Calcium and signal transduction in plants

Environmental and hormonal signals control diverse physiological processes in plants. The mechanisms by which plant cells perceive and transduce these signals are poorly understood. Understanding biochemical and molecular events involved in signal transduction pathways has become one of the most active areas of plant research. Research during the last 15 years has established that Ca2+ acts as a messenger in transducing external signals. The evidence in support of Ca2+ as a messenger is unequivocal and fulfills all the requirements of a messenger. The role of Ca2+ becomes even more important because it is the only messenger known so far in plants. Since our last review on the Ca2+ messenger system in 1987, there has been tremendous progress in elucidating various aspects of Ca(2+) -signaling pathways in plants. These include demonstration of signal-induced changes in cytosolic Ca2+, calmodulin and calmodulin-like proteins, identification of different Ca2+ channels, characterization of Ca(2+) -dependent protein kinases (CDPKs) both at the biochemical and molecular levels, evidence for the presence of calmodulin-dependent protein kinases, and increased evidence in support of the role of inositol phospholipids in the Ca(2+) -signaling system. Despite the progress in Ca2+ research in plants, it is still in its infancy and much more needs to be done to understand the precise mechanisms by which Ca2+ regulates a wide variety of physiological processes. The purpose of this review is to summarize some of these recent developments in Ca2+ research as it relates to signal transduction in plants.     

Messenger-specific role for nicotinic acid adenine dinucleotide phosphate in neuronal differentiation

Cells possess several Ca2+-mobilizing messengers, which couple stimulation at the cell surface by a multitude of extracellular cues to the regulation of intracellular Ca2+-sensitive targets. Recent studies suggest that agonists differentially select from this molecular palette to generate their characteristic Ca2+ signals but it is still unclear whether different messengers mediate different functions or whether they act in a redundant fashion. In this study, we compared the effects of nicotinic acid adenine dinucleotide phosphate (NAADP), a novel Ca2+-mobilizing messenger, with that of the prototypical messenger inositol trisphosphate on cytosolic Ca2+ levels and differentiation status of PC12 cells. We demonstrate that liposomal delivery of NAADP mediated release of Ca2+ from acidic Ca2+ stores and that this stimulus was sufficient to drive differentiation of the cells to a neuronal-like phenotype. In sharp contrast, cell fate was unaffected by more transient Ca2+ signals generated by inositol trisphosphate-evoked release of endoplasmic reticulum Ca2+ stores. Our data establish for the first time (i) the presence of novel NAADP-sensitive Ca2+ stores in PC12 cells, (ii) a role for NAADP in differentiation, and (iii) that Ca2+-dependent function can be messenger-specific. Thus, differential recruitment of intracellular Ca2+-mobilizing messengers and their target Ca2+ stores may represent a robust means of maintaining stimulus fidelity in the control of Ca2+-dependent cell function.     

Inositol tetrakisphosphate as a second messenger: confusions, contradictions, and a potential resolution.

The second messenger function of inositol 1,4,5-trisphosphate (InsP3) is now well-defined--it mobilizes Ca2+ from intracellular stores so that cystolic Ca2+ increases. However, the function of inositol 1,3,4,5-tetrakisphosphate (InsP4) has proved much more difficult to fathom, as it has been reported to exert a wide variety of effects in a collection of experimental systems. In this review, a proposed molecular mechanism for InsP4's actions is discussed; it is suggested that InsP4 is the second messenger that controls Ca2+ entry into cells, and that it does so by binding to a receptor which itself interacts, directly or indirectly, with the receptor for InsP3. It is proposed that this is InsP4's true physiological function, but the mechanism by which it exerts this function has led to confusing data concerning its action, and also to some misconceptions about how inositol phosphates control Ca2+ entry.     

