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Brenda Russell - One of the best experts on this subject based on the ideXlab platform.

  • CapZ actua como integrador de senalamiento intracelular en respuesta a estimulos mecanicos en el miofilamento cardiaco
    Memorias del I Congreso Internacional de Ciencias Exactas y Naturales, 2019
    Co-Authors: Christopher Solis Ocampo, Brenda Russell
    Abstract:

    Los miocitos del ventriculo cardiaco pueden modificar la composicion y la morfologia estructural de su citoesqueleto (sarcomero) ante estimulos externos que tiene como objetivo regular las propiedades contractiles necesarias para mantener el funcionamiento adecuado. La hipotesis planteada es que uno de los elementos de regulacion citoesquleticos corresponde a la proteina CapZ, la cual controla el dinamismo del sarcomero en respuesta a estimulos mecanicos. Cardiomiocitos fueron cultivados en substratos con grados de rigidez fisiologica (10 kPa) o patologica (100 kPa). El dinamismo de CapZ fue probado mediante FRAP y la interaccion de CapZ con PIP2 (fosfatidilinositol 4,5-bifosfato) fue determinada mediante FRET. Se encontro que CapZ es mas dinamica en substratos patologicos que en substratos fisiologicos. A la vez, la interaccion entre CapZ y PIP2 no varia con la rigidez del substrato.

  • CapZ integrates several signaling pathways in response to mechanical stiffness.
    The Journal of general physiology, 2019
    Co-Authors: Christopher Solís, Brenda Russell
    Abstract:

    Muscle adaptation is a response to physiological demand elicited by changes in mechanical load, hormones, or metabolic stress. Cytoskeletal remodeling processes in many cell types are thought to be primarily regulated by thin filament formation due to actin-binding accessory proteins, such as the actin-capping protein. Here, we hypothesize that in muscle, the actin-capping protein (named CapZ) integrates signaling by a variety of pathways, including phosphorylation and phosphatidylinositol 4,5-bisphosphate (PIP2) binding, to regulate muscle fiber growth in response to mechanical load. To test this hypothesis, we assess mechanotransduction signaling that regulates muscle growth using neonatal rat ventricular myocytes cultured on substrates with the stiffness of the healthy myocardium (10 kPa), fibrotic myocardium (100 kPa), or glass. We investigate how PIP2 signaling affects CapZ using the PIP2 sequestering agent neomycin and the effect of PKC-mediated CapZ phosphorylation using the PKC-activating drug phorbol 12-myristate 13-acetate (PMA). Molecular simulations suggest that close interactions between PIP2 and the β-tentacle of CapZ are modified by phosphorylation at T267. Fluorescence recovery after photobleaching (FRAP) demonstrates that the kinetic binding constant of CapZ to sarcomeric thin filaments in living muscle cells increases with stiffness or PMA treatment but is diminished by PIP2 reduction. Furthermore, CapZ with a deletion of the β-tentacle that lacks the phosphorylation site T267 shows increased FRAP kinetics with lack of sensitivity to PMA treatment or PIP2 reduction. Forster resonance energy transfer (FRET) probes the molecular interactions between PIP2 and CapZ, which are decreased by PIP2 availability or by the β-tentacle truncation. These data suggest that CapZ is bound to actin tightly in the idle, locked state, with little phosphorylation or PIP2 binding. However, this tight binding is loosened in growth states triggered by mechanical stimuli such as substrate stiffness, which may have relevance to fibrotic heart disease.

