H-89, a Non-Specific Inhibitor of Protein Kinase A, Promotes Post-Ischemic Cardiac Contractile Recovery and Reduces Infarct Size

2005 ◽  
Vol 45 (4) ◽  
pp. 341-347 ◽  
Author(s):  
Siyanda Makaula ◽  
Amanda Lochner ◽  
Sonia Genade ◽  
Michael N Sack ◽  
Moneeb M Awan ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Infarct Size ◽  
Protein Kinase A ◽  
Specific Inhibitor ◽  
Cardiac Contractile ◽  
Contractile Recovery ◽  
Kinase A

scholarly journals Negative Regulation of Parathyroid Hormone (PTH)-Activated Phospholipase C by PTH/PTH-Related Peptide Receptor Phosphorylation and Protein Kinase A

Endocrinology ◽  
2008 ◽  
Vol 149 (8) ◽  
pp. 4016-4023 ◽  
Author(s):  
Hesham A. W. Tawfeek ◽  
Abdul B. Abou-Samra
Keyword(s):  
Protein Kinase ◽  
Adenylate Cyclase ◽  
Protein Kinase A ◽  
Phospholipase C ◽  
Specific Inhibitor ◽  
Dependent Manner ◽  
Receptor Phosphorylation ◽  
Pthrp Receptor ◽  
Dose Dependent ◽  
Kinase A

PTH binding to the PTH/PTHrP receptor activates adenylate cyclase/protein kinase A (PKA) and phospholipase C (PLC) pathways and increases receptor phosphorylation. The mechanisms regulating PTH activation of PLC signaling are poorly understood. In the current study, we explored the role of PTH/PTHrP receptor phosphorylation and PKA in PTH activation of PLC. When treated with PTH, LLCPK-1 cells stably expressing a green fluorescent protein (GFP)-tagged wild-type (WT) PTH/PTHrP receptor show a small dose-dependent increase in PLC signaling as measured by inositol trisphosphate accumulation assay. In contrast, PTH treatment of LLCPK-1 cells stably expressing a GFP-tagged receptor mutated in its carboxyl-terminal tail so that it cannot be phosphorylated (PD-GFP) results in significantly higher PLC activation (P < 0.001). The effects of PTH on PLC activation are dose dependent and reach maximum at the 100 nm PTH dose. When WT receptor-expressing cells are pretreated with H89, a specific inhibitor of PKA, PTH activation of PLC signaling is enhanced in a dose-dependent manner. H89 pretreatment in PD-GFP cells causes a further increase in PLC activation in response to PTH treatment. Interestingly, PTH and forskolin (adenylate cyclase/PKA pathway activator) treatment causes an increase in PLCβ3 phosphorylation at the Ser1105 inhibitory site and that increase is blocked by the PKA inhibitor, H89. Expression of a mutant PLCβ3 in which Ser1105 was mutated to alanine (PLCβ3-SA), in WT or PD cells increases PTH stimulation of inositol 1,4,5-trisphosphate formation. Altogether, these data suggest that PTH signaling to PLC is negatively regulated by PTH/PTHrP receptor phosphorylation and PKA. Furthermore, phosphorylation at Ser1105 is demonstrated as a regulatory mechanism of PLCβ3 by PKA.

Protein kinase A phosphorylation enhances sodium channel currents in Xenopus oocytes

1992 ◽  
Vol 263 (3) ◽  
pp. C660-C666 ◽  
Author(s):  
R. D. Smith ◽  
A. L. Goldin
Keyword(s):  
Protein Kinase ◽  
Sodium Channel ◽  
Protein Kinase A ◽  
Rat Brain ◽  
Xenopus Oocytes ◽  
Kinase Inhibitor ◽  
Sodium Current ◽  
Specific Inhibitor ◽  
Channel Currents ◽  
Kinase A

The voltage-sensitive rat brain sodium channel is known to be phosphorylated by adenosine 3',5'-cyclic monophosphate (cAMP)-dependent protein kinase A (PKA), but the functional significance of that phosphorylation is unknown. We have shown that rat brain sodium channel currents expressed in Xenopus oocytes were enhanced by induction of PKA activity. Stimulation of the beta 2-adrenergic receptor or treatment with dibutyryl cAMP resulted in increased sodium current amplitudes without affecting the voltage dependence of channel activation or inactivation. These increases were completely blocked by preinjection of protein kinase inhibitor, a specific inhibitor of PKA. Injection of phosphatase into the oocytes resulted in a significant decrease in sodium current amplitude, indicating that phosphorylation is important for basal levels of sodium channel activity in oocytes. The enhancement was specific for the rat brain IIA sodium channel, because currents expressed from the rat muscle microI sodium channel were not enhanced by the same procedures. These data demonstrate a modulatory role of PKA phosphorylation on brain sodium channel function and suggest a means by which the electrical excitability of cells may be regulated.

scholarly journals Ser-2030, but not Ser-2808, is the major phosphorylation site in cardiac ryanodine receptors responding to protein kinase A activation upon β-adrenergic stimulation in normal and failing hearts

