Opposing roles of gsk3α and gsk3β phosphorylation in platelet function and thrombosis

Samantha Frances Moore; Ejaife O Agbani; J A Wersall; Alastair W Poole; Christopher M Williams; Xiaojuan Zhao; Yong Li; J L Hutchinson; Roger W Hunter; Ingeborg Hers

doi:10.3390/ijms221910656

Opposing roles of gsk3α and gsk3β phosphorylation in platelet function and thrombosis

Samantha Frances Moore, Ejaife O Agbani, J A Wersall, Alastair W Poole, Christopher M Williams , Xiaojuan Zhao, Yong Li, J L Hutchinson, Roger W Hunter, Ingeborg Hers^*

^*Corresponding author for this work

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Abstract

One of the mechanisms by which PI3 kinase can regulate platelet function is through phosphorylation of downstream substrates, including glycogen synthase kinase-3 (GSK3) and GSK3. Platelet activation results in the phosphorylation of an N-terminal serine residue in GSK3 (Ser21) and GSK3(Ser9), which competitively inhibits substrate phosphorylation. However, the role of phosphorylation of these paralogs is still largely unknown. Here we employed GSK3/ phosphorylation-resistant mouse models to explore the role of this inhibitory phosphorylation in regulating platelet activation. Expression of phosphorylation-resistant GSK3α/β reduced thrombin-mediated platelet aggregation, integrin αIIbβ3 activation, and α-granule secretion, whereas platelet responses to the GPVI agonist collagen-related peptide (CRP-XL) were significantly enhanced. GSK3 single knock-in lines revealed that this divergence is due to differential roles of GSK3α and GSK3β phosphorylation in regulating platelet function. Expression of phosphorylation-resistant GSK3α resulted in enhanced GPVI-mediated platelet activation, whereas expression of phosphorylation-resistant GSK3β resulted in a reduction in PAR-mediated platelet activation and impaired in vitro thrombus formation under flow. Interestingly, the latter was normalised in double GSK3α/β KI mice, indicating that GSK3α KI can compensate for the impairment in thrombosis caused by GSK3β KI. In conclusion, our data indicate that GSK3α and GSK3β have differential roles in regulating platelet function.

Original language	English
Article number	10656
Pages (from-to)	10656
Number of pages	20
Journal	International Journal of Molecular Sciences
Volume	22
Issue number	19
Early online date	30 Sept 2021
DOIs	https://doi.org/10.3390/ijms221910656
Publication status	Published - 1 Oct 2021

Bibliographical note

Funding Information:
Funding: This work was supported by the British Heart Foundation (PG/16/3/31833,

Funding Information:
Acknowledgments: We thank the University of Bristol’s Animal Services Unit (ASU), Wolfson Bi-oimaging Facility and BrisSynBio, a BBSRC/EPSRC-funded Synthetic Biology Research Centre (Grant Number: L01386X). We also thank Elizabeth Aitken for technical assistance and the blood donors within the Biomedical Sciences building at the University of Bristol. We are grateful to Kei Sakamoto (University of Copenhagen) for providing us with the GSK3α/β “knock-in” line.

Funding Information:
This work was supported by the British Heart Foundation (PG/16/3/31833, PG/16/21/32083, PG/16/102/32647, RG/15/16/31758, FS/14/23/30756), Welcome Trust Investigatorship (219472/Z/19/Z) and Elizabeth Blackwell Institute for Health Research, University of Bristol and Medical Research Council.

Publisher Copyright:
© 2021 by the authors. Licensee MDPI, Basel, Switzerland.

Keywords

GSK3; PI3 kinase; Akt; integrin activation; P-selectin expression; thrombosis; knock-in mouse models; p110β; phosphorylation.

