Adenovirus-mediated Expression of an Olfactory Cyclic Nucleotide-gated Channel Regulates the Endogenous Ca2+-inhibitable Adenylyl Cyclase in C6-2B Glioma Cells

Fagan, Kent A.; Rich, Thomas C.; Tolman, Shawna; Schaack, Jerome; Karpen, Jeffrey W.; Cooper, Dermot M.F.

doi:10.1074/jbc.274.18.12445

Cited by 38 publications

(29 citation statements)

References 35 publications

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“…We took advantage of the method developed by Rich and coworkers 27,31 in HEK293 cells to follow in real time cGMP changes beneath the sarcolemmal membrane using the WT-CNGA2 channel as a readout. We confirmed by immunocytochemistry (see online-only Data Supplement) that this channel is not normally expressed in native rat cardiomyocytes 35 but becomes functionally expressed 24 hours after infection of the myocytes with the Ad-CNGA2 construct developed by Fagan et al 32 A similar method was used recently to follow cAMP changes in the same preparation, 16 with the use of mutants of the CNGA2 channel with a higher sensitivity toward cAMP versus cGMP.The WT-CNGA2 channel responds to cGMP changes with a threshold at 0.1 to 0.5 mol/L concentration, a K 1/2 of 1.4 mol/L, and a maximal amplitude obtained at 5 to 10 mol/L cGMP. 27 With the use of these parameters and the amplitude of the CNG current measured at the end of each experiment after application of a saturating concentration (100 mol/L) of the cGMP analogue Sp-8, it was possible to give a rough estimate of the subsarcolemmal cGMP concentration reached in each of the experimental conditions tested here.…”

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confidence: 51%

Section: Discussionmentioning

confidence: 61%

“…27,31,32 In the present study we have applied this methodology to differentiated adult cardiomyocytes in primary culture to measure in real time the changes in subsarcolemmal cGMP. Our study reveals major differences in the spatiotemporal distribution of intracellular cGMP on activation of its 2 main routes of synthesis.…”

Section: Discussionmentioning

confidence: 99%

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Cyclic Guanosine Monophosphate Compartmentation in Rat Cardiac Myocytes

et al. 2006

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Background-Cyclic guanosine monophosphate (cGMP) is the common second messenger for the cardiovascular effects of nitric oxide (NO) and natriuretic peptides, such as atrial or brain natriuretic peptide, which activate the soluble and particulate forms of guanylyl cyclase, respectively. However, natriuretic peptides and NO donors exert different effects on cardiac and vascular smooth muscle function. We therefore tested whether these differences are due to an intracellular compartmentation of cGMP and evaluated the role of phosphodiesterase (PDE) subtypes in this process. Methods and Results-Subsarcolemmal cGMP signals were monitored in adult rat cardiomyocytes by expression of the rat olfactory cyclic nucleotide-gated (CNG) channel ␣-subunit and recording of the associated cGMP-gated current (I CNG ). Atrial natriuretic peptide (10 nmol/L) or brain natriuretic peptide (10 nmol/L) induced a clear activation of I CNG , whereas NO donors (S-nitroso-N-acetyl-penicillamine, diethylamine NONOate, 3-morpholinosydnonimine, and spermine NO, all at 100 mol/L) had little effect. The I CNG current was strongly potentiated by nonselective PDE inhibition with isobutyl methylxanthine (100 mol/L) and by the PDE2 inhibitors erythro-9-(2-hydroxy-3-nonyl)adenine (10 mol/L) and Bay 60-7550 (50 nmol/L). Surprisingly, sildenafil, a PDE5 inhibitor, produced a dose-dependent increase of I CNG activated by NO donors but had no effect (at 100 nmol/L) on the current elicited by atrial natriuretic peptide. Conclusions-These results indicate that in rat cardiomyocytes (1) the particulate cGMP pool is readily accessible at the plasma membrane, whereas the soluble pool is not; and (2) PDE5 controls the soluble but not the particulate pool, whereas the latter is under the exclusive control of PDE2. Differential spatiotemporal distributions of cGMP may therefore contribute to the specific effects of natriuretic peptides and NO donors on cardiac function. Key Words: cyclic GMP Ⅲ natriuretic peptides Ⅲ nitric oxide Ⅲ phosphodiesterases Ⅲ sildenafil C yclic guanosine monophosphate (cGMP) is a ubiquitous intracellular second messenger in the cardiovascular system. In the heart, acute elevation of cGMP concentration usually exerts negative metabolic as well as inotropic effects, 1,2 whereas chronic elevation prevents and reverses cardiac hypertrophy. 3-5 cGMP synthesis is controlled by 2 types of guanylyl cyclases (GC) that differ in their cellular location and activation by specific ligands: a particulate GC (pGC) present at the plasma membrane, which is activated by natriuretic peptides such as atrial (ANP), brain (BNP), and C-type natriuretic peptide 6 -8 ; and a soluble guanylyl cyclase (sGC) present in the cytosol and activated by nitric oxide (NO). 8,9 Clinical Perspective p 2228Although NO and natriuretic peptides use cGMP as a common second messenger, there are many instances in which activation of pGC and sGC leads to different functional effects. 10 -14 One explanation for these divergent effects is that cGMP rises in specific subcellular...

