Role of protein kinase C in phospholemman mediated regulation of α2β1 isozyme of Na+/K+-ATPase in caveolae of pulmonary artery smooth muscle cells

Dey, Kuntal; Roy, Subhasish; Ghosh, Biswarup; Chakraborti, Sajal

doi:10.1016/j.biochi.2011.12.020

Cited by 14 publications

(19 citation statements)

References 44 publications

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“…PKC has been reported to increase Na + /K + -ATPase activity in other cell types including arterial smooth muscle [53] and cardiac myocytes [54]. However, unlike with SERCA, there is no evidence for phosphorylation of this protein in the resting platelet phosphoproteome [47], although given the need for activation with a strong platelet activator for the Na + /K + -ATPase to be influenced by PKC, phosphorylation of this protein might not be expected under resting conditions.…”

Section: Discussionmentioning

confidence: 99%

Conventional protein kinase C isoforms differentially regulate ADP- and thrombin-evoked Ca2+ signalling in human platelets

et al. 2015

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Rises in cytosolic Ca 2+ concentration ([Ca 2+ ] cyt ) are central in platelet activation, yet many aspects of the underlying mechanisms are poorly understood. Most studies examine how experimental manipulations affect agonist-evoked rises in [Ca 2+ ] cyt , but these only monitor the net effect of manipulations on the processes controlling [Ca 2+ ] cyt (Ca 2+ buffering, sequestration, release, entry and removal), and cannot resolve the source of the Ca 2+ or the transporters or channels affected. To investigate the effects of protein kinase C (PKC) on platelet Ca 2+ signalling, we here monitor Ca 2+ flux around the platelet by measuring net Ca 2+ fluxes to or from the extracellular space and the intracellular Ca 2+ stores, which act as the major sources and sinks for Ca 2+ influx into and efflux from the cytosol, as well as monitoring the cytosolic Na + concentration ([Na + ] cyt ), which influences platelet Ca 2+ fluxes via Na + /Ca 2+ exchange. The intracellular store Ca 2+ concentration ([Ca 2+ ] st ) was monitored using Fluo-5N, the extracellular Ca 2+ concentration ([Ca 2+ ] ext ) was monitored using Fluo-4 whilst [Ca 2+ ] cyt and [Na + ] cyt were monitored using Fura-2 and SFBI respectively. PKC inhibition using Ro-31-8220 or bisindolaemide I potentiated ADP-and thrombin-evoked rises in [Ca 2+ ] cyt in the absence of extracellular Ca 2+ . PKC inhibition potentiated ADP-evoked but reduced thrombin-evoked intracellular Ca 2+ release and Ca 2+ removal into the extracellular medium. SERCA inhibition using thapsigargin and 2,5-di(tert-butyl) l,4-benzohydroquinone abolished the effect of PKC inhibitors on ADP-evoked changes in [Ca 2+ ] cyt but only reduced the effect on thrombin-evoked responses. Thrombin evokes substantial rises in [Na + ] cyt which would be expected to reduce Ca 2+ removal via the Na + /Ca 2+ exchanger (NCX). Thrombinevoked rises in [Na + ] cyt were potentiated by PKC inhibition, an effect which was not due to altered changes in non-selective cation permeability of the plasma membrane as assessed by Mn 2+ quench of Fura-2 fluorescence. PKC inhibition was without effect on thrombin-evoked rises in [Ca 2+ ] cyt following SERCA inhibition and either removal of extracellular Na + or inhibition of Na + /K + -ATPase activity by removal of extracellular K + or treatment with digoxin. These data suggest that PKC limits ADP-evoked rises in [Ca 2+ ] cyt by acceleration of SERCA activity, whilst rises in [Ca 2+ ] cyt evoked by the stronger platelet activator thrombin are limited by PKC through acceleration of both SERCA and Na + /K + -ATPase activity, with the latter limiting the effect of thrombin on rises in [Na + ] cyt and so forward mode NCX activity. The use of selective PKC inhibitors indicated that conventional and not novel PKC isoforms are responsible for the inhibition of agonist-evoked Ca 2+ signalling.

