13, TIMP-1, and TGF-β1 mRNA levels in the liver were studied using reverse transcriptase polymerase chain reaction. Results Liver histopathology was significantly better in rats given thalidomide than in the untreated model group. The levels of TIMP-1 and TGF-β1 mRNA and protein expressions were decreased significantly and MMP-13 mRNA and protein in the liver were significantly elevated in the thalidomide-treated group. Conclusion Thalidomide may exert its effects on the regulation of MMP-13 and TIMP-1 via inhibition of the TGF-β1 signaling pathway, which enhances the degradation of extracellular matrix and accelerates the regression of hepatic cirrhosis in rats. PMID:26446639

  1. Sulforaphane attenuates hepatic fibrosis via NF-E2-related factor 2-mediated inhibition of transforming growth factor-β/Smad signaling.

    PubMed

    Oh, Chang Joo; Kim, Joon-Young; Min, Ae-Kyung; Park, Keun-Gyu; Harris, Robert A; Kim, Han-Jong; Lee, In-Kyu

    2012-02-01

    Sulforaphane (SFN) is a dietary isothiocyanate that exerts chemopreventive effects via NF-E2-related factor 2 (Nrf2)-mediated induction of antioxidant/phase II enzymes, such as heme oxygenase-1 (HO-1) and NAD(P)H quinone oxidoreductase 1 (NQO1). This work was undertaken to evaluate the effects of SFN on hepatic fibrosis and profibrotic transforming growth factor (TGF)-β/Smad signaling, which are closely associated with oxidative stress. SFN suppressed TGF-β-enhanced expression of α-smooth muscle actin (α-SMA), a marker of hepatic stellate cell (HSC) activation, and profibrogenic genes such as type I collagen, fibronectin, tissue inhibitor of matrix metalloproteinase (TIMP)-1, and plasminogen activator inhibitor (PAI)-1 in hTERT, an immortalized human HSC line. SFN inhibited TGF-β-stimulated activity of a PAI-1 promoter construct and (CAGA)(9) MLP-Luc, an artificial Smad3/4-specific reporter, in addition to reducing phosphorylation and nuclear translocation of Smad3. Nrf2 overexpression was sufficient to inhibit the TGF-β/Smad signaling and PAI-1 expression. Conversely, knockdown of Nrf2, but not inhibition of HO-1 or NQO1 activity, significantly abolished the inhibitory effect of SFN on (CAGA)(9) MLP-Luc activity. However, inhibition of NQO1 activity reversed repression of TGF-β-stimulated expression of type I collagen by SFN, suggesting the involvement of antioxidant activity of SFN in the suppression of Smad-independent fibrogenic gene expression. Finally, SFN treatment attenuated the development and progression of early stage hepatic fibrosis induced by bile duct ligation in mice, accompanied by reduced expression of type I collagen and α-SMA. Collectively, these results show that SFN elicits an antifibrotic effect on hepatic fibrosis through Nrf2-mediated inhibition of the TGF-β/Smad signaling and subsequent suppression of HSC activation and fibrogenic gene expression. PMID:22155056

  2. High-Mobility Group Box 1 Mediates Epithelial-to-Mesenchymal Transition in Pulmonary Fibrosis Involving Transforming Growth Factor-β1/Smad2/3 Signaling.

    PubMed

    Li, Liu-Cheng; Li, De-Lin; Xu, Liang; Mo, Xiao-Ting; Cui, Wen-Hui; Zhao, Ping; Zhou, Wen-Cheng; Gao, Jian; Li, Jun

    2015-09-01

    Epithelial-to-mesenchymal transition (EMT) is a crucial event in the cellular origin of myofibroblasts that secrete extracellular matrix in the progression of pulmonary fibrosis (PF). High-mobility group box 1 (HMGB1) is a novel mediator of EMT. However, whether this process involves the recognized transforming growth factor-β1 (TGF-β1)/Smad signaling that also contributes to EMT in PF has not yet been elucidated. Here, we developed a model of PF induced by bleomycin (BLM) in rats and conducted several simulation experiments in A549 (human) and RLE-6TN (rat) alveolar epithelial cell (AEC) lines to unravel the role of TGF-β1/Smad2/3 signaling in HMGB1-mediated EMT. We found that the levels of serum HMGB1 and lung hydroxyproline were severely elevated after BLM administration. Moreover, the protein expression of HMGB1, TGF-β1, phosphorylated Smad2/3 (p-Smad2/3), and mesenchymal markers including α-smooth muscle actin, vimentin, and type I collagen were significantly increased with the reduced protein expression of an epithelial marker (E-cadherin) in the rat model by Western blot or immunohistochemical analysis. In addition, the uptake of both exogenous TGF-β1 and HMGB1 by AECs could induce EMT; meanwhile, HMGB1 dramatically enhanced TGF-β1 expression and triggered Smad2/3 phosphorylation. In contrast, TGF-β1 deficiency evidently ameliorated HMGB1-mediated EMT with reduced p-Smad2/3 in A549 cells. It provides new insights that HMGB1 release from injured lungs promotes AEC damage through induction of the EMT process, in which TGF-β1/Smad2/3 signaling is activated and contributes to PF. These results suggest that HMGB1 may constitute a therapeutic target for developing antifibrotic agents for abnormal lung remodeling. PMID:26126535

  3. Induction of epithelial-mesenchymal transition in alveolar epithelial cells by transforming growth factor-beta1: potential role in idiopathic pulmonary fibrosis.

    PubMed

    Willis, Brigham C; Liebler, Janice M; Luby-Phelps, Katherine; Nicholson, Andrew G; Crandall, Edward D; du Bois, Roland M; Borok, Zea

    2005-05-01

    The hallmark of idiopathic pulmonary fibrosis (IPF) is the myofibroblast, the cellular origin of which in the lung is unknown. We hypothesized that alveolar epithelial cells (AECs) may serve as a source of myofibroblasts through epithelial-mesenchymal transition (EMT). Effects of chronic exposure to transforming growth factor (TGF)-beta1 on the phenotype of isolated rat AECs in primary culture and a rat type II cell line (RLE-6TN) were evaluated. Additionally, tissue samples from patients with IPF were evaluated for cells co-expressing epithelial (thyroid transcription factor (TTF)-1 and pro-surfactant protein-B (pro-SP-B), and mesenchymal (alpha-smooth muscle actin (alpha-SMA)) markers. RLE-6TN cells exposed to TGF-beta1 for 6 days demonstrated increased expression of mesenchymal cell markers and a fibroblast-like morphology, an effect augmented by tumor necrosis factor-alpha (TNF-alpha). Exposure of rat AECs to TGF-beta1 (100 pmol/L) resulted in increased expression of alpha-SMA, type I collagen, vimentin, and desmin, with concurrent transition to a fibroblast-like morphology and decreased expression of TTF-1, aquaporin-5 (AQP5), zonula occludens-1 (ZO-1), and cytokeratins. Cells co-expressing epithelial markers and alpha-SMA were abundant in lung tissue from IPF patients. These results suggest that AECs undergo EMT when chronically exposed to TGF-beta1, raising the possibility that epithelial cells may serve as a novel source of myofibroblasts in IPF. PMID:15855634

  4. Inhibition of liver fibrosis by solubilized coenzyme Q10: Role of Nrf2 activation in inhibiting transforming growth factor-beta1 expression

    SciTech Connect

    Choi, Hoo-Kyun; Pokharel, Yuba Raj; Lim, Sung Chul; Han, Hyo-Kyung; Ryu, Chang Seon; Kim, Sang Kyum; Kwak, Mi Kyong; Kang, Keon Wook

    2009-11-01

    Coenzyme Q10 (CoQ10), an endogenous antioxidant, is important in oxidative phosphorylation in mitochondria. It has anti-diabetic and anti-cardiovascular disease effects, but its ability to protect against liver fibrosis has not been studied. Here, we assessed the ability of solubilized CoQ10 to improve dimethylnitrosamine (DMN)-induced liver fibrogenesis in mice. DMN treatments for 3 weeks produced a marked liver fibrosis as assessed by histopathological examination and tissue 4-hydroxyproline content. Solubilized CoQ10 (10 and 30 mg/kg) significantly inhibited both the increases in fibrosis score and 4-hydroxyproline content induced by DMN. Reverse transcription-polymerase chain reaction and Western blot analyses revealed that solubilized CoQ10 inhibited increases in the transforming growth factor-beta1 (TGF-beta1) mRNA and alpha-smooth muscle actin (alpha-SMA) protein by DMN. Interestingly, hepatic glutamate-cysteine ligase (GCL) and glutathione S-transferase A2 (GSTA2) were up-regulated in mice treated with CoQ10. Solubilized CoQ10 also up-regulated antioxidant enzymes such as catalytic subunits of GCL and GSTA2 via activating NF-E2 related factor2 (Nrf2)/antioxidant response element (ARE) in H4IIE hepatoma cells. Moreover, CoQ10's inhibition of alpha-SMA and TGF-beta1 expressions disappeared in Nrf2-null MEF cells. In contrast, Nrf2 overexpression significantly decreased the basal expression levels of alpha-SMA and TGF-beta1 in Nrf2-null MEF cells. These results demonstrated that solubilized CoQ10 inhibited DMN-induced liver fibrosis through suppression of TGF-beta1 expression via Nrf2/ARE activation.

  5. Essential Roles of RNA-binding Protein HuR in Activation of Hepatic Stellate Cells Induced by Transforming Growth Factor-β1

    PubMed Central

    Ge, Jingjing; Chang, Na; Zhao, Zhongxin; Tian, Lei; Duan, Xianghui; Yang, Lin; Li, Liying

    2016-01-01

    RNA-binding protein HuR mediates transforming growth factor (TGF)-β1-induced profibrogenic actions. Up-regulation of Sphingosine kinase 1 (SphK1) is involved in TGF-β1-induced activation of hepatic stellate cells (HSCs) in liver fibrogenesis. However, the molecular mechanism of TGF-β1 regulates SphK1 remains unclear. This study was designed to investigate the role of HuR in TGF-β1-induced SphK1 expression and identify a new molecular mechanism in liver fibrogenensis. In vivo, HuR expression was increased, translocated to cytoplasm, and bound to SphK1 mRNA in carbon tetrachloride- and bile duct ligation-induced mouse fibrotic liver. HuR mRNA expression had a positive correlation with mRNA expressions of SphK1 and fibrotic markers, α-smooth muscle actin (α-SMA) and Collagen α1(I), respectively. In vitro, up-regulation of SphK1 and activation of HSCs stimulated by TGF-β1 depended on HuR cytoplasmic accumulation. The effects of TGF-β1 were diminished when HuR was silenced or HuR cytoplasmic translocation was blocked. Meanwhile, overexpression of HuR mimicked the effects of TGF-β1. Furthermore, TGF-β1 prolonged half-life of SphK1 mRNA by promoting its binding to HuR. Pharmacological or siRNA-induced SphK1 inhibition abrogated HuR-mediated HSC activation. In conclusion, our data suggested that HuR bound to SphK1 mRNA and played a crucial role in TGF-β1-induced HSC activation. PMID:26912347

  6. Attenuation of Tubular Injury and Renal Fibrosis by TI-HU-YIN via Reduction in Transforming Growth Factor-β1 Expression in Unilateral Ureteral Obstruction Mice.

    PubMed

    Tarng, Der-Cherng; Liu, I-Shan; Lin, Lie-Chwen; Chen, Nien-Jung

    2015-12-31

    TI-HU-YIN (JCKD), a compound composed of many Chinese herbs, is hypothesized to attenuate renal tubular injury and interstitial fibrosis. Moreover, its renoprotective effects were assessed in animal and in vitro studies. First, male C57BL/6 mice were under sham operation or unilateral ureteral obstruction (UUO) surgery, and then treated with phosphate buffer solution (PBS), aliskirin and valsartan (A+V), and JCKD for 14 days. At 7 and 14 days, mice were sacrificed and the kidney tissues were assessed for histopathological changes and transforming growth factor (TGF)-β1 expression. As compared to sham group, UUO-PBS group had more serious tubular dilatation and injury, α-smooth muscle actin-positive areas, F4/80-positive macrophages, and interstitial fibrosis. Impressively, these pathologic changes were significantly attenuated in UUO mice both treated with JCKD and A+V as compared to UUO-PBS group. At 14 days, TGF-β1 expression was significantly suppressed in kidney tissues of UUO-JCKD group as well as in UUO-A+V group. Second, TGF-β1 production was increased in macrophage J774 cells and NRK-52E proximal tubular cells stimulated by angiotensin (Ang)-II at 10 nM for 24 h and at 1 nM for 48 h, respectively. JCKD (≥ 400 μg/ml) inhibited the TGF-β1 production at baseline and stimulated by Ang II in both cell lines. Our study showed that JCKD reduced renal injury, macrophage infiltration and interstitial fibrosis possibly through suppressing the TGF-β1 expression in UUO mice. Accordingly, JCKD is potential to retard the progression of chronic kidney disease. Further studies are needed to validate its renoprotective effects in the inhibition of TGF-β1 expression and the amelioration of renal fibrosis. PMID:26717915

  7. Transforming growth factor beta 3 involved in the pathogenesis of synovial chondromatosis of temporomandibular joint

    PubMed Central

    Li, Yingjie; El.Mozen, Loaye Abdelaziz; Cai, Hengxing; Fang, Wei; Meng, Qinggong; Li, Jian; Deng, Mohong; Long, Xing

    2015-01-01

    Synovial chondromatosis (SC) of temporomandibular joint is rare proliferative disorder featured by the formation of cartilaginous nodules in synovium and joint space. Transforming growth factor beta 3 (TGF-β3) is closely related to chondrogenic differentiation, and might participate in pathogenesis of SC. We discovered that increased quantity of synoviocytes and blood vessels were observed in SC synovium. The vessel wall and sublining fibroblasts were stained positively by the antibodies against TGF-β3, fibroblast growth factor 2 (FGF-2), and CD34. In loose bodies (LBs), TGF-β3 was mainly expressed in chondrocytes and FGF-2 was expressed in chondrocytes, fibroblasts, and vessel walls. Expressions of TGF-β1, TGF-β3, FGF-2, Sox9, Wnt-4, Foxc2, and VEGF-A mRNA were significantly higher in SC synovium. Stimulation of TGF-β3 on synoviocytes increased alkaline phosphatase (ALP) activity and expressions of chondrogenic genes (Sox9, Col2α1, Aggrecan, Wnt-4, and Wnt-11), osteogenic genes (Runx2, Foxc2, osteocalcin, and Col1α1), and VEGF-A, but failed to influence FGF-2 expression. However, the addition of FGF-2 increased TGF-β3 expression. In conclusion, TGF-β3 existed in synovium and LBs of SC, and was responsible for the pathogenesis of SC. PMID:25742744

  8. Transforming growth factor beta 3 involved in the pathogenesis of synovial chondromatosis of temporomandibular joint.

    PubMed

    Li, Yingjie; El Mozen, Loaye Abdelaziz; Cai, Hengxing; Fang, Wei; Meng, Qinggong; Li, Jian; Deng, Mohong; Long, Xing

    2015-01-01

    Synovial chondromatosis (SC) of temporomandibular joint is rare proliferative disorder featured by the formation of cartilaginous nodules in synovium and joint space. Transforming growth factor beta 3 (TGF-β3) is closely related to chondrogenic differentiation, and might participate in pathogenesis of SC. We discovered that increased quantity of synoviocytes and blood vessels were observed in SC synovium. The vessel wall and sublining fibroblasts were stained positively by the antibodies against TGF-β3, fibroblast growth factor 2 (FGF-2), and CD34. In loose bodies (LBs), TGF-β3 was mainly expressed in chondrocytes and FGF-2 was expressed in chondrocytes, fibroblasts, and vessel walls. Expressions of TGF-β1, TGF-β3, FGF-2, Sox9, Wnt-4, Foxc2, and VEGF-A mRNA were significantly higher in SC synovium. Stimulation of TGF-β3 on synoviocytes increased alkaline phosphatase (ALP) activity and expressions of chondrogenic genes (Sox9, Col2α1, Aggrecan, Wnt-4, and Wnt-11), osteogenic genes (Runx2, Foxc2, osteocalcin, and Col1α1), and VEGF-A, but failed to influence FGF-2 expression. However, the addition of FGF-2 increased TGF-β3 expression. In conclusion, TGF-β3 existed in synovium and LBs of SC, and was responsible for the pathogenesis of SC. PMID:25742744

  9. Tumor-promoting functions of transforming growth factor-β in progression of cancer

    PubMed Central

    2012-01-01

    Transforming growth factor-β (TGF-β) elicits both tumor-suppressive and tumor-promoting functions during cancer progression. Here, we describe the tumor-promoting functions of TGF-β and how these functions play a role in cancer progression. Normal epithelial cells undergo epithelial-mesenchymal transition (EMT) through the action of TGF-β, while treatment with TGF-β and fibroblast growth factor (FGF)-2 results in transdifferentiation into activated fibroblastic cells that are highly migratory, thereby facilitating cancer invasion and metastasis. TGF-β also induces EMT in tumor cells, which can be regulated by oncogenic and anti-oncogenic signals. In addition to EMT promotion, invasion and metastasis of cancer are facilitated by TGF-β through other mechanisms, such as regulation of cell survival, angiogenesis, and vascular integrity, and interaction with the tumor microenvironment. TGF-β also plays a critical role in regulating the cancer-initiating properties of certain types of cells, including glioma-initiating cells. These findings thus may be useful for establishing treatment strategies for advanced cancer by inhibiting TGF-β signaling. PMID:22111550

  10. alpha1B-Adrenergic receptor phosphorylation and desensitization induced by transforming growth factor-beta.

    PubMed Central

    Romero-Avila, M Teresa; Flores-Jasso, C Fabián; García-Sáinz, J Adolfo

    2002-01-01

    Transforming growth factor-beta (TGF-beta) induced alpha(1B)-adrenergic receptor phosphorylation in Rat-1 fibroblasts stably expressing these adrenoceptors. This effect of TGF-beta was rapid, reaching a maximum within 30 min and decreasing thereafter, and concentration-dependent (EC(50) 0.3 pM). The phosphoinositide 3-kinase inhibitors wortmannin and LY294002, and the protein kinase C inhibitors staurosporine, Ro 318220 and bisindolylmaleimide, blocked the effect of this growth factor. alpha(1B)-Adrenergic receptor phosphorylation was associated with desensitization, as indicated by a reduction in the adrenergic-mediated production of [(3)H]inositol phosphates. Phosphorylation of alpha(1B)-adrenergic receptors by TGF-beta was also observed in Cos-1 cells transfected with the receptor. Co-transfection of the dominant-negative mutant of the regulatory subunit of phosphoinositide 3-kinase (Deltap85) inhibited the phosphorylation of alpha(1B)-adrenergic receptors induced by TGF-beta. Our results indicate that activation of TGF-beta receptors induces alpha(1B)-adrenergic receptor phosphorylation and desensitization. The data suggest that phosphoinositide 3-kinase and protein kinase C play key roles in this effect of TGF-beta. PMID:12234252

  11. alpha1B-Adrenergic receptor phosphorylation and desensitization induced by transforming growth factor-beta.

    PubMed

    Romero-Avila, M Teresa; Flores-Jasso, C Fabián; García-Sáinz, J Adolfo

    2002-12-01

    Transforming growth factor-beta (TGF-beta) induced alpha(1B)-adrenergic receptor phosphorylation in Rat-1 fibroblasts stably expressing these adrenoceptors. This effect of TGF-beta was rapid, reaching a maximum within 30 min and decreasing thereafter, and concentration-dependent (EC(50) 0.3 pM). The phosphoinositide 3-kinase inhibitors wortmannin and LY294002, and the protein kinase C inhibitors staurosporine, Ro 318220 and bisindolylmaleimide, blocked the effect of this growth factor. alpha(1B)-Adrenergic receptor phosphorylation was associated with desensitization, as indicated by a reduction in the adrenergic-mediated production of [(3)H]inositol phosphates. Phosphorylation of alpha(1B)-adrenergic receptors by TGF-beta was also observed in Cos-1 cells transfected with the receptor. Co-transfection of the dominant-negative mutant of the regulatory subunit of phosphoinositide 3-kinase (Deltap85) inhibited the phosphorylation of alpha(1B)-adrenergic receptors induced by TGF-beta. Our results indicate that activation of TGF-beta receptors induces alpha(1B)-adrenergic receptor phosphorylation and desensitization. The data suggest that phosphoinositide 3-kinase and protein kinase C play key roles in this effect of TGF-beta. PMID:12234252

  12. Demonstration of single crystal growth via solid-solid transformation of a glass

    DOE PAGESBeta

    Savytskii, Dmytro; Knorr, Brian; Dierolf, Volkmar; Jain, Himanshu

    2016-03-18

    Many advanced technologies have relied on the availability of single crystals of appropriate material such as silicon for microelectronics or superalloys for turbine blades. Similarly, many promising materials could unleash their full potential if they were available in a single crystal form. However, the current methods are unsuitable for growing single crystals of these oftentimes incongruently melting, unstable or metastable materials. Here we demonstrate a strategy to overcome this hurdle by avoiding the gaseous or liquid phase, and directly converting glass into a single crystal. Specifically, Sb2S3 single crystals are grown in Sb-S-I glasses as an example of this approach.more » In this first unambiguous demonstration of an all-solid-state glass → crystal transformation, extraneous nucleation is avoided relative to crystal growth via spatially localized laser heating and inclusion of a suitable glass former in the composition. Lastly, the ability to fabricate patterned single-crystal architecture on a glass surface is demonstrated, providing a new class of micro-structured substrate for low cost epitaxial growth, active planar devices, etc.« less

  13. Melanoma-associated expression of transforming growth factor-beta isoforms.

    PubMed Central

    Van Belle, P.; Rodeck, U.; Nuamah, I.; Halpern, A. C.; Elder, D. E.

    1996-01-01

    Melanocytic neoplasia is characterized by the aberrant overproduction of multiple cytokines in vitro. However, it is largely unknown how cytokine expression relates to melanoma progression in vivo. Transforming growth factor beta (TGF-beta) is a multifunctional cytokine commonly produced by cultured melanoma cells. The in situ expression of all three TGF-beta isoforms (TGF-beta1, -2, and -3) was determined immunohistochemically in melanocytes and in 51 melanocytic lesions using isoform-specific antibodies. Significant linear trends of expression were observed from melanocytes through nevi and primary and metastatic melanomas for all three isoforms. TGF-beta1 was expressed by some melanocytes and almost uniformly by nevi and melanomas. TGF-beta2 and -3 were not expressed in normal melanocytes but were expressed in nevi and early and advanced primary (radial and vertical growth phase) and metastatic melanomas in a tumor-progression-related manner. TGF-beta2 was heterogeneously expressed in advanced primary and metastatic melanomas, whereas TGF-beta3 was uniformly and highly expressed in these lesions. Thus, expression of TGF-beta isoforms in melanocytes and melanocytic lesions is heterogeneous and related to tumor progression, and expression of TGF-beta2 and TGF-beta3 commences at critical junctures during progression of nevi to primary melanomas. Images Figure 2 Figure 3 PMID:8669474

  14. Changes in Maternal Serum Transforming Growth Factor Beta-1 during Pregnancy: A Cross-Sectional Study

    PubMed Central

    Orazulike, Ngozi C.; Ashmore, Jill; Konje, Justin C.

    2013-01-01

    Changes in circulating levels of maternal serum transforming growth factor beta-1 (TGF-β1), collected from 98 women (AGA) at different gestational ages (10–38 weeks) were measured and comparisons were made between levels in pregnant and nonpregnant controls and also between 10 women with small-for-gestational age (SGA) and 7 with appropriate-for-gestational age (AGA) fetuses. Maternal serum TGF-β1 levels at all stages of pregnancy were higher than those in normal healthy nonpregnant adults. The mean TGF-β1 levels in SGA pregnancies at 34-week gestation (32.5 + 3.2 ng/mL) were significantly less than those in AGA pregnancies (39.2 + 9.8 ng/mL) while at 38-week gestation, the levels were similar in the two groups (36.04 + 4.3 versus 36.7 + 7.0 ng/mL). This differential change in TGF-β1 levels is probably an important modulating factor in the aetiopathogenesis of abnormal intrauterine fetal growth. PMID:24350258

  15. Demonstration of single crystal growth via solid-solid transformation of a glass.

    PubMed

    Savytskii, Dmytro; Knorr, Brian; Dierolf, Volkmar; Jain, Himanshu

    2016-01-01

    Many advanced technologies have relied on the availability of single crystals of appropriate material such as silicon for microelectronics or superalloys for turbine blades. Similarly, many promising materials could unleash their full potential if they were available in a single crystal form. However, the current methods are unsuitable for growing single crystals of these oftentimes incongruently melting, unstable or metastable materials. Here we demonstrate a strategy to overcome this hurdle by avoiding the gaseous or liquid phase, and directly converting glass into a single crystal. Specifically, Sb2S3 single crystals are grown in Sb-S-I glasses as an example of this approach. In this first unambiguous demonstration of an all-solid-state glass → crystal transformation, extraneous nucleation is avoided relative to crystal growth via spatially localized laser heating and inclusion of a suitable glass former in the composition. The ability to fabricate patterned single-crystal architecture on a glass surface is demonstrated, providing a new class of micro-structured substrate for low cost epitaxial growth, active planar devices, etc. PMID:26988919

  16. Altered expression of transforming growth factor-beta isoforms in bovine cystic ovarian disease.

    PubMed

    Matiller, V; Stangaferro, M L; Díaz, P U; Ortega, H H; Rey, F; Huber, E; Salvetti, N R

    2014-10-01

    Cystic ovarian disease (COD) is one of the main causes of infertility in dairy cattle. It has been shown that intra-ovarian factors may contribute to follicular persistence. Transforming growth factor-beta (TGFB) isoforms are important paracrine and autocrine signalling molecules that regulate ovarian follicle growth and physiology. Considering the importance of these factors in the ovarian physiology, in this study, we examined the expression of TGFB isoforms (TGFB1, TGFB2 and TGFB3) in the ovary of healthy cows and animals with spontaneous and adrenocorticotrophic hormone (ACTH)-induced COD. In the oestrous-synchronized control group, the expression of TGFB1 in granulosa and theca cells was higher in spontaneous cysts than in atretic or tertiary follicles. When we compared TGFB2 expression in granulosa cells from atretic or tertiary follicles from the oestrous-synchronized control group with that in ACTH-induced or spontaneous follicular cysts, we found a higher expression in the latter. The expression of the TGFB isoforms studied was also altered during folliculogenesis in both the spontaneous and ACTH-induced COD groups. As it has been previously shown that TGFB influences steroidogenesis, ovarian follicular proliferation and apoptosis, an alteration in its expression may contribute to the pathogenesis of this disease. PMID:25112788

  17. Demonstration of single crystal growth via solid-solid transformation of a glass

    NASA Astrophysics Data System (ADS)

    Savytskii, Dmytro; Knorr, Brian; Dierolf, Volkmar; Jain, Himanshu

    2016-03-01

    Many advanced technologies have relied on the availability of single crystals of appropriate material such as silicon for microelectronics or superalloys for turbine blades. Similarly, many promising materials could unleash their full potential if they were available in a single crystal form. However, the current methods are unsuitable for growing single crystals of these oftentimes incongruently melting, unstable or metastable materials. Here we demonstrate a strategy to overcome this hurdle by avoiding the gaseous or liquid phase, and directly converting glass into a single crystal. Specifically, Sb2S3 single crystals are grown in Sb-S-I glasses as an example of this approach. In this first unambiguous demonstration of an all-solid-state glass → crystal transformation, extraneous nucleation is avoided relative to crystal growth via spatially localized laser heating and inclusion of a suitable glass former in the composition. The ability to fabricate patterned single-crystal architecture on a glass surface is demonstrated, providing a new class of micro-structured substrate for low cost epitaxial growth, active planar devices, etc.

  18. Down-regulated expression of transforming growth factor beta 1 mRNA in endometrial carcinoma.

    PubMed Central

    Perlino, E.; Loverro, G.; Maiorano, E.; Giannini, T.; Cazzolla, A.; Napoli, A.; Fiore, M. G.; Ricco, R.; Marra, E.; Selvaggi, L.

    1998-01-01

    Transforming growth factor beta1 (TGF-beta1) is a potent modulator of cell proliferation in vitro, and recent studies have demonstrated its overexpression in several different tumours; nevertheless, the molecular mechanisms of TGF-beta1 action on cell growth and differentiation have not been fully elucidated. To clarify the role of TGF-beta and its receptor in human endometrial proliferation and differentiation, TGF-beta1 expression at both the mRNA and protein levels has been evaluated by using Northern blotting and immunohistochemistry, in both normal (atrophic, proliferative and secretory) and neoplastic (adenocarcinoma) endometrial samples. This study demonstrates that TGF-beta1 mRNA expression is dramatically reduced in endometrial carcinomas with respect to non-neoplastic tissues, whereas the immunohistochemical expression of TGF-beta1 is enhanced in the epithelial component of endometrial carcinomas compared with non-neoplastic tissues. These data suggest that TGF-beta1 acts as a paracrine regulator of endometrial cell proliferation and that it may contribute to the carcinogenic mechanisms of endometrial carcinoma. Images Figure 1 Figure 5 Figure 6 Figure 8 PMID:9579831

  19. Down-regulated expression of transforming growth factor beta 1 mRNA in endometrial carcinoma.

    PubMed

    Perlino, E; Loverro, G; Maiorano, E; Giannini, T; Cazzolla, A; Napoli, A; Fiore, M G; Ricco, R; Marra, E; Selvaggi, L

    1998-04-01

    Transforming growth factor beta1 (TGF-beta1) is a potent modulator of cell proliferation in vitro, and recent studies have demonstrated its overexpression in several different tumours; nevertheless, the molecular mechanisms of TGF-beta1 action on cell growth and differentiation have not been fully elucidated. To clarify the role of TGF-beta and its receptor in human endometrial proliferation and differentiation, TGF-beta1 expression at both the mRNA and protein levels has been evaluated by using Northern blotting and immunohistochemistry, in both normal (atrophic, proliferative and secretory) and neoplastic (adenocarcinoma) endometrial samples. This study demonstrates that TGF-beta1 mRNA expression is dramatically reduced in endometrial carcinomas with respect to non-neoplastic tissues, whereas the immunohistochemical expression of TGF-beta1 is enhanced in the epithelial component of endometrial carcinomas compared with non-neoplastic tissues. These data suggest that TGF-beta1 acts as a paracrine regulator of endometrial cell proliferation and that it may contribute to the carcinogenic mechanisms of endometrial carcinoma. PMID:9579831

  20. Mutant forms of growth factor-binding protein-2 reverse BCR-ABL-induced transformation.

    PubMed Central

    Gishizky, M L; Cortez, D; Pendergast, A M

    1995-01-01

    Growth factor-binding protein 2 (Grb2) is an adaptor protein that links tyrosine kinases to Ras. BCR-ABL is a tyrosine kinase oncoprotein that is implicated in the pathogenesis of Philadelphia chromosome (Ph1)-positive leukemias. Grb2 forms a complex with BCR-ABL and the nucleotide exchange factor Sos that leads to the activation of the Ras protooncogene. In this report we demonstrate that Grb2 mutant proteins lacking amino- or carboxyl-terminal src homology SH3 domains suppress BCR-ABL-induced Ras activation and reverse the oncogenic phenotype. The Grb2 SH3-deletion mutant proteins bind to BCR-ABL and do not impair tyrosine kinase activity. Expression of the Grb2 SH3-deletion mutant proteins in BCR-ABL-transformed Rat-1 fibroblasts and in the human Ph1-positive leukemic cell line K562 inhibits their ability to grow as foci in soft agar and form tumors in nude mice. Furthermore, expression of the Grb2 SH3-deletion mutants in K562 cells induced their differentiation. Because Ras plays an important role in signaling by receptor and nonreceptor tyrosine kinases, the use of interfering mutant Grb2 proteins may be applied to block the proliferation of other cancers that depend in part on activated tyrosine kinases for growth. Images Fig. 1 Fig. 2 Fig. 3 PMID:7479904

  1. Demonstration of single crystal growth via solid-solid transformation of a glass

    PubMed Central

    Savytskii, Dmytro; Knorr, Brian; Dierolf, Volkmar; Jain, Himanshu

    2016-01-01

    Many advanced technologies have relied on the availability of single crystals of appropriate material such as silicon for microelectronics or superalloys for turbine blades. Similarly, many promising materials could unleash their full potential if they were available in a single crystal form. However, the current methods are unsuitable for growing single crystals of these oftentimes incongruently melting, unstable or metastable materials. Here we demonstrate a strategy to overcome this hurdle by avoiding the gaseous or liquid phase, and directly converting glass into a single crystal. Specifically, Sb2S3 single crystals are grown in Sb-S-I glasses as an example of this approach. In this first unambiguous demonstration of an all-solid-state glass → crystal transformation, extraneous nucleation is avoided relative to crystal growth via spatially localized laser heating and inclusion of a suitable glass former in the composition. The ability to fabricate patterned single-crystal architecture on a glass surface is demonstrated, providing a new class of micro-structured substrate for low cost epitaxial growth, active planar devices, etc. PMID:26988919

  2. Abrogation of growth arrest signals by human papillomavirus type 16 E7 is mediated by sequences required for transformation.

