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N-wasp is essential for the negative regulation of B cell receptor signaling.

Liu C, Bai X, Wu J, Sharma S, Upadhyaya A, Dahlberg CI, Westerberg LS, Snapper SB, Zhao X, Song W - PLoS Biol. (2013)

Bottom Line: The increased signaling in N-WASP knockout B cells is concurrent with increased accumulation of F-actin at the B-cell surface, enhanced B-cell spreading on the antigen-presenting membrane, delayed B-cell contraction, inhibition in the merger of signaling active BCR microclusters into signaling inactive central clusters, and a blockage of BCR internalization.The activation of N-WASP is suppressed by Bruton's tyrosine kinase-induced WASP activation, and is restored by the activation of SH2 domain-containing inositol 5-phosphatase that inhibits WASP activation.Our results reveal a new mechanism for the negative regulation of BCR signaling and broadly suggest an actin-mediated mechanism for signaling down-regulation.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology & Molecular Genetics, University of Maryland, College Park, Maryland, United States of America.

ABSTRACT
Negative regulation of receptor signaling is essential for controlling cell activation and differentiation. In B-lymphocytes, the down-regulation of B-cell antigen receptor (BCR) signaling is critical for suppressing the activation of self-reactive B cells; however, the mechanism underlying the negative regulation of signaling remains elusive. Using genetically manipulated mouse models and total internal reflection fluorescence microscopy, we demonstrate that neuronal Wiskott-Aldrich syndrome protein (N-WASP), which is coexpressed with WASP in all immune cells, is a critical negative regulator of B-cell signaling. B-cell-specific N-WASP gene deletion causes enhanced and prolonged BCR signaling and elevated levels of autoantibodies in the mouse serum. The increased signaling in N-WASP knockout B cells is concurrent with increased accumulation of F-actin at the B-cell surface, enhanced B-cell spreading on the antigen-presenting membrane, delayed B-cell contraction, inhibition in the merger of signaling active BCR microclusters into signaling inactive central clusters, and a blockage of BCR internalization. Upon BCR activation, WASP is activated first, followed by N-WASP in mouse and human primary B cells. The activation of N-WASP is suppressed by Bruton's tyrosine kinase-induced WASP activation, and is restored by the activation of SH2 domain-containing inositol 5-phosphatase that inhibits WASP activation. Our results reveal a new mechanism for the negative regulation of BCR signaling and broadly suggest an actin-mediated mechanism for signaling down-regulation.

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A working model for the coordination of N-WASP with WASP in the regulation of BCR signaling.(A) Antigen binding to the BCR induces an early activation of Btk. (B) Activated Btk in turn activates and translocates WASP to the cell surface. Activated WASP (W) stimulates actin polymerization and reorganization by binding to Arp2/3 and suppresses N-WASP activation, which drives B-cell spreading and facilitates BCR clustering and signaling. (C) The activation of SHIP-1 induced by the BCR after initial signaling inhibits Btk and WASP activation, consequently releasing N-WASP from WASP suppression. Active N-WASP decreases actin accumulation at the B-cell contact zone, which enables B-cell contraction and promotes the coalescence of BCR microclusters into a central cluster and BCR internalization. The formation of BCR central clusters and BCR endocytosis lead to the down-regulation of BCR signaling at the cell surface.
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pbio-1001704-g009: A working model for the coordination of N-WASP with WASP in the regulation of BCR signaling.(A) Antigen binding to the BCR induces an early activation of Btk. (B) Activated Btk in turn activates and translocates WASP to the cell surface. Activated WASP (W) stimulates actin polymerization and reorganization by binding to Arp2/3 and suppresses N-WASP activation, which drives B-cell spreading and facilitates BCR clustering and signaling. (C) The activation of SHIP-1 induced by the BCR after initial signaling inhibits Btk and WASP activation, consequently releasing N-WASP from WASP suppression. Active N-WASP decreases actin accumulation at the B-cell contact zone, which enables B-cell contraction and promotes the coalescence of BCR microclusters into a central cluster and BCR internalization. The formation of BCR central clusters and BCR endocytosis lead to the down-regulation of BCR signaling at the cell surface.

Mentions: Taking the results of this study and previous studies together enables us to propose a working model for the functional coordination of WASP and N-WASP during BCR activation (Figure 9). Antigen binding to the BCR induces an early activation of Btk (Figure 9A) that in turn activates and translocates WASP to the cell surface. Activated WASP stimulates actin polymerization by binding to Arp2/3, which modulates BCR lateral mobility and drives B-cell spreading. Together, these facilitate BCR clustering and signaling (Figure 9B). The activation of SHIP-1 after the initial signaling inhibits Btk activation, which decreases the level of active WASP and releases N-WASP from the suppression of WASP. The activated N-WASP reduces the surface level of F-actin probably by depolymerizing and/or transferring F-actin to BCR containing budding vesicles. Reductions in F-actin at the B-cell contact zone allow B-cells to contract, which promotes the coalescence of BCR microclusters into a central cluster. Tethering F-actin to endocytosing vesicles by N-WASP is essential for the endocytosis of antigen–BCR complexes. The formation of BCR central clusters and BCR endocytosis lead to the down-regulation of BCR signaling at the cell surface (Figure 9C). While molecular details of this working model require further investigation, this study reveals a novel function of N-WASP in the down-regulation of BCR signaling and a unique functional coordination between WASP and N-WASP during receptor signaling.


