Thrombin, tyrosine phosphorylation

Du J, Sperling LS, Marrero MB, Phillips L, Delafontaine P. 1996. G-protein and tyrosine kinase receptor cross-talk in rat aortic smooth muscle cells thrombin- and angiotensin Il-induced tyrosine phosphorylation of insulin receptor substrate-1 and insulin-like growth factor 1 receptor. Biochem Biophys Res Commun 218 934-939. 

Dillcm AM, Heath MF. Protein tyrosine phosphorylation in equine platelets the effect of stimulation by thrombin and platelet-activating factor. Equine Veterin J 1995 27 448-458... 

Figure 9.4S.1. Time coarees of tyrosine phosphorylation in intact platelets stimulated with thrombin (1 U/ml) and collagen (50 ig/ml) in tiie absence of inhibitors of autocrine stimulation. Thrombin induce a fast and transient (maximal at 1 min) tyrosine phosphorylation (IP), while collagen is a slow inducer of TF (maximal around 3-5 min). Collagen induces specifically TF ofproteins of40,140 kDa, where the 40 kDa protein after 10 ruin of incubation still have not reached its maximal levels. Thrombin and collagen induces most of tire same proteins.

Caution should however be taken in determining the proteins that are specifically tyrosine phosphoiylated in response to various agonists. This can be illustrated with thrombin, where studies have shown that removal of the autocrine stimulation in platelets during activation strongly reduces the tyrosine phosphorylation stimulated by thrombin (Figure 9.4.9.2). 

RYNINGEN a, HOLMSEN H. Thrombin per se does not induce tyrosine phosphorylation in human platelets as judged by Western blotting, while collagen does the significance of synergistic, autocrine stimulation. Biochem BiophysRes Commun. In Press, 1998. 

Grabham, P. and Cunningham, D. B., Thrombin receptor activation stimulates astrocyte proliferation and reversal of stellatation by distinct pathways. Involvement of tyrosine phosphorylation, / Neurochem., 64, 583, 1995. 

Immunoblotting-based experiments using an anti-phosphotyrosine antibody showed that incubation of platelets with ajoene enhanced the phosphorylation of at least four proteins (estimated molecular weights 76, 80, 84 and 120 kDa), both in resting platelets and in platelets subsequently stimulated with thrombin (0.1 U/ml). This effect was both dose- and incubation-time-dependent. High concentrations of ajoene (50 pM) or long periods of incubation (10 min) led to nonselective hyperphosphorylation of numerous proteins. Also, protein tyrosine phosphatase activity was inhibited when platelets were incubated with ajoene before lysis, but not when ajoene was added to lysates of platelets which had not been pre-exposed to ajoene [99],... 

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