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Proximal Tyrosine Kinases That Initiate T Cell Activation 10
Proximal Tyrosine Kinases That Initiate T Cell Activation 10
TCR signalling
The TCR is a multi-subunit complex that initiates an intricate series of signalling events. In a resting state, the LAT and SLP76, which have a central role in nucleating the recruitment of downstream effector molecules.
ITK SRC family kinase LCK is usually associated with the co-receptor CD4 or CD8. Following engagement of During this process, a third protein tyrosine kinase, ITK, is recruited to a LAT-containing multimolecular CSK
ITK is held in an inactive conformation by an SH2–SH3 the TCR and a co-receptor by a peptide-bound MHC molecule, LCK phosphorylates ITAMs present in the signalling complex at the plasma membrane. ITK phosphorylates and activates PLCγ1, inducing the production When anchored to the plasma membrane by association
domain interaction. Following TCR stimulation, the cytoplasmic regions of the CD3 chains and TCR ζ-chains. ZAP70 then binds to doubly phosphorylated ITAMs of the crucial second messengers InsP3 and DAG, which lead to an increase in cytoplasmic Ca2+ concentration with PAG, CSK is active owing to the conformation of both
PH domain of ITK binds PtdIns(3,4,5)P3, the result of resulting in a conformational change that allows for its phosphorylation by LCK. Phosphorylation of ZAP70 and to the activation of RAS and PKCs, respectively. These events activate further downstream signalling linker regions with respect to the kinase domain and an
the activation of PI3K, inducing plasma membrane promotes its stabilization in an active conformation. Activated ZAP70 phosphorylates the adaptor molecules pathways leading to T cell activation. SH2–kinase domain interaction. TCR stimulation induces
localization of ITK. InsP4, generated by the dephosphorylation of PAG through unknown
action of InsP3 kinase B, augments this mechanisms, resulting in the release and
PtdIns(3,4,5)P3 interaction. Once at +6- %5- inactivation of CSK. Importantly, PAG-
the membrane, ITK is activated by MHC CD4 deficient mice have no appreciable
LCK-mediated phosphorylation on PtdIns(3,4,5)P3 PH PAG defect in CSK function indicating
N SH3 P
Tyr511. This is then followed by InsP4 PP Peptide that functionally redundant molecules
ITK trans- N PTPN22
trans-autophosphorylation on N PH SH3 Tyr180 CD45 P are likely to exist. Interestingly,
autophosphorylation CD3 α β TCR
Tyr180, which induces the opening SH2 SH3 the PTPN22 (also known as LYP)
of ITK and brings the SH2 domain SH2 TCR stimulation SH2 SH3 N cytoplasmic phosphatase contains a
P PtdIns(3,4,5)P3
and PI3K activation Kinase TCR-induced P SH2
into contact with the kinase domain, PP proline-rich sequence that interacts
Kinase C dephosphorylation
allowing the SH2 domain to bind Pro287
Tyr511 P P ITAM LCK of PAG
with the SH3 domain of CSK. PTPN22
C SLP76 P P PAG P
phosphorylated SLP76. ITK is also SH2 SH2 LAT PP P
P P P dephosphorylates the activation loop
Phosphorylation InsP4 Kinase Kinase
regulated by prolyl isomerization at Cis Trans by LCK ITK P PLCγ1 PP
P P P P P tyrosine of SRC family kinases. Thus,
P P
Pro287 in the SH2 domain: the trans Inactive γε ε δ
P P
C CSK and PTPN22 act in tandem to
conformation has a greater affinity P P Active P
P P P P negatively regulate SRC family
SH2 SH2 P Inactive C Active
for phosphorylated tyrosine residues P
P
P P
ZAP70 PTPN22 kinases. The interaction of PTPN22
P P P ζ ζ
compared with the cis conformation. Prolyl isomerization with CSK is disrupted in the allele
P P of the phosphatase associated
SLP76 P
with autoimmunity.
DAG InsP3 P
P
Experimental inhibitors of tyrosine kinases P CSK
RAS-
Inhibitor PKCs Ca2+
GRP GADS P
RAS
Inhibitor Inhibitor P
T cell activation
ATP analogue inhibitor Allosteric inhibitor stabilizing the Competitive inhibitor RNA interference-mediated
of the ATP-binding pocket kinase domain in the inactive state of binding partners knockdown of kinase expression <#2 .%-
Most current inhibitors target the ATP-binding region of the kinase domain; however this strategy poses many limitations owing
to the conserved catalytic mechanism of all tyrosine kinases. In light of more recent structural data, allosteric inhibitors are now Phosphorylation N N N Recruitment of proline-
by LCK Dephosphorylation TCR-induced trans- containing substrate
being designed to stabilize the inactive conformation of the kinase. Also, some inhibitors are being designed to inhibit the SH3
by CD45 autophosphorylation
recruitment of the kinase to its proper location by blocking its association with binding partners. More recently, RNA Tyr315 P P Tyr319 SH3 SH3
interference-mediated knockdown of kinase expression has shown some promise, although this approach is still in its infancy. Kinase Phosphorylation Kinase Dephosphorylation P Tyr394
P P by CSK by PTPN22 (and/or CD45) SH2
SH2 SH2 P SH2
ITAM SH2 Kinase
P Kinase P Tyr505 Recruitment of
Tyrosine kinase Human mutation Mouse knockout P LCK or ZAP70 trans- C
SH2 SH2 C phosphotyrosine-
Kinase Tyr493 autophosphorylation
LCK • CD4+ T cell lymphopenia • Decreased number of DP and SP thymocytes, as well as peripheral Inactive Primed containing substrate C
• Diminished in vitro response to mitogens T cells N
C N
• Generally preserved early TCR signalling • Decreased TCR signalling and responses TCR ζ-chain Active
• Complete block at DN thymocyte stage when combined with C
Fyn knockout
ZAP70 • Absence of peripheral CD8+ T cells • Arrest at DP to SP thymocyte stage affecting both CD4+ and CD8+
Inactive Active LCK
• CD4+ T cells present but have a TCR signalling defect T cells When LCK is phosphorylated on Tyr505, intramolecular interactions (between the SH2 domain and phosphorylated
(compensation by SYK, expressed by thymocytes but not • Complete block at DN thymocyte stage when combined with Tyr505 and between the SH3 domain and interdomain) hold LCK in a closed, inactive conformation. Dephosphorylation
peripheral CD4+ T cells) Syk knockout ZAP70 of Tyr505 by CD45 ‘unlocks’ LCK into a primed state. Trans-autophosphorylation of Tyr394, which can result from
ITK • Immunodeficiency syndrome • Mild immunodeficiency owing to redundancy with other TEC family
• Fatal inadequate response to EBV kinases Binding of the tandem SH2 domains of ZAP70 to a doubly phosphorylated ITAM clustering of co-receptors or decreased CSK function, results in full activation of LCK. LCK is inactivated by
• Similar phenotype to XLP caused by defects in SAP or XIAP • Population of non-conventional, innate-like CD8+ T cells (Yxx(L/I)x7–8Yxx(L/I)) in the cytoplasmic regions of the CD3 chains or TCR ζ-chains induces dephosphorylation of Tyr394 by the phosphatase PTPN22 (and/or CD45) and phosphorylation of Tyr505 by CSK.
CSK • No known human mutations • Early embryonic lethal phenotype a conformational change in ZAP70 that increases activity of the kinase and results in The phosphorylation status of LCK is probably a dynamically controlled process subject to relocalization of CD45
• PTPN22 polymorphism that decreases association with CSK • Depletion in early immature thymocytes bypasses need for pre-TCR accessibility of Tyr315 and Tyr319 for phosphorylation by LCK. Activation of ZAP70 is further and/or CSK from their substrate LCK. Note that in some cases the related SRC family kinase FYN can substitute for
predisposes to autoimmunity and TCR signalling as a result of increased basal LCK and FYN activity potentiated by Tyr493 phosphorylation by LCK or by ZAP70 trans-autophosphorylation. LCK function or may be activated by LCK to promote downstream substrate phosphorylation.