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Original Research ARTICLE

Front. Immunol., 08 April 2013 | http://dx.doi.org/10.3389/fimmu.2013.00084

Ezrin/radixin/moesin proteins and flotillins cooperate to promote uropod formation in T cells

Sibylla Martinelli1, Emily J. H. Chen2, Fiona Clarke2, Ruth Lyck3, Sarah Affentranger1, Janis K. Burkhardt2 and Verena Niggli1*
  • 1Department of Pathology, University of Bern, Bern, Switzerland
  • 2Department of Pathology and Laboratory Medicine, Children’s Hospital of Philadelphia and The Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, USA
  • 3Theodor Kocher Institute, University of Bern, Bern, Switzerland

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.

Keywords: ezrin, radixin, moesin, T-lymphocyte, flotillin, cell polarization, cell migration

Citation: Martinelli S, Chen EJH, Clarke F, Lyck R, Affentranger S, Burkhardt JK and Niggli V (2013) Ezrin/radixin/moesin proteins and flotillins cooperate to promote uropod formation in T cells. Front. Immunol. 4:84. doi: 10.3389/fimmu.2013.00084

Received: 26 November 2012; Accepted: 24 March 2013;
Published online: 08 April 2013.

Edited by:

Francisco Sánchez, Universidad Autónoma de Madrid, Spain

Reviewed by:

Cosima T. Baldari, University of Siena, Italy
Juan M. Serrador, Consejo Superior de Investigaciones Cientificas, Spain

Copyright: © 2013 Martinelli, Chen, Clarke, Lyck, Affentranger, Burkhardt and Niggli. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in other forums, provided the original authors and source are credited and subject to any copyright notices concerning any third-party graphics etc.

*Correspondence: Verena Niggli, Department of Pathology, University of Bern, Murtenstr. 31, CH-3010 Bern, Switzerland. e-mail: verena.niggli@pathology.unibe.ch