Generation of Calcium Oscillations in Fibroblasts by Positive Feedback Between Calcium and IP
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Harootunian, A.; Kao, Joseph P. Y.; Paranjape, Suman M.; Tsien, Roger Y.

doi:10.1126/science.1986413

Cited by 307 publications

(163 citation statements)

References 33 publications

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“…Our results imply that Ca 2ϩ is acting within this pathway to regulate phospholipase C␤. Indeed many studies have described positive feedback by Ca 2ϩ on G q -coupled receptor signaling pathways (25)(26)(27)(28)(29). Our findings do not rule out a role of Ca 2ϩ released via IP 3 receptor stimulation acting on a distinct phospholipase C subtype, such as phospholipase C␦, but they confirm a role of Ca 2ϩ in regulating DAG production as a whole.…”

Section: Discussionsupporting

confidence: 75%

Calcium-dependent Regulation of Protein Kinase D Revealed by a Genetically Encoded Kinase Activity Reporter

Kunkel¹,

Toker

Tsien

et al. 2007

Journal of Biological Chemistry

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Protein kinase D (PKD) regulates many diverse cellular functions in response to diacylglycerol. To monitor PKD signaling in live cells, we generated a genetically encoded fluorescent reporter for PKD activity, DKAR (D kinase activity reporter). DKAR expressed in mammalian cells undergoes reversible fluorescence resonance energy transfer changes upon activation and inhibition of endogenous PKD. Surprisingly, we find that agonist-evoked activation of PKD is driven not only by diacylglycerol production, but by Ca 2؉ . Furthermore, elevation of intracellular Ca 2؉ , in the absence of any other stimulus, is sufficient to activate PKD. Concurrent imaging of Ca 2؉ , diacylglycerol, and PKD activity reveals that thapsigargin-mediated elevation of intracellular Ca 2؉ is closely followed by a robust increase in diacylglycerol production, in turn followed by PKD activation. The Ca 2؉ -induced production of diacylglycerol and accompanying PKD activation is dependent on phospholipase C activity. These data reveal that Ca 2؉ is a major contributor to the initiation of PKD signaling through positive feedback regulation of diacylglycerol production, unveiling a new mechanism in PKD activation.Protein kinase D (PKD) 2 comprises a family of three isoforms belonging to the Ca 2ϩ /calmodulin-dependent kinase group of serine/threonine protein kinases. PKD plays a role in numerous processes, including cell proliferation, cell survival, immune cell signaling, gene expression, vesicle trafficking, and neuronal development (1). PKD transduces signals that generate the second messenger diacylglycerol (DAG). This ligand has two roles in the activation of PKD: it activates novel protein kinase C (PKC) family members, which catalyze an activating phosphorylation of PKD, and it directly binds PKD thus recruiting it to the membrane.PKD isoforms comprise a conserved catalytic core and N-terminal regulatory moiety. The regulatory region contains two cysteine-rich (C1) domains and a pleckstrin homology domain, and this region as a whole acts in an inhibitory manner on the kinase (2). C1 domains are membrane-targeting modules that typically bind DAG and the functional analogues, phorbol esters (3). They are found in a number of proteins, most notably PKC, and provide a mechanism for proteins to be reversibly recruited to membranes in response to DAG. In the case of PKD, binding to either phorbol ester or DAG results in its membrane recruitment and activation.In addition to membrane recruitment by DAG, activation of PKD requires phosphorylation at two sites within its catalytic core (4). Thus, although DAG production leads to activation of PKD, it is not simply through the C1-mediated membrane binding and removal of autoinhibition by the regulatory region by which PKD becomes active. In addition, the upstream kinases, the novel PKCs, must phosphorylate PKD within its activation loop at Ser-744 and Ser-748 to promote its activity. This phosphorylation event is the rate-limiting step in PKD activation, and once phosphorylated, PKD remains active eve...

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Section: Discussionsupporting

confidence: 75%

Calcium-dependent Regulation of Protein Kinase D Revealed by a Genetically Encoded Kinase Activity Reporter

Kunkel¹,

Toker

Tsien

et al. 2007

Journal of Biological Chemistry

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“…[Ca2+]i oscillations, the majority of which involve the repetitive emptying and refilling of intracellular calcium stores initiated by Ins 1,4,5P3 (Berridge & Galione, 1990;Berridge, 1990;Harootunian et al, 1991). The maintained response induced by collagen may therefore reflect a supramaximal and sustained formation of Ins 1,4,5P3.…”

