The Experts below are selected from a list of 1941 Experts worldwide ranked by ideXlab platform
Mary C Beckerle - One of the best experts on this subject based on the ideXlab platform.
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LIM domains target actin regulators paxillin and Zyxin to sites of stress fiber strain.
PloS one, 2013Co-Authors: Mark A. Smith, Elizabeth Blankman, Christopher C. Jensen, Laura M. Hoffman, Nicholas O. Deakin, Christopher E. Turner, Mary C BeckerleAbstract:Contractile actomyosin stress fibers are critical for maintaining the force balance between the interior of the cell and its environment. Consequently, the actin cytoskeleton undergoes dynamic mechanical loading. This results in spontaneous, stochastic, highly localized strain events, characterized by thinning and elongation within a discrete region of stress fiber. Previous work showed the LIM-domain adaptor protein, Zyxin, is essential for repair and stabilization of these sites. Using live imaging, we show paxillin, another LIM-domain adaptor protein, is also recruited to stress fiber strain sites. Paxillin recruitment to stress fiber strain sites precedes Zyxin recruitment. Zyxin and paxillin are each recruited independently of the other. In cells lacking paxillin, actin recovery is abrogated, resulting in slowed actin recovery and increased incidence of catastrophic stress fiber breaks. For both paxillin and Zyxin, the LIM domains are necessary and sufficient for recruitment. This work provides further evidence of the critical role of LIM-domain proteins in responding to mechanical stress in the actin cytoskeleton.
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Stretch-induced actin remodeling requires targeting of Zyxin to stress fibers and recruitment of actin regulators.
Molecular biology of the cell, 2012Co-Authors: Laura M. Hoffman, Christopher C. Jensen, Aashi Chaturvedi, Masaaki Yoshigi, Mary C BeckerleAbstract:Reinforcement of actin stress fibers in response to mechanical stimulation depends on a posttranslational mechanism that requires the LIM protein Zyxin. The C-terminal LIM region of Zyxin directs the force-sensitive accumulation of Zyxin on actin stress fibers. The N-terminal region of Zyxin promotes actin reinforcement even when Rho kinase is inhibited. The mechanosensitive integrin effector p130Cas binds Zyxin but is not required for mitogen-activated protein kinase–dependent Zyxin phosphorylation or stress fiber remodeling in cells exposed to uniaxial cyclic stretch. α-Actinin and Ena/VASP proteins bind to the stress fiber reinforcement domain of Zyxin. Mutation of their docking sites reveals that Zyxin is required for recruitment of both groups of proteins to regions of stress fiber remodeling. Zyxin-null cells reconstituted with Zyxin variants that lack either α-actinin or Ena/VASP-binding capacity display compromised response to mechanical stimulation. Our findings define a bipartite mechanism for stretch-induced actin remodeling that involves mechanosensitive targeting of Zyxin to actin stress fibers and localized recruitment of actin regulatory machinery.
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The LIM Protein Zyxin Binds CARP-1 and Promotes Apoptosis
Genes & cancer, 2010Co-Authors: Martial Hervy, Laura M. Hoffman, Christopher C. Jensen, Mark A. Smith, Mary C BeckerleAbstract:Zyxin is a dual-function LIM domain protein that regulates actin dynamics in response to mechanical stress and shuttles between focal adhesions and the cell nucleus. Here we show that Zyxin contributes to UV-induced apoptosis. Exposure of wild-type fibroblasts to UV-C irradiation results in apoptotic cell death, whereas cells harboring a homozygous disruption of the Zyxin gene display a statistically significant survival advantage. To gain insight into the molecular mechanism by which Zyxin promotes apoptotic signaling, we expressed an affinity-tagged Zyxin variant in Zyxin-null cells and isolated Zyxinassociated proteins from cell lysates under physiological conditions. A 130-kDa protein that was co-isolated with Zyxin was identified by microsequence analysis as the Cell Cycle and Apoptosis Regulator Protein-1 (CARP-1). CARP-1 associates with the LIM region of Zyxin. Zyxin lacking the CARP-1 binding region shows reduced proapoptotic activity in response to UV-C irradiation. We demonstrate that CARP-1 is a nuclear protein. Zyxin is modified by phosphorylation in cells exposed to UV-C irradiation, and nuclear accumulation of Zyxin is induced by UV-C exposure. These findings highlight a novel mechanism for modulating the apoptotic response to UV irradiation.
