Integrin cytoplasmic tyrosine motif is required for outside-in αIIbβ3 signalling and platelet function

Abstract

Integrins not only bind adhesive ligands, they also act as signalling receptors. Both functions allow the integrin alphaIIbbeta3 to mediate platelet aggregation. Platelet agonists activate alphaIIbbeta3 (inside-out signalling) to allow the binding of soluble fibrinogen. Subsequent platelet aggregation leads to outside-in alphaIIbbeta3 signalling, which results in calcium mobilization, tyrosine phosphorylation of numerous proteins including beta3 itself, increased cytoskeletal reorganisation and further activation of alphaIIbbeta3. Thus, outside-in signals enhance aggregation, although the mechanisms and functional consequences of specific signalling events remain unclear. Here we describe a mouse that expresses an alphaIIbbeta3 in which the tyrosines in the integrin cytoplasmic tyrosine motif have been mutated to phenylalanines. These mice are selectively impaired in outside-in alphaIIbbeta3 signalling, with defective aggregation and clot-retraction responses in vitro, and an in vivo bleeding defect which is characterized by a pronounced tendency to rebleed. These data provide evidence for an important role of outside-in signalling in platelet physiology. Furthermore, they identify the integrin cytoplasmic tyrosine motif as a key mediator of beta-integrin signals and a potential target for new therapeutic agents.

Restoration of positioning control following Disabled-2 expression in ovarian and breast tumor cells

“…Alternatively, all of the integrin beta subunits also contain NPXY motif in their intracellular domain. The NPXY motif in beta-integrin subunit is important in intracellular signaling (Law et al, 1999;Vignoud et al, 1997;Tahiliani et al, 1997;O'Toole et al, 1995), and Dab2 may mediate contact signaling of integrin. Whether Dab2 bind to these NPXY motifs and is involved in LDL-family receptors or integrin signaling is to be determined.…”

Integrins in regulation of tissue development and function

“…Indeed, mice in which the two conserved tyrosines in the cytoplasmic tail of the β3 subunit were mutated to phenylalanine show defective platelet aggregation and clot retraction in vitro and suffer from a bleeding disorder in vivo, indicating that αIIbβ3-mediated 'outside-in' signalling is crucial to proper platelet function in vivo [114]. Because of the central role of this integrin in haemostasis, αIIbβ3 antagonists are currently being tested in clinical trials [115].…”

In vivo β1 integrin function requires phosphorylation-independent regulation by cytoplasmic tyrosines

“…Following inside-out activation and fibrinogen binding, the cytoplasmic tail of the ␤3 subunit is tyrosine phosphorylated, an event that initiates a secondary wave of integrin signaling referred to as outside-in. In vivo replacement of the ␤3 cytoplasmic tyrosines with phenylalanine does not interrupt inside-out activation of platelet integrins, but results in defective hemostasis due to the loss of outside-in integrin signals (Law et al 1999). Mouse and human platelets express the ␣2␤1 integrin, which functions as a receptor for vessel wall collagen.…”

“…␤3 integrins in platelets are critical for hemostasis, and substitution of the two ␤3 NPxY motifs with NPxF to block phosphorylation-dependent integrin signaling results in a hemostatic defect (Law et al 1999). Whether this phenotype reflects a broad role for ␤ integrin tyrosine phosphorylation or a more restricted role for ␤3 integrins in platelets is unknown.…”