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Section: Discussionmentioning

confidence: 99%

Conventional protein kinase C isoforms differentially regulate ADP- and thrombin-evoked Ca2+ signalling in human platelets

et al. 2015

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“…NKA b 1 -subunit and a-subunit [19]. Custom made anti-PLM antibody was prepared according to Dey et al [11].…”

Section: Methodsmentioning

confidence: 99%

“…The purified fraction containing both the a 1 b 1 and a 2 b 1 isozymes of NKA was reconstituted with or without PLM into the liposomes which were prepared using the lipid DOPC by following the procedure of Dey et al [11]. Two other liposomal systems were prepared which were reconstituted with PLM, 70 kDa inhibitor and either with a 1 b 1 or with a 2 b 1 isozymes of NKA.…”

Section: Reconstitution Of Nka and Plm Into Liposomesmentioning

confidence: 99%

“…Recently, we have reported the presence of PLM in smooth muscle cells [11] and hence we investigated its involvement in this scenario. Huang et al [18] showed that different isoforms of NKA are differentially inhibited by oxidants.…”

Section: Introductionmentioning

confidence: 99%

“…BPASMCs contain both the a 1 and a 2 isoforms of NKA [11], therefore, one of our aims was to investigate the isoform specific regulation of NKA by oxidants and to explore the underlying mechanisms. In view of our recent report about the presence of a 70 kDa inhibitor of NKA [16], we also determined its role in regulating the enzyme activity upon stimulation of the cells with U46619.…”

Section: Introductionmentioning

confidence: 99%

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Role of phospholemman and the 70 kDa inhibitor protein in regulating Na⁺/K⁺ ATPase activity in pulmonary artery smooth muscle cells under U46619 stimulation

Dey¹,

Rahaman²,

Chakraborti³

et al. 2013

FEBS Letters

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a b s t r a c tTreatment of bovine pulmonary smooth muscle cells with U46619 inhibited the Na + /K + ATPase activity in two parallel pathways: one of which is mediated via glutathionylation of the pump and the other by augmenting the inhibitory activity of the 70 kDa inhibitor protein of Na + /K + ATPase. Although phospholemman deglutathionylates the pump leading to its activation, the inhibitor is responsible for irreversible inhibition of Na + /K + ATPase in an isoform specific manner during treatment of the cells with U46619.

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Na+/K+-ATPase: A Perspective

Chakraborti

Rahaman

Alam

et al. 2015

Regulation of Membrane Na+-K+ ATPase

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Role of protein kinase C in phospholemman mediated regulation of α2β1 isozyme of Na+/K+-ATPase in caveolae of pulmonary artery smooth muscle cells

Cited by 14 publications

References 44 publications

Conventional protein kinase C isoforms differentially regulate ADP- and thrombin-evoked Ca2+ signalling in human platelets

Conventional protein kinase C isoforms differentially regulate ADP- and thrombin-evoked Ca2+ signalling in human platelets

Role of phospholemman and the 70 kDa inhibitor protein in regulating Na⁺/K⁺ ATPase activity in pulmonary artery smooth muscle cells under U46619 stimulation

Na+/K+-ATPase: A Perspective

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Role of protein kinase C in phospholemman mediated regulation of α2β1 isozyme of Na+/K+-ATPase in caveolae of pulmonary artery smooth muscle cells

Cited by 14 publications

References 44 publications

Conventional protein kinase C isoforms differentially regulate ADP- and thrombin-evoked Ca2+ signalling in human platelets

Conventional protein kinase C isoforms differentially regulate ADP- and thrombin-evoked Ca2+ signalling in human platelets

Role of phospholemman and the 70 kDa inhibitor protein in regulating Na+/K+ ATPase activity in pulmonary artery smooth muscle cells under U46619 stimulation

Na+/K+-ATPase: A Perspective

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Role of phospholemman and the 70 kDa inhibitor protein in regulating Na⁺/K⁺ ATPase activity in pulmonary artery smooth muscle cells under U46619 stimulation