Regulation of cardiac ryanodine receptor function by the cyclic-GMP dependent protein kinase G
- Autores
- Gonano, Luis Alberto; Aitken-Buck, Hamish M.; Chakraborty, Akash D.; Worthington, Luke P. I.; Cully, Tanya R.; Lamberts, Regis R.; Vila Petroff, Martín Gerardo; Jones, Peter P.
- Año de publicación
- 2022
- Idioma
- inglés
- Tipo de recurso
- artículo
- Estado
- versión publicada
- Descripción
- Background: The cGMP-dependent protein kinase G (PKG) phosphorylates the cardiac ryanodine receptor (RyR2) in vitro. We aimed to determine whether modulation of endogenous PKG alters RyR2-mediated spontaneous Ca²⁺ release and whether this effect is linked to a change in RyR2 phosphorylation. Methods: & Results: Human embryonic kidney (HEK293) cells with inducible RyR2 expression were treated with the cGMP analogue 8-Br-cGMP (100 μM) to activate endogenous PKG. In cells transfected with luminal Ca²⁺ sensor, D1ER, PKG activation significantly reduced the threshold for RyR2-mediated spontaneous Ca²⁺ release (93.9 ± 0.4% of store size with vehicle vs. 91.7 ± 0.8% with 8-Br-cGMP, P = 0.04). Mutation of the proposed PKG phosphorylation sites, S2808 and S2030, either individually or as a combination, prevented the decrease in Ca²⁺ release threshold induced by endogenous PKG activation. Interestingly, despite a functional dependence on expression of RyR2 phosphorylation sites, 8-Br-cGMP activation of PKG did not promote a detectable change in S2808 phosphorylation (P = 0.9). Paradoxically, pharmacological inhibition of PKG with KT 5823 (1 μM) also reduced the threshold for spontaneous Ca²⁺ release through RyR2 without affecting S2808 phosphorylation. Silencing RNA knockdown of endogenous PKG expression also had no quantifiable effect on RyR2 S2808 phosphorylation (P = 0.9). However, unlike PKG inhibition with KT 5823, PKG knockdown did not alter spontaneous Ca²⁺ release propensity or luminal Ca²⁺ handling. Conclusion: In an intact cell model, activation of endogenous PKG reduces the threshold for RyR2-mediated spontaneous Ca²⁺ release in a manner dependent on the RyR2 phosphorylation sites S2808 and S2030. This study clarifies the regulation of RyR2 Ca²⁺ release by endogenous PKG and functionally implicates the role of RyR2 phosphorylation.
Facultad de Ciencias Médicas
Centro de Investigaciones Cardiovasculares - Materia
-
Medicina
Protein kinase G
Cardiac ryanodine receptor
Phosphorylation
Calcium
Store overload-induced calcium release
KT 5823 - Nivel de accesibilidad
- acceso abierto
- Condiciones de uso
- http://creativecommons.org/licenses/by-nc-nd/4.0/
- Repositorio
- Institución
- Universidad Nacional de La Plata
- OAI Identificador
- oai:sedici.unlp.edu.ar:10915/154468
Ver los metadatos del registro completo
id |
SEDICI_ccf71c09c578720c78347ef4827ad8e0 |
---|---|
oai_identifier_str |
oai:sedici.unlp.edu.ar:10915/154468 |
network_acronym_str |
SEDICI |
repository_id_str |
1329 |
network_name_str |
SEDICI (UNLP) |
spelling |
Regulation of cardiac ryanodine receptor function by the cyclic-GMP dependent protein kinase GGonano, Luis AlbertoAitken-Buck, Hamish M.Chakraborty, Akash D.Worthington, Luke P. I.Cully, Tanya R.Lamberts, Regis R.Vila Petroff, Martín GerardoJones, Peter P.MedicinaProtein kinase GCardiac ryanodine receptorPhosphorylationCalciumStore overload-induced calcium releaseKT 5823Background: The cGMP-dependent protein kinase G (PKG) phosphorylates the cardiac ryanodine receptor (RyR2) in vitro. We aimed to determine whether modulation of endogenous PKG alters RyR2-mediated spontaneous Ca²⁺ release and whether this effect is linked to a change in RyR2 phosphorylation. Methods: & Results: Human embryonic kidney (HEK293) cells with inducible RyR2 expression were treated with the cGMP analogue 8-Br-cGMP (100 μM) to activate endogenous PKG. In cells transfected with luminal Ca²⁺ sensor, D1ER, PKG activation significantly reduced the threshold for RyR2-mediated spontaneous Ca²⁺ release (93.9 ± 0.4% of store size with vehicle vs. 91.7 ± 0.8% with 8-Br-cGMP, P = 0.04). Mutation of the proposed PKG phosphorylation sites, S2808 and S2030, either individually or as a combination, prevented the decrease in Ca²⁺ release threshold induced by endogenous PKG activation. Interestingly, despite a functional dependence on expression of RyR2 phosphorylation sites, 8-Br-cGMP activation of PKG did not promote a detectable change in S2808 phosphorylation (P = 0.9). Paradoxically, pharmacological inhibition of PKG with KT 5823 (1 μM) also reduced