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α-Actinin links LPP, but not Zyxin, to cadherin-based junctions
Biochemical and biophysical research communications, 2008Co-Authors: Marc D.h. Hansen, Mary C BeckerleAbstract:The actin regulator VASP localizes to cell-cell junctions and has been implicated in cell-cell adhesion. VASP is recruited to sites of actin dynamics by interactions with proline rich FPPPPP motifs. Zyxin and its relative LPP use FPPPPP motifs to recruit VASP to specific cellular locations, thus directing changes in actin dynamics. It has been proposed that Zyxin and LPP localize to cell-cell junctions by binding α-actinin. However, the role of α-actinin in recruiting Zyxin and LPP to cell-cell contacts has not been experimentally tested. Here we use Zyxin and LPP fragments to demonstrate that the α-actinin binding site of both proteins independently targets to cell-cell junctions. While the α-actinin binding site is required for LPP localization and function at cell-cell contacts, Zyxin localization and function at cell-cell contacts is independent of the α-actinin binding site. Perturbation of LPP function, but not that of Zyxin, results in changes in anchoring of α-actinin to detergent-insoluble networks at cell-cell contacts.
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opposing roles of Zyxin lpp acta repeats and the lim domain region in cell cell adhesion
Journal of Biological Chemistry, 2006Co-Authors: Marc D.h. Hansen, Mary C BeckerleAbstract:Cadherins mediate cell-cell adhesion by linking cell junctions to actin networks. Although several actin regulatory systems have been implicated in cell-cell adhesion, it remains unclear how such systems drive cadherin-actin network formation and how they are regulated to coincide with initiation of adhesion. Previous work implicated VASP in assembly of cell-cell junctions in keratinocytes and the VASP-binding protein Zyxin colocalizes with VASP at cell-cell junctions. Here we examine how domains in Zyxin and its relative LPP contribute to cell-cell junction assembly. Using a quantitative assay for cell-cell adhesion, we demonstrate that Zyxin and LPP function to increase the rate of early cell-cell junction assembly through the VASP-binding ActA repeat region. We also identify the LIM region of Zyxin and LPP to be a regulatory domain that blocks function of these proteins. Deletion of the LIM domains drives adhesion and increases VASP level in detergent insoluble cadherin-actin. Dominant-negative Zyxin/LPP mutants reduce the rate of adhesion, lower VASP levels in detergent-insoluble cadherin-actin networks, and allow for the accumulation of capping protein at cell-cell contacts. These data implicate the LIM domains of Zyxin and LPP in regulating cell-cell junction assembly through VASP.
Marc D.h. Hansen - One of the best experts on this subject based on the ideXlab platform.
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Zyxin-VASP interactions alter actin regulatory activity in Zyxin-VASP complexes
Cellular & Molecular Biology Letters, 2013Co-Authors: Jacob Grange, James D. Moody, Marc P.a. Ascione, Marc D.h. HansenAbstract:Cell-cell and cell-substrate adhesions are sites of dramatic actin rearrangements and where actin-membrane connections are tightly regulated. Zyxin-VASP complexes localize to sites of cell-cell and cell-substrate adhesion and function to regulate actin dynamics and actin-membrane connections at these sites. To accomplish these functions, Zyxin recruits VASP to cellular sites via proline-rich binding sites near Zyxin’s amino terminus. While the prevailing thought has been that Zyxin simply acts as a scaffold protein for VASP binding, the identification of a LIM domain-VASP interaction could complicate this view. Here we assess how Zyxin-VASP binding through both the proline rich motifs and the LIM domains alters specific VASP functions. We find that neither individual interaction alters VASP’s actin regulatory activities. In contrast, however, we find that full-length Zyxin dramatically reduces VASPmediated actin bundling and actin assembly. Taken together, these results suggest a model where Zyxin-VASP complexes occur in complex organizations with suppressed actin regulatory activity.
