CMLS, Cell. Mol. Life Sci. 55 (1999) 214–232 1420-682X/99/020214-19 $ 1.50+0.20/0 © Birkha¨user Verlag, Basel, 1999 Calcium signaling in plants J. J. Rudd and V. E. Franklin-Tong* Wolfson Laboratory for Plant Molecular Biology, School of Biological Sciences, University of Birmingham, Edgbaston, Birmingham, B15 2TT (UK), Fax +44 121 414 5925, e-mail: [email protected] 2+ Abstract. Changes in the cytosolic concentration of cal- Characteristic changes in [Ca ]i have been seen to 2+ cium ions ([Ca ]i) play a key second messenger role in precede the responses of plant cells and whole plants to signal transduction. These changes are visualized by physiological stimuli. This has had a major impact on making use of either Ca2+-sensitive fluorescent dyes or our understanding of cell signaling in plants. The next the Ca2+-sensitive photoprotein, aequorin. Here we challenge will be to establish how the Ca2+ signals are describe the advances made over the last 10 years or so, encrypted and decoded in order to provide specificity, which have conclusively demonstrated a second messen- and we discuss the current understanding of how this 2+ ger role for [Ca ]i in a few model plant systems. may be achieved. Key words. Cytosolic calcium; signaling; Ca2+ imaging; angiosperms. 2+ 2+ Signal transduction in plant cells has rapidly become a cytosolic free Ca ([Ca ]i) is generally regulated in major topic of research that has emerged from the mammalian and plant cells. We will consider the major disciplines of genetics, molecular biology, physiology advances achieved in this field over the last 10 years or and biochemistry and now largely combines these disci- so, which have been made possible by the new genera- 2+ plines in order to advance our knowledge of how plants tion of dyes which accurately report [Ca ]i, together sense, and respond to, the diversity of extracellular with the advent of use of aequorin in transformed stimuli they are exposed to. These studies have often plants. These new technologies, coupled with the drawn comparisons with how a mammalian cell pro- availability of a handful of model systems which have cesses the information it receives in the form of extracel- clearly identifiable responses to defined physiologically lular signals, and how this information is then decoded relevant stimuli, have had a major impact on our cur- to produce an appropriate response. The concept of rent understanding of cell signaling in plants. Although signal perception followed by intracellular signal trans- lagging behind the mammalian field with respect to the duction, which precedes the cellular response, has detailed knowledge of calcium signaling, it has clearly proven a consistent formula adapted by many eukary- been established that plants use similar methods of otic cells, including those of plants. signaling to animal cells. Nevertheless, despite the ad- The exact mechanisms whereby this is achieved from vances made, there is much more to be learned about start to finish is obviously beyond the scope of any calcium signaling in plants. single review article. We must, therefore, focus in on specific mechanisms of particular interest. This article focuses on what is now regarded as an extremely impor- Calcium dynamics in animal and plant cells tant ‘second messenger’ in plant cells, the Ca2+ ion. 2+ 2+ The focus of the discussion will be on the characteristics The cytosolic free Ca ([Ca ]i) in the cell is under of changes in the cytoplasmic concentration of Ca2+ strict biochemical and physiological control. Cells gen- 2+ observed in intact, living cells that have been exposed to erally operate to keep [Ca ]i low ( 100–200 nM) in a physiological stimulus. We will briefly summarize how the ‘resting’ or quiescent state (see fig. 1). Increases in 2+ [Ca ]i can be used as a ‘second messenger’ following 2+ * Corresponding author. cell stimulation, and in this way [Ca ]i can bring about CMLS, Cell. Mol. Life Sci. Vol. 55, 1999 Multi-Author Review Article 215 Figure 1. The localization and mobilization of Ca2+ during signal transduction in animal and plant cells. This cartoon of a generalized cell uses a pseudocolor scale (as indicated in the scale bar), in order to indicate the calcium concentration within the cytoplasm and organelles of this cell. It contains organelles that are known to store Ca2+ in the mM to mM range. These organelles are illustrated in 2+ 2+ 2+ red to indicate their high [Ca ] (see scale bar). The intracellular concentration of Ca ([Ca ]i) of a quiescent, or unstimulated, cell 2+ is generally accepted to be 100–200 nM (shown as blue; see scale bar). This low [Ca ]i is maintained, at least in part, by the action 2+ 2+ 2+ 2+ of Ca -ATPases which act to