    PubMed Central

    Demers, G W; Espling, E; Harry, J B; Etscheid, B G; Galloway, D A

    1996-01-01

    Cells arrest in the G1 or G0 phase of the cell cycle in response to a variety of negative growth signals that induce arrest by different molecular pathways. The ability of human papillomavirus (HPV) oncogenes to bypass these signals and allow cells to progress into the S phase probably contributes to the neoplastic potential of the virus. The E7 protein of HPV-16 was able to disrupt the response of epithelial cells to three different negative growth arrest signals: quiescence imposed upon suprabasal epithelial cells, G1 arrest induced by DNA damage, and inhibition of DNA synthesis caused by treatment with transforming growth factor beta. The same set of mutated E7 proteins was able to abrogate all three growth arrest signals. Mutant proteins that failed to abrogate growth arrest signals were transformation deficient and included E7 proteins that bound retinoblastoma protein in vitro. In contrast, HPV-16 E6 was able to bypass only DNA damage-induced G1 arrest, not suprabasal quiescence or transforming growth factor beta-induced arrest. The E6 and E7 proteins from the low-risk virus HPV-6 were not able to bypass any of the growth arrest signals. PMID:8794328

  3. Cross-Talk Between Nitric Oxide and Transforming Growth Factor-β1 in Malaria

    PubMed Central

    Vodovotz, Yoram; Zamora, Ruben; Lieber, Matthew J.; Luckhart, Shirley

    2008-01-01

    Malaria has re-emerged as a global health problem, leading to an increased focus on the cellular and molecular biology of the mosquito Anopheles and the parasite Plasmodium with the goal of identifying novel points of intervention in the parasite life cycle. Anti-parasite defenses mounted by both mammalian hosts and Anopheles can suppress the growth of Plasmodium. Nonetheless, the parasite is able to escape complete elimination in vivo, perhaps by thwarting or co-opting these mechanisms for its own survival, as do numerous other pathogens. Among the defense systems used by the mammalian host against Plasmodium is the synthesis of nitric oxide (NO), catalyzed by an inducible NO synthase (iNOS). Nitric oxide produced by the action of an inducible Anopheles stephensi NO synthase (AsNOS) may be central to the anti-parasitic arsenal of this mosquito. In mammals, iNOS can be modulated by members of the transforming growth factor-β (TGF-β) cytokine superfamily. Transforming growth factor-β is produced as an inactive precursor that is activated following dissociation of certain inhibitory proteins, a process that can be promoted by reaction products of NO as well as by hemin. Ingestion by Anopheles of blood containing Plasmodium initiates parasite development, blood digestion which results in the accumulation of hematin (hemin) in the insect midgut, and induction of both AsNOS and TGF-β-like (As60A) gene expression in the midgut epithelium. Active mammalian TGF-β1 can be detected in the A. stephensi midgut up to 48h post-ingestion and latent TGF-β1 can be activated by midgut components in vitro, a process that is potentiated by NO and that may involve hematin. Further, mammalian TGF-β1 is perceived as a cytokine by A. stephensi cells in vitro and can alter Plasmodium development in vivo. Bloodfeeding by Anopheles, therefore, results in a juxtaposition of evolutionarily conserved mosquito and mammalian TGF-β superfamily homologs that may influence transmission

  4. ACD toxin-produced actin oligomers poison formin-controlled actin polymerization

    PubMed Central

    Heisler, David B.; Kudryashova, Elena; Grinevich, Dmitry O.; Suarez, Cristian; Winkelman, Jonathan D.; Birukov, Konstantin G.; Kotha, Sainath R.; Parinandi, Narasimham L.; Vavylonis, Dimitrios; Kovar, David R.; Kudryashov, Dmitri S.

    2015-01-01

    The actin crosslinking domain (ACD) is an actin-specific toxin produced by several pathogens, including life-threatening spp. of Vibrio cholerae, Vibrio vulnificus, and Aeromonas hydrophila. Actin crosslinking by ACD is thought to lead to slow cytoskeleton failure owing to a gradual sequestration of actin in the form of nonfunctional oligomers. Here we found that ACD converted cytoplasmic actin into highly toxic oligomers that potently “poisoned” the ability of major actin assembly proteins, formins, to sustain actin polymerization. Thus, ACD can target the most abundant cellular protein by employing actin oligomers as secondary toxins to efficiently subvert cellular functions of actin while functioning at very low doses. PMID:26228148

  5. Dynamics of Transforming Growth Factor Beta Signaling in Wound Healing and Scarring

    PubMed Central

    Finnson, Kenneth W.; McLean, Sarah; Di Guglielmo, Gianni M.; Philip, Anie

    2013-01-01

    Significance Wound healing is an intricate biological process in which the skin, or any other tissue, repairs itself after injury. Normal wound healing relies on the appropriate levels of cytokines and growth factors to ensure that cellular responses are mediated in a coordinated manner. Among the many growth factors studied in the context of wound healing, transforming growth factor beta (TGF-β) is thought to have the broadest spectrum of effects. Recent Advances Many of the molecular mechanisms underlying the TGF-β/Smad signaling pathway have been elucidated, and the role of TGF-β in wound healing has been well characterized. Targeting the TGF-β signaling pathway using therapeutic agents to improve wound healing and/or reduce scarring has been successful in pre-clinical studies. Critical Issues Although TGF-β isoforms (β1, β2, β3) signal through the same cell surface receptors, they display distinct functions during wound healing in vivo through mechanisms that have not been fully elucidated. The challenge of translating preclinical studies targeting the TGF-β signaling pathway to a clinical setting may require more extensive preclinical research using animal models that more closely mimic wound healing and scarring in humans, and taking into account the spatial, temporal, and cell-type–specific aspects of TGF-β isoform expression and function. Future Directions Understanding the differences in TGF-β isoform signaling at the molecular level and identification of novel components of the TGF-β signaling pathway that critically regulate wound healing may lead to the discovery of potential therapeutic targets for treatment of impaired wound healing and pathological scarring. PMID:24527343

  6. Mesenchymal stem cells promote tumor angiogenesis via the action of transforming growth factor β1

    PubMed Central

    LI, GUO-CAI; ZHANG, HONG-WEI; ZHAO, QING-CHUN; SUN, LI; YANG, JIAN-JUN; HONG, LIU; FENG, FAN; CAI, LEI

    2016-01-01

    Mesenchymal stem cells (MSCs) may influence the growth and metastasis of various human malignancies, including hepatocellular carcinoma (HCC). Therefore, the underlying mechanisms via which MSCs are able to affect malignancies require investigation. In the present study, the potential role of MSC in the angiogenesis of HCC was investigated. A total of 17 nude mouse models exhibiting human HCC were used to evaluate the effects of MSC on angiogenesis. A total of 8 mice were injected with human MSCs via the tail vein, and the remaining 9 mice were injected with phosphate-buffered saline as a control. A total of 35 days subsequent to the injection of MSCs, the microvessel density (MVD) of tumors was evaluated by immunostaining, using cluster of differentiation 31 antibody. The mRNA levels of transforming growth factor (TGF)β1, Smad2 and Smad7 were detected using reverse transcription-quantitative polymerase chain reaction. Protein expression levels of TGFβ1 and vascular endothelial growth factor (VEGF) in tumor tissues were analyzed using ELISA. Compared with controls, MVD in MSC-treated mice was significantly increased (28.00±9.19 vs. 18.11±3.30; P=0.006). The levels of TGFβ1 mRNA in the MSC-treated group were 2.15-fold higher compared with the control group (1.27±0.61 vs. 0.59±0.39; P=0.033), and MVD was higher in the group exhibiting increased TGFβ1 mRNA levels compared with the control group (26.50±9.11 vs. 19.44±6.14; P=0.038). In addition, a close correlation between the expression levels of TGFβ1 and VEGF was identified. The results of the present study suggested that MSCs may be capable of enhancing the angiogenesis of HCC, which may be partly due to the involvement of TGFβ1. PMID:26893697

  7. Analysis of Epithelial-Mesenchymal Transition Induced by Transforming Growth Factor β.

    PubMed

    Valcourt, Ulrich; Carthy, Jonathon; Okita, Yukari; Alcaraz, Lindsay; Kato, Mitsuyasu; Thuault, Sylvie; Bartholin, Laurent; Moustakas, Aristidis

    2016-01-01

    In recent years, the importance of the cell biological process of epithelial-mesenchymal transition (EMT) has been established via an exponentially growing number of reports. EMT has been documented during embryonic development, tissue fibrosis, and cancer progression in vitro, in animal models in vivo and in human specimens. EMT relates to many molecular and cellular alterations that occur when epithelial cells undergo a switch in differentiation that generates mesenchymal-like cells with newly acquired migratory and invasive properties. In addition, EMT relates to a nuclear reprogramming similar to the one occurring in the generation of induced pluripotent stem cells. Via such a process, EMT is gradually established to promote the generation and maintenance of adult tissue stem cells which under disease states such as cancer, are known as cancer stem cells. EMT is induced by developmental growth factors, oncogenes, radiation, and hypoxia. A prominent growth factor that causes EMT is transforming growth factor β (TGF-β).A series of molecular and cellular techniques can be applied to define and characterize the state of EMT in diverse biological samples. These methods range from DNA and RNA-based techniques that measure the expression of key EMT regulators and markers of epithelial or mesenchymal differentiation to functional assays of cell mobility, invasiveness and in vitro stemness. This chapter focuses on EMT induced by TGF-β and provides authoritative protocols and relevant reagents and citations of key publications aiming at assisting newcomers that enter this prolific area of biomedical sciences, and offering a useful reference tool to pioneers and aficionados of the field. PMID:26520123

  8. Effect of transforming growth factor beta on synthesis of glycosaminoglycans by human lung fibroblasts

    SciTech Connect

    Dubaybo, B.A.; Thet, L.A. )

    1990-09-01

    The processes of lung growth, injury, and repair are characterized by alterations in fibroblast synthesis and interstitial distribution of extracellular matrix components. Transforming growth factor beta (TGF-beta), which is postulated to play a role in modulating lung repair, alters the distribution of several matrix components such as collagen and fibronectin. We studied the effect of TGF-beta on the synthesis and distribution of the various glycosaminoglycans (GAGs) and whether these effects may explain its role in lung repair. Human diploid lung fibroblasts (IMR-90) were exposed to various concentrations of TGF-beta (0-5 nM) for variable periods of time (0-18 h). Newly synthesized GAGs were labeled with either (3H)glucosamine or (35S)sulfate. Individual GAGs were separated by size exclusion chromatography after serial enzymatic and chemical digestions and quantitated using scintillation counting. There was a dose-dependent increase in total GAG synthesis with maximal levels detected after 6 h of exposure. This increase was noted in all individual GAG types measured and was observed in both the cell associated GAGs (cell-matrix fraction) as well as the GAGs released into the medium (medium fraction). In the cell-matrix fraction, TGF-beta increased the proportion of heparan sulfate that was membrane bound as well as the proportion of dermatan sulfate in the intracellular compartment. In the medium fraction, TGF-beta increased the proportion of hyaluronic acid, chondroitin sulfate and dermatan sulfate released. We conclude that the role of TGF-beta in lung growth and repair may be related to increased synthesis of GAGs by human lung fibroblasts as well as alterations in the distribution of individual GAGs.

  9. Chemotaxis and Actin Oscillations

    NASA Astrophysics Data System (ADS)

    Bodenschatz, Eberhard; Hsu, Hsin-Fang; Negrete, Jose; Beta, Carsten; Pumir, Alain; Gholami, Azam; Tarantola, Marco; Westendorf, Christian; Zykov, Vladimir

    Recently, self-oscillations of the cytoskeletal actin have been observed in Dictyostelium, a model system for studying chemotaxis. Here we report experimental results on the self-oscillation mechanism and the role of regulatory proteins and myosin II. We stimulate cells rapidly and periodically by using photo un-caging of the chemoattractant in a micro-fluidic device and measured the cellular responses. We found that the response amplitude grows with stimulation strength only in a very narrow region of stimulation, after which the response amplitude reaches a plateau. Moreover, the frequency-response is not constant but rather varies with the strength of external stimuli. To understand the underlying mechanism, we analyzed the polymerization and de-polymerization time in the single cell level. Despite of the large cell-to-cell variability, we found that the polymerization time is independent of external stimuli and the de-polymerization time is prolonged as the stimulation strength increases. Our conclusions will be summarized and the role of noise in the signaling network will be discussed. German Science Foundation CRC 937.

  10. Growth and phase transformations: Insights from hydrothermal coarsening experiements using nanocrystalline titanium dioxide

    NASA Astrophysics Data System (ADS)

    Penn, Rona Lee

    Controlling the morphology and size of crystallizing materials is of special interest in Materials Science as particle morphology can affect many materials characteristics, including magnetism, reactivity, and densification. High-resolution transmission electron microscopy (HRTEM), X-ray diffraction (XRD) peak broadening (Scherrer) analysis, and determination of room temperature adsorption of organic acids by ion chromatography were used to study adsorption and hydrothermal coarsening and morphology development in nanocrystalline anatase (TiOsb2). HRTEM micrographs show that nanocrystalline anatase coarsens by two growth mechanisms: single crystal growth, via subnanometer-high surface steps, and a new growth mechanism known as oriented attachment, which involves topotactic joining of crystallites with subsequent coarsening to form single crystals of unique morphology. Addition of hydrochloric acid (pH = 3.0) to suspensions prior to hydrothermal treatment results in the formation of extremely elongate single crystals often comprised of more than 7 primary anatase crystallites. Both decreasing the pH of solutions below 3 and addition of organic acids resulted in a reduction in the frequency of observed attachments. Because attachment is only constrained by two dimensional match at the interface, attachment can lead to twinning and intergrowths on \\{112\\}, \\{101\\}, and (001). Slight misorientations between particles arising from surface topography at attaching surfaces, produces dislocations of edge, screw, and mixed character. Furthermore, anatase \\{112\\} twin boundaries consist of structural elements common to rutile and may serve as nucleation sites for the anatase-to-rutile transformation. These results have broad relevance for nucleation and growth models as they suggest that twinning, polytypism, and linear defects in macroscopic crystals can originate at oriented interfaces between primary nanocrystalline particles early in their crystallization history

  11. Mechanical Detection of a Long-Range Actin Network Emanating from a Biomimetic Cortex

    PubMed Central

    Bussonnier, Matthias; Carvalho, Kevin; Lemière, Joël; Joanny, Jean-François; Sykes, Cécile; Betz, Timo

    2014-01-01

    Actin is ubiquitous globular protein that polymerizes into filaments and forms networks that participate in the force generation of eukaryotic cells. Such forces are used for cell motility, cytokinesis, and tissue remodeling. Among those actin networks, we focus on the actin cortex, a dense branched network beneath the plasma membrane that is of particular importance for the mechanical properties of the cell. Here we reproduce the cellular cortex by activating actin filament growth on a solid surface. We unveil the existence of a sparse actin network that emanates from the surface and extends over a distance that is at least 10 times larger than the cortex itself. We call this sparse actin network the “actin cloud” and characterize its mechanical properties with optical tweezers. We show, both experimentally and theoretically, that the actin cloud is mechanically relevant and that it should be taken into account because it can sustain forces as high as several picoNewtons (pN). In particular, it is known that in plant cells, actin networks similar to the actin cloud have a role in positioning the nucleus; in large oocytes, they play a role in driving chromosome movement. Recent evidence shows that such networks even prevent granule condensation in large cells. PMID:25140420

  12. Interleukin-6, vascular endothelial growth factor and transforming growth factor beta 1 in canine steroid responsive meningitis-arteritis

    PubMed Central

    2013-01-01

    Background Steroid Responsive Meningitis-Arteritis (SRMA) is a common cause of inflammation of the canine central nervous system (CNS). To investigate if transforming growth factor beta 1 (TGF-β1), interleukin-6 (IL-6) and vascular endothelial growth factor (VEGF) are involved in the production of excessive immunoglobulin A (IgA), the induction of acute phase proteins and in the development of a systemic necrotizing vasculitis, characteristic of SRMA, these three signalling proteins were evaluated. Results Cerebrospinal fluid (CSF) and serum samples of dogs during the acute phase of SRMA (SRMA) were tested for IL-6, VEGF and TGF- β1. Results were compared to those of dogs affected with SRMA during treatment (SRMA Th) and during relapse (SRMA R), to dogs with other meningoencephalomyelitides (ME), with miscellaneous non-inflammatory diseases of the CNS (CNS-Mix), with idiopathic epilepsy (IE), with systemic inflammatory diseases (Syst. Infl.) and with healthy dogs (Healthy). Concentrations of IL-6 and VEGF in CSF were significantly elevated in the SRMA group compared to the other disease categories (p < 0.05). The CSF concentrations of TGF-β1 were increased in SRMA group, but statistically significant differences were found only in comparison with Healthy and CNS-Mix groups. No differences were detected in the serum concentrations of TGF-β1 between the different groups. In untreated SRMA patients, a positive correlation (rSpear = 0.3549; P = 0.0337) between concentrations of TGF-β1 and IgA concentration was found in CSF, while concentrations of IL-6 and VEGF in CSF positively correlated with the degree of pleocytosis (rSpear = 0.8323; P < 0.0001 and rSpear = 0.5711; P = 0.0166, respectively). Conclusions Our results suggest that these three signalling proteins are biomarkers of disease activity in SRMA. VEGF might play an important role in the development of a systemic arteritis. TGF-β1 is considered to be involved in the excessive

  13. Cloning and characterization of the actin gene from Puccinia striiformis f. sp. tritici.

    PubMed

    Liu, Jie; Zhang, Qiong; Chang, Qing; Zhuang, Hua; Huang, Li-Li; Kang, Zhen-Sheng

    2012-06-01

    The fungus Puccinia striiformis f. sp. tritici, the causal agent of wheat stripe rust, is an obligate biotrophic basidiomycete. Urediniospores are the most common spore type involved in the epidemiology of this disease. Tip growth of germ tubes of germinated urediniospores is a key step during infection of wheat, but few studies have investigated it so far. Recent research has found that actin is closely associated with hyphal tip growth. In this study, we have cloned and obtained the full-length actin cDNA from P. striiformis f. sp. tritici and characterized its expression. Furthermore, actin filament (F-actin) patterns were visualized microscopically during germ tube formation. The most conspicuous actin-containing structures were actin patches. They were mainly concentrated near the hyphal tip and scattered throughout the cortex. By using cytochalasin B, we observed that depolymerization of F-actin greatly reduced the germination rate of urediniospores and disrupted the transport of vesicles to the germ tube tip, indicating that F-actin played a key role in the tip growth of P. striiformis f. sp. tritici. This work helps us to understand the tip growth mechanism of P. striiformis f. sp. tritici, and may provide a theoretical framework for designing novel pesticides. PMID:22806107

  14. HopW1 from Pseudomonas syringae Disrupts the Actin Cytoskeleton to Promote Virulence in Arabidopsis

    PubMed Central

    Cecchini, Nicolas M.; Li, Yujie; Lee, Min Woo; Kovar, David R.; Greenberg, Jean T.

    2014-01-01

    A central mechanism of virulence of extracellular bacterial pathogens is the injection into host cells of effector proteins that modify host cellular functions. HopW1 is an effector injected by the type III secretion system that increases the growth of the plant pathogen Pseudomonas syringae on the Columbia accession of Arabidopsis. When delivered by P. syringae into plant cells, HopW1 causes a reduction in the filamentous actin (F-actin) network and the inhibition of endocytosis, a known actin-dependent process. When directly produced in plants, HopW1 forms complexes with actin, disrupts the actin cytoskeleton and inhibits endocytosis as well as the trafficking of certain proteins to vacuoles. The C-terminal region of HopW1 can reduce the length of actin filaments and therefore solubilize F-actin in vitro. Thus, HopW1 acts by disrupting the actin cytoskeleton and the cell biological processes that depend on actin, which in turn are needed for restricting P. syringae growth in Arabidopsis. PMID:24968323

  15. Synthetic Chondramide A Analogues Stabilize Filamentous Actin and Block Invasion by Toxoplasma gondii

    PubMed Central

    2013-01-01

    Apicomplexan parasites such as Toxoplasma gondii rely on actin-based motility to cross biological barriers and invade host cells. Key structural and biochemical differences in host and parasite actins make this an attractive target for small-molecule inhibitors. Here we took advantage of recent advances in the synthesis of cyclic depsipeptide compounds that stabilize filamentous actin to test the ability of chondramides to disrupt growth of T. gondii in vitro. Structural modeling of chondramide A (2) binding to an actin filament model revealed variations in the binding site between host and parasite actins. A series of 10 previously synthesized analogues (2b–k) with substitutions in the β-tyrosine moiety blocked parasite growth on host cell monolayers with EC50 values that ranged from 0.3 to 1.3 μM. In vitro polymerization assays using highly purified recombinant actin from T. gondii verified that synthetic and natural product chondramides target the actin cytoskeleton. Consistent with this, chondramide treatment blocked parasite invasion into host cells and was more rapidly effective than pyrimethamine, a standard therapeutic agent. Although the current compounds lack specificity for parasite vs host actin, these studies provide a platform for the future design and synthesis of synthetic cyclic peptide inhibitors that selectively disrupt actin dynamics in parasites. PMID:24020843

  16. Atomic Force Microscopy and Light Scattering of Small Unilamellar Actin-Containing Liposomes

    PubMed Central

    Palmer, Andre F.; Wingert, Philip; Nickels, Jonathan

    2003-01-01

    Three-dimensional networks of filamentous actin (F-actin) encapsulated inside phosphatidylcholine liposomes are currently being used in an effort to model the cytoskeleton and plasma membrane of eukaryotic cells. In this article, unilamellar lipid vesicles consisting of egg yolk-derived phosphatidylcholine encapsulating monomeric actin (G-actin) were made via extrusion in low ionic strength buffer (G-buffer). Vesicle shape and structure in these dispersions was studied using a combination of fluid-tapping atomic force microscopy, and multiangle static light scattering. After subjecting the liposome dispersion to high ionic strength polymerization buffer (F-buffer) containing K+ ions, atomic force microscopy imaging and light scattering of these liposomes indicated the formation of specialized structures, including an overall liposome structure transformation from spherical to torus, disk-shaped geometries and tubular assemblies. Several atomic force microscopy control measurements were made to ascertain that the specialized structures formed were not due to free G-actin and F-actin self-assembling on the sample surface, plain liposomes exposed to G- and F-buffer, or liposomes encapsulating G-actin. Liposomes encapsulating G-actin assumed mostly thin disk shapes and some large irregularly shaped aggregates. In contrast, liposomes encapsulating polymerized actin assumed mostly torus or disk shapes along with some high aspect ratio tubular structures. PMID:12885667

  17. Nuclear translocation of type I transforming growth factor β receptor confers a novel function in RNA processing.

    PubMed

    Chandra, Manasa; Zang, Shengbing; Li, Haiqing; Zimmerman, Lisa J; Champer, Jackson; Tsuyada, Akihiro; Chow, Amy; Zhou, Weiying; Yu, Yang; Gao, Harry; Ren, Xiubao; Lin, Ren-Jang; Wang, Shizhen Emily

    2012-06-01

    Signaling of transforming growth factor β (TGF-β) is redirected in cancer to promote malignancy, but how TGF-β function is altered in a transformed cell is not fully understood. We investigated TGF-β signaling by profiling proteins that differentially bound to type I TGF-β receptor (TβRI) in nontransformed, HER2-transformed, and HER2-negative breast cancer cells using immunoprecipitation followed by protein identification. Interestingly, several nuclear proteins implicated in posttranscriptional RNA processing were uniquely identified in the TβRI coprecipitates from HER2-transformed cells. Ligand-inducible nuclear translocation of TβRI was observed only in transformed cells, and the translocation required importin β1, nucleolin, and Smad2/3. This trafficking was dependent on the high Ran GTPase activity resulting from oncogenic transformation. In the nucleus, TβRI associated with purine-rich RNA sequences in a synergistic manner with the RNA-binding factor hnRNP A1. We further found that nuclear translocation of TβRI specifically induced epidermal growth factor receptor (EGFR) transcript isoform c, which encodes a soluble EGFR protein, through alternative splicing or 3'-end processing. Our study confirms a cancer-specific nuclear translocation of TβRI and demonstrates its potential function in regulating nuclear RNA processing, as well as a novel gain-of-function mechanism of TGF-β signaling in cancer. PMID:22473997

  18. Cell-autonomous cytokinin-independent growth of tobacco cells transformed by Agrobacterium tumefaciens strains lacking the cytokinin biosynthesis gene.

    PubMed Central

    Black, R C; Binns, A N; Chang, C F; Lynn, D G

    1994-01-01

    Mutations at the cytokinin biosynthesis locus (tmr) of Agrobacterium tumefaciens usually result in strains that induce tumors exhibiting the rooty phenotype associated with high auxin-to-cytokinin ratios. However, tobacco (Nicotiana tabacum cv Havana 425) leaf disc explants responded to tmr- mutant strain A356 by producing rapidly growing, unorganized tumors, indicating that these lines can grow in a cytokinin-independent fashion despite the absence of a functional tmr gene. Several methods have been used to characterize the physiological and cellular basis of this phenotype. The results indicate that tmr- tumors have a physiologically distinct mechanism for cytokinin-independent growth in comparison to tumors induced by wild-type bacteria. The cytokinin-independent phenotype of the tmr- transformants appears to be cell autonomous in nature: only the transformed cells and their progeny were capable of cytokinin-independent growth. Specifically, the tmr- tumors did not accumulate cytokinin, and clonal analysis indicated the tmr- transformed cells were not capable of stimulating the growth of neighboring nontransformed cells. Finally, the cytokinin-independent phenotype of the tmr- transformants was shown to be cold sensitive, whereas the wild-type tumors exhibited a cold-resistant cytokinin-independent phenotype. Potential mechanisms for this novel form of cytokinin-independent growth, including the role of the dehydrodiconiferyl alcohol glucosides found in both tumor types, are discussed. PMID:8058843

  19. Growth habit and photo-synthetic activity of shoot cultures of Medicago sativa L. transformed with the oryzacystatin II gene

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In vitro maintained shoot cultures of alfalfa (Medicago sativa L. cv. Zajeÿarska 83) that were transformed with the oryzacystatin II (OCII) gene and propagated on growth regulator-free medium were subjected to analysis of morphological characteristics and photosynthetic activity. The most striking f...

  20. From wavy hair to naked proteins: the role of transforming growth factor alpha in health and disease.

    PubMed

    Singh, Bhuminder; Coffey, Robert J

    2014-04-01

    Since its discovery in 1978 and cloning in 1984, transforming growth factor-alpha (TGF-α, TGFA) has been one of the most extensively studied EGF receptor (EGFR) ligands. In this review, we provide a historical perspective on TGFA-related studies, highlighting what we consider important advances related to its function in normal and disease states. PMID:24631356

  1. GAP JUNCTION COMMUNICATION MEDIATES TRANSFORMING GROWTH FACTOR-BETA ACTIVATION AND ENDOTHELIAL-INDUCED MURAL CELL DIFFERENTIATION

    Technology Transfer Automated Retrieval System (TEKTRAN)

    During blood vessel assembly, endothelial cells recruit mesenchymal progenitors and induce their differentiation into mural cells via contact-dependent transforming growth factor-beta (TGF-beta) activation. We investigated whether gap junction channels are formed between endothelial cells and recrui...

  2. Increased susceptibility to atrial fibrillation secondary to atrial fibrosis in transgenic goats expressing transforming growth factor - B1

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Atrial fibrillation (AF) is the most common sustained cardiac arrhythmia in people with significant morbidity and mortality. There is a strong association between atrial fibrosis and AF. Transforming growth factor B1 (TGF-B1) is an essential mediator of atrial fibrosis in animal models and human pat...

  3. Regulation of water flow by actin-binding protein-induced actin gelatin.

    PubMed Central

    Ito, T; Suzuki, A; Stossel, T P

    1992-01-01

    Actin filaments inhibit osmotically driven water flow (Ito, T., K.S. Zaner, and T.P. Stossel. 1987. Biophys. J. 51: 745-753). Here we show that the actin gelation protein, actin-binding protein (ABP), impedes both osmotic shrinkage and swelling of an actin filament solution and reduces markedly the concentration of actin filaments required for this inhibition. These effects depend on actin filament immobilization, because the ABP concentration that causes initial impairment of water flow by actin filaments corresponds to the gel point measured viscometrically and because gelsolin, which noncovalently severs actin filaments, solates actin gels and restores water flow in a solution of actin cross-linked by ABP. Since ABP gels actin filaments in the periphery of many eukaryotic cells, such actin networks may contribute to physiological cell volume regulation. PMID:1318095

  4. Active Chemical Thermodynamics promoted by activity of cortical actin

    NASA Astrophysics Data System (ADS)

    Bhattacharya, Bhaswati; Chaudhuri, Abhishek; Gowrishankar, Kripa; Rao, Madan

    2011-03-01

    The spatial distribution and dynamics of formation and breakup of the nanoclusters of cell surface proteins is controlled by the active remodeling dynamics of the underlying cortical actin. To explain these observations, we have proposed a novel mechanism of nanoclustering, involving the transient binding to and advection along constitutively occuring ``asters'' of cortical actin. We study the consequences of such active actin-based clustering, in the context of chemical reactions involving conformational changes of cell surface proteins. We find that the active remodeling of cortical actin, can give rise to a dramatic increase in efficiency and extent of conformational spread, even at low levels of expression at the cell surface. We define a activity temperature (τa) arising due to actin activities which can be used to describe chemical thermodynamics of the system. We plot TTT (time-temparature-transformation) curves and compute the Arrhenius factors which depend on τa . With this, the active asters can be treated as enzymes whose enzymatic reaction rate can be related to the activity.

  5. Long Non-coding RNAs (LncRNA) Regulated by Transforming Growth Factor (TGF) β

    PubMed Central

    Richards, Edward J.; Zhang, Gu; Li, Zhu-Peng; Permuth-Wey, Jennifer; Challa, Sridevi; Li, Yajuan; Kong, William; Dan, Su; Bui, Marilyn M.; Coppola, Domenico; Mao, Wei-Min; Sellers, Thomas A.; Cheng, Jin Q.

    2015-01-01

    Long noncoding RNAs (lncRNAs) are emerging as key regulators in various biological processes. Epithelial-to-mesenchymal transition (EMT) is a developmental process hijacked by tumor cells to depart from the primary tumor site, invade surrounding tissue, and establish distant metastases. Transforming growth factor β (TGFβ) signaling has been shown to be a major inducer of EMT and to facilitate breast cancer metastasis. However, the role of lncRNAs in this process remains largely unknown. Here we report a genome-wide lncRNA profile in mouse mammary epithelial NMuMG cells upon TGFβ induction of EMT. Among 10,802 lncRNAs profiled, over 600 were up-regulated and down-regulated during the EMT, respectively. Furthermore, we identify that lncRNA-HIT (HOXA transcript induced by TGFβ) mediates TGFβ function, i.e. depletion of lncRNA-HIT inhibits TGFβ-induced migration, invasion, and EMT in NMuMG. LncRNA-HIT is also significantly elevated in the highly metastatic 4T1 cells. Knockdown of lncRNA-HIT in 4T1 results in decrease of cell migration, invasion, tumor growth, and metastasis. E-cadherin was identified as a major target of lncRNA-HIT. Moreover, lncRNA-HIT is conserved in humans and elevated expression associates with more invasive human primary breast carcinoma. Collectively, these data suggest that a subset of lncRNAs such as lncRNA-HIT play a significant role in regulation of EMT and breast cancer invasion and metastasis, and could be potential therapeutic targets in breast cancers. PMID:25605728

  6. Potassium Inhibits Dietary Salt-Induced Transforming Growth Factor-β Production

    PubMed Central

    Ying, Wei-Zhong; Aaron, Kristal; Wang, Pei-Xuan; Sanders, Paul W.

