Phosphoprotein Phosphatase 1 but Not 2A Activity Modulates Coupled-Clock Mechanisms to Impact on Intrinsic Automaticity of Sinoatrial Nodal Pacemaker Cells

Spontaneous AP (action potential) firing of sinoatrial nodal cells (SANC) is critically dependent on protein kinase A (PKA) and Ca<sup>2+</sup>/calmodulin-dependent protein kinase II (CaMKII)-dependent protein phosphorylation, which are required for the generation of spontaneous, diastol...

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Autores principales:	Syevda Tagirova Sirenko, Ihor Zahanich, Yue Li, Yevgeniya O. Lukyanenko, Alexey E. Lyashkov, Bruce D. Ziman, Kirill V. Tarasov, Antoine Younes, Daniel R. Riordon, Yelena S. Tarasova, Dongmei Yang, Tatiana M. Vinogradova, Victor A. Maltsev, Edward G. Lakatta
Formato:	article
Lenguaje:	EN
Publicado:	MDPI AG 2021
Materias:	sinoatrial node cells local Ca<sup>2+</sup> releases phosphoprotein phosphatase endogenous phosphatase inhibitors calyculin A okadaic acid Biology (General) QH301-705.5
Acceso en línea:	https://doaj.org/article/d34e3e9a556b4610af29e09b6ffdc8e6
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spelling	oai:doaj.org-article:d34e3e9a556b4610af29e09b6ffdc8e62021-11-25T17:11:34ZPhosphoprotein Phosphatase 1 but Not 2A Activity Modulates Coupled-Clock Mechanisms to Impact on Intrinsic Automaticity of Sinoatrial Nodal Pacemaker Cells10.3390/cells101131062073-4409https://doaj.org/article/d34e3e9a556b4610af29e09b6ffdc8e62021-11-01T00:00:00Zhttps://www.mdpi.com/2073-4409/10/11/3106https://doaj.org/toc/2073-4409Spontaneous AP (action potential) firing of sinoatrial nodal cells (SANC) is critically dependent on protein kinase A (PKA) and Ca<sup>2+</sup>/calmodulin-dependent protein kinase II (CaMKII)-dependent protein phosphorylation, which are required for the generation of spontaneous, diastolic local Ca<sup>2+</sup> releases (LCRs). Although phosphoprotein phosphatases (PP) regulate protein phosphorylation, the expression level of PPs and phosphatase inhibitors in SANC and the impact of phosphatase inhibition on the spontaneous LCRs and other players of the oscillatory coupled-clock system is unknown. Here, we show that rabbit SANC express both PP1, PP2A, and endogenous PP inhibitors I-1 (PPI-1), dopamine and cyclic adenosine 3′,5′-monophosphate (cAMP)-regulated phosphoprotein (DARPP-32), kinase C-enhanced PP1 inhibitor (KEPI). Application of Calyculin A, (CyA), a PPs inhibitor, to intact, freshly isolated single SANC: (1) significantly increased phospholamban (PLB) phosphorylation (by 2–3-fold) at both CaMKII-dependent Thr<sup>17</sup> and PKA-dependent Ser<sup>16</sup> sites, in a time and concentration dependent manner; (2) increased ryanodine receptor (RyR) phosphorylation at the Ser<sup>2809</sup> site; (3) substantially increased sarcoplasmic reticulum (SR) Ca<sup>2+</sup> load; (4) augmented L-type Ca<sup>2+</sup> current amplitude; (5) augmented LCR’s characteristics and decreased LCR period in intact and permeabilized SANC, and (6) increased the spontaneous basal AP firing rate. In contrast, the selective PP2A inhibitor okadaic acid (100 nmol/L) had no significant effect on spontaneous AP firing, LCR parameters, or PLB phosphorylation. Application of purified PP1 to permeabilized SANC suppressed LCR, whereas purified PP2A had no effect on LCR characteristics. Our numerical model simulations demonstrated that PP inhibition increases AP firing rate via a coupled-clock mechanism, including respective increases in the SR Ca<sup>2+</sup> pumping rate, L-type Ca<sup>2+</sup> current, and Na<sup>+</sup>/Ca<sup>2+</sup>-exchanger current. Thus, PP1 and its endogenous inhibitors modulate the basal spontaneous firing rate of cardiac pacemaker cells by suppressing SR Ca<sup>2+</sup> cycling protein phosphorylation, the SR Ca<sup>2+</sup> load and LCRs, and L-type Ca<sup>2+</sup> current.Syevda Tagirova SirenkoIhor ZahanichYue LiYevgeniya O. LukyanenkoAlexey E. LyashkovBruce D. ZimanKirill V. TarasovAntoine YounesDaniel R. RiordonYelena S. TarasovaDongmei YangTatiana M. VinogradovaVictor A. MaltsevEdward G. LakattaMDPI AGarticlesinoatrial node cellslocal Ca<sup>2+</sup> releasesphosphoprotein phosphataseendogenous phosphatase inhibitorscalyculin Aokadaic acidBiology (General)QH301-705.5ENCells, Vol 10, Iss 3106, p 3106 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	sinoatrial node cells local Ca<sup>2+</sup> releases phosphoprotein phosphatase endogenous phosphatase inhibitors calyculin A okadaic acid Biology (General) QH301-705.5
