Bacitracin reveals a role for multiple thiol isomerases in platelet function.
The platelet-specific integrin alphaIIb beta3 has endogenous thiol isomerase activity associated with the CXXC motifs within the beta subunit. Using a highly purified form of bacitracin, a thiol isomerase inhibitor, we now provide further evidence of the functional significance of this enzymatic activity in integrin activation. In addition, we demonstrate a role for multiple thiol isomerases in platelet function. This bacitracin prevented platelet aggregation to thrombin and collagen, and directly inhibited alphaIIb beta3 activation, as detected by PAC-1 binding. In parallel, bacitracin inhibited the endogenous thiol isomerase activity of purified alphaIIb beta3 with a 50% inhibitory concentration of 15.5 micromol/l. In order to determine whether the effects of bacitracin are solely mediated by inhibition of integrin enzymatic activity, we examined integrin-independent indices of platelet activation. We found bacitracin inhibited both platelet secretion (CD62P and CD63) and thromboxane (TxA2) production, with complete inhibition at different concentrations. Thus, we demonstrated a role for multiple thiol isomerases in platelet function. Taken together, these studies support a role for the endogenous integrin thiol isomerase activity in activation of alphaIIb beta3 and highlight the novel regulation of platelet function by other, as yet undefined thiol isomerases.
Robinson
Aisling
A
Department of Clinical Pharmacology, Royal College of Surgeons in Ireland, Dublin 2, Ireland.
O'Neill
Sarah
S
Kiernan
Aoife
A
O'Donoghue
Niaobh
N
Moran
Niamh
N
D001414:Bacitracin / Q000302:isolation & purification / Q000494:pharmacology*; D001792:Blood Platelets / Q000187:drug effects* / Q000201:enzymology / Q000502:physiology; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000302:isolation & purification / Q000494:pharmacology*; D005434:Flow Cytometry / Q000379:methods; D006801:Humans; D015539:Platelet Activation / Q000187:drug effects; D010973:Platelet Adhesiveness / Q000187:drug effects; D010974:Platelet Aggregation / Q000187:drug effects; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000378:metabolism; D019704:Protein Disulfide-Isomerases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods; D013928:Thromboxane A2 / Q000096:biosynthesis
D004791:Enzyme Inhibitors; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D001414:Bacitracin; D013928:Thromboxane A2; D019704:Protein Disulfide-Isomerases