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a Zyxin nectin interaction facilitates Zyxin localization to cell cell adhesions
Biochemical and Biophysical Research Communications, 2011Co-Authors: Gregory S Call, Dan Brereton, Jace T. Bullard, Jarom Y. Chung, Kristen L. Meacham, David J. Morrell, David J. Reeder, Jeffrey T. Schuler, Austen D. Slade, Marc D.h. HansenAbstract:Abstract Cell–cell junction remodeling is associated with dramatic actin reorganizations. Several actin regulatory systems have been implicated in actin remodeling events as cell–cell contacts are assembled and disassembled, including Zyxin/LPP–VASP complexes. These complexes facilitate strong cell–cell adhesion by maintaining actin-membrane connections. It has been proposed that Zyxin and LPP localize to cell–cell junctions via a well-defined interaction with alpha-actinin. This was recently confirmed for LPP, but Zyxin localization at cell–cell contacts occurs independently of alpha-actinin binding. Here we seek to map the Zyxin sequence responsible for localization to cell–cell contacts and identify the protein that docks Zyxin at this cellular location. Previous results have shown that a Zyxin fragment excluding the alpha-actin binding site and the LIM domains (amino acids 51–392) can independently localize to cell–cell contacts. Here, expression of smaller Zyxin fragments show that Zyxin localization requires amino acids 230–280. A yeast-two-hybrid screen, using the central region of Zyxin as bait, resulted in the identification of the cell–cell adhesion receptor nectin-4 as a Zyxin binding partner. Further demonstrating Zyxin–nectin interactions, Zyxin binds the intracellular domain of nectin-2 in vitro. Depletion of nectin-2 from L cells expressing E-cadherin results in a loss of Zyxin localization to cell–cell contacts, demonstrating that the Zyxin–nectin interaction plays a critical role in Zyxin targeting to these sites.
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A Zyxin–nectin interaction facilitates Zyxin localization to cell–cell adhesions
Biochemical and biophysical research communications, 2011Co-Authors: S. Gregory Call, Dan Brereton, Jace T. Bullard, Jarom Y. Chung, Kristen L. Meacham, David J. Morrell, David J. Reeder, Jeffrey T. Schuler, Austen D. Slade, Marc D.h. HansenAbstract:Abstract Cell–cell junction remodeling is associated with dramatic actin reorganizations. Several actin regulatory systems have been implicated in actin remodeling events as cell–cell contacts are assembled and disassembled, including Zyxin/LPP–VASP complexes. These complexes facilitate strong cell–cell adhesion by maintaining actin-membrane connections. It has been proposed that Zyxin and LPP localize to cell–cell junctions via a well-defined interaction with alpha-actinin. This was recently confirmed for LPP, but Zyxin localization at cell–cell contacts occurs independently of alpha-actinin binding. Here we seek to map the Zyxin sequence responsible for localization to cell–cell contacts and identify the protein that docks Zyxin at this cellular location. Previous results have shown that a Zyxin fragment excluding the alpha-actin binding site and the LIM domains (amino acids 51–392) can independently localize to cell–cell contacts. Here, expression of smaller Zyxin fragments show that Zyxin localization requires amino acids 230–280. A yeast-two-hybrid screen, using the central region of Zyxin as bait, resulted in the identification of the cell–cell adhesion receptor nectin-4 as a Zyxin binding partner. Further demonstrating Zyxin–nectin interactions, Zyxin binds the intracellular domain of nectin-2 in vitro. Depletion of nectin-2 from L cells expressing E-cadherin results in a loss of Zyxin localization to cell–cell contacts, demonstrating that the Zyxin–nectin interaction plays a critical role in Zyxin targeting to these sites.
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Zyxin phosphorylation at serine 142 modulates the Zyxin head tail interaction to alter cell cell adhesion
Biochemical and Biophysical Research Communications, 2011Co-Authors: Gregory S Call, Jarom Y. Chung, John A. Davis, Braden D. Price, Tyler S. Primavera, Nick C. Thomson, Mark V. Wagner, Marc D.h. HansenAbstract:Zyxin is an actin regulatory protein that is concentrated at sites of actin–membrane association, particularly cell junctions. Zyxin participates in actin dynamics by binding VASP, an interaction that occurs via proline-rich N-terminal ActA repeats. An intramolecular association of the N-terminal LIM domains at or near the ActA repeats can prevent VASP and other binding partners from binding full-length Zyxin. Such a head–tail interaction likely accounts for how Zyxin function in actin dynamics, cell adhesion, and cell migration can be regulated by the cell. Since Zyxin binding to several partners, via the LIM domains, requires phosphorylation, it seems likely that Zyxin phosphorylation might alter the head–tail interaction and, thus, Zyxin activity. Here we show that Zyxin point mutants at a known phosphorylation site, serine 142, alter the ability of a Zyxin fragment to directly bind a separate Zyxin LIM domains fragment protein. Further, expression of the Zyxin phosphomimetic mutant results in increased localization to cell–cell contacts of MDCK cells and generates a cellular phenotype, namely inability to disassemble cell–cell contacts, precisely like that produced by expression of Zyxin mutants that lack the entire regulatory LIM domain region. These data suggest that Zyxin phosphorylation at serine 142 results in release of the head–tail interaction, changing Zyxin activity at cell–cell contacts.
