CHARACTERIZATION OF THE HUMAN ADENYLYL CYCLASE GENE FAMILY: cDNA, GENE STRUCTURE, AND TISSUE DISTRIBUTION OF THE NINE ISOFORMS

Ludwig, Marie-Gabrielle; Seuwen, Klaus

doi:10.1081/rrs-120014589

Cited by 93 publications

(68 citation statements)

References 41 publications

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“…15 Western blots using the selective antibody for the AC-VI isoform ( Figure 1B) did not show a band at 130 kDa 16 where it would be expected. A very faint band was barely apparent in this lane just below the 130-kDa site.…”

Section: Ac-v and Ac-vi Isoform Expression In Jg Cells Using Western mentioning

confidence: 99%

“…Proteins were electrophoretically transferred to a polyvinylidene difluoride membrane (polyvinylidene fluoride, Invitrogen) overnight at 4°C. The membranes were blocked with 5% milk in tris-buffered saline tween for 1 hour at room temperature and then incubated with either an antibody selective for the AC-V isoform (Fabgennix International Inc) 15 or an antibody selective for the AC-VI isoform (Abcam) 16 in 1:1000 dilutions in 5% albumin overnight at 4°C. Primary antibodies were labeled with a horseradish peroxidase-conjugated goat anti-rabbit IgG secondary antibody at a dilution of 1:2000.…”

Section: Ac-v and Ac-vi Isoform Expression In Jg Cells Using Western mentioning

confidence: 99%

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Adenylyl Cyclase Isoform V Mediates Renin Release From Juxtaglomerular Cells

et al. 2007

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Abstract-We have shown previously that decreasing intracellular calcium in the juxtaglomerular cells increases both cAMP formation and renin release. We hypothesized that this is because of an interaction between intracellular calcium and the calcium-inhibitable isoform of adenylyl cyclase, type-V. We used primary cultures of juxtaglomerular cells isolated from C-57/B6 mice at 70% to 80% confluence. Western blots were performed on isolated juxtaglomerular cells using antibodies against either of the 2 calcium inhibitable isoforms of adenylyl cyclase, types-V and -VI. Only the antibody against adenylyl cyclase-V gave us a strong band at 120 kDa as expected. There are 9 isoforms of adenylyl cyclase. They all share a similar structure that includes an intracellular N terminus followed by 2 membrane-spanning domains alternating with 2 catalytic cytoplasmic loops. 7 There are 2 adenylyl cyclase isoforms, type-V (AC-V) and type-VI (AC-VI), which are both inhibited by micromolar concentrations of [Ca ϩ2 ] i . 8 Both isoforms are found in lipid rafts in either the cell or intracellular membrane regions. 9 In addition, AC-V and AC-VI share a high amino acid sequence identity, particularly in the regions that form the enzymes catalytic unit. 10 However, the N termini of the 2 isoforms are quite divergent. 7 It has been reported that, in nonexcitable cells, AC-VI colocalizes with the calcium entry channels in the cell membrane. 11 Thus, whereas the 2 enzymes share many properties, they may also have different interactions within the cell and have different compartmentalization (such as in the surface or on intracellular membranes).Previously, 6 using an antibody that does not discriminate between the 2 isoforms, we found that either 1 or both of these calcium-inhibitable adenylyl cyclases existed within the JG cell, and their presence could explain the paradoxical effect of how decreasing [Ca ϩ2 ] i could result in a cAMPmediated stimulation of renin secretion. Although we observed a provocative colocalization of our antibody with renin on what appeared to be granule-like foci within the cytoplasm, 6 we were unable to distinguish between the 2 isoforms to answer the critical question of which calciuminhibitable isoform of adenylyl cyclase is in the JG cell and

