Date of Award

1-1-2018

Embargo Period

1-1-2020

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

College

College of Graduate Studies

First Advisor

Zihai Li

Second Advisor

Bei Liu

Third Advisor

Carl Atkinson

Fourth Advisor

Gary Gilkeson

Fifth Advisor

Philip Howe

Sixth Advisor

Chenthamarakshan Vasu

Abstract

Glycoprotein-A repetitions predominant (GARP, encoded by Lrrc32) is a cell surface docking receptor for latent TGFβ, with no known direct signaling mechanism. GARP is highly expressed by megakaryocytes, platelets, and activated regulatory T cells and is implicated in immune evasion by cancer. While the pleotropic roles of active TGF-β on immune cell functions have been largely elucidated, the function of the membrane latent TGF-β – GARP axis has only been investigated in regulatory T cells and platelets. We discovered that GARP is expressed on the surface of highly activated human and mouse B cells by ligands for multiple Toll-like receptors. We hypothesized that GARP may be a regulatory mechanism for maintaining B cell peripheral tolerance. This was investigated in two specific aims that probed 1) the induction and expression of GARP-LTGF-β in homeostatic proliferation and activation-induced responses in B cells and 2) the role of the GARP-LTGF-β axis on B cells in the pathogenesis of lupus and B cell-driven tolerance. First, we found that GARP is upregulated by TLRs on multiple B cell subsets, and GARP+ cells are more highly activated than GARP- cells. In gain-of-function studies, B cell surface GARP reduced B cell proliferation, induced IgA class-switching, and dampened in vivo and in vitro antibody responses. In a B cell-specific GARP knockout mouse model we observed an increase in B cell proliferation at baseline, particularly in the Peyer’s patches. However, GARP is required to constrain TLR-induced IL-6 cytokine production. The results from both the overexpression and loss-of-function studies were consistent with the known roles of active TGF-β signaling in B cells. Secondly, we observed that GARP is expressed on B cells in patients with systemic lupus erythematosus and multiple murine lupus models. Strikingly, loss of GARP on B cells leads to loss of tolerance in aged mice, as well as increased autoimmunity in the pristane-induced lupus model. We made the intriguing discovery that Peyer’s patch B cells upregulate GARP more readily with enhanced canonical TGF-β signaling comparing with splenic B cells. Furthermore, we demonstrated that B cell GARP is required for the induction of oral tolerance. Lastly, we began investigating the use of soluble GARP as a therapeutic in murine lupus models; early analysis revealed that in vivo GARP-Fc decreased B cell activation and increased systemic IgA levels. The equilibrium between self-tolerance and tumor surveillance is delicately maintained between regulatory and effector immune cells. During the course of investigating the role of B cell GARP in tolerance, we also hypothesized that this mechanism may be detrimental in the cancer microenvironment. Using a B cell-specific GARP KO mouse and MC38 colon carcinoma model we found that B cell GARP deficiency reduces primary tumor growth and promotes effector T cell functionality in the tumor microenvironment. The results in this dissertation reveal for the first time that cell surface GARP-TGF-β is a key checkpoint for regulating B cell peripheral tolerance, highlighting a novel mechanism of autoimmune diseases and a new strategy for clinical intervention.

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