cAMP and Ca signaling in secretory epithelia: Crosstalk and synergism

The Ca²⁺ and cAMP/PKA pathways are the primary signaling systems in secretory epithelia that control virtually all secretory gland functions. Interaction and crosstalk in Ca²⁺ and cAMP signaling occur at multiple levels to control and tune the activity of each other. Physiologically, Ca²⁺ and cAMP signaling operate at 5–10% of maximal strength, but synergize to generate the maximal response. Although synergistic action of the Ca²⁺ and cAMP signaling is the common mode of signaling and has been known for many years, we know very little of the molecular mechanism and mediators of the synergism. In this review, we discuss crosstalk between the Ca²⁺ and cAMP signaling and the function of IRBIT (IP₃ receptors binding protein release with IP₃) as a third messenger that mediates the synergistic action of the Ca²⁺ and cAMP signaling.     

Compartmentalized crosstalk of CFTR and TMEM16A (ANO1) through EPAC1 and ADCY1

Airway epithelial cells express both Ca²⁺ activated TMEM16A/ANO1 and cAMP activated CFTR anion channels. Previous work suggested a significant crosstalk of intracellular Ca²⁺ and cAMP signaling pathways, leading to activation of both chloride channels. We demonstrate that in airway epithelial cells, stimulation of purinergic or muscarinic G-protein coupled receptors (GPCRs) activates TMEM16A and CFTR. Additional expression of G_q/11 and phospholipase C coupled GPCRs strongly enhanced the crosstalk between Ca²⁺- and cAMP-dependent signaling. Knockdown of endogenous GRCRs attenuated crosstalk and functional coupling between TMEM16A and CFTR. The number of receptors did not affect expression or membrane localization of TMEM16A or CFTR, but controlled assembly of the local signalosome. GPCRs translocate Ca²⁺-sensitive adenylate cyclase type 1 (ADCY1) and exchange protein directly activated by cAMP (EPAC1) to particular plasma membrane domains containing GPCRs, CFTR and TMEM16A, thereby producing compartmentalized Ca²⁺ and cAMP signals and significant crosstalk. While biosynthesis and membrane trafficking of CFTR requires a functional Golgi apparatus, maturation and membrane trafficking of TMEM16A may occur independent of the Golgi. Because Ca²⁺ activated TMEM16A currents are only transient, continuous Cl⁻ secretion by airway epithelial cells requires CFTR. The present data also explain why receptor-dependent activation of TMEM16A is more efficient than direct stimulation by Ca²⁺.     

Spine neck geometry determines spino-dendritic cross-talk in the presence of mobile endogenous calcium binding proteins

Dendritic spines are thought to compartmentalize second messengers like Ca²⁺. The notion of isolated spine signaling, however, was challenged by the recent finding that under certain conditions mobile endogenous Ca²⁺-binding proteins may break the spine limit and lead to activation of Ca²⁺-dependent dendritic signaling cascades. Since the size of spines is variable, the spine neck may be an important regulator of this spino-dendritic crosstalk. We tested this hypothesis by using an experimentally defined, kinetic computer model in which spines of Purkinje neurons were coupled to their parent dendrite by necks of variable geometry. We show that Ca²⁺ signaling and calmodulin activation in spines with long necks is essentially isolated from the dendrite, while stubby spines show a strong coupling with their dendrite, mediated particularly by calbindin D28k. We conclude that the spine neck geometry, in close interplay with mobile Ca²⁺-binding proteins, regulates the spino-dendritic crosstalk.     

Chemical signaling under abiotic stress environment in plants

Many chemicals are critical for plant growth and development and play an important role in integrating various stress signals and controlling downstream stress responses by modulating gene expression machinery and regulating various transporters/pumps and biochemical reactions. These chemicals include calcium (Ca²⁺), cyclic nucleotides, polyphosphoinositides, nitric oxide (NO), sugars, abscisic acid (ABA), jasmonates (JA), salicylic acid (SA) and polyamines. Ca²⁺ is one of the very important ubiquitous second messengers in signal transduction pathways and usually its concentration increases in response to the stimuli including stress signals. Many Ca²⁺ sensors detect the Ca²⁺ signals and direct them to downstream signaling pathways by binding and activating diverse targets. cAMP or cGMP protects the cell with ion toxicity. Phosphoinositides are known to be involved both in transmission of signal across the plasma membrane and in intracellular signaling. NO activates various defense genes and acts as a developmental regulator in plants. Sugars affect the expression of many genes involved in photosynthesis, glycolysis, nitrogen metabolism, sucrose and starch metabolism, defense mechanisms and cell cycle regulation. ABA, JA, SA and polyamines are also involved in many stress responses. Cross-talk between these chemical signaling pathways is very common in plant responses to abiotic and bitotic factors. In this article we have described the role of these chemicals in initiating signaling under stress conditions mainly the abiotic stress.Key words: ABA, abiotic stress, Ca²⁺ binding proteins, calcium signaling, cyclic nucleotides, nitric oxide, phosphoinositides signaling, signal transduction, sugar signaling     

