Phosphorylation of PLCγ2 by collagen in the presence of TPO.

PLCγ2 tyrosine phosphorylation induced by TPO and collagen. Platelets were stimulated with stirring by collagen (10 μg/mL) for 90 seconds; TPO (50 ng/mL) for 300 seconds; TPO (50 ng/mL) for 300 seconds, followed by collagen for 90 seconds. Wortmannin (100 nmol/L) and LY294002 (50 μmol/L) are preincubated 3 minutes before the addition of TPO or collagen. PLCγ2 was immunoprecipitated and probed for tyrosine phosphorylation using the monoclonal antibody 4G10 (upper panel). Membranes are stripped and reprobed for PLCγ2 (lower panel). Results are from 1 experiment representative of 3.