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confidence: 51%

Section: Discussionmentioning

confidence: 61%

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Cyclic Guanosine Monophosphate Compartmentation in Rat Cardiac Myocytes

et al. 2006

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“…The methodology was developed in a series of elegant studies in model cell lines for the measurement of intracellular cAMP. 115,143,143,144 This method uses wild-type or genetically modified ␣ subunits of rat olfactory CNG channel (CNGA2), which form a cationic channel directly opened by cyclic nucleotides. Adult cardiac myocytes infected with an adenovirus encoding the native or modified channels elicit a nonselective cation current when, respectively, cGMP 145 or cAMP concentration 146,147 rises beneath the sarcolemmal membrane.…”

Section: Methods To Study Cyclic Nucleotide Compartmentation In Intacmentioning

confidence: 99%

Compartmentation of Cyclic Nucleotide Signaling in the Heart

Fischmeister

Castro

Abi-Gerges

et al. 2006

Circulation Research

328

251

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Abstract-A current challenge in cellular signaling is to decipher the complex intracellular spatiotemporal organization that any given cell type has developed to discriminate among different external stimuli acting via a common signaling pathway. This obviously applies to cAMP and cGMP signaling in the heart, where these cyclic nucleotides determine the regulation of cardiac function by many hormones and neuromediators. Key Words: cAMP Ⅲ cGMP Ⅲ heart Ⅲ G protein-coupled receptor Ⅲ phosphodiesterase T he cyclic nucleotides cyclic adenosine 3Ј,5Ј-monophosphate (cAMP) and cyclic guanosine 3Ј,5Ј-monophosphate (cGMP) were identified more than 4 decades ago. 1,2 Since then, many studies have appeared on how these 2 second messengers are synthesized or degraded, what makes their level go up or down, what they do to target effectors by either covalent (phosphorylation) or noncovalent (direct binding to proteins, such as ion channels or guanine-nucleotide-exchange factors) mechanisms, and how they affect a countless number of cellular functions. [3][4][5][6] In certain tissues and organs, the cyclic nucleotide pathways have been so fully explored over the years that one can wonder what else is there to be found. This is the case for cAMP in the heart, where it plays a key role in the sympathetic nerve/␤-adrenergic receptor (␤-AR)/adenylyl cyclase (AC)/protein kinase A (PKA) axis that serves to stimulate cardiac rhythm (chronotropy) as well as contractile force (inotropy) and relaxation (lusitropy). 7 Yet, there are a number of questions that have always made us wonder but have only lately begun to receive the attention they deserve: how so many different receptors coupled to cAMP or cGMP signaling pathway manage to achieve specific cellular responses? What is the purpose of the different adenylyl and guanylyl cyclases present in the same cell? Why do Original

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“…Ca 2C influx through CNG ion channels is stimulated in response to an increase in cAMP. When coupled with the Ca 2C -sensitive indicator Fura-2 and PDE inhibitors, mutants of cAMP-selective CNG channels function as real-time biosensors of cAMP accumulation at the plasma membrane [71][72][73] (Figure 4c). Likewise, the cAMPbinding domain of the Epac-GEF has been used to generate fluorescent cAMP biosensors that detect compartmentalized accumulation of the second messenger [74,75] (Figure 4d).…”

Section: Phosphorylation In Akap Complexesmentioning

confidence: 99%

The where's and when's of kinase anchoring

Smith

Langeberg

Scott

2006

Trends in Biochemical Sciences

130

132

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Kinase anchoring has gained acceptance as a means to synchronize spatial and temporal aspects of cell signaling. A-kinase anchoring proteins (AKAPs) are a diverse group of functionally related proteins that target protein kinase A and other enzymes to coordinate a range of signaling events. Recent advances in this field have shown that incorporating phosphodiesterases into AKAP signaling complexes exerts local control of cAMP metabolism, that phosphorylation of some AKAPs potentiates downstream signaling events, that anchoring of distinct enzyme combinations functions as a mechanism to expand the repertoire of cellular events controlled by a single AKAP, and that fluorescent biosensors can be used to visualize dynamic aspects of localized cAMP signaling. IntroductionA half-century of work has proclaimed protein phosphorylation as a principal means of reversibly controlling biochemical events. It began when Fischer and Krebs [1] demonstrated that conversion of inactive muscle phosphorylase b into active phosphorylase a requires ATP and is catalyzed by a phosphorylase b kinase. They then showed that phosphorylase b kinase itself is controlled by a serine kinase that is responsive to the second messenger cAMP. Together with their colleagues, they subsequently discovered protein kinase A (PKA) and the concept of a 'kinase cascade' [2].From these humble beginnings the field of protein kinase research was born, paving the way for the discovery of phosphotyrosine by Hunter and co-workers [3], the identification of Src tyrosine kinase by Brugge and Ericsson et al. [4], the seminal work of Pawson and associates on phosphotyrosine recognition motifs [5], and the resolution of a crystal structure of the PKA catalytic subunit by Taylor and colleagues [6]. Consequently, we now recognize that the w518 members of the human kinome represent a superfamily of enzymes that participate in all aspects of cellular regulation [7].Protein kinase research is now focusing on how these enzymes are organized in relation to their effectors and substrates within the three dimensions of the cell. Interestingly, the study of PKA is shedding new light on the complexity of these protein-protein interactions.

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Adenovirus-mediated Expression of an Olfactory Cyclic Nucleotide-gated Channel Regulates the Endogenous Ca2+-inhibitable Adenylyl Cyclase in C6-2B Glioma Cells

Cited by 38 publications

References 35 publications

Cyclic Guanosine Monophosphate Compartmentation in Rat Cardiac Myocytes

Cyclic Guanosine Monophosphate Compartmentation in Rat Cardiac Myocytes

Compartmentation of Cyclic Nucleotide Signaling in the Heart

The where's and when's of kinase anchoring

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