Atypical properties of a conventional calcium channel β subunit from the platyhelminth Schistosoma mansoni

Background

Serotonin induces fluid secretion from Calliphora salivary glands by the parallel activation of the InsP₃/Ca²⁺ and cAMP signaling pathways. We investigated whether cAMP affects 5-HT-induced Ca²⁺ signaling and InsP₃-induced Ca²⁺ release from the endoplasmic reticulum (ER).

Results

Increasing intracellular cAMP level by bath application of forskolin, IBMX or cAMP in the continuous presence of threshold 5-HT concentrations converted oscillatory [Ca²⁺]_i changes into a sustained increase. Intraluminal Ca²⁺ measurements in the ER of β-escin-permeabilized glands with mag-fura-2 revealed that cAMP augmented InsP₃-induced Ca²⁺ release in a concentration-dependent manner. This indicated that cAMP sensitized the InsP₃ receptor Ca²⁺ channel for InsP₃. By using cAMP analogs that activated either protein kinase A (PKA) or Epac and the application of PKA-inhibitors, we found that cAMP-induced augmentation of InsP₃-induced Ca²⁺ release was mediated by PKA not by Epac. Recordings of the transepithelial potential of the glands suggested that cAMP sensitized the InsP₃/Ca²⁺ signaling pathway for 5-HT, because IBMX potentiated Ca²⁺-dependent Cl^- transport activated by a threshold 5-HT concentration.

Conclusion

This report shows, for the first time for an insect system, that cAMP can potentiate InsP₃-induced Ca²⁺ release from the ER in a PKA-dependent manner, and that this crosstalk between cAMP and InsP₃/Ca²⁺ signaling pathways enhances transepithelial electrolyte transport.     

Crosstalk between purinergic receptors and canonical signaling pathways in the mouse salivary gland

Isolated clusters of mouse parotid acinar cells in combination with live cell imaging were used to explore the crosstalk in molecular signaling between purinergic, cholinergic and adrenergic pathways that integrate to control fluid and protein secretion. This crosstalk was manifested by (1) β-adrenergic receptor activation and amplification of P2X4R evoked Ca²⁺ signals, (2) β-adrenergic-induced amplification of P2X7R-evoked Ca²⁺ signals and (3) muscarinic receptor induced activation of P2X7Rs via exocytotic activity. The findings from our study reveal that purinoceptor-mediated Ca²⁺ signaling is modulated by crosstalk with canonical signaling pathways in parotid acinar cells. Integration of these signals are likely important for dynamic control of saliva secretion to match physiological demand in the parotid gland.     

Lipids at membrane contact sites: cell signaling and ion transport

Communication between organelles is essential to coordinate cellular functions and the cell's response to physiological and pathological stimuli. Organellar communication occurs at membrane contact sites (MCSs), where the endoplasmic reticulum (ER) membrane is tethered to cellular organelle membranes by specific tether proteins and where lipid transfer proteins and cell signaling proteins are located. MCSs have many cellular functions and are the sites of lipid and ion transfer between organelles and generation of second messengers. This review discusses several aspects of MCSs in the context of lipid transfer, formation of lipid domains, generation of Ca²⁺ and cAMP second messengers, and regulation of ion transporters by lipids.     

