Dual regulation of Glycogen Synthase Kinase 3 (GSK3)alpha/beta by Protein Kinase C (PKC)alpha and Akt promotes Thrombin-mediated Integrin alpha(IIb)beta(3) activation and granule secretion in platelets

TY - JOUR

T1 - Dual regulation of Glycogen Synthase Kinase 3 (GSK3)alpha/beta by Protein Kinase C (PKC)alpha and Akt promotes Thrombin-mediated Integrin alpha(IIb)beta(3) activation and granule secretion in platelets

AU - Moore, Samantha F.

AU - van den Bosch, Marion T. J.

AU - Hunter, Roger W.

AU - Sakamoto, Kei

AU - Poole, Alastair W.

AU - Hers, Ingeborg

PY - 2013

Y1 - 2013

N2 - Glycogen synthase kinase-3 is a Ser/Thr kinase, tonically active in resting cells but inhibited by phosphorylation of an N-terminal Ser residue (Ser(21) in GSK3 alpha and Ser(9) in GSK3 beta) in response to varied external stimuli. Recent work suggests that GSK3 functions as a negative regulator of platelet function, but how GSK3 is regulated in platelets has not been examined in detail. Here, we show that early thrombin-mediated GSK3 phosphorylation (0-30 s) was blocked by PKC inhibitors and largely absent in platelets from PKC alpha knock-out mice. In contrast, late (2-5 min) GSK3 phosphorylation was dependent on the PI3K/Akt pathway. Similarly, early thrombin-mediated inhibition of GSK3 activity was blocked in PKC alpha knock-out platelets, whereas the Akt inhibitor MK2206 reduced late thrombin-mediated GSK3 inhibition and largely prevented GSK3 inhibition in PKC alpha knock-out platelets. More importantly, GSK3 phosphorylation contributes to platelet function as knock-in mice where GSK3 alpha Ser(21) and GSK3 beta Ser(9) were mutated to Ala showed a significant reduction in PAR4-mediated platelet aggregation, fibrinogen binding, and P-selectin expression, whereas the GSK3 inhibitor CHIR99021 enhanced these responses. Together, these results demonstrate that PKC alpha and Akt modulate platelet function by phosphorylating and inhibiting GSK3 alpha/beta, thereby relieving the negative effect of GSK3 alpha/beta on thrombin-mediated platelet activation.

AB - Glycogen synthase kinase-3 is a Ser/Thr kinase, tonically active in resting cells but inhibited by phosphorylation of an N-terminal Ser residue (Ser(21) in GSK3 alpha and Ser(9) in GSK3 beta) in response to varied external stimuli. Recent work suggests that GSK3 functions as a negative regulator of platelet function, but how GSK3 is regulated in platelets has not been examined in detail. Here, we show that early thrombin-mediated GSK3 phosphorylation (0-30 s) was blocked by PKC inhibitors and largely absent in platelets from PKC alpha knock-out mice. In contrast, late (2-5 min) GSK3 phosphorylation was dependent on the PI3K/Akt pathway. Similarly, early thrombin-mediated inhibition of GSK3 activity was blocked in PKC alpha knock-out platelets, whereas the Akt inhibitor MK2206 reduced late thrombin-mediated GSK3 inhibition and largely prevented GSK3 inhibition in PKC alpha knock-out platelets. More importantly, GSK3 phosphorylation contributes to platelet function as knock-in mice where GSK3 alpha Ser(21) and GSK3 beta Ser(9) were mutated to Ala showed a significant reduction in PAR4-mediated platelet aggregation, fibrinogen binding, and P-selectin expression, whereas the GSK3 inhibitor CHIR99021 enhanced these responses. Together, these results demonstrate that PKC alpha and Akt modulate platelet function by phosphorylating and inhibiting GSK3 alpha/beta, thereby relieving the negative effect of GSK3 alpha/beta on thrombin-mediated platelet activation.

KW - INSULIN

KW - INHIBITORS

KW - COLLAGEN

KW - MICE

KW - AGGREGATION

KW - PHOSPHORYLATION

KW - PHOSPHOINOSITIDE 3-KINASE

KW - G(I) SIGNALING PATHWAYS

U2 - 10.1074/jbc.M112.429936

DO - 10.1074/jbc.M112.429936

M3 - Article

C2 - 23239877

SN - 0021-9258

VL - 288

SP - 3918

EP - 3928

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

IS - 6

ER -