the threshold for spontaneous Ca²⁺ release through RyR2 without affecting S2808 phosphorylation. Silencing RNA knockdown of endogenous PKG expression also had no quantifiable effect on RyR2 S2808 phosphorylation (P = 0.9). However, unlike PKG inhibition with KT 5823, PKG knockdown did not alter spontaneous Ca²⁺ release propensity or luminal Ca²⁺ handling. Conclusion: In an intact cell model, activation of endogenous PKG reduces the threshold for RyR2-mediated spontaneous Ca²⁺ release in a manner dependent on the RyR2 phosphorylation sites S2808 and S2030. This study clarifies the regulation of RyR2 Ca²⁺ release by endogenous PKG and functionally implicates the role of RyR2 phosphorylation.Facultad de Ciencias MédicasCentro de Investigaciones Cardiovasculares2022info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionArticulohttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdf171-178http://sedici.unlp.edu.ar/handle/10915/154468enginfo:eu-repo/semantics/altIdentifier/issn/2665-9441info:eu-repo/semantics/altIdentifier/doi/10.1016/j.crphys.2022.03.004info:eu-repo/semantics/openAccesshttp://creativecommons.org/licenses/by-nc-nd/4.0/Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International (CC BY-NC-ND 4.0)reponame:SEDICI (UNLP)instname:Universidad Nacional de La Platainstacron:UNLP2025-09-29T11:40:03Zoai:sedici.unlp.edu.ar:10915/154468Institucionalhttp://sedici.unlp.edu.ar/Universidad públicaNo correspondehttp://sedici.unlp.edu.ar/oai/snrdalira@sedici.unlp.edu.arArgentinaNo correspondeNo correspondeNo correspondeopendoar:13292025-09-29 11:40:03.391SEDICI (UNLP) - Universidad Nacional de La Platafalse |
dc.title.none.fl_str_mv |
Regulation of cardiac ryanodine receptor function by the cyclic-GMP dependent protein kinase G |
title |
Regulation of cardiac ryanodine receptor function by the cyclic-GMP dependent protein kinase G |
spellingShingle |
Regulation of cardiac ryanodine receptor function by the cyclic-GMP dependent protein kinase G Gonano, Luis Alberto Medicina Protein kinase G Cardiac ryanodine receptor Phosphorylation Calcium Store overload-induced calcium release KT 5823 |
title_short |
Regulation of cardiac ryanodine receptor function by the cyclic-GMP dependent protein kinase G |
title_full |
Regulation of cardiac ryanodine receptor function by the cyclic-GMP dependent protein kinase G |
title_fullStr |
Regulation of cardiac ryanodine receptor function by the cyclic-GMP dependent protein kinase G |
title_full_unstemmed |
Regulation of cardiac ryanodine receptor function by the cyclic-GMP dependent protein kinase G |
title_sort |
Regulation of cardiac ryanodine receptor function by the cyclic-GMP dependent protein kinase G |
dc.creator.none.fl_str_mv |
Gonano, Luis Alberto Aitken-Buck, Hamish M. Chakraborty, Akash D. Worthington, Luke P. I. Cully, Tanya R. Lamberts, Regis R. Vila Petroff, Martín Gerardo Jones, Peter P. |
author |
Gonano, Luis Alberto |
author_facet |
Gonano, Luis Alberto Aitken-Buck, Hamish M. Chakraborty, Akash D. Worthington, Luke P. I. Cully, Tanya R. Lamberts, Regis R. Vila Petroff, Martín Gerardo Jones, Peter P. |
author_role |
author |
author2 |
Aitken-Buck, Hamish M. Chakraborty, Akash D. Worthington, Luke P. I. Cully, Tanya R. Lamberts, Regis R. Vila Petroff, Martín Gerardo Jones, Peter P. |
author2_role |
author author author author author author author |
dc.subject.none.fl_str_mv |
Medicina Protein kinase G Cardiac ryanodine receptor Phosphorylation Calcium Store overload-induced calcium release KT 5823 |
topic |
Medicina Protein kinase G Cardiac ryanodine receptor Phosphorylation Calcium Store overload-induced calcium release KT 5823 |
dc.description.none.fl_txt_mv |
Background: The cGMP-dependent protein kinase G (PKG) phosphorylates the cardiac ryanodine receptor (RyR2) in vitro. We aimed to determine whether modulation of endogenous PKG alters RyR2-mediated spontaneous Ca²⁺ release and whether this effect is linked to a change in RyR2 phosphorylation. Methods: & Results: Human embryonic kidney (HEK293) cells with inducible RyR2 expression were treated with the cGMP analogue 8-Br-cGMP (100 μM) to activate endogenous PKG. In cells transfected with luminal Ca²⁺ sensor, D1ER, PKG activation significantly reduced the threshold for RyR2-mediated spontaneous Ca²⁺ release (93.9 ± 0.4% of store size with vehicle vs. 91.7 ± 0.8% with 8-Br-cGMP, P = 0.04). Mutation of the proposed PKG phosphorylation sites, S2808 and S2030, either individually or as a combination, prevented the decrease