Selective inhibition of histamine-evoked Ca2+ signals by compartmentalized cAMP in human bronchial airway smooth muscle cells

Intracellular Ca²⁺ and cAMP typically cause opposing effects on airway smooth muscle contraction. Receptors that stimulate these pathways are therapeutic targets in asthma and chronic obstructive pulmonary disease. However, the interactions between different G protein-coupled receptors (GPCRs) that evoke cAMP and Ca²⁺ signals in human bronchial airway smooth muscle cells (hBASMCs) are poorly understood. We measured Ca²⁺ signals in cultures of fluo-4-loaded hBASMCs alongside measurements of intracellular cAMP using mass spectrometry or [³H]-adenine labeling. Interactions between the signaling pathways were examined using selective ligands of GPCRs, and inhibitors of Ca²⁺ and cAMP signaling pathways. Histamine stimulated Ca²⁺ release through inositol 1,4,5-trisphosphate (IP₃) receptors in hBASMCs. β₂-adrenoceptors, through cAMP and protein kinase A (PKA), substantially inhibited histamine-evoked Ca²⁺ signals. Responses to other Ca²⁺-mobilizing stimuli were unaffected by cAMP (carbachol and bradykinin) or minimally affected (lysophosphatidic acid). Prostaglandin E₂ (PGE₂), through EP₂ and EP₄ receptors, stimulated formation of cAMP and inhibited histamine-evoked Ca²⁺ signals. There was no consistent relationship between the inhibition of Ca²⁺ signals and the amounts of intracellular cAMP produced by different stimuli. We conclude that β-adrenoceptors, EP₂ and EP₄ receptors, through cAMP and PKA, selectively inhibit Ca²⁺ signals evoked by histamine in hBASMCs, suggesting that PKA inhibits an early step in H₁ receptor signaling. Local delivery of cAMP within hyperactive signaling junctions mediates the inhibition.     

Mechanisms of Regulation of Olfactory Transduction and Adaptation in the Olfactory Cilium

Olfactory adaptation is a fundamental process for the functioning of the olfactory system, but the underlying mechanisms regulating its occurrence in intact olfactory sensory neurons (OSNs) are not fully understood. In this work, we have combined stochastic computational modeling and a systematic pharmacological study of different signaling pathways to investigate their impact during short-term adaptation (STA). We used odorant stimulation and electroolfactogram (EOG) recordings of the olfactory epithelium treated with pharmacological blockers to study the molecular mechanisms regulating the occurrence of adaptation in OSNs. EOG responses to paired-pulses of odorants showed that inhibition of phosphodiesterases (PDEs) and phosphatases enhanced the levels of STA in the olfactory epithelium, and this effect was mimicked by blocking vesicle exocytosis and reduced by blocking cyclic adenosine monophosphate (cAMP)-dependent protein kinase (PKA) and vesicle endocytosis. These results suggest that G-coupled receptors (GPCRs) cycling is involved with the occurrence of STA. To gain insights on the dynamical aspects of this process, we developed a stochastic computational model. The model consists of the olfactory transduction currents mediated by the cyclic nucleotide gated (CNG) channels and calcium ion (Ca²⁺)-activated chloride (CAC) channels, and the dynamics of their respective ligands, cAMP and Ca²⁺, and it simulates the EOG results obtained under different experimental conditions through changes in the amplitude and duration of cAMP and Ca²⁺ response, two second messengers implicated with STA occurrence. The model reproduced the experimental data for each pharmacological treatment and provided a mechanistic explanation for the action of GPCR cycling in the levels of second messengers modulating the levels of STA. All together, these experimental and theoretical results indicate the existence of a mechanism of regulation of STA by signaling pathways that control GPCR cycling and tune the levels of second messengers in OSNs, and not only by CNG channel desensitization as previously thought.     

Melatonin as a central molecule connecting neural development and calcium signaling

Melatonin (MEL) is a neuroendocrine hormone secreted by the pineal gland in association with the suprachiasmatic nucleus and peripheral tissues. MEL has been observed to play a critical role in the reproductive process and in the fetomaternal interface. Extrapineal synthesis has been reported in mammalian models during pregnancy, especially by the placenta tissue. MEL can regulate intracellular processes (e.g., G-proteins) and the activity of second messengers (e.g., cAMP, IP_3, Ca²⁺). During neurodevelopment, these activities regulated by melatonin have an important role as an intracellular signaling for gene expression regulation. To review the role of MEL in neurodevelopment, we built interactome networks of different proteins that act in these processes using systems biology tools. The analyses of interactome networks revealed that MEL could modulate neurodevelopment through the regulation of Ca²⁺ intracellular levels and influencing BMP/SMAD signaling, thus affecting neural gene responses and neuronal differentiation.     

