Tyrosine phosphorylation of the beta3 cytoplasmic domain mediates integrin-cytoskeletal interactions.
Tyrosine phosphorylation of the beta3 subunit of the major platelet integrin alphaIIb beta3 has been shown to occur during thrombin-induced platelet aggregation (1). We now show that a wide variety of platelet stimuli induced beta3 tyrosine phosphorylation, but that this phosphorylation occurred only following platelet aggregation. Several lines of evidence suggest that the beta3 cytoplasmic domain tyrosine residues and/or their phosphorylation function to mediate interactions between beta3 integrins and cytoskeletal proteins. First, phospho-beta3 was retained preferentially in a Triton X-100 insoluble cytoskeletal fraction of thrombin-aggregated platelets. Second, in vitro experiments show that the cytoskeletal protein, myosin, associated in a phosphotyrosine-dependent manner with a diphosphorylated peptide corresponding to residues 740-762 of beta3. Third, mutation of both tyrosines in the beta3 cytoplasmic domain to phenylalanines markedly reduced beta3-dependent fibrin clot retraction. Thus, our data indicate that platelet aggregation is both necessary and sufficient for beta3 tyrosine phosphorylation, and this phosphorylation results in the physical linkage of alphaIIb beta3 to the cytoskeleton. We hypothesize that this linkage may involve direct binding of the phosphorylated integrin to the contractile protein myosin in order to mediate transmission of force to the fibrin clot during the process of clot retraction.
Jenkins
A L
AL
COR Therapeutics, Inc., South San Francisco, California 94080, USA.
Nannizzi-Alaimo
L
L
Silver
D
D
Sellers
J R
JR
Ginsberg
M H
MH
Law
D A
DA
Phillips
D R
DR
D000595:Amino Acid Sequence; D000818:Animals; D015703:Antigens, CD / Q000737:chemistry / Q000378:metabolism*; D016466:CHO Cells; D006224:Cricetinae; D003599:Cytoskeleton / Q000378:metabolism*; D006801:Humans; D039661:Integrin beta3; D008969:Molecular Sequence Data; D009218:Myosins / Q000378:metabolism; D010766:Phosphorylation; D010974:Platelet Aggregation; D010980:Platelet Membrane Glycoproteins / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D014443:Tyrosine / Q000378:metabolism*
D015703:Antigens, CD; D039661:Integrin beta3; D010980:Platelet Membrane Glycoproteins; D014443:Tyrosine; D009218:Myosins