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Zyxin phosphorylation at serine 142 modulates the Zyxin head–tail interaction to alter cell–cell adhesion
Biochemical and biophysical research communications, 2010Co-Authors: S. Gregory Call, Jarom Y. Chung, John A. Davis, Braden D. Price, Tyler S. Primavera, Nick C. Thomson, Mark V. Wagner, Marc D.h. HansenAbstract:Zyxin is an actin regulatory protein that is concentrated at sites of actin–membrane association, particularly cell junctions. Zyxin participates in actin dynamics by binding VASP, an interaction that occurs via proline-rich N-terminal ActA repeats. An intramolecular association of the N-terminal LIM domains at or near the ActA repeats can prevent VASP and other binding partners from binding full-length Zyxin. Such a head–tail interaction likely accounts for how Zyxin function in actin dynamics, cell adhesion, and cell migration can be regulated by the cell. Since Zyxin binding to several partners, via the LIM domains, requires phosphorylation, it seems likely that Zyxin phosphorylation might alter the head–tail interaction and, thus, Zyxin activity. Here we show that Zyxin point mutants at a known phosphorylation site, serine 142, alter the ability of a Zyxin fragment to directly bind a separate Zyxin LIM domains fragment protein. Further, expression of the Zyxin phosphomimetic mutant results in increased localization to cell–cell contacts of MDCK cells and generates a cellular phenotype, namely inability to disassemble cell–cell contacts, precisely like that produced by expression of Zyxin mutants that lack the entire regulatory LIM domain region. These data suggest that Zyxin phosphorylation at serine 142 results in release of the head–tail interaction, changing Zyxin activity at cell–cell contacts.
Andrey G. Zaraisky - One of the best experts on this subject based on the ideXlab platform.
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Cytoskeletal Protein Zyxin Inhibits the Activity of Genes Responsible for Embryonic Stem Cell Status.
Cell reports, 2020Co-Authors: Elena A. Parshina, Fedor M. Eroshkin, Eugeny E. Оrlov, Fatima K. Gyoeva, Arina G. Shokhina, Dmitry B. Staroverov, Vsevolod V. Belousov, Nadezhda Zhigalova, Egor Prokhortchouk, Andrey G. ZaraiskyAbstract:Zyxin is a cytoskeletal LIM-domain protein that regulates actin cytoskeleton assembly and gene expression. In the present work, we find that Zyxin downregulation in Xenopus laevis embryos reduces the expression of numerous genes that regulate cell differentiation, but it enhances that of several genes responsible for embryonic stem cell status, specifically klf4, pou5f3.1, pou5f3.2, pou5f3.3, and vent2.1/2. For pou5f3 family genes (mammalian POU5F1/OCT4 homologs), we show that this effect is the result of mRNA stabilization due to complex formation with the Y-box protein Ybx1. When bound to Ybx1, Zyxin interferes with the formation of these complexes, thereby stimulating pou5f3 mRNA degradation. In addition, in zebrafish embryos and human HEK293 cells, Zyxin downregulation increases mRNA levels of the pluripotency genes KLF4, NANOG, and POU5F1/OCT4. Our findings indicate that Zyxin may play a role as a switch among morphogenetic cell movement, differentiation, and embryonic stem cell status.
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The cytoskeletal protein Zyxin interacts with the zinc-finger transcription factor Zic1 and plays the role of a scaffold for Gli1 and Zic1 interactions during early development of Xenopus laevis.
Biochemical and biophysical research communications, 2018Co-Authors: N. Y. Martynova, Elena A. Parshina, L. V. Ermolina, Andrey G. ZaraiskyAbstract:We have shown recently that the cytoskeletal protein Zyxin participates in the fine tuning of the neural plate pattering in Xenopus laevis embryos by modulating activity of one of the effectors of Hedgehog (Shh) signaling cascade, the transcription factor Gli1. In the present work, we show that Zyxin can also interact with the potential modulator of the Shh pathway, the transcription factor Zic1. The interaction of proteins occurs primarily by mean of the zinc-finger domain of Zic1 and 2nd LIM domain of Zyxin. Moreover, we have also revealed the ability of the Zyxin, Zic1 and Gli1 to form a ternary complex. The activity of this complex resembles that of the previously described by other authors protein complex formed by Gli1 and Zic1, amplifying effect of the latter. The data obtained provide evidence for the scaffolding role of Zyxin for Gli1 and Zic1 interactions and confirm its role in the regulation of Shh signaling cascade.