Segregation and Crosstalk of D1 Receptor-Mediated Activation of ERK in Striatal Medium Spiny Neurons upon Acute Administration of Psychostimulants

The convergence of corticostriatal glutamate and dopamine from the midbrain in the striatal medium spiny neurons (MSN) triggers synaptic plasticity that underlies reinforcement learning and pathological conditions such as psychostimulant addiction. The increase in striatal dopamine produced by the acute administration of psychostimulants has been found to activate not only effectors of the AC5/cAMP/PKA signaling cascade such as GluR1, but also effectors of the NMDAR/Ca²⁺/RAS cascade such as ERK. The dopamine-triggered effects on both these cascades are mediated by D1R coupled to Golf but while the phosphorylation of GluR1 is affected by reductions in the available amount of Golf but not of D1R, the activation of ERK follows the opposite pattern. This segregation is puzzling considering that D1R-induced Golf activation monotonically increases with DA and that there is crosstalk from the AC5/cAMP/PKA cascade to the NMDAR/Ca²⁺/RAS cascade via a STEP (a tyrosine phosphatase). In this work, we developed a signaling model which accounts for this segregation based on the assumption that a common pool of D1R and Golf is distributed in two D1R/Golf signaling compartments. This model integrates a relatively large amount of experimental data for neurons in vivo and in vitro. We used it to explore the crosstalk topologies under which the sensitivities of the AC5/cAMP/PKA signaling cascade to reductions in D1R or Golf are transferred or not to the activation of ERK. We found that the sequestration of STEP by its substrate ERK together with the insensitivity of STEP activity on targets upstream of ERK (i.e. Fyn and NR2B) to PKA phosphorylation are able to explain the experimentally observed segregation. This model provides a quantitative framework for simulation based experiments to study signaling required for long term potentiation in MSNs.     

Molecular Properties of Sodium/Dicarboxylate Cotransporters

Cells possess multiple Ca²⁺ stores and multiple messengers for mobilizing them. In addition to inositol trisphosphate and cyclic ADP-ribose, nicotinic acid adenine dinucleotide phosphate (NAADP), a metabolite of NADP, is shown to be a potent Ca²⁺ signaling molecule in both invertebrate and mammalian cells. This article summarizes the recent results of this newly discovered Ca²⁺ signaling mechanism and explores the implications of the apparent proliferation of Ca²⁺ messengers. Received: 11 August 1999/Revised: 21 September 1999     

Role of calcineurin biosignaling in cell secretion and the possible regulatory mechanisms

Cyclic adenosine monophosphate (cAMP) and calcium ions (Ca²⁺) are two chemical molecules that play a central role in the stimulus-dependent secretion processes within cells. Ca²⁺ acts as the basal signaling molecule responsible to initiate cell secretion. cAMP primarily acts as an intracellular second messenger in a myriad of cellular processes by activating cAMP-dependent protein kinases through association with such kinases in order to mediate post-translational phosphorylation of those protein targets. Put succinctly, both Ca²⁺ and cAMP act by associating or activating other proteins to ensure successful secretion. Calcineurin is one such protein regulated by Ca²⁺; its action depends on the intracellular levels of Ca²⁺. Being a phosphatase, calcineurin dephosphorylate and other proteins, as is the case with most other phosphatases, such as protein phosphatase 2A (PP2A), PP2C, and protein phosphatase-1 (PP1), will likely be activated by phosphorylation. Via this process, calcineurin is able to affect different intracellular signaling with clinical importance, some of which has been the basis for development of different calcineurin inhibitors. In this review, the cAMP-dependent calcineurin bio-signaling, protein-protein interactions and their physiological implications as well as regulatory signaling within the context of cellular secretion are explored.     

Crosstalk between calcium and reactive oxygen species signaling in cancer

The interplay between Ca²⁺ and reactive oxygen species (ROS) signaling pathways is well established, with reciprocal regulation occurring at a number of subcellular locations. Many Ca²⁺ channels at the cell surface and intracellular organelles, including the endoplasmic reticulum and mitochondria are regulated by redox modifications. In turn, Ca²⁺ signaling can influence the cellular generation of ROS, from sources such as NADPH oxidases and mitochondria. This relationship has been explored in great depth during the process of apoptosis, where surges of Ca²⁺ and ROS are important mediators of cell death. More recently, coordinated and localized Ca²⁺ and ROS transients appear to play a major role in a vast variety of pro-survival signaling pathways that may be crucial for both physiological and pathophysiological functions. While much work is required to firmly establish this Ca²⁺-ROS relationship in cancer, existing evidence from other disease models suggests this crosstalk is likely of significant importance in tumorigenesis. In this review, we describe the regulation of Ca²⁺ channels and transporters by oxidants and discuss the potential consequences of the ROS-Ca²⁺ interplay in tumor cells.     

