PKA-mediated phosphorylation and inhibition of Na+-K+-ATPase in response to β-adrenergic hormone

Xian Jun Cheng, Gilberto Fisone, Oleg Aizman, Roman Aizman, Robert Levenson, Paul Greengard, Anita Aperia

Research output: Contribution to journalArticle

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Abstract

The activity of Na+-K+-ATPase can be regulated by hormones that activate adenosine 3',5'-cyclic monophosphate-dependent protein kinase (PKA). Here, using a site-directed phosphorylation state-specific antibody, we show that hormonal regulation of Na+-K+-ATPase can occur via phosphorylation of Ser-943 on its α-subunit. cDNAs coding for wild-type rat Na+-K+-ATPase and Na+-K+-ATPase in which the PKA phosphorylation site Ser-943 was mutated to Ala were stably and transiently transfected into COS cells. In COS cells expressing wild-type Na+-K+-ATPase the β-adrenergic agonist isoproterenol (1 μM) significantly increased the level of phosphorylation of the α- subunit. Phosphorylation was accompanied by a significant inhibition of the enzyme activity, as reflected by a decrease in ATP hydrolysis and 86Rb+ transport. The effect of isoproterenol was reproduced by the PKA activator forskolin used in combination with the phosphodiesterase inhibitor 3- isobutyl-1-methylxanthine and was abolished by the specific PKA inhibitor H- 89. Okadaic acid, an inhibitor of protein phosphatases 1 and 2A, enhanced phosphorylation and inhibition of Na+-K+-ATPase induced by isoproterenol. The changes in activity of Na+-K+-ATPase linearly correlated with the extent of the α-subunit of Na+-K+-ATPase being phosphorylated. When Ser- 943 was replaced by alanine, stimulation of the phosphorylation and inhibition of the activity of Na+-K+-ATPase induced by isoproterenol, alone or in combination with okadaic acid, were not observed. These results indicate that, in intact cells, modulation of the activity of Na+-K+- ATPase can be achieved by regulation of the state of phosphorylation of Ser- 943. Moreover, they provide a biochemical mechanism by which β-adrenergic agonists can regulate Na+-K+-ATPase activity.

Original languageEnglish (US)
Pages (from-to)C893-C901
JournalAmerican Journal of Physiology - Cell Physiology
Volume273
Issue number3 42-3
StatePublished - Sep 1 1997

Fingerprint

Adrenergic Agents
Protein Kinases
Phosphorylation
Hormones
Isoproterenol
Okadaic Acid
Adrenergic Agonists
COS Cells
sodium-translocating ATPase
Phospho-Specific Antibodies
1-Methyl-3-isobutylxanthine
Protein Phosphatase 2
Phosphodiesterase Inhibitors
Colforsin
Protein Kinase Inhibitors
Alanine
Cyclic AMP
Hydrolysis
Complementary DNA
Adenosine Triphosphate

All Science Journal Classification (ASJC) codes

  • Physiology
  • Cell Biology

Cite this

Cheng, X. J., Fisone, G., Aizman, O., Aizman, R., Levenson, R., Greengard, P., & Aperia, A. (1997). PKA-mediated phosphorylation and inhibition of Na+-K+-ATPase in response to β-adrenergic hormone. American Journal of Physiology - Cell Physiology, 273(3 42-3), C893-C901.
Cheng, Xian Jun ; Fisone, Gilberto ; Aizman, Oleg ; Aizman, Roman ; Levenson, Robert ; Greengard, Paul ; Aperia, Anita. / PKA-mediated phosphorylation and inhibition of Na+-K+-ATPase in response to β-adrenergic hormone. In: American Journal of Physiology - Cell Physiology. 1997 ; Vol. 273, No. 3 42-3. pp. C893-C901.
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Cheng, XJ, Fisone, G, Aizman, O, Aizman, R, Levenson, R, Greengard, P & Aperia, A 1997, 'PKA-mediated phosphorylation and inhibition of Na+-K+-ATPase in response to β-adrenergic hormone', American Journal of Physiology - Cell Physiology, vol. 273, no. 3 42-3, pp. C893-C901.

PKA-mediated phosphorylation and inhibition of Na+-K+-ATPase in response to β-adrenergic hormone. / Cheng, Xian Jun; Fisone, Gilberto; Aizman, Oleg; Aizman, Roman; Levenson, Robert; Greengard, Paul; Aperia, Anita.

