The actin and tetraspanin networks organize receptor nanoclusters to regulate B cell receptor-mediated signaling.

A key role is emerging for the cytoskeleton in coordinating receptor signaling, although the underlying molecular requirements remain unclear. Here we show that cytoskeleton disruption triggered signaling requiring not only the B cell receptor (BCR), but also the coreceptor CD19 and tetraspanin CD81, thus providing a mechanism for signal amplification upon surface-bound antigen stimulation. By using superresolution microscopy, we demonstrated that endogenous IgM, IgD, and CD19 exhibited distinct nanoscale organization within the plasma membrane of primary B cells. Upon stimulation, we detect a local convergence of receptors, although their global organization was not dramatically altered. Thus, we postulate that cytoskeleton reorganization releases BCR nanoclusters, which can interact with CD19 held in place by the tetraspanin network. These results not only suggest that receptor compartmentalization regulates antigen-induced activation but also imply a potential role for CD19 in mediating ligand-independent "tonic" BCR signaling necessary for B cell survival.

[1]  R. Geha,et al.  Impaired CD19 expression and signaling, enhanced antibody response to type II T independent antigen and reduction of B-1 cells in CD81-deficient mice. , 1997, Proceedings of the National Academy of Sciences of the United States of America.

[2]  Dan R. Littman,et al.  Signal transduction by lymphocyte antigen receptors , 1994, Cell.

[3]  D. Campana,et al.  Primary B cell immunodeficiencies: comparisons and contrasts. , 2009, Annual review of immunology.

[4]  A. Bruckbauer,et al.  The Membrane Skeleton Controls Diffusion Dynamics and Signaling through the B Cell Receptor , 2010, Immunity.

[5]  B. Alarcón,et al.  Coexistence of multivalent and monovalent TCRs explains high sensitivity and wide range of response , 2005, The Journal of experimental medicine.

[6]  R. DePinho,et al.  PI3 Kinase Signals BCR-Dependent Mature B Cell Survival , 2009, Cell.

[7]  W. Bornmann,et al.  B Cell Signaling Is Regulated by Induced Palmitoylation of CD81* , 2004, Journal of Biological Chemistry.

[8]  M. Reth,et al.  The dissociation activation model of B cell antigen receptor triggering , 2010, FEBS letters.

[9]  M. Reth.  Antigen receptor tail clue , 1989, Nature.

[10]  S. Steinberg,et al.  Characterizing the topography of membrane receptors and signaling molecules from spatial patterns obtained using nanometer-scale electron-dense probes and electron microscopy. , 2006, Micron.

[11]  Klaus Rajewsky,et al.  Impairment of T-cell-dependent B-cell responses and B-l cell development in CD19-deficient mice , 1995, Nature.

[12]  Kathy W. K. Tse,et al.  The rap GTPases regulate B cell morphology, immune-synapse formation, and signaling by particulate B cell receptor ligands. , 2008, Immunity.

[13]  V. Tybulewicz,et al.  CD19 is essential for B cell activation by promoting B cell receptor–antigen microcluster formation in response to membrane-bound ligand , 2008, Nature Immunology.

[14]  M. Heilemann,et al.  Subdiffraction-resolution fluorescence imaging with conventional fluorescent probes. , 2008, Angewandte Chemie.

[15]  Facundo D Batista,et al.  Early events in B cell activation. , 2010, Annual review of immunology.

[16]  D. Vaux,et al.  Enforced BCL2 expression in B-lymphoid cells prolongs antibody responses and elicits autoimmune disease. , 1991, Proceedings of the National Academy of Sciences of the United States of America.

[17]  S. Smith‐Gill,et al.  Altered immunoglobulin expression and functional silencing of self-reactive B lymphocytes in transgenic mice , 1988, Nature.

[18]  Bridget S. Wilson,et al.  Plasma membrane-associated proteins are clustered into islands attached to the cytoskeleton , 2006, Proceedings of the National Academy of Sciences.

[19]  A. Miller,et al.  Regulation of B lymphocyte development and activation by the CD19/CD21/CD81/Leu 13 complex requires the cytoplasmic domain of CD19. , 1997, Journal of immunology.

[20]  S. Pierce,et al.  The initiation of antigen-induced B cell antigen receptor signaling viewed in living cells by fluorescence resonance energy transfer , 2005, Nature Immunology.

[21]  M. Gold,et al.  Cofilin-Mediated F-Actin Severing Is Regulated by the Rap GTPase and Controls the Cytoskeletal Dynamics That Drive Lymphocyte Spreading and BCR Microcluster Formation , 2011, The Journal of Immunology.

[22]  D. Rawlings,et al.  Wiskott-Aldrich syndrome protein deficiency in B cells results in impaired peripheral homeostasis , 2008, Blood.

[23]  M. Reth,et al.  Oligomeric organization of the B-cell antigen receptor on resting cells , 2010, Nature.

[24]  M. Reth,et al.  Monomeric and oligomeric complexes of the B cell antigen receptor. , 2000, Immunity.

[25]  Shiang-Jong Tzeng,et al.  Live Cell Imaging Reveals that the Inhibitory FcγRIIB Destabilizes B Cell Receptor Membrane-Lipid Interactions and Blocks Immune Synapse Formation1 , 2008, The Journal of Immunology.

[26]  S. Levy,et al.  Normal Lymphocyte Development but Delayed Humoral Immune Response in CD81-null Mice , 1997, The Journal of experimental medicine.

[27]  L. Samelson,et al.  Signal transduction mediated by the T cell antigen receptor: the role of adapter proteins. , 2002, Annual review of immunology.

[28]  T. Rothstein,et al.  Cytochalasin induces an increase in cytosolic free calcium in murine B lymphocytes. , 1987, Journal of immunology.

[29]  D. Fearon,et al.  Regulation of B lymphocyte responses to foreign and self-antigens by the CD19/CD21 complex. , 2000, Annual review of immunology.

[30]  M. Fujimoto,et al.  Modulation of B lymphocyte antigen receptor signal transduction by a CD19/CD22 regulatory loop. , 1999, Immunity.

[31]  K. Rajewsky,et al.  In Vivo Ablation of Surface Immunoglobulin on Mature B Cells by Inducible Gene Targeting Results in Rapid Cell Death , 1997, Cell.

[32]  K. Campbell,et al.  Normal development but differentially altered proliferative responses of lymphocytes in mice lacking CD81 , 1997, The EMBO journal.

[33]  T. Kurosaki,et al.  Phospholipase C-γ2 and Vav cooperate within signaling microclusters to propagate B cell spreading in response to membrane-bound antigen , 2008, The Journal of Experimental Medicine.

[34]  Bridget S. Wilson,et al.  Observing Fcεri Signaling from the Inside of the Mast Cell Membrane , 2000, The Journal of cell biology.

[35]  Akihiro Kusumi,et al.  Paradigm shift of the plasma membrane concept from the two-dimensional continuum fluid to the partitioned fluid: high-speed single-molecule tracking of membrane molecules. , 2005, Annual review of biophysics and biomolecular structure.

[36]  V. Dixit,et al.  Identification of a novel receptor for B lymphocyte stimulator that is mutated in a mouse strain with severe B cell deficiency , 2001, Current Biology.

[37]  S. Smith‐Gill,et al.  Altered immunoglobulin expression and functional silencing of self-reactive B lymphocytes in transgenic mice. , 2009, Journal of immunology.

[38]  J. Cyster.  B cell follicles and antigen encounters of the third kind , 2010, Nature Immunology.

[39]  E. Unanue,et al.  LIGAND-INDUCED MOVEMENT OF LYMPHOCYTE MEMBRANE MACROMOLECULES , 1972, The Journal of experimental medicine.

[40]  Facundo D. Batista,et al.  Dynamic cortical actin remodeling by ERM proteins controls BCR microcluster organization and integrity , 2011, The Journal of experimental medicine.

[41]  Mark M Davis,et al.  TCR and Lat are expressed on separate protein islands on T cell membranes and concatenate during activation , 2010, Nature Immunology.

[42]  M. van der Burg,et al.  CD81 gene defect in humans disrupts CD19 complex formation and leads to antibody deficiency. , 2010, The Journal of clinical investigation.

[43]  T. Kurosaki,et al.  Phospholipase C-γ2 and Vav cooperate within signaling microclusters to propagate B cell spreading in response to membrane-bound antigen , 2008 .

[44]  J. Goldman,et al.  B Cell Ligand Discrimination Through a Spreading and Contraction Response , 2006, Science.

[45]  Suliana Manley,et al.  Functional nanoscale organization of signaling molecules downstream of the T cell antigen receptor. , 2011, Immunity.