    2009-01-01

    Human and animal studies demonstrate an untoward effect of excess dietary NaCl (salt) intake on cardiovascular function and life span. The endothelium in particular augments the production of transforming growth factor (TGF)-β, a fibrogenic growth factor, in response to excess dietary salt intake. This study explored the initiating mechanism that regulates salt-induced endothelial cell production of TGF-β. Male Sprague-Dawley rats were given diets containing different amounts of NaCl and potassium for 4 days. A bioassay for TGF-β demonstrated increased (35.2%) amounts of active TGF-β in the medium of aortic ring segments from rats on the high-salt diet compared with rats maintained on a 0.3% NaCl diet. Inhibition of the large-conductance, calcium-activated potassium channel inhibited dietary salt-induced vascular production of TGF-β but did not affect production of TGF-β by ring segments from rats on the low-salt diet. Immunohistochemical and Western analyses demonstrated the α subunit of the calcium-activated potassium channel in endothelial cells. Increasing medium [K+] inhibited production of dietary salt-induced vascular production levels of total and active TGF-β but did not alter TGF-β production by aortic rings from rats on the 0.3% NaCl diet. Increasing dietary potassium content decreased urinary active TGF-β in animals receiving the high-salt diet but did not change urinary active TGF-β in animals receiving the low-salt diet. The findings demonstrated an interesting interaction between the dietary intake of potassium and excess NaCl and further showed the fundamental role of the endothelial calcium-activated potassium channel in the vascular response to excess salt intake. PMID:19738156

  7. Transforming growth factor-beta during carcinogenesis: the shift from epithelial to mesenchymal signaling.

    PubMed

    Matsuzaki, Koichi; Okazaki, Kazuichi

    2006-04-01

    Transforming growth factor-beta (TGF-beta) activates not only TGF-beta type I receptor (TbetaRI) but also c-Jun N-terminal kinase (JNK), changing unphosphorylated Smad3 to its phosphoisoforms: C-terminally phosphorylated Smad3 (pSmad3C) and linker phosphorylated Smad3 (pSmad3L). While the TbetaRI/pSmad3C pathway inhibits growth of normal epithelial cells, JNK/pSmad3L-mediated signaling is involved in invasion by activated mesenchymal cells. During sporadic human colorectal carcinogenesis, TGF-beta signaling confers a selective advantage on tumor cells by shifting from the TbetaRI/pSmad3C pathway characteristic of mature epithelial cells to the JNK/pSmad3L pathway, which is more characteristic of the state of flux shown by the activated mesenchymal cells. JNK acts as a regulator of TGF-beta signaling by increasing the basal level of pSmad3L available for action in the nuclei of the invasive adenocarcinoma, in the meantime shutting down TGF-beta-dependent nuclear activity of pSmad3C. Loss of epithelial homeostasis and acquisition of a migratory, mesenchymal phenotype are essential for tumor invasion. From the viewpoint of TGF-beta signaling, a key therapeutic aim in cancer would be restoration of the lost tumor suppressor function observed in normal colorectal epithelial cells at the expense of effects promoting aggressive behavior of the adenocarcinoma. Specific inhibitors of the JNK/pSmad3L pathway might prove useful in this respect. In the case of molecularly targeted therapy for human cancer, pSmad3L and pSmad3C could be assessed as biomarkers to evaluate the likely benefit from specific inhibition of the JNK/pSmad3L pathway. PMID:16741607

  8. Hypertrophic stimuli induce transforming growth factor-beta 1 expression in rat ventricular myocytes.

    PubMed Central

    Takahashi, N; Calderone, A; Izzo, N J; Mäki, T M; Marsh, J D; Colucci, W S

    1994-01-01

    Transforming growth factor-beta 1 (TGF-beta 1) is a peptide growth factor that may play a role in the myocardial response to hypertrophic stimuli. However, the cellular distribution, mechanism of induction, and source of increased TGF-beta 1 in response to hypertrophic stimuli are not known. We tested the hypothesis that the cardiac myocyte responds to hypertrophic stimuli with the increased expression of TGF-beta 1. In adult rat ventricular myocardium freshly dissociated into myocyte and nonmyocyte cellular fractions, the preponderance of TGF-beta 1 mRNA visualized by Northern hybridization was in the nonmyocyte fraction. Abdominal aortic constriction (7 d) and subcutaneous norepinephrine infusion (36 h) each caused ventricular hypertrophy associated with 3.1-fold and 3.8-fold increases, respectively, in TGF-beta 1 mRNA in the myocyte fraction, but had no effect on the level of TGF-beta 1 mRNA in the nonmyocyte fraction. In ventricular myocytes, norepinephrine likewise caused a 4.1-fold increase in TGF-beta 1 mRNA associated with an increase in TGF-beta bioactivity. This induction of TGF-beta 1 mRNA occurred at norepinephrine concentrations as low as 1 nM and was blocked by prazosin, but not propranolol. NE did not increase the TGF-beta 1 mRNA level in nonmyocytes, primarily fibroblasts, cultured from neonatal rat ventricle. Thus, the cardiac myocyte responds to two hypertrophic stimuli, pressure overload and norepinephrine, with the induction of TGF-beta 1. These data support the view that TGF-beta 1, released by myocytes and acting in an autocrine and/or paracrine manner, is involved in myocardial remodeling by hypertrophic stimuli. Images PMID:7929822

  9. Transforming growth factor: beta signaling is essential for limb regeneration in axolotls.

    PubMed

    Lévesque, Mathieu; Gatien, Samuel; Finnson, Kenneth; Desmeules, Sophie; Villiard, Eric; Pilote, Mireille; Philip, Anie; Roy, Stéphane

    2007-01-01

    Axolotls (urodele amphibians) have the unique ability, among vertebrates, to perfectly regenerate many parts of their body including limbs, tail, jaw and spinal cord following injury or amputation. The axolotl limb is the most widely used structure as an experimental model to study tissue regeneration. The process is well characterized, requiring multiple cellular and molecular mechanisms. The preparation phase represents the first part of the regeneration process which includes wound healing, cellular migration, dedifferentiation and proliferation. The redevelopment phase represents the second part when dedifferentiated cells stop proliferating and redifferentiate to give rise to all missing structures. In the axolotl, when a limb is amputated, the missing or wounded part is regenerated perfectly without scar formation between the stump and the regenerated structure. Multiple authors have recently highlighted the similarities between the early phases of mammalian wound healing and urodele limb regeneration. In mammals, one very important family of growth factors implicated in the control of almost all aspects of wound healing is the transforming growth factor-beta family (TGF-beta). In the present study, the full length sequence of the axolotl TGF-beta1 cDNA was isolated. The spatio-temporal expression pattern of TGF-beta1 in regenerating limbs shows that this gene is up-regulated during the preparation phase of regeneration. Our results also demonstrate the presence of multiple components of the TGF-beta signaling machinery in axolotl cells. By using a specific pharmacological inhibitor of TGF-beta type I receptor, SB-431542, we show that TGF-beta signaling is required for axolotl limb regeneration. Treatment of regenerating limbs with SB-431542 reveals that cellular proliferation during limb regeneration as well as the expression of genes directly dependent on TGF-beta signaling are down-regulated. These data directly implicate TGF-beta signaling in the

  10. Transforming Growth Factor: β Signaling Is Essential for Limb Regeneration in Axolotls

    PubMed Central

    Lévesque, Mathieu; Gatien, Samuel; Finnson, Kenneth; Desmeules, Sophie; Villiard, Éric; Pilote, Mireille; Philip, Anie; Roy, Stéphane

    2007-01-01

    Axolotls (urodele amphibians) have the unique ability, among vertebrates, to perfectly regenerate many parts of their body including limbs, tail, jaw and spinal cord following injury or amputation. The axolotl limb is the most widely used structure as an experimental model to study tissue regeneration. The process is well characterized, requiring multiple cellular and molecular mechanisms. The preparation phase represents the first part of the regeneration process which includes wound healing, cellular migration, dedifferentiation and proliferation. The redevelopment phase represents the second part when dedifferentiated cells stop proliferating and redifferentiate to give rise to all missing structures. In the axolotl, when a limb is amputated, the missing or wounded part is regenerated perfectly without scar formation between the stump and the regenerated structure. Multiple authors have recently highlighted the similarities between the early phases of mammalian wound healing and urodele limb regeneration. In mammals, one very important family of growth factors implicated in the control of almost all aspects of wound healing is the transforming growth factor-beta family (TGF-β). In the present study, the full length sequence of the axolotl TGF-β1 cDNA was isolated. The spatio-temporal expression pattern of TGF-β1 in regenerating limbs shows that this gene is up-regulated during the preparation phase of regeneration. Our results also demonstrate the presence of multiple components of the TGF-β signaling machinery in axolotl cells. By using a specific pharmacological inhibitor of TGF-β type I receptor, SB-431542, we show that TGF-β signaling is required for axolotl limb regeneration. Treatment of regenerating limbs with SB-431542 reveals that cellular proliferation during limb regeneration as well as the expression of genes directly dependent on TGF-β signaling are down-regulated. These data directly implicate TGF-β signaling in the initiation and control

  11. Diclofenac Topical (actinic keratosis)

    MedlinePlus

    ... growths on the skin caused by too much sun exposure). Diclofenac is in a class of medications called ... side effects.talk to your doctor before applying sunscreen or cosmetics to skin that is being treated ...

  12. Organized F-actin is essential for normal trichome morphogenesis in Arabidopsis.

    PubMed Central

    Szymanski, D B; Marks, M D; Wick, S M

    1999-01-01

    Actin microfilaments form a three-dimensional cytoskeletal network throughout the cell and constitute an essential throughway for organelle and vesicle transport. Development of Arabidopsis trichomes, unicellular structures derived from the epidermis, is being used as a genetic system in which to study actin-dependent growth in plant cells. The present study indicates that filamentous actin (F-actin) plays an important role during Arabidopsis trichome morphogenesis. For example, immunolocalization of actin filaments during trichome morphogenesis identified rearrangements of the cytoskeletal structure during the development of the mature cell. Moreover, pharmacological experiments indicate that there are distinct requirements for actin- and microtubule-dependent function during trichome morphogenesis. The F-actin-disrupting drug cytochalasin D does not affect the establishment of polarity during trichome development; however, maintenance and coordination of the normal pattern of cell growth are very sensitive to this drug. In contrast, oryzalin, an agent that depolymerizes microtubules, severely inhibits cell polarization. Furthermore, cytochalasin D treatment phenocopies a known class of mutations that cause distorted trichome morphology. Results of an analysis of cell shape and microfilament structure in wild-type, mutant, and drug-treated trichomes are consistent with a role for actin in the maintenance and coordination of an established growth pattern. PMID:10590162

  13. Effect of Flumorph on F-Actin Dynamics in the Potato Late Blight Pathogen Phytophthora infestans.

    PubMed

    Hua, Chenlei; Kots, Kiki; Ketelaar, Tijs; Govers, Francine; Meijer, Harold J G

    2015-04-01

    Oomycetes are fungal-like pathogens that cause notorious diseases. Protecting crops against oomycetes requires regular spraying with chemicals, many with an unknown mode of action. In the 1990s, flumorph was identified as a novel crop protection agent. It was shown to inhibit the growth of oomycete pathogens including Phytophthora spp., presumably by targeting actin. We recently generated transgenic Phytophthora infestans strains that express Lifeact-enhanced green fluorescent protein (eGFP), which enabled us to monitor the actin cytoskeleton during hyphal growth. For analyzing effects of oomicides on the actin cytoskeleton in vivo, the P. infestans Lifeact-eGFP strain is an excellent tool. Here, we confirm that flumorph is an oomicide with growth inhibitory activity. Microscopic analyses showed that low flumorph concentrations provoked hyphal tip swellings accompanied by accumulation of actin plaques in the apex, a feature reminiscent of tips of nongrowing hyphae. At higher concentrations, swelling was more pronounced and accompanied by an increase in hyphal bursting events. However, in hyphae that remained intact, actin filaments were indistinguishable from those in nontreated, nongrowing hyphae. In contrast, in hyphae treated with the actin depolymerizing drug latrunculin B, no hyphal bursting was observed but the actin filaments were completely disrupted. This difference demonstrates that actin is not the primary target of flumorph. PMID:25496300

  14. Protective effect of transforming growth factor beta 1 on experimental autoimmune diseases in mice.

    PubMed

    Kuruvilla, A P; Shah, R; Hochwald, G M; Liggitt, H D; Palladino, M A; Thorbecke, G J

    1991-04-01

    Interleukin 1 (IL-1) and tumor necrosis factor alpha are thought to contribute to the inflammatory response associated with autoimmune diseases. Transforming growth factor beta 1 (TGF-beta 1) counteracts many effects of these cytokines and has various immunosuppressive properties. In the present study, it is shown that microgram amounts of TGF-beta 1, injected daily for 1-2 weeks, protect against collagen-induced arthritis (CIA) and relapsing experimental allergic encephalomyelitis (REAE), the animal models for rheumatoid arthritis and multiple sclerosis, respectively. When administered during induction of the disease, TGF-beta 1 prevents CIA but only delays the onset of REAE by 2-3 days. However, when administered during a remission. TGF-beta 1 prevents the occurrence of relapses in REAE. The results suggest that TGF-beta 1 has powerful anti-inflammatory effects, mimicking in some respects the beneficial effects of immunosuppressive drugs in these experimental models of autoimmune disease, but without discernable adverse effects. PMID:2011600

  15. Loss of transforming growth factor-beta 2 leads to impairment of central synapse function

    PubMed Central

    Heupel, Katharina; Sargsyan, Vardanush; Plomp, Jaap J; Rickmann, Michael; Varoqueaux, Frédérique; Zhang, Weiqi; Krieglstein, Kerstin

    2008-01-01

    Background The formation of functional synapses is a crucial event in neuronal network formation, and with regard to regulation of breathing it is essential for life. Members of the transforming growth factor-beta (TGF-β) superfamily act as intercellular signaling molecules during synaptogenesis of the neuromuscular junction of Drosophila and are involved in synaptic function of sensory neurons of Aplysia. Results Here we show that while TGF-β2 is not crucial for the morphology and function of the neuromuscular junction of the diaphragm muscle of mice, it is essential for proper synaptic function in the pre-Bötzinger complex, a central rhythm organizer located in the brainstem. Genetic deletion of TGF-β2 in mice strongly impaired both GABA/glycinergic and glutamatergic synaptic transmission in the pre-Bötzinger complex area, while numbers and morphology of central synapses of knock-out animals were indistinguishable from their wild-type littermates at embryonic day 18.5. Conclusion The results demonstrate that TGF-β2 influences synaptic function, rather than synaptogenesis, specifically at central synapses. The functional alterations in the respiratory center of the brain are probably the underlying cause of the perinatal death of the TGF-β2 knock-out mice. PMID:18854036

  16. Association of interleukin 10 and transforming growth factor β gene polymorphisms with chronic idiopathic urticaria.

    PubMed

    Tavakol, Marzieh; Movahedi, Masoud; Amirzargar, Ali Akbar; Aryan, Zahra; Bidoki, Alireza Zare; Heidari, Kimia; Soltani, Samaneh; Gharagozlou, Mohammad; Aghamohammadi, Asghar; Nabavi, Mohammad; Nasiri, Rasoul; Ahmadvand, Alireza; Rezaei, Nima

    2014-01-01

    Transforming growth factor β (TGF-β) and interleukin 10 (IL-10) are two anti-inflammatory cytokines that are implicated in the pathogenesis of urticaria. The goal of this study was to examine the possible association of polymorphisms of TGF-β and IL-10 genes with susceptibility to chronic idiopathic urticaria (CIU). This study was conducted on 90 patients with CIU. Polymerase chain reaction (PCR) was done to determine the genotype at 5 polymorphic sites; TGF-β (codon10C/T and codon25G/C) and IL-10 (-1082G/A, -819C/T, and -592C/A). The C allele at codon 25 of TGF-β was more prevalent in CIU patients compared to controls (OR = 9.5, 95% CI = 5.4-16.8, P<0.001). Genotypes of CT and CG at 10 and 25 codons of TGF-β gene, respectively, and AG, CT, and CA for loci of -1082, -819, and -592 of IL-10 gene were significantly higher in CIU patients (P<0.001). In haplotype analysis, frequency of TGF-β haplotypes differed between patients with CIU and controls; CC haplotype was overrepresented, while CG and TG haplotypes were underrepresented (P<0.001). These results suggest that TGF-β and IL-10 genetic variability could contribute to susceptibility to CIU. Additionally, patients with CIU seem to have genotypes leading to high production of TGF-β and IL-10. PMID:25580782

  17. Mitotic Asynchrony Induces Transforming Growth Factor-β1 Secretion from Airway Epithelium

    PubMed Central

    Alcala, Sarah E.; Benton, Angela S.; Watson, Alan M.; Kureshi, Suraiya; Reeves, Erica M. K.; Damsker, Jesse; Wang, Zuyi; Nagaraju, Kanneboyina; Anderson, Julia; Williams, Aaron M.; Lee, Amber J. Y.; Hayes, Kathleen; Rose, Mary C.; Hoffman, Eric P.

    2014-01-01

    We recently proposed that mitotic asynchrony in repairing tissue may underlie chronic inflammation and fibrosis, where immune cell infiltration is secondary to proinflammatory cross-talk among asynchronously repairing adjacent tissues. Building on our previous finding that mitotic asynchrony is associated with proinflammatory/fibrotic cytokine secretion (e.g., transforming growth factor [TGF]-β1), here we provide evidence supporting cause-and-effect. Under normal conditions, primary airway epithelial basal cell populations undergo mitosis synchronously and do not secrete proinflammatory or profibrotic cytokines. However, when pairs of nonasthmatic cultures were mitotically synchronized at 12 hours off-set and then combined, the mixed cell populations secreted elevated levels of TGF-β1. This shows that mitotic asynchrony is not only associated with but is also causative of TGF-β1 secretion. The secreted cytokines and other mediators from asthmatic cells were not the cause of asynchronous regeneration; synchronously mitotic nonasthmatic epithelia exposed to conditioned media from asthmatic cells did not show changes in mitotic synchrony. We also tested if resynchronization of regenerating asthmatic airway epithelia reduces TGF-β1 secretion and found that pulse-dosed dexamethasone, simvastatin, and aphidicolin were all effective. We therefore propose a new model for chronic inflammatory and fibrotic conditions where an underlying factor is mitotic asynchrony. PMID:24669775

  18. Transforming growth factor-β1 in carcinogenesis, progression, and therapy in cervical cancer.

    PubMed

    Zhu, Haiyan; Luo, Hui; Shen, Zhaojun; Hu, Xiaoli; Sun, Luzhe; Zhu, Xueqiong

    2016-06-01

    Transforming growth factor β1 (TGF-β1) is a multifunctional cytokine that plays important roles in cervical tumor formation, invasion, progression, and metastasis. TGF-β1 functions as a tumor inhibitor in precancerous lesions and early stage cancers of cervix whereas as a tumor promoter in later stage. This switch from a tumor inhibitor to a tumor promoter might be due to various alterations in TGF-β signaling pathway, such as mutations or loss of expression of TGF-β receptors and SMAD proteins. Additionally, the oncoproteins of human papillomaviruses have been shown to stimulate TGF-β1 expression, which in turn suppresses host immune surveillance. Thus, in addition to driving tumor cell migration and metastasis, TGF-β1 is believed to play a key role in promoting human papillomavirus infection by weakening host immune defense. In this article, we will discuss the role of TGF-β1 in the expression, carcinogenesis, progression, and therapy in cervical cancers. A better understanding of this cytokine in cervical carcinogenesis is essential for critical evaluation of this cytokine as a potential prognostic marker and therapeutic target. PMID:27010470

  19. Metformin is a novel suppressor for transforming growth factor (TGF)-β1.

    PubMed

    Xiao, Han; Zhang, Jianshu; Xu, Zhonghe; Feng, Yenan; Zhang, Mingliang; Liu, Jianli; Chen, Ruifei; Shen, Jing; Wu, Jimin; Lu, Zhizhen; Fang, Xiaohong; Li, Jingyuan; Zhang, Youyi

    2016-01-01

    Metformin is a widely used first-line antidiabetic drug that has been shown to protect against a variety of specific diseases in addition to diabetes, including cardiovascular disorders, polycystic ovary syndrome, and cancer. However, the precise mechanisms underlying the diverse therapeutic effects of metformin remain elusive. Here, we report that transforming growth factor-β1 (TGF-β1), which is involved in the pathogenesis of numerous diseases, is a novel target of metformin. Using a surface plasmon resonance-based assay, we identified the direct binding of metformin to TGF-β1 and found that metformin inhibits [(125)I]-TGF-β1 binding to its receptor. Furthermore, based on molecular docking and molecular dynamics simulations, metformin was predicted to interact with TGF-β1 at its receptor-binding domain. Single-molecule force spectroscopy revealed that metformin reduces the binding probability but not the binding force of TGF-β1 to its type II receptor. Consequently, metformin suppresses type II TGF-β1 receptor dimerization upon exposure to TGF-β1, which is essential for downstream signal transduction. Thus, our results indicate that metformin is a novel TGF-β suppressor with therapeutic potential for numerous diseases in which TGF-β1 hyperfunction is indicated. PMID:27349853

  20. Overexpressed homeobox B9 regulates oncogenic activities by transforming growth factor-β1 in gliomas

    SciTech Connect

    Fang, Liping; Xu, Yinghui; Zou, Lijuan

    2014-03-28

    Highlights: • HOXB9 is overexpressed in gliomas. • HOXB9 over expression had shorter survival time than down expression in gliomas. • HOXB9 stimulated the proliferation, migration and sphere formation of glioma cells. • Activation of TGF-β1 contributed to HOXB9-induced oncogenic activities. - Abstract: Glioma is the leading cause of deaths related to tumors in the central nervous system. The mechanisms of gliomagenesis remain elusive to date. Homeobox B9 (HOXB9) has a crucial function in the regulation of gene expression and cell survival, but its functions in glioma formation and development have yet to be elucidated. This study showed that HOXB9 expression in glioma tissues was significantly higher than that in nontumor tissues. Higher HOXB9 expression was also significantly associated with advanced clinical stage in glioma patients. HOXB9 overexpression stimulated the proliferation, migration, and sphere formation of glioma cells, whereas HOXB9 knockdown elicited an opposite effect. HOXB9 overexpression also increased the tumorigenicity of glioma cells in vivo. Moreover, the activation of transforming growth factor-β1 contributed to HOXB9-induced oncogenic activities. HOXB9 could be used as a predictable biomarker to be detected in different pathological and histological subtypes in glioma for diagnosis or prognosis.

  1. Uptake of Host Cell Transforming Growth Factor-β by Trypanosoma cruzi Amastigotes in Cardiomyocytes

    PubMed Central

    Waghabi, Mariana C.; Keramidas, Michelle; Bailly, Sabine; Degrave, Wim; Mendonça-Lima, Leila; Soeiro, Maria de Nazaré C.; Meirelles, Maria de Nazareth L.; Paciornik, Sidnei; Araújo-Jorge, Tania C.; Feige, Jean-Jacques

    2005-01-01

    The cytokine transforming growth factor-β (TGF-β) plays various functions in the control of Trypanosoma cruzi infectivity and in the progression of Chagas’ disease. When we immunostained T. cruzi-infected cardiomyocytes (after either in vivo or in vitro infections) for TGF-β, we observed stronger immunoreactivity in parasites than in host cells. TGF-β immunoreactivity evolved during parasite cycle progression, with intense staining in amastigotes versus very faint staining in trypomastigotes. TGF-β was present on the surface of amastigotes, in the flagellar pocket, and in intraparasitic vesicles as revealed by electron microscopy. However, no ortholog TGF-β gene could be identified in the genome of T. cruzi by in silico analysis or by extensive polymerase chain reaction and reverse transcriptase-polymerase chain reaction studies. Immunoreactive TGF-β was most probably taken up by the parasite from the host cell cytoplasm because such an internalization process of biotinylated TGF-β could be observed in axenic amastigotes in vitro. These observations represent the first example of a novel mechanism by which a primitive unicellular protozoan can use host cell TGF-β to control its own intracellular life cycle. PMID:16192635

  2. Novel alleles of the transforming growth factor β-1 regulatory region and exon 1.

    PubMed

    Arrieta-Bolaños, E; Madrigal, J A; Shaw, B E

    2015-06-01

    Transforming growth factor β-1, encoded by the TGFB1 gene, is a cytokine that plays a central role in many physiologic and pathogenic processes with pleiotropic effects. Regulatory activity for this gene has been shown for 3.0 kb between positions -2665 and +423 from its translational start site. At least 17 TGFB1 regulatory region and exon 1 alleles have been defined on the basis of 18 polymorphic sites. Polymorphisms in TGFB1's regulatory region have been associated with differential levels of expression of this cytokine and to genetic risk in cancer and transplantation. In this report, we present 19 novel TGFB1 regulatory region and exon 1 alleles: p018-p036. Amplification of TGFB1's regulatory region was performed with an in-house protocol, and novel alleles were defined by either allele-specific amplification and/or molecular cloning of the amplicons, followed by sequencing in isolation. Three of these novel alleles (p018, p019, and p020) are shown to be formed by novel combinations of the aforementioned known polymorphic positions. Another 16 novel alleles are shown to carry additional known and unknown single-nucleotide polymorphisms. Polymorphism in TGFB1's regulatory region could have an impact on important processes, including embryogenesis, hematopoiesis, carcinogenesis, angiogenesis, fibrosis, immune responses, and transplantation, making its characterization necessary. PMID:25808355

  3. Transforming growth factor β1 inhibition protects from noise-induced hearing loss

    PubMed Central

    Murillo-Cuesta, Silvia; Rodríguez-de la Rosa, Lourdes; Contreras, Julio; Celaya, Adelaida M.; Camarero, Guadalupe; Rivera, Teresa; Varela-Nieto, Isabel

    2015-01-01

    Excessive exposure to noise damages the principal cochlear structures leading to hearing impairment. Inflammatory and immune responses are central mechanisms in cochlear defensive response to noise but, if unregulated, they contribute to inner ear damage and hearing loss. Transforming growth factor β (TGF-β) is a key regulator of both responses and high levels of this factor have been associated with cochlear injury in hearing loss animal models. To evaluate the potential of targeting TGF-β as a therapeutic strategy for preventing or ameliorating noise-induced hearing loss (NIHL), we studied the auditory function, cochlear morphology, gene expression and oxidative stress markers in mice exposed to noise and treated with TGF-β1 peptidic inhibitors P17 and P144, just before or immediately after noise insult. Our results indicate that systemic administration of both peptides significantly improved both the evolution of hearing thresholds and the degenerative changes induced by noise-exposure in lateral wall structures. Moreover, treatments ameliorated the inflammatory state and redox balance. These therapeutic effects were dose-dependent and more effective if the TGF-β1 inhibitors were administered prior to inducing the injury. In conclusion, inhibition of TGF-β1 actions with antagonistic peptides represents a new, promising therapeutic strategy for the prevention and repair of noise-induced cochlear damage. PMID:25852546

  4. Transforming Growth Factor-β Signaling Guides the Differentiation of Innate Lymphoid Cells in Salivary Glands.

    PubMed

    Cortez, Victor S; Cervantes-Barragan, Luisa; Robinette, Michelle L; Bando, Jennifer K; Wang, Yaming; Geiger, Theresa L; Gilfillan, Susan; Fuchs, Anja; Vivier, Eric; Sun, Joe C; Cella, Marina; Colonna, Marco

    2016-05-17

    The signals guiding differentiation of innate lymphoid cells (ILCs) within tissues are not well understood. Salivary gland (SG) ILCs as well as liver and intestinal intraepithelial ILC1 have markers that denote tissue residency and transforming growth factor-β (TGF-β) imprinting. We deleted Tgfbr2 in cells expressing the ILC and NK marker NKp46 and found that SG ILCs were reduced in number. They lost distinct tissue markers, such as CD49a, and the effector molecules TRAIL and CD73. Expression of the transcription factor Eomes, which promotes NK cell differentiation, was elevated. Conversely, Eomes deletion in NKp46(+) cells enhanced TGF-β-imprinting of SG ILCs. Thus, TGF-β induces SG ILC differentiation by suppressing Eomes. TGF-β acted through a JNK-dependent, Smad4-independent pathway. Transcriptome analysis demonstrated that SG ILCs had characteristic of both NK cells and ILC1. Finally, TGF-β imprinting of SG ILCs was synchronized with SG development, highlighting the impact of tissue microenvironment on ILC development. PMID:27156386

  5. Transforming growth factor β1 inhibition protects from noise-induced hearing loss.

    PubMed

    Murillo-Cuesta, Silvia; Rodríguez-de la Rosa, Lourdes; Contreras, Julio; Celaya, Adelaida M; Camarero, Guadalupe; Rivera, Teresa; Varela-Nieto, Isabel

    2015-01-01

    Excessive exposure to noise damages the principal cochlear structures leading to hearing impairment. Inflammatory and immune responses are central mechanisms in cochlear defensive response to noise but, if unregulated, they contribute to inner ear damage and hearing loss. Transforming growth factor β (TGF-β) is a key regulator of both responses and high levels of this factor have been associated with cochlear injury in hearing loss animal models. To evaluate the potential of targeting TGF-β as a therapeutic strategy for preventing or ameliorating noise-induced hearing loss (NIHL), we studied the auditory function, cochlear morphology, gene expression and oxidative stress markers in mice exposed to noise and treated with TGF-β1 peptidic inhibitors P17 and P144, just before or immediately after noise insult. Our results indicate that systemic administration of both peptides significantly improved both the evolution of hearing thresholds and the degenerative changes induced by noise-exposure in lateral wall structures. Moreover, treatments ameliorated the inflammatory state and redox balance. These therapeutic effects were dose-dependent and more effective if the TGF-β1 inhibitors were administered prior to inducing the injury. In conclusion, inhibition of TGF-β1 actions with antagonistic peptides represents a new, promising therapeutic strategy for the prevention and repair of noise-induced cochlear damage. PMID:25852546

  6. E1A inhibits transforming growth factor-beta signaling through binding to Smad proteins.

    PubMed

    Nishihara, A; Hanai, J; Imamura, T; Miyazono, K; Kawabata, M

    1999-10-01

    Smads form a recently identified family of proteins that mediate intracellular signaling of the transforming growth factor (TGF)-beta superfamily. Smads bind to DNA and act as transcriptional regulators. Smads interact with a variety of transcription factors, and the interaction is likely to determine the target specificity of gene induction. Smads also associate with transcriptional coactivators such as p300 and CBP. E1A, an adenoviral oncoprotein, inhibits TGF-beta-induced transactivation, and the ability of E1A to bind p300/CBP is required for the inhibition. Here we determined the Smad interaction domain (SID) in p300 and found that two adjacent regions are required for the interaction. One of the regions is the C/H3 domain conserved between p300 and CBP, and the other is a nonconserved region. p300 mutants containing SID inhibit transactivation by TGF-beta in a dose-dependent manner. E1A inhibits the interaction of Smad3 with a p300 mutant that contains SID but lacks the E1A binding domain. We found that E1A interacts specifically with receptor-regulated Smads, suggesting a novel mechanism whereby E1A antagonizes TGF-beta signaling. PMID:10497242

  7. Targeting the Transforming Growth Factor-β Signaling Pathway in Human Cancer

    PubMed Central

    Nagaraj, Nagathihalli S

    2009-01-01

    The transforming growth factor-β (TGF-β) signaling pathway plays a pivotal role in diverse cellular processes. TGF-β switches its role from tumor suppressor in normal or dysplastic cells to a tumor promoter in advanced cancers. It is widely believed that Smad-dependent pathway is involved in TGF-β tumor suppressive functions, whereas activation of Smad-independent pathways coupled with the loss of tumor suppressor functions of TGF-β is important for its pro-oncogenic functions. TGF-β signaling has been considered as a very suitable therapeutic target. The discovery of oncogenic actions of TGF-β has generated a great deal of enthusiasm for developing TGF-β signaling inhibitors for the treatment of cancer. The challenge is to identify the group of patients where targeted tumors are not only refractory to TGF-β-induced tumor suppressor functions but also responsive to tumor promoting effects of TGF-β. TGF-β pathway inhibitors including small and large molecules have now entered clinical trials. Preclinical studies with these inhibitors have shown promise in a variety of different tumor models. Here we emphasize on the mechanisms of signaling and specific targets of the TGF-β pathway that are critical effectors of tumor progression and invasion. This report also focuses on the therapeutic intervention of TGF-β signaling in human cancers. PMID:20001556

  8. Roles of transforming growth factor β in hyperoxia-induced lung injury.

    PubMed

    Liu, Y; Mao, Y F; Zheng, J; Liu, K; Han, C H; Liu, W W

    2016-01-01

    Hyperoxia induced lung injury (HILI) refers to the acute lung injury secondary to prolonged exposure to hyperoxia at elevated partial pressure. With the advent of efficient systems for delivery of high concentrations of oxygen in hospitals, the population at risk for this condition has been markedly increased. Although numerous studies have been conducted to investigate the pathogenesis of HILI, the specific mechanism is still poorly understood and some hypotheses have been proposed. Transforming growth factor β (TGF-β) is a secreted protein that controls proliferation, cellular differentiation and other functions in most cells and is a type of cytokine that plays a role in many diseases. In this mini-review, we summarize the role of TGF-β in HILI according to its relationships with reactive oxygen species (ROS), pro-inflammatory cytokines, cell apoptosis and pulmonary fibrosis. We hope it may help the understanding of pathogenesis of HILI and provide a greater understanding for the target therapy of HILI. PMID:27416690

  9. Role of Ras signaling in the induction of snail by transforming growth factor-beta.

    PubMed

    Horiguchi, Kana; Shirakihara, Takuya; Nakano, Ayako; Imamura, Takeshi; Miyazono, Kohei; Saitoh, Masao

    2009-01-01

    The epithelial-mesenchymal transition (EMT) is a crucial morphological event that occurs during the progression of epithelial tumors. EMT can be induced by transforming growth factor (TGF)-beta in some tumor cells. Here, we demonstrate the molecular mechanism whereby Snail, a key regulator of EMT, is induced by TGF-beta in tumor cells. Snail induction by TGF-beta was highly dependent on cooperation with active Ras signals, and silencing of Ras abolished Snail induction by TGF-beta in pancreatic cancer Panc-1 cells. Transfection of constitutively active Ras into HeLa cells led to induction of Snail by TGF-beta, while representative direct targets of TGF-beta, including Smad7 and PAI-1, were not affected by Ras signaling. Using mitogen-activated protein kinase inhibitors or Smad3 or Smad2 mutants, we found that phosphorylation at the linker region of Smad2/3 was not required for the induction of Snail by TGF-beta. Taken together, these findings indicate that Ras and TGF-beta-Smad signaling selectively cooperate in the induction of Snail, which occurs in a Smad-dependent manner, but independently of phosphorylation at the linker region of R-Smads by Ras signaling. PMID:19010789

  10. Effect of transforming growth factor-β3 on mono and multilayer chondrocytes.

    PubMed

    Sefat, Farshid; Youseffi, Mansour; Khaghani, Seyed Ali; Soon, Chin Fhung; Javid, Farideh

    2016-07-01

    Articular cartilage is an avascular and flexible connective tissue found in joints. It produces a cushioning effect at the joints and provides low friction to protect the ends of the bones from wear and tear/damage. It has poor repair capacity and any injury can result pain and loss of mobility. Transforming growth factor-beta (TGF-β), a cytokine superfamily, regulates cell function, including differentiation and proliferation. Although the function of the TGF-βs in various cell types has been investigated, their function in cartilage repair is as yet not fully understood. The effect of TGF-β3 in biological regulation of primary chondrocyte was investigated in this work. TGF-β3 provided fibroblastic morphology to chondrocytes and therefore overall reduction in cell proliferation was observed. The length of the cells supplemented with TGF-β3 were larger than the cells without TGF-β3 treatment. This was caused by the fibroblast like cells (dedifferentiated chondrocytes) which occupied larger areas compared to cells without TGF-β3 addition. The healing process of the model wound closure assay of chondrocyte multilayer was slowed down by TGF-β3, and this cytokine negatively affected the strength of chondrocyte adhesion to the cell culture surface. PMID:27108397

  11. Immunocytochemical Localization of Latent Transforming Growth Factor-B1 Activation by Stimulated Macrophages

    SciTech Connect

    Chong, Hyonkyong; Vodovotz, Yoram; Cox, G.W.; Barcellos-Hoff, M.H.