pump Ca into intracellular stores. The extracellular Ca concentration ([Ca ]e) is significantly 2+ higher than the [Ca ]i of the cytosol of a quiescent cell. Interaction between signal molecules and receptors (shown at the cell surface here) may result in Ca2+ influx. This is achieved by ‘opening’ of Ca2+ channels that are present in the peripheral plasma membrane, 2+ 2+ 2+ and this leads to local increases in [Ca ]i, as illustrated by the color coding. Ca -ATPases act to return [Ca ]i to basal levels. Signal molecules can also interact with receptors to result in the mobilization of Ca2+ from intracellular stores. The organelles identified as the predominant pool for mobilizable Ca2+ in animal and plant cells are thought to be different. Animal cells are thought to mainly release Ca2+ from the ER and/or the SR. Plant cells, however, appear to use the vacuole as the major mobilizable Ca2+ store, although there is some emerging evidence that they also use Ca2+ stored in the ER. Ca2+ in intracellular stores can be mobilized by the generation of IP3 and cADPR, as indicated. These second messengers interact with specific receptors (IP3R and RyR) located on intracellular Ca2+ stores. These receptors form ion channels through which Ca2+ is released into the cytosol, resulting in spatially (and 2+ temporally) defined increases in [Ca ]i which are indicated in the diagram in pseudocolor. The dotted arrows linking the plasma 2+ membrane receptor to the IP3R indicates that the pathway(s) are not well characterized in plants. These increases in [Ca ]i mediate the elicitation of specific cellular responses to the extracellular signals. The pseudocolors used in this figure may be broadly related to 2+ the images in figure 2, and in this way one can begin to envisage how some of the alterations in [Ca ]i may be brought about. 2+ a cellular response to the stimulus. We attempt to As indicated in figure 1, the [Ca ]i in both these locales explain some of the dynamics and controls involved in is generally accepted to be in the micromolar to mil- regulating Ca2+ in a ‘generalized’ cell, shown in figure limolar range, which is much higher than in the cytosol 1, which encompasses features found in both animal (100–200 nM). This is due, at least in part, to the 2+ 2+ and plant cells. The actual increases in [Ca ]i vary activity of Ca -ATPases (as shown in fig. 1) which with cell types and the stimulus applied, but it is gener- actively pump the Ca2+ into these stores, thereby main- 2+ 2+ ally accepted that the [Ca ]i can increase up to 1–2 taining low [Ca ]i. Figure 1 illustrates that following mM in stimulated cells. The nature of the downstream stimulation by receptor binding, this stored Ca2+ is effectors are quite diverse and will not be described in permitted to move down its concentration gradient and 2+ any detail here, but interested readers should refer to [1] into the cell cytoplasm, increasing [Ca ]i. This can for a good general discussion. involve extracellular or intracellular Ca2+, or both. 2+ 2+ The biochemical means by which [Ca ]i increases Ca that originates from outside the cell often enters varies with the source of the mobilizable Ca2+ store. It via selective ion channels located in the plasma mem- may arise from outside the cell or from an internal pool. brane (as indicated in fig. 1). Intracellular Ca2+ can be 216 J. J. Rudd and V. E. Franklin-Tong Calcium signaling in plant cells released in a number of ways. The first mechanism is cauliflower inflorescences that were not highly vacuo- 2+ the classical example of receptor-mediated increase in lated, it has been demonstrated that IP3-induced Ca 2+ [Ca ]i, resulting from the operation of the phospho- release may take place from nonvacuolar membranes inositide-signaling pathway. This involves phospho- [14]. This suggests that plant cells also have the ability 2+ inositidase C (PIC)-mediated generation of to mobilize Ca stored in the ER as well as the inositol(1,4,5)-trisphosphate (IP3) from the membrane vacuole, for use in cell signaling. phospholipid phosphatidylinositol(4,5)bisphosphate Figure 1 illustrates how these mechanisms may operate 2+ (PIP2). IP3 interacts with specific IP3 receptors (IP3R) in a cell to produce spatially localized high [Ca ]i as a that form Ca2+ channels in the membranes of intracel- result of the activity of both intracellular Ca2+ ion lular Ca2+ stores (see fig. 1). Once bound to its recep- channels and those that are distributed on the periph- tor, IP3 induces channel opening, allowing the efflux of eral plasma membrane. It is thought that messages 2+ 2+ 2+ Ca into the cytosol, through IP3-induced Ca -re- carried by [Ca ]i are decoded to give an appropriate lease (IICR) [2, 3].