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10.3390/ijms221910656

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The Role of the Water Channel Aquaporin-1 in the Regulation of Platelet Function, Procoagulant activity and Thrombosis
Poole, A. W.
10/11/17 → 9/11/20
Project: Research

Cite this

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title = "Opposing roles of gsk3α and gsk3β phosphorylation in platelet function and thrombosis",

abstract = "One of the mechanisms by which PI3 kinase can regulate platelet function is through phosphorylation of downstream substrates, including glycogen synthase kinase-3 (GSK3) and GSK3. Platelet activation results in the phosphorylation of an N-terminal serine residue in GSK3 (Ser21) and GSK3(Ser9), which competitively inhibits substrate phosphorylation. However, the role of phosphorylation of these paralogs is still largely unknown. Here we employed GSK3/ phosphorylation-resistant mouse models to explore the role of this inhibitory phosphorylation in regulating platelet activation. Expression of phosphorylation-resistant GSK3α/β reduced thrombin-mediated platelet aggregation, integrin αIIbβ3 activation, and α-granule secretion, whereas platelet responses to the GPVI agonist collagen-related peptide (CRP-XL) were significantly enhanced. GSK3 single knock-in lines revealed that this divergence is due to differential roles of GSK3α and GSK3β phosphorylation in regulating platelet function. Expression of phosphorylation-resistant GSK3α resulted in enhanced GPVI-mediated platelet activation, whereas expression of phosphorylation-resistant GSK3β resulted in a reduction in PAR-mediated platelet activation and impaired in vitro thrombus formation under flow. Interestingly, the latter was normalised in double GSK3α/β KI mice, indicating that GSK3α KI can compensate for the impairment in thrombosis caused by GSK3β KI. In conclusion, our data indicate that GSK3α and GSK3β have differential roles in regulating platelet function.",

keywords = "GSK3; PI3 kinase; Akt; integrin activation; P-selectin expression; thrombosis; knock-in mouse models; p110β; phosphorylation.",

author = "Moore, {Samantha Frances} and Agbani, {Ejaife O} and Wersall, {J A} and Poole, {Alastair W} and Williams, {Christopher M} and Xiaojuan Zhao and Yong Li and Hutchinson, {J L} and Hunter, {Roger W} and Ingeborg Hers",

note = "Funding Information: Funding: This work was supported by the British Heart Foundation (PG/16/3/31833, Funding Information: Acknowledgments: We thank the University of Bristol{\textquoteright}s Animal Services Unit (ASU), Wolfson Bi-oimaging Facility and BrisSynBio, a BBSRC/EPSRC-funded Synthetic Biology Research Centre (Grant Number: L01386X). We also thank Elizabeth Aitken for technical assistance and the blood donors within the Biomedical Sciences building at the University of Bristol. We are grateful to Kei Sakamoto (University of Copenhagen) for providing us with the GSK3α/β “knock-in” line. Funding Information: This work was supported by the British Heart Foundation (PG/16/3/31833, PG/16/21/32083, PG/16/102/32647, RG/15/16/31758, FS/14/23/30756), Welcome Trust Investigatorship (219472/Z/19/Z) and Elizabeth Blackwell Institute for Health Research, University of Bristol and Medical Research Council. Publisher Copyright: {\textcopyright} 2021 by the authors. Licensee MDPI, Basel, Switzerland.",

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T1 - Opposing roles of gsk3α and gsk3β phosphorylation in platelet function and thrombosis

AU - Moore, Samantha Frances

AU - Agbani, Ejaife O

AU - Wersall, J A

AU - Poole, Alastair W

AU - Williams , Christopher M

AU - Zhao, Xiaojuan

AU - Li, Yong

AU - Hutchinson, J L

AU - Hunter, Roger W

AU - Hers, Ingeborg

N1 - Funding Information: Funding: This work was supported by the British Heart Foundation (PG/16/3/31833, Funding Information: Acknowledgments: We thank the University of Bristol’s Animal Services Unit (ASU), Wolfson Bi-oimaging Facility and BrisSynBio, a BBSRC/EPSRC-funded Synthetic Biology Research Centre (Grant Number: L01386X). We also thank Elizabeth Aitken for technical assistance and the blood donors within the Biomedical Sciences building at the University of Bristol. We are grateful to Kei Sakamoto (University of Copenhagen) for providing us with the GSK3α/β “knock-in” line. Funding Information: This work was supported by the British Heart Foundation (PG/16/3/31833, PG/16/21/32083, PG/16/102/32647, RG/15/16/31758, FS/14/23/30756), Welcome Trust Investigatorship (219472/Z/19/Z) and Elizabeth Blackwell Institute for Health Research, University of Bristol and Medical Research Council. Publisher Copyright: © 2021 by the authors. Licensee MDPI, Basel, Switzerland.