in Ca²⁺ release threshold induced by endogenous PKG activation. Interestingly, despite a functional dependence on expression of RyR2 phosphorylation sites, 8-Br-cGMP activation of PKG did not promote a detectable change in S2808 phosphorylation (P = 0.9). Paradoxically, pharmacological inhibition of PKG with KT 5823 (1 μM) also reduced the threshold for spontaneous Ca²⁺ release through RyR2 without affecting S2808 phosphorylation. Silencing RNA knockdown of endogenous PKG expression also had no quantifiable effect on RyR2 S2808 phosphorylation (P = 0.9). However, unlike PKG inhibition with KT 5823, PKG knockdown did not alter spontaneous Ca²⁺ release propensity or luminal Ca²⁺ handling. Conclusion: In an intact cell model, activation of endogenous PKG reduces the threshold for RyR2-mediated spontaneous Ca²⁺ release in a manner dependent on the RyR2 phosphorylation sites S2808 and S2030. This study clarifies the regulation of RyR2 Ca²⁺ release by endogenous PKG and functionally implicates the role of RyR2 phosphorylation. Facultad de Ciencias Médicas Centro de Investigaciones Cardiovasculares |
description |
Background: The cGMP-dependent protein kinase G (PKG) phosphorylates the cardiac ryanodine receptor (RyR2) in vitro. We aimed to determine whether modulation of endogenous PKG alters RyR2-mediated spontaneous Ca²⁺ release and whether this effect is linked to a change in RyR2 phosphorylation. Methods: & Results: Human embryonic kidney (HEK293) cells with inducible RyR2 expression were treated with the cGMP analogue 8-Br-cGMP (100 μM) to activate endogenous PKG. In cells transfected with luminal Ca²⁺ sensor, D1ER, PKG activation significantly reduced the threshold for RyR2-mediated spontaneous Ca²⁺ release (93.9 ± 0.4% of store size with vehicle vs. 91.7 ± 0.8% with 8-Br-cGMP, P = 0.04). Mutation of the proposed PKG phosphorylation sites, S2808 and S2030, either individually or as a combination, prevented the decrease in Ca²⁺ release threshold induced by endogenous PKG activation. Interestingly, despite a functional dependence on expression of RyR2 phosphorylation sites, 8-Br-cGMP activation of PKG did not promote a detectable change in S2808 phosphorylation (P = 0.9). Paradoxically, pharmacological inhibition of PKG with KT 5823 (1 μM) also reduced the threshold for spontaneous Ca²⁺ release through RyR2 without affecting S2808 phosphorylation. Silencing RNA knockdown of endogenous PKG expression also had no quantifiable effect on RyR2 S2808 phosphorylation (P = 0.9). However, unlike PKG inhibition with KT 5823, PKG knockdown did not alter spontaneous Ca²⁺ release propensity or luminal Ca²⁺ handling. Conclusion: In an intact cell model, activation of endogenous PKG reduces the threshold for RyR2-mediated spontaneous Ca²⁺ release in a manner dependent on the RyR2 phosphorylation sites S2808 and S2030. This study clarifies the regulation of RyR2 Ca²⁺ release by endogenous PKG and functionally implicates the role of RyR2 phosphorylation. |
publishDate |
2022 |
dc.date.none.fl_str_mv |
2022 |
dc.type.none.fl_str_mv |
info:eu-repo/semantics/article info:eu-repo/semantics/publishedVersion Articulo http://purl.org/coar/resource_type/c_6501 info:ar-repo/semantics/articulo |
format |
article |
status_str |
publishedVersion |
dc.identifier.none.fl_str_mv |
http://sedici.unlp.edu.ar/handle/10915/154468 |
url |
http://sedici.unlp.edu.ar/handle/10915/154468 |
dc.language.none.fl_str_mv |
eng |
language |
eng |
dc.relation.none.fl_str_mv |
info:eu-repo/semantics/altIdentifier/issn/2665-9441 info:eu-repo/semantics/altIdentifier/doi/10.1016/j.crphys.2022.03.004 |
dc.rights.none.fl_str_mv |
info:eu-repo/semantics/openAccess http://creativecommons.org/licenses/by-nc-nd/4.0/ Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International (CC BY-NC-ND 4.0) |
eu_rights_str_mv |
openAccess |
rights_invalid_str_mv |
http://creativecommons.org/licenses/by-nc-nd/4.0/ Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International (CC BY-NC-ND 4.0) |
dc.format.none.fl_str_mv |
application/pdf 171-178 |
dc.source.none.fl_str_mv |
reponame:SEDICI (UNLP) instname:Universidad Nacional de La Plata instacron:UNLP |
reponame_str |
SEDICI (UNLP) |
collection |
SEDICI (UNLP) |
instname_str |
Universidad Nacional de La Plata |
instacron_str |
UNLP |
institution |
UNLP |
repository.name.fl_str_mv |
SEDICI (UNLP) - Universidad Nacional de La Plata |
repository.mail.fl_str_mv |
alira@sedici.unlp.edu.ar |
_version_ |
1844616273721294848 |
score |
13.070432 |