Search tips
Search criteria 


Logo of plantsigLink to Publisher's site
Plant Signal Behav. 2009 October; 4(10): 989–991.
PMCID: PMC2801370

Extracellular ATP activates an Arabidopsis plasma membrane Ca2+-permeable conductance


Extracellular ATP has been found to elevate cytosolic free Ca2+ in Arabidopsis thaliana and trigger gene transcription, suggesting that it acts as a plant cell regulator. Recent findings place extracellular ATP upstream of Arabidopsis thaliana NADPH oxidase activity and plasma membrane Ca2+-permeable channels in the root epidermis. Here we show that increasing extracellular ATP concentration evokes a larger but more irregular Ca2+ influx conductance in root epidermal protoplasts. This may help modulate changes in cytosolic free Ca2+ as a second messenger and help explain the dose-dependent effects of extracellular ATP on cell function. The receptors for ATP and the downstream plasma membrane Ca2+ channels remain unknown at the protein or gene level. No equivalents of animal ATP receptors have been identified in higher plant genomes. We propose here that annexins could perceive extracellular ATP and participate in Ca2+ influx.

Key words: ADP, annexin, ATP, calcium, channel

Extracellular purine nucleotides are well established as animal cell regulators, acting in a variety of process from cell death to neurotransmission.1 Signaling is effected by activation of ionotropic or metabotropic plasma membrane receptors (purinoceptors) and can result in elevation of cytosolic free calcium ([Ca2+]cyt) as a second messenger.1 It is now clear that, although apparently lacking homologues of purinoceptors, plant cells also respond to extracellular ATP. The latter is now implicated in regulation of cell viability,2 growth,35 gravitropism6 and stress responses.710 In both roots and leaves, extracellular ATP increases transcription of stress-responsive genes such as MPK3 (mitogen-activated protein kinase 3).9,10 There may be cell- and dose-dependent effects on growth and development.4

Very early work on plant responses to extracellular ATP interpreted effects (such as enhanced closure rate of Venus fly trap) as the uptake of ATP to act as an additional energy source.11,12 Indeed tracer experiments confirmed ATP influx. Establishing a bona fide effect of extracellular ATP is made difficult by the ability of ATP to chelate cations, which could in turn trigger a signaling response. Chelation of extracellular Ca2+ promotes production of reactive oxygen species (ROS) by Arabidopsis thaliana roots13 and low cation availability is known to trigger adaptive responses by roots, often involving ROS.14

Such processes could readily confound the delineation of purine nucleotide effects. By ensuring that extracellular Ca2+ is held constant or is in excess, it has been shown that extracellular ATP can induce a transient increase in [Ca2+]cyt of Arabidopsis roots and leaves.8,15 Recently it was found to elevate [Ca2+]cyt in root epidermal protoplasts from Arabidopsis which may indicate that, in this tissue, the cell wall is not involved in ATP perception.10 The current model is that an initial event downstream of ATP perception is release of Ca2+ from intracellular stores. This can then activate the root epidermal plasma membrane (PM) NADPH oxidase AtRBOHC, resulting in production of reactive oxygen species (ROS). ROS then activate PM Ca2+-permeable channels which can further contribute to [Ca2+]cyt elevation.10

Low levels (20 µM) of extracellular ATP evoke a hyperpolarization-activated, time-dependent and regular inwardly-directed PM Ca2+ conductance in root epidermal protoplasts.10 Here we show that increasing the ATP concentration to 2 mM (reviewed in ref. 10) changes the conductance profile, making it increasingly irregular over the course of the voltage clamping step as membrane voltage becomes more hyperpolarized (Fig. 1). The control mean ± SE current at −200 mV was −177 ± 40 pA and after ATP addition it increased to −610 ± 87 pA (n = 4; Fig. 2). This represents an approximate threefold increase, compared to the two fold increase observed at this voltage in response to 20 µM ATP.10 This result illustrates the point that the [Ca2+]cyt response to changing extracellular [ATP] could be fine-tuned at the level of the PM Ca2+ influx pathway.

Figure 1
Extracellular ATP at high concentration evokes an irregular Ca2+ conductance in root epidermal plasma membrane. Protoplasts were isolated from the mature epidermis.10 Representative current traces obtained with the whole cell patch clamp recording configuration ...
Figure 2
Extracellular ATP evokes a hyperpolarization-activated conductance. Mean ± SE current-voltage relationships for control and +2 mM extracellular ATP (n = 4) obtained using experimental conditions described in Figure 1 and ref. 10. Negative current ...

It is held that the combined but differentially regulated activities of passive and active transporters plus Ca2+-binding proteins effect stimulus-specific spatiotemporal patterns of [Ca2+]cyt as a second messenger, known as the “Ca2+ signature”.16 The PM Ca2+ influx response to extracellular ATP does now appear to be dose-dependent with the possibility of pulses of Ca2+ entry at higher [ATP] were the PM voltage to remain constant. The molecular identity of the underlying channels remains unknown. However, it is feasible that annexins could contribute to some or all of the ATP-evoked Ca2+ conductance or even be the “receptor”. Annexins are soluble proteins capable of Ca2+-dependent membrane association with or insertion into membranes.17 Animal and now plant annexins have been found to facilitate passive Ca2+ transport in vitro.18 Two annexins of the Arabidopsis root epidermis, AtANN1 and AtANN2, are possible candidates as they are predicted to be extracellular18 and have been found to be in the cell wall in proteomic studies on other cell types.19,20 AtANN1 can reside in the plasma membrane21 and, as it can bind ATP,22 could insert into the membrane to form a transport route. AtANN1 has cation transport capacity in vitro23 although its ability to translocate Ca2+ is unknown. AtANN2 is predicted to bind nucleotide triphosphates24 and could also be a transport route or could regulate levels of extracellular ATP. With loss of function mutants now available, these possibilities could be addressed.