NAADP receptors

Nicotinic acid adenine dinucleotide phosphate (NAADP) is a recently described Ca2+ mobilizing messenger. First described in the sea urchin egg, it has been shown to mobilize Ca2+ from intracellular stores. It is a remarkably potent molecule, and recent reports show that its cellular levels change in response to a variety of agonists confirming its role as a Ca2+ mobilizing messenger. In many cases NAADP interacts with other Ca2+ mobilizing messengers such as inositol 1,4,5 trisphosphate (IP3 and cyclic adenosine diphosphate ribose (cADPR) in shaping cytosolic Ca2+ signals. What is not clear is the molecular nature of the NAADP-sensitive Ca2+ release mechanism and its sub-cellular localization. In this review we focus on the recent progress made in sea urchin eggs, which indicates that NAADP activates a novel Ca2+ release channel distinct from the relatively well-characterized IP3 and ryanodine receptors. Furthermore, in the sea urchin egg, the NAADP-sensitive store appears to be separate from the endoplasmic reticulum (ER) and is most likely an acidic store. These findings have also been reinforced by similar findings by some in mammalian cells. Finally, we discuss ongoing strategies to characterise NAADP-binding proteins which will greatly enhance our understanding of NAADP-mediated Ca2+ signalling, and lead to the development of more selective tools to probe the role of this messenger.     

Micro-injection of inositol 1,3,4,5-tetrakisphosphate activates sea urchin eggs by a mechanism dependent on external Ca2+.

Micro-injection of submicromolar concentrations of inositol 1,3,4,5-tetrakisphosphate caused a raising of the fertilization envelope in eggs of the sea urchin Lytechinus variegatus. This effect was dependent both on the presence of extracellular Ca2+ and on co-injection with a Ca2+-mobilizing compound, inositol 2,4,5-trisphosphate. Inositol 1,3,4,5-tetrakisphosphate was the most potent compound tested in this assay; removal of the 3- or 5-phosphates or randomization of the phosphates in the inositol ring decreased its potency. These results show that inositol 1,3,4,5-tetrakisphosphate is an intracellular second messenger, and suggest that its function is to control cellular Ca2+ homoeostasis at the plasma membrane.     

Investigations of calcium homeostasis mechanisms in nerve cells and their alterations during brain pathology

The paper summarises new data about the molecular mechanisms of calcium homeostasis maintenance in nerve cells and generation of intracellular calcium transients--the most general secondary messenger triggering or modulating all steps of neuronal life cycle and its main functions. It describes the low- and high-voltage activated plasmalemmal ion channels injecting Ca2+ into the cell, cytosolic buffering systems which rapidly bind the main part of injected ions, properties of intracellular stores accumulating Ca2+ ions due to the activity of CERCA-pumps and releasing them back into the cytosol via the CICR mechanism, possible participation of mitochondria in this process, extrusion of Ca2+ from the cell by PMCA-pumps. By introducing new techniques, quantitative characteristics are obtained of these mechanisms and of their participation in determining the amplitude and kinetics of calcium signals in different neurons, as well as their changes during ageing and some forms of brain pathology.     

Fertilisation calcium signals in the ascidian egg

The egg of ascidians (urochordate), as virtually all animal and plant species, displays Ca2+ signals upon fertilisation. These Ca2+ signals are repetitive Ca2+ waves that initiate in the cortex of the egg and spread through the whole egg interior. Two series of Ca2+ waves triggered from two distinct Ca2+ wave pacemakers entrain the two meiotic divisions preceding entry into the first interphase. The second messenger inositol (1,4,5) trisphosphate (IP3) is the main mediator of these global Ca2+ waves. Other Ca2+ signalling pathways (RyR and NAADPR) are functional in the egg but they mediate localised cortical Ca2+ signals whose physiological significance remains unclear. The meiosis I Ca2+ wave pacemaker is mobile and relies on intracellular Ca2+ release from the endoplasmic reticulum (ER) induced by a large production of IP3 at the sperm aster site. The meiosis II Ca2+ wave pacemaker is stably localised in a vegetal protrusion called the contraction pole. It is probable that a local production of IP3 in the contraction pole determines the site of this second pacemaker while functional interactions between ER and mitochondria regulate its activity. Finally, a third ectopic pacemaker can be induced by a global increase in IP3, making the ascidian egg a unique system where three different Ca2+ wave pacemakers coexist in the same cell.     