    1998-09-22

    Transforming growth factor-{beta}1 (TGF-{beta}) is secreted in a latent form consisting of mature TGF-{beta} noncovalently associated with its amino-terminal propeptide, which is called latency associated peptide (LAP). Biological activity depends upon the release of TGF-{beta} from the latent complex following extracellular activation, which appears to be the key regulatory mechanism controlling TGF-{beta} action. We have identified two events associated with latent TGF-{beta} (LTGF-{beta}) activation in vivo: increased immunoreactivity of certain antibodies that specifically detect TGF-{beta} concomitant with decreased immunoreactivity of antibodies to LAP. Macrophages stimulated in vitro with interferon-{gamma} and lipopolysaccharide reportedly activate LTGF-{beta} via cell membrane-bound protease activity. We show through dual immunostaining of paraformaldehyde-fixed macrophages that such physiological TGF-{beta} activation is accompanied by a loss of LAP immunoreactivity with concomitant revelation of TGF-{beta} epitopes. The induction of TGF-{beta} immunoreactivity colocalized with immunoreactive betaglycan/RIII in activated macrophages, suggesting that LTGF-{beta} activation occurs on the cell surface. Confocal microscopy of metabolically active macrophages incubated with antibodies to TGF-{beta} and betaglycan/RIII prior to fixation supported the localization of activation to the cell surface. The ability to specifically detect and localize LTGF-{beta} activation provides an important tool for studies of its regulation.

  12. Transforming growth factor-beta 1 in experimental autoimmune neuritis. Cellular localization and time course.

    PubMed Central

    Kiefer, R.; Funa, K.; Schweitzer, T.; Jung, S.; Bourde, O.; Toyka, K. V.; Hartung, H. P.

    1996-01-01

    Experimental autoimmune neuritis (EAN) is a monophasic inflammatory disorder of the peripheral nervous system that resolves spontaneously by molecular mechanisms as yet unknown. We have investigated whether the immunosuppressive cytokine transforming growth factor-beta 1 (TGF-beta 1) might be endogenously expressed in the peripheral nervous system of Lewis rats with actively induced and adoptive transfer EAN. TGF-beta 1 mRNA was upregulated to high levels in sensory and motor roots, spinal ganglia, and sciatic nerve as revealed by quantitative Northern blot analysis and in situ hybridization histochemistry, with peak levels just preceding the first signs of clinical recovery. TGF-beta 1 mRNA was localized to scattered round cells and dense cellular infiltrates, but only rarely to Schwann cell profiles. Double labeling studies revealed macrophages and subpopulations of T cells as the major cellular source of TGF-beta 1 mRNA. TGF-beta 1 protein was visualized immunocytochemically and localized to infiltrating mononuclear cells with peak expression around the same time as mRNA, in addition to some constitutive expression in axons and Schwann cells. Our studies suggest that the spontaneous recovery observed in Lewis rat EAN might be mediated by the endogenous elaboration of TGF-beta 1 within the peripheral nerve, and that macrophages might control their own cytotoxicity by expressing TGF-beta 1. Images Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 PMID:8546208

  13. Immunocytochemical localization of latent transforming growth factor-beta1 activation by stimulated macrophages

    NASA Technical Reports Server (NTRS)

    Chong, H.; Vodovotz, Y.; Cox, G. W.; Barcellos-Hoff, M. H.; Chatterjee, A. (Principal Investigator)

    1999-01-01

    Transforming growth factor-beta1 (TGF-beta) is secreted in a latent form consisting of mature TGF-beta noncovalently associated with its amino-terminal propeptide, which is called latency associated peptide (LAP). Biological activity depends upon the release of TGF-beta from the latent complex following extracellular activation, which appears to be the key regulatory mechanism controlling TGF-beta action. We have identified two events associated with latent TGF-beta (LTGF-beta) activation in vivo: increased immunoreactivity of certain antibodies that specifically detect TGF-beta concomitant with decreased immunoreactivity of antibodies to LAP. Macrophages stimulated in vitro with interferon-gamma and lipopolysaccharide reportedly activate LTGF-beta via cell membrane-bound protease activity. We show through dual immunostaining of paraformaldehyde-fixed macrophages that such physiological TGF-beta activation is accompanied by a loss of LAP immunoreactivity with concomitant revelation of TGF-beta epitopes. The induction of TGF-beta immunoreactivity colocalized with immunoreactive betaglycan/RIII in activated macrophages, suggesting that LTGF-beta activation occurs on the cell surface. Confocal microscopy of metabolically active macrophages incubated with antibodies to TGF-beta and betaglycan/RIII prior to fixation supported the localization of activation to the cell surface. The ability to specifically detect and localize LTGF-beta activation provides an important tool for studies of its regulation.

  14. Tissue localization of transforming growth factor-beta1 in pulmonary eosinophilic granuloma.

    PubMed

    Asakura, S; Colby, T V; Limper, A H

    1996-11-01

    Pulmonary eosinophilic granuloma is characterized by infiltration of the lungs with fibronodular lesions containing specialized Langerhans' cells. In some patients, progressive pulmonary fibrosis leads to significant respiratory impairment. Transforming growth factor-beta1 (TGF-beta1) promotes fibrosis by enhancing the synthesis of extracellular matrix components. The role of TGF-beta1 in promoting fibrosis in the setting of pulmonary eosinophilic granuloma is currently unknown. We used immunohistochemistry to evaluate the extent and distribution of TGF-beta1 and the extracellular matrix components type I collagen and decorin, a TGF-beta1-binding proteoglycan. Lung biopsies from 11 patients with pulmonary eosinophilic granuloma were evaluated. In biopsies with active inflammatory lesions containing Langerhans' cells, hyperplastic type 2 pneumocytes and alveolar macrophages within and surrounding the fibronodular lesions contained abundant TGF-beta1. Langerhans' cells were consistently devoid of immunoreactive TGF-beta1. Active inflammatory lesions also exhibited staining for decorin, in a loosely organized distribution. Advanced fibrotic lesions of eosinophilic granuloma, containing minimal inflammatory cells and few or no Langerhans' cells, exhibited weak or absent staining for TGF-beta1 within either hyperplastic type 2 pneumocytes or alveolar macrophages. The fibroconnective tissues of these advanced fibrotic lesions consistently revealed dense staining for decorin. Through their actions on extracellular matrix protein accumulation, TGF-beta1 and the TGF-beta1-binding proteoglycan decorin may modulate fibrotic repair accompanying pulmonary eosinophilic granuloma. PMID:8912775

  15. Elevated Transforming Growth Factor β1 in Plasma of Primary Open-Angle Glaucoma Patients

    PubMed Central

    Kuchtey, John; Kunkel, Jessica; Burgess, L. Goodwin; Parks, Megan B.; Brantley, Milam A.; Kuchtey, Rachel W.

    2014-01-01

    Purpose. To test the hypothesis that primary open-angle glaucoma (POAG) patients have a systemic elevation of transforming growth factor β1 (TGFβ1). Methods. Plasma was prepared from blood samples drawn from patients of the Vanderbilt Eye Institute during clinic visits. Concentrations of total TGFβ1 and thrombospondin-1 (TSP1) in plasma were determined by ELISA. Statistical significance of differences between POAG and control samples was evaluated by Mann-Whitney test. Regression analysis was used to evaluate correlations between plasma TGFβ1 and patient age and between plasma TGFβ1 and TSP1. Results. Plasma samples were obtained from 148 POAG patients and 150 controls. Concentration of total TGFβ1 in the plasma of POAG patients (median = 3.25 ng/mL) was significantly higher (P < 0.0001) than in controls (median = 2.46 ng/mL). Plasma TGFβ1 was not correlated with age of patient (P = 0.17). Thrombospondin-1 concentration was also significantly higher (P < 0.0001) in POAG patients (median = 0.774 μg/mL) as compared to controls (median = 0.567 μg/mL). Plasma total TGFβ1 and TSP1 concentrations were linearly correlated (P < 0.0001). Conclusions. Plasma samples from POAG patients display elevated total TGFβ1 compared to controls, consistent with elevated systemic TGFβ1 in POAG patients. PMID:25061114

  16. Requirement of HDAC6 for Transforming Growth Factor-β1-induced Epithelial-Mesenchymal Transition*

    PubMed Central

    Shan, Bin; Yao, Tso-pang; Nguyen, Hong T.; Zhuo, Ying; Levy, Dawn R.; Klingsberg, Ross C.; Tao, Hui; Palmer, Michael L.; Holder, Kevin N.; Lasky, Joseph A.

    2008-01-01

    The aberrant expression of transforming growth factor (TGF)-β1 in the tumor microenvironment and fibrotic lesions plays a critical role in tumor progression and tissue fibrosis by inducing epithelial-mesenchymal transition (EMT). EMT promotes tumor cell motility and invasiveness. How EMT affects motility and invasion is not well understood. Here we report that HDAC6 is a novel modulator of TGF-β1-induced EMT. HDAC6 is a microtubule-associated deacetylase that predominantly deacetylates nonhistone proteins, including α-tubulin, and regulates cell motility. We showed that TGF-β1-induced EMT is accompanied by HDAC6-dependent deacetylation of α-tubulin. Importantly, inhibition of HDAC6 by small interfering RNA or the small molecule inhibitor tubacin attenuated the TGF-β1-induced EMT markers, such as the aberrant expression of epithelial and mesenchymal peptides, as well as the formation of stress fibers. Reduced expression of HDAC6 also impaired the activation of SMAD3 in response to TGF-β1. Conversely, inhibition of SMAD3 activation substantially impaired HDAC6-dependent deacetylation of α-tubulin as well as the expression of EMT markers. These findings reveal a novel function of HDAC6 in EMT by intercepting the TGF-β-SMAD3 signaling cascade. Our results identify HDAC6 as a critical regulator of EMT and a potential therapeutic target against pathological EMT, a key event for tumor progression and fibrogenesis. PMID:18499657

  17. Transforming growth factor beta (TGF-β) isomers influence cell detachment of MG-63 bone cells.

    PubMed

    Sefat, Farshid; Khaghani, Seyed Ali; Nejatian, Touraj; Genedy, Mohammed; Abdeldayem, Ali; Moghaddam, Zoha Salehi; Denyer, Morgan C T; Youseffi, Mansour

    2015-12-01

    Bone repair and wound healing are modulated by different stimuli. There is evidence that Transforming Growth Factor-beta (TGF-β) super-family of cytokines have significant effects on bone structure by regulating the replication and differentiation of chondrocytes, osteoblasts and osteoclasts. There is also significant evidence that interactions with extracellular matrix molecules influence cell behaviour. In this study cell surface attachment was examined via a trypsinization assay using various TGF-β isomers in which the time taken to trypsinize cells from the surface provided a means of assessing the strength of attachment. Three TGF-β isomers (TGF-β1, 2 and 3), four combined forms (TGF-β(1+2), TGF-β(1+3), TGF-β(2+3) and TGF-β(1+2+3)) along with four different controls (BSA, HCl, BSA/HCl and negative control) were investigated in this study. The results indicated that treatment with TGF-β1, 2, 3 and HCl decreased cell attachment, however, this effect was significantly greater in the case of TGF-β3 (p<0.001) indicating perhaps that TGF-β3 does not act alone in cell detachment, but instead functions synergistically with signalling pathways that are dependent on the availability of hydrogen ions. Widefield Surface Plasmon Resonance (WSPR) microscope was also used to investigate cell surface interactions. PMID:26372305

  18. Effects of transforming growth factor beta 1 on the regulation of osteoclastic development and function

    SciTech Connect

    Hattersley, G.; Chambers, T.J. )

    1991-02-01

    Transforming growth factor (TGF) beta 1 is a multifunctional cytokine with powerful effects on osteoblastic cells. Its role in the regulation of osteoclast generation and function, however, is unclear. It has been reported both to stimulate and to inhibit resorption in organ culture and to inhibit multinuclear cell formation in bone marrow cultures. We tested the effects of TGF-beta 1 on bone resorption by osteoclasts isolated from neonatal rat long bones. We found potent stimulation of osteoclastic bone resorption, mediated by osteoblastic cells, with an EC50 of 10 pg/ml, considerably lower than that of well-documented osteotropic hormones. Stimulation was not mediated by Swiss mouse 3T3 cells, a nonosteoblastic cell line. TGF-beta 1 strongly inhibited the generation of calcitonin receptor (CTR)-positive cells in mouse bone marrow cultures, but as for isolated osteoclasts, bone resorption per CTR-positive cell was increased. The inhibition of CTR-positive cell formation was associated with suppression of maturation of other bone marrow derivatives and may be related more to the known ability of TGF-beta 1 to suppress the proliferation of primitive hematopoietic cells than to a specific role of TGF-beta 1 in osteoclast generation.

  19. The type III transforming growth factor beta receptor regulates vascular and osteoblast development during palatogenesis

    PubMed Central

    Hill, Cynthia R.; Jacobs, Britni H.; Brown, Christopher B.; Barnett, Joey V.; Goudy, Steven L.

    2015-01-01

    Background Cleft palate occurs in up to 1:1000 live births and is associated with mutations in multiple genes. Palatogenesis involves a complex choreography of palatal shelf elongation, elevation, and fusion. Transforming growth factor β (TGFβ) and bone morphogenetic protein 2 (BMP2) canonical signaling is required during each stage of palate development. The type III TGFβ receptor (TGFβR3) binds all three TGFβ ligands and BMP2, but its contribution to palatogenesis is unknown. Results The role of TGFβR3 during palate formation was found to be during palatal shelf elongation and elevation. Tgfbr3-/- embryos displayed reduced palatal shelf width and height, changes in proliferation and apoptosis, and reduced vascular and osteoblast differentiation. Abnormal vascular plexus organization as well as aberrant expression of arterial (Notch1, Alk1), venous (EphB4), and lymphatic (Lyve1) markers was also observed. Decreased osteoblast differentiation factors (Runx2, alk phos, osteocalcin, col1A1, and col1A2) demonstrated poor mesenchymal cell commitment to the osteoblast lineage within the maxilla and palatal shelves in Tgfbr3-/- embryos. Additionally, in vitro bone mineralization induced by osteogenic medium (OM+BMP2) was insufficient in Tgfbr3-/- palatal mesenchyme, but mineralization was rescued by overexpression of TGFβR3. Conclusions These data reveal a critical, previously unrecognized role for TGFβR3 in vascular and osteoblast development during palatogenesis. PMID:25382630

  20. Transforming Growth Factor {beta} Can Stimulate Smad1 Phosphorylation Independently of Bone Morphogenic Protein Receptors.

    PubMed

    Wrighton, Katharine H; Lin, Xia; Yu, Paul B; Feng, Xin-Hua

    2009-04-10

    Transforming growth factor-beta (TGFbeta) superfamily ligands control a diverse set of cellular processes by activating type I and type II serine-threonine receptor kinases. Canonical TGFbeta signaling is mediated via the TbetaRI/ALK5 type I receptor that phosphorylates Smad2 and Smad3 in their SXS motif to facilitate their activation and subsequent role in transcriptional regulation. Canonical bone morphogenic protein (BMP) signaling is mediated via the ALK1/2/3/6 type I receptors that phosphorylate Smad1, Smad5, and Smad8 in their SXS motif. However, studies in endothelial cells have shown that TGFbeta can also lead to the phosphorylation of Smad1, dependent on ALK1 receptor activity. Here we present data showing that TGFbeta can significantly induce Smad1 phosphorylation in several non-endothelial cell lineages. Additionally, by using chemical inhibitors specific for the TGFbeta/activin/nodal (ALK4/5/7) and BMP (ALK1/2/3/6) type I receptors, we show that in some cell types TGFbeta induces Smad1 phosphorylation independently of the BMP type I receptors. Thus, TGFbeta-mediated Smad1 phosphorylation appears to occur via different receptor complexes in a cell type-specific manner. PMID:19224917

  1. MTMR4 attenuates transforming growth factor beta (TGFbeta) signaling by dephosphorylating R-Smads in endosomes.

    PubMed

    Yu, Junjing; Pan, Lei; Qin, Xincheng; Chen, Hua; Xu, Youli; Chen, Yeguang; Tang, Hong

    2010-03-12

    Homeostasis of Smad phosphorylation at its C-terminal SXS motif is essential for transforming growth factor beta (TGFbeta) signaling. Whereas it is known that TGFbeta signaling can be terminated by phosphatases, which dephosphorylate R-Smads in the nucleus, it is unclear whether there are any cytoplasmic phosphatase(s) that can attenuate R-Smad phosphorylation and nuclear translocation. Here we demonstrate that myotubularin-related protein 4 (MTMR4), a FYVE domain-containing dual-specificity protein phosphatase (DSP), attenuates TGFbeta signaling by reducing the phosphorylation level of R-Smads in early endosomes. Co-immunoprecipitation experiments showed that endogenous MTMR4 interacts with phosphorylated R-Smads, and that this interaction is correlated with dephosphorylation of R-Smads. Further analysis showed that overexpression of MTMR4 resulted in the sequestration of activated Smad3 in the early endosomes, thus reducing its nuclear translocation. However, both point mutations at the conserved catalytic site of the phosphatase (MTMR4-C407S) and small interference RNA of endogenous Mtmr4 expression led to sustained Smad3 activation. This work therefore suggests that MTMR4 plays an important role in preventing the overactivation of TGFbeta signaling by dephosphorylating activated R-Smads that have been trafficked to early endosomes. PMID:20061380

  2. Hepatic transforming growth factor-β 1 stimulated clone-22 D1 controls systemic cholesterol metabolism☆

    PubMed Central

    Jäger, Julia; Greiner, Vera; Strzoda, Daniela; Seibert, Oksana; Niopek, Katharina; Sijmonsma, Tjeerd P.; Schäfer, Michaela; Jones, Allan; De Guia, Roldan; Martignoni, Marc; Dallinga-Thie, Geesje M.; Diaz, Mauricio B.; Hofmann, Thomas G.; Herzig, Stephan

    2014-01-01

    Disturbances in lipid homeostasis are hallmarks of severe metabolic disorders and their long-term complications, including obesity, diabetes, and atherosclerosis. Whereas elevation of triglyceride (TG)-rich very-low-density lipoproteins (VLDL) has been identified as a risk factor for cardiovascular complications, high-density lipoprotein (HDL)-associated cholesterol confers atheroprotection under obese and/or diabetic conditions. Here we show that hepatocyte-specific deficiency of transcription factor transforming growth factor β 1-stimulated clone (TSC) 22 D1 led to a substantial reduction in HDL levels in both wild-type and obese mice, mediated through the transcriptional down-regulation of the HDL formation pathway in liver. Indeed, overexpression of TSC22D1 promoted high levels of HDL cholesterol in healthy animals, and hepatic expression of TSC22D1 was found to be aberrantly regulated in disease models of opposing energy availability. The hepatic TSC22D1 transcription factor complex may thus represent an attractive target in HDL raising strategies in obesity/diabetes-related dyslipidemia and atheroprotection. PMID:24634828

  3. Association of Transforming Growth Factor Alpha Polymorphisms with Nonsyndromic Cleft Lip and Palate in Iranian Population

    PubMed Central

    Ebadifar, Asghar; Hamedi, Roya; Khorram Khorshid, Hamid Reza; Saliminejad, Kioomars; Kamali, Koorosh; Aghakhani Moghadam, Fatemeh; Esmaeili Anvar, Nazanin; Ameli, Nazilla

    2015-01-01

    Background: Cleft lip with or without cleft palate (CL/P) is one of the most common congenital anomalies and the etiology of orofacial clefts is multifactorial. Transforming growth factor alpha (TGFA) is expressed at the medial edge epithelium of fusing palatal shelves during craniofacial development. In this study, the association of two important TGFA gene polymorphisms, BamHI (rs11466297) and RsaI (rs3732248), with CL/P was evaluated in an Iranian population. Methods: The frequencies of BamHI and RsaI variations were determined in 105 unrelated Iranian subjects with nonsyndromic CL/P and 218 control subjects using PCR and RFLP methods, and the results were compared with healthy controls. A p-value of <0.05 was considered statistically significant. Results: The BamHI AC genotype was significantly higher (p=0.016) in the patients (12.4%) than the control group (5.0%). The BamHI C allele was significantly higher (p=0.001; OR=3.4, 95% CI: 1.6–7.4) in the cases (8.0%) compared with the control group (2.5%). Conclusion: Our study showed that there was an association between the TGFA BamHI variation and nonsyndromic CL/P in Iranian population. PMID:26605011

  4. Transforming growth factor receptor type II (ec-TβR II) behaves as a halophile.

    PubMed

    Saini, Komal; Khan, M Ashhar I; Chakrapani, Sumit; Deep, Shashank

    2015-01-01

    The members of transforming growth factor β family (TGF-β) are multifunctional proteins but their main role is to control cell proliferation and differentiation. Polypeptides of TGF-β family function by binding to two related, functionally distinct transmembrane receptor kinases, first to the type II (TβR II) followed by type I receptor (TβR I). The paper describes, in details, the stability of wt-ec-TβR II under different conditions. The stability of wt-ec-TβR II was observed at different pH and salt concentration using fluorescence spectroscopy. Stability of ec-TβR II decreases with decrease in pH. Interestingly, the addition of salt increases the stability of the TβRII at pH 5.0 as observed for halophiles. Computational analysis using DELPHI suggests that this is probably due to the decrease in repulsion between negatively charged residues at surface on the addition of salt. This is further confirmed by the change in the stability of receptor on mutation of some of the residues (D32A) at surface. PMID:25316422

  5. Transforming growth factor-beta. Murine glomerular receptors and responses of isolated glomerular cells.

    PubMed Central

    MacKay, K; Striker, L J; Stauffer, J W; Doi, T; Agodoa, L Y; Striker, G E

    1989-01-01

    Proliferation of resident glomerular cells and the accumulation of mesangial matrix are histologic abnormalities which are observed in the course of many progressive glomerular diseases. We explored the potential regulatory effects of transforming growth factor-beta (TGF-beta) on these processes. We found that cultured mouse glomerular endothelial, mesangial, and epithelial cells as well as isolated intact rat glomeruli possess high-affinity receptors for TGF-beta. We also found that, although TGF-beta consistently inhibited the proliferation of glomerular endothelial and epithelial cells, it acted as a bifunctional regulator of mesangial cell proliferation. TGF-beta significantly increased the production of collagen and fibronectin by glomerular mesangial cells whereas only fibronectin production was augmented in glomerular epithelial cells. The presence of TGF-beta receptors on intact glomeruli and on each glomerular cell type and the demonstrated responsiveness of these cells to TGF-beta combine to suggest that potentially important interactions may occur between resident glomerular cells and TGF-beta in vivo. Images PMID:2539392

  6. Integration of sexual trauma in a religious narrative: Transformation, resolution and growth among contemplative nuns

    PubMed Central

    Littlewood, Roland; Leavey, Gerard

    2013-01-01

    The psychological consequences of sexual abuse are generally serious and enduring, particularly when the perpetrator is known and trusted by the survivor. This paper explores the experiences of five contemplative nuns who were sexually abused by priests and the spiritual journeys that followed. In the context of an ethnographic study of contemplative practice, participant observation and in-depth interviews were used to examine the ways that the nuns sought to make sense of their experiences through a long process of solitary introspection. The pursuit of meaning was shaped by religious beliefs relating to forgiveness, sacrifice, and salvation. Thus, trauma was transformed into a symbolic religious narrative that shaped their sense of identity. They were able to restructure core beliefs and to manage their current relationships with priests more securely. They described regaining their spiritual well-being in ways that suggest a form of posttraumatic spiritual growth. We conclude by discussing the findings in the light of the existing literature on the interaction of trauma and spirituality. PMID:23296289

  7. Metformin is a novel suppressor for transforming growth factor (TGF)-β1

    NASA Astrophysics Data System (ADS)

    Xiao, Han; Zhang, Jianshu; Xu, Zhonghe; Feng, Yenan; Zhang, Mingliang; Liu, Jianli; Chen, Ruifei; Shen, Jing; Wu, Jimin; Lu, Zhizhen; Fang, Xiaohong; Li, Jingyuan; Zhang, Youyi

    2016-06-01

    Metformin is a widely used first-line antidiabetic drug that has been shown to protect against a variety of specific diseases in addition to diabetes, including cardiovascular disorders, polycystic ovary syndrome, and cancer. However, the precise mechanisms underlying the diverse therapeutic effects of metformin remain elusive. Here, we report that transforming growth factor-β1 (TGF-β1), which is involved in the pathogenesis of numerous diseases, is a novel target of metformin. Using a surface plasmon resonance-based assay, we identified the direct binding of metformin to TGF-β1 and found that metformin inhibits [125I]-TGF-β1 binding to its receptor. Furthermore, based on molecular docking and molecular dynamics simulations, metformin was predicted to interact with TGF-β1 at its receptor-binding domain. Single-molecule force spectroscopy revealed that metformin reduces the binding probability but not the binding force of TGF-β1 to its type II receptor. Consequently, metformin suppresses type II TGF-β1 receptor dimerization upon exposure to TGF-β1, which is essential for downstream signal transduction. Thus, our results indicate that metformin is a novel TGF-β suppressor with therapeutic potential for numerous diseases in which TGF-β1 hyperfunction is indicated.

  8. Association of transforming growth-factor alpha gene polymorphisms with nonsyndromic cleft palate only (CPO)

    SciTech Connect

    Shiang, R. ); Lidral, A.C.; Ardinger, H.H.; Murray, J.C.; Romitti, P.A.; Munger, R.G.; Buetow, K.H.

    1993-10-01

    Genetic analysis and tissue-specific expression studies support a role for transforming growth-factor alpha (TGFA) in craniofacial development. Previous studies have confirmed an association of alleles for TGFA with nonsyndromic cleft lip with or without cleft palate (CL/P) in humans. The authors carried out a retrospective association study to determine whether specific allelic variants of the TGFA gene are also associated with cleft palate only (CPO). The PCR products from 12 overlapping sets of primers to the TGFA cDNA were examined by using single-strand conformational polymorphism analysis. Four DNA polymorphic sites for TGFA were identified in the 3[prime] untranslated region of the TGFA gene. These variants, as well as previously identified RFLPs for TGFA, were characterized in case and control populations for CPO by using X[sup 2] analysis. A significant association between alleles of TGFA and CPO was identified which further supports a role for this gene as one of the genetic determinants of craniofacial development. Sequence analysis of the variants disclosed a cluster of three variable sites within 30 bp of each other in the 3[prime] untranslated region previously associated with an antisense transcript. These studies extend the role for TGFA in craniofacial morphogenesis and support an interrelated mechanism underlying nonsyndromic forms of CL/P. 46 refs., 3 figs., 3 tabs.

  9. Generation of Mice With a Conditional Allele for the Transforming Growth Factor Beta3 Gene

    PubMed Central

    Doetschman, Thomas; Georgieva, Teodora; Li, Hongqi; Reed, Thomas D.; Grisham, Christina; Friel, Jacqueline; Estabrook, Mark A.; Gard, Connie; Sanford, L.P.; Azhar, Mohamad

    2013-01-01

    Summary The transforming growth factor beta (TGFβ) pathway is involved in embryonic development and several inherited and acquired human diseases. The gene for TGFβ3 (Tgfb3) encodes one of the three ligands for TGF b receptors. It is widely expressed in the embryo and its mutation or misexpression is found in human diseases. Tgfb3−/− mice die at birth from cleft palate, precluding functional studies in adults. Here, we generated mice in which exon 6 of Tgfb3 was flanked with LoxP sites (Tgfb3flox/flox). The adult mice were normal and fertile. EIIa-Cre-mediated deletion of exon 6 in Tgfb3flox/flox mice efficiently generated Tgfb3 conditional knockout (Tgfb3cko/cko) mice which died at birth from the same cleft palate defect as Tgfb3−/− mice, indicating that the conditional and knockout alleles are functionally equivalent. This Tgfb3cko allele will now enable studies of TGFβ3 function in different cell or tissue types in embryonic development and during adulthood. genesis 50:59-66, 2012. PMID:22223248

  10. Integration of sexual trauma in a religious narrative: transformation, resolution and growth among contemplative nuns.

    PubMed

    Durà-Vilà, Glòria; Littlewood, Roland; Leavey, Gerard

    2013-02-01

    The psychological consequences of sexual abuse are generally serious and enduring, particularly when the perpetrator is known and trusted by the survivor. This paper explores the experiences of five contemplative nuns who were sexually abused by priests and the spiritual journeys that followed. In the context of an ethnographic study of contemplative practice, participant observation and in-depth interviews were used to examine the ways that the nuns sought to make sense of their experiences through a long process of solitary introspection. The pursuit of meaning was shaped by religious beliefs relating to forgiveness, sacrifice, and salvation. Thus, trauma was transformed into a symbolic religious narrative that shaped their sense of identity. They were able to restructure core beliefs and to manage their current relationships with priests more securely. They described regaining their spiritual well-being in ways that suggest a form of posttraumatic spiritual growth. We conclude by discussing the findings in the light of the existing literature on the interaction of trauma and spirituality. PMID:23296289

  11. Metformin is a novel suppressor for transforming growth factor (TGF)-β1

    PubMed Central

    Xiao, Han; Zhang, Jianshu; Xu, Zhonghe; Feng, Yenan; Zhang, Mingliang; Liu, Jianli; Chen, Ruifei; Shen, Jing; Wu, Jimin; Lu, Zhizhen; Fang, Xiaohong; Li, Jingyuan; Zhang, Youyi

    2016-01-01

    Metformin is a widely used first-line antidiabetic drug that has been shown to protect against a variety of specific diseases in addition to diabetes, including cardiovascular disorders, polycystic ovary syndrome, and cancer. However, the precise mechanisms underlying the diverse therapeutic effects of metformin remain elusive. Here, we report that transforming growth factor-β1 (TGF-β1), which is involved in the pathogenesis of numerous diseases, is a novel target of metformin. Using a surface plasmon resonance-based assay, we identified the direct binding of metformin to TGF-β1 and found that metformin inhibits [125I]-TGF-β1 binding to its receptor. Furthermore, based on molecular docking and molecular dynamics simulations, metformin was predicted to interact with TGF-β1 at its receptor-binding domain. Single-molecule force spectroscopy revealed that metformin reduces the binding probability but not the binding force of TGF-β1 to its type II receptor. Consequently, metformin suppresses type II TGF-β1 receptor dimerization upon exposure to TGF-β1, which is essential for downstream signal transduction. Thus, our results indicate that metformin is a novel TGF-β suppressor with therapeutic potential for numerous diseases in which TGF-β1 hyperfunction is indicated. PMID:27349853

  12. Localization of transforming growth factor-beta at the human fetal-maternal interface: role in trophoblast growth and differentiation.

    PubMed

    Graham, C H; Lysiak, J J; McCrae, K R; Lala, P K

    1992-04-01

    We examined the localization of transforming growth factor (TGF)-beta in first-trimester and term human decidua and chorionic villi and explored the role of this factor on the proliferation and differentiation of cultured trophoblast cells. Two antibodies, 1D11.16.8, a mouse monoclonal neutralizing antibody capable of recognizing both TGF-beta 1 and TGF-beta 2 and CL-B1/29, a rabbit polyclonal antibody capable of recognizing TGF-beta 2, were used to immunolocalize TGF-beta in fixed, paraffin-embedded, or fixed, frozen sections of placenta and decidua, providing similar results. Intense labeling was observed in the extracellular matrix (ECM) of the first-trimester decidua and cytoplasm of term decidual cells. Syncytiotrophoblast cell cytoplasm as well as the ECM in the core of the chorionic villi of both first-trimester and term placentas exhibited a moderate degree of labeling. Strong cytoplasmic labeling was observed in the cytotrophoblastic shell of the term placenta. To examine the role of TGF-beta on trophoblast proliferation and differentiation, early passage cultures of first-trimester and primary cultures of term trophoblast cells were established and characterized on the basis of numerous immunocytochemical and functional markers. These cells expressed cytokeratin, placental alkaline phosphatase, urokinase-type plasminogen activator, and pregnancy-specific beta glycoprotein, but not factor VIII or 63D3; they also produced hCG and collagenase type IV. Exposure of first-trimester trophoblast cultures to TGF-beta 1 significantly inhibited proliferation in a dose-dependent manner. An antiproliferative effect was also noted in the presence of TGF-beta 2. These effects were abrogated in the presence of the neutralizing anti-TGF-beta antibody (1D11.16.8) in a concentration-dependent manner. In a 3-day culture, exogenous TGF-beta 1 stimulated formation of multinucleated cells by the first trimester as well as term trophoblast cells. Addition of neutralizing anti

  13. Transforming Growth Factor Beta 1 Augments Calvarial Defect Healing and Promotes Suture Regeneration

    PubMed Central

    Shakir, Sameer; MacIsaac, Zoe M.; Naran, Sanjay; Smith, Darren M.; Bykowski, Michael R.; Cray, James J.; Craft, Timothy K.; Wang, Dan; Weiss, Lee; Campbell, Phil G.; Mooney, Mark P.; Losee, Joseph E.

