Premed · Premed · Immunology
Lecture 15: T Cell Activation and Co-stimulation
Immunology
Learning Objectives
By the end of this lecture, students will be able to:
- Describe the three signals required for full T cell activation
- Explain the role of the TCR complex and co-receptors (CD4, CD8) in antigen recognition
- Describe the major co-stimulatory and co-inhibitory pathways
- Outline the key signaling cascades downstream of TCR engagement
- Explain the consequences of T cell activation without co-stimulation (anergy)
Lecture Content
I. The Three-Signal Model of T Cell Activation
Full T cell activation requires the convergence of three distinct signals. Signal 1 (antigen recognition) is provided when the TCR binds a peptide-MHC complex on the surface of an antigen-presenting cell. Signal 2 (co-stimulation) occurs when co-stimulatory molecules on the APC engage their receptors on the T cell. Signal 3 (cytokine signaling) is delivered by cytokines from the APC and surrounding environment that direct T cell differentiation into specific effector subsets. Signal 1 alone, without the co-stimulatory support of Signal 2, drives the T cell into a state of anergy -- functional unresponsiveness to subsequent stimulation. Only when all three signals are provided together does the T cell undergo full activation, proliferation, and differentiation into an effector cell.
II. Signal 1: TCR-Peptide/MHC Interaction
The TCR is a heterodimer consisting of alpha and beta chains in the vast majority of T cells (approximately 95 percent bear alpha-beta TCRs), while a smaller population uses gamma and delta chains. Each chain has a variable (V) domain and a constant (C) domain. The complementarity-determining regions (CDRs), especially CDR3, make direct contact with the peptide-MHC complex: CDR1 and CDR2 of the alpha-beta TCR contact the MHC helices, while CDR3 contacts the peptide itself.
The TCR itself has no intrinsic signaling capacity and relies on the CD3 complex to transduce activation signals. The TCR associates with CD3-gamma, CD3-delta, and CD3-epsilon chains (each containing one ITAM) and a CD3-zeta-zeta homodimer (each zeta chain containing three ITAMs), for a total of 10 immunoreceptor tyrosine-based activation motifs (ITAMs) per TCR-CD3 complex. These ITAMs become phosphorylated upon TCR engagement, initiating the downstream signaling cascade.
The co-receptors CD4 and CD8 play dual roles: they stabilize the TCR-peptide-MHC interaction and recruit the tyrosine kinase Lck. CD4 binds the beta-2 domain of MHC class II and is expressed on helper T cells, while CD8 (usually an alpha-beta heterodimer) binds the alpha-3 domain of MHC class I and is expressed on cytotoxic T cells. Both co-receptors associate with Lck on their cytoplasmic tails, and by bringing Lck into close proximity with the CD3 ITAMs, they are essential for efficient signaling.
III. TCR Signaling Cascade
When the TCR engages its cognate peptide-MHC complex and the co-receptor binds, a carefully orchestrated signaling cascade unfolds. Lck, associated with CD4 or CD8, phosphorylates the ITAMs on CD3-zeta and CD3-epsilon. Phosphorylated ITAMs then recruit ZAP-70 (zeta-chain associated protein kinase 70), which is itself phosphorylated and activated by Lck. Activated ZAP-70 phosphorylates the scaffold proteins LAT (linker for activation of T cells) and SLP-76, which serve as platforms for the assembly of three major downstream signaling pathways.
The PLC-gamma-1 pathway is particularly important. PLC-gamma-1 cleaves PIP2 into DAG and IP3. DAG activates PKC-theta, which triggers the NF-kappa-B pathway (promoting survival and cytokine gene expression), and also activates RasGRP, feeding into the Ras-Raf-MEK-ERK (MAPK) cascade that leads to the transcription factor Fos (part of AP-1). IP3, meanwhile, triggers calcium release from the ER, which activates calmodulin and then calcineurin; calcineurin dephosphorylates NFAT, allowing it to translocate into the nucleus. The Ras/MAPK pathway converges on the AP-1 transcription factor (Fos/Jun), and the PI3K pathway activates Akt, promoting cell survival and metabolic reprogramming.
Three key transcription factors are therefore activated: NFAT (the target of the immunosuppressive drugs cyclosporine and tacrolimus, which inhibit calcineurin), NF-kappa-B (driving survival genes and cytokines), and AP-1 (promoting proliferation and cytokine expression). All three converge on the IL-2 promoter, driving the production of IL-2, which acts in an autocrine and paracrine fashion to stimulate T cell proliferation.
<image>A detailed signaling diagram of the TCR signaling cascade. At the top, a T cell TCR-CD3 complex engages a peptide-MHC complex on an APC, with CD4 (or CD8) binding the MHC molecule and bringing Lck to the ITAMs. Step 1: Lck phosphorylates CD3 zeta ITAMs. Step 2: ZAP-70 is recruited to phospho-ITAMs and activated by Lck. Step 3: ZAP-70 phosphorylates LAT and SLP-76 scaffold proteins. Three pathways branch from LAT/SLP-76: (A) PLCgamma1 → PIP2 cleavage into IP3 (→ Ca2+ release → calcineurin → NFAT nuclear translocation) and DAG (→ PKC-theta → NF-kB; → RasGRP → Ras-MAPK → AP-1). (B) PI3K → Akt → survival. All three transcription factors (NFAT, NF-kB, AP-1) converge on the IL-2 gene promoter in the nucleus, leading to IL-2 transcription. Cyclosporine and tacrolimus are shown blocking calcineurin, preventing NFAT activation.</image>
IV. Signal 2: Co-stimulation
The principal co-stimulatory pathway is the CD28-B7 pathway. CD28 on naive T cells binds B7-1 (CD80) and B7-2 (CD86) on APCs. B7-2 is constitutively expressed at low levels, while B7-1 is upregulated upon APC activation. CD28 signaling enhances IL-2 production by stabilizing IL-2 mRNA, promotes T cell survival by upregulating Bcl-xL, enhances glucose metabolism through mTOR activation, and lowers the threshold of TCR signaling needed for activation. Without CD28 co-stimulation, the T cell becomes anergic -- functionally unresponsive to subsequent stimulation.