Section: Discussionmentioning

confidence: 99%

Regulation of cytosolic calcium by collagen in single human platelets

Poole

Watson

1995

British J Pharmacology

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There is controversy in the literature as to whether collagen is able to induce directly a rise in cytosolic calcium concentration ([Ca2+]i) in human platelets. We have addressed this question by observing the cytosolic calcium response of single fura‐2‐loaded human platelets settling onto a collagen‐coated surface using dynamic fluorescence ratio imaging. Following a short lag phase after adherence to collagen fibres, platelets underwent a rapid rise in cytosolic calcium from basal values of 80 ± 13 nM (n = 24) to a peak of 475 ± 42 nM (n = 24) which was sustained for the remaining period of the experiment. The tyrphostin protein tyrosine kinase inhibitor, ST271, reduced substantially the proportion of platelets which exhibited a rise in [Ca2+]i on adherence to collagen and transformed the response in remaining cells to one of oscillations. In contrast, and as a control for collagen, laminin‐coated surfaces induced adherence of human platelets without elevating intracellular [Ca2+]; the cells however remained responsive to ADP. We conclude that collagen directly induces a rise in cytosolic calcium in single human platelets through a tyrosine kinase‐mediated pathway.

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“…It has been reported before that fibroblasts can possess L-type Ca2+ channels [8,9,11,12], but a function for such channels in these cells has so far been unclear. Here, we show that NRK fibroblasts also possess L-type voltage-dependent Ca2+ channels (Fig.…”

Section: The Involvement Of L-type Ca2+ Channels In the Upstroke Of Tmentioning

confidence: 99%

“…Although considered to be inexcitable, it has been reported that fibroblasts do possess voltage-dependent Ca2+ channels [8][9][10][11][12], but their function in these cells has so far been unclear. Fibroblasts can also be electrically coupled via gap junctions [13][14][15][16][17].…”

Section: Introductionmentioning

confidence: 99%

Synchronized calcium spiking resulting from spontaneous calcium action potentials in monolayers of NRK fibroblasts

et al. 1997

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SummaryThe correlation between the intracellular Ca2+ concentration ([Ca2+].) and membrane potential in monolayers of density-arrested normal rat kidney (NRK) fibroblasts was investigated. Using the fluorescent probe Fura-2, spontaneous repetitive spike-like increases in [Ca2+]. (Ca2+ spikes) were observed that were synchronised throughout the entire monolayer. Ca2+ spikes disappeared in Ca2+-free solutions and could be blocked by the L-type Ca2* channel antagonist felodipine. Simultaneous measurements of [Ca2+], and membrane potential showed that these Ca2+ spikes were paralleled by depolarisations of the plasma membrane. Using patch clamp measurements, action potential-like depolarisations consisting of a fast spike depolarisation followed by a plateau phase were seen with similar kinetics as the Ca2+ spikes. The action potentials could be blocked by L-type Ca2+ channel blockers and were dependent on extracellular Ca2+. The plateau phase was predominantly determined by a Cl-conductance and was dependent on intracellular Ca2+. The presence of voltage-dependent L-type Ca2+ channels in NRK cells was confirmed by patch clamp measurements in single cells. It is concluded that monolayers of density-arrested NRK fibroblasts exhibit spontaneous Ca2+ action potentials leading to synchronised Ca2+ spiking. This excitability of monolayers of fibroblasts may represent a novel Ca2+ signaling pathway in electrically coupled fibroblasts, cells that were hitherto considered to be inexcitable.

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Generation of Calcium Oscillations in Fibroblasts by Positive Feedback Between Calcium and IP ₃

Cited by 307 publications

References 33 publications

Calcium-dependent Regulation of Protein Kinase D Revealed by a Genetically Encoded Kinase Activity Reporter

Calcium-dependent Regulation of Protein Kinase D Revealed by a Genetically Encoded Kinase Activity Reporter

Regulation of cytosolic calcium by collagen in single human platelets

Synchronized calcium spiking resulting from spontaneous calcium action potentials in monolayers of NRK fibroblasts

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Generation of Calcium Oscillations in Fibroblasts by Positive Feedback Between Calcium and IP 3

Cited by 307 publications

References 33 publications

Calcium-dependent Regulation of Protein Kinase D Revealed by a Genetically Encoded Kinase Activity Reporter

Calcium-dependent Regulation of Protein Kinase D Revealed by a Genetically Encoded Kinase Activity Reporter

Regulation of cytosolic calcium by collagen in single human platelets

Synchronized calcium spiking resulting from spontaneous calcium action potentials in monolayers of NRK fibroblasts

Contact Info

Product

Resources

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Generation of Calcium Oscillations in Fibroblasts by Positive Feedback Between Calcium and IP ₃