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Section: Ac-v and Ac-vi Isoform Expression In Jg Cells Using Western mentioning

confidence: 99%

Section: Ac-v and Ac-vi Isoform Expression In Jg Cells Using Western mentioning

confidence: 99%

Adenylyl Cyclase Isoform V Mediates Renin Release From Juxtaglomerular Cells

et al. 2007

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“…There are nine types of adenylyl cyclases, which are all transmembrane proteins, synthesizing cAMP at the plasma membrane (73,74). This results in a gradient in which higher concentrations of cAMP are found at the membrane and lower concentrations in the cytosol (73,(75)(76)(77)(78).…”

Section: Role Of Alveolar Epithelial B 2 -Adrenergic Receptorsmentioning

confidence: 99%

Alveolar Epithelial β₂-Adrenergic Receptors

Mutlu

Factor²

2008

Am J Respir Cell Mol Biol

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b 2 -adrenergic receptors are present throughout the lung, including the alveolar airspace, where they play an important role for regulation of the active Na 1 transport needed for clearance of excess fluid out of alveolar airspace. b 2 -adrenergic receptor signaling is required for up-regulation of alveolar epithelial active ion transport in the setting of excess alveolar edema. The positive, protective effects of b 2 -adrenergic receptor signaling on alveolar active Na 1 transport in normal and injured lungs provide substantial support for the use of b-adrenergic agonists to accelerate alveolar fluid clearance in patients with cardiogenic and noncardiogenic pulmonary edema. In this review, we summarize the role of b 2 -adrenergic receptors in the alveolar epithelium with emphasis on their role in the regulation of alveolar active Na 1 transport in normal and injured lungs.Keywords: pulmonary edema; acute respiratory distress syndrome; acute lung injury; alveoli; albuterol b 2 -adrenergic receptors (b 2 AR) are present throughout the lung. In the alveolar airspace they are important for regulation of the active Na 1 transport needed for clearance of excess fluid out of alveolar airspace (1). Both experimental and limited clinical data suggest that b-adrenergic agonists working via the b 2 AR accelerate clearance of excess fluid from the alveolar airspace, creating the possibility of their use for treatment of pulmonary edema and acute lung injury (ALI).In this review, we summarize the role of b 2 -adrenergic receptors in the alveolar epithelium with emphasis on their role in regulation of alveolar active Na 1 transport in normal and injured lungs. We also overview data regarding b 2 -agonist therapy for pulmonary edema and lung injury. In the lung, b2-adrenergic receptor (b 2 AR) expression increases with each airway generation, with the greatest total amounts in the distal airways and alveoli (3). Greater than 90% of all b-adrenergic receptors in human lung are located in the alveoli (4). Although both b 1 and b 2 subtypes coexist and are distributed uniformly in the alveolar walls, the b 2 -subtype predominates (70%) (4). Isolated rat alveolar type II cells possess b 2 AR and data from autoradiographic and immunohistochemical studies support their presence in the alveolar type 1 cells (4, 5). -Adrenergic Receptor Structure and FunctionThe b 2 -adrenergic receptor is a 1,200-base pair, single-copy, intronless gene located on the long arm of human chromosome 5 that encodes a 413-amino acid protein with a molecular mass of approximately 46.5 kD (1). The b 2 -receptor is a prototypical G protein-coupled receptor (GPCR) with seven-transmembrane domains, an extracellular amino terminus, an intracellular carboxyl terminus, three interconnecting extracellular loops, and three intracellular loops.b 2 -adrenergic receptors exist in the plasma membrane in an equilibrium between at least two structural conformations; inactive and active forms that are defined based on their ability to associate with the stimulatory guano...