In: American Journal of Physiology - Cell Physiology, Vol. 273, No. 3 42-3, 01.09.1997, p. C893-C901.

Research output: Contribution to journalArticle

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T1 - PKA-mediated phosphorylation and inhibition of Na+-K+-ATPase in response to β-adrenergic hormone

AU - Cheng, Xian Jun

AU - Fisone, Gilberto

AU - Aizman, Oleg

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AU - Greengard, Paul

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N2 - The activity of Na+-K+-ATPase can be regulated by hormones that activate adenosine 3',5'-cyclic monophosphate-dependent protein kinase (PKA). Here, using a site-directed phosphorylation state-specific antibody, we show that hormonal regulation of Na+-K+-ATPase can occur via phosphorylation of Ser-943 on its α-subunit. cDNAs coding for wild-type rat Na+-K+-ATPase and Na+-K+-ATPase in which the PKA phosphorylation site Ser-943 was mutated to Ala were stably and transiently transfected into COS cells. In COS cells expressing wild-type Na+-K+-ATPase the β-adrenergic agonist isoproterenol (1 μM) significantly increased the level of phosphorylation of the α- subunit. Phosphorylation was accompanied by a significant inhibition of the enzyme activity, as reflected by a decrease in ATP hydrolysis and 86Rb+ transport. The effect of isoproterenol was reproduced by the PKA activator forskolin used in combination with the phosphodiesterase inhibitor 3- isobutyl-1-methylxanthine and was abolished by the specific PKA inhibitor H- 89. Okadaic acid, an inhibitor of protein phosphatases 1 and 2A, enhanced phosphorylation and inhibition of Na+-K+-ATPase induced by isoproterenol. The changes in activity of Na+-K+-ATPase linearly correlated with the extent of the α-subunit of Na+-K+-ATPase being phosphorylated. When Ser- 943 was replaced by alanine, stimulation of the phosphorylation and inhibition of the activity of Na+-K+-ATPase induced by isoproterenol, alone or in combination with okadaic acid, were not observed. These results indicate that, in intact cells, modulation of the activity of Na+-K+- ATPase can be achieved by regulation of the state of phosphorylation of Ser- 943. Moreover, they provide a biochemical mechanism by which β-adrenergic agonists can regulate Na+-K+-ATPase activity.

AB - The activity of Na+-K+-ATPase can be regulated by hormones that activate adenosine 3',5'-cyclic monophosphate-dependent protein kinase (PKA). Here, using a site-directed phosphorylation state-specific antibody, we show that hormonal regulation of Na+-K+-ATPase can occur via phosphorylation of Ser-943 on its α-subunit. cDNAs coding for wild-type rat Na+-K+-ATPase and Na+-K+-ATPase in which the PKA phosphorylation site Ser-943 was mutated to Ala were stably and transiently transfected into COS cells. In COS cells expressing wild-type Na+-K+-ATPase the β-adrenergic agonist isoproterenol (1 μM) significantly increased the level of phosphorylation of the α- subunit. Phosphorylation was accompanied by a significant inhibition of the enzyme activity, as reflected by a decrease in ATP hydrolysis and 86Rb+ transport. The effect of isoproterenol was reproduced by the PKA activator forskolin used in combination with the phosphodiesterase inhibitor 3- isobutyl-1-methylxanthine and was abolished by the specific PKA inhibitor H- 89. Okadaic acid, an inhibitor of protein phosphatases 1 and 2A, enhanced phosphorylation and inhibition of Na+-K+-ATPase induced by isoproterenol. The changes in activity of Na+-K+-ATPase linearly correlated with the extent of the α-subunit of Na+-K+-ATPase being phosphorylated. When Ser- 943 was replaced by alanine, stimulation of the phosphorylation and inhibition of the activity of Na+-K+-ATPase induced by isoproterenol, alone or in combination with okadaic acid, were not observed. These results indicate that, in intact cells, modulation of the activity of Na+-K+- ATPase can be achieved by regulation of the state of phosphorylation of Ser- 943. Moreover, they provide a biochemical mechanism by which β-adrenergic agonists can regulate Na+-K+-ATPase activity.

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