Triggering of Ca2+ signals by NAADP-gated two-pore channels: a role for membrane contact sites?

NAADP (nicotinic acid-adenine dinucleotide phosphate) is a potent Ca2+-mobilizing messenger implicated in many Ca2+-dependent cellular processes. It is highly unusual in that it appears to trigger Ca2+ release from acidic organelles such as lysosomes. These signals are often amplified by archetypal Ca2+ channels located in the endoplasmic reticulum. Recent studies have converged on the TPCs (two-pore channels) which localize to the endolysosomal system as the likely primary targets through which NAADP mediates its effects. 'Chatter' between TPCs and endoplasmic reticulum Ca2+ channels is disrupted when TPCs are directed away from the endolysosomal system. This suggests that intracellular Ca2+ release channels may be closely apposed, possibly at specific membrane contact sites between acidic organelles and the endoplasmic reticulum.     

Painful nerve injury increases plasma membrane Ca2+ -ATPase activity in axotomized sensory neurons

ABSTRACT: BACKGROUND: The plasma membrane Ca2+-ATPase (PMCA) is the principal means by which sensory neurons expel Ca 2+and thereby regulate the concentration of cytoplasmic Ca 2+and the processes controlled by this critical second messenger. We have previously found that painful nerve injury decreases resting cytoplasmic Ca2+ levels and activity-induced cytoplasmic Ca2+accumulation in axotomized sensory neurons. Here we examine the contribution of PMCA after nerve injury in a rat model of neuropathic pain. RESULTS: PMCA function was isolated in dissociated sensory neurons by blocking intracellular Ca2+sequestration with thapsigargin, and cytoplasmic Ca2+concentration was recorded with Fura-2 fluorometry. Compared to control neurons, the rate at which depolarization-induced Ca 2+transients resolved was increased in axotomized neurons after spinal nerve ligation, indicating accelerated PMCA function. Electrophysiological recordings showed that blockade of PMCA by vanadate prolonged the action potential afterhyperpolarization, and also decreased the rate at which neurons could fire repetitively. CONCLUSION: We found that PMCA function is elevated in axotomized sensory neurons, which contributes to neuronal hyperexcitability. Accelerated PMCA function in the primary sensory neuron may contribute to the generation of neuropathic pain, and thus its modulation could provide a new pathway for peripheral treatment of post-traumatic neuropathic pain.     

Calcium signaling in liver

In hepatocytes, hormones linked to the formation of the second messenger inositol 1,4,5-trisphosphate (InsP3) evoke transient increases or spikes in cytosolic free calcium ([Ca2+]i), that increase in frequency with the agonist concentration. These oscillatory Ca2+ signals are thought to transmit the information encoded in the extracellular stimulus to down-stream Ca2+-sensitive metabolic processes. We have utilized both confocal and wide field fluorescence microscopy techniques to study the InsP3-dependent signaling pathway at the cellular and subcellular levels in the intact perfused liver. Typically InsP3-dependent [Ca2+]i spikes manifest as Ca2+ waves that propagate throughout the entire cytoplasm and nucleus, and in the intact liver these [Ca2+]i increases are conveyed through gap junctions to encompass entire lobular units. The translobular movement of Ca2+ provides a means to coordinate the function of metabolic zones of the lobule and thus, liver function. In this article, we describe the characteristics of agonist-evoked [Ca2+]i signals in the liver and discuss possible mechanisms to explain the propagation of intercellular Ca2+ waves in the intact organ.     

钙信号在卵母细胞激活中的作用

钙信号是胞内主要的第二信使之一,发挥广泛的作用如细胞分裂、细胞凋亡等,对细胞的生命活动起着非常重要的作用。在精子和卵母细胞中,钙信号对精子获能、顶体反应、卵母细胞成熟、受精及卵裂等一系列复杂的过程有非常重要的影响。现就Ca2 在卵母细胞中的释放机制、信号转导途径、调控功能作一综述。     

Extracellular calcium acts as a "third messenger" to regulate enzyme and alkaline secretion

