American Society of Hematology

Figure 6

$Figure 6. Mutation of tyrosine 136 of LAT does not affect GPVI-dependent membranes/cytoskeleton recruitment of PLCγ2. WT, LatY136F (A), and Lat3YF (B) platelets treated or not with the PI3K inhibitor wortmannin (50 nM) were stimulated by 5 nM Cvx for the indicated times. (A,B) Half of the cells were analyzed for the total amount of PLC-γ2 (bottom panels), and the other half were immediately permeabilized by saponin for cytosol depletion. After centrifugation (12 000g for 40 seconds), the pellet (corresponding to the membranes and cytoskeleton fraction; Mbr/Csk) was suspended in Laemli sample buffer, and the amount of PLC-γ2 was analyzed by immunoblotting (top panels). The effect of PI3K inhibition by wortmannin on PtdOH formation (C) and PLCγ2 tyrosine phosphorylation (D) of platelets from WT mice was then analyzed. Results are means plus or minus SEM of 3 experiments.$

Mutation of tyrosine 136 of LAT does not affect GPVI-dependent membranes/cytoskeleton recruitment of PLCγ2. WT, LatY136F (A), and Lat3YF (B) platelets treated or not with the PI3K inhibitor wortmannin (50 nM) were stimulated by 5 nM Cvx for the indicated times. (A,B) Half of the cells were analyzed for the total amount of PLC-γ2 (bottom panels), and the other half were immediately permeabilized by saponin for cytosol depletion. After centrifugation (12 000g for 40 seconds), the pellet (corresponding to the membranes and cytoskeleton fraction; Mbr/Csk) was suspended in Laemli sample buffer, and the amount of PLC-γ2 was analyzed by immunoblotting (top panels). The effect of PI3K inhibition by wortmannin on PtdOH formation (C) and PLCγ2 tyrosine phosphorylation (D) of platelets from WT mice was then analyzed. Results are means plus or minus SEM of 3 experiments.

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