Protein Kinase A Increases Type-2 Inositol 1,4,5-Trisphosphate Receptor Activity by Phosphorylation of Serine 937
Department of Pharmacology and Physiology, School of Medicine and Dentistry, University of Rochester Medical Center, Rochester, New York 14642, USA. Journal of Biological Chemistry
(Impact Factor: 4.57).
08/2009; 284(37):25116-25. DOI: 10.1074/jbc.M109.010132
Protein kinase A (PKA) phosphorylation of inositol 1,4,5-trisphosphate receptors (InsP(3)Rs) represents a mechanism for shaping intracellular Ca(2+) signals following a concomitant elevation in cAMP. Activation of PKA results in enhanced Ca(2+) release in cells that express predominantly InsP(3)R2. PKA is known to phosphorylate InsP(3)R2, but the molecular determinants of this effect are not known. We have expressed mouse InsP(3)R2 in DT40-3KO cells that are devoid of endogenous InsP(3)R and examined the effects of PKA phosphorylation on this isoform in unambiguous isolation. Activation of PKA increased Ca(2+) signals and augmented the single channel open probability of InsP(3)R2. A PKA phosphorylation site unique to the InsP(3)R2 was identified at Ser(937). The enhancing effects of PKA activation on this isoform required the phosphorylation of Ser(937), since replacing this residue with alanine eliminated the positive effects of PKA activation. These results provide a mechanism responsible for the enhanced Ca(2+) signaling following PKA activation in cells that express predominantly InsP(3)R2.
Available from: Syed Zahid Ali Shah
- "Controlled tyrosine-digested mutational analysis revealed a novel protein kinase A (PKA) phosphorylation site S937 in IP3R2 (Fig. 1) (Betzenhauser et al. 2009a, b). DT40- TKO cell study revealed that S937 is a bonafide phosphorylation site for protein kinase A (PKA) in IP3R2 (Betzenhauser et al. 2009a, b). The pancreatic acinar cells AR4-2 J also showed increased activity of IP3R2 and inositol-induced calcium release (IICR) (Regimbald-Dumas et al. 2007). "
Available from: James Kevin Foskett
- "Furthermore, nuclear patch-clamping eliminates cumbersome procedures required to extract InsP 3 R and enables rapid, reversible, and reliable exchange of solutions on one side of the InsP 3 R channel. Recently, the discovery that, in some cell types, InsP 3 R channels are localized to the plasma membrane at very low densities (<5 per cell) has made it possible to perform single-channel studies on these channels by whole-cell patch-clamping (Dellis et al. 2006Dellis et al. , 2008Betzenhauser et al. 2008Betzenhauser et al. , 2009aSchug et al. 2008;Wagner et al. 2008;Gin et al. 2009). However, the channels studied were in the plasma membrane, with protein and lipid environments different from those in the ER. "
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ABSTRACT: The modulation of cytoplasmic free Ca(2+) concentration ([Ca(2+)]i) is a universal intracellular signaling pathway that regulates numerous cellular physiological processes. Ubiquitous intracellular Ca(2+)-release channels localized to the endoplasmic/sarcoplasmic reticulum-inositol 1,4,5-trisphosphate receptor (InsP3R) and ryanodine receptor (RyR) channels-play a central role in [Ca(2+)]i signaling in all animal cells. Despite their intracellular localization, electrophysiological studies of the single-channel permeation and gating properties of these Ca(2+)-release channels using the powerful patch-clamp approach have been possible by application of this technique to isolated nuclei because the channels are present in membranes of the nuclear envelope. Here we provide a concise description of how nuclear patch-clamp experiments have been used to study single-channel properties of different InsP3R channels in the outer nuclear membrane. We compare this with other methods for studying intracellular Ca(2+) release. We also briefly describe application of the technique to InsP3R channels in the inner nuclear membrane and to channels in the outer nuclear membrane of HEK293 cells expressing recombinant RyR.
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