Evidence against a direct role of the integrin alpha2beta1 in collagen-induced tyrosine phosphorylation in human platelets.
In the present study we have investigated whether the collagen receptor alpha2beta1 (GPIa-IIa; GP, glycoprotein) regulates protein tyrosine phosphorylation in platelets directly through activation of tyrosine kinases or indirectly through modification of the response to GPVI. The interaction of collagen with alpha2beta1 was inhibited in two distinct ways, using the metalloprotease jararhagin, which cleaves the beta1 subunit, or the antibody P1E6 which competes with binding of collagen to the integrin. The two inhibitors caused a shift to the right in the collagen concentration response curves for protein tyrosine phosphorylation and platelet activation consistent with a causal relationship between the two events. There was no change in the overall pattern of tyrosine phosphorylation in response to high concentrations of collagen in the presence of alpha2beta1 blockade demonstrating that the integrin is not required for this event. In contrast, jararhagin and P1E6 had a small, almost negligible inhibitory effect against responses to the GPVI-selective agonist collagen-related peptide (CRP) and the G protein-coupled receptor agonist thrombin. Crosslinking of alpha2beta1 in solution or by adhesion to a monolayer using a variety of antibodies to either subunit of the integrin did not induce detectable protein tyrosine phosphorylation in whole cell lysates. The snake venom toxin trimucytin-stimulated a similar pattern of tyrosine phosphorylation to that induced by crosslinking of GPVI which was maintained in the presence of jararhagin. Trimucytin may therefore induce activation via GPVI rather than alpha2beta1 as previously thought. These observations show that the integrin alpha2beta1 is not required for regulation of tyrosine phosphorylation by collagen.
Hers
I
I
Department of Pharmacology, University of Oxford, UK.
Berlanga
O
O
Tiekstra
M J
MJ
Kamiguti
A S
AS
Theakston
R D
RD
Watson
S P
SP
D000328:Adult; D000911:Antibodies, Monoclonal / Q000276:immunology; D001792:Blood Platelets / Q000187:drug effects / Q000378:metabolism*; D003094:Collagen / Q000378:metabolism*; D003435:Crotalid Venoms / Q000737:chemistry / Q000494:pharmacology; D006801:Humans; D016023:Integrins / Q000276:immunology / Q000378:metabolism*; D008666:Metalloendopeptidases / Q000494:pharmacology; D010766:Phosphorylation; D010975:Platelet Aggregation Inhibitors / Q000378:metabolism; D011509:Proteoglycans / Q000494:pharmacology; D038981:Receptors, Collagen; D015398:Signal Transduction; D014443:Tyrosine / Q000378:metabolism*; D000097146:Bothrops jararaca Venom
D000911:Antibodies, Monoclonal; D003094:Collagen; D003435:Crotalid Venoms; D016023:Integrins; D008666:Metalloendopeptidases; D010975:Platelet Aggregation Inhibitors; D011509:Proteoglycans; D038981:Receptors, Collagen; D014443:Tyrosine; D000097146:Bothrops jararaca Venom; C026064:aggregoserpentin