This work was supported by the Royal Society and the University of Cambridge Brooks Fund.



1. Khakh BS, North RA. P2X receptors as cell-surface ATP sensors in health and disease. Nature. 2006;442:527–532. [PubMed]
2. Chivasa S, Ndimba BK, Simon WJ, Lindsey K, Slabas AR. Extracellular ATP functions as an endogenous external metabolite regulating plant cell viability. Plant Cell. 2005;17:3019–3034. [PubMed]
3. Kim SY, Sivaguru M, Stacey G. Extracellular ATP in plants: Visualisation, localization and analysis of physiological significance in growth and signaling. Plant Physiol. 2006;142:984–992. [PubMed]
4. Roux SJ, Steinebrunner I. Extracellular ATP: an unexpected role as a signaler in plants. Trends Plant Sci. 2007;12:1380–1385. [PubMed]
5. Wu J, Steinebrunner I, Sun Y, Butterfield T, Torres J, Arnold D, et al. Apyrases (nucleoside triphosphatediphosphohydrolases) play a key role in growth control in Arabidopsis. Plant Physiol. 2007;144:961–975. [PubMed]
6. Tang W, Brady SR, Sun Y, Muday GK, Roux SJ. Extracellular ATP inhibits root gravitropism at concentrations that inhibit polar auxin transport. Plant Physiol. 2003;131:147–154. [PubMed]
7. Thomas C, Rajagopal A, Windsor B, Dudler R, Lloyd A, Roux SJ. A role for ectoapyrase in xenobiotic resistance. Plant Cell. 2000;12:519–533. [PubMed]
8. Jeter CR, Tang W, Henaff E, Butterfield T, Roux SJ. Evidence of a novel cell signalling role for extracellular adenosine triphosphates and diphosphates in Arabidopsis. Plant Cell. 2004;16:2652–2664. [PubMed]
9. Song CJ, Steinebrunner I, Wang X, Stout SC, Roux SJ. Extracellular ATP induces the accumulation of superoxide via NADPH oxidases in Arabidopsis. Plant Physiol. 2006;140:1222–1232. [PubMed]
10. Demidchik V, Shang Z, Shin R, Thompson E, Rubio L, Laohavisit A, et al. Plant extracellular ATP signaling by plasma membrane NADPH oxidase and Ca2+ channels. Plant J. 2009;58:903–913. [PubMed]
11. Jaffe MJ. The role of ATP in mechanically-stimulated rapid closure of the Venus's Fly trap. Plant Physiol. 1973;51:17–18. [PubMed]
12. Lüttge U, Schöch EV, Ball E. Can externally applied ATP supply energy to active ion uptake mechanisms of intact plant cells? Aust J Plant Physiol. 1974;1:211–220.
13. Mortimer JC, Laohavisit A, Miedema H, Davies JM. Voltage, reactive oxygen species and the influx of calcium. Plant Signal Behav. 2008;3:698–699. [PMC free article] [PubMed]
14. Shin R, Schachtman DP. Hydrogen peroxide mediates plant root cell response to nutrient deprivation. Proc Natl Acad Sci USA. 2004;101:8827–8832. [PubMed]
15. Demidchik V, Nichols C, Oliynyk M, Dark A, Glover BJ, Davies JM. Is ATP a signaling agent in plants? Plant Physiol. 2003;133:456–461. [PubMed]
16. McAinsh MR, Pittman JL. Shaping the calcium signature. New Phytol. 2009;181:275–294. [PubMed]
17. Hofmann A. Annexins in the plant kingdom, perspectives and potentials. Annexins. 2004;1:51–61.
18. Laohavisit A, Mortimer JC, Demidchik V, Coxon KM, Stancombe MA, Macpherson N, et al. Zea mays annexins modulate cytosolic free Ca2+ and generate a Ca2+-permeable conductance. Plant Cell. 2009;21:479–493. [PubMed]
19. Bayer EM, Bottrill AR, Walshaw J, et al. Arabidopsis cell wall proteome defined using multidimensional protein identification technology. Proteomics. 2006;6:301–311. [PubMed]
20. Kwon HK, Yokoyama R, Nishitani K. A proteomic approach to apoplastic proteins involved in cell wall regeneration in protoplasts of Arabidopsis suspension-culture cells. Plant Cell Physiol. 2005;46:843–857. [PubMed]
21. Bayer EM, Bottrill AR, Walshaw J, Vigouroux M, Naldrett MJ, Thomas CL, et al. Use of a proteome strategy for tagging proteins present at the plasma membrane. Plant J. 1998;16:633–641. [PubMed]
22. Ito J, Heazlewood JL, Millar AH. Analysis of the soluble ATP-binding proteome of plant mitochondria identifies new proteins and nucleotide triphosphate interactions within the matrix. J Prot Res. 2006;5:3459–3469. [PubMed]
23. Gorecka KM, Thouverey C, Buchet CR, Pikula S. Potential role of annexin AtANN1 from Arabidopsis thaliana in pH-mediated cellular response to environment stimuli. Plant Cell Physiol. 2007;48:792–803. [PubMed]
24. Clark GB, Sessions A, Eastburn DJ, Roux SJ. Differential expression of members of the annexin multigene family in Arabidopsis. Plant Physiol. 2001;126:1072–1084. [PubMed]

Articles from Plant Signaling & Behavior are provided here courtesy of Taylor & Francis