Page 2111 - Hematology_ Basic Principles and Practice ( PDFDrive )

1874 Part XII Hemostasis and Thrombosis

Heterotrimeric G Proteins Are Early Response PI-4P and PI-4,5P 2 to yield PI-3,4P 2 and PI-3,4,5P 3, respectively.
Inner membrane leaflet PIP 3 functions in signaling by interacting
Elements for Most Soluble Platelet Agonists 24,28 with Akt via binding motifs termed pleckstrin homology domains
(PH), thereby activating Akt to function as a serine/threonine kinase.
The GPCRs for soluble platelet agonists are constitutively associated
at the inner plasma membrane with specific heterotrimeric G proteins Serine/Threonine Kinases Regulate the Activity of
(consisting of α, β, and γ subunits) of the families G q, G 12/13 , and G i
(Table 125.2A). The receptor for the potent platelet inhibitory Other Proteins and Enable the Development of
molecule, prostaglandin I 2 (PGI 2; prostacyclin) is also a GPCR and Signaling Complexes 31,33
is coupled to G s.
In the basal, resting state, GDP is bound to the α subunit of the PKC is a central protein kinase in the PLC signaling pathway that,
2+
G protein. Upon ligand binding, a GPCR acts as a guanine nucleotide upon activation by DAG and Ca , phosphorylates the serine and
exchange factor, promoting exchange of GDP for GTP on different threonine residues of many platelet proteins, including pleckstrin.
classes of the α subunit. Sites on the α and βγ subunits are thereby The serine/threonine kinase Akt is an effector of PI3K, activating
exposed, allowing for activation of effector molecules including the Rap1B. PKC is important in platelet secretion and both PKC and
β isoform of phospholipase C (PLC-β), the γ isoform of phosphati- Akt are involved in αIIbβ3 activation, resulting in aggregation.
dylinositol 3-kinase (PI3K), and a Rho-specific guanine nucleotide
exchange factor, p115-RhoGEF (Table 125.2B). Adenylate cyclase is
activated via G s and inhibited by G i . Intrinsic GTPase activity of the Cytosolic Ca 2+28,34
α subunit hydrolyzes the bound GTP to GDP, thus restoring the G
2+
2+
protein to its resting conformation. An increase in the intracellular Ca concentration ([Ca ] i) is a key
2+
event that triggers platelet activation and aggregation; Ca -dependent
Protein Tyrosine Kinases Modulate Enzyme Activity responses include shape change, secretion, procoagulant surface
2+
and Allow for the Formation of Signaling exposure, TxA 2 formation, and αIIbβ3 activation. The basal [Ca ] i
is maintained at approximately 0.1 µM and can increase to greater
Complexes 28–30 than 1 µM with strong stimulation. The initial increase results from
2+
IP 3 -mediated release of Ca from the dense tubular system (DTS).
2+
2+
GPVI is the most potent signaling collagen receptor. Clustering of Upon depletion of this Ca pool, store-operated Ca (SOC) influx
GPVI upon binding to collagen results in phosphorylation of its occurs from the platelet exterior. Stromal interaction molecule 1
constitutively-associated FcRγ by tyrosine kinases of the Src family (STIM1), a protein in the DTS membrane, undergoes a conforma-
2+
(Fyn and Lyn) that are associated with the cytoplasmic domain of tional change when DTS Ca is depleted, allowing it to bind and
2+
2+
GPVI. Phosphorylation of tandem ITAM motifs enables binding of activate Orai1, the major SOC Ca -selective release-activated Ca
Syk via Src homology 2 (SH2) domains. Activation of the tyrosine (CRAC) channel in the platelet plasma membrane.
kinase Syk leads to the phosphorylation of the adaptor protein LAT,
which serves as the platform for the assembly of a signalosome—a
complex of multiple signaling enzymes and adaptor molecules (e.g., Phospholipase A 2 Is Responsible for the Synthesis
SLP 76, Btk, Gads, Gab1) within membrane lipid rafts. The signalo- of TxA 2 28
some serves as a membrane scaffold for phosphorylation/activation
2+
of effectors including phospholipase C-γ (PLC-γ) and the α, β, and An increase in [Ca ] i is one of the main triggers for activation of
δ isoforms of PI3K. cytosolic phospholipase A 2 (cPLA2), resulting in the formation of the
The platelet receptor platelet-endothelial cell adhesion molecule proaggregatory prostanoid TxA 2 . cPLA2 hydrolyzes the polyunsatu-
(PECAM)1 negatively regulates GPVI signaling by recruiting the rated fatty acid arachidonate from the C2 position of inner plasma
tyrosine phosphatase SHP2, and reducing LAT signalosome assembly. membrane leaflet phospholipids, and arachidonate is converted to
The signaling pathway initiated by binding of GPIb–IX–V to the cyclic endoperoxide intermediates prostaglandin (PG)G 2 and
VWF is similar to that of GPVI in that both involve activation of PGH 2 by cyclooxygenase (COX)-1. Thromboxane synthase in the
PLC-γ via FcRγ, Fyn and Lyn, and Syk. Binding of platelets to platelet cytosol metabolizes PGH 2 to TxA 2 , which diffuses out of
podoplanin via CLEC2 also induces signaling events similar to those the platelet and is available to activate additional platelets via the TP
initiated through GPVI. However, rather than (tandem) ITAM receptor.
phosphorylation, it involves hemi-ITAM phosphorylation. 29
cAMP 28
Phospholipase C Is Responsible for the Hydrolysis
28
of Membrane PI-4,5P 2 Activation of adenylate cyclase via G s increases the intracellular level
of cAMP, which activates protein kinase A (PKA). This serine/
Activation of PLC, either the β-isoform (by thrombin, ADP, or threonine kinase is responsible for phosphorylating proteins includ-
TxA 2 via G q ) or the γ-isoform (by collagen, protein tyrosine kinases, ing GPIbβ, filamin, myosin light chain (MLC), and Rap1B. Elevated
2+
and scaffold molecules) results in the hydrolysis of the minor inner levels of cAMP result in sequestration of intracellular Ca in the
plasma membrane leaflet phospholipid phosphatidylinositol-4,5- DTS, and inhibition of cytoskeletal rearrangement, granule secretion,
bisphosphate (PI-4,5P 2 ). The two second messengers that are the and aggregation. Inhibition of adenylate cyclase via G i is important
formed, soluble inositol-1,4,5-trisphosphate (IP 3 ) and membrane- in facilitating activation of platelets by lowering cAMP concentra-
associated diacylglycerol (DAG), lead, respectively, to increased tions that are raised above basal levels by PGI 2 produced by the
2+
cytosolic Ca concentrations and activation of protein kinase C endothelium.
(PKC), a serine/threonine kinase.
Reorganization of the Actin Cytoskeleton
Phosphatidylinositol 3-Kinase Forms
3-Phosphorylated Phosphoinositides 28,32 Platelet plug formation requires platelets to undergo a rapid change
from their resting discoid shape to active forms spread over the
Activation of PI3K isoforms, either the γ (via G i) or the α, β, and δ damaged subendothelium that then recruit additional platelets
(via the collagen signaling pathway), results in phosphorylation of by providing an enlarged surface area for platelet-platelet or