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The cytoskeletal protein Zyxin inhibits Shh signaling during the CNS patterning in Xenopus laevis through interaction with the transcription factor Gli1.
Developmental biology, 2013Co-Authors: N. Y. Martynova, Fedor M. Eroshkin, Fatima K. Gyoeva, L. V. Ermolina, Galina V. Ermakova, Natalia S. Baturina, Andrey G. ZaraiskyAbstract:Zyxin is a cytoskeletal protein that controls cell movements by regulating actin filaments assembly, but it can also modulate gene expression owing to its interactions with the proteins involved in signaling cascades. Therefore, identification of proteins that interact with Zyxin in embryonic cells is a promising way to unravel mechanisms responsible for coupling of two major components of embryogenesis: morphogenetic movements and cell differentiation. Now we show that in Xenopus laevis embryos Zyxin can bind to and suppress activity of the primary effector of Sonic hedgehog (Shh) signaling cascade, the transcription factor Gli1. By using loss- and gain-of-function approaches, we demonstrate that Zyxin is essential for reduction of Shh signaling within the dorsal part of the neural tube of X. laevis embryo. Thus, our finding discloses a novel function of Zyxin in fine tuning of the central neural system patterning which is based on the ventral-to-dorsal gradient of Shh signaling.
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The cytoskeletal protein Zyxin--a universal regulator of cell adhesion and gene expression
Bioorganicheskaia khimiia, 2010Co-Authors: L. V. Ermolina, N. Y. Martynova, Andrey G. ZaraiskyAbstract:The attachment of a cell to the extracellular matrix or the surface of another cells affects not only the cell motility, but also gene expression. In view of this, an important problem is to establish the molecular mechanisms of signal transduction from the receptors of cell adhesion to the nucleus, in particular, to identify and investigate the protein transducers of these signals. One of these transducers, the LIM domain protein Zyxin, is predominantly localized at the sites of cell adhesion, where it participates in the assembly of actin filaments. Owing to its location near the inner surface of the membrane, Zyxin can interact with the transmembrane receptors of some signaling cascades and affect the signal transduction from the extracellular ligands of these receptors. Furthermore, under certain conditions, Zyxin moves from the sites of cell contacts to the nucleus, where it directly participates in the regulation of gene expression. Of particular interest is the function of Zyxin as a possible coordinator of gene expression and morphogenetic movements in embryogenesis. The published data discussed in the present review indicate the important role of Zyxin in transmitting information from the regions of cell contacts to the genetic apparatus of the cell.
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Transcriptional factor Xanf1 interacts with the focal adhesion protein Zyxin at early development stage of the Xenopus laevis brain
Russian Journal of Bioorganic Chemistry, 2008Co-Authors: N. Y. Martynova, Fedor M. Eroshkin, L. V. Ermolina, F. K. Gioeva, Andrey G. ZaraiskyAbstract:We searched for potential protein partners of the homeodomain transcription repressor Xanf1, a key transcriptional regulator of the early development of the forebrain, using yeast two-hybrid system and identified for the first time the LIM domain protein Zyxin from the African clawed frog Xenopus laevis. This protein is interesting, because it can play role of a signal transmitter from cell receptors to the cell nucleus and, thus, participate in the regulation of the cell morphogenetic movements and gene expression. The Zyxin interaction with Xanf1 was confirmed by the immunoprecipitation of the complex of endogenous Zyxin with the hybrid myc-Xanf1 protein from the lysate of X. laevis embryos. Using a set of deletion mutants of both proteins, we found that binding of the LIM2 domain of Zyxin with the Engrailed Homology 1 repressor domain of Xanf1 is responsible for the interaction of these proteins.
Jarom Y. Chung - One of the best experts on this subject based on the ideXlab platform.