  • Cyclic mechanical strain of myocytes modifies CapZβ1 post translationally via PKCε
    Journal of Muscle Research and Cell Motility, 2015
    Co-Authors: Ying-hsi Lin, Michael A. Mkrtschjan, Erik R. Swanson, Brenda Russell
    Abstract:

    The heart is exquisitely sensitive to mechanical stimuli and adapts to increased demands for work by enlarging the cardiomyocytes. In order to determine links between mechano-transduction mechanisms and hypertrophy, neonatal rat ventricular myocytes (NRVM) were subjected to physiologic strain for analysis of the dynamics of the actin capping protein, CapZ, and its post-translational modifications (PTM). CapZ binding rates were assessed after strain by fluorescence recovery after photobleaching (FRAP) of green fluorescent protein (GFP) expressed by a GFP-CapZβ1 adenovirus. To assess the role of the protein kinase C epsilon isoform (PKCε), rest or cyclic strain were combined with specific PKCε activation by constitutively active PKCε, or by inhibition with dominant negative PKCε (dnPKCε) expression. Significant increases of CapZ FRAP kinetics with strain were blunted by dnPKCε, suggesting that PKCε is involved in mechano-transduction signaling. Similar combinations of strain and PKC regulation in NRVMs were studied by PTM profiles of CapZβ1 using quantitative two-dimensional gel electrophoresis. The significantly increased charge on CapZ seen with mechanical strain was reversed by the addition of dnPKCε. Potential clinical relevance was confirmed in vivo by PTMs of CapZ in the failing heart of one-year old transgenic mice over-expressing PKCε. Furthermore, with strain there was significant PKCε translocation to the Z-disc and co-localization with CapZβ1 or α-actinin, which was quantified on confocal images. A hypothetical model is presented proposing that one destination of the mechanotransduction signaling pathways might be for PTMs of CapZ thereby regulating actin capping and filament assembly.

  • Cyclic mechanical strain of myocytes modifies CapZβ1 post translationally via PKCε.
    Journal of Muscle Research and Cell Motility, 2015
    Co-Authors: Ying-hsi Lin, Michael A. Mkrtschjan, Erik R. Swanson, Brenda Russell
    Abstract:

    The heart is exquisitely sensitive to mechanical stimuli and adapts to increased demands for work by enlarging the cardiomyocytes. In order to determine links between mechano-transduction mechanisms and hypertrophy, neonatal rat ventricular myocytes (NRVM) were subjected to physiologic strain for analysis of the dynamics of the actin capping protein, CapZ, and its post-translational modifications (PTM). CapZ binding rates were assessed after strain by fluorescence recovery after photobleaching (FRAP) of green fluorescent protein (GFP) expressed by a GFP-CapZβ1 adenovirus. To assess the role of the protein kinase C epsilon isoform (PKCe), rest or cyclic strain were combined with specific PKCe activation by constitutively active PKCe, or by inhibition with dominant negative PKCe (dnPKCe) expression. Significant increases of CapZ FRAP kinetics with strain were blunted by dnPKCe, suggesting that PKCe is involved in mechano-transduction signaling. Similar combinations of strain and PKC regulation in NRVMs were studied by PTM profiles of CapZβ1 using quantitative two-dimensional gel electrophoresis. The significantly increased charge on CapZ seen with mechanical strain was reversed by the addition of dnPKCe. Potential clinical relevance was confirmed in vivo by PTMs of CapZ in the failing heart of one-year old transgenic mice over-expressing PKCe. Furthermore, with strain there was significant PKCe translocation to the Z-disc and co-localization with CapZβ1 or α-actinin, which was quantified on confocal images. A hypothetical model is presented proposing that one destination of the mechanotransduction signaling pathways might be for PTMs of CapZ thereby regulating actin capping and filament assembly.