It is generally assumed that the functional consequences of stimulation with Ca2+ -mobilizing agonists are derived exclusively from the second messenger action of intracellular Ca2+, acting on targets inside the cells. However, during Ca2+ signaling events, Ca2+ moves in and out of the cell, causing changes not only in intracellular Ca2+, but also in local extracellular Ca2+. The fact that numerous cell types possess an extracellular Ca2+ "sensor" raises the question of whether these dynamic changes in external [Ca2+] may serve some sort of messenger function. We found that in intact gastric mucosa, the changes in extracellular [Ca2+] secondary to carbachol-induced increases in intracellular [Ca2+] were sufficient and necessary to elicit alkaline secretion and pepsinogen secretion, independent of intracellular [Ca2+] changes. These findings suggest that extracellular Ca2+ can act as a "third messenger" via Ca2+ sensor(s) to regulate specific subsets of tissue function previously assumed to be under the direct control of intracellular Ca2+.     

Ca2+ homeostasis in apoptotic resistance of prostate cancer cells

Ca2+ is a universal messenger regulating many physiological functions including such an important one, as the ability of the cell to undergo orderly self-destruction upon completion of its mission, called apoptosis. If this function is compromised unwanted cells may eventually take over the tissue turning it into a cancer. Ca2+ dependency of apoptosis, when its all aspects are learned and understood and key molecular players identified, may provide a good opportunity for controlling tumor growth. In the present mini-review we describe the major molecular determinants of Ca2+ homeostasis in prostate cancer cells and establish their role in the transformation to apoptosis-resistant cell phenotypes typical of advanced androgen-independent prostate cancer. We show that the hallmark of such transformation is the inhibition of apoptosis pathway associated with endoplasmic reticulum Ca2+ store depletion.     

CAPRI and RASAL impose different modes of information processing on Ras due to contrasting temporal filtering of Ca2+

The versatility of Ca2+ as a second messenger lies in the complex manner in which Ca2+ signals are generated. How information contained within the Ca2+ code is interpreted underlies cell function. Recently, we identified CAPRI and RASAL as related Ca2+-triggered Ras GTPase-activating proteins. RASAL tracks agonist-stimulated Ca2+ oscillations by repetitively associating with the plasma membrane, yet CAPRI displays a long-lasting Ca2+-triggered translocation that is refractory to cytosolic Ca2+ oscillations. CAPRI behavior is Ca2+- and C2 domain-dependent but sustained recruitment is predominantly Ca2+ independent, necessitating integration of Ca2+ by the C2 domains with agonist-evoked plasma membrane interaction sites for the pleckstrin homology domain. Using an assay to monitor Ras activity in real time, we correlate the spatial and temporal translocation of CAPRI with the deactivation of H-Ras. CAPRI seems to low-pass filter the Ca2+ signal, converting different intensities of stimulation into different durations of Ras activity in contrast to the preservation of Ca2+ frequency information by RASAL, suggesting sophisticated modes of Ca2+-regulated Ras deactivation.     

PPADS is a reversible competitive antagonist of the NAADP receptor

NAADP has been shown to act as a second messenger in a wide range of systems from plants to mammalian cells. Although it had always been considered as a canonical second messenger, recent work has shown that it is also active when applied extracellularly. It has also been suggested that NAADP might have a direct action on P2 receptors, based on the action of a pharmacological agent, PPADS, on Ca2+ signals in response to extracellular NAADP. We have therefore investigated whether PPADS can act directly on the intracellular NAADP-induced Ca2+-release system in the well characterised sea urchin egg homogenate system. Indeed, PPADS, and its structural analogue PPNDS were able to compete with [32P]NAADP for the binding site and binding curves revealed that both compounds display affinities in the low micromolar range. The binding of PPADS was reversible in contrast to that of NAADP. In fluorimetric Ca2+-release experiments, PPADS was able to competitively antagonise NAADP-induced Ca2+-release with an IC50 of 20 microM, while it did not affect the other Ca2+-release channels. This is the first report of a reversible, competitive antagonist of the sea urchin NAADP receptor. Furthermore, PPADS might reveal itself as an invaluable tool to investigate NAADP signalling and is a lead compound for the synthesis of potent and specific antagonists.     