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“…The C1 and C2 regions are further subdivided in to "a" and "b" regions. The C1a and C2a regions interact to form the catalytic domain of adenylyl cyclase (Tang and Hurley, 1998;Ludwig and Seuwen, 2002). The transmembrane spanning domains have been suggested to be important in the regulation of protein assembly and membrane trafficking of the enzyme (Gu et al, 2001).…”

mentioning

confidence: 99%

Isoform-Specific Regulation of Adenylyl Cyclase Function by Disruption of Membrane Trafficking

Ding

Gros²,

Chorazyczewski³

et al. 2004

Mol Pharmacol

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Oligomerization plays an important role in endoplasmic reticulum processing and membrane insertion (and ultimately in regulation of function) of a number of transmembrane spanning proteins. Furthermore, it is known that adenylyl cyclases (ACs), critical regulators of cellular functions, associate into higher order (dimeric) forms. However, the importance of these higher order aggregates in regulating adenylyl cyclase activity or trafficking to the cell membrane is unclear. Therefore, we examined the potential role of oligomerization in the membrane trafficking of adenylyl cyclase. For this purpose, the ability of full-length adenylyl cyclase and various truncation mutants to self-assemble and to be targeted to the cell membrane was assessed. A truncation mutant comprised of the initial six transmembrane spanning domains and half of the C1 catalytic domain coimmunoprecipitated with full-length AC VI. Using both biotinylation assays and assessment of enzyme distribution using sucrose density gradients, we demonstrate that expression of this mutant in human embryonic kidney 293 cells impaired the ability of AC VI to traffic to the plasma membrane. Furthermore, mutant expression resulted in a significant reduction in adenylyl cyclase activity. The decrease in AC VI membrane expression was not caused by alterations in enzyme transcription. The effect of the mutant was specific for the AC V and VI isoforms and expression of the transmembrane M1 domain but not the C1a domain was required for the mutant to affect adenylyl cyclase activity. In aggregate, these data suggest that alterations in the ability of adenylyl cyclases to form higher order forms regulate both enzyme trafficking and enzyme activity.Adenylyl cyclases comprise a large family of enzymes that catalyze the synthesis of cyclic AMP from ATP. Adenylyl cyclases constitute the "effector" component of transmembrane signaling pathways that are both positively and negatively regulated by the activity of heterotrimeric G protein and G protein-coupled receptors (GPCRs). The activity of the enzyme adenylyl cyclase is itself regulated by both extracellular stimuli (via GPCR receptor activation) and intracellular stimuli (Cooper et al., 1995;Hanoune and Defer, 2001;Patel et al., 2001). Studies of the regulation of adenylyl cyclases initially focused "upstream" of the enzyme (i.e., on characterization of regulation of enzyme function via G protein and GPCR function/expression) or on the regulation of substrate (Mg 2ϩ /ATP) availability. More recently, the importance of regulation of enzyme activity by covalent modifications (i.e., enzyme phosphorylation) has increasingly been appreciated

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CHARACTERIZATION OF THE HUMAN ADENYLYL CYCLASE GENE FAMILY: cDNA, GENE STRUCTURE, AND TISSUE DISTRIBUTION OF THE NINE ISOFORMS

Cited by 93 publications

References 41 publications

Adenylyl Cyclase Isoform V Mediates Renin Release From Juxtaglomerular Cells

Adenylyl Cyclase Isoform V Mediates Renin Release From Juxtaglomerular Cells

Alveolar Epithelial β₂-Adrenergic Receptors

Isoform-Specific Regulation of Adenylyl Cyclase Function by Disruption of Membrane Trafficking

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CHARACTERIZATION OF THE HUMAN ADENYLYL CYCLASE GENE FAMILY: cDNA, GENE STRUCTURE, AND TISSUE DISTRIBUTION OF THE NINE ISOFORMS

Cited by 93 publications

References 41 publications

Adenylyl Cyclase Isoform V Mediates Renin Release From Juxtaglomerular Cells

Adenylyl Cyclase Isoform V Mediates Renin Release From Juxtaglomerular Cells

Alveolar Epithelial β2-Adrenergic Receptors

Isoform-Specific Regulation of Adenylyl Cyclase Function by Disruption of Membrane Trafficking

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Product

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Alveolar Epithelial β₂-Adrenergic Receptors