Dissertation
Roles of TRAF3 in regulation of T lymphocyte biology
University of Iowa
Doctor of Philosophy (PhD), University of Iowa
Spring 2023
DOI: 10.25820/etd.007107
Abstract
The versatile intracellular adaptor protein Tumor necrosis factor receptor (TNFR) associated factor 3 (TRAF3) plays an important role in T cell biology. Mice with T cell-specific TRAF3 deficiency (T-Traf3-/-) have normal numbers of conventional T cells, but fail to clear infection with sublethal doses of the intracellular pathogen Listeria monocytogenes, and do not mount an effective antibody response following immunization with a T-dependent antigen. Further examination of TRAF3-/- T cells revealed that defective T cell receptor (TCR)/CD28-mediated signaling, and subsequently impaired proliferation and cytokine production, contribute to the failure of T-Traf3-/- mice to mount effective immune responses upon antigenic challenge.
TRAF3 is recruited to the TCR/CD28 complex upon engagement of both CD3 and CD28, and enhances TCR/CD28 signaling. The studies described in Chapters 3 and 4 of this dissertation show that TRAF3 is recruited to the TCR/CD28 complex via interactions with the adaptor protein Linker for activated T cells (LAT), promoting TCR/CD28 signaling via restraint of the LAT-associated negative regulator Downstream of kinase 1 (Dok1). The interaction between TRAF3 and LAT requires the TRAF-C domain of TRAF3, as well as a TRAF2/3 binding motif in LAT that we newly identified. TRAF3-mediated restraint of Dok1 activity and stability requires activation of Breast tumor kinase (Brk), the tyrosine kinase responsible for inhibitory phosphorylation of Dok1. The sustained activity of Dok1 in TRAF3-/- T cells is due to enhanced dephosphorylation of Brk at an activating tyrosine residue by the tyrosine phosphatase Protein tyrosine phosphatase 1B (PTP1B). The role of PTP1B in negative regulation of Brk activation and TCR/CD28 signaling was confirmed by pharmacological inhibition of PTP1B, which increased phosphorylation of Brk at an activating tyrosine residue and restored TCR/CD28-mediated Src family kinase activation in TRAF3-/- T cells. These results identify a novel mechanism of TRAF3-mediated regulation of TCR/CD28 signaling.
TCR/CD28 signaling promotes T cell metabolic reprogramming and cell fate decisions via activation of Mammalian target of rapamycin (mTOR). In Chapter 5, we hypothesized that both T cell metabolism and T cell fate decisions would be altered in TRAF3-/- T cells. Interestingly, TRAF3 deficiency impacted expression of molecules involved in amino acid uptake and catabolism, subsequently impairing TCR/CD28-mediated activation of mTOR. This is reflected by altered phosphorylation of p70S6K and Akt S473, targets of mTORC1 and mTORC2, respectively. As mTOR activity impacts T cell fate decisions, we examined CD4 T helper subset differentiation, finding that differentiation into TH1, TH2, and TH17 subsets was reduced in the absence of TRAF3. Consistent with defective TH2 differentiation, activation of STAT6 was reduced in CD3/CD28-stimulated TRAF3-/- T cells, although early IL-4R signaling was normal. These findings suggest that TRAF3 regulates TH2 differentiation via IL-4-independent mechanisms of GATA3 upregulation.
In Chapter 6, we tested the prediction that TRAF3 is a resident nuclear protein in T cells. Consistent with this prediction, TRAF3 was present in the nuclear fraction of resting human and mouse T cells. TRAF3 translocated to the nuclear fraction upon stimulation through CD3/CD28, and regulated the nuclear localization of TRAF2, as there was more TRAF2 in the nuclear fraction in resting and CD3/CD28-stimulated TRAF3-/- T cells. These findings suggest that nuclear TRAF3 promotes T cell function via regulation of TRAF2/cellular inhibitor of apoptosis protein (cIAP)-mediated degradation of target proteins. Taken together, the findings presented in this dissertation provide significant new insights into the mechanisms by which TRAF3 uniquely regulates T cell activation, fate, and function, and have implications for the manipulation of T cell function for therapeutic purposes.
Details
- Title: Subtitle
- Roles of TRAF3 in regulation of T lymphocyte biology
- Creators
- Tina Arkee
- Contributors
- Gail A Bishop (Advisor)John Colgan (Committee Member)Jon Houtman (Committee Member)Scott Lieberman (Committee Member)Fred Quelle (Committee Member)
- Resource Type
- Dissertation
- Degree Awarded
- Doctor of Philosophy (PhD), University of Iowa
- Degree in
- Immunology
- Date degree season
- Spring 2023
- DOI
- 10.25820/etd.007107
- Publisher
- University of Iowa
- Number of pages
- xviii, 176 pages
- Copyright
- Copyright 2023 Tina Arkee
- Language
- English
- Date approved
- 12/30/2021
- Description illustrations
- illustrations, graphs
- Description bibliographic
- Includes bibliographical references (pages 153-176).
- Public Abstract (ETD)
- T cells are an important component of the immune system, which normally detects and clears infections. When circulating immune cells called antigen-presenting cells (APCs) encounter microbes, they communicate their findings to T cells through interactions with the T cell antigen receptor (TCR) and its co-receptor CD28. This message is converted into a signal that is relayed throughout the cell, resulting in T cell activation. Activated T cells rapidly divide and produce molecules that directly and indirectly promote clearance of microbes. Relay of the initial signal to the nucleus of a T cell, which contains the genetic information required to make effector molecules, is facilitated by the signal transduction protein TRAF3. T cells lacking TRAF3 are not activated properly, and thus cannot effectively clear infections. The goal of this dissertation was to determine how TRAF3 promotes T cell activation, fate, and function. We found that TRAF3 associates with LAT, a signaling protein that integrates messages from TCR and CD28 and transmits them throughout the T cell. This association allows TRAF3 to regulate the protein Dok1, which is an inhibitor of TCR signaling. TRAF3 also promotes expression of a cellular amino acid transporter. This activates pathways that contribute to T cell differentiation into more specialized T cells. Finally, we found that TRAF3 is present in the T cell nucleus, where it may promote expression of genes important for T cell function. This work demonstrates that TRAF3 is an important regulator of T cell biology, and thus contributes to a balanced immune system.
- Academic Unit
- Immunology Graduate Program
- Record Identifier
- 9984428943702771
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