2006 ◽  
Vol 396 (1) ◽  
pp. 7-16 ◽  
Author(s):  
Bailong Xiao ◽  
Guofeng Zhong ◽  
Masakazu Obayashi ◽  
Dongmei Yang ◽  
Keyun Chen ◽  
...  
Keyword(s):  
Heart Failure ◽  
Protein Kinase ◽  
Protein Kinase A ◽  
Ryanodine Receptors ◽  
Phosphorylation Site ◽  
Specific Inhibitor ◽  
Dependent Manner ◽  
Adrenergic Stimulation ◽  
Rat Hearts ◽  
Kinase A

We have recently shown that RyR2 (cardiac ryanodine receptor) is phosphorylated by PKA (protein kinase A/cAMP-dependent protein kinase) at two major sites, Ser-2030 and Ser-2808. In the present study, we examined the properties and physiological relevance of phosphorylation of these two sites. Using site- and phospho-specific antibodies, we demonstrated that Ser-2030 of both recombinant and native RyR2 from a number of species was phosphorylated by PKA, indicating that Ser-2030 is a highly conserved PKA site. Furthermore, we found that the phosphorylation of Ser-2030 responded to isoproterenol (isoprenaline) stimulation in rat cardiac myocytes in a concentration- and time-dependent manner, whereas Ser-2808 was already substantially phosphorylated before β-adrenergic stimulation, and the extent of the increase in Ser-2808 phosphorylation after β-adrenergic stimulation was much less than that for Ser-2030. Interestingly, the isoproterenol-induced phosphorylation of Ser-2030, but not of Ser-2808, was markedly inhibited by PKI, a specific inhibitor of PKA. The basal phosphorylation of Ser-2808 was also insensitive to PKA inhibition. Moreover, Ser-2808, but not Ser-2030, was stoichiometrically phosphorylated by PKG (protein kinase G). In addition, we found no significant phosphorylation of RyR2 at the Ser-2030 PKA site in failing rat hearts. Importantly, isoproterenol stimulation markedly increased the phosphorylation of Ser-2030, but not of Ser-2808, in failing rat hearts. Taken together, these observations indicate that Ser-2030, but not Ser-2808, is the major PKA phosphorylation site in RyR2 responding to PKA activation upon β-adrenergic stimulation in both normal and failing hearts, and that RyR2 is not hyperphosphorylated by PKA in heart failure. Our results also suggest that phosphorylation of RyR2 at Ser-2030 may be an important event associated with altered Ca2+ handling and cardiac arrhythmia that is commonly observed in heart failure upon β-adrenergic stimulation.

scholarly journals Regulation of renal Na(+),K(+)-ATPase and ouabain-sensitive H(+),K(+)-ATPase by the cyclic AMP-protein kinase A signal transduction pathway.

2003 ◽  
Vol 50 (1) ◽  
pp. 103-114 ◽  
Author(s):  
Jerzy Bełtowski ◽  
Andrzej Marciniak ◽  
Grazyna Wójcicka ◽  
Dionizy Górny
Keyword(s):  
Cytochrome P450 ◽  
Protein Kinase ◽  
Cyclic Amp ◽  
Atpase Activity ◽  
Protein Kinase A ◽  
Plasma Membranes ◽  
Renal Cortex ◽  
Signal Transduction Pathway ◽  
Specific Inhibitor ◽  
Kinase A

We investigated the effect of the cyclic AMP-protein kinase A (PKA) signalling pathway on renal Na(+),K(+)-ATPase and ouabain-sensitive H(+),K(+)-ATPase. Male Wistar rats were anaesthetized and catheter was inserted through the femoral artery into the abdominal aorta proximally to the renal arteries for infusion of the investigated substances. Na(+),K(+)-ATPase activity was measured in the presence of Sch 28080 to block ouabain-sensitive H(+),K(+)-ATPase and improve specificity of the assay. Dibutyryl-cyclic AMP (db-cAMP) administered at a dose of 10(-7) mol/kg per min and 10(-6) mol/kg per min increased Na(+),K(+)-ATPase activity in the renal cortex by 34% and 42%, respectively, and decreased it in the renal medulla by 30% and 44%, respectively. db-cAMP infused at 10(-6) mol/kg per min increased the activity of cortical ouabain-sensitive H(+),K(+)-ATPase by 33%, and medullary ouabain-sensitive H(+),K(+)-ATPase by 30%. All the effects of db-cAMP were abolished by a specific inhibitor of protein kinase A, KT 5720. The stimulatory effect on ouabain-sensitive H(+),K(+)-ATPase and on cortical Na(+),K(+)-ATPase was also abolished by brefeldin A which inhibits the insertion of proteins into the plasma membranes, whereas the inhibitory effect on medullary Na(+),K(+)-ATPase was partially attenuated by 17-octadecynoic acid, an inhibitor of cytochrome p450-dependent arachidonate metabolism. We conclude that the cAMP-PKA pathway stimulates Na(+),K(+)-ATPase in the renal cortex as well as ouabain-sensitive H(+),K(+)-ATPase in the cortex and medulla by a mechanism requiring insertion of proteins into the plasma membrane. In contrast, medullary Na(+),K(+)-ATPase is inhibited by cAMP through a mechanism involving cytochrome p450-dependent arachidonate metabolites.

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