    2015-01-01

    Background: Repair of complex cranial defects is hindered by a paucity of appropriate donor tissue. Bone morphogenetic protein 2 (BMP2) and transforming growth factor beta 1 (TGFβ1) have been shown separately to induce bone formation through physiologically distinct mechanisms and potentially improve surgical outcome for cranial defect repair by obviating the need for donor tissue. We hypothesize that a combination of BMP2 and TGFβ1 would improve calvarial defect healing by augmenting physiologic osteogenic mechanisms. Methods/Results: Coronal suturectomies (3×15 mm) were performed in 10-day-old New Zealand White rabbits. DermaMatrix™ (3×15mm) patterned with four treatments (vehicle, 350 ng BMP2, 200 ng TGFβ1, or 350 ng BMP2+200 ng TGFβ1) was placed in suturectomy sites and rabbits were euthanized at 6 weeks of age. Two-dimensional (2D) defect healing, bone volume, and bone density were quantified by computed tomography. Regenerated bone was qualitatively assessed histologically. One-way analysis of variance revealed significant group main effects for all bone quantity measures. Analysis revealed significant differences in 2D defect healing, bone volume, and bone density between the control group and all treatment groups, but no significant differences were detected among the three growth factor treatment groups. Qualitatively, TGFβ1 treatment produced bone with morphology most similar to native bone. TGFβ1-regenerated bone contained a suture-like tissue, growing from the lateral edge of the defect margin toward the midline. Unique to the BMP2 treatment group, regenerated bone contained lacunae with chondrocytes, demonstrating the presence of endochondral ossification. Conclusions/Significance: Total healing in BMP2 and TGFβ1 treatment groups is not significantly different. The combination of BMP2+TGFβ1 did not significantly increase bone healing compared with treatment with BMP2 or TGFβ1 alone postoperatively at 4 weeks. We highlight the

  14. Follicle-restricted compartmentalization of transforming growth factor beta superfamily ligands in the feline ovary.

    PubMed

    Bristol, Sarah K; Woodruff, Teresa K

    2004-03-01

    Ovarian follicular development, follicle selection, and the process of ovulation remain poorly understood in most species. Throughout reproductive life, follicle fate is balanced between growth and apoptosis. These opposing forces are controlled by numerous endocrine, paracrine, and autocrine factors, including the ligands represented by the transforming growth factor beta (TGFbeta) superfamily. TGFbeta, activin, inhibin, bone morphometric protein (BMP), and growth differentiation factor 9 (GDF-9) are present in the ovary of many animals; however, no comprehensive analysis of the localization of each ligand or its receptors and intracellular signaling molecules during folliculogenesis has been done. The domestic cat is an ideal model for studying ovarian follicle dynamics due to an abundance of all follicle populations, including primordial stage, and the amount of readily available tissue following routine animal spaying. Additionally, knowledge of the factors involved in feline follicular development could make an important impact on in vitro maturation/in vitro fertilization (IVM/IVF) success for endangered feline species. Thus, the presence and position of TGFbeta superfamily members within the feline ovary have been evaluated in all stages of follicular development by immunolocalization. The cat inhibin alpha subunit protein is present in all follicle stages but increases in intensity within the mural granulosa cells in large antral follicles. The inhibin betaA and betaB subunit proteins, in addition to the activin type I (ActRIB) and activin type II receptor (ActRIIB), are produced in primordial and primary follicle granulosa cells. Additionally, inhibin betaA subunit is detected in the theca cells from secondary through large antral follicle size classes. GDF-9 is restricted to the oocyte of preantral and antral follicles, whereas the type II BMP receptor (BMP-RII) protein is predominantly localized to primordial- and primary-stage follicles. TGFbeta1, 2

  15. Boolean gates on actin filaments

    NASA Astrophysics Data System (ADS)

    Siccardi, Stefano; Tuszynski, Jack A.; Adamatzky, Andrew

    2016-01-01

    Actin is a globular protein which forms long polar filaments in the eukaryotic cytoskeleton. Actin networks play a key role in cell mechanics and cell motility. They have also been implicated in information transmission and processing, memory and learning in neuronal cells. The actin filaments have been shown to support propagation of voltage pulses. Here we apply a coupled nonlinear transmission line model of actin filaments to study interactions between voltage pulses. To represent digital information we assign a logical TRUTH value to the presence of a voltage pulse in a given location of the actin filament, and FALSE to the pulse's absence, so that information flows along the filament with pulse transmission. When two pulses, representing Boolean values of input variables, interact, then they can facilitate or inhibit further propagation of each other. We explore this phenomenon to construct Boolean logical gates and a one-bit half-adder with interacting voltage pulses. We discuss implications of these findings on cellular process and technological applications.

  16. Technical advance: identification of plant actin-binding proteins by F-actin affinity chromatography

    NASA Technical Reports Server (NTRS)

    Hu, S.; Brady, S. R.; Kovar, D. R.; Staiger, C. J.; Clark, G. B.; Roux, S. J.; Muday, G. K.

    2000-01-01

    Proteins that interact with the actin cytoskeleton often modulate the dynamics or organization of the cytoskeleton or use the cytoskeleton to control their localization. In plants, very few actin-binding proteins have been identified and most are thought to modulate cytoskeleton function. To identify actin-binding proteins that are unique to plants, the development of new biochemical procedures will be critical. Affinity columns using actin monomers (globular actin, G-actin) or actin filaments (filamentous actin, F-actin) have been used to identify actin-binding proteins from a wide variety of organisms. Monomeric actin from zucchini (Cucurbita pepo L.) hypocotyl tissue was purified to electrophoretic homogeneity and shown to be native and competent for polymerization to actin filaments. G-actin, F-actin and bovine serum albumin affinity columns were prepared and used to separate samples enriched in either soluble or membrane-associated actin-binding proteins. Extracts of soluble actin-binding proteins yield distinct patterns when eluted from the G-actin and F-actin columns, respectively, leading to the identification of a putative F-actin-binding protein of approximately 40 kDa. When plasma membrane-associated proteins were applied to these columns, two abundant polypeptides eluted selectively from the F-actin column and cross-reacted with antiserum against pea annexins. Additionally, a protein that binds auxin transport inhibitors, the naphthylphthalamic acid binding protein, which has been previously suggested to associate with the actin cytoskeleton, was eluted in a single peak from the F-actin column. These experiments provide a new approach that may help to identify novel actin-binding proteins from plants.

  17. How cofilin severs an actin filament.

    PubMed

    De La Cruz, Enrique M

    2009-05-15

    The actin regulatory protein, cofilin, promotes actin assembly dynamics by severing filaments and increasing the number of ends from which subunits add and dissociate. Recent studies provide biophysical descriptions of cooperative filament interactions in energetic, mechanical and structural terms. A one-dimensional Ising model with nearest-neighbor interactions permits thermodynamic analysis of cooperative binding and indicates that one or a few cofilin molecules can sever a filament. Binding and cooperative interactions are entropically driven. A significant fraction of the binding free energy results from the linked dissociation of filament-associated ions (polyelectrolyte effect), which modulate filament structure, stability and mechanics. The remaining binding free energy and essentially all of the cooperative free energy arise from the enhanced conformational dynamics of the cofilactin complex. Filament mechanics are modulated by cofilin such that cofilin-saturated filaments are approximately 10- to 20-fold more compliant in bending and twisting than bare filaments. Cofilin activity is well described by models in which discontinuities in topology, mechanics and conformational dynamics generate stress concentration and promote fracture at junctions of bare and decorated segments, analogous to the grain boundary fracture of crystalline materials and the thermally driven formation of shear transformation zones in colloidal glass. PMID:20700473

  18. Actin binding proteins, spermatid transport and spermiation*

    PubMed Central

    Qian, Xiaojing; Mruk, Dolores D.; Cheng, Yan-Ho; Tang, Elizabeth I.; Han, Daishu; Lee, Will M.; Wong, Elissa W. P.; Cheng, C. Yan

    2014-01-01

    The transport of germ cells across the seminiferous epithelium is composed of a series of cellular events during the epithelial cycle essential to the completion of spermatogenesis. Without the timely transport of spermatids during spermiogenesis, spermatozoa that are transformed from step 19 spermatids in the rat testis fail to reach the luminal edge of the apical compartment and enter the tubule lumen at spermiation, thereby entering the epididymis for further maturation. Step 19 spermatids and/or sperms that remain in the epithelium will be removed by the Sertoli cell via phagocytosis to form phagosomes and be degraded by lysosomes, leading to subfertility and/or infertility. However, the biology of spermatid transport, in particular the final events that lead to spermiation remain elusive. Based on recent data in the field, we critically evaluate the biology of spermiation herein by focusing on the actin binding proteins (ABPs) that regulate the organization of actin microfilaments at the Sertoli-spermatid interface, which is crucial for spermatid transport during this event. The hypothesis we put forth herein also highlights some specific areas of research that can be pursued by investigators in the years to come. PMID:24735648

  19. Bacterial Actins? An Evolutionary Perspective

    NASA Technical Reports Server (NTRS)

    Doolittle, Russell F.; York, Amanda L.

    2003-01-01

    According to the conventional wisdom, the existence of a cytoskeleton in eukaryotes and its absence in prokaryotes constitute a fundamental divide between the two domains of life. An integral part of the dogma is that a cytoskeleton enabled an early eukaryote to feed upon prokaryotes, a consequence of which was the occasional endosymbiosis and the eventual evolution of organelles. Two recent papers present compelling evidence that actin, one of the principal components of a cytoskeleton, has a homolog in Bacteria that behaves in many ways like eukaryotic actin. Sequence comparisons reveml that eukaryotic actin and the bacterial homolog (mreB protein), unlike many other proteins common to eukaryotes and Bacteria, have very different and more highly extended evolutionary histories.

  20. Actin engine in immunological synapse.

    PubMed

    Piragyte, Indre; Jun, Chang-Duk

    2012-06-01

    T cell activation and function require physical contact with antigen presenting cells at a specialized junctional structure known as the immunological synapse. Once formed, the immunological synapse leads to sustained T cell receptor-mediated signalling and stabilized adhesion. High resolution microscopy indeed had a great impact in understanding the function and dynamic structure of immunological synapse. Trends of recent research are now moving towards understanding the mechanical part of immune system, expanding our knowledge in mechanosensitivity, force generation, and biophysics of cell-cell interaction. Actin cytoskeleton plays inevitable role in adaptive immune system, allowing it to bear dynamic and precise characteristics at the same time. The regulation of mechanical engine seems very complicated and overlapping, but it enables cells to be very sensitive to external signals such as surface rigidity. In this review, we focus on actin regulators and how immune cells regulate dynamic actin rearrangement process to drive the formation of immunological synapse. PMID:22916042

  1. Synthetic polyamines: new compounds specific to actin dynamics for mammalian cell and fission yeast

    PubMed Central

    Riveline, Daniel; Thiagarajan, Raghavan; Lehn, Jean-Marie; Carlier, Marie-France

    2014-01-01

    Actin is a major actor in the determination of cell shape. On the one hand, site-directed assembly/disassembly cycles of actin filaments drive protrusive force leading to lamellipodia and filopodia dynamics. Force produced by actin similarly contributes in membrane scission in endocytosis or Golgi remodeling. On the other hand, cellular processes like adhesion, immune synapse, cortex dynamics or cytokinesis are achieved by combining acto-myosin contractility and actin assembly in a complex and not fully understood manner. New chemical compounds are therefore needed to disentangle acto-myosin and actin dynamics. We have found that synthetic, cell permeant, short polyamines are promising new actin regulators in this context. They generate growth and stabilization of lamellipodia within minutes by slowing down the actin assembly/disassembly cycle and facilitating nucleation. We now report that these polyamines also slow down cytokinetic ring closure in fission yeast. This shows that these synthetic compounds are active also in yeasts, and these experiments specifically highlight that actin depolymerization is involved in the ring closure. Thus, synthetic polyamines appear to be potentially powerful agents in a quantitative approach to the role of actin in complex processes in cell biology, developmental biology and potentially cancer research. PMID:25664996

  2. F-actin distribution and function during sexual development in Eimeria maxima.

    PubMed

    Frölich, Sonja; Wallach, Michael

    2015-06-01

    To determine the involvement of the actin cytoskeleton in macrogametocyte growth and oocyst wall formation, freshly purified macrogametocytes and oocysts were stained with Oregon Green 514 conjugated phalloidin to visualize F-actin microfilaments, while Evans blue staining was used to detect type 1 wall forming bodies (WFB1s) and the outer oocyst wall. The double-labelled parasites were then analysed at various stages of sexual development using three-dimensional confocal microscopy. The results showed F-actin filaments were distributed throughout the entire cytoplasm of mature Eimeria maxima macrogametocytes forming a web-like meshwork of actin filaments linking the type 1 WFBs together into structures resembling 'beads on a string'. At the early stages of oocyst wall formation, F-actin localization changed in alignment with the egg-shaped morphology of the forming oocysts with F-actin microfilaments making direct contact with the WFB1s. In tissue oocysts, the labelled actin cytoskeleton was situated underneath the forming outer layer of the oocyst wall. Treatment of macrogametocytes in vitro with the actin depolymerizing agents, Cytochalasin D and Latrunculin, led to a reduction in the numbers of mature WFB1s in the cytoplasm of the developing macrogametocytes, indicating that the actin plays an important role in WFB1 transport and oocyst wall formation in E. maxima. PMID:25800683

  3. Synthetic polyamines: new compounds specific to actin dynamics for mammalian cell and fission yeast.

    PubMed

    Riveline, Daniel; Thiagarajan, Raghavan; Lehn, Jean-Marie; Carlier, Marie-France

    2014-01-01

    Actin is a major actor in the determination of cell shape. On the one hand, site-directed assembly/disassembly cycles of actin filaments drive protrusive force leading to lamellipodia and filopodia dynamics. Force produced by actin similarly contributes in membrane scission in endocytosis or Golgi remodeling. On the other hand, cellular processes like adhesion, immune synapse, cortex dynamics or cytokinesis are achieved by combining acto-myosin contractility and actin assembly in a complex and not fully understood manner. New chemical compounds are therefore needed to disentangle acto-myosin and actin dynamics. We have found that synthetic, cell permeant, short polyamines are promising new actin regulators in this context. They generate growth and stabilization of lamellipodia within minutes by slowing down the actin assembly/disassembly cycle and facilitating nucleation. We now report that these polyamines also slow down cytokinetic ring closure in fission yeast. This shows that these synthetic compounds are active also in yeasts, and these experiments specifically highlight that actin depolymerization is involved in the ring closure. Thus, synthetic polyamines appear to be potentially powerful agents in a quantitative approach to the role of actin in complex processes in cell biology, developmental biology and potentially cancer research. PMID:25664996

  4. TWISTED DWARF1 Mediates the Action of Auxin Transport Inhibitors on Actin Cytoskeleton Dynamics.

    PubMed

    Zhu, Jinsheng; Bailly, Aurelien; Zwiewka, Marta; Sovero, Valpuri; Di Donato, Martin; Ge, Pei; Oehri, Jacqueline; Aryal, Bibek; Hao, Pengchao; Linnert, Miriam; Burgardt, Noelia Inés; Lücke, Christian; Weiwad, Matthias; Michel, Max; Weiergräber, Oliver H; Pollmann, Stephan; Azzarello, Elisa; Mancuso, Stefano; Ferro, Noel; Fukao, Yoichiro; Hoffmann, Céline; Wedlich-Söldner, Roland; Friml, Jiří; Thomas, Clément; Geisler, Markus

    2016-04-01

    Plant growth and architecture is regulated by the polar distribution of the hormone auxin. Polarity and flexibility of this process is provided by constant cycling of auxin transporter vesicles along actin filaments, coordinated by a positive auxin-actin feedback loop. Both polar auxin transport and vesicle cycling are inhibited by synthetic auxin transport inhibitors, such as 1-N-naphthylphthalamic acid (NPA), counteracting the effect of auxin; however, underlying targets and mechanisms are unclear. Using NMR, we map the NPA binding surface on the Arabidopsis thaliana ABCB chaperone TWISTED DWARF1 (TWD1). We identify ACTIN7 as a relevant, although likely indirect, TWD1 interactor, and show TWD1-dependent regulation of actin filament organization and dynamics and that TWD1 is required for NPA-mediated actin cytoskeleton remodeling. The TWD1-ACTIN7 axis controls plasma membrane presence of efflux transporters, and as a consequence act7 and twd1 share developmental and physiological phenotypes indicative of defects in auxin transport. These can be phenocopied by NPA treatment or by chemical actin (de)stabilization. We provide evidence that TWD1 determines downstream locations of auxin efflux transporters by adjusting actin filament debundling and dynamizing processes and mediating NPA action on the latter. This function appears to be evolutionary conserved since TWD1 expression in budding yeast alters actin polarization and cell polarity and provides NPA sensitivity. PMID:27053424

  5. Profilin-Dependent Nucleation and Assembly of Actin Filaments Controls Cell Elongation in Arabidopsis1[OPEN

    PubMed Central

    Cao, Lingyan; Blanchoin, Laurent; Staiger, Christopher J.

    2016-01-01

    Actin filaments in plant cells are incredibly dynamic; they undergo incessant remodeling and assembly or disassembly within seconds. These dynamic events are choreographed by a plethora of actin-binding proteins, but the exact mechanisms are poorly understood. Here, we dissect the contribution of Arabidopsis (Arabidopsis thaliana) PROFILIN1 (PRF1), a conserved actin monomer-binding protein, to actin organization and single filament dynamics during axial cell expansion of living epidermal cells. We found that reduced PRF1 levels enhanced cell and organ growth. Surprisingly, we observed that the overall frequency of nucleation events in prf1 mutants was dramatically decreased and that a subpopulation of actin filaments that assemble at high rates was reduced. To test whether profilin cooperates with plant formin proteins to execute actin nucleation and rapid filament elongation in cells, we used a pharmacological approach. Here, we used Small Molecule Inhibitor of Formin FH2 (SMIFH2), after validating its mode of action on a plant formin in vitro, and observed a reduced nucleation frequency of actin filaments in live cells. Treatment of wild-type epidermal cells with SMIFH2 mimicked the phenotype of prf1 mutants, and the nucleation frequency in prf1-2 mutant was completely insensitive to these treatments. Our data provide compelling evidence that PRF1 coordinates the stochastic dynamic properties of actin filaments by modulating formin-mediated actin nucleation and assembly during plant cell expansion. PMID:26574597

  6. Actin polymerization is stimulated by actin cross-linking protein palladin.

    PubMed

    Gurung, Ritu; Yadav, Rahul; Brungardt, Joseph G; Orlova, Albina; Egelman, Edward H; Beck, Moriah R

    2016-02-15

    The actin scaffold protein palladin regulates both normal cell migration and invasive cell motility, processes that require the co-ordinated regulation of actin dynamics. However, the potential effect of palladin on actin dynamics has remained elusive. In the present study, we show that the actin-binding immunoglobulin-like domain of palladin, which is directly responsible for both actin binding and bundling, also stimulates actin polymerization in vitro. Palladin eliminated the lag phase that is characteristic of the slow nucleation step of actin polymerization. Furthermore, palladin dramatically reduced depolymerization, slightly enhanced the elongation rate, and did not alter the critical concentration. Microscopy and in vitro cross-linking assays reveal differences in actin bundle architecture when palladin is incubated with actin before or after polymerization. These results suggest a model whereby palladin stimulates a polymerization-competent form of globular or monomeric actin (G-actin), akin to metal ions, either through charge neutralization or through conformational changes. PMID:26607837

  7. Actin polymerization or myosin contraction: two ways to build up cortical tension for symmetry breaking.

    PubMed

    Carvalho, Kevin; Lemière, Joël; Faqir, Fahima; Manzi, John; Blanchoin, Laurent; Plastino, Julie; Betz, Timo; Sykes, Cécile

    2013-01-01

    Cells use complex biochemical pathways to drive shape changes for polarization and movement. One of these pathways is the self-assembly of actin filaments and myosin motors that together produce the forces and tensions that drive cell shape changes. Whereas the role of actin and myosin motors in cell polarization is clear, the exact mechanism of how the cortex, a thin shell of actin that is underneath the plasma membrane, can drive cell shape changes is still an open question. Here, we address this issue using biomimetic systems: the actin cortex is reconstituted on liposome membranes, in an 'outside geometry'. The actin shell is either grown from an activator of actin polymerization immobilized at the membrane by a biotin-streptavidin link, or built by simple adsorption of biotinylated actin filaments to the membrane, in the presence or absence of myosin motors. We show that tension in the actin network can be induced either by active actin polymerization on the membrane via the Arp2/3 complex or by myosin II filament pulling activity. Symmetry breaking and spontaneous polarization occur above a critical tension that opens up a crack in the actin shell. We show that this critical tension is reached by growing branched networks, nucleated by the Arp2/3 complex, in a concentration window of capping protein that limits actin filament growth and by a sufficient number of motors that pull on actin filaments. Our study provides the groundwork to understanding the physical mechanisms at work during polarization prior to cell shape modifications. PMID:24062578

  8. Genetic transformation of Gentiana macrophylla with Agrobacterium rhizogenes: growth and production of secoiridoid glucoside gentiopicroside in transformed hairy root cultures.

    PubMed

    Tiwari, Rajesh Kumar; Trivedi, Mala; Guang, Zhang Chun; Guo, Guang-Qin; Zheng, Guo-Chang

    2007-02-01

    Hairy root cultures of Gentiana macrophylla were established by infecting the different explants four Agrobacterium rhizogenes strains namely A(4)GUS, R1000, LBA 9402 and ATCC11325, and hairy root lines were established with A. rhizogenes strain R1000 in 1/2 MS + B(5) medium. Initially, 42 independent hairy root clones were maintained and seven clones belongs to different category were evaluated for growth, morphology, integration and expression of Ri T-DNA genes, and alkaloid contents in dry root samples. On the basis of total root elongation, lateral root density and biomass accumulation on solid media, hairy root clones were separated into three categories. PCR and Southern hybridization analysis revealed both left and right T-DNA integration in the root clones and RT-PCR analysis confirmed the expression of hairy root inducible gene. GUS assay was also performed to confirm the integration of left T-DNA. The accumulation of considerable amounts of the root-specific secoiridoid glucosides gentiopicroside was observed in GM1 (T +/L and T +/R) and the GM2 (T +/L and T -/R DNA) type clones in considerably higher amount whether as two T -/L but T +/R callus-type clones (GM3) accumulated much less or only very negligible amounts of gentiopicroside. Out of four media composition the 1/2 MS + B(5) vitamin media was found most suitable. We found that initial establishment of root cultures largely depends on root:media ratio. Maximum growth rate was recorded in 1:50 root:media ratio. The maximum biomass in terms of fresh weight (33-fold) was achieved in 1/2 MS + B(5) media composition after 35 days in comparison to sixfold increase in control. The biomass increase was most abundant maximum from 15 to 30 days. Influence of A. rhizogenes strains and Ri plasmid of hairy root induction, the possible role of the T(L)-DNA and T(R)-DNA genes on growth pattern of hairy root, initial root inoculum:media ratio and effect of media composition is discussed. PMID:16972092

  9. Interactions between the yeast SM22 homologue Scp1 and actin demonstrate the importance of actin bundling in endocytosis.

    PubMed

    Gheorghe, Dana M; Aghamohammadzadeh, Soheil; Smaczynska-de Rooij, Iwona I; Allwood, Ellen G; Winder, Steve J; Ayscough, Kathryn R

    2008-05-30

    The yeast SM22 homologue Scp1 has previously been shown to act as an actin-bundling protein in vitro. In cells, Scp1 localizes to the cortical actin patches that form as part of the invagination process during endocytosis, and its function overlaps with that of the well characterized yeast fimbrin homologue Sac6p. In this work we have used live cell imaging to demonstrate the importance of key residues in the Scp1 actin interface. We have defined two actin binding domains within Scp1 that allow the protein to both bind and bundle actin without the need for dimerization. Green fluorescent protein-tagged mutants of Scp1 also indicate that actin localization does not require the putative phosphorylation site Ser-185 to be functional. Deletion of SCP1 has few discernable effects on cell growth and morphology. However, we reveal that scp1 deletion is compensated for by up-regulation of Sac6. Furthermore, Scp1 levels are increased in the absence of sac6. The presence of compensatory pathways to up-regulate Sac6 or Scp1 levels in the absence of the other suggest that maintenance of sufficient bundling activity is critical within the cell. Analysis of cortical patch assembly and movement during endocytosis reveals a previously undetected role for Scp1 in movement of patches away from the plasma membrane. Additionally, we observe a dramatic increase in patch lifetime in a strain lacking both sac6 and scp1, demonstrating the central role played by actin-bundling proteins in the endocytic process. PMID:18400761

  10. Differential regulation of actin microfilaments by human MICAL proteins

    PubMed Central

    Giridharan, Sai Srinivas Panapakkam; Rohn, Jennifer L.; Naslavsky, Naava; Caplan, Steve

    2012-01-01

    The Drosophila melanogaster MICAL protein is essential for the neuronal growth cone machinery that functions through plexin- and semaphorin-mediated axonal signaling. Drosophila MICAL is also involved in regulating myofilament organization and synaptic structures, and serves as an actin disassembly factor downstream of plexin-mediated axonal repulsion. In mammalian cells there are three known isoforms, MICAL1, MICAL2 and MICAL3, as well as the MICAL-like proteins MICAL-L1 and MICAL-L2, but little is known of their function, and information comes almost exclusively from neural cells. In this study we show that in non-neural cells human MICALs are required for normal actin organization, and all three MICALs regulate actin stress fibers. Moreover, we provide evidence that the generation of reactive oxygen species by MICAL proteins is crucial for their actin-regulatory function. However, although MICAL1 is auto-inhibited by its C-terminal coiled-coil region, MICAL2 remains constitutively active and affects stress fibers. These data suggest differential but complementary roles for MICAL1 and MICAL2 in actin microfilament regulation. PMID:22331357

  11. Cortactin promotes exosome secretion by controlling branched actin dynamics.

    PubMed

    Sinha, Seema; Hoshino, Daisuke; Hong, Nan Hyung; Kirkbride, Kellye C; Grega-Larson, Nathan E; Seiki, Motoharu; Tyska, Matthew J; Weaver, Alissa M

    2016-07-18

    Exosomes are extracellular vesicles that influence cellular behavior and enhance cancer aggressiveness by carrying bioactive molecules. The mechanisms that regulate exosome secretion are poorly understood. Here, we show that the actin cytoskeletal regulatory protein cortactin promotes exosome secretion. Knockdown or overexpression of cortactin in cancer cells leads to a respective decrease or increase in exosome secretion, without altering exosome cargo content. Live-cell imaging revealed that cortactin controls both trafficking and plasma membrane docking of multivesicular late endosomes (MVEs). Regulation of exosome secretion by cortactin requires binding to the branched actin nucleating Arp2/3 complex and to actin filaments. Furthermore, cortactin, Rab27a, and coronin 1b coordinately control stability of cortical actin MVE docking sites and exosome secretion. Functionally, the addition of purified exosomes to cortactin-knockdown cells rescued defects of those cells in serum-independent growth and invasion. These data suggest a model in which cortactin promotes exosome secretion by stabilizing cortical actin-rich MVE docking sites. PMID:27402952

  12. MICAL-Family Proteins: Complex Regulators of the Actin Cytoskeleton

    PubMed Central

    Giridharan, Sai Srinivas Panapakkam

    2014-01-01

    Abstract Significance: The molecules interacting with CasL (MICAL) family members participate in a multitude of activities, including axonal growth cone repulsion, membrane trafficking, apoptosis, and bristle development in flies. An interesting feature of MICAL proteins is the presence of an N-terminal flavo-mono-oxygenase domain. This mono-oxygenase domain generates redox potential with which MICALs can either oxidize proteins or produce reactive oxygen species (ROS). Actin is one such protein that is affected by MICAL function, leading to dramatic cytoskeletal rearrangements. This review describes the MICAL-family members, and discusses their mechanisms of actin-binding and regulation of actin cytoskeleton organization. Recent Advances: Recent studies show that MICALs directly induce oxidation of actin molecules, leading to actin depolymerization. ROS production by MICALs also causes oxidation of collapsin response mediator protein-2, a microtubule assembly promoter, which subsequently undergoes phosphorylation. Critical Issues: MICAL proteins oxidize proteins through two mechanisms: either directly by oxidizing methionine residues or indirectly via the production of ROS. It remains unclear whether MICAL proteins employ both mechanisms or whether the activity of MICAL-family proteins might vary with different substrates. Future Directions: The identification of additional substrates oxidized by MICAL will shed new light on MICAL protein function. Additional directions include expanding studies toward the MICAL-like homologs that lack flavin adenine dinucleotide domains and oxidation activity. Antioxid. Redox Signal. 20, 2059–2073. PMID:23834433

  13. Effects of transforming growth factor-beta on long-term human cord blood monocyte cultures

    SciTech Connect

    Orcel, P.; Bielakoff, J.; De Vernejoul, M.C. )

    1990-02-01

    Transforming growth factor-beta (TGF-beta) modulates growth and differentiation in many cell types and is abundant in bone matrix. We recently showed that human cord blood monocytes cultured in the presence of 1,25(OH)2D3 acquire some features of osteoclast precursors. Since TGF-beta has been shown to influence bone resorption in organ culture, we have studied the effect of TGF-beta (1-1,000 pg/ml) on cord blood monocyte cultures. These cells were cultured on plastic substrate during 3 weeks in the presence of 20% horse serum and 10(-9) M 1,25(OH)2D3. TGF-beta, from a concentration of 10 pg/ml in the culture medium, decreased in a dose dependent manner the formation of multinucleated cells. At a concentration of TGF-beta of 1 ng/ml, the multinucleated cells were reduced to 2.1% +/- 0.3%, compared to 19.3% +/- 1.5% in control cultures. TGF-beta inhibited in a dose-dependent manner the proliferation of cord blood monocytes as assessed by 3H-thymidine incorporation at 7 and 14 days of culture. The fusion index was also decreased by 3 weeks of treatment with TGF-beta. Indomethacin did not reverse the inhibitory effects of TGF-beta. The expression of the osteoclastic phenotype was assessed using two different antibodies: 23C6, a monoclonal antibody directed against the vitronectin receptor, which is highly expressed by osteoclasts but not by adult monocytes, and an antibody to HLA-DR, which is not present on osteoclast. TGF-beta decreased the expression of HLA-DR and increased in a dose-dependent manner the proportion of 23C6-labeled cells; these results suggest that TGF-beta could modulate a differentiation effect to the osteoclastic phenotype. However, when cord blood monocytes were cultured on devitalized rat calvariae prelabeled with 45Ca, TGF-beta did not induce any 45Ca release from bone cultured with monocytes.

  14. Transforming growth factor-beta1 upregulates myostatin expression in mouse C2C12 myoblasts.

    PubMed

    Budasz-Rwiderska, M; Jank, M; Motyl, T

    2005-06-01

    Myostatin (MSTN) and transforming growth factor-beta1 (TGF-beta1) belong to the same TGF-beta superfamily of proteins. They are involved in regulation of skeletal muscle growth and development as well as muscle catabolism. The aim of the present study was to investigate the relationship between MSTN and TGF-beta1 expression in proliferating and differentiating mouse C2C12 myoblasts cultured in normal and catabolic conditions and to evaluate the effect of exogenous TGF-beta1 as well as "knock down" of TGF-beta1 receptor type II on MSTN expression in proliferating and differentiating myogenic cells. The direct effect of TGF-beta1 on myostatin was also examined. Myostatin expression increased gradually with cell confluency in proliferating cultures, while the level of TGF-beta1, detected in the form of a 100 kDa small latent complex diminished. Myostatin expression was accompanied by a partial cell cycle arrest. Three forms of myostatin were found: a 52 kDa precursor, a 40 kDa latency associated propeptide, and a 26 kDa active peptide. A decrease in myostatin and TGF-beta1 levels was observed during the first three days of differentiation, which was subsequently followed by significant increase of their expression during next three to four days of differentiation. Catabolic state induced by dexamethasone significantly increased the level of all forms of myostatin as well as latent (100 kDa) and active (25 kDa) forms of TGF-beta1 in differentiating myoblasts in a dose dependent manner. Exogenous TGF-beta1 (2 ng/ml) significantly increased myostatin levels both in proliferating and differentiating C2C12 myoblasts, whereas silencing of the TGF-beta1 receptor II gene significantly lowered myostatin level in examined cells. The presented results indicate that TGF-beta1 may control myostatin-related regulation of myogenesis through up-regulation of myostatin, predominantly in the course of terminal differentiation and glucocorticoid-dependent catabolic stimulation. PMID

  15. Cellular localization of transforming growth factor-beta expression in bleomycin-induced pulmonary fibrosis.

    PubMed Central

    Zhang, K.; Flanders, K. C.; Phan, S. H.