  • is dedicated to innovative approaches to the study of cell and molecular physiology. It is published 12 timesAJP- Cell Physiology o n
    2015
    Co-Authors: First Published Mar, Brenda Russell, Thomas J. Hartman, Jody L. Martin, Am Physiol Cell J Physiol, John R. Solaro, Allen M. Samarel
    Abstract:

    mechanisms phenylephrine via PIP2- and PKC-dependent CapZ dynamics are altered by endothelin-1 and You might find this additional information useful... 36 articles, 23 of which you can access free at: This article cite

W. Glen Pyle - One of the best experts on this subject based on the ideXlab platform.

  • Reduced cardiac CapZ protein protects hearts against acute ischemia–reperfusion injury and enhances preconditioning
    Journal of molecular and cellular cardiology, 2011
    Co-Authors: Feng Hua Yang, W. Glen Pyle
    Abstract:

    The Z-disc protein CapZ has historically been classified as a structural element, anchoring sarcomeric actin. Our previous work expanded its role to include signal transduction, as CapZ transgenic myofilaments are less sensitive to protein kinase C (PKC). Myocardial PKC has paradoxical effects, mediating both preconditioning and ischemia-reperfusion (IR) injury. Our objective was to determine how decreased CapZ affects IR injury and cardiac preconditioning. Mouse hearts were subjected to 20 min global ischemia and 60 min reperfusion. Some hearts were preconditioned with intermittent IR (IPC). Left ventricular function was assessed and myocardial tissue collected post-IR for molecular analysis and tissue staining. Post-ischemic function was significantly better and infarct size smaller in CapZ transgenic hearts, as compared to wildtype. IPC decreased IR damage in both wildtype and CapZ transgenic hearts, although CapZ transgenic hearts performed significantly better than wildtype. Immunoblotting revealed increased myofilament-associated PKC-α and -e following IR in wildtype hearts, but no change in PKC-δ or -ζ. By contrast, post-IR myofilament-associated PKC-α was significantly higher in CapZ transgenic mice but the rise in PKC-e was attenuated. Both PKC-δ and PKC-ζ decreased in CapZ transgenic myofilaments following IR. IPC increased myofilament-associated PKC-α and -e, while decreasing PKC-δ in wildtype hearts. Preconditioned CapZ IPC hearts showed attenuated increases in myofilament PKC-α and -e, but also a significant decrease in myofilament PKC-δ and -ζ. These data demonstrate significant differences in post-IR myofilament PKC in untreated and preconditioned CapZ transgenic mice. CapZ reduction did not dramatically affect post-IR myofilament function, nor did preconditioning. These results demonstrate that CapZ deficiency decreases IR injury, while providing enhanced cardioprotection with IPC. The cardioprotected phenotype of CapZ transgenic mice is associated with an altered translocation of PKC-isoforms to cardiac myofilaments.

  • Cardiac actin capping protein reduction and protein kinase C inhibition maintain myofilament function during cardioplegic arrest.
    Cellular physiology and biochemistry : international journal of experimental cellular physiology biochemistry and pharmacology, 2011
    Co-Authors: Feng Hua Yang, W. Glen Pyle
    Abstract:

    Background: Heart transplantation is associated with cold, cardioplegic arrest that impairs myocardial function. Protein Kinase C (PKC) suppression of myofilaments may contribute to this dysfunction. CapZ-deficient cardiac myofilaments are unresponsive to PKC. We hypothesized that myofilaments from CapZ-deficient transgenic hearts are resistant to cardioplegic dysfunction and that PKC inhibition improves function. Methods: Heart function was assessed using a Langendorff apparatus. Myofilaments isolated from murine hearts were assessed with an actomyosin MgATPase assay and protein phosphorylation gels. PKC activation was examined by immunoblotting. Results: Wildtype hearts showed impaired function after cardioplegic arrest. CapZ-deficient transgenic mouse hearts performed significantly better after 1 h cardioplegia than wildtype hearts, but not after 4 h cardioplegic arrest. Wildtype myofilaments had depressed activation at 1 and 4 h cardioplegic arrest, as demonstrated by reduced actomyosin MgATPase activity. CapZ-deficient myofilaments showed no reduced actomyosin MgATPase activity at either time. Troponin I (TnI) phosphorylation increased by approximately 20% at 1 and 4 h in wildtype mice. Myosin binding protein C (MyBP-C),and troponin T (TnT) phosphorylation increased by less than 10% at 1 h, and tended to rise at 4 h. Myofilament protein phosphorylation was largely unchanged in CapZ-deficient hearts at 1 h, but MyBP-C tended to be dephosphorylated at 4 h cardioplegic arrest. Myofilament-associated PKC-D , -EII, -G, and -H increased at 1 and 4 h cardioplegia in wildtype hearts, whereas only PKC-D increased in transgenic myofilaments at 1 h. PKC inhibition abolished the cardioplegic-dependent changes in actomyosin MgATPase activity and TnI phosphorylation of wildtype myofilaments. Conclusions: We demonstrate a direct link between PKC activation and myofilament dysfunction associated with cold, cardioplegic arrest. Moreover, we show for the first time a cardioprotective benefit of decreased cardiac CapZ.

