Date

June 2009

Document Type

Dissertation

Degree Name

Ph.D.

Department

Dept. of Molecular Microbiology and Immunology

Institution

Oregon Health & Science University

Abstract

The immunological synapse (IS) is a large-scale rearrangement of molecules at the interface between T cells and antigen presenting cells (APCs). The structure of the ‘mature’ IS is well defined, with a ring of adhesion molecules surrounding a centralized accumulation of T cell receptor (TCR)-peptide major histocompatibility complex (pMHC) interactions. However, there is evidence that the structure of the IS differs depending on experimental conditions and the differentiation state of the cells used. Given that IS are hypothesized to play a crucial role in the delivery of effector molecules, further studies are needed to determine the relationship between the phenotype of effector T cells and IS structure. The aim of my thesis research was to test the hypothesis that CD4 T cell subsets possessing specialized effector functions form IS with distinct structures. To achieve this goal, I carried out experiments designed to examine the IS formed by Th1 and Th2 cells interacting with transfected fibroblast APCs and supported planar bilayers, and induced T regulatory cells (iTregs) interacting with bilayers and dendritic cells (DCs). I provide evidence that Th1 and Th2 cells form morphologically distinct IS. Th1 cells form IS with adhesion molecules surrounding TCR-pMHC interactions, while Th2 cells form multifocal IS with adhesion molecules interspersed among multiple small accumulations of TCR-pMHC. There are also differences in the distribution of the phosphatase CD45 and phosphorylated signaling molecules in Th1 and Th2 IS. I hypothesize that these structural differences are related to differences in the effector molecules secreted by Th1 and Th2 cells, specifically that the mature IS formed by Th1 cells act as gaskets for the directed secretion of cytotoxic molecules and CD40 ligand. I also show that iTregs form either immunological kinapses or stable IS depending on the level of CD80 on the surface of APCs. Induced Tregs are capable of downmodulating the costimulatory ligand CD80 on the surface of DCs in an antigen specific manner. On the same timescale as CD80 downmodulation, iTregs transition from stable to motile contacts with DCs. Experiments utilizing supported planar bilayers containing pMHC, ICAM-1 and CD80 showed that iTregs form motile immunological kinapses with a highly polarized cell shape in the presence of low levels of CD80. However, iTregs arrest and form stable IS when interacting with bilayers containing high levels of CD80. Consistent with CD80 modulating the stop signals iTregs receive upon antigen recognition, intracellular calcium flux is more sustained in the presence of high levels of CD80. I hypothesize that mature IS formation is required for downmodulation of costimulatory molecules by iTregs.

Identifier

doi:10.6083/M4RV0KP8

School

School of Medicine

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