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a Zyxin nectin interaction facilitates Zyxin localization to cell cell adhesions
Biochemical and Biophysical Research Communications, 2011Co-Authors: Gregory S Call, Dan Brereton, Jace T. Bullard, Jarom Y. Chung, Kristen L. Meacham, David J. Morrell, David J. Reeder, Jeffrey T. Schuler, Austen D. Slade, Marc D.h. HansenAbstract:Abstract Cell–cell junction remodeling is associated with dramatic actin reorganizations. Several actin regulatory systems have been implicated in actin remodeling events as cell–cell contacts are assembled and disassembled, including Zyxin/LPP–VASP complexes. These complexes facilitate strong cell–cell adhesion by maintaining actin-membrane connections. It has been proposed that Zyxin and LPP localize to cell–cell junctions via a well-defined interaction with alpha-actinin. This was recently confirmed for LPP, but Zyxin localization at cell–cell contacts occurs independently of alpha-actinin binding. Here we seek to map the Zyxin sequence responsible for localization to cell–cell contacts and identify the protein that docks Zyxin at this cellular location. Previous results have shown that a Zyxin fragment excluding the alpha-actin binding site and the LIM domains (amino acids 51–392) can independently localize to cell–cell contacts. Here, expression of smaller Zyxin fragments show that Zyxin localization requires amino acids 230–280. A yeast-two-hybrid screen, using the central region of Zyxin as bait, resulted in the identification of the cell–cell adhesion receptor nectin-4 as a Zyxin binding partner. Further demonstrating Zyxin–nectin interactions, Zyxin binds the intracellular domain of nectin-2 in vitro. Depletion of nectin-2 from L cells expressing E-cadherin results in a loss of Zyxin localization to cell–cell contacts, demonstrating that the Zyxin–nectin interaction plays a critical role in Zyxin targeting to these sites.
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A Zyxin–nectin interaction facilitates Zyxin localization to cell–cell adhesions
Biochemical and biophysical research communications, 2011Co-Authors: S. Gregory Call, Dan Brereton, Jace T. Bullard, Jarom Y. Chung, Kristen L. Meacham, David J. Morrell, David J. Reeder, Jeffrey T. Schuler, Austen D. Slade, Marc D.h. HansenAbstract:Abstract Cell–cell junction remodeling is associated with dramatic actin reorganizations. Several actin regulatory systems have been implicated in actin remodeling events as cell–cell contacts are assembled and disassembled, including Zyxin/LPP–VASP complexes. These complexes facilitate strong cell–cell adhesion by maintaining actin-membrane connections. It has been proposed that Zyxin and LPP localize to cell–cell junctions via a well-defined interaction with alpha-actinin. This was recently confirmed for LPP, but Zyxin localization at cell–cell contacts occurs independently of alpha-actinin binding. Here we seek to map the Zyxin sequence responsible for localization to cell–cell contacts and identify the protein that docks Zyxin at this cellular location. Previous results have shown that a Zyxin fragment excluding the alpha-actin binding site and the LIM domains (amino acids 51–392) can independently localize to cell–cell contacts. Here, expression of smaller Zyxin fragments show that Zyxin localization requires amino acids 230–280. A yeast-two-hybrid screen, using the central region of Zyxin as bait, resulted in the identification of the cell–cell adhesion receptor nectin-4 as a Zyxin binding partner. Further demonstrating Zyxin–nectin interactions, Zyxin binds the intracellular domain of nectin-2 in vitro. Depletion of nectin-2 from L cells expressing E-cadherin results in a loss of Zyxin localization to cell–cell contacts, demonstrating that the Zyxin–nectin interaction plays a critical role in Zyxin targeting to these sites.
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Zyxin phosphorylation at serine 142 modulates the Zyxin head tail interaction to alter cell cell adhesion
Biochemical and Biophysical Research Communications, 2011Co-Authors: Gregory S Call, Jarom Y. Chung, John A. Davis, Braden D. Price, Tyler S. Primavera, Nick C. Thomson, Mark V. Wagner, Marc D.h. HansenAbstract:Zyxin is an actin regulatory protein that is concentrated at sites of actin–membrane association, particularly cell junctions. Zyxin participates in actin dynamics by binding VASP, an interaction that occurs via proline-rich N-terminal ActA repeats. An intramolecular association of the N-terminal LIM domains at or near the ActA repeats can prevent VASP and other binding partners from binding full-length Zyxin. Such a head–tail interaction likely accounts for how Zyxin function in actin dynamics, cell adhesion, and cell migration can be regulated by the cell. Since Zyxin binding to several partners, via the LIM domains, requires phosphorylation, it seems likely that Zyxin phosphorylation might alter the head–tail interaction and, thus, Zyxin activity. Here we show that Zyxin point mutants at a known phosphorylation site, serine 142, alter the ability of a Zyxin fragment to directly bind a separate Zyxin LIM domains fragment protein. Further, expression of the Zyxin phosphomimetic mutant results in increased localization to cell–cell contacts of MDCK cells and generates a cellular phenotype, namely inability to disassemble cell–cell contacts, precisely like that produced by expression of Zyxin mutants that lack the entire regulatory LIM domain region. These data suggest that Zyxin phosphorylation at serine 142 results in release of the head–tail interaction, changing Zyxin activity at cell–cell contacts.