    1995-01-01

    Bleomycin-induced pulmonary fibrosis is associated with increased lung transforming growth factor-beta (TGF-beta) gene expression, but cellular localization of the source of this expression has not been unequivocally established. In this study, lung fibrosis was induced in rats by endotracheal bleomycin injection on day 0 and, on selected days afterwards, lungs were harvested for in situ hybridization, immunohistochemical and histochemical analyses for TGF-beta 1 mRNA and protein expression, and cell identification. The results show that control lungs express essentially no detectable TGF-beta 1 mRNA or protein in the parenchyma. Before day 3 after bleomycin treatment, scattered bronchiolar epithelial cells, mononuclear cells, and eosinophils expressed elevated levels of TGF-beta 1. Between days 3 and 14, there was a major increase in the number of eosinophils, myofibroblasts, and fibroblasts strongly expressing TGF-beta 1 mRNA and protein. TGF-beta 1-producing cells were predominantly localized within areas of injury and active fibrosis. After day 14, the intensity and number of TGF-beta 1-expressing cells significantly declined and were predominantly found in fibroblasts in fibrotic areas. The expression of TGF-beta 1 protein was generally coincident with that for mRNA with the exception of bronchiolar epithelial cells in which strong protein expression was unaccompanied by a commensurate increase in mRNA. The study demonstrates that myofibroblasts, fibroblasts, and eosinophils represent the major sources of increased lung TGF-beta 1 expression in this model of pulmonary fibrosis. Images Figure 2 Figure 3 Figure 4 PMID:7543734

  16. Protection of transforming growth factor-beta 1 activity by heparin and fucoidan.

    PubMed

    McCaffrey, T A; Falcone, D J; Vicente, D; Du, B; Consigli, S; Borth, W

    1994-04-01

    The transforming growth factor-beta (TGF-beta) family of proteins exert diverse and potent effects on proliferation, differentiation, and extracellular matrix synthesis. However, relatively little is known about the stability or processing of endogenous TGF-beta activity in vitro or in vivo. Our previous work indicated that 1) TGF-beta 1 has strong heparin-binding properties that were not previously recognized because of neutralization by iodination, and 2) heparin, and certain other polyanions, could block the binding of TGF-beta 1 to alpha 2-macroglobulin (alpha 2-M). The present studies investigated the influence of heparin-like molecules on the stability of the TGF-beta 1 signal in the pericellular environment. The results indicate that heparin and fucoidan, a naturally occurring sulfated L-fucose polymer, suppress the formation of an initial non-covalent interaction between 125I-TGF-beta 1 and activated alpha 2-M. Electrophoresis of 125I-TGF-beta 1 showed that fucoidan protects TGF-beta 1 from proteolytic degradation by plasmin and trypsin. While plasmin caused little, if any, activation of latent TGF-beta derived from vascular smooth muscle cells (SMC), plasmin degraded acid-activated TGF-beta, and purified TGF-beta 1, and this degradation was inhibited by fucoidan. In vitro, heparin and fucoidan tripled the half-life of 125I-TGF-beta 1 and doubled the amount of cell-associated 125I-TGF-beta 1. Consistent with this protective effect, heparin- and fucoidan-treated SMC demonstrated elevated levels of active, but not latent, TGF-beta activity. PMID:7511146

  17. 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

  18. Early defect of transforming growth factor β1 formation in Huntington’s disease

    PubMed Central

    Battaglia, Giuseppe; Cannella, Milena; Riozzi, Barbara; Orobello, Sara; Maat-Schieman, Marion L; Aronica, Eleonora; Busceti, Carla Letizia; Ciarmiello, Andrea; Alberti, Silvia; Amico, Enrico; Sassone, Jenny; Sipione, Simonetta; Bruno, Valeria; Frati, Luigi; Nicoletti, Ferdinando; Squitieri, Ferdinando

    2011-01-01

    Abstract A defective expression or activity of neurotrophic factors, such as brain- and glial-derived neurotrophic factors, contributes to neuronal damage in Huntington’s disease (HD). Here, we focused on transforming growth factor-β (TGF-β1), a pleiotropic cytokine with an established role in mechanisms of neuroprotection. Asymptomatic HD patients showed a reduction in TGF-β1 levels in the peripheral blood, which was related to trinucleotide mutation length and glucose hypometabolism in the caudate nucleus. Immunohistochemical analysis in post-mortem brain tissues showed that TGF-β1 was reduced in cortical neurons of asymptomatic and symptomatic HD patients. Both YAC128 and R6/2 HD mutant mice showed a reduced expression of TGF-β1 in the cerebral cortex, localized in neurons, but not in astrocytes. We examined the pharmacological regulation of TGF-β1 formation in asymptomatic R6/2 mice, where blood TGF-β1 levels were also reduced. In these R6/2 mice, both the mGlu2/3 metabotropic glutamate receptor agonist, LY379268, and riluzole failed to increase TGF-β1 formation in the cerebral cortex and corpus striatum, suggesting that a defect in the regulation of TGF-β1 production is associated with HD. Accordingly, reduced TGF-β1 mRNA and protein levels were found in cultured astrocytes transfected with mutated exon 1 of the human huntingtin gene, and in striatal knock-in cell lines expressing full-length huntingtin with an expanded glutamine repeat. Taken together, our data suggest that serum TGF-β1 levels are potential biomarkers of HD development during the asymptomatic phase of the disease, and raise the possibility that strategies aimed at rescuing TGF-β1 levels in the brain may influence the progression of HD. PMID:20082658

  19. Immunohistochemical detection of active transforming growth factor-beta in situ using engineered tissue

    NASA Technical Reports Server (NTRS)

    Barcellos-Hoff, M. H.; Ehrhart, E. J.; Kalia, M.; Jirtle, R.; Flanders, K.; Tsang, M. L.; Chatterjee, A. (Principal Investigator)

    1995-01-01

    The biological activity of transforming growth factor-beta 1 (TGF-beta) is governed by dissociation from its latent complex. Immunohistochemical discrimination of active and latent TGF-beta could provide insight into TGF-beta activation in physiological and pathological processes. However, evaluation of immunoreactivity specificity in situ has been hindered by the lack of tissue in which TGF-beta status is known. To provide in situ analysis of antibodies to differentiate between these functional forms, we used xenografts of human tumor cells modified by transfection to overexpress latent TGF-beta or constitutively active TGF-beta. This comparison revealed that, whereas most antibodies did not differentiate between TGF-beta activation status, the immunoreactivity of some antibodies was activation dependent. Two widely used peptide antibodies to the amino-terminus of TGF-beta, LC(1-30) and CC(1-30) showed marked preferential immunoreactivity with active TGF-beta versus latent TGF-beta in cryosections. However, in formalin-fixed, paraffin-embedded tissue, discrimination of active TGF-beta by CC(1-30) was lost and immunoreactivity was distinctly extracellular, as previously reported for this antibody. Similar processing-dependent extracellular localization was found with a neutralizing antibody raised to recombinant TGF-beta. Antigen retrieval recovered cell-associated immunoreactivity of both antibodies. Two antibodies to peptides 78-109 showed mild to moderate preferential immunoreactivity with active TGF-beta only in paraffin sections. LC(1-30) was the only antibody tested that discriminated active from latent TGF-beta in both frozen and paraffin-embedded tissue. Thus, in situ discrimination of active versus latent TGF-beta depends on both the antibody and tissue preparation. We propose that tissues engineered to express a specific form of a given protein provide a physiological setting in which to evaluate antibody reactivity with specific functional forms of a

  20. Hydrogen Sulfide Inhibits Transforming Growth Factor-β1-Induced EMT via Wnt/Catenin Pathway

    PubMed Central

    Tao, Jie; Lan, Zhen; Hei, Hongya; Tian, Lulu; Pan, Wanma; Wang, Li; Zhang, Xuemei

    2016-01-01

    Hydrogen sulfide (H2S) has anti-fibrotic potential in lung, kidney and other organs. The exogenous H2S is released from sodium hydrosulfide (NaHS) and can influence the renal fibrosis by blocking the differentiation of quiescent renal fibroblasts to myofibroblasts. But whether H2S affects renal epithelial-to-mesenchymal transition (EMT) and the underlying mechanisms remain unknown. Our study is aimed at investigating the in vitro effects of H2S on transforming growth factor-β1 (TGF-β1)-induced EMT in renal tubular epithelial cells (HK-2 cells) and the associated mechanisms. The induced EMT is assessed by Western blotting analysis on the expressions of α-SMA, E-cadherin and fibronectin. HK-2 cells were treated with NaHS before incubating with TGF-β1 to investigate its effect on EMT and the related molecular mechanism. Results demonstrated that NaHS decreased the expression of α-SMA and fibronectin, and increased the expression of E-cadherin. NaHS reduced the expression of TGF-β receptor type I (TβR I) and TGF-β receptor type II (TβR II). In addition, NaHS attenuated TGF-β1-induced increase of β-catenin expression and ERK phosphorylation. Moreover, it inhibited the TGF-β1-induced nuclear translocation of ββ-catenin. These effects of NaHS on fibronectin, E-cadherin and TβR I were abolished by the ERK inhibitor U0126 or β-catenin inhibitor XAV939, or β-catenin siRNA interference. We get the conclusion that NaHS attenuated TGF-β1-induced EMT in HK-2 cells through both ERK-dependent and β-catenin-dependent pathways. PMID:26760502

  1. Genetic variation in Transforming Growth Factor beta 1 and mammographic density in Singapore Chinese women

    PubMed Central

    Lee, Eunjung; Van den Berg, David; Hsu, Chris; Ursin, Giske; Koh, Woon-Puay; Yuan, Jian-Min; Stram, Daniel O.; Yu, Mimi C.; Wu, Anna H.

    2013-01-01

    Transforming growth factor-beta (TGF-β) plays a critical role in normal mammary development and morphogenesis. Decreased TGF-β signaling has been associated with increased mammographic density. Percent mammographic density (PMD) adjusted for age and body mass index (BMI) is a strong risk factor and predictor of breast cancer risk. PMD is highly heritable, but few genetic determinants have been identified. We investigated the association between genetic variation in TGFB1 and PMD using a cross-sectional study of 2,038 women who were members of the population-based Singapore Chinese Health Study cohort. We assessed PMD using a computer-assisted method. We used linear regression to examine the association between 9 tagging SNPs of TGFB1 and PMD and their interaction with parity, adjusting for age, BMI, and dialect group. We calculated ‘P-values adjusted for correlated tests’ (PACT) to account for multiple testing. The strongest association was observed for rs2241716. Adjusted PMD was higher by 1.5% per minor allele (PACT =0.04). When stratifying by parity, this association was limited to nulliparous women. For nulliparous women, adjusted PMD was higher by 8.6% per minor allele (PACT=0.003; P for interaction with parity=0.002). Three additional TGFB1 tagging SNPs, which were in linkage disequilibrium with rs2241716, were statistically significantly associated with adjusted PMD (PACT<0.05) for nulliparous women. However, none of these three SNPs showed statistically significant association after adjusting for rs2241716. Our data support that TGFB1 genetic variation may be an important genetic determinant of mammographic density measure that predicts breast cancer risk, particularly in nulliparous women. PMID:23333936

  2. Transforming growth factor-β mediates endothelial dysfunction in rats during high salt intake.

    PubMed

    Feng, Wenguang; Ying, Wei-Zhong; Aaron, Kristal J; Sanders, Paul W

    2015-12-15

    Endothelial dysfunction has been shown to be predictive of subsequent cardiovascular events and death. Through a mechanism that is incompletely understood, increased dietary salt intake promotes endothelial dysfunction in healthy, salt-resistant humans. The present study tested the hypothesis that dietary salt-induced transforming growth factor (TGF)-β promoted endothelial dysfunction and salt-dependent changes in blood pressure (BP). Sprague-Dawley rats that received diets containing 0.3% NaCl [low salt (LS)] or 8.0% NaCl [high salt (HS)] were treated with vehicle or SB-525334, a specific inhibitor of TGF-β receptor I/activin receptor-like kinase 5, beginning on day 5. BP was monitored using radiotelemetry in four groups of rats (LS, LS + SB-525334, HS, and HS + SB-525334) for up to 14 days. By day 14 of the study, mean daytime systolic BP and mean pulse pressure of the HS group treated with vehicle was greater than those in the other three groups; mean daytime systolic BP and pulse pressure of the HS + SB-525334 group did not differ from the LS and LS + SB-525334-treated groups. Whereas mean systolic BP, mean diastolic BP, and mean arterial pressure did not differ among the groups on the seventh day of the study, endothelium-dependent vasorelaxation was impaired specifically in the HS group; treatment with the activin receptor-like kinase 5 inhibitor prevented the dietary HS intake-induced increases in phospho-Smad2 (Ser(465/467)) and NADPH oxidase-4 in endothelial lysates and normalized endothelial function. These findings suggest that HS-induced endothelial dysfunction and the development of salt-dependent increases in BP were related to endothelial TGF-β signaling. PMID:26447221

  3. Transforming growth factor-β3 promotes facial nerve injury repair in rabbits

    PubMed Central

    WANG, YANMEI; ZHAO, XINXIANG; HUOJIA, MUHTER; XU, HUI; ZHUANG, YOUMEI

    2016-01-01

    The present study investigated the effects of transforming growth factor (TGF)-β3 on the regeneration of facial nerves in rabbits. A total of 20 adult rabbits were randomly divided into three equal groups: Normal control (n=10), surgical control (n=10) and TGF-β3 treatment (n=10). The total number and diameter of the regenerated nerve fibers was significantly increased in the TGF-β3 treatment group, as compared with in the surgical control group (P<0.01). Furthermore, in the TGF-β3 treatment group, the epineurial repair of the facial nerves was intact and the nerve fibers, which were arranged in neat rows, were morphologically intact with visible myelin swelling. However, in the surgical control group, the epineurial repair was incomplete, as demonstrated by: Atrophic nerve fibers, partially disappeared axons and myelin of uneven thickness with fuzzy borders. Electron microscopy demonstrated that the regenerated fibers in the TGF-β3 treatment group were predominantly myelinated, with clear-layered myelin sheath structures and axoplasms rich in organelles. Although typical layered myelin sheath structures were observed in the surgical control group, the myelin sheaths of the myelinated nerve fibers were poorly developed and few organelles were detected in the axoplasms. Neuro-electrophysiological examination demonstrated that, as compared with the surgical control group, the latency period of the action potentials in the TGF-β3 treatment group were shorter, whereas the stimulus amplitudes of the action potentials were significantly increased (P<0.01). The results of the present study suggest that TGF-β3 may improve the regeneration of facial nerves following trauma or injury. PMID:26997982

  4. Disruption of transforming growth factor beta signaling by a novel ligand-dependent mechanism.

    PubMed

    Fernandez, Tania; Amoroso, Stephanie; Sharpe, Shellyann; Jones, Gary M; Bliskovski, Valery; Kovalchuk, Alexander; Wakefield, Lalage M; Kim, Seong-Jin; Potter, Michael; Letterio, John J

    2002-05-20

    Transforming growth factor (TGF)-beta is the prototype in a family of secreted proteins that act in autocrine and paracrine pathways to regulate cell development and function. Normal cells typically coexpress TGF-beta receptors and one or more isoforms of TGF-beta, thus the synthesis and secretion of TGF-beta as an inactive latent complex is considered an essential step in regula-ting the activity of this pathway. To determine whether intracellular activation of TGF-beta results in TGF-beta ligand-receptor interactions within the cell, we studied pristane-induced plasma cell tumors (PCTs). We now demonstrate that active TGF-beta1 in the PCT binds to intracellular TGF-beta type II receptor (TbetaRII). Disruption of the expression of TGF-beta1 by antisense TGF-beta1 mRNA restores localization of TbetaRII at the PCT cell surface, indicating a ligand-induced impediment in receptor trafficking. We also show that retroviral expression of a truncated, dominant-negative TbetaRII (dnTbetaRII) effectively competes for intracellular binding of active ligand in the PCT and restores cell surface expression of the endogenous TbetaRII. Analysis of TGF-beta receptor-activated Smad2 suggests the intracellular ligand-receptor complex is not capable of signaling. These data are the first to demonstrate the formation of an intracellular TGF-beta-receptor complex, and define a novel mechanism for modulating the TGF-beta signaling pathway. PMID:12021305

  5. Transforming Growth Factor-β2 Induces Bronchial Epithelial Mucin Expression in Asthma

    PubMed Central

    Chu, Hong Wei; Balzar, Silvana; Seedorf, Gregory J.; Westcott, Jay Y.; Trudeau, John B.; Silkoff, Phil; Wenzel, Sally E.

    2004-01-01

    The transforming growth factor (TGF)-β family is important for tissue repair in pathological conditions including asthma. However, little is known about the impact of either TGF-β1 or TGF-β2 on asthmatic airway epithelial mucin expression. We evaluated bronchial epithelial TGF-β1 and TGF-β2 expression and their effects on mucin expression, and the role of TGF-β1 or TGF-β2 in interleukin (IL)-13-induced mucin expression. Epithelial TGF-β1, TGF-β2, and mucin expression were evaluated in endobronchial biopsies from asthmatics and normal subjects. The effects of TGF-β1 or TGF-β2 on mucin MUC5AC protein and mRNA expression, and the impact of IL-13 on epithelial TGF-β1, TGF-β2, and MUC5AC were determined in cultured bronchial epithelial cells from endobronchial brushings of both subject groups. In biopsy tissue, epithelial TGF-β2 expression levels were higher than TGF-β1 in both asthmatics and normals. TGF-β2, but not TGF-β1, was increased in asthmatics compared with normals, and significantly correlated with mucin expression. TGF-β2, but not TGF-β1, increased mucin expression in cultured epithelial cells from both subject groups. IL-13 increased the release of TGF-β2, but not TGF-β1, from epithelial cells. A neutralizing TGF-β2 antibody partially inhibited IL-13-induced mucin expression. These data suggest that TGF-β2 production by asthmatic bronchial epithelial cells may increase airway mucin expression. IL-13-induced mucin expression may occur in part through TGF-β2 up-regulation. PMID:15466377

  6. Transforming growth factor-beta2 induces bronchial epithelial mucin expression in asthma.

    PubMed

    Chu, Hong Wei; Balzar, Silvana; Seedorf, Gregory J; Westcott, Jay Y; Trudeau, John B; Silkoff, Phil; Wenzel, Sally E

    2004-10-01

    The transforming growth factor (TGF)-beta family is important for tissue repair in pathological conditions including asthma. However, little is known about the impact of either TGF-beta1 or TGF-beta2 on asthmatic airway epithelial mucin expression. We evaluated bronchial epithelial TGF-beta1 and TGF-beta2 expression and their effects on mucin expression, and the role of TGF-beta1 or TGF-beta2 in interleukin (IL)-13-induced mucin expression. Epithelial TGF-beta1, TGF-beta2, and mucin expression were evaluated in endobronchial biopsies from asthmatics and normal subjects. The effects of TGF-beta1 or TGF-beta2 on mucin MUC5AC protein and mRNA expression, and the impact of IL-13 on epithelial TGF-beta1, TGF-beta2, and MUC5AC were determined in cultured bronchial epithelial cells from endobronchial brushings of both subject groups. In biopsy tissue, epithelial TGF-beta2 expression levels were higher than TGF-beta1 in both asthmatics and normals. TGF-beta2, but not TGF-beta1, was increased in asthmatics compared with normals, and significantly correlated with mucin expression. TGF-beta2, but not TGF-beta1, increased mucin expression in cultured epithelial cells from both subject groups. IL-13 increased the release of TGF-beta2, but not TGF-beta1, from epithelial cells. A neutralizing TGF-beta2 antibody partially inhibited IL-13-induced mucin expression. These data suggest that TGF-beta2 production by asthmatic bronchial epithelial cells may increase airway mucin expression. IL-13-induced mucin expression may occur in part through TGF-beta2 up-regulation. PMID:15466377

  7. Redox-mediated activation of latent transforming growth factor-beta 1.

    PubMed

    Barcellos-Hoff, M H; Dix, T A

    1996-09-01

    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

  8. Orofacial clefts, parental cigarette smoking, and transforming growth factor-alpha gene variants

    SciTech Connect

    Shaw, G.M.; Wasserman, C.R.; O`Malley, C.D.

    1996-03-01

    Results of studies determine whether women who smoke during early pregnancy are at increased risk of delivering infants with orofacial clefts have been mixed, and recently a gene-environment interaction between maternal smoking, transforming growth factor-alpha (TGFa), and clefting has been reported. Using a large population-based case-control study, we investigated whether parental periconceptional cigarette smoking was associated with an increased risk for having offspring with orofacial clefts. We also investigated the influence of genetic variation of the TGFa locus on the relation between smoking and clefting. Parental smoking information was obtained from telephone interviews with mothers of 731 (84.7% of eligible) orofacial cleft case infants and with mothers of 734 (78.2%) nonmalformed control infants. DNA was obtained from newborn screening blood spots and genotyped for the allelic variants of TGFa. We found that risks associated with maternal smoking were most elevated for isolated cleft lip with or without cleft palate, (odds ratio 2.1 [95% confidence interval 1.3-3.6]) and for isolated cleft palate (odds ratio 2.2 [1.1-4.5]) when mothers smoked {ge} 20 cigarrettes/d. These risks for white infants ranged from 3-fold to 11-fold across phenotypic groups. Paternal smoking was not associated with clefting among the offspring of nonsmoking mothers, and passive smoke exposures were associated with at most slightly increased risks. This study offers evidence that the risk for orofacial clefting in infants may be influenced by maternal smoke exposures alone as well as in combination (gene-environment interaction) with the presence of the uncommon TGFa allele. 56 refs., 5 tabs.

  9. Regulation of Transforming Growth Factor-β1–driven Lung Fibrosis by Galectin-3

    PubMed Central

    MacKinnon, Alison C.; Gibbons, Michael A.; Farnworth, Sarah L.; Leffler, Hakon; Nilsson, Ulf J.; Delaine, Tamara; Simpson, A. John; Forbes, Stuart J.; Hirani, Nik; Gauldie, Jack

    2012-01-01

    Rationale: Idiopathic pulmonary fibrosis (IPF) is a chronic dysregulated response to alveolar epithelial injury with differentiation of epithelial cells and fibroblasts into matrix-secreting myofibroblasts resulting in lung scaring. The prognosis is poor and there are no effective therapies or reliable biomarkers. Galectin-3 is a β-galactoside binding lectin that is highly expressed in fibrotic tissue of diverse etiologies. Objectives: To examine the role of galectin-3 in pulmonary fibrosis. Methods: We used genetic deletion and pharmacologic inhibition in well-characterized murine models of lung fibrosis. Further mechanistic studies were performed in vitro and on samples from patients with IPF. Measurements and Main Results: Transforming growth factor (TGF)-β and bleomycin-induced lung fibrosis was dramatically reduced in mice deficient in galectin-3, manifest by reduced TGF-β1–induced EMT and myofibroblast activation and collagen production. Galectin-3 reduced phosphorylation and nuclear translocation of β-catenin but had no effect on Smad2/3 phosphorylation. A novel inhibitor of galectin-3, TD139, blocked TGF-β–induced β-catenin activation in vitro and in vivo and attenuated the late-stage progression of lung fibrosis after bleomycin. There was increased expression of galectin-3 in the bronchoalveolar lavage fluid and serum from patients with stable IPF compared with nonspecific interstitial pneumonitis and controls, which rose sharply during an acute exacerbation suggesting that galectin-3 may be a marker of active fibrosis in IPF and that strategies that block galectin-3 may be effective in treating acute fibrotic exacerbations of IPF. Conclusions: This study identifies galectin-3 as an important regulator of lung fibrosis and provides a proof of principle for galectin-3 inhibition as a potential novel therapeutic strategy for IPF. PMID:22095546

  10. Transforming growth factor-β and atherosclerosis: interwoven atherogenic and atheroprotective aspects

    PubMed Central

    Toma, Ian

    2016-01-01

    Age-related progression of cardiovascular disease is by far the largest health problem in the US and involves vascular damage, progressive vascular fibrosis and the accumulation of lipid-rich atherosclerotic lesions. Advanced lesions can restrict flow to key organs and can trigger occlusive thrombosis resulting in a stroke or myocardial infarction. Transforming growth factor-beta (TGF-β) is a major orchestrator of the fibroproliferative response to tissue damage. In the early stages of repair, TGF-β is released from platelets and activated from matrix reservoirs; it then stimulates the chemotaxis of repair cells, modulates immunity and inflammation and induces matrix production. At later stages, it negatively regulates fibrosis through its strong antiproliferative and apoptotic effects on fibrotic cells. In advanced lesions, TGF-β might be important in arterial calcification, commonly referred to as “hardening of the arteries”. Because TGF-β can signal through multiple pathways, namely the SMADs, a MAPK pathway and the Rho/ROCK pathways, selective defects in TGF-β signaling can disrupt otherwise coordinated pathways of tissue regeneration. TGF-β is known to control cell proliferation, cell migration, matrix synthesis, wound contraction, calcification and the immune response, all being major components of the atherosclerotic process. However, many of the effects of TGF-β are essential to normal tissue repair and thus, TGF-β is often thought to be “atheroprotective”. The present review attempts to parse systematically the known effects of TGF-β on both the major risk factors for atherosclerosis and to isolate the role of TGF-β in the many component pathways involved in atherogenesis. PMID:21626289

  11. Verprolin function in endocytosis and actin organization. Roles of the Las17p (yeast WASP)-binding domain and a novel C-terminal actin-binding domain.

    PubMed

    Thanabalu, Thirumaran; Rajmohan, Rajamuthiah; Meng, Lei; Ren, Gang; Vajjhala, Parimala R; Munn, Alan L

    2007-08-01

    Vrp1p (verprolin, End5p) is the yeast ortholog of human Wiskott-Aldrich syndrome protein (WASP)-interacting protein (WIP). Vrp1p localizes to the cortical actin cytoskeleton, is necessary for its polarization to sites of growth and is also essential for endocytosis. At elevated temperature, Vrp1p becomes essential for growth. A C-terminal Vrp1p fragment (C-Vrp1p) retains the ability to localize to the cortical actin cytoskeleton and function in actin-cytoskeleton polarization, endocytosis and growth. Here, we demonstrate that two submodules in C-Vrp1p are required for actin-cytoskeleton polarization: a novel C-terminal actin-binding submodule (CABS) that contains a novel G-actin-binding domain, which we call a verprolin homology 2 C-terminal (VH2-C) domain; and a second submodule comprising the Las17p-binding domain (LBD) that binds Las17p (yeast WASP). The LBD localizes C-Vrp1p to membranes and the cortical actin cytoskeleton. Intriguingly, the LBD is sufficient to restore endocytosis and growth at elevated temperature to Vrp1p-deficient cells. The CABS also restores these functions, but only if modified by a lipid anchor to provide membrane association. Our findings highlight the role of Las17p binding for Vrp1p membrane association, suggest general membrane association may be more important than specific targeting to the cortical actin cytoskeleton for Vrp1p function in endocytosis and cell growth, and suggest that Vrp1p binding to individual effectors may alter their physiological activity. PMID:17635585

  12. Expression of transfected mutant beta-actin genes: transitions toward the stable tumorigenic state.

    PubMed Central

    Leavitt, J; Ng, S Y; Varma, M; Latter, G; Burbeck, S; Gunning, P; Kedes, L

    1987-01-01

    Mutant human beta-actin genes were introduced into normal human (KD) fibroblasts and the derivative cell line HuT-12, which is immortalized but nontumorigenic, to test their ability to promote conversion to the tumorigenic state. Transfected substrains of HuT-12 fibroblasts that expressed abundant levels of mutant beta-actin (Gly-244----Asp-244) produced subcutaneous tumors in athymic mice after long latent periods (1.5 to 3 months). However, transfected substrains of KD fibroblasts retained their normal finite life span in culture and consequently were incapable of producing tumors. Substrains of HuT-12 cells transfected with the wild-type beta-actin gene and some transfected strains that expressed low or undetectable levels of mutant beta-actin did not produce tumors. Cell lines derived from transfectant cell tumors always exhibited elevated synthesis of the mutant beta-actin, ranging from 145 to 476% of the level expressed by the transfected cells that were inoculated to form the tumor. In general, primary transfectant cells that expressed the highest levels of mutant beta-actin were more tumorigenic than strains that expressed lower levels. The tumor-derived strains were stable in tumorigenicity and produced tumors with shortened latent periods of only 2 to 4 weeks. These findings imply that the primary transfectant strains develop subpopulations of cells that are selected to form tumors because of their elevated rate of exogenous mutant beta-actin synthesis. Actin synthesis and accumulation of gamma-actin mRNA from the endogenous beta- and gamma-actin genes were diminished in tumor-derived strains, apparently to compensate for elevated mutant beta-actin synthesis and maintain the normal cellular concentration of actin. Synthesis of the transformation-sensitive tropomyosin isoforms was decreased along with mutant beta-actin expression. Such modulations in tropomyosin synthesis are characteristically seen in transformation of avian, rodent, and human fibroblasts

  13. Notch1 signaling antagonizes transforming growth factor-β pathway and induces apoptosis in rabbit trophoblast stem cells.

    PubMed

    Tan, Tao; Lu, Bin; Zhang, Jing; Niu, Yuyu; Si, Wei; Wei, Qiang; Ji, Weizhi

    2014-04-15

    During mammalian development, placental growth needs to be tightly controlled by apoptosis. However, despite the potentially significant problems, the strategies used to balance growth and apoptosis have remained elusive. Here we report that activation of the Notch1 signal pathway inhibits transduction of transforming growth factor (TGF)-β signaling, which leads to cell cycle arrest and apoptosis in rabbit trophoblast stem cells (TSCs). The subcellular location of notch intracellular domain 1 (NICD1) appears to determine whether TGF-β signaling will be inhibited or not. Moreover, changes in NICD1 subcellular location are regulated by intracellular calcium distribution. Collectively, these results establish a potential mechanism whereby TSCs can balance growth and apoptosis, and thus guarantee the development of the fetus. PMID:24251382

  14. ER sheet persistence is coupled to myosin 1c-regulated dynamic actin filament arrays.

    PubMed

    Joensuu, Merja; Belevich, Ilya; Rämö, Olli; Nevzorov, Ilya; Vihinen, Helena; Puhka, Maija; Witkos, Tomasz M; Lowe, Martin; Vartiainen, Maria K; Jokitalo, Eija

    2014-04-01

    The endoplasmic reticulum (ER) comprises a dynamic three-dimensional (3D) network with diverse structural and functional domains. Proper ER operation requires an intricate balance within and between dynamics, morphology, and functions, but how these processes are coupled in cells has been unclear. Using live-cell imaging and 3D electron microscopy, we identify a specific subset of actin filaments localizing to polygons defined by ER sheets and tubules and describe a role for these actin arrays in ER sheet persistence and, thereby, in maintenance of the characteristic network architecture by showing that actin depolymerization leads to increased sheet fluctuation and transformations and results in small and less abundant sheet remnants and a defective ER network distribution. Furthermore, we identify myosin 1c localizing to the ER-associated actin filament arrays and reveal a novel role for myosin 1c in regulating these actin structures, as myosin 1c manipulations lead to loss of the actin filaments and to similar ER phenotype as observed after actin depolymerization. We propose that ER-associated actin filaments have a role in ER sheet persistence regulation and thus support the maintenance of sheets as a stationary subdomain of the dynamic ER network. PMID:24523293

  15. ER sheet persistence is coupled to myosin 1c–regulated dynamic actin filament arrays

    PubMed Central

    Joensuu, Merja; Belevich, Ilya; Rämö, Olli; Nevzorov, Ilya; Vihinen, Helena; Puhka, Maija; Witkos, Tomasz M.; Lowe, Martin; Vartiainen, Maria K.; Jokitalo, Eija

    2014-01-01

    The endoplasmic reticulum (ER) comprises a dynamic three-dimensional (3D) network with diverse structural and functional domains. Proper ER operation requires an intricate balance within and between dynamics, morphology, and functions, but how these processes are coupled in cells has been unclear. Using live-cell imaging and 3D electron microscopy, we identify a specific subset of actin filaments localizing to polygons defined by ER sheets and tubules and describe a role for these actin arrays in ER sheet persistence and, thereby, in maintenance of the characteristic network architecture by showing that actin depolymerization leads to increased sheet fluctuation and transformations and results in small and less abundant sheet remnants and a defective ER network distribution. Furthermore, we identify myosin 1c localizing to the ER-associated actin filament arrays and reveal a novel role for myosin 1c in regulating these actin structures, as myosin 1c manipulations lead to loss of the actin filaments and to similar ER phenotype as observed after actin depolymerization. We propose that ER-associated actin filaments have a role in ER sheet persistence regulation and thus support the maintenance of sheets as a stationary subdomain of the dynamic ER network. PMID:24523293

  16. Rapid Glucose Depletion Immobilizes Active Myosin-V on Stabilized Actin Cables

    PubMed Central

    Xu, Li; Bretscher, Anthony

    2014-01-01

    Summary Polarization of eukaryotic cells requires organelles and protein complexes to be transported to their proper destinations along the cytoskeleton [1]. When nutrients are abundant, budding yeast grows rapidly transporting secretory vesicles for localized growth and actively segregating organelles [2, 3]. This is mediated by myosin-Vs transporting cargos along F-actin bundles known as actin cables [4]. Actin cables are dynamic structures regulated by assembly, stabilization and disassembly [5]. Polarized growth and actin filament dynamics consume energy. For most organisms, glucose is the preferred energy source and generally represses alternative carbon source usage [6]. Thus upon abrupt glucose depletion, yeast shuts down pathways consuming large amounts of energy, including the vacuolar-ATPase [7, 8], translation [9] and phosphoinositide metabolism [10]. Here we show that glucose withdrawal rapidly (<1 min) depletes ATP levels and the yeast myosin V, Myo2, responds by relocalizing to actin cables, making it the fastest response documented. Myo2 immobilized on cables releases its secretory cargo, defining a new rigor-like state of a myosin-V in vivo. Only actively transporting Myo2 can be converted to the rigor-like state. Glucose depletion has differential effects on the actin cytoskeleton resulting in disassembly of actin patches with concomitant inhibition of endocytosis, and strong stabilization of actin cables, thereby revealing a selective and previously unappreciated ATP requirement for actin cable disassembly. A similar response is seen in HeLa cells to ATP depletion. These findings reveal a new fast-acting energy conservation strategy halting growth by immobilizing myosin-V in a newly described state on selectively stabilized actin cables. PMID:25308080

  17. Association of actin with alpha crystallins

    NASA Technical Reports Server (NTRS)

    Gopalakrishnan, S.; Boyle, D.; Takemoto, L.; Spooner, B. S. (Principal Investigator)

    1993-01-01

    The alpha crystallins are cytosolic proteins that co-localize and co-purify with actin-containing microfilaments. Affinity column chromatography employing both covalently-coupled actin or alpha crystallin was used to demonstrate specific and saturable binding of actin with alpha crystallin. This conclusion was confirmed by direct visualization of alpha aggregates bound to actin polymerized in vitro. The significance of this interaction in relation to the functional properties of these two polypeptides will be discussed.

  18. An actin cytoskeleton with evolutionarily conserved functions in the absence of canonical actin-binding proteins

    PubMed Central

    Paredez, Alexander R.; Assaf, Zoe June; Sept, David; Timofejeva, Ljudmilla; Dawson, Scott C.; Wang, Chung-Ju Rachel; Cande, W. Z.

