Transcript PowerPoint Presentation - How T cells recognize antigen

Signal Transduction
Transmission of external signals across the
membrane and conversion of these signals into
intracellular biochemical events
Peptide:MHC
TCR
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Changes in gene expression
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TCR signaling: Big Picture
RECEPTOR
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Alterations in
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Gene expression
General principle of signal transduction:
Ligand induced dimerization is the initiating step in signal transduction
(or clustering)
Example: Kit signaling
A kinase is an enzyme that
attaches phosphate groups
to specific amino acids of
proteins (Tyr, Ser, Thr)
General principle:
Phosphorylation is a
dynamic, reversible way of
modulating protein function
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How does the TCR transduce signals with such a little tail?
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TCR associates with the CD3 Complex
Analogous to the Ig
and Ig proteins that
associate with BCR on
B cells
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ITAM
ITAMs are the essential basis of TCR signal transduction
ITAM: Immune Tyrosine Activation Motif
ITAM structure is Y- X - X - I/L - X 6-8 - Y - X - X -I/L
When a tyrosine within an ITAM is phosphorylated, it
creates a binding site for other signaling molecules
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SH2 domains bind to specific
phosphotyrosines within ITAMs
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Protein Tyrosine Kinases in initial TCR signaling
Lck and Fyn - Src family kinases
Y
SH3
SH2
Y
Kinase
Regulatory Tyrosines
Zap-70 - Syk family kinase
Y
SH2
SH2
Kinase
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How do the ITAMs on CD3 get phosphorylated?
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Lck binds to the tails of CD4 and CD8
Lck
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Initiation of TCR signaling
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Initial events in TCR signaling
1.
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TCR + CD4 or CD8 bind peptide:MHC (clustering)
Lck and Fyn phosphorylate ITAMs on CD3 complex
ZAP-70 binds to P-ITAMs on CD3zeta chains
Lck and Fyn phosphorylate and activate ZAP-70
ZAP-70 initiates downstream signaling events…
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Initial TCR signaling
RECEPTOR
Protein tyrosine kinases
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Regulation of early TCR signaling
Lck or Fyn
Phosphorylation is a
dynamic, reversible way of
modulating protein function
• Activating and inhibitory Tyrphosphorylation sites
• CD45 (B220) is a phosphatase
which activates PTKs by
dephosphorylating inhibitory P-Tyr.
• Csk is a kinase which inhibits PTKs
by phosphorylating inhibitory Tyr
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The phosphatase CD45 regulates PTK activity
CD45 removes inhibitory phosphates from Lck and Fyn
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Regulation of early TCR signaling
• The plasma membrane is not homogeneous
• Lipid rafts are enriched for saturated lipids and cholesterol making
them more rigid
• This composition regulates which types of proteins can localize to
these rafts
-most transmembrane proteins (e.g. TCR) are excluded
-Lck and Fyn are enriched in lipid rafts
TCR
Fyn
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Lipid rafts help regulate T cell activation
If rafts are disrupted (by depleting cholesterol), T cell activation is blocked
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Downstream TCR signaling
RECEPTOR
Protein tyrosine kinases
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ZAP-70 phosphorylates SLP-76 and LAT which leads to
activation of 3 downstream signaling pathways
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2
3
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TCR signaling: Big Picture
RECEPTOR
Protein tyrosine kinases
Phospholipase C
Intracellular Ca2+
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Protein kinase C
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Transcription factors
Ras (G-proteins)
MAP kinases
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Alterations in
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Gene expression
Zap-70 phosphorylates PLC,
activating it to cleave PIP2
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IP3 binds to Ca2+ channels and opens them
2nd messengers amplify initial signals
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TCR signaling: Big Picture
RECEPTOR
Protein tyrosine kinase
Phospholipase C
Intracellular Ca2+
1
Ras (G-proteins)
Protein kinase C
2
NFAT
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MAP kinases
3
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Alterations in
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Gene expression
Immunosuppressive drugs block NF-AT activation
FK506 and cyclosporin block
NF-AT activation by inhibiting
calcineurin
Essential drugs to prevent
transplant rejection
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Cyclosporin
Tolypocladium inflatum, the white mold from Norway
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TCR signaling: Big Picture
RECEPTOR
Protein tyrosine kinase
Phospholipase C
Intracellular Ca2+
1
Ras (G-proteins)
Protein kinase C
2
NFAT
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MAP kinases
3
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Alterations in
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Gene expression
IP3 binds to Ca2+ channels and opens them
2nd messengers amplify initial signals
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Activating NF-kB
Ca++
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TCR signaling: Big Picture
RECEPTOR
Protein tyrosine kinase
Phospholipase C
Intracellular Ca2+
1
Ras (G-proteins)
Protein kinase C
2
NFAT NF-kB
MAP kinases
3
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Alterations in
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Gene expression
ZAP-70 phosphorylates SLP-76 and LAT which leads to
activation of 3 downstream signaling pathways
1
2
3
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Ras is a small G-protein
GTPase
activating
protein
GEF
Guanine nucleotide
exchange factor
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Ras activates the MAP kinase pathway
TCR activation
Raf (MAP kinase kinase kinase)
MEK (MAP kinase kinase)
MAP kinase
AP-1 (Transcription factor)
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RECEPTOR
Protein tyrosine kinase
Phospholipase C
Intracellular Ca2+
NF-AT
Ras (G-proteins)
Protein kinase C
NF-kB
AP-1
MAP kinases
The IL-2 gene
promoter
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Example of T cell activation
TCR binds peptide:MHC
Induces expression of genes
T cells divides many times
creating more T cells with
the same TCR
T cells differentiate
and acquire new
characteristics
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IL-2 is one of hundreds of genes induced by TCR signaling
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Mysteries in T cell antigen recognition
• Affinity of TCR for peptide:MHC is quite low
• How is T cell-APC interaction maintained?
• How are low frequency peptide:MHC
complexes ever recognized?
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Initial T cell/APC contact is
mediated by adhesion molecules
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Clustering leads to formation
of an “immunological synapse”
B-Cell
(Peptide-pulsed)
Qui ckTime™ and a Vi deo decompressor are needed to see this pictur e.
T-Cell
minutes in 20 seconds
of video cluster at
TCR, MHC & 8adhesion
molecules
interface between T cell and APC
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Many additional receptor interactions contribute
to T cell activation
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Summary of TCR signaling
• TCR signaling is initiated by TCR binding peptide:MHC and
clustering of TCR with CD4/CD8 and other accessory molecules.
• Clustering leads to activation of Src family protein tyrosine kinases
(Lck and Fyn) that phosphorylate ITAMs of the CD3 complex
• In naïve T cells, the TCR is excluded from lipid rafts. TCR activation
allows the TCR to enter lipid rafts and associate with PTKs.
• T cell signaling activates 3 general downstream pathways:
PLC - Ca2+ - NFAT
PLC - PKC - NFkB
Ras - MAPKs - AP-1
• Coreceptors and adhesion molecules contribute to T cell:APC
interactions. These molecules are organized into an immunological
synapse which facilitates prolonged T cell signaling.
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