spellingShingle	sinoatrial node cells local Ca<sup>2+</sup> releases phosphoprotein phosphatase endogenous phosphatase inhibitors calyculin A okadaic acid Biology (General) QH301-705.5 Syevda Tagirova Sirenko Ihor Zahanich Yue Li Yevgeniya O. Lukyanenko Alexey E. Lyashkov Bruce D. Ziman Kirill V. Tarasov Antoine Younes Daniel R. Riordon Yelena S. Tarasova Dongmei Yang Tatiana M. Vinogradova Victor A. Maltsev Edward G. Lakatta Phosphoprotein Phosphatase 1 but Not 2A Activity Modulates Coupled-Clock Mechanisms to Impact on Intrinsic Automaticity of Sinoatrial Nodal Pacemaker Cells
description	Spontaneous AP (action potential) firing of sinoatrial nodal cells (SANC) is critically dependent on protein kinase A (PKA) and Ca<sup>2+</sup>/calmodulin-dependent protein kinase II (CaMKII)-dependent protein phosphorylation, which are required for the generation of spontaneous, diastolic local Ca<sup>2+</sup> releases (LCRs). Although phosphoprotein phosphatases (PP) regulate protein phosphorylation, the expression level of PPs and phosphatase inhibitors in SANC and the impact of phosphatase inhibition on the spontaneous LCRs and other players of the oscillatory coupled-clock system is unknown. Here, we show that rabbit SANC express both PP1, PP2A, and endogenous PP inhibitors I-1 (PPI-1), dopamine and cyclic adenosine 3′,5′-monophosphate (cAMP)-regulated phosphoprotein (DARPP-32), kinase C-enhanced PP1 inhibitor (KEPI). Application of Calyculin A, (CyA), a PPs inhibitor, to intact, freshly isolated single SANC: (1) significantly increased phospholamban (PLB) phosphorylation (by 2–3-fold) at both CaMKII-dependent Thr<sup>17</sup> and PKA-dependent Ser<sup>16</sup> sites, in a time and concentration dependent manner; (2) increased ryanodine receptor (RyR) phosphorylation at the Ser<sup>2809</sup> site; (3) substantially increased sarcoplasmic reticulum (SR) Ca<sup>2+</sup> load; (4) augmented L-type Ca<sup>2+</sup> current amplitude; (5) augmented LCR’s characteristics and decreased LCR period in intact and permeabilized SANC, and (6) increased the spontaneous basal AP firing rate. In contrast, the selective PP2A inhibitor okadaic acid (100 nmol/L) had no significant effect on spontaneous AP firing, LCR parameters, or PLB phosphorylation. Application of purified PP1 to permeabilized SANC suppressed LCR, whereas purified PP2A had no effect on LCR characteristics. Our numerical model simulations demonstrated that PP inhibition increases AP firing rate via a coupled-clock mechanism, including respective increases in the SR Ca<sup>2+</sup> pumping rate, L-type Ca<sup>2+</sup> current, and Na<sup>+</sup>/Ca<sup>2+</sup>-exchanger current. Thus, PP1 and its endogenous inhibitors modulate the basal spontaneous firing rate of cardiac pacemaker cells by suppressing SR Ca<sup>2+</sup> cycling protein phosphorylation, the SR Ca<sup>2+</sup> load and LCRs, and L-type Ca<sup>2+</sup> current.
format	article
author	Syevda Tagirova Sirenko Ihor Zahanich Yue Li Yevgeniya O. Lukyanenko Alexey E. Lyashkov Bruce D. Ziman Kirill V. Tarasov Antoine Younes Daniel R. Riordon Yelena S. Tarasova Dongmei Yang Tatiana M. Vinogradova Victor A. Maltsev Edward G. Lakatta
author_facet	Syevda Tagirova Sirenko Ihor Zahanich Yue Li Yevgeniya O. Lukyanenko Alexey E. Lyashkov Bruce D. Ziman Kirill V. Tarasov Antoine Younes Daniel R. Riordon Yelena S. Tarasova Dongmei Yang Tatiana M. Vinogradova Victor A. Maltsev Edward G. Lakatta
author_sort	Syevda Tagirova Sirenko
title	Phosphoprotein Phosphatase 1 but Not 2A Activity Modulates Coupled-Clock Mechanisms to Impact on Intrinsic Automaticity of Sinoatrial Nodal Pacemaker Cells
title_short	Phosphoprotein Phosphatase 1 but Not 2A Activity Modulates Coupled-Clock Mechanisms to Impact on Intrinsic Automaticity of Sinoatrial Nodal Pacemaker Cells
title_full	Phosphoprotein Phosphatase 1 but Not 2A Activity Modulates Coupled-Clock Mechanisms to Impact on Intrinsic Automaticity of Sinoatrial Nodal Pacemaker Cells
title_fullStr	Phosphoprotein Phosphatase 1 but Not 2A Activity Modulates Coupled-Clock Mechanisms to Impact on Intrinsic Automaticity of Sinoatrial Nodal Pacemaker Cells
title_full_unstemmed	Phosphoprotein Phosphatase 1 but Not 2A Activity Modulates Coupled-Clock Mechanisms to Impact on Intrinsic Automaticity of Sinoatrial Nodal Pacemaker Cells
title_sort	phosphoprotein phosphatase 1 but not 2a activity modulates coupled-clock mechanisms to impact on intrinsic automaticity of sinoatrial nodal pacemaker cells
publisher	MDPI AG
publishDate	2021
url	https://doaj.org/article/d34e3e9a556b4610af29e09b6ffdc8e6
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Phosphoprotein Phosphatase 1 but Not 2A Activity Modulates Coupled-Clock Mechanisms to Impact on Intrinsic Automaticity of Sinoatrial Nodal Pacemaker Cells

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