  • Cardiac myofilament regulation by protein phosphatase type 1α and CapZ
    Biochemistry and cell biology = Biochimie et biologie cellulaire, 2008
    Co-Authors: Fenghua Yangf. Yang, David L. Aiellod.l. Aiello, W. Glen Pyle
    Abstract:

    Myofilament regulation by protein kinases is well characterized, but relatively little is known about protein phosphatase control of myofilaments. Increased protein phosphatase type 1 (PP1) activity observed in failing hearts underscores the need for investigation of this intracellular signal, including the elements that regulate its activity. The Z-disc protein CapZ controls protein kinase C (PKC) regulation of cardiac myofilaments, but whether this effect is specific to PKC, or CapZ plays a general role in intracellular signalling, is not known. We sought to determine how the alpha isoform of PP1 (PP1alpha) regulates murine cardiac myofilaments and whether CapZ influences PP1alpha-dependent regulation of cardiac myofilaments. Immunoblot analysis showed PP1alpha binding to cardiac myofilaments. Exogenous PP1alpha increased myofilament Ca2+ sensitivity and maximal actomyosin Mg2+-ATPase activity while dephosphorylating myosin binding protein C, troponin T, troponin I, and myosin light chain 2. Extraction of CapZ decreased myofilament-associated PP1alpha and attenuated the effects of PP1alpha on myofilament activation. PP1alpha-dependent dephosphorylation of myofilament proteins was reduced with CapZ extraction, except for troponin I. Extracting CapZ after PP1alpha treatment allowed most of the PP1alpha-dependent effects on myofilament activation to remain, indicating that CapZ removal modestly desensitizes cardiac myofilaments to dephosphorylation. Our results demonstrate myofilament regulation by PP1alpha and support the concept that cardiac Z-discs are vital components in intracellular signalling.

  • Control of cardiac myofilament activation and PKC-βII signaling through the actin capping protein, CapZ☆
    Journal of molecular and cellular cardiology, 2006
    Co-Authors: W. Glen Pyle, Gustavo La Rotta, Pieter P. De Tombe, Marius P. Sumandea, R. John Solaro
    Abstract:

    Actin capping protein (CapZ) anchors the barbed ends of sarcomeric actin to the Z-disc. Myofilaments from transgenic mice (TG-CapZ) expressing a reduced amount of CapZ demonstrate altered function and protein kinase C (PKC) signaling [Pyle WG, Hart MC, Cooper JA, Sumandea MP, de Tombe PP, and Solaro RJ., Circ. Res. 90 (2002) 1299-306]. The aims of the current study were to determine the direct effects of CapZ on myofilament function and on PKC signaling to the myofilaments. Our studies compared mechanical properties of single myocytes from TG-CapZ mouse hearts to wild-type myocytes from which CapZ was extracted using PIP(2). We found that myofilaments from CapZ-deficient transgenic myocardium exhibited increased Ca(2+) sensitivity and maximum isometric tension. The extraction of CapZ from wild-type myofilaments replicated the increase in maximum isometric tension, but had no effect on myofilament Ca(2+) sensitivity. Immunoblot analysis revealed that the extraction of CapZ was associated with a reduction in myofilament-associated PKC-beta(II) and that CapZ-deficient transgenic myofilaments also lacked PKC-beta(II). Treatment of wild-type myofilaments with recombinant PKC-beta(II) reduced myofilament Ca(2+) sensitivity, whereas this effect was attenuated in myofilaments from TG-CapZ mice. Our results indicate that cardiac CapZ directly controls maximum isometric tension generation, and establish CapZ as an important component in anchoring PKC-beta(II) at the myofilaments, and for mediating the effects of PKC-beta(II) on myofilament function.