Receptor-activated cytoplasmic Ca2+ oscillations in pancreatic acinar cells: generation and spreading of Ca2+ signals.

Receptor-activated cytoplasmic Ca2+ oscillations have been investigated using both single cell microfluorometry and voltage-clamp recording of Ca(2+)-dependent Cl- current in single internally perfused acinar cells. In these cells there is direct experimental evidence showing that the ACh-evoked [Ca2+]i fluctuations are due to an inositol trisphosphate-induced small steady Ca2+ release which in turn evokes repetitive Ca2+ spikes via a caffeine-sensitive Ca(2+)-induced Ca2+ release process. There is indirect evidence suggesting that receptor-activation in addition to generating the Ca2+ releasing messenger, inositol trisphosphate, also produces another regulator involved in the control of Ca2+ signal spreading. Intracellular inositol trisphosphate or Ca2+ infusion produce short duration repetitive spikes confined to the cytoplasmic area close to the plasma membrane, but these signals can be made to progress throughout the cell by addition of caffeine or by receptor activation.     

Ca2+ Dynamics during Membrane Excitation of Green Alga Chara: Model Simulations and Experimental Data

Kinetic investigations of stimulus response coupling in the green alga Chara have revealed that an intermediate second messenger is formed in the process of membrane excitation. This second messenger links electrical stimulation to the mobilization of Ca2+ from internal stores. In the present work, the experimentally based kinetic model, which describes the stimulus-dependent production of the second messenger and Ca2+ mobilization, is combined with a model for inositol 1,4,5-trisphosphate (IP3)-and Ca2+-sensitive gating of a Ca2+-release channel in endomembranes of animal cells. The combination of models allows a good simulation of experimental data, including the all-or-none-type dependence of the Ca2+ response on stimulus duration and complex phase locking phenomena for the dependence of the Ca2+ response on stimulation frequency. The model offers a molecular explanation for the refractory phenomenon in Chara, assigning it to the life time of an inactive state of the Ca2+-release channel. The model furthermore explains the steep dependence of excitation on strength/duration of electrical stimulation as a consequence of an interplay of the dynamical variables in the model.     

Second messenger function of inositol 1,4,5-trisphosphate. Early changes in inositol phosphates, cytosolic Ca2+, and insulin release in carbamylcholine-stimulated RINm5F cells

The second messenger function of inositol 1,4,5-trisphosphate (Ins-1,4,5-P3) was investigated in carbamylcholine-stimulated RINm5F cells by analysis of the early changes in inositol phosphates, cytosolic free Ca2+ concentration ([Ca2+]i), and insulin secretion. After a lag of 2 s, [Ca2+]i rose to a peak at 13 +/- 2 s, a response which was due mainly to mobilization from intracellular stores since it persisted even in the absence of extracellular Ca2+. The Ca2+ response had already declined toward prestimulatory levels by the time insulin secretion reached its maximal rate (2-3 min). Although the rises in inositol trisphosphate preceded those of both inositol bisphosphate and monophosphate, all three attained maximal concentrations after 1 min and remained elevated for at least 10 min. The accumulation of inositol trisphosphate was truly Ca2+-independent since it persisted under conditions in which the rise in [Ca2+]i was abolished by prior depletion of intracellular Ca2+ pools. Further analysis by high performance liquid chromatography revealed the presence of the two isomers, Ins-1,4,5-P3 and Ins-1,3,4-P3 in stimulated cells. The latter was virtually absent under nonstimulatory conditions but started to accumulate after a 5-s lag and reached maximal levels after 30 s of stimulation. Ins-1,4,5-P3 doubled within 1 s of carbamylcholine addition, reached a peak after 5 s, and, although declining thereafter, remained slightly elevated for at least 3 min. Hence, both the onset and peak of the rise of Ins-1,4,5-P3 preceded that of [Ca2+]i, which in turn preceded the peak in insulin release. These results strongly suggest that Ins-1,4,5-P3 acts as the second messenger by which carbamylcholine mobilizes intracellular Ca2+ during the initiation of insulin release.