Download T cell receptors, T cell function and signaling

T and B cell activation and
signaling
Leslie Berg
S3-143B
6-8371
[email protected]
Activation of naïve lymphocytes
• T cell development in the thymus and B cell
development in the bone marrow produce
populations of naïve circulating lymphocytes
• Naïve T and B cells are cells that have not yet
encountered their antigen
• For any particular antigen (pathogen), the
frequency of naïve T and B cells is very low
• Naïve T and B cells do not produce effector
molecules
B cell activation, differentiation,
and antibody production
1. Antigen recognition and activation
2. Proliferation and new gene expression (differentiation)
3. Effector response to clear infection (antibody production)
T cell activation, proliferation, and
differentiation
1. Antigen recognition and activation
2. Proliferation and new gene expression (differentiation)
3. Effector response to clear infection
Two major subsets of effector T
cells
• Cytolytic T cells: naïve CD8+ T cells differentiate
into effector cytolytic T cells (CTLs) that can kill
infected target cells
• Helper T cells: naïve CD4+ T cells differentiate
into one of two types of effector cells
– TH1 cells that activate macrophages
– TH2 cells that help B cells produce antibodies
CD8+
CD4+
Summary of key points
• Naïve T and B cells are activated after
encounter with antigen, then proliferate and
differentiate into effector cells
• B cells differentiate into antibody-secreting
cells
• CD8+ T cells differentiate into effector CTLs,
CD4+ T cells into effector “helper” (TH2) or
“inflammatory” (TH1) T cells
Biochemical analysis of antigen
receptor complexes
• Lyse T or B cells in a mild detergent (e.g.,
digitonin)
– Retains protein-protein interactions more efficiently
• Two other polypeptides immunoprecipitate with
the BCR (membrane-bound IgM)
– Igα and Igβ
• Four other polypeptides immuno-precipitate with
the TCR αβ dimer
– CD3ε, δ, γ, and TCR ζ
BCR complex
TCR complex
TCR or BCR aggregation lead to
signaling
APC
T cell
TCR or BCR aggregation lead to
signaling
Can mimic with
Ab cross-linking
Secondary Ab
(or Streptavidin)
Anti-CD3 Ab
T cell
Y
Co-receptor complex
for B cells
• Consists of three chains
– CR2/CD21 is a complement receptor
– CD19 is the signaling component
– CD81 may help traffic complex to cell
surface
• Co-receptor is engaged when
antigen has complement proteins
attached to it
• Co-receptor is aggregated
together with BCR and increases
signaling
Co-receptor for T cells: CD4 or CD8
• CD4 is expressed on helper T cells
– CD4 binds to MHC class II
• CD8 is expressed on cytotoxic T cells
– CD8 binds to MHC class I
• CD4 and CD8 have two functions
– Adhesion (binding to MHC)
– Signaling
• Cytoplasmic tail associated with tyrosine kinase, p56lck
• CD4 >>> CD8
TCR - coreceptor MHC interaction
T cell
BCR - coreceptor Ag interaction
B cell
TCR
BCR
CD4
MHC
Class II
APC
C
’
Coreceptor
Summary of key points
• Membrane-bound IgM is in a complex with invariant chains
called Igα and Igβ, forming the BCR
• The TCR is in a complex with invariant transmembrane proteins
called CD3ε, δ, γ, and TCRζ
• Igα, Igβ, CD3ε, CD3δ, CD3γ, and TCRζ each contain ITAM motifs
• B cells express a co-receptor composed of CD21, CD18, and
CD81 that binds complement, increasing B cell signaling
• T cells also express a co-receptor, CD4 or CD8, that aids in
MHC/peptide recognition and provides signaling function
Naïve B and T cells need two
signals for activation
• B cell activation requires two independent
signals
– BCR binding to antigen
– “co-stimulatory” signal provided by either T cell help or
by TLR signaling
• Activation of a naïve T cell requires two
independent signals
– TCR binding to specific MHC/peptide complex
– “co-stimulatory” signal
• CD28 binding to B7 on the antigen-presenting-cell
• Ensures B and T cells only activated during an
infection
Two signals for B cell activation
1. Antigen
binding to
BCR
2. T cell
help via
CD40 and
cytokines
1. Antigen binding to BCR
2. TLR ligand binding to TLR
Only “professional” APCs express costimulatory signals for naive T cells
• Since naïve T cells need both TCR signals and
co-stimulatory signals for activation, only APCs
expressing B7 are able to activate a naïve T cell
• Three types of cells express B7
– Dendritic cells
– Activated macrophages
– Activated B cells
• All three cell types upregulate B7 after
encounter with pathogens
Activation of naïve B cells
• Resting naïve B cells express surface IgM
• B cells also express receptors to interact with T
cells and their products
– CD40
– Cytokine receptors
• Binding of antigen to IgM plus T cell help leads to
B cell activation, including antibody secretion
Activation of naïve
B cells
Activation of naïve T cells
• Resting naïve T cells express low affinity
receptors for interleukin-2 (IL-2)
• Activation of naïve T cells through TCR induces
transcription of cytokine and cytokine receptor
genes
– IL-2
– High affinity chain of IL-2 receptor
• IL-2 is a T cell growth and differentiation factor
that promotes T cell proliferation and the
production of “effector” molecules needed to
combat infection
Signal transduction pathways
leading to B cell and T cell
activation
• TCR and BCR complexes use nearly
identical signal transduction pathways
• Some proteins come in two flavors: a T
cell version and a B cell version
• Details below focus on the TCR, but
similar concepts and mechanisms apply
to B cell activation
Naïve T cell activation
• Leads to IL-2 secretion and synthesis of
high affinity IL-2 receptor
– Drives T cell proliferation and differentiation
• Biochemical signaling pathways well
worked out (using “sledgehammer” to
activate TCR)
– Aggregation of TCR using antibodies to
conserved CD3 proteins
T cell signaling
Stimulus
TCRαβ
TCR/CD3 complex
signaling
proteins
(CD3)
?