PY - 2021/10/1

Y1 - 2021/10/1

N2 - One of the mechanisms by which PI3 kinase can regulate platelet function is through phosphorylation of downstream substrates, including glycogen synthase kinase-3 (GSK3) and GSK3. Platelet activation results in the phosphorylation of an N-terminal serine residue in GSK3 (Ser21) and GSK3(Ser9), which competitively inhibits substrate phosphorylation. However, the role of phosphorylation of these paralogs is still largely unknown. Here we employed GSK3/ phosphorylation-resistant mouse models to explore the role of this inhibitory phosphorylation in regulating platelet activation. Expression of phosphorylation-resistant GSK3α/β reduced thrombin-mediated platelet aggregation, integrin αIIbβ3 activation, and α-granule secretion, whereas platelet responses to the GPVI agonist collagen-related peptide (CRP-XL) were significantly enhanced. GSK3 single knock-in lines revealed that this divergence is due to differential roles of GSK3α and GSK3β phosphorylation in regulating platelet function. Expression of phosphorylation-resistant GSK3α resulted in enhanced GPVI-mediated platelet activation, whereas expression of phosphorylation-resistant GSK3β resulted in a reduction in PAR-mediated platelet activation and impaired in vitro thrombus formation under flow. Interestingly, the latter was normalised in double GSK3α/β KI mice, indicating that GSK3α KI can compensate for the impairment in thrombosis caused by GSK3β KI. In conclusion, our data indicate that GSK3α and GSK3β have differential roles in regulating platelet function.

AB - One of the mechanisms by which PI3 kinase can regulate platelet function is through phosphorylation of downstream substrates, including glycogen synthase kinase-3 (GSK3) and GSK3. Platelet activation results in the phosphorylation of an N-terminal serine residue in GSK3 (Ser21) and GSK3(Ser9), which competitively inhibits substrate phosphorylation. However, the role of phosphorylation of these paralogs is still largely unknown. Here we employed GSK3/ phosphorylation-resistant mouse models to explore the role of this inhibitory phosphorylation in regulating platelet activation. Expression of phosphorylation-resistant GSK3α/β reduced thrombin-mediated platelet aggregation, integrin αIIbβ3 activation, and α-granule secretion, whereas platelet responses to the GPVI agonist collagen-related peptide (CRP-XL) were significantly enhanced. GSK3 single knock-in lines revealed that this divergence is due to differential roles of GSK3α and GSK3β phosphorylation in regulating platelet function. Expression of phosphorylation-resistant GSK3α resulted in enhanced GPVI-mediated platelet activation, whereas expression of phosphorylation-resistant GSK3β resulted in a reduction in PAR-mediated platelet activation and impaired in vitro thrombus formation under flow. Interestingly, the latter was normalised in double GSK3α/β KI mice, indicating that GSK3α KI can compensate for the impairment in thrombosis caused by GSK3β KI. In conclusion, our data indicate that GSK3α and GSK3β have differential roles in regulating platelet function.

KW - GSK3; PI3 kinase; Akt; integrin activation; P-selectin expression; thrombosis; knock-in mouse models; p110β; phosphorylation.

U2 - 10.3390/ijms221910656

DO - 10.3390/ijms221910656

M3 - Article (Academic Journal)

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SN - 1661-6596

VL - 22

SP - 10656

JO - International Journal of Molecular Sciences

JF - International Journal of Molecular Sciences

IS - 19

M1 - 10656

ER -

Opposing roles of gsk3α and gsk3β phosphorylation in platelet function and thrombosis

Abstract

Bibliographical note

Keywords

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The Role of the Water Channel Aquaporin-1 in the Regulation of Platelet Function, Procoagulant activity and Thrombosis

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