Abstract
BackgroundPaxillin is a LIM domain protein localized at integrin-mediated focal adhesions. Although paxillin is thought to modulate the functions of integrins, little is known about the contribution of paxillin to signaling pathways in platelets. Here, we studied the role of paxillin in platelet activation in vitro and in vivo.Methods and resultsWe generated paxillin knockdown (Pxn-KD) platelets in mice by transplanting bone marrow cells transduced with a lentiviral vector carrying a short hairpin RNA sequence, and confirmed that paxillin expression was significantly reduced in platelets derived from the transduced cells. Pxn-KD platelets showed a slight increased in size and augmented integrin αIIbβ3 activation following stimulation of multiple receptors including glycoprotein VI and G protein-coupled receptors. Thromboxane A2 biosynthesis and the release of α-granules and dense granules in response to agonist stimulation were also enhanced in Pxn-KD platelets. However, Pxn-KD did not increase tyrosine phosphorylation or intracellular calcium mobilization. Intravital imaging confirmed that Pxn-KD enhanced thrombus formation in vivo.ConclusionsOur findings suggest that paxillin negatively regulates several common platelet signaling pathways, resulting in the activation of integrin αIIbβ3 and release reactions.
Highlights
Paxillin is a LIM domain protein localized at integrin-mediated focal adhesions
Our findings suggest that paxillin negatively regulates several common platelet signaling pathways, resulting in the activation of integrin αIIbβ3 and release reactions
We previously reported that lentiviral vector-mediated short hairpin RNA expression in hematopoietic stem cells greatly reduces the expression of the target protein in platelets [21]
Summary
Paxillin is thought to modulate the functions of integrins, little is known about the contribution of paxillin to signaling pathways in platelets. We studied the role of paxillin in platelet activation in vitro and in vivo. A breakdown of normal platelet function results in either unexpected bleeding or thrombotic events [1]. Activated platelets release several classes of agonists, including ADP and thromboxane (Tx) A2, which promote further platelet activation [3]. These steps increase the affinity of integrin αIIbβ for its ligands and induce platelet aggregation [4]. The intracellular signaling that increases the affinity of integrins is known as inside-out signaling [4].
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