A DOCK8-WIP-WASp complex links T cell receptors to the actin cytoskeleton
Erin Janssen, Mira Tohme, Mona Hedayat, Marion Leick, Sudha Kumari, Narayanaswamy Ramesh, Michel J. Massaad, Sumana Ullas, Veronica Azcutia, Christopher C. Goodnow, Katrina L. Randall, Qi Qiao, Hao Wu, Waleed Al-Herz, Dianne Cox, John Hartwig, Darrell J. Irvine, Francis W. Luscinskas, Raif S. Geha
Erin Janssen, Mira Tohme, Mona Hedayat, Marion Leick, Sudha Kumari, Narayanaswamy Ramesh, Michel J. Massaad, Sumana Ullas, Veronica Azcutia, Christopher C. Goodnow, Katrina L. Randall, Qi Qiao, Hao Wu, Waleed Al-Herz, Dianne Cox, John Hartwig, Darrell J. Irvine, Francis W. Luscinskas, Raif S. Geha
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Research Article Immunology

A DOCK8-WIP-WASp complex links T cell receptors to the actin cytoskeleton

Abstract

Wiskott-Aldrich syndrome (WAS) is associated with mutations in the WAS protein (WASp), which plays a critical role in the initiation of T cell receptor–driven (TCR-driven) actin polymerization. The clinical phenotype of WAS includes susceptibility to infection, allergy, autoimmunity, and malignancy and overlaps with the symptoms of dedicator of cytokinesis 8 (DOCK8) deficiency, suggesting that the 2 syndromes share common pathogenic mechanisms. Here, we demonstrated that the WASp-interacting protein (WIP) bridges DOCK8 to WASp and actin in T cells. We determined that the guanine nucleotide exchange factor activity of DOCK8 is essential for the integrity of the subcortical actin cytoskeleton as well as for TCR-driven WASp activation, F-actin assembly, immune synapse formation, actin foci formation, mechanotransduction, T cell transendothelial migration, and homing to lymph nodes, all of which also depend on WASp. These results indicate that DOCK8 and WASp are in the same signaling pathway that links TCRs to the actin cytoskeleton in TCR-driven actin assembly. Further, they provide an explanation for similarities in the clinical phenotypes of WAS and DOCK8 deficiency.

Authors

Erin Janssen, Mira Tohme, Mona Hedayat, Marion Leick, Sudha Kumari, Narayanaswamy Ramesh, Michel J. Massaad, Sumana Ullas, Veronica Azcutia, Christopher C. Goodnow, Katrina L. Randall, Qi Qiao, Hao Wu, Waleed Al-Herz, Dianne Cox, John Hartwig, Darrell J. Irvine, Francis W. Luscinskas, Raif S. Geha

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Figure 3

WIP bridges DOCK8 to actin.

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WIP bridges DOCK8 to actin. (A) Map of the WIPΔABD protein. (B) Represen...
(A) Map of the WIPΔABD protein. (B) Representative immunoblot of the co-IP of WIP-EGFP and WIPΔABD-EGFP with Myc-tagged DOCK8 in 293T cell transfectants. LRRC8A-Myc and EGFP transfectants were used as negative controls, and an aliquot of the lysates used for the coprecipitation was probed for Myc and EGFP to ensure equal loading. The * denotes a non-specific band. (C and D) Representative immunoblot (C) and quantitative analysis (D) of the co-IP of WIP, WASp, and actin with DOCK8 in splenic T cells from WT and WIPΔABD knockin mice. An aliquot of the lysates used for the co-IP was probed for DOCK8, WIP, WASp, and actin as a loading control. Quantification of the results was performed by calculating the relative ratio of actin/DOCK8, WIP/DOCK8, and WASp/DOCK8 in DOCK8 immunoprecipitates relative to lysates and normalizing the values to those obtained in WT T cells by setting the WT ratio to 1. Ctrl, control. Data are representative of 3 independent experiments in B and 2 independent experiments in C and D. Error bars in D represent the mean ± SEM.