  • Searching for the missing link: a role for the actin capping protein in heart failure.
    The Canadian journal of cardiology, 2004
    Co-Authors: W. Glen Pyle
    Abstract:

    Les disques Z cardiaques ont toujours ete classes parmi les elements myocardiques passifs. Les disques Z se trouvent a la jonction entre le cytosquelette et les myofilaments, assurant un lien physique entre lesarcomere, le noyau, la membrane et le reticulum sarcoplasmique. De plus, de nombreux messagers moleculaires s'assemblent au disque Z. La combinaison de signaux physiques et chimiques traversant le disque Z fait de cet element myocardique une station de commutation essentielle du coeur et indique un potentiel regulateur marque. A l'aide de la proteine de coiffe de l'actine (CapZ) pour representer le disque Z, on a decouvert que le fait de diminuer la CapZ accroit le developpement de la force et inhibe la proteine kinase C, un messager d'insuffisance cardiaque. Ces resultats revelent le potentiel de la CapZ comme cible therapeutique dans la prise en charge de l'insuffisance cardiaque. Des recherches s'imposent pour determiner les mecanismes par lesquels les modifications a la CapZ influent sur la fonction myocardique et la signalisation intracellulaire, et pour mettre au point des strategies faisables qui peuvent manipuler la CapZ dans le coeur intact.

Sofia Khaitlina - One of the best experts on this subject based on the ideXlab platform.

  • 1 REGULATION OF SODIUM CHANNEL ACTIVITY BY CAPPING OF ACTIN FILAMENTS
    2013
    Co-Authors: Ekaterina Shumilina, Yuri A. Negulyaev, Elena A. Morachevskaya, Sofia Khaitlina
    Abstract:

    cell. 2 Ion transport in various tissues can be regulated by the cortical actin cytoskeleton. Specifically, involvement of actin dynamics in the regulation of non-voltage-gated sodium channels has been shown. Here, inside-out patch clamp experiments were performed to study the effect of the heterodimeric actin capping protein CapZ on sodium channel regulation in leukemia K562 cells. The channels were activated by cytochalasininduced disruption of actin filaments and inactivated by G-actin under ionic conditions promoting rapid actin polymerization. CapZ had no direct effect on channel activity. However, being added together with G-actin, CapZ prevented actin-induced channel inactivation, and this effect occurred at CapZ/actin molar ratios from 1:5 to 1:100. When actin was allowed to polymerize at the plasma membrane to induce partial channel inactivation, subsequent addition of CapZ restored the channel activity. These results can be explained by CapZ-induced inhibition of further assembly of actin filaments at the plasma membrane due to the modification of actin dynamics by CapZ. No effect on the channel activity was observed in response to F-actin confirming that the mechanism of channel inactivation does not involve interaction of the channel with preformed filaments. Our data show that actin-capping protein can participate in the cytoskeletonassociated regulation of sodium transport in nonexcitable cells

  • Regulation of Sodium Channel Activity by Capping of Actin Filaments
    Molecular biology of the cell, 2003
    Co-Authors: Ekaterina Shumilina, Yuri A. Negulyaev, Elena A. Morachevskaya, Horst Hinssen, Sofia Khaitlina
    Abstract:

    Ion transport in various tissues can be regulated by the cortical actin cytoskeleton. Specifically, involvement of actin dynamics in the regulation of nonvoltage-gated sodium channels has been shown. Herein, inside-out patch clamp experiments were performed to study the effect of the heterodimeric actin capping protein CapZ on sodium channel regulation in leukemia K562 cells. The channels were activated by cytochalasin-induced disruption of actin filaments and inactivated by G-actin under ionic conditions promoting rapid actin polymerization. CapZ had no direct effect on channel activity. However, being added together with G-actin, CapZ prevented actin-induced channel inactivation, and this effect occurred at CapZ/actin molar ratios from 1:5 to 1:100. When actin was allowed to polymerize at the plasma membrane to induce partial channel inactivation, subsequent addition of CapZ restored the channel activity. These results can be explained by CapZ-induced inhibition of further assembly of actin filaments at the plasma membrane due to the modification of actin dynamics by CapZ. No effect on the channel activity was observed in response to F-actin, confirming that the mechanism of channel inactivation does not involve interaction of the channel with preformed filaments. Our data show that actin-capping protein can participate in the cytoskeleton-associated regulation of sodium transport in nonexcitable cells.

Kwang Seok Ahn - One of the best experts on this subject based on the ideXlab platform.

  • Capsazepine inhibits JAK/STAT3 signaling, tumor growth, and cell survival in prostate cancer.
    Oncotarget, 2016
    Co-Authors: Jong Hyun Lee, Chulwon Kim, Seung Ho Baek, Seok-geun Lee, Woong Mo Yang, Gautam Sethi, Kwang Seok Ahn
    Abstract:

    // Jong Hyun Lee 1 , Chulwon Kim 1 , Seung Ho Baek 1 , Jeong-Hyeon Ko 1 , Seok Geun Lee 1 , Woong Mo Yang 1 , Jae-Young Um 1 , Gautam Sethi 2 , Kwang Seok Ahn 1 1 College of Korean Medicine, Kyung Hee University, Seoul 130-701, Republic of Korea 2 Department of Pharmacology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117597 Correspondence to: Gautam Sethi, email: phcgs@nus.edu.sg Kwang Seok Ahn, email: ksahn@khu.ac.kr Keywords: capsazepine, STAT3, PTPe, apoptosis, prostate cancer Received: January 6, 2016      Accepted: July 14, 2016      Published: July 22, 2016 ABSTRACT Persistent STAT3 activation is seen in many tumor cells and promotes malignant transformation. Here, we investigated whether capsazepine (CapZ), a synthetic analogue of capsaicin, exerts anticancer effects by inhibiting STAT3 activation in prostate cancer cells. CapZ inhibited both constitutive and induced STAT3 activation in human prostate carcinoma cells. CapZ also inhibited activation of the upstream kinases JAK1/2 and c-Src. The phosphatase inhibitor pervanadate reversed CapZ-induced STAT3 inhibition, indicating that the effect of CapZ depends on a protein tyrosine phosphatase. CapZ treatment increased PTPe protein and mRNA levels. Moreover, siRNA-mediated knockdown of PTPe reversed the CapZ-induced induction of PTPe and inhibition of STAT3 activation, indicating that PTPe is crucial for CapZ-dependent STAT3 dephosphorylation. CapZ also decreased levels of the protein products of various oncogenes, which in turn inhibited proliferation and invasion and induced apoptosis. Finally, intraperitoneal CapZ administration decreased tumor growth in a xenograft mouse prostate cancer model and reduced p-STAT3 and Ki-67 expression. These data suggest that CapZ is a novel pharmacological inhibitor of STAT3 activation with several anticancer effects in prostate cancer cells.

