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Ezrin and moesin are required for efficient T cell adhesion and homing to lymphoid organs.

Chen EJ, Shaffer MH, Williamson EK, Huang Y, Burkhardt JK - PLoS ONE (2013)

Bottom Line: Members of the ezrin, radixin and moesin (ERM) family of actin-binding proteins have been implicated in several aspects of this process, but studies have yielded conflicting results.In vivo, ERM-deficient T cells showed defects in homing to lymphoid organs.Taken together, these results show that ERM proteins are largely dispensable for T cell chemotaxis, but are important for β1 integrin function and homing to lymphoid organs.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology and Laboratory Medicine, The Children's Hospital of Philadelphia, Pennsylvania, USA.

ABSTRACT
T cell trafficking between the blood and lymphoid organs is a complex, multistep process that requires several highly dynamic and coordinated changes in cyto-architecture. Members of the ezrin, radixin and moesin (ERM) family of actin-binding proteins have been implicated in several aspects of this process, but studies have yielded conflicting results. Using mice with a conditional deletion of ezrin in CD4+ cells and moesin-specific siRNA, we generated T cells lacking ERM proteins, and investigated the effect on specific events required for T cell trafficking. ERM-deficient T cells migrated normally in multiple in vitro and in vivo assays, and could undergo efficient diapedesis in vitro. However, these cells were impaired in their ability to adhere to the β1 integrin ligand fibronectin, and to polarize appropriately in response to fibronectin and VCAM-1 binding. This defect was specific for β1 integrins, as adhesion and polarization in response to ICAM-1 were normal. In vivo, ERM-deficient T cells showed defects in homing to lymphoid organs. Taken together, these results show that ERM proteins are largely dispensable for T cell chemotaxis, but are important for β1 integrin function and homing to lymphoid organs.

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ERM proteins are required for T cell homing to peripheral lymphoid organs.Wild-type and ERM-deficient T cells were differentially labeled with CFSE and CMTMR, mixed in a 1∶1 ratio and injected into the tail veins of C57Bl/6 hosts. (A) Blood, spleen, and peripheral and mesenteric lymph nodes were collected 1 hour after injection, and cell suspensions were analyzed by flow cytometry. The ratio of adoptively transferred ERM-deficient to wild type T cells is shown. Data represent mean ± StDev from 5 mice in one experiment, representative of four experiments. * p<0.05, **p<0.005. (B and C) Lymph nodes were harvested 1 hour after injection for multi-photon imaging, and cell migration was tracked. (B) The percentage of cells showing active migration, defined as detailed in Materials and Methods. (C) Tracks, average velocities and average track lengths from one representative lymph node. Data represent mean ± StDev of three experiments.
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pone-0052368-g005: ERM proteins are required for T cell homing to peripheral lymphoid organs.Wild-type and ERM-deficient T cells were differentially labeled with CFSE and CMTMR, mixed in a 1∶1 ratio and injected into the tail veins of C57Bl/6 hosts. (A) Blood, spleen, and peripheral and mesenteric lymph nodes were collected 1 hour after injection, and cell suspensions were analyzed by flow cytometry. The ratio of adoptively transferred ERM-deficient to wild type T cells is shown. Data represent mean ± StDev from 5 mice in one experiment, representative of four experiments. * p<0.05, **p<0.005. (B and C) Lymph nodes were harvested 1 hour after injection for multi-photon imaging, and cell migration was tracked. (B) The percentage of cells showing active migration, defined as detailed in Materials and Methods. (C) Tracks, average velocities and average track lengths from one representative lymph node. Data represent mean ± StDev of three experiments.

Mentions: T cells trafficking in vivo involves the coordination of adhesion and chemotaxis, as well as the ability to squeeze through tissue barriers. To assess the ability of ERM-deficient T cells to migrate to secondary lymphoid organs in vivo, we used a competitive homing assay. Wild-type and ERM-deficient T cells were each fluorescently labeled, mixed in a 1∶1 ratio, and co-injected intravenously into wild-type recipient mice. After 1 hour, recipient mice were sacrificed, and blood, peripheral lymph nodes and spleens were harvested and analyzed by flow cytometry. As shown in Figure 5A, significantly reduced numbers of ERM-deficient cells were found in peripheral lymph nodes and spleen compared to wild-type cells. Conversely, the proportion of ERM-deficient cells remaining in the blood was significantly higher. These data show that expression of ERM proteins is required for efficient homing of T cells to secondary lymphoid organs.


Ezrin and moesin are required for efficient T cell adhesion and homing to lymphoid organs.

Chen EJ, Shaffer MH, Williamson EK, Huang Y, Burkhardt JK - PLoS ONE (2013)

ERM proteins are required for T cell homing to peripheral lymphoid organs.Wild-type and ERM-deficient T cells were differentially labeled with CFSE and CMTMR, mixed in a 1∶1 ratio and injected into the tail veins of C57Bl/6 hosts. (A) Blood, spleen, and peripheral and mesenteric lymph nodes were collected 1 hour after injection, and cell suspensions were analyzed by flow cytometry. The ratio of adoptively transferred ERM-deficient to wild type T cells is shown. Data represent mean ± StDev from 5 mice in one experiment, representative of four experiments. * p<0.05, **p<0.005. (B and C) Lymph nodes were harvested 1 hour after injection for multi-photon imaging, and cell migration was tracked. (B) The percentage of cells showing active migration, defined as detailed in Materials and Methods. (C) Tracks, average velocities and average track lengths from one representative lymph node. Data represent mean ± StDev of three experiments.
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Related In: Results  -  Collection

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pone-0052368-g005: ERM proteins are required for T cell homing to peripheral lymphoid organs.Wild-type and ERM-deficient T cells were differentially labeled with CFSE and CMTMR, mixed in a 1∶1 ratio and injected into the tail veins of C57Bl/6 hosts. (A) Blood, spleen, and peripheral and mesenteric lymph nodes were collected 1 hour after injection, and cell suspensions were analyzed by flow cytometry. The ratio of adoptively transferred ERM-deficient to wild type T cells is shown. Data represent mean ± StDev from 5 mice in one experiment, representative of four experiments. * p<0.05, **p<0.005. (B and C) Lymph nodes were harvested 1 hour after injection for multi-photon imaging, and cell migration was tracked. (B) The percentage of cells showing active migration, defined as detailed in Materials and Methods. (C) Tracks, average velocities and average track lengths from one representative lymph node. Data represent mean ± StDev of three experiments.
Mentions: T cells trafficking in vivo involves the coordination of adhesion and chemotaxis, as well as the ability to squeeze through tissue barriers. To assess the ability of ERM-deficient T cells to migrate to secondary lymphoid organs in vivo, we used a competitive homing assay. Wild-type and ERM-deficient T cells were each fluorescently labeled, mixed in a 1∶1 ratio, and co-injected intravenously into wild-type recipient mice. After 1 hour, recipient mice were sacrificed, and blood, peripheral lymph nodes and spleens were harvested and analyzed by flow cytometry. As shown in Figure 5A, significantly reduced numbers of ERM-deficient cells were found in peripheral lymph nodes and spleen compared to wild-type cells. Conversely, the proportion of ERM-deficient cells remaining in the blood was significantly higher. These data show that expression of ERM proteins is required for efficient homing of T cells to secondary lymphoid organs.

Bottom Line: Members of the ezrin, radixin and moesin (ERM) family of actin-binding proteins have been implicated in several aspects of this process, but studies have yielded conflicting results.In vivo, ERM-deficient T cells showed defects in homing to lymphoid organs.Taken together, these results show that ERM proteins are largely dispensable for T cell chemotaxis, but are important for β1 integrin function and homing to lymphoid organs.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology and Laboratory Medicine, The Children's Hospital of Philadelphia, Pennsylvania, USA.

ABSTRACT
T cell trafficking between the blood and lymphoid organs is a complex, multistep process that requires several highly dynamic and coordinated changes in cyto-architecture. Members of the ezrin, radixin and moesin (ERM) family of actin-binding proteins have been implicated in several aspects of this process, but studies have yielded conflicting results. Using mice with a conditional deletion of ezrin in CD4+ cells and moesin-specific siRNA, we generated T cells lacking ERM proteins, and investigated the effect on specific events required for T cell trafficking. ERM-deficient T cells migrated normally in multiple in vitro and in vivo assays, and could undergo efficient diapedesis in vitro. However, these cells were impaired in their ability to adhere to the β1 integrin ligand fibronectin, and to polarize appropriately in response to fibronectin and VCAM-1 binding. This defect was specific for β1 integrins, as adhesion and polarization in response to ICAM-1 were normal. In vivo, ERM-deficient T cells showed defects in homing to lymphoid organs. Taken together, these results show that ERM proteins are largely dispensable for T cell chemotaxis, but are important for β1 integrin function and homing to lymphoid organs.

Show MeSH
Related in: MedlinePlus