    2011-01-01

    Giardia intestinalis, a human intestinal parasite and member of what is perhaps the earliest-diverging eukaryotic lineage, contains the most divergent eukaryotic actin identified to date and is the first eukaryote known to lack all canonical actin-binding proteins (ABPs). We sought to investigate the properties and functions of the actin cytoskeleton in Giardia to determine whether Giardia actin (giActin) has reduced or conserved roles in core cellular processes. In vitro polymerization of giActin produced filaments, indicating that this divergent actin is a true filament-forming actin. We generated an anti-giActin antibody to localize giActin throughout the cell cycle. GiActin localized to the cortex, nuclei, internal axonemes, and formed C-shaped filaments along the anterior of the cell and a flagella-bundling helix. These structures were regulated with the cell cycle and in encysting cells giActin was recruited to the Golgi-like cyst wall processing vesicles. Knockdown of giActin demonstrated that giActin functions in cell morphogenesis, membrane trafficking, and cytokinesis. Additionally, Giardia contains a single G protein, giRac, which affects the Giardia actin cytoskeleton independently of known target ABPs. These results imply that there exist ancestral and perhaps conserved roles for actin in core cellular processes that are independent of canonical ABPs. Of medical significance, the divergent giActin cytoskeleton is essential and commonly used actin-disrupting drugs do not depolymerize giActin structures. Therefore, the giActin cytoskeleton is a promising drug target for treating giardiasis, as we predict drugs that interfere with the Giardia actin cytoskeleton will not affect the mammalian host. PMID:21444821

  19. Unidirectional movement of an actin filament taking advantage of temperature gradients.

    PubMed

    Kawaguchi, Tomoaki; Honda, Hajime

    2007-01-01

    An actin filament with heat acceptors attached to its Cys374 residue in each actin monomer could move unidirectionally even under heat pulsation alone, while in the total absence of both ATP and myosin. The prime driver for the movement was temperature gradients operating between locally heated portions on an actin filament and its cooler surroundings. In this report, we investigated how the mitigation of the temperature gradients induces a unidirectional movement of an actin filament. We then observed the transversal fluctuations of the filament in response to heat pulsation and their transition into longitudinally unidirectional movement. The transition was significantly accelerated when Cys374 and Lys336 were simultaneously excited within an actin monomer. These results suggest that the mitigation of the temperature gradients within each actin monomer first went through the energy transformation to transversal fluctuations of the filament, and then followed by the transformation further down to longitudinal movements of the filament. The faster mitigation of temperature gradients within actin monomer helps build up the transition from the transversal to longitudinal movements of the filament by coordinating the interaction between the neighboring monomers. PMID:17030086

  20. 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. PMID:20082218

  1. Actin dynamics: from nanoscale to microscale.

    PubMed

    Carlsson, Anders E

    2010-01-01

    The dynamic nature of actin in cells manifests itself constantly. Polymerization near the cell edge is balanced by depolymerization in the interior, externally induced actin polymerization is followed by depolymerization, and spontaneous oscillations of actin at the cell periphery are frequently seen. I discuss how mathematical modeling relates quantitative measures of actin dynamics to the rates of underlying molecular level processes. The dynamic properties addressed include the rate of actin assembly at the leading edge of a moving cell, the disassembly rates of intracellular actin networks, the polymerization time course in externally stimulated cells, and spontaneous spatiotemporal patterns formed by actin. Although several aspects of actin assembly have been clarified by increasingly sophisticated models, our understanding of rapid actin disassembly is limited, and the origins of nonmonotonic features in externally stimulated actin polymerization remain unclear. Theory has generated several concrete, testable hypotheses for the origins of spontaneous actin waves and cell-edge oscillations. The development and use of more biomimetic systems applicable to the geometry of a cell will be key to obtaining a quantitative understanding of actin dynamics in cells. PMID:20462375

  2. Yersinia effector YopO uses actin as bait to phosphorylate proteins that regulate actin polymerization

    PubMed Central

    Lee, Wei Lin; Grimes, Jonathan M; Robinson, Robert C

    2016-01-01

    Pathogenic Yersinia species evade host immune systems through the injection of Yersinia outer proteins (Yops) into phagocytic cells. One Yop, YopO, also known as YpkA, induces actin-filament disruption, impairing phagocytosis. Here we describe the X-ray structure of Yersinia enterocolitica YopO in complex with actin, which reveals that YopO binds to an actin monomer in a manner that blocks polymerization yet allows the bound actin to interact with host actin-regulating proteins. SILAC-MS and biochemical analyses confirm that actin-polymerization regulators such as VASP, EVL, WASP, gelsolin and the formin diaphanous 1 are directly sequestered and phosphorylated by YopO through formation of ternary complexes with actin. This leads to a model in which YopO at the membrane sequesters actin from polymerization while using the bound actin as bait to recruit, phosphorylate and misregulate host actin-regulating proteins to disrupt phagocytosis. PMID:25664724

  3. Ingenol mebutate gel treatment for actinic cheilitis: report of four cases.

    PubMed

    Barrado Solís, Nerea; Molés Poveda, Paula; Lloret Ruiz, César; Pont Sanjuan, Virginia; Velasco Pastor, Manel; Quecedo Estébanez, Esther; Miquel Miquel, Javier

    2015-01-01

    Actinic cheilitis (AC) are premalignant lesions that have an increased risk of malignant transformation. Their treatment, therefore, is essential to prevent carcinogenesis. However, optimal therapy is not well established and different modalities yield variable results. Ingenol mebutate gel has recently been approved by the US Food and Drug Administration for topical treatment of actinic keratosis, with high clearance rates. On the basis of these findings, we report our experience with this drug for the treatment of AC. PMID:25545762

  4. Aluminum Induces Rigor within the Actin Network of Soybean Cells.

    PubMed Central

    Grabski, S.; Schindler, M.

    1995-01-01

    Aluminum is toxic to both plants and animals. Root growth and pollen-tube extension are inhibited after aluminum stress in acidic environments. Incubation of cultured neurons with aluminum results in the formation of neurofibrillar tangles reminiscent of the neural pathology observed in Alzheimer's disease. The present communication demonstrates that aluminum induces a rapid and dramatic increase in the rigidity of the actin network in soybean (Glycine max) root cells. This rigidity can be prevented by either co-incubation with sodium fluoride or magnesium, or pretreatment with cytochalasin D. It is proposed that the growth-inhibitory activity and cytotoxicity of aluminum in plants may be a consequence of a global rigor that is induced within the actin network. This rigor may result from the formation of nonhydrolyzable [Al3+-ADP] or [Al3+-ATP] complexes whose binding to actin/myosin can modify contraction. Additionally, Al3+-mediated interference with the normal kinetics of F-actin filament assembly/disassembly could precipitate subsequent disorganization of associated cytoskeletal structures and promote altered expression of cytoskeletal proteins. PMID:12228515

  5. Induction of Transforming Growth Factor Beta Receptors following Focal Ischemia in the Rat Brain

    PubMed Central

    Pál, Gabriella; Lovas, Gábor; Dobolyi, Arpád

    2014-01-01

    Transforming growth factor-βs (TGF-βs) regulate cellular proliferation, differentiation, and survival. TGF-βs bind to type I (TGF-βRI) and II receptors (TGF-βRII), which are transmembrane kinase receptors, and an accessory type III receptor (TGF-βRIII). TGF-β may utilize another type I receptor, activin-like kinase receptor (Alk1). TGF-β is neuroprotective in the middle cerebral artery occlusion (MCAO) model of stroke. Recently, we reported the expression pattern of TGF-β1-3 after MCAO. To establish how TGF-βs exert their actions following MCAO, the present study describes the induction of TGF-βRI, RII, RIII and Alk1 at 24 h, 72 h and 1 mo after transient 1 h MCAO as well as following 24 h permanent MCAO using in situ hybridization histochemistry. In intact brain, only TGF-βRI had significant expression: neurons in cortical layer IV contained TGF-βRI. At 24 h after the occlusion, no TGF-β receptors showed induction. At 72 h following MCAO, all four types of TGF-β receptors were induced in the infarct area, while TGF-βRI and RII also appeared in the penumbra. Most cells with elevated TGF-βRI mRNA levels were microglia. TGF-βRII co-localized with both microglial and endothelial markers while TGF-βRIII and Alk1 were present predominantly in endothels. All four TGF-β receptors were induced within the lesion 1 mo after the occlusion. In particular, TGF-βRIII was further induced as compared to 72 h after MCAO. At this time point, TGF-βRIII signal was predominantly not associated with blood vessels suggesting its microglial location. These data suggest that TGF-β receptors are induced after MCAO in a timely and spatially regulated fashion. TGF-β receptor expression is preceded by increased TGF-β expression. TGF-βRI and RII are likely to be co-expressed in microglial cells while Alk1, TGF-βRII, and RIII in endothels within the infarct where TGF-β1 may be their ligand. At later time points, TGF-βRIII may also appear in glial cells to potentially

  6. Expression of transforming growth factor β receptor II in mesenchymal stem cells from systemic sclerosis patients

    PubMed Central

    Vanneaux, Valérie; Farge-Bancel, Dominique; Lecourt, Séverine; Baraut, Julie; Cras, Audrey; Jean-Louis, Francette; Brun, Cécilia; Verrecchia, Franck; Larghero, Jérôme; Michel, Laurence

    2013-01-01

    Objectives The present work aimed to evaluate the expression of transforming growth factor-β (TGF-β) receptors on bone marrow-derived multipotent mesenchymal stromal cells (MSCs) in patients with systemic sclerosis (SSc) and the consequences of TGF-β activation in these cells, since MSC have potential therapeutic interest for SSc patients and knowing that TGF-β plays a critical role during the development of fibrosis in SSc. Design This is a prospective research study using MSC samples obtained from SSc patients and compared with MSC from healthy donors. Setting One medical hospital involving collaboration between an internal medicine department for initial patient recruitment, a clinical biotherapeutic unit for MSC preparation and an academic laboratory for research. Participants 9 patients with diffuse SSc for which bone marrow (BM) aspiration was prescribed by sternum aspiration before haematopoietic stem cell transplantation, versus nine healthy donors for normal BM. Primary and secondary outcome measures TGF-β, TGF-β receptor types I (TBRI) and II (TBRII) mRNA and protein expression were assessed by quantitative PCR and flow cytometry, respectively, in MSC from both SSc patients and healthy donors. MSC were exposed to TGF-β and assessed for collagen 1α2 synthesis and Smad expression. As positive controls, primary cultures of dermal fibroblasts were also analysed. Results Compared with nine controls, MSC from nine SSc patients showed significant increase in mRNA levels (p<0.002) and in membrane expression (p<0.0001) of TBRII. In response to TGF-β activation, a significant increase in collagen 1α synthesis (p<0.05) and Smad-3 phosphorylation was upregulated in SSc MSC. Similar results were obtained on eight SSc-derived dermal fibroblasts compared to six healthy controls. Conclusions TBRII gene and protein expression defect in MSC derived from SSc patients may have pathological significance. These findings should be taken into account when considering

  7. Regulation of type II transforming-growth-factor-beta receptors by protein kinase C iota.

    PubMed Central

    Chuang, Lea-Yea; Guh, Jinn-Yuh; Liu, Shu-Fen; Hung, Min-Yuan; Liao, Tung-Nan; Chiang, Tai-An; Huang, Jau-Shyang; Huang, Yu-Lun; Lin, Chi-Fong; Yang, Yu-Lin

    2003-01-01

    TGF-beta (transforming growth factor-beta) is implicated in the pathogenesis of diabetic nephropathy. We previously demonstrated that up-regulation of type II TGF-beta receptor (TbetaRII) induced by high glucose might contribute to distal tubular hypertrophy [Yang, Guh, Yang, Lai, Tsai, Hung, Chang and Chuang (1998) J. Am. Soc. Nephrol. 9, 182-193]. We have elucidated the mechanism by using cultured Madin-Darby canine kidney cells. Enhancer assay and electrophoretic-mobility-shift assay were used to estimate the involvement of transcription factors. Western blotting and an in vitro kinase assay were used to evaluate the level and activity of protein kinase. We showed that glucose (100-900 mg/dl) induced an increase in mRNA level and promoter activity of TbetaRII (note: 'mg/dl' are the units commonly used in diabetes studies). The promoter region -209 to -177 appeared to contribute to positive transactivation of TbetaRII promoter by comparing five TbetaRII-promoter-CAT (chloramphenicol acetyl-transferase) plasmids. Moreover, the transcription factor AP-1 (activator protein 1) was significantly activated and specifically binds to TbetaRII promoter (-209 to -177). More importantly, we found that atypical PKC iota might be pivotal for high glucose-induced increase in both AP-1 binding and TbetaRII promoter activity. First, high glucose induced cytosolic translocation, activation and autophosphorylation of PKC iota. Secondly, antisense PKC iota expression plasmids attenuated high-glucose-induced increase in AP-1 binding and TbetaRII promoter activity; moreover, sense PKC iota expression plasmids enhanced these instead. Finally, we showed that antisense PKC iota expression plasmids might partly attenuate a high-glucose/TGF-beta1-induced increase in fibronectin. We conclude that PKC iota might mediate high-glucose-induced increase in TbetaRII promoter activity. In addition, antisense PKC iota expression plasmid effectively suppressed up-regulation of TbetaRII and

  8. Angiotensin II stimulated expression of transforming growth factor-beta1 in cardiac fibroblasts and myofibroblasts.

    PubMed

    Campbell, S E; Katwa, L C

    1997-07-01

    Angiotensin II (Ang II) stimulates pathologic myocardial fibrosis. Cardiac fibroblasts (CFb) and myofibroblasts mediate this response, perhaps in part by indirect production of specific cytokines. We sought to determine if Ang II could stimulate transforming growth factor-beta1 (TGF-beta1) gene expression and protein production in adult rat CFb and two cardiac myofibroblast cell types, scar myofibroblasts (MyoFb) and valvular interstitial cells (VIC). Confluent CFb, MyoFb, and VIC in serum-deprived (0.4% FCS) media were treated with Ang II (10(-7) m for CFb; 10(-9) m for MyoFb, VIC) for 24 h. Untreated cells served as controls. Culture media was collected and TGF-beta1 levels determined in triplicate using a sandwich ELISA. Reverse transcriptase-polymerase chain reaction (RT-PCR) analysis was performed to determine TGF-beta1 mRNA expression. Ang II increased CFb (P<0.02) and VIC (P<0.04) TGF-beta1 mRNA expression, while the increase in MyoFb was not statistically significant. MyoFb produced the highest TGF-beta1 levels under control conditions compared to VIC and CFb. Ang II stimulated further TGF-beta1 secretion in VIC and CFb, but not MyoFb. The AT1 receptor antagonist Losartan (10(-7) m) greatly attenuated Ang II-stimulated TGF-B1 secretion and decreased TGF-beta1 immunostaining in VIC. The AT2 receptor antagonist PD123177 (10(-7) m) also decreased secretion and immunostaining of TGF-beta1 in VIC, but to a lesser extent than Losartan. TGF-beta1 secretion by MyoFb was unaffected by Losartan and PD123177, although TGF-B1 immunostaining was absent or greatly decreased, respectively, compared to Ang II-treated MyoFb. Ang II stimulates TGF-beta1 gene expression and/or protein production in cardiac fibroblast-like cells which may act as an autocrine/paracrine stimulus to collagen formation. Furthermore, TGF-beta1 production and secretion in these cells can be modulated by specific Ang II receptor antagonists, suggesting a potential benefit in preventing

  9. Expression of transforming growth factors beta-1, beta 2 and beta 3 in human bladder carcinomas.

    PubMed Central

    Eder, I. E.; Stenzl, A.; Hobisch, A.; Cronauer, M. V.; Bartsch, G.; Klocker, H.

    1997-01-01

    We previously detected elevated transforming growth factor beta-1 (TGF-beta1) serum levels in patients with invasive bladder carcinomas. In this study, we therefore investigated whether elevated serum levels correlate with enhanced TGF-beta expression in human bladder tumours. mRNA levels of TGF-beta1, -beta2 and -beta3 were reduced in bladder tumour tissue to 86%, 68% and 56%, respectively, of the levels in normal urothelium. On the other hand, TGF-beta1 protein levels were found to be higher in superficial tumours (Ta-T1) (mean level of 0.153 ng mg(-1)) and in invasive T2/T3 tumours (mean level of 0.104 ng mg(-1)) compared with normal urothelium (mean level of 0.065 ng mg(-1)). Invasive T4 tumours, however, contained only low amounts of TGF-beta1 (mean level of 0.02 ng mg(-1)). Neither in mean nor in individual patients were serum and tissue TGF-beta levels correlated with each other. Cell culture experiments on primary bladder cells revealed a 57% decrease in TGF-beta1 mRNA levels in tumour compared with normal epithelial cells. Tumour epithelial cells contained about two times higher levels of TGF-beta2 and TGF-beta3 mRNA than normal epithelial cells. Fibroblasts expressed about the same amount of TGF-beta1 or TGF-beta2 as epithelial cells. Yet, fibroblasts released only 19% and 13% of the amount secreted by tumour epithelial cells into the supernatant. TGF-beta3, on the other hand, was expressed by fibroblasts with higher levels than by epithelial cells. TGF-beta1 was the predominent isoform in bladder tissue and cells at protein as well as on mRNA levels indicating that TGFs-beta2 and -beta3 are of minor importance in bladder cancer. In summary, there is a lack of correlation between TGF-beta serum levels and TGF-beta expression in tumour tissue in bladder cancer. Images Figure 1 PMID:9192977

  10. Transforming growth factor-betas and their signaling receptors are coexpressed in Crohn's disease.

    PubMed Central

    di Mola, F F; Friess, H; Scheuren, A; Di Sebastiano, P; Graber, H; Egger, B; Zimmermann, A; Korc, M; Büchler, M W

    1999-01-01

    OBJECTIVE: To evaluate mechanisms that contribute to tissue repair and tissue remodeling in Crohn's disease (CD). SUMMARY BACKGROUND DATA: Transforming growth factor-betas (TGF-betas) are involved in different chronic inflammatory disorders. They function by binding to two receptors, type I (TbetaR-I) subtype ALK5 and type II (TbetaR-II), which are concomitantly required for signal transduction. METHODS: Tissues were obtained from 18 patients with CD (10 female patients, 8 male patients, median age 38.7 years [range 16 to 58 years]) undergoing surgery because of CD-related complications. Tissue samples of 18 healthy organ donors (10 female subjects, 8 male subjects, median age 50.3 years [range 15 to 65 years]) served as controls. The expression and localization of TGF-beta1, TGF-beta2, TGF-beta3, TbetaR-IALK5, TbetaR-II, and TbetaR-III were studied by Northern blot analysis, in situ hybridization, and immunohistochemistry. RESULTS: On Northern blot analysis, 94% of the CD samples exhibited enhanced TGF-beta1, TGF-beta3, and TbetaR-II mRNA expression compared with controls. TGF-beta2 was increased in 72%, TbetaR-IALK5 in 72%, and TbetaR-III in 82% of the patients with CD. On in situ hybridization and immunohistochemical analysis, TGF-beta1, TbetaR-IALK5, and TbetaR-II were seen to be colocalized in the lamina propria cells and in the lymphocytes closest to the luminal surface, but also in the remaining epithelial cells, and in fibroblasts of CD tissue samples. CONCLUSIONS: The concomitant overexpression of TGF-betas and their signaling receptors in CD points to a potential role of these regulatory molecules in the pathophysiology of CD. Activation of TGF-beta-mediated pathways might promote the repair of mucosal injury by enhancing the process of reepithelization, but might also contribute to extracellular matrix generation and subsequently to intramural fibrosis and intestinal obstruction. Images Figure 1. Figure 3. Figure 4. Figure 5. PMID:9923802

  11. Urinary active transforming growth factor β in feline chronic kidney disease.

    PubMed

    Lawson, J S; Syme, H M; Wheeler-Jones, C P D; Elliott, J

    2016-08-01

    The cytokine transforming growth factor beta 1 (TGF-β1) has been widely implicated in the development and progression of renal fibrosis in chronic kidney disease (CKD) in humans and in experimental models. The aims of this study were to assess the association between urinary active TGF-β1 and (a) development of CKD in a cross-sectional study, (b) deterioration of renal function over 1 year in a longitudinal study, and (c) renal histopathological parameters in cats. A human active TGF-β1 ELISA was validated for use in feline urine. Cross-sectional analysis revealed no significant difference in urinary active TGF-β1:creatinine ratio (aTGF-β1:UCr) between groups with differing renal function. Longitudinally, non-azotaemic cats that developed CKD demonstrated a significant (P = 0.028) increase in aTGF-β1:UCr approximately 6 months before the development of azotaemia, which remained elevated (P = 0.046) at diagnosis (approximately 12 months prior, 8.4 pg/mg; approximately 6 months prior, 22.2 pg/mg; at CKD diagnosis, 24.6 pg/mg). In the histopathology study, aTGF-β1:UCr was significantly higher in cats with moderate (P = 0.02) and diffuse (P = 0.005) renal fibrosis than in cats without fibrosis. Cats with moderate renal inflammation had significantly higher urinary active aTGF-β1 concentrations than cats with mild (P = 0.035) or no inflammatory change (P = 0.004). The parameter aTGF-β1:UCr was independently associated with Log urine protein:creatinine ratio in a multivariable analysis of clinicopathological parameters and interstitial fibrosis score in a multivariable analysis of histopathological features. These results suggest that urinary aTGF-β1 reflects the severity of renal pathology. Increases in urinary aTGF-β1 followed longitudinally in individual cats may indicate the development of CKD. PMID:27387717

  12. A RhoA and Rnd3 cycle regulates actin reassembly during membrane blebbing.

    PubMed

    Aoki, Kana; Maeda, Fumiyo; Nagasako, Tomoya; Mochizuki, Yuki; Uchida, Seiichi; Ikenouchi, Junichi

    2016-03-29

    The actin cytoskeleton usually lies beneath the plasma membrane. When the membrane-associated actin cytoskeleton is transiently disrupted or the intracellular pressure is increased, the plasma membrane detaches from the cortex and protrudes. Such protruded membrane regions are called blebs. However, the molecular mechanisms underlying membrane blebbing are poorly understood. This study revealed that epidermal growth factor receptor kinase substrate 8 (Eps8) and ezrin are important regulators of rapid actin reassembly for the initiation and retraction of protruded blebs. Live-cell imaging of membrane blebbing revealed that local reassembly of actin filaments occurred at Eps8- and activated ezrin-positive foci of membrane blebs. Furthermore, we found that a RhoA-ROCK-Rnd3 feedback loop determined the local reassembly sites of the actin cortex during membrane blebbing. PMID:26976596

  13. A formin-nucleated actin aster concentrates cell wall hydrolases for cell fusion in fission yeast

    PubMed Central

    Dudin, Omaya; Bendezú, Felipe O.; Groux, Raphael; Laroche, Thierry; Seitz, Arne

    2015-01-01

    Cell–cell fusion is essential for fertilization. For fusion of walled cells, the cell wall must be degraded at a precise location but maintained in surrounding regions to protect against lysis. In fission yeast cells, the formin Fus1, which nucleates linear actin filaments, is essential for this process. In this paper, we show that this formin organizes a specific actin structure—the actin fusion focus. Structured illumination microscopy and live-cell imaging of Fus1, actin, and type V myosins revealed an aster of actin filaments whose barbed ends are focalized near the plasma membrane. Focalization requires Fus1 and type V myosins and happens asynchronously always in the M cell first. Type V myosins are essential for fusion and concentrate cell wall hydrolases, but not cell wall synthases, at the fusion focus. Thus, the fusion focus focalizes cell wall dissolution within a broader cell wall synthesis zone to shift from cell growth to cell fusion. PMID:25825517

  14. A Steric Antagonism of Actin Polymerization by a Salmonella Virulence Protein

    SciTech Connect

    Margarit,S.; Davidson, W.; Frego, L.; Stebbins, F.

    2006-01-01

    Salmonella spp. require the ADP-ribosyltransferase activity of the SpvB protein for intracellular growth and systemic virulence. SpvB covalently modifies actin, causing cytoskeletal disruption and apoptosis. We report here the crystal structure of the catalytic domain of SpvB, and we show by mass spectrometric analysis that SpvB modifies actin at Arg177, inhibiting its ATPase activity. We also describe two crystal structures of SpvB-modified, polymerization-deficient actin. These structures reveal that ADP-ribosylation does not lead to dramatic conformational changes in actin, suggesting a model in which this large family of toxins inhibits actin polymerization primarily through steric disruption of intrafilament contacts.

  15. Transforming growth factor-β-sphingosine kinase 1/S1P signaling upregulates microRNA-21 to promote fibrosis in renal tubular epithelial cells.

    PubMed

    Liu, Xiujuan; Hong, Quan; Wang, Zhen; Yu, Yanyan; Zou, Xin; Xu, Lihong

    2016-02-01

    Renal fibrosis is a progressive pathological change characterized by tubular cell apoptosis, tubulointerstitial fibroblast proliferation, and excessive deposition of extracellular matrix (ECM). miR-21 has been implicated in transforming growth factor-β (TGF-β)-stimulated tissue fibrosis. Recent studies showed that sphingosine kinase/sphingosine-1-phosphate (SphK/S1P) are also critical for TGF-β-stimulated tissue fibrosis; however, it is not clear whether SphK/S1P interacts with miR-21 or not. In this study, we hypothesized that SphK/S1P signaling is linked to upregulation of miR-21 by TGF-β. To verify this hypothesis, we first determined that miR-21 was highly expressed in renal tubular epithelial cells (TECs) stimulated with TGF-β by using qRT-PCR and Northern blotting. Simultaneously, inhibition of miR-21, mediated by the corresponding antimir, markedly decreased the expression and deposition of type I collagen, fibronectin (Fn), cysteine-rich protein 61 (CCN1), α-smooth muscle actin, and fibroblast-specific protein1 in TGF-β-treated TECs. ELISA and qRT-PCR were used to measure the S1P and SphK1 levels in TECs. S1P production was induced by TGF-β through activation of SphK1. Furthermore, it was observed that TGF-β-stimulated upregulation of miR-21 was abolished by SphK1 siRNA and was restored by the addition of exogenous S1P. Blocking S1PR2 also inhibited upregulation of miR-21. Additionally, miR-21 overexpression attenuated the repression of TGF-β-stimulated ECM deposition and epithelial-mesenchymal transition by SphK1 and S1PR2 siRNA. In summary, our study demonstrates a link between SphK1/S1P and TGF-β-induced miR-21 in renal TECs and may represent a novel therapeutic target in renal fibrosis. PMID:26376826

  16. Distinct roles for mammalian target of rapamycin complexes in the fibroblast response to transforming growth factor-beta.

    PubMed

    Rahimi, Rod A; Andrianifahanana, Mahefatiana; Wilkes, Mark C; Edens, Maryanne; Kottom, Theodore J; Blenis, John; Leof, Edward B

    2009-01-01

    Transforming growth factor-beta (TGF-beta) promotes a multitude of diverse biological processes, including growth arrest of epithelial cells and proliferation of fibroblasts. Although the TGF-beta signaling pathways that promote inhibition of epithelial cell growth are well characterized, less is known about the mechanisms mediating the positive response to this growth factor. Given that TGF-beta has been shown to promote fibrotic diseases and desmoplasia, identifying the fibroblast-specific TGF-beta signaling pathways is critical. Here, we investigate the role of mammalian target of rapamycin (mTOR), a known effector of phosphatidylinositol 3-kinase (PI3K) and promoter of cell growth, in the fibroblast response to TGF-beta. We show that TGF-beta activates mTOR complex 1 (mTORC1) in fibroblasts but not epithelial cells via a PI3K-Akt-TSC2-dependent pathway. Rapamycin, the pharmacologic inhibitor of mTOR, prevents TGF-beta-mediated anchorage-independent growth without affecting TGF-beta transcriptional responses or extracellular matrix protein induction. In addition to mTORC1, we also examined the role of mTORC2 in TGF-beta action. mTORC2 promotes TGF-beta-induced morphologic transformation and is required for TGF-beta-induced Akt S473 phosphorylation but not mTORC1 activation. Interestingly, both mTOR complexes are necessary for TGF-beta-mediated growth in soft agar. These results define distinct and overlapping roles for mTORC1 and mTORC2 in the fibroblast response to TGF-beta and suggest that inhibitors of mTOR signaling may be useful in treating fibrotic processes, such as desmoplasia. PMID:19117990

  17. Transforming growth factor-beta in the chicken fundal layers: an immunohistochemical study.

    PubMed

    Mathis, Ute; Schaeffel, Frank

    2010-06-01

    In the chicken model of myopia, it has first been shown that imposing defocus to the retina results in active remodelling of the sclera which, in turn, results in axial length changes of the eye. Transforming growth factor-beta (TGF-beta) is one of the scleral growth modulators but its cellular localization in the fundal layers, colocalization and function are not well known. The aim of the current study was to investigate the cellular distribution of the three isoforms TGF-beta1, 2 and 3 by immunohistochemical labelling. Furthermore, the effects of visual experience that induces refractive errors on TGF-beta2 labelling were examined. Transversal cryostat sections of the fundal layers were analyzed by indirect immunofluorescent labelling and cell counts. Visual experience was changed by having the chicks wear either diffusers, or positive or negative lenses of 7D power in front of the right eyes for various periods of time. Left eyes served as uncovered controls. All TGF-beta isoforms were localized in both scleral layers. In choroid, diffuse labelling of all isoforms was found. In retina, TGF-beta1 and 3 were detected in bipolar, amacrine and ganglion cells and TGF-beta2 in amacrine and ganglion cells. To further characterize these cells, double-labelling with known amacrine and bipolar cell markers was performed (calbindin, cellular retinoic acid binding protein (CRABP), Islet1, Lim3 and protein kinase C (PKC)). TGF-beta1, 2 and 3 could be colocalized with calbindin and CRABP in single amacrine cells. TGF-beta1-positive bipolar cells were immunoreactive to Lim3. TGF-beta1 and 3 were never colocalized with PKC in bipolar cells. Also, colocalization with peptides known to be involved in myopia development in chicks, such as glucagon, or vasointestinal polypeptide and the key enzyme for dopamine synthesis, tyrosine hydroxylase, was not observed. Lenses or diffusers, worn by the chicks for various periods of time, had no effect on TGF-beta2 immunoreactivity in

  18. Effects of transforming growth factor-beta on growth and differentiation of the continuous rat thyroid follicular cell line, FRTL-5

    SciTech Connect

    Morris, J.C. III; Ranganathan, G.; Hay, I.D.; Nelson, R.E.; Jiang, N.S.

    1988-09-01

    Transforming growth factor-beta (TGF beta) has been shown to influence the growth and differentiation of many widely varied cell types in vitro, including some that are endocrinologically active. We have investigated the previously unknown effects of this unique growth factor in the differentiated rat thyroid follicular cell line FRTL-5. The cells demonstrated specific, high affinity binding of TGF beta, and as with other epithelial cells, the growth of these thyroid follicular cells was potently inhibited by addition of TGF beta to the culture medium. TGF beta caused a significant reduction in TSH-sensitive adenylate cyclase activity in the cells. The addition of (Bu)2cAMP along with the growth factor to cultures partially reversed the characteristic morphological changes seen with TGF beta, but did not reverse the growth inhibition. To further investigate the possible mechanisms of the effects of TGF beta on the cells, we measured the influence of the growth factor on (125I)TSH binding. TGF beta did not compete for specific TSH-binding sites; however, exposure of the cells to TGF beta for 12 or more h resulted in a dose-dependent down-regulation of TSH receptors that was fully reversible. While cellular proliferation was potently inhibited by TGF beta, differentiated function, as manifest by iodine-trapping ability, was stimulated by the growth factor. This stimulation of iodine uptake was independent of, and additive to, the stimulatory effects of TSH. Finally, FRTL-5 cells in serum-free medium and in response to TSH were shown to secrete TGF beta-like activity that competed for (125I)TGF beta in a RRA. These studies suggest that TGF beta may represent an autocrine mechanism of controlling the growth response to TSH in thyroid follicular cells, while allowing the continuance of differentiated function.

  19. Effect of intestinal ischemia-reperfusion on expressions of endogenous basic fibroblast growth factor and transforming growth factor betain lung and its relation with lung repair.

    PubMed

    Fu, Xiao-Bing; Yang, Yin-Hui; Sun, Tong-Zhu; Gu, Xiao-Man; Jiang, Li-Xian; Sun, Xiao-Qing; Sheng, Zhi-Yong

    2000-06-01

    AIM:To study the changes of endogenous transforming growth factor beta(TGFbeta) and basic fibroblast growth factor (bFGF) in lung following intestinal ischemia and reperfusion injury and their effects on lung injury and repair.METHODS:Sixty Wistar rats were divided into five groups, which underwent sham-operation, ischemia (45 minutes), and reperfusion (6, 24 and 48 hours, respectively) after ischemia (45 minutes). Immunohistochemical method was used to observe the localization and amounts of both growth factors.RESULTS:Positive signals of both growth factors could be found in normal lung, mainly in alveolar cells and endothelial cells of vein. After ischemia and reperfusion insult, expressions of both growth factors were increased and their amounts at 6 hours were larger than those of normal control or of 24 and 48 hours after insult.CONCLUSION:The endogenous bFGF and TGF beta expression appears to be upregulated in the lung following intestinal ischemia and reperfusion, suggesting that both growth factors may be involved in the process of lung injury and repair. PMID:11819596

  20. Mechanism of Actin-Based Motility

    NASA Astrophysics Data System (ADS)

    Pantaloni, Dominique; Le Clainche, Christophe; Carlier, Marie-France

    2001-05-01

    Spatially controlled polymerization of actin is at the origin of cell motility and is responsible for the formation of cellular protrusions like lamellipodia. The pathogens Listeria monocytogenes and Shigella flexneri, which undergo actin-based propulsion, are acknowledged models of the leading edge of lamellipodia. Actin-based motility of the bacteria or of functionalized microspheres can be reconstituted in vitro from only five pure proteins. Movement results from the regulated site-directed treadmilling of actin filaments, consistent with observations of actin dynamics in living motile cells and with the biochemical properties of the components of the synthetic motility medium.