  • Abstract 3807: Capsazepine attenuates JAK/STAT3 signaling pathway, proliferation, survival of prostate carcinoma DU145 cells through induction of the protein tyrosine phosphatase epsilon (PTP-epsilon)
    Molecular and Cellular Biology, 2015
    Co-Authors: Jong Hyun Lee, Chulwon Kim, Seok-geun Lee, Junhee Lee, Jung-woo Lee, Kwang Seok Ahn
    Abstract:

    Constitutive activation of signal transducers and activators of transcription (STAT) 3 is a major distinguishing feature of various human cancer cells and its activation leads to survival, proliferation, and metastasis of tumor cells. Whether the anti-proliferative, pro-apoptotic, and anti-invasive effects of capsazepine (CapZ), a synthetic analog of capsaicin, are linked to its capability to inhibit STAT3 activation was investigated. We found that CapZ suppressed both constitutive and IL-6-inducible STAT3 activation in human prostate carcinoma cells. CapZ also suppressed nuclear translocation and DNA binding activity of STAT3. The suppression was mediated through the inhibition of activation of the upstream kinases JAK1/2 and c-Src. Treatment with the protein tyrosine phosphatase (PTP) inhibitor pervanadate treatment reversed the CapZ-induced down-regulation of STAT3, thereby suggesting the involvement of a PTP. CapZ induced the expression of PTPe protein and mRNA. Moreover, knockdown of PTPe by small interfering RNA suppressed the induction of PTPe, reversed the inhibition of STAT3 activation, suggesting the critical role of PTPe in its possible mechanism of action. CapZ downregulated the expression of STAT3-regulated proliferative, antiapoptotic, angiogenetic, and metastatic gene products; and this correlated with suppression of cell proliferation and invasion, the accumulation of cells in sub-G1 phase of cell cycle, and induction of apoptosis. Overexpression of phosphorylated STAT3 led to the attenuation of CapZ-mediated cleavage of PARP as compared to the control. Thus, overall, our results suggest that CapZ is a novel inhibitor of STAT3 activation and thus may have a potential in negative regulation of growth, metastasis, and angiogenesis of tumor cells. Citation Format: Jong Hyun Lee, Chulwon Kim, Seok-Geun Lee, Junhee Lee, Jung-woo Lee, Kwang Seok Ahn. Capsazepine attenuates JAK/STAT3 signaling pathway, proliferation, survival of prostate carcinoma DU145 cells through induction of the protein tyrosine phosphatase epsilon (PTP-epsilon). [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3807. doi:10.1158/1538-7445.AM2015-3807

Yves Benyamin - One of the best experts on this subject based on the ideXlab platform.

  • Interaction of actin with the capping protein, CapZ from sea bass (Dicentrarchus labrax) white skeletal muscle.
    Comparative biochemistry and physiology. Part B Biochemistry & molecular biology, 2000
    Co-Authors: Olivier Kwiateck, Iris Papa, Marie Christine Lebart, Yves Benyamin, Claude Roustan
    Abstract:

    We have compared the functional properties of CapZ from fish white skeletal muscle with those of CapZ from chicken muscle. CapZ is a heterodimer, which enhances actin nucleation and inhibits the depolymerization process by binding to the barbed ends of microfilaments. Here, we report the interaction of CapZ not only with F-actin, but also with monomeric actin. The affinity of sea bass CapZ for G-actin estimated by enzyme-linked immunosorbent assay (ELISA) was in the microM range. This association was PIP2 dependent. Binding contacts with the barbed end of actin were delimited by both ELISA and fluorescence approaches. One site (actin sequence 338-348) was located in a helical region of the subdomain 1, region already implicated in the interaction with other actin binding proteins such as gelsolin. Another site implicates the C-terminal region (sequence 360-372) of actin. Finally, the partial competition of antibodies directed against CapZ alpha or beta-subunits towards CapZ interaction with actin filaments suggests both subunits participate in the complex with actin.