Regulation of bile secretion by calcium signaling in health and disease

Calcium (Ca²⁺) signaling controls secretion in many types of cells and tissues. In the liver, Ca²⁺ regulates secretion in both hepatocytes, which are responsible for primary formation of bile, and cholangiocytes, which line the biliary tree and further condition the bile before it is secreted. Cholestatic liver diseases, which are characterized by impaired bile secretion, may result from impaired Ca²⁺ signaling mechanisms in either hepatocytes or cholangiocytes. This review will discuss the Ca²⁺ signaling machinery and mechanisms responsible for regulation of secretion in both hepatocytes and cholangiocytes, and the pathophysiological changes in Ca²⁺ signaling that can occur in each of these cell types to result in cholestasis.     

Termination and activation of store‐operated cyclic AMP production

Diverse pathophysiological processes (e.g. obesity, lifespan determination, addiction and male fertility) have been linked to the expression of specific isoforms of the adenylyl cyclases (AC1‐AC10), the enzymes that generate cyclic AMP (cAMP). Our laboratory recently discovered a new mode of cAMP production, prominent in certain cell types, that is stimulated by any manoeuvre causing reduction of free [Ca²⁺] within the lumen of the endoplasmic reticulum (ER) calcium store. Activation of this ‘store‐operated’ pathway requires the ER Ca²⁺ sensor, STIM1, but the identity of the enzymes responsible for cAMP production and how this process is regulated is unknown. Here, we used sensitive FRET‐based sensors for cAMP in single cells combined with silencing and overexpression approaches to show that store‐operated cAMP production occurred preferentially via the isoform AC3 in NCM460 colonic epithelial cells. Ca²⁺ entry via the plasma membrane Ca²⁺ channel, Orai1, suppressed cAMP production, independent of store refilling. These findings are an important first step towards defining the functional significance and to identify the protein composition of this novel Ca²⁺/cAMP crosstalk system.     

Anoctamin 1 in secretory epithelia

Fluid and electrolyte releasing from secretory epithelia are elaborately regulated by orchestrated activity of ion channels. The activity of chloride channel at the apical membrane decides on the direction and the rate of secretory fluid and electrolyte. Chloride-dependent secretion is conventionally associated with intracellular increases in two second messengers, cAMP and Ca²⁺, responding to luminal purinergic and basolateral adrenergic or cholinergic stimulation. While it is broadly regarded that cAMP-dependent Cl⁻ secretion is regulated by cystic fibrosis transmembrane conductance regulator (CFTR), Ca²⁺-activated Cl⁻ channel (CaCC) had been veiled for quite some time. Now, Anoctamin 1 (ANO1 or TMEM16A) confers Ca²⁺-activated Cl⁻ currents. Ano 1 and its paralogs have been actively investigated for multiple functions underlying Ca²⁺-activated Cl⁻ efflux and fluid secretion in a variety of secretory epithelial cells. In this review, we will discuss recent advances in the secretory function and signaling of ANO1 in the secretory epithelia, such as airways, intestines, and salivary glands.     

Cell adhesion and intracellular calcium signaling in neurons

Cell adhesion molecules (CAMs) play indispensable roles in the developing and mature brain by regulating neuronal migration and differentiation, neurite outgrowth, axonal fasciculation, synapse formation and synaptic plasticity. CAM-mediated changes in neuronal behavior depend on a number of intracellular signaling cascades including changes in various second messengers, among which CAM-dependent changes in intracellular Ca²⁺ levels play a prominent role. Ca²⁺ is an essential secondary intracellular signaling molecule that regulates fundamental cellular functions in various cell types, including neurons. We present a systematic review of the studies reporting changes in intracellular Ca²⁺ levels in response to activation of the immunoglobulin superfamily CAMs, cadherins and integrins in neurons. We also analyze current experimental evidence on the Ca²⁺ sources and channels involved in intracellular Ca²⁺ increases mediated by CAMs of these families, and systematically review the role of the voltage-dependent Ca²⁺ channels (VDCCs) in neurite outgrowth induced by activation of these CAMs. Molecular mechanisms linking CAMs to VDCCs and intracellular Ca²⁺ stores in neurons are discussed.