  1. Microtubule and Actin Interplay Drive Intracellular c-Src Trafficking.

    PubMed

    Arnette, Christopher; Frye, Keyada; Kaverina, Irina

    2016-01-01

    The proto-oncogene c-Src is involved in a variety of signaling processes. Therefore, c-Src spatiotemporal localization is critical for interaction with downstream targets. However, the mechanisms regulating this localization have remained elusive. Previous studies have shown that c-Src trafficking is a microtubule-dependent process that facilitates c-Src turnover in neuronal growth cones. As such, microtubule depolymerization lead to the inhibition of c-Src recycling. Alternatively, c-Src trafficking was also shown to be regulated by RhoB-dependent actin polymerization. Our results show that c-Src vesicles primarily exhibit microtubule-dependent trafficking; however, microtubule depolymerization does not inhibit vesicle movement. Instead, vesicular movement becomes both faster and less directional. This movement was associated with actin polymerization directly at c-Src vesicle membranes. Interestingly, it has been shown previously that c-Src delivery is an actin polymerization-dependent process that relies on small GTPase RhoB at c-Src vesicles. In agreement with this finding, microtubule depolymerization induced significant activation of RhoB, together with actin comet tail formation. These effects occurred downstream of GTP-exchange factor, GEF-H1, which was released from depolymerizing MTs. Accordingly, GEF-H1 activity was necessary for actin comet tail formation at the Src vesicles. Our results indicate that regulation of c-Src trafficking requires both microtubules and actin polymerization, and that GEF-H1 coordinates c-Src trafficking, acting as a molecular switch between these two mechanisms. PMID:26866809

  2. Microtubule and Actin Interplay Drive Intracellular c-Src Trafficking

    PubMed Central

    Arnette, Christopher; Frye, Keyada; Kaverina, Irina

    2016-01-01

    The proto-oncogene c-Src is involved in a variety of signaling processes. Therefore, c-Src spatiotemporal localization is critical for interaction with downstream targets. However, the mechanisms regulating this localization have remained elusive. Previous studies have shown that c-Src trafficking is a microtubule-dependent process that facilitates c-Src turnover in neuronal growth cones. As such, microtubule depolymerization lead to the inhibition of c-Src recycling. Alternatively, c-Src trafficking was also shown to be regulated by RhoB-dependent actin polymerization. Our results show that c-Src vesicles primarily exhibit microtubule-dependent trafficking; however, microtubule depolymerization does not inhibit vesicle movement. Instead, vesicular movement becomes both faster and less directional. This movement was associated with actin polymerization directly at c-Src vesicle membranes. Interestingly, it has been shown previously that c-Src delivery is an actin polymerization-dependent process that relies on small GTPase RhoB at c-Src vesicles. In agreement with this finding, microtubule depolymerization induced significant activation of RhoB, together with actin comet tail formation. These effects occurred downstream of GTP-exchange factor, GEF-H1, which was released from depolymerizing MTs. Accordingly, GEF-H1 activity was necessary for actin comet tail formation at the Src vesicles. Our results indicate that regulation of c-Src trafficking requires both microtubules and actin polymerization, and that GEF-H1 coordinates c-Src trafficking, acting as a molecular switch between these two mechanisms. PMID:26866809

  3. Cloning the promoter for transforming growth factor-beta type III receptor. Basal and conditional expression in fetal rat osteoblasts

    NASA Technical Reports Server (NTRS)

    Ji, C.; Chen, Y.; McCarthy, T. L.; Centrella, M.

    1999-01-01

    Transforming growth factor-beta binds to three high affinity cell surface molecules that directly or indirectly regulate its biological effects. The type III receptor (TRIII) is a proteoglycan that lacks significant intracellular signaling or enzymatic motifs but may facilitate transforming growth factor-beta binding to other receptors, stabilize multimeric receptor complexes, or segregate growth factor from activating receptors. Because various agents or events that regulate osteoblast function rapidly modulate TRIII expression, we cloned the 5' region of the rat TRIII gene to assess possible control elements. DNA fragments from this region directed high reporter gene expression in osteoblasts. Sequencing showed no consensus TATA or CCAAT boxes, whereas several nuclear factors binding sequences within the 3' region of the promoter co-mapped with multiple transcription initiation sites, DNase I footprints, gel mobility shift analysis, or loss of activity by deletion or mutation. An upstream enhancer was evident 5' proximal to nucleotide -979, and a silencer region occurred between nucleotides -2014 and -2194. Glucocorticoid sensitivity mapped between nucleotides -687 and -253, whereas bone morphogenetic protein 2 sensitivity co-mapped within the silencer region. Thus, the TRIII promoter contains cooperative basal elements and dispersed growth factor- and hormone-sensitive regulatory regions that can control TRIII expression by osteoblasts.

  4. Structural Variability of Tropospheric Growth Factors Transforming Mid-latitude Cyclones to Severe Storms over the North Atlantic

    NASA Astrophysics Data System (ADS)

    Wild, Simon; Befort, Daniel J.; Leckebusch, Gregor C.

    2015-04-01

    The development of European surface wind storms out of normal mid-latitude cyclones is substantially influenced by upstream tropospheric growth factors over the Northern Atlantic. The main factors include divergence and vorticity advection in the upper troposphere, latent heat release and the presence of instabilities of short baroclinic waves of suitable wave lengths. In this study we examine a subset of these potential growth factors and their related influences on the transformation of extra-tropical cyclones into severe damage prone surface storm systems. Previous studies have shown links between specific growth factors and surface wind storms related to extreme cyclones. In our study we investigate in further detail spatial and temporal variability patterns of these upstream processes at different vertical levels of the troposphere. The analyses will comprise of the three growth factors baroclinicity, latent heat release and upper tropospheric divergence. Our definition of surface wind storms is based on the Storm Severity Index (SSI) alongside a wind tracking algorithm identifying areas of exceedances of the local 98th percentile of the 10m wind speed. We also make use of a well-established extra-tropical cyclone identification and tracking algorithm. These cyclone tracks form the base for a composite analysis of the aforementioned growth factors using ERA-Interim Reanalysis from 1979 - 2014 for the extended winter season (ONDJFM). Our composite analysis corroborates previous similar studies but extends them by using an impact based algorithm for the identification of strong wind systems. Based on this composite analysis we further identify variability patterns for each growth factor most important for the transformation of a cyclone into a surface wind storm. We thus also address the question whether the link between storm intensity and related growth factor anomaly taking into account its spatial variability is stable and can be quantified. While the

  5. Quantifying actin wave modulation on periodic topography

    NASA Astrophysics Data System (ADS)

    Guven, Can; Driscoll, Meghan; Sun, Xiaoyu; Parker, Joshua; Fourkas, John; Carlsson, Anders; Losert, Wolfgang

    2014-03-01

    Actin is the essential builder of the cell cytoskeleton, whose dynamics are responsible for generating the necessary forces for the formation of protrusions. By exposing amoeboid cells to periodic topographical cues, we show that actin can be directionally guided via inducing preferential polymerization waves. To quantify the dynamics of these actin waves and their interaction with the substrate, we modify a technique from computer vision called ``optical flow.'' We obtain vectors that represent the apparent actin flow and cluster these vectors to obtain patches of newly polymerized actin, which represent actin waves. Using this technique, we compare experimental results, including speed distribution of waves and distance from the wave centroid to the closest ridge, with actin polymerization simulations. We hypothesize the modulation of the activity of nucleation promotion factors on ridges (elevated regions of the surface) as a potential mechanism for the wave-substrate coupling. Funded by NIH grant R01GM085574.

  6. Allyl Isothiocyanate Inhibits Actin-Dependent Intracellular Transport in Arabidopsis thaliana.

    PubMed

    Sporsheim, Bjørnar; Øverby, Anders; Bones, Atle Magnar

    2015-01-01

    Volatile allyl isothiocyanate (AITC) derives from the biodegradation of the glucosinolate sinigrin and has been associated with growth inhibition in several plants, including the model plant Arabidopsis thaliana. However, the underlying cellular mechanisms of this feature remain scarcely investigated in plants. In this study, we present evidence of an AITC-induced inhibition of actin-dependent intracellular transport in A. thaliana. A transgenic line of A. thaliana expressing yellow fluorescent protein (YFP)-tagged actin filaments was used to show attenuation of actin filament movement by AITC. This appeared gradually in a time- and dose-dependent manner and resulted in actin filaments appearing close to static. Further, we employed four transgenic lines with YFP-fusion proteins labeling the Golgi apparatus, endoplasmic reticulum (ER), vacuoles and peroxisomes to demonstrate an AITC-induced inhibition of actin-dependent intracellular transport of or, in these structures, consistent with the decline in actin filament movement. Furthermore, the morphologies of actin filaments, ER and vacuoles appeared aberrant following AITC-exposure. However, AITC-treated seedlings of all transgenic lines tested displayed morphologies and intracellular movements similar to that of the corresponding untreated and control-treated plants, following overnight incubation in an AITC-absent environment, indicating that AITC-induced decline in actin-related movements is a reversible process. These findings provide novel insights into the cellular events in plant cells following exposure to AITC, which may further expose clues to the physiological significance of the glucosinolate-myrosinase system. PMID:26690132

  7. Plastins regulate ectoplasmic specialization via its actin bundling activity on microfilaments in the rat testis.

    PubMed

    Li, Nan; Wong, Chris Kc; Cheng, C Yan

    2016-01-01

    Plastins are a family of actin binding proteins (ABPs) known to cross-link actin microfilaments in mammalian cells, creating actin microfilament bundles necessary to confer cell polarity and cell shape. Plastins also support cell movement in response to changes in environment, involved in cell/tissue growth and development. They also confer plasticity to cells and tissues in response to infection or other pathological conditions (e.g., inflammation). In the testis, the cell-cell anchoring junction unique to the testis that is found at the Sertoli cell-cell interface at the blood-testis barrier (BTB) and at the Sertoli-spermatid (e.g., 8-19 spermatids in the rat testis) is the basal and the apical ectoplasmic specialization (ES), respectively. The ES is an F-actin-rich anchoring junction constituted most notably by actin microfilament bundles. A recent report using RNAi that specifically knocks down plastin 3 has yielded some insightful information regarding the mechanism by which plastin 3 regulates the status of actin microfilament bundles at the ES via its intrinsic actin filament bundling activity. Herein, we provide a brief review on the role of plastins in the testis in light of this report, which together with recent findings in the field, we propose a likely model by which plastins regulate ES function during the epithelial cycle of spermatogenesis via their intrinsic activity on actin microfilament organization in the rat testis. PMID:26608945

  8. α-Synuclein and Its A30P Mutant Affect Actin Cytoskeletal Structure and Dynamics

    PubMed Central

    Sousa, Vítor L.; Bellani, Serena; Giannandrea, Maila; Yousuf, Malikmohamed; Valtorta, Flavia; Meldolesi, Jacopo

    2009-01-01

    The function of α-synuclein, a soluble protein abundant in the brain and concentrated at presynaptic terminals, is still undefined. Yet, α-synuclein overexpression and the expression of its A30P mutant are associated with familial Parkinson's disease. Working in cell-free conditions, in two cell lines as well as in primary neurons we demonstrate that α-synuclein and its A30P mutant have different effects on actin polymerization. Wild-type α-synuclein binds actin, slows down its polymerization and accelerates its depolymerization, probably by monomer sequestration; A30P mutant α-synuclein increases the rate of actin polymerization and disrupts the cytoskeleton during reassembly of actin filaments. Consequently, in cells expressing mutant α-synuclein, cytoskeleton-dependent processes, such as cell migration, are inhibited, while exo- and endocytic traffic is altered. In hippocampal neurons from mice carrying a deletion of the α-synuclein gene, electroporation of wild-type α-synuclein increases actin instability during remodeling, with growth of lamellipodia-like structures and apparent cell enlargement, whereas A30P α-synuclein induces discrete actin-rich foci during cytoskeleton reassembly. In conclusion, α-synuclein appears to play a major role in actin cytoskeletal dynamics and various aspects of microfilament function. Actin cytoskeletal disruption induced by the A30P mutant might alter various cellular processes and thereby play a role in the pathogenesis of neurodegeneration. PMID:19553474

  9. Allyl Isothiocyanate Inhibits Actin-Dependent Intracellular Transport in Arabidopsis thaliana

    PubMed Central

    Sporsheim, Bjørnar; Øverby, Anders; Bones, Atle Magnar

    2015-01-01

    Volatile allyl isothiocyanate (AITC) derives from the biodegradation of the glucosinolate sinigrin and has been associated with growth inhibition in several plants, including the model plant Arabidopsis thaliana. However, the underlying cellular mechanisms of this feature remain scarcely investigated in plants. In this study, we present evidence of an AITC-induced inhibition of actin-dependent intracellular transport in A. thaliana. A transgenic line of A. thaliana expressing yellow fluorescent protein (YFP)-tagged actin filaments was used to show attenuation of actin filament movement by AITC. This appeared gradually in a time- and dose-dependent manner and resulted in actin filaments appearing close to static. Further, we employed four transgenic lines with YFP-fusion proteins labeling the Golgi apparatus, endoplasmic reticulum (ER), vacuoles and peroxisomes to demonstrate an AITC-induced inhibition of actin-dependent intracellular transport of or, in these structures, consistent with the decline in actin filament movement. Furthermore, the morphologies of actin filaments, ER and vacuoles appeared aberrant following AITC-exposure. However, AITC-treated seedlings of all transgenic lines tested displayed morphologies and intracellular movements similar to that of the corresponding untreated and control-treated plants, following overnight incubation in an AITC-absent environment, indicating that AITC-induced decline in actin-related movements is a reversible process. These findings provide novel insights into the cellular events in plant cells following exposure to AITC, which may further expose clues to the physiological significance of the glucosinolate-myrosinase system. PMID:26690132

  10. Plastins regulate ectoplasmic specialization via its actin bundling activity on microfilaments in the rat testis

    PubMed Central

    Li, Nan; Wong, Chris KC; Cheng, C Yan

    2016-01-01

    Plastins are a family of actin binding proteins (ABPs) known to cross-link actin microfilaments in mammalian cells, creating actin microfilament bundles necessary to confer cell polarity and cell shape. Plastins also support cell movement in response to changes in environment, involved in cell/tissue growth and development. They also confer plasticity to cells and tissues in response to infection or other pathological conditions (e.g., inflammation). In the testis, the cell-cell anchoring junction unique to the testis that is found at the Sertoli cell-cell interface at the blood-testis barrier (BTB) and at the Sertoli-spermatid (e.g., 8–19 spermatids in the rat testis) is the basal and the apical ectoplasmic specialization (ES), respectively. The ES is an F-actin-rich anchoring junction constituted most notably by actin microfilament bundles. A recent report using RNAi that specifically knocks down plastin 3 has yielded some insightful information regarding the mechanism by which plastin 3 regulates the status of actin microfilament bundles at the ES via its intrinsic actin filament bundling activity. Herein, we provide a brief review on the role of plastins in the testis in light of this report, which together with recent findings in the field, we propose a likely model by which plastins regulate ES function during the epithelial cycle of spermatogenesis via their intrinsic activity on actin microfilament organization in the rat testis. PMID:26608945

  11. Size distribution of linear and helical polymers in actin solution analyzed by photon counting histogram.

    PubMed

    Terada, Naofumi; Shimozawa, Togo; Ishiwata, Shin'ichi; Funatsu, Takashi

    2007-03-15

    Actin is a ubiquitous protein that is a major component of the cytoskeleton, playing an important role in muscle contraction and cell motility. At steady state, actin monomers and filaments (F-actin) coexist, and actin subunits continuously attach and detach at the filament ends. However, the size distribution of actin oligomers in F-actin solution has never been clarified. In this study, we investigated the size distribution of actin oligomers using photon-counting histograms. For this purpose, actin was labeled with a fluorescent dye, and the emitted photons were detected by confocal optics (the detection volume was of femtoliter (fL) order). Photon-counting histograms were analyzed to obtain the number distribution of actin oligomers in the detection area from their brightness, assuming that the brightness of an oligomer was proportional to the number of protomers. We found that the major populations at physiological ionic strength were 1-5mers. For data analysis, we successfully applied the theory of linear and helical aggregations of macromolecules. The model postulates three states of actin, i.e., monomers, linear polymers, and helical polymers. Here we obtained three parameters: the equilibrium constants for polymerization of linear polymers, K(l)=(5.2 +/- 1.1) x 10(6) M(-1), and helical polymers, K(h)=(1.6 +/- 0.5) x 10(7) M(-1); and the ratio of helical to linear trimers, gamma = (3.6 +/- 2.3) x 10(-2). The excess free energy of transforming a linear trimer to a helical trimer, which is assumed to be a nucleus for helical polymers, was calculated to be 2.0 kcal/mol. These analyses demonstrate that the oligomeric phase at steady state is predominantly composed of linear 1-5mers, and the transition from linear to helical polymers occurs on the level of 5-7mers. PMID:17172301

  12. Transforming growth factor alpha may be a physiological regulator of liver regeneration by means of an autocrine mechanism.

    PubMed Central

    Mead, J E; Fausto, N

    1989-01-01

    We investigated whether transforming growth factor alpha (TGF-alpha) is involved in hepatocyte growth responses both in vivo and in culture. During liver regeneration after partial hepatectomy in rats, TGF-alpha mRNA increased; it reached a maximum (approximately 9-fold higher than normal) at the peak of DNA synthesis. The message and the peptide were localized in hepatocytes and found in higher amounts in hepatocytes obtained from regenerating liver. TGF-alpha caused a 13-fold elevation of DNA synthesis in hepatocytes in primary culture and was slightly more effective than epidermal growth factor. TGF-beta blocked TGF-alpha stimulation when added either simultaneously with TGF-alpha or a day later. TGF-alpha message increased in hepatocytes stimulated to undergo DNA synthesis by TGF-alpha or epidermal growth factor, and the peptide was detected in the culture medium by RIA. In the regenerating liver, the increase in TGF-alpha mRNA during the first day after partial hepatectomy coincided with an increase in epidermal growth factor/TGF-alpha receptor mRNA and a decrease (already reported) in the number of these receptors. We conclude that TGF-alpha may function as a physiological inducer of hepatocyte DNA synthesis during liver regeneration by means of an autocrine mechanism and that its stimulatory effects in this growth process are balanced by the inhibitory action of TGF-beta 1. Images PMID:2922399

  13. Stimulation of glycolysis and amino acid uptake in NRK-49F cells by transforming growth factor beta and epidermal growth factor.

    PubMed Central

    Boerner, P; Resnick, R J; Racker, E

    1985-01-01

    Glycolysis in normal resting rat kidney cells (NRK-49F) was stimulated by a 2-hr exposure to transforming growth factors prior to assay. Transforming growth factor beta (TGF-beta) was effective when added alone, and further addition of epidermal growth factor (EGF) had little effect. The stimulation by TGF-beta was abolished when cycloheximide was present during the incubation, suggesting that protein synthesis is required for the effect. Incubation of the cells with 25 mM methionine abolished the stimulation of glycolysis by TGF-beta. The uptake of methylaminoisobutyrate via system A was stimulated by either TGF-beta or EGF. The greater than 3-fold stimulation of uptake by 1 ng of pure TGF-beta per ml was usually somewhat enhanced on addition of 0.5 ng of EGF per ml. Moreover, an antiserum against EGF receptor partially depressed the response to TGF-beta, suggesting some overlapping interactions of EGF and TGF-beta. Images PMID:3871948

  14. Reactive oxygen species (ROS)-induced actin glutathionylation controls actin dynamics in neutrophils

    PubMed Central

    Sakai, Jiro; Li, Jingyu; Subramanian, Kulandayan K.; Mondal, Subhanjan; Bajrami, Besnik; Hattori, Hidenori; Jia, Yonghui; Dickinson, Bryan C.; Zhong, Jia; Ye, Keqiang; Chang, Christopher J; Ho, Ye-Shih; Zhou, Jun; Luo, Hongbo R.

    2012-01-01

    Summary The regulation of actin dynamics is pivotal for cellular processes such as cell adhesion, migration, and phagocytosis, and thus is crucial for neutrophils to fulfill their roles in innate immunity. Many factors have been implicated in signal-induced actin polymerization, however the essential nature of the potential negative modulators are still poorly understood. Here we report that NADPH oxidase-dependent physiologically generated reactive oxygen species (ROS) negatively regulate actin polymerization in stimulated neutrophils via driving reversible actin glutathionylation. Disruption of glutaredoxin 1 (Grx1), an enzyme that catalyzes actin deglutathionylation, increased actin glutathionylation, attenuated actin polymerization, and consequently impaired neutrophil polarization, chemotaxis, adhesion, and phagocytosis. Consistently, Grx1-deficient murine neutrophils showed impaired in vivo recruitment to sites of inflammation and reduced bactericidal capability. Together, these results present a physiological role for glutaredoxin and ROS- induced reversible actin glutathionylation in regulation of actin dynamics in neutrophils. PMID:23159440

  15. Platelet-derived growth factor and transforming growth factor-beta in invertebrate immune and neuroendocrine interactions: another sign of conservation in evolution.

    PubMed

    Ottaviani, E; Franchini, A; Kletsas, D

    2001-08-01

    Growth factor-like molecules have been found in various invertebrate species. In particular, we have reported the presence of platelet-derived growth factor (PDGF)-AB and transforming growth factor-beta (TGF-beta)1 immunoreactive molecules in molluscs, insects and annelids. Moreover, PDGF-AB and TGF-beta1 affect the main immune functions, such as phagocytosis, chemotaxis and cell motility. Changes in cell shape are induced via interactions of growth factors with their respective specific receptors. The extracellular signals are transduced by the activation of classical signal transduction pathways, such as those involving PKA and PKC, and pivotal transcription regulators, i.e. the Fos, Jun and SMAD proteins. The two growth factors intervene in stress responses by activating the CRH-ACTH-biogenic amine axis. Exogenous administration of PDGF-AB and TGF-beta1 in a molluscan wound provokes an accelerated migration of immunocytes and fibroblasts to the injured area, stimulating granulation tissue formation and wound re-epithelialization. These findings suggest that these molecules are ancestral and that their function is well conserved and crucial in the maintenance of invertebrate homeostasis. PMID:11489427

  16. Radial expansion rates and tumor growth kinetics predict malignant transformation in contrast-enhancing low-grade diffuse astrocytoma

    PubMed Central

    Hathout, Leith; Pope, Whitney B; Lai, Albert; Nghiemphu, Phioanh L; Cloughesy, Timothy F; Ellingson, Benjamin M

    2015-01-01

    Summary Background Contrast-enhancing low-grade diffuse astrocytomas are an understudied, aggressive subtype at increased risk because of few radiographic indications of malignant transformation. In the current study, we tested whether tumor growth kinetics could identify tumors that undergo malignant transformation to higher grades. Methods Thirty patients with untreated diffuse astrocytomas (WHO II) that underwent tumor progression were enrolled. Contrast-enhancing and T2 hyperintense tumor regions were segmented and the radius of tumor at two time points leading to progression was estimated. Radial expansion rates were used to estimate proliferation and invasion rates using a biomathematical model. Results Radial expansion rates for both contrast-enhancing (p = 0.0040) and T2 hyperintense regions (p = 0.0016) were significantly higher in WHO II–IV tumors compared with nontransformers. Similarly, model estimates showed a significantly higher proliferation (p = 0.0324) and invasion rate (p = 0.0050) in WHO II–IV tumors compared with nontransformers. Conclusion Tumor growth kinetics can identify contrast-enhancing diffuse astrocytomas undergoing malignant transformation. PMID:26095141

  17. Profilin Interaction with Actin Filament Barbed End Controls Dynamic Instability, Capping, Branching, and Motility

    PubMed Central

    Pernier, Julien; Shekhar, Shashank; Jegou, Antoine; Guichard, Bérengère; Carlier, Marie-France

    2016-01-01

    Summary Cell motility and actin homeostasis depend on the control of polarized growth of actin filaments. Profilin, an abundant regulator of actin dynamics, supports filament assembly at barbed ends by binding G-actin. Here, we demonstrate how, by binding and destabilizing filament barbed ends at physiological concentrations, profilin also controls motility, cell migration, and actin homeostasis. Profilin enhances filament length fluctuations. Profilin competes with Capping Protein at barbed ends, which generates a lower amount of profilin-actin than expected if barbed ends were tightly capped. Profilin competes with barbed end polymerases, such as formins and VopF, and inhibits filament branching by WASP-Arp2/3 complex by competition for filament barbed ends, accounting for its as-yet-unknown effects on motility and metastatic cell migration observed in this concentration range. In conclusion, profilin is a major coordinator of polarized growth of actin filaments, controlled by competition between barbed end cappers, trackers, destabilizers, and filament branching machineries. PMID:26812019

  18. Protopanaxatirol type ginsenoside Re promotes cyclic growth of hair follicles via inhibiting transforming growth factor β signaling cascades.

    PubMed

    Li, Zheng; Ryu, Seung-Wook; Lee, Jungsul; Choi, Kyungsun; Kim, Sunchang; Choi, Chulhee

    2016-02-19

    Ginsenosides, the major bio-active ingredients included in Panax ginseng, have been known for the hair growth activity and used to treat patients who suffer from hair loss; however, the detailed mechanisms of this action are still largely unknown. This study was conducted to investigate the molecular and cellular mechanisms responsible for hair growth promoting effect of ginsenoside Re (GRe) in vitro and in vivo. Different doses of minoxidil and GRe were administered topically to the back regions of nude mice for up to 45 days, and hair shaft length and hair cycles were determined for hair promoting activities. Topical treatment of GRe significantly increased the hair shaft length and hair existent time, which was comparable to the action of minoxidil. We also demonstrated that GRe stimulated hair shaft elongation in the ex vivo cultures of vibrissa hair follicles isolated from C57BL/6 mouse. Systemic transcriptome analysis by next generation sequencing demonstrated that TGF-β-pathway related genes were selectively down-regulated by treatment of GRe in vivo, and the same treatment suppressed TGF-β-induced phosphorylation of ERK in HeLa cells. The results clearly indicated that GRe is the effective constituent in the ginseng on hair promotion via selective inhibition of the hair growth phase transition related signaling pathways, TGF-β signaling cascades. PMID:26820528

  19. Althaea rosea Cavanil and Plantago major L. suppress neoplastic cell transformation through the inhibition of epidermal growth factor receptor kinase.

    PubMed

    Choi, Eun-Sun; Cho, Sung-Dae; Shin, Ji-Ae; Kwon, Ki Han; Cho, Nam-Pyo; Shim, Jung-Hyun

    2012-10-01

    For thousands of years in Asia, Althaea rosea Cavanil (ARC) and Plantago major L. (PML) have been used as powerful non-toxic therapeutic agents that inhibit inflammation. However, the anticancer mechanisms and molecular targets of ARC and PML are poorly understood, particularly in epidermal growth factor (EGF)-induced neoplastic cell transformation. The aim of this study was to evaluate the chemopreventive effects and mechanisms of the methanol extracts from ARC (MARC) and PML (MPML) in EGF-induced neoplastic cell transformation of JB6 P+ mouse epidermal cells using an MTS assay, anchorage-independent cell transformation assay and western blotting. Our results showed that MARC and MPML significantly suppressed neoplastic cell transformation by inhibiting the kinase activity of the EGF receptor (EGFR). The activation of EGFR by EGF was suppressed by MARC and MPML treatment in EGFR(+/+) cells, but not in EGFR(-/-) cells. In addition, MARC and MPML inhibited EGF-induced cell proliferation in EGFR-expressing murine embryonic fibroblasts (EGFR(+/+)). These results strongly indicate that EGFR targeting by MARC and MPML may be a good strategy for chemopreventive or chemotherapeutic applications. PMID:22767187

  20. Assessing spiritual growth and spiritual decline following a diagnosis of cancer: reliability and validity of the spiritual transformation scale.

    PubMed

    Cole, Brenda S; Hopkins, Clare M; Tisak, John; Steel, Jennifer L; Carr, Brian I

    2008-02-01

    This study assessed the factor structure, reliability, and validity of an instrument designed to assess spiritual transformations following a diagnosis of cancer-the Spiritual Transformation Scale (STS). The instrument was administering to 253 people diagnosed with cancer within the previous 2 years. Two underlying factors emerged (spiritual growth (SG) and spiritual decline (SD)) with adequate internal reliability (alpha = 0.98 and 0.86, respectively) and test-retest reliability (r = 0.85 and 0.73, respectively). Validity was supported by correlations between SG and the Positive and Negative Affect Scale (PANAS) Positive Affect Subscale (r = 0.23, p < 0.001), the Daily Spiritual Experiences Scale (r = 0.57, p < 0.001), and the Post-traumatic Growth Inventory (r = 0.68, p < 0.001). SD was associated with higher scores on the Center for Epidemiological Studies Depression scale (r = 0.38, p < 0.001) and PANAS-Negative Affect Subscale (r = 0.40, p < 0.001), and lower scores on the PANAS-Positive Affect Subscale (r = -0.23, p < 0.001), and the Daily Spiritual Experiences Scale (r = -0.30, p < 0.001). Hierarchical regression analyses indicated that the subscales uniquely predicted adjustment beyond related constructs (intrinsic religiousness, spiritual coping, and general post-traumatic growth). The results indicate that the STS is psychometrically sound, with SG predicting better, and SD predicting poorer, mental and spiritual well-being following a diagnosis of cancer. PMID:17458862

  1. Apurinic/apyrimidinic endonuclease 1 regulates angiogenesis in a transforming growth factor β-dependent manner in human osteosarcoma

    PubMed Central

    Jiang, Xuan; Shan, Jinlu; Dai, Nan; Zhong, Zhaoyang; Qing, Yi; Yang, Yuxing; Zhang, Shiheng; Li, Chongyi; Sui, Jiangdong; Ren, Tao; Li, Mengxia; Wang, Dong

    2015-01-01

    Angiogenesis plays an important role in tumor growth and metastasis and has been reported to be inversely correlated with overall survival of osteosarcoma patients. It has been shown that apurinic/apyrimidinic endonuclease 1 (APE1), a dually functional protein possessing both base excision repair and redox activities, is involved in tumor angiogenesis, although these mechanisms are not fully understood. Our previous study showed that the expression of transforming growth factor β (TGFβ) was significantly reduced in APE1-deficient osteosarcoma cells. Transforming growth factor β promotes cancer metastasis through various mechanisms including immunosuppression, angiogenesis, and invasion. In the current study, we initially revealed that APE1, TGFβ, and microvessel density (MVD) have pairwise correlation in osteosarcoma tissue samples, whereas TGFβ, tumor size, and MVD were inversely related to the prognosis of the cohort. We found that knocking down APE1 in osteosarcoma cells resulted in TGFβ downregulation. In addition, APE1-siRNA led to suppression of angiogenesis in vitro based on HUVECs in Transwell and Matrigel tube formation assays. Reduced secretory protein level of TGFβ of culture medium also resulted in decreased phosphorylation of Smad3 of HUVECs. In a mouse xenograft model, siRNA-mediated silencing of APE1 downregulated TGFβ expression, tumor size, and MVD. Collectively, the current evidence indicates that APE1 regulates angiogenesis in osteosarcoma by controlling the TGFβ pathway, suggesting a novel target for anti-angiogenesis therapy in human osteosarcoma. PMID:26250694

  2. The Design of MACs (Minimal Actin Cortices)

    PubMed Central

    Vogel, Sven K; Heinemann, Fabian; Chwastek, Grzegorz; Schwille, Petra

    2013-01-01

    The actin cell cortex in eukaryotic cells is a key player in controlling and maintaining the shape of cells, and in driving major shape changes such as in cytokinesis. It is thereby constantly being remodeled. Cell shape changes require forces acting on membranes that are generated by the interplay of membrane coupled actin filaments and assemblies of myosin motors. Little is known about how their interaction regulates actin cell cortex remodeling and cell shape changes. Because of the vital importance of actin, myosin motors and the cell membrane, selective in vivo experiments and manipulations are often difficult to perform or not feasible. Thus, the intelligent design of minimal in vitro systems for actin-myosin-membrane interactions could pave a way for investigating actin cell cortex mechanics in a detailed and quantitative manner. Here, we present and discuss the design of several bottom-up in vitro systems accomplishing the coupling of actin filaments to artificial membranes, where key parameters such as actin densities and membrane properties can be varied in a controlled manner. Insights gained from these in vitro systems may help to uncover fundamental principles of how exactly actin-myosin-membrane interactions govern actin cortex remodeling and membrane properties for cell shape changes. © 2013 Wiley Periodicals, Inc. PMID:24039068

  3. Mesoscopic model of actin-based propulsion.

    PubMed

    Zhu, Jie; Mogilner, Alex

    2012-01-01

    Two theoretical models dominate current understanding of actin-based propulsion: microscopic polymerization ratchet model predicts that growing and writhing actin filaments generate forces and movements, while macroscopic elastic propulsion model suggests that deformation and stress of growing actin gel are responsible for the propulsion. We examine both experimentally and computationally the 2D movement of ellipsoidal beads propelled by actin tails and show that neither of the two models can explain the observed bistability of the orientation of the beads. To explain the data, we develop a 2D hybrid mesoscopic model by reconciling these two models such that individual actin filaments undergoing nucleation, elongation, attachment, detachment and capping are embedded into the boundary of a node-spring viscoelastic network representing the macroscopic actin gel. Stochastic simulations of this 'in silico' actin network show that the combined effects of the macroscopic elastic deformation and microscopic ratchets can explain the observed bistable orientation of the actin-propelled ellipsoidal beads. To test the theory further, we analyze observed distribution of the curvatures of the trajectories and show that the hybrid model's predictions fit the data. Finally, we demonstrate that the model can explain both concave-up and concave-down force-velocity relations for growing actin networks depending on the characteristic time scale and network recoil. To summarize, we propose that both microscopic polymerization ratchets and macroscopic stresses of the deformable actin network are responsible for the force and movement generation. PMID:23133366

  4. Affinity chromatography of immobilized actin and myosin.

    PubMed Central

    Bottomley, R C; Trayer, I P

    1975-01-01

    Actin