  • Alpha actinin–CapZ, an anchoring complex for thin filaments in Z-line
    Journal of Muscle Research & Cell Motility, 1999
    Co-Authors: Iris Papa, Marie Christine Lebart, Claude Roustan, Catherine Astier, Olivier Kwiatek, Fabrice Raynaud, Chantal Bonnal, Yves Benyamin
    Abstract:

    CapZ is a widely distributed and highly conserved, heterodimeric protein, that nucleates actin polymerization and binds to the barbed ends of actin filaments, preventing the addition or loss of actin monomers. CapZ interaction with actin filaments was shown to be of high affinity and decreased in the presence of PIP2. CapZ was located in nascent Z-lines during skeletal muscle myofibrillogenesis before the striated appearance of thin filaments in sarcomers. In this study, the stabilization and the anchorage of thin filaments were explored through identification of CapZ partners in the Z-line. Fish (sea bass) striated white muscle and its related Z-line proteins were selected since they correspond to the simplest Z-line organization. We report here the interaction between purified CapZ and α-actinin, a major component of Z filaments and polar links in Z-discs. Affinity of CapZ for α-actinin, estimated by fluorescence and immunochemical assays, is in the μ m range. This association was found to be independent of actin and shown to be weakened in the presence of phosphoinositides. Binding contacts on the α-actinin molecule lie in the 55 kDa repetitive domain. A model including CapZ/α-actinin/titin/actin interactions is proposed considering Luther's 3D Z-line reconstruction.

  • ALPHA ACTININ-CapZ, AN ANCHORING COMPLEX FOR THIN FILAMENTS IN Z-LINE
    Journal of muscle research and cell motility, 1999
    Co-Authors: Iris Papa, Marie Christine Lebart, Claude Roustan, Catherine Astier, Olivier Kwiatek, Fabrice Raynaud, Chantal Bonnal, Yves Benyamin
    Abstract:

    CapZ is a widely distributed and highly conserved, heterodimeric protein, that nucleates actin polymerization and binds to the barbed ends of actin filaments, preventing the addition or loss of actin monomers. CapZ interaction with actin filaments was shown to be of high affinity and decreased in the presence of PIP2. CapZ was located in nascent Z-lines during skeletal muscle myofibrillogenesis before the striated appearance of thin filaments in sarcomers. In this study, the stabilization and the anchorage of thin filaments were explored through identification of CapZ partners in the Z-line. Fish (sea bass) striated white muscle and its related Z-line proteins were selected since they correspond to the simplest Z-line organization. We report here the interaction between purified CapZ and α-actinin, a major component of Z filaments and polar links in Z-discs. Affinity of CapZ for α-actinin, estimated by fluorescence and immunochemical assays, is in the μ m range. This association was found to be independent of actin and shown to be weakened in the presence of phosphoinositides. Binding contacts on the α-actinin molecule lie in the 55 kDa repetitive domain. A model including CapZ/α-actinin/titin/actin interactions is proposed considering Luther's 3D Z-line reconstruction.

  • Use of a chaotropic anion iodide in the purification of Z-line proteins: isolation of CapZ from fish white muscle.
    Protein expression and purification, 1999
    Co-Authors: Iris Papa, Marie Christine Lebart, Yves Benyamin, Catherine Astier, Olivier Kwiatek, Fabrice Raynaud, Claude Roustan
    Abstract:

    Abstract In the present study, we have described an improved method allowing the isolation of proteins which form tightly associated complexes in organized structures such as Z line in skeletal muscle. This procedure is based on both extraction and chromatography in the presence of a chaotropic agent. KI at medium concentration (0.6 M) was selected, taking into account its dissociating activity and mild effect on the native state of proteins. This procedure was applied to purify and to characterize for the first time a CapZ from fish white muscle, a protein involved in the stabilization of the filaments in Z line. The α and β CapZ subunits were identified using anti-synthetic peptide antibodies directed against conserved sequences derived from chicken CapZ. The protocol can be also used for the isolation of other muscular proteins such as α-actinin and actin. Finally this technique may be utilized to obtain a good amount of capping protein which could be employed in experiments of microfilament dynamics.