Lee, J.H. et al. Roles of p-ERM and Rho-ROCK signaling in lymphocyte polarity and uropod formation. J. Cell Biol. 167, 327-337

Center for Molecular Biology and Genetics, Kyoto University, Sakyo-ku, Kyoto 606-8507, Japan.
The Journal of Cell Biology (Impact Factor: 9.83). 11/2004; 167(2):327-37. DOI: 10.1083/jcb.200403091
Source: PubMed


Front-rear asymmetry in motile cells is crucial for efficient directional movement. The uropod in migrating lymphocytes is a posterior protrusion in which several proteins, including CD44 and ezrin/radixin/moesin (ERM), are concentrated. In EL4.G8 T-lymphoma cells, Thr567 phosphorylation in the COOH-terminal domain of ezrin regulates the selective localization of ezrin in the uropod. Overexpression of the phosphorylation-mimetic T567D ezrin enhances uropod size and cell migration. T567D ezrin also induces construction of the CD44-associated polar cap, which covers the posterior cytoplasm in staurosporine-treated, uropod-disrupted EL4.G8 cells or in naturally unpolarized X63.653 myeloma cells in an actin cytoskeleton-dependent manner. Rho-associated coiled coil-containing protein kinase (ROCK) inhibitor Y-27632 disrupts the uropod but not the polar cap, indicating that Rho-ROCK signaling is required for posterior protrusion but not for ERM phosphorylation. Phosphorylated ezrin associates with Dbl through its NH2-terminal domain and causes Rho activation. Moreover, constitutively active Q63L RhoA is selectively localized in the rear part of the cells. Thus, phosphorylated ERM has a potential function in establishing plasma membrane "posteriority" in the induction of the uropod in T lymphocytes.

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Available from: Tomoya Katakai, Apr 11, 2014
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    • "ERM proteins are activated by phosphorylation of a conserved C-terminal threonine residue, which enables them to crosslink membrane receptors with the underlying actin cytoskeleton (Fehon et al., 2010). ERM protein activation is crucial for uropod formation (Ivetic and Ridley, 2004; Lee et al., 2004; Martinelli et al., 2013) and for β1-integrin-dependent T cell adhesion and polarization (Chen et al., 2013; Liu et al., 2002). Therefore, the inhibition of ERM phosphorylation in the absence of PrP likely contributes to both the lack of uropods and the reduced β1 integrin adhesion observed for PrP-deficient monocytes. "
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    ABSTRACT: The broad tissue distribution and evolutionary conservation of the GPI-anchored protein PrP suggests that it plays a role in cellular homeostasis. Since integrin adhesion determines cell behavior, the proposed role of PrP in cell adhesion may underlie the various in vitro and in vivo effects associated to PrP loss-of-function, including the immune phenotypes described in PrP(-/-) mice. We have investigated the role of PrP in the adhesion and (transendothelial) migration of human (pro)monocytes. We found that PrP regulates β1 integrin-mediated adhesion of monocytes. Additionally, PrP controls cell morphology and migratory behavior of monocytes: PrP-silenced cells show deficient uropod formation on immobilized VCAM and display bleb-like protrusions on the endothelium. Our data further show that PrP regulates ligand-induced integrin activation. Finally, we found that PrP controls the activation of several proteins involved in cell adhesion and migration, including RhoA and its effector cofilin as well as proteins of the ERM family. We propose that PrP modulates β1 integrin adhesion and migration of monocytes through RhoA-induced actin remodeling by cofilin and through the regulation of ERM-mediated membrane-cytoskeleton linkage. © 2015. Published by The Company of Biologists Ltd.
    Full-text · Article · Jul 2015 · Journal of Cell Science
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    • "The Ezrin-radixin-moesin (ERM) family of actin regulatory proteins regulate membrane tension and have been hypothesized to be important in uropod formation [6]. ERM proteins do so by specific localization of proteins to the uropods of migrating T cells [33]. ERM proteins are also responsible for localizing transmembrane proteins including CD43 to this region [8], [10]. "
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    ABSTRACT: Cell motility is a fundamental process crucial for function in many cell types, including T cells. T cell motility is critical for T cell-mediated immune responses, including initiation, activation, and effector function. While many extracellular receptors and cytoskeletal regulators have been shown to control T cell migration, relatively few signaling mediators have been identified that can modulate T cell motility. In this study, we find a previously unknown role for PKCθ in regulating T cell migration to lymph nodes. PKCθ localizes to the migrating T cell uropod and regulates localization of the MTOC, CD43 and ERM proteins to the uropod. Furthermore, PKCθ-deficient T cells are less responsive to chemokine induced migration and are defective in migration to lymph nodes. Our results reveal a novel role for PKCθ in regulating T cell migration and demonstrate that PKCθ signals downstream of CCR7 to regulate protein localization and uropod formation.
    Full-text · Article · Nov 2013 · PLoS ONE
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    • "The latter deletion mutant is thought to act as a dominant-negative mutant, by competing with endogenous ERM proteins for binding sites on receptors, thus uncoupling the receptors from the cortical actin network (Martin et al., 1995; Faure et al., 2004). Lee et al. (2004) propose that ERM proteins accumulate at the site of the future uropod and locally activate Rho, resulting in formation of a contracted uropod. In line with these findings, naïve T cells derived from mice lacking moesin show reduced in vitro chemotaxis to CXCL12 and CCL21 (Hirata et al., 2012). "
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    ABSTRACT: T cell uropods are enriched in specific proteins including adhesion receptors such as P-selectin glycoprotein ligand-1 (PSGL-1), lipid raft-associated proteins such as flotillins and ezrin/radixin/moesin (ERM) proteins which associate with cholesterol-rich raft domains and anchor adhesion receptors to the actin cytoskeleton. Using dominant mutants and siRNA technology we have tested the interactions among these proteins and their role in shaping the T cell uropod. Expression of wild type (WT) ezrin-EGFP failed to affect the morphology of human T cells or chemokine-induced uropod recruitment of PSGL-1 and flotillin-1 and -2. In contrast, expression of constitutively active T567D ezrin-EGFP induced a motile, polarized phenotype in some of the transfected T cells, even in the absence of chemokine. These cells featured F-actin-rich ruffles in the front and uropod enrichment of PSGL-1 and flotillins. T567D ezrin-EGFP was itself strongly enriched in the rear of the polarized T cells. Uropod formation induced by T567D ezrin-EGFP was actin-dependent as it was attenuated by inhibition of Rho-kinase or myosin II, and abolished by disruption of actin filaments. While expression of constitutively active ezrin enhanced cell polarity, expression of a dominant-negative deletion mutant of ezrin, 1-310 ezrin-EGFP, markedly reduced uropod formation induced by the chemokine SDF-1, T cell front-tail polarity, and capping of PSGL-1 and flotillins. Transfection of T cells with WT or T567D ezrin did not affect chemokine-mediated chemotaxis whereas 1-310 ezrin significantly impaired spontaneous 2D migration and chemotaxis. siRNA-mediated downregulation of flotillins in murine T cells attenuated moesin capping and uropod formation, indicating that ERM proteins and flotillins cooperate in uropod formation. In summary, our results indicate that activated ERM proteins function together with flotillins to promote efficient chemotaxis of T cells by structuring the uropod of migrating T cells.
    Full-text · Article · Apr 2013 · Frontiers in Immunology
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