CTLA-4 (CD152) is the major co-inhibitory counterpart to CD28. Although structurally homologous to CD28, CTLA-4 binds B7-1 and B7-2 with much higher affinity. It is upregulated on T cells after activation and is constitutively expressed on Tregs. CTLA-4 suppresses T cell responses by competing with CD28 for B7 binding, delivering inhibitory signals through the recruitment of phosphatases PP2A and SHP-2, and performing trans-endocytosis -- physically capturing and removing B7 molecules from the APC surface. The importance of CTLA-4 is underscored by the fact that CTLA-4 knockout mice develop fatal lymphoproliferation. In clinical oncology, ipilimumab (anti-CTLA-4) is used to block this checkpoint and enhance antitumor immunity.
The ICOS-ICOSL pathway provides co-stimulation to activated T cells through the inducible co-stimulator (ICOS), which binds ICOSL on APCs and non-immune cells. This pathway is particularly important for T follicular helper (Tfh) cell function, germinal center reactions, and antibody responses, and it promotes IL-21, IL-4, and IL-10 production.
The PD-1-PD-L1/PD-L2 pathway is another major co-inhibitory axis. PD-1, expressed on activated T cells, engages PD-L1 (broadly expressed on many cell types) or PD-L2 (primarily on APCs). PD-1 signaling recruits SHP-2 phosphatase, which dephosphorylates TCR signaling molecules, inhibiting T cell activation and effector function. Chronic PD-1 engagement promotes T cell exhaustion in the settings of chronic infection and cancer, making PD-1 and PD-L1 important therapeutic targets: nivolumab and pembrolizumab block PD-1, while atezolizumab and durvalumab block PD-L1.
Several additional co-stimulatory receptors of the TNF receptor family further modulate T cell responses. OX40 (CD134) promotes T cell survival and memory. 4-1BB (CD137) enhances T cell proliferation and survival and is incorporated into CAR-T cell constructs to improve persistence. The CD40L (CD154)-CD40 interaction between T cells and APCs provides bidirectional activation and is essential for isotype switching and DC licensing.
V. Signal 3: Cytokine-Directed Differentiation
The cytokines produced by APCs and the surrounding microenvironment during T cell activation determine which effector subset the T cell will become. IL-12 combined with IFN-gamma drives Th1 differentiation (targeting intracellular pathogens). IL-4 drives Th2 differentiation (addressing parasites and allergy). TGF-beta combined with IL-6 promotes Th17 development (for extracellular bacteria and fungi), while TGF-beta combined with IL-2 favors Treg induction (for immune regulation). IL-6 with IL-21 supports Tfh cell development (providing B cell help in germinal centers). These subset-specific differentiation pathways are detailed further in Lecture 16.
VI. The Immunological Synapse
When a T cell engages an APC, the interface between the two cells becomes organized into a highly structured contact zone known as the immunological synapse. The central supramolecular activation cluster (cSMAC) contains the TCR-peptide-MHC complexes, CD28-B7 pairs, and PKC-theta. Surrounding it, the peripheral SMAC (pSMAC) forms an adhesion ring composed of LFA-1 bound to ICAM-1. The outermost distal SMAC (dSMAC) contains CD45 (a tyrosine phosphatase) and is enriched in actin. Sustained signaling through this synapse for several hours is required for full T cell activation. The synapse also allows directed secretion of cytokines and, in the case of CTLs, lytic granules, precisely toward the APC or target cell.
VII. T Cell Anergy
Anergy results when a T cell receives Signal 1 (TCR engagement) without Signal 2 (co-stimulation). Anergic T cells remain alive but are functionally unresponsive. At the molecular level, anergy is maintained by a failure to produce IL-2, upregulation of E3 ubiquitin ligases (GRAIL, Cbl-b, and Itch) that degrade key signaling molecules, and epigenetic silencing of the IL-2 locus. Anergy serves as an important peripheral tolerance mechanism, preventing autoimmunity when self-antigens are presented by non-professional APCs that lack B7 co-stimulatory molecules.
<image>A diagram of the immunological synapse between a T cell and an APC, viewed both from the side and from the T cell face (en face view). Side view: T cell and APC are closely apposed with a narrow cleft. Molecular interactions are shown: TCR-pMHC and CD28-B7 in the center, LFA-1-ICAM-1 forming a ring, CD4 or CD8 bridging to MHC. En face view (looking at the T cell contact surface): concentric ring structure -- central SMAC (cSMAC, yellow) containing clustered TCR-CD3, CD28, PKC-theta; peripheral SMAC (pSMAC, green ring) containing LFA-1-ICAM-1 adhesion molecules and talin; distal SMAC (dSMAC, outer red ring) containing CD45 phosphatase and F-actin. Arrows indicate directed cytokine secretion through the synapse toward the APC.</image>
