Plant K+ Transport: Not Just an UphillStruggle

Dispatch

Colin Brownlee

Most plant cells can accumulate K+ to concentra-tions much higher than those in the external medium.Recent studies are providing new insights into therelative roles of channels and transporters in K+

accumulation. These studies are also pointing tospecific roles for K+ channels and transporters inpolarized development.

Potassium is an essential element in all cells. It isrequired for osmotic regulation and the electrical prop-erties of membranes, and provides an ionic environ-ment that is compatible with a very wide range ofenzymatic activities and metabolic processes. For manyhigher plants, K+ is present in the external medium atconcentrations that are several orders of magnitudelower (µM) than those in the cell (>100 mM). The exter-nal K+ concentration can also vary within a wide range,and it is not surprising that higher plants possess avariety of K+ accumulation mechanisms [1–3].

The conventional ‘pump and leak’ view of membraneion transport considers active uptake to occur via theaction of pumps or transporters. These can couplechemical energy from the hydrolysis of ATP (ATPases)or the energy stored in other ionic gradients (sym-porters or antiporters) to the movement of a particularion against its electrochemical potential gradient. Ionfluxes down their respective electrochemical potentialgradients are mediated by channels. K+ uptake by plantroots shows both low and high affinity uptake kinetics[2], indicating the operation of different types of K+

uptake transporter.Recent studies, however, have blurred the distinc-

tion between the roles of channels or transporters,indicating that K+ uptake though channels may beinvolved in both high and low affinity uptake. It is alsobecoming clear from work with polarized cells such as root hairs and pollen tubes that K+ channels ortransporters may play cell-specific roles related topolarized growth. External K+ is absolutely required forpolarized growth [4,5] and it has long been recognizedthat K+ accumulation is required for the generation ofturgor that is necessary for polarized growth. It nowseems likely that K+ uptake through particular chan-nels or transporters may have functions in addition toproviding the osmotic driving force for polarized growth.

Higher plants possess a subfamily of K+-selectivechannels that belong to the Shaker family that allowK+ influx [1,2]. A recent report by Mouline et al. [5] hasdescribed an Arabidopsis T-DNA insertion mutantdefective in the Shaker-like K+ channel gene Shakerpollen inward K+ channel (SPIK). This gene showed a

pollen-specific expression pattern and seemed likelyto encode the main K+ channel of pollen cells. Pollenfrom mutant plants could germinate but showedsignificantly reduced pollen tube growth and reducedability to compete with wild-type pollen in fertilizationassays in vivo. Heterologous expression of SPIK inanimal COS cells and electrophysiological studies ofwild-type and mutant pollen confirmed that the SPIKgene product forms K+ channels which activate uponmembrane hyperpolarization to values more negativethan about –150 mV.

It is perhaps not surprising that a K+ channel isrequired for pollen tube growth. The results reportedby Mouline et al. [5], however, go further in showingthat spik mutants have defective pollen tube growth atall external K+ concentrations tested between 5 µMand 1 mM. These concentrations cover the range ofboth high and low affinity K+ transporters, implying thatSPIK channels may be involved in high affinity K+

uptake at sufficiently negative membrane potentials.The SPIK channel also showed strong sensitivity toexternal pH suggesting a close coupling between K+

and H+ fluxes. Measurement of the current carried bySPIK channels suggested that they could carry themain K+ influx associated with growth. This contrastswith the indication from other studies ([6,7] for example)that the pollen membrane potential is insufficientlynegative to allow channel-mediated K+ uptake.

There is, however, good evidence from other studiesshowing that, in other cell types at least, K+ channelsmay mediate high-affinity uptake. The growth charac-teristics of Arabidopsis mutants defective for anotherinwardly rectifying K+ channel, AKT1, indicated that theywere disrupted in high-affinity K+ uptake [8]. Furtherdissection of AKT1 function, comparing K+ fluxes inakt1 mutant and wild-type plants, provided furthersupport for the involvement of AKT1 in high-affinity K+

uptake [9]. The generation of single and double knock-out mutants of AKT1 and AKT2 showed that AKT1, butnot AKT2, mediates growth-sustaining K+ uptake byroots under rate-limiting K+ availability [10].

The cell-specificity of SPIK expression and its require-ment for pollen tube elongation over a wide range of K+ concentrations strongly suggests a role that may be specific to polarized pollen tube growth. Aspecific role for K+ transport in polarized growth hasalso been indicated by studies of Arabidopsis T-DNAinsertion mutants that are defective in root hair elon-gation. Root hairs are formed from specialized tri-choblast cells in the root epidermis. Hairs initiate as abulge of the trichoblast cell and subsequently elon-gate by tip growth. Mutants for the gene tiny root hair(Trh) are able to initiate root hairs, but are unable toelongate the cell into a fully formed root hair [5].Sequence data indicate that TRH belongs to theAtKT/AtKup/HAK family of K+ transporters. TRH isexpressed throughout the plant, but the effects of its

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Marine Biological Association of the UK, The Laboratory,Citadel Hill, Plymouth, PL1 2PB, UK. E-mail: cbr@mba.ac.uk

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deletion were only apparent in root hair growth underthe conditions studied, suggesting that its role may beredundant in other cell types. TRH was able tocomplement a yeast strain deficient in high-affinity K+

transport and trh plants were shown to be deficient inhigh-affinity K+ uptake.

