CD4 T cell affinity evolution under acute and chronic antigen exposure 公开

Abstract

T cell receptor (TCR) interaction with pMHC on the respective cell surfaces of a T cell and an antigen-presenting cell can elicit a multitude of functional differentiation pathways or transpire without an outcome. The result hinges on affinity and other biophysical interaction parameters that govern TCR:pMHC binding and consequently modulate the degree of the T cell response. It is unclear what TCR affinities are represented within an antigen-specific polyclonal CD4 T cell population, and if TCR affinity diversity differs in the context of acute and chronic infections. Relative to in-solution measurements with recombinant proteins, single cell in-situ analysis of TCR affinity for membrane-anchored pMHC provides a more relevant assessment of this interaction and better correlates with a T cells’ functional response. Here, using the 2-dimensional micropipette adhesion frequency assay (2D-MP), we measured the range and average TCR affinity of polyclonal IA^b GP66-77 specific CD4 T cells in the B6 acute (Armstrong) and chronic (CL13) models of LCMV infection. In both responses, the presence of antigen maintained an effector population with a high average TCR affinity that later declined at memory with antigen clearance. Greater than 1000-fold affinity ranges were measured in both responses and population average 2D affinity at peak and late time points were equivalent between the two infections. These results indicate that intrinsic and extrinsic factors regulate T cell function in chronic infections without altering TCR affinity diversity. To further elucidate the effects of antigen stimulation on 2D affinity and TCR:pMHC bond lifetime, we used a reductionist approach, tracking activation-induced changes in IA^b GP66-77 specific SMARTA transgenic CD4 T cells. In vitro, peptide-stimulated cells exhibited dynamic TCR:pMHC 2D affinities and bond lifetimes that decreased from naïve levels within the first 24 hours of antigen encounter but recovered at the 48-hour mark with the start of cell division. This occurred independently of antigen dose and degree of activation-induced TCR downregulation. Hence, 2D analysis of monoclonal and polyclonal T cells’ affinities can represent the sum total of single cell TCR:pMHC interaction dynamics within the respective cellular environments. 

Table of Contents

Chapter 1. 

Introduction ………………………………………......…...…... 1 

Chapter 2. CD4 T cell affinity diversity is equally maintained during acute and 

chronic infection 

Abstract …………………………………………..........……… 31 

Introduction ……………………………………...........…….… 32 

Materials and Methods ….……………………...........……...… 36 

Results ………………………………………………............… 43 

Discussion ……………………………………...……...........… 53 

Figure 1 ……………………………………………..........…… 60 

Figure 2 ……………………………………………..........…… 62 

Figure 3 …………………………………………..........……… 64 

Figure 4 …………………………………………..........……… 65 

Figure 5 ………………………………………..........………… 67 

Figure 6 ………………………………………..........………… 68 

Supplemental Figure 1 ...……………………………………… 70 

Supplemental Figure 2 ...……………………………………… 71 

Supplemental Figure 3 ...……………………………………… 73 

Chapter 3. Activation induced decrease and recovery of TCR 2D affinity and bond lifetime 

Abstract …………………………………………….........…… 74 

Introduction ……………………………………...…………… 75 

Materials and Methods ….…………………………………… 79 

Results …………………………………………..........……… 84 

Discussion ………………………………………………....… 89 

Figure 1 ………………………………………………........… 93 

Figure 2 ……………………………………………........…… 95 

Figure 3 ……………………………………………........…… 97 

Figure 4 ………………………………………………........… 98 

Supplemental Figure 1 ...…………………………………..… 99 

Chapter 4. Discussion ……………………………………… 100 

Figure 1 ..……………………………………………......…… 109 

References ………..………………………………………… 111 

About this Dissertation

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School	Laney Graduate School
Department	Biological and Biomedical Sciences
Degree	Ph.D.
Submission	Dissertation
Language	English
Research Field	Health Sciences, Immunology
关键词	2D Affinity micropipette Avidity CD4 T cell Polyclonal Acute Viral Infection Chronic Viral Infection LCMV clone 13 Affinity TCR Affinity Armstrong 2-dimensional micropipette adhesion frequency assay Tetramer
Committee Chair / Thesis Advisor	Evavold, Brian, Emory University
Committee Members	Ford, Mandy, Emory University Kohlmeier, Jacob, Emory University Rengarajan, Jyothi, Emory University Altman, John, Emory University

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