T cell signaling
Stimulus
TCR/CD3 complex
Tyrosine motifs
(ITAMs)
phosphorylated
mIgM
TCR
(T Cells)
αβ
δε
γε
ζ−ζ
-S-S-
-S-S-
Immune receptors
that contain the
ITAM: Immunoreceptor
Tyrosine-based
Activation Motif (ITAM;
YxxL/Ix6-8YxxL/I)
-S-S-
Igα
-S-S-
KIR2DS
β
γ
(NK Cells)
-S-S-S-S-
(also FcαR, FcγRII/III associate
with ζ-like subunits)
Igβ
α
FcεRI
(Mast Cells
and Basophils)
(B Cells)
DAP12
Big picture
• Goals of Ag receptor signaling on B
and T cells
–
–
–
–
Entry into cell cycle
Increase metabolism
Establish differentiation pathways
Elicit effector functions
transcription
TCR
actin polymerization
survival
metabolism
adhesion
TCR stimulation leading to gene
expression
• T cell is “activated”
– T cell secretes cytokines (growth
factors)
– T cell proliferates
– T cell differentiates into an effector cell
• Mechanism: TCR signal turns on
“latent” transcription factors
leading to new gene expression
TCR aggregation leads to tyrosine
phosphorylation of the TCR
CD4
Lck
TCR/CD3
ZAP-70
Phosphorylation of adapter proteins leading to formation
of signaling complex
Tyrosine phosphorylation of
receptor or adaptor proteins
recruits new substrates
TCR phosphorylation recruits Zap70
Adapter complex assembled
following TCR signaling
BCR phosphorylation recruits Syk
TCR and BCR signaling lead to PLC-γ
activation, and the production of two
critical second messengers
TCR signaling activates
Phospholipase C-γ (similar for
BCR)
Can mimic with PMA +
Ionomycin
Map-kinase
PKC +
Ras-GRP
Activate
pre-existing TFs
Summary of key points
• TCR and BCR signaling activate latent
transcription factors leading to new gene
expression
• TCR and BCR signaling activate tyrosine
kinases
• A key event downstream of the TCR and
BCR is the activation of PLCγ
• PLCγ cleaves PIP2 into two important
second messengers, DAG and IP3
TCR or BCR Signaling
• Activated Zap-70 or Syk phosphorylates
downstream targets
– Second messengers are generated, e.g., Ca+2, small
lipid co-factors
– Additional protein kinases and phosphatases are
activated
• Outcome is the activation of latent transcription
factors, leading to new gene transcription
• Examples of latent transcription factors
activated by TCR signaling: Elk-1, c-Jun, NF-kB,
NFAT
Mechanisms of transcription
factor activation
• Phosphorylation activates transcription factor
activity
– Elk, Jun family members
• Phosphorylation leads to dimerization and
nuclear transport
– STAT proteins (activated by IL-2R signaling)
• Degradation of inhibitory subunit
– NF-kB
• Dephosphorylation leads to nuclear transport
– NFAT
The MAP-kinases Erk and Jnk phosphorylate
transcription factors, thereby activating their
transcriptional activity
Cytokine receptor signaling activates Jak kinases
that phosphorylate the STAT transcription factors,
thereby inducing dimerization and nuclear
transport of the STAT dimers
TLR signaling induces degradation of IkB,
thereby releasing NF-kB to enter the
nucleus and activate transcription
Activation of NFAT
• The NFAT transcription factors are
phosphorylated and inactive in resting T and B
cells
• T cell activation leads to increased activity of a
protein phosphatase, Calcineurin
• Calcineurin dephosphorylates NFAT, leading to
NFAT migration into the nucleus
T cell
CD4
TCR/CD3
CD28
Zap-70
NFAT
Nucleus
Lck
IL-2 gene
Calcineurin
Ca+2
Summary of key points
• Latent transcription factors are activated by TCR or
BCR signaling and mediate new gene expression
• Mechanisms for activating latent transcription
factors include phosphorylation, dephosphorylation,
and degradation of an inhbitor
• This strategy allows cells to have the necessary
transcription factors present, but to keep them in an
inactive state until TCR or BCR signals occur