The mechanism of action of high-affinity K+ trans-porters is not well understood. Direct K+/H+ symporthas been suggested, but this mechanism has not beenconfirmed in Arabidopsis [11] and different affinitieshave been proposed for different K+ transporters.Thus, AtKT1 (AtKUP) has been reported to exhibithigh-affinity or dual-affinity uptake [12,13], while AtKT2has been suggested to mediate low-affinity K+ uptake[14]. As trh plants show only a root hair phenotype, itseems likely that TRH has an essential role related tolocalized K+ transport. TRH may be involved in gener-ating a sufficiently high root hair K+ concentration forturgor generation.

A specific role for TRH in polarized growth is sug-gested by experiments showing that the trh mutantphenotype is retained even at high (up to 50 mM)external K+, under which conditions high-affinity uptakewould not be required for K+ accumulation. So a simplenon-vectorial role for TRH in K+ accumulation appearsto be insufficient to explain the trh phenotype. TRHmay mediate asymmetric K+ fluxes which may to be essential for polarized root hair growth. Support for this comes from the observation that trh plantsoccasionally initiate multiple sites of abortive root hairgrowth from a single epidermal trichome cell [5].

These recent findings are in accord with both earlierand more recent studies of the essential roles of polar-ized and temporally regulated ion fluxes in the controlof polarized growth in root hairs and pollen tubes.Earlier experiments with external voltage-sensitive self-referencing electrodes [15] have shown that the growingroot hair generates a polarized current flux along itslength. Positive current carried mainly by H+ ionsenters the growing apex and leaves from sub-apicalregions. Moreover, localized elevations of H+ and Ca2+

are intimately associated with root hair initiation orgrowth [16,17], and probably serve to regulate inter aliaexocytosis at the growing apex.

Exactly how K+ fluxes interact with H+ and Ca2+

fluxes in the regulation of root hair growth is not

clear. Measurements of K+ fluxes around root hairs withK+-selective self-referencing electrodes have not shownany distinct localization of K+ entry [18]. One possibil-ity is that K+ entry through either TRH-like transportersor K+ channels may regulate the activity of hyperpo-larization-activated calcium channels that have beenshown to be present in Arabidopsis root hair apices[19] by limiting the membrane hyperpolarization broughtabout by the activity of the H+ pump. Hyperpolariza-tion-activated calcium channels allow Ca2+ influx inresponse to membrane hyperpolarization and theiractivity, shown in patch clamp studies to be higher atthe root hair apex, may underlie the elevated cytoso-lic Ca2+ at the root hair apex.

Elevations of Ca2+ and H+ at the growing apex, as inroot hairs, appear to be essential features of pollentube polarized growth. Ca2+ and pH in this region bothshow regular oscillations in relation to growth rate[7,8,20]. Recently Messerli et al. [7], using external ion-selective self-referencing electrodes, recorded inwardK+ and H+ fluxes at the pollen tube apex that were bal-anced by H+ efflux from the pollen grain. The K+ andH+ influxes occurred as pulses that were in phase withone another, and their magnitude was dictated by thesize of the preceding growth pulse. Similarly, influx ofCa2+ was also shown to be coincident with, but anorder of magnitude lower than, the H+ and K+ fluxes.While parallel measurements have not been madewith Arabidopsis pollen, it is tempting to conclude thatSPIK channels may underlie a polarized K+ influxsimilar to that found in lily pollen.

The cellular localization of SPIK and TRH in thepollen tube or root hair respectively, together withmeasurements of K+ and H+ fluxes in mutants com-pared with wild-type plants and consequent effects onapical Ca2+ and pH gradients will be required tounderstand more fully the essential roles of these K+

transport processes in polarized cell growth.

References1. Maser, P.S., Thomine, S., Schroeder, J.I., Ward, J.M., Hirschi, K.,

Sze, H., Talke, I., Amtmann, A., Maathuis, F.J., Sanders, D. et al.(2001). Phylogenetic relationships within cation transporter familiesof Arabidopsis. Plant Physiol. 126, 1446–1467.

2. Zimmermann, S. and Sentenac, H. (1999). Plant ion channels: frommolecular structures to physiological functions. Curr. Opin. PlantBiol. 2, 477–482.

Figure 1. Models of the possibleinvolvement of K+ fluxes in pollen tubeand root hair polarized growth.

