B lymphocytes confer immune tolerance via cell surface GARP-TGF-β complex
Caroline H. Wallace, Bill X. Wu, Mohammad Salem, Ephraim A. Ansa-Addo, Alessandra Metelli, Shaoli Sun, Gary Gilkeson, Mark J. Shlomchik, Bei Liu, Zihai Li
Caroline H. Wallace, Bill X. Wu, Mohammad Salem, Ephraim A. Ansa-Addo, Alessandra Metelli, Shaoli Sun, Gary Gilkeson, Mark J. Shlomchik, Bei Liu, Zihai Li
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Research Article Immunology

B lymphocytes confer immune tolerance via cell surface GARP-TGF-β complex

Abstract

GARP, a cell surface docking receptor for binding and activating latent TGF-β, is highly expressed by platelets and activated Tregs. While GARP is implicated in immune invasion in cancer, the roles of the GARP-TGF-β axis in systemic autoimmune diseases are unknown. Although B cells do not express GARP at baseline, we found that the GARP-TGF-β complex is induced on activated human and mouse B cells by ligands for multiple TLRs, including TLR4, TLR7, and TLR9. GARP overexpression on B cells inhibited their proliferation, induced IgA class-switching, and dampened T cell–independent antibody production. In contrast, B cell–specific deletion of GARP-encoding gene Lrrc32 in mice led to development of systemic autoimmune diseases spontaneously as well as worsening of pristane-induced lupus-like disease. Canonical TGF-β signaling more readily upregulates GARP in Peyer patch B cells than in splenic B cells. Furthermore, we demonstrated that B cells are required for the induction of oral tolerance of T cell–dependent antigens via GARP. Our studies reveal for the first time to our knowledge that cell surface GARP-TGF-β is an important checkpoint for regulating B cell peripheral tolerance, highlighting a mechanism of autoimmune disease pathogenesis.

Authors

Caroline H. Wallace, Bill X. Wu, Mohammad Salem, Ephraim A. Ansa-Addo, Alessandra Metelli, Shaoli Sun, Gary Gilkeson, Mark J. Shlomchik, Bei Liu, Zihai Li

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Figure 3

Mice with GARP-deficient B cells develop spontaneous lupus-like disease.

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Mice with GARP-deficient B cells develop spontaneous lupus-like disease....
(A) Schematic of the generation of B cell–specific Lrrc32-KO mice using a mixed bone marrow chimera (BM chimera) strategy. (B) Confirmation of effective GARP KO in B cells 3 months after bone marrow reconstitution. Mice were bled and PBMCs were cultured with LPS for 72 hours to induce GARP expression. GARP and LAP expression was analyzed by flow cytometry. Representative of n = 5 biological replicates. (C) IgM-specific ANAs (1:80 dilution) were analyzed 6 months after bone marrow reconstitution (n = 5 WT and n = 6 KO). (D) IgG-specific ANAs in the sera were quantified at the indicated dilutions 6 months after bone marrow reconstitution using Hep-2 slides. Representative images are shown (n = 5 WT and n = 6 KO). Scale bar: 50 μm. (E and F) Five-μm kidney sections were stained with anti-IgM-FITC (E) and anti-IgG-FITC (F) to detect Ig deposition in the glomeruli. Quantification and representative images are shown (n = 5 WT and n = 6 KO). Scale bar: 50 μm. (G) Total Ig in the sera from mice 3 months after bone marrow reconstitution was measured by ELISA (n = 15 WT and n = 16 KO). (H) IgM, IgG1, and IgG2b levels in the sera from mice 3 months after bone marrow reconstitution were measured by ELISA. (I) Total IgA from the sera of BM chimera mice was measured by ELISA. (J) Gut bacteria-specific IgA was measured in the sera of mice 6 months after bone marrow reconstitution (n = 5 WT and n = 6 KO). Statistical analysis was performed by 2-way ANOVA; *P < 0.05. (K) CD19+ cells were detected in the spleen and mesenteric lymph node (mLN) by flow cytometry (n = 5). All statistics performed by 2-tailed t test, unless otherwise indicated; *P < 0.05, **P < 0.01, ***P < 0.001. Error bars represent SD.