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Zyxin phosphorylation at serine 142 modulates the Zyxin head–tail interaction to alter cell–cell adhesion
Biochemical and biophysical research communications, 2010Co-Authors: S. Gregory Call, Jarom Y. Chung, John A. Davis, Braden D. Price, Tyler S. Primavera, Nick C. Thomson, Mark V. Wagner, Marc D.h. HansenAbstract:Zyxin is an actin regulatory protein that is concentrated at sites of actin–membrane association, particularly cell junctions. Zyxin participates in actin dynamics by binding VASP, an interaction that occurs via proline-rich N-terminal ActA repeats. An intramolecular association of the N-terminal LIM domains at or near the ActA repeats can prevent VASP and other binding partners from binding full-length Zyxin. Such a head–tail interaction likely accounts for how Zyxin function in actin dynamics, cell adhesion, and cell migration can be regulated by the cell. Since Zyxin binding to several partners, via the LIM domains, requires phosphorylation, it seems likely that Zyxin phosphorylation might alter the head–tail interaction and, thus, Zyxin activity. Here we show that Zyxin point mutants at a known phosphorylation site, serine 142, alter the ability of a Zyxin fragment to directly bind a separate Zyxin LIM domains fragment protein. Further, expression of the Zyxin phosphomimetic mutant results in increased localization to cell–cell contacts of MDCK cells and generates a cellular phenotype, namely inability to disassemble cell–cell contacts, precisely like that produced by expression of Zyxin mutants that lack the entire regulatory LIM domain region. These data suggest that Zyxin phosphorylation at serine 142 results in release of the head–tail interaction, changing Zyxin activity at cell–cell contacts.
Christopher C. Jensen - One of the best experts on this subject based on the ideXlab platform.
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The actin regulator Zyxin reinforces airway smooth muscle and accumulates in airways of fatal asthmatics.
PloS one, 2017Co-Authors: Sonia R. Rosner, Christopher D. Pascoe, Elizabeth Blankman, Christopher C. Jensen, Ramaswamy Krishnan, John G. Elliot, Alan James, Francis H. Y. Green, Jeffrey C.-y. Liu, Chun Y. SeowAbstract:Bronchospasm induced in non-asthmatic human subjects can be easily reversed by a deep inspiration (DI) whereas bronchospasm that occurs spontaneously in asthmatic subjects cannot. This physiological effect of a DI has been attributed to the manner in which a DI causes airway smooth muscle (ASM) cells to stretch, but underlying molecular mechanisms-and their failure in asthma-remain obscure. Using cells and tissues from wild type and Zyxin-/- mice we report responses to a transient stretch of physiologic magnitude and duration. At the level of the cytoskeleton, Zyxin facilitated repair at sites of stress fiber fragmentation. At the level of the isolated ASM cell, Zyxin facilitated recovery of contractile force. Finally, at the level of the small airway embedded with a precision cut lung slice, Zyxin slowed airway dilation. Thus, at each level Zyxin stabilized ASM structure and contractile properties at current muscle length. Furthermore, when we examined tissue samples from humans who died as the result of an asthma attack, we found increased accumulation of Zyxin compared with non-asthmatics and asthmatics who died of other causes. Together, these data suggest a biophysical role for Zyxin in fatal asthma.
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Zyxin is recruited to sites of actin stress fiber strain following single isotropic stretch, and is responsible for decreasing the cytoskeleton remodeling rate.
2017Co-Authors: Sonia R. Rosner, Christopher D. Pascoe, Elizabeth Blankman, Christopher C. Jensen, Ramaswamy Krishnan, Alan L. James, John G. Elliot, Francis H. Green, Jeffrey C. Liu, Chun Y. SeowAbstract:(A) Kymographic analysis of Zyxin and actin on a representative stress fiber following a single isotropic stretch. The kymograph is a time lapse montage of images of an actin SF captured in a Zyxin-/- MEF rescued with Zyxin-GFP and also expressing actin-mApple Zyxin localizes to sites of stress fiber fragmentation and facilitates stress fiber repair. (B) Quantification of actin and Zyxin intensity on a representative stress fiber following a single isotropic stretch showing Zyxin’s localization to a site of stress fiber fragmentation and the subsequent recovery of the stress fiber. Kymograph (C) and intensity analysis (D) of Zyxin recruitment to an actin strain site in a HASM cells. (E) Mean square displacements (MSD) of microbeads adherent to the cytoskeleton in Zyxin-/- and GFP-Zyxin-rescued MEFs. Zyxin-/- cells have an increased rate of remodeling, as evidenced by the upward shift of the MSD curve. (F) qPCR shows successful knockdown of Zyxin in HASM cells of approximately 90% by 48 hours. (G) Western blot shows large reduction in Zyxin in HASM cells by 72 hours post-transfection. Two replicates of this knockdown are shown as the two lanes for each condition in the western blot. (H) Cytoskeletal remodeling rate in HASM cells is increased after Zyxin knockdown as indicated by the upward shift of the MSD curve.
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Zyxin mediates post-fluidization resolidification in MASM.
2017Co-Authors: Sonia R. Rosner, Christopher D. Pascoe, Elizabeth Blankman, Christopher C. Jensen, Ramaswamy Krishnan, Alan L. James, John G. Elliot, Francis H. Green, Jeffrey C. Liu, Chun Y. SeowAbstract:(A) Representative traction maps of wild type and Zyxin-/- primary ASM cells at baseline and 10, 50 and 300 seconds after a single transient isotropic stretch. Maps show colorized distribution of forces. The legend shows the color values in KPa. (B) Net contractile moment is similar in both Zyxin-/- and wild type primary MASM cells in isometric conditions. (C) Normalized changes in net contractile moment in wild type (n = 25) and Zyxin-/- (n = 26) primary MASM cells following a transient 5–10% stretch at time = 0 seconds. (D) Normalized changes in net contractile moment in wild type (n = 22) and Zyxin-/- (n = 23) primary ASM cells following a series of three transient 5–10% stretches at 0, 310 and 620 seconds.
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Quantification of Zyxin protein expression in the airways of asthmatic and non-asthmatic bronchi.
2017Co-Authors: Sonia R. Rosner, Christopher D. Pascoe, Elizabeth Blankman, Christopher C. Jensen, Ramaswamy Krishnan, Alan L. James, John G. Elliot, Francis H. Green, Jeffrey C. Liu, Chun Y. SeowAbstract:A) Representative image of Zyxin IHC staining. Black arrow indicates ASM bands and black star indicates epithelial layer. Scale bar is 100um. B) Comparison of the percent positivity for Zyxin in the ASM layer of asthmatics and non-asthmatics expressed as a fraction of the positive control. C) Comparison of percent positivity for Zyxin in the ASM of non-asthmatic controls (n = 27), non-fatal asthmatics (n = 21) and fatal asthmatics (n = 22) in the replication cohort expressed as a fraction of the positive control. p
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LIM domains target actin regulators paxillin and Zyxin to sites of stress fiber strain.
PloS one, 2013Co-Authors: Mark A. Smith, Elizabeth Blankman, Christopher C. Jensen, Laura M. Hoffman, Nicholas O. Deakin, Christopher E. Turner, Mary C BeckerleAbstract:Contractile actomyosin stress fibers are critical for maintaining the force balance between the interior of the cell and its environment. Consequently, the actin cytoskeleton undergoes dynamic mechanical loading. This results in spontaneous, stochastic, highly localized strain events, characterized by thinning and elongation within a discrete region of stress fiber. Previous work showed the LIM-domain adaptor protein, Zyxin, is essential for repair and stabilization of these sites. Using live imaging, we show paxillin, another LIM-domain adaptor protein, is also recruited to stress fiber strain sites. Paxillin recruitment to stress fiber strain sites precedes Zyxin recruitment. Zyxin and paxillin are each recruited independently of the other. In cells lacking paxillin, actin recovery is abrogated, resulting in slowed actin recovery and increased incidence of catastrophic stress fiber breaks. For both paxillin and Zyxin, the LIM domains are necessary and sufficient for recruitment. This work provides further evidence of the critical role of LIM-domain proteins in responding to mechanical stress in the actin cytoskeleton.