In a pollen tube (left), localized K+ influx atthe growth tube apex contributes to thetotal inward cation flux in this region. Theessential role of K+ influx for pollen tubegrowth is revealed by the defectivegrowth characteristics of spik1 mutantsdefective in an inward rectifying K+

channel, raising the possibility that SPIK1channels may carry the K+ influx at thetube apex. In a root hair (right), K+ influxby the K+ transporter TRH may serve toregulate the activity of hyperpolarization-

activated Ca2+ channels (HACC) at the growing apex via regulation of the membrane potential. In both pollen tubes and root hairs, ahyperpolarized membrane potential results from the activity of the electrogenic H+-extruding ATPase (blue circles). Preferential influxof H+ and Ca2+ at the growing apices leads to the formation of localized elevated concentrations of these ions.

H+

H+

H+

H+H+

+

SPIK1?K+

H+

Ca2+––

+

+

–

++

+

+

–

–

––

v

?TRH

H+

Ca2+

(HACC)K+

K+

–

K+

K+K+

H+

H+H+

H+

–-

––

–

+

Pollen tube Root hair

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3. Santa-Maria, G.E., Rubio, F., Dubkovsky, J. and Rodriguez-Navarro,A. (1997). The HAK1 gene of barley is a member of a large genefamily and encodes a high affinity potassium transporter. Plant Cell9, 2281–2289.

4. Rigas, S., Debrosses, G., Harampidis, K., Vicente-Agullo, F., Feld-mann, K.A., Grabov, A., Dolan, L. and Hatzopoulos, P. (2001). TRH1Encodes a potassium transporter required for tip growth in Ara-bidospsis root hairs. Plant Cell 13, 139–151.

5. Mouline, K., Verey, A.-E., Gaymard, F., Boucherez, J., Pilot, G.,Devic, M., Bouchez, D., Thibaud, J.-B. and Sentenac, H. (2002).Pollen tube development and competitive ability are impaired bydisruption of a Shaker K+ channel in Arabidopsis. Genes Dev. 16,339–350.

6. Messerli, M.A., Creton, R., Jaffe, L.F. and Robinson, K.R. (2000).Periodic increases in elongation rate precede increases in cytoso-lic Ca2+ during pollen tube growth. Dev. Biol. 222, 84–98.

7. Messerli, M.A., Danuser, G. and Robinson, K.R. (1999). Pulsatileinfluxes of H+, K+ and Ca2+ lag growth of Lilium longiflorum pollentubes. J. Cell Sci. 112, 1497–1509.

8. Hisch, R.E., Lewis, B.D., Spalding, E.P. and Sussman, M.R. (1998).A role for the AKT1 potassium channel in plant nutrition. Science280, 918–921.

9. Spalding, E.P., Hirsch, R.E., Lewis, D.R., Qi, Z., Sussman, M.R. andLewis, B.D. (1999). Potassium uptake supporting plant growth in theabsence of AKT1 channel activity - inhibition by ammonium andstimulation by sodium. J. Gen. Physiol. 113, 909–918.

10. Dennison, K.L., Robertson, W.R., Lewis, B.D., Hirsch, R.E.,Sussman, M.R. and Spalding, E.P. (2001). Functions of AKT1 andAKT2 potassium channels determined by studies of single anddouble mutants of Arabidopsis. Plant Physiol. 127, 1012–1019.

11. Rubio, F., Gassmann, W. and Schroeder, J.I. (1995). Sodium-drivenpotassium uptake by the plant potassium transporter HKT1 andmutations conferring salt tolerance. Science 270, 1660–1663.

12. Kim, E.J., Kwak, J.M., Uozumi, N. and Schroeder, J.I. (1998). AnArabidopsis gene encoding high affinity potassium transport activ-ity. Plant Cell 10, 51–62.

13. Fu, H.H. and Luan, S. (1998). AtKUP1: A dual affinity K+ transporterfrom Arabidopsis. Plant Cell 10, 63–73.

14. Quintero, F.J. and Blatt, M.R. (1997). A new family of K+ transportersfrom Arabidopsis that are conserved across phyla. FEBS Lett. 415,206–211.

15. Weisenseel, M.H., Dorn, A. and Jaffe, L.F. (1979). Natural H+ cur-rents traverse growing roots and root hairs of barley (Hordeumvulgare L.). Plant Physiol. 64, 512–518.

16. Bibikova, T.N., Jacob, T., Dahse, I. and Gilroy, S. (1998). Localizedchanges in apoplastic and cytoplasmic pH are associated with roothair development in Arabidopsis thaliana. Development 125,2925–2934.

17. Wymer, C.L., Bibikova, T.N. and Gilroy, S. (1997). Cytoplasmic freecalcium distributions during the development of root hairs of Ara-bidopsis thaliana. Plant J. 12, 427–439.

18. Jones, D.L., Shaff, J.E. and Kochian, L.V. (1995). Role of calciumand other ions in directing root hair tip growth in Limobiumstoloniferum. 1. Inhibition of tip growth by aluminium. Planta 197,672–680.

19. Very, A.A. and Davies, J.M. (2000). Hyperpolarization-activatedcalcium channels at the tip of Arabidopsis root hairs. Proc. Natