Thrombin receptor activation and integrin engagement stimulate tyrosine phosphorylation of the proto-oncogene product, p95vav, in platelets.
The vav proto-oncogene product, p95vav or Vav, is primarily expressed in hematopoietic cells and has been shown to be a substrate for tyrosine kinases. Although its function is unknown, Vav shares a region of homology with DBL, an exchange factor for the Rho family of GTP-binding proteins. The presence of this domain and the observation that cells transformed with Vav display prominent stress fibers and focal adhesions similar to those that are observed in RhoA transformed cells suggests that Vav may play a role in regulating the actin cytoskeleton. We have, therefore, examined Vav phosphorylation in platelets, which undergo dramatic cytoskeletal reorganization in response to agonists. Two potent platelet agonists, thrombin (via its G protein-coupled receptor) and collagen (via its interaction with the alpha2beta1 integrin), caused Vav to become phosphorylated on tyrosine. Weaker platelet agonists, including ADP, epinephrine and the thromboxane A2 analog, U46619, did not. The phosphorylation of Vav in response to thrombin was maximal within 15 s and was unaffected by aspirin, inhibitors of aggregation, or the presence of the ADP scavenger, apyrase. Vav phosphorylation was also observed when platelets became adherent to immobilized collagen (via integrin alpha2beta1), fibronectin (via integrin alpha5beta1), and fibrinogen (via integrin alphaIIbbeta3). These results show that Vav phosphorylation by tyrosine kinases 1) occurs during platelet activation by potent agonists, 2) also occurs when platelets adhere to biologically relevant matrix proteins, 3) requires neither platelet aggregation nor the release of secondary agonists such as ADP and TxA2, and 4) can be initiated by at least some members of two additional classes of receptors, G protein-coupled receptors and integrins, providing further evidence that both of these can couple to tyrosine kinases.
Cichowski
K
K
Department of Medicine, University of Pennsylvania, Philadelphia, 19104, USA.
Brugge
J S
JS
Brass
L F
LF
D000595:Amino Acid Sequence; D000906:Antibodies / Q000494:pharmacology; D001241:Aspirin / Q000494:pharmacology; D001792:Blood Platelets / Q000187:drug effects / Q000378:metabolism* / Q000502:physiology; D018797:Cell Cycle Proteins; D003094:Collagen / Q000494:pharmacology; D016326:Extracellular Matrix Proteins; D019204:GTP-Binding Proteins / Q000378:metabolism; D006801:Humans; D015151:Immunoblotting; D016023:Integrins / Q000502:physiology*; D007700:Kinetics; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000494:pharmacology; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000032:analysis / Q000378:metabolism; D015539:Platelet Activation; D010973:Platelet Adhesiveness; D000090063:Proto-Oncogene Mas; D011518:Proto-Oncogene Proteins / Q000097:blood* / Q000302:isolation & purification; D051567:Proto-Oncogene Proteins c-vav; D018179:Receptors, Thrombin / Q000502:physiology*; D013917:Thrombin / Q000494:pharmacology; D018909:src Homology Domains
D000906:Antibodies; D018797:Cell Cycle Proteins; D016326:Extracellular Matrix Proteins; D016023:Integrins; C000715470:MAS1 protein, human; D010446:Peptide Fragments; D000090063:Proto-Oncogene Mas; D011518:Proto-Oncogene Proteins; D051567:Proto-Oncogene Proteins c-vav; D018179:Receptors, Thrombin; C496618:VAV1 protein, human; C082835:thrombin receptor peptide (42-47); D019000:Phosphotyrosine; D003094:Collagen; D013917:Thrombin; D019204:GTP-Binding Proteins; D001241:Aspirin