N-wasp is essential for the negative regulation of B cell receptor signaling.

Liu C, Bai X, Wu J, Sharma S, Upadhyaya A, Dahlberg CI, Westerberg LS, Snapper SB, Zhao X, Song W - PLoS Biol. (2013)

A working model for the coordination of N-WASP with WASP in the regulation of BCR signaling.(A) Antigen binding to the BCR induces an early activation of Btk. (B) Activated Btk in turn activates and translocates WASP to the cell surface. Activated WASP (W) stimulates actin polymerization and reorganization by binding to Arp2/3 and suppresses N-WASP activation, which drives B-cell spreading and facilitates BCR clustering and signaling. (C) The activation of SHIP-1 induced by the BCR after initial signaling inhibits Btk and WASP activation, consequently releasing N-WASP from WASP suppression. Active N-WASP decreases actin accumulation at the B-cell contact zone, which enables B-cell contraction and promotes the coalescence of BCR microclusters into a central cluster and BCR internalization. The formation of BCR central clusters and BCR endocytosis lead to the down-regulation of BCR signaling at the cell surface.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3818172&req=5

pbio-1001704-g009: A working model for the coordination of N-WASP with WASP in the regulation of BCR signaling.(A) Antigen binding to the BCR induces an early activation of Btk. (B) Activated Btk in turn activates and translocates WASP to the cell surface. Activated WASP (W) stimulates actin polymerization and reorganization by binding to Arp2/3 and suppresses N-WASP activation, which drives B-cell spreading and facilitates BCR clustering and signaling. (C) The activation of SHIP-1 induced by the BCR after initial signaling inhibits Btk and WASP activation, consequently releasing N-WASP from WASP suppression. Active N-WASP decreases actin accumulation at the B-cell contact zone, which enables B-cell contraction and promotes the coalescence of BCR microclusters into a central cluster and BCR internalization. The formation of BCR central clusters and BCR endocytosis lead to the down-regulation of BCR signaling at the cell surface.
Mentions: Taking the results of this study and previous studies together enables us to propose a working model for the functional coordination of WASP and N-WASP during BCR activation (Figure 9). Antigen binding to the BCR induces an early activation of Btk (Figure 9A) that in turn activates and translocates WASP to the cell surface. Activated WASP stimulates actin polymerization by binding to Arp2/3, which modulates BCR lateral mobility and drives B-cell spreading. Together, these facilitate BCR clustering and signaling (Figure 9B). The activation of SHIP-1 after the initial signaling inhibits Btk activation, which decreases the level of active WASP and releases N-WASP from the suppression of WASP. The activated N-WASP reduces the surface level of F-actin probably by depolymerizing and/or transferring F-actin to BCR containing budding vesicles. Reductions in F-actin at the B-cell contact zone allow B-cells to contract, which promotes the coalescence of BCR microclusters into a central cluster. Tethering F-actin to endocytosing vesicles by N-WASP is essential for the endocytosis of antigen–BCR complexes. The formation of BCR central clusters and BCR endocytosis lead to the down-regulation of BCR signaling at the cell surface (Figure 9C). While molecular details of this working model require further investigation, this study reveals a novel function of N-WASP in the down-regulation of BCR signaling and a unique functional coordination between WASP and N-WASP during receptor signaling.

Bottom Line: The increased signaling in N-WASP knockout B cells is concurrent with increased accumulation of F-actin at the B-cell surface, enhanced B-cell spreading on the antigen-presenting membrane, delayed B-cell contraction, inhibition in the merger of signaling active BCR microclusters into signaling inactive central clusters, and a blockage of BCR internalization.The activation of N-WASP is suppressed by Bruton's tyrosine kinase-induced WASP activation, and is restored by the activation of SH2 domain-containing inositol 5-phosphatase that inhibits WASP activation.Our results reveal a new mechanism for the negative regulation of BCR signaling and broadly suggest an actin-mediated mechanism for signaling down-regulation.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology & Molecular Genetics, University of Maryland, College Park, Maryland, United States of America.

ABSTRACT
Negative regulation of receptor signaling is essential for controlling cell activation and differentiation. In B-lymphocytes, the down-regulation of B-cell antigen receptor (BCR) signaling is critical for suppressing the activation of self-reactive B cells; however, the mechanism underlying the negative regulation of signaling remains elusive. Using genetically manipulated mouse models and total internal reflection fluorescence microscopy, we demonstrate that neuronal Wiskott-Aldrich syndrome protein (N-WASP), which is coexpressed with WASP in all immune cells, is a critical negative regulator of B-cell signaling. B-cell-specific N-WASP gene deletion causes enhanced and prolonged BCR signaling and elevated levels of autoantibodies in the mouse serum. The increased signaling in N-WASP knockout B cells is concurrent with increased accumulation of F-actin at the B-cell surface, enhanced B-cell spreading on the antigen-presenting membrane, delayed B-cell contraction, inhibition in the merger of signaling active BCR microclusters into signaling inactive central clusters, and a blockage of BCR internalization. Upon BCR activation, WASP is activated first, followed by N-WASP in mouse and human primary B cells. The activation of N-WASP is suppressed by Bruton's tyrosine kinase-induced WASP activation, and is restored by the activation of SH2 domain-containing inositol 5-phosphatase that inhibits WASP activation. Our results reveal a new mechanism for the negative regulation of BCR signaling and broadly suggest an actin-mediated mechanism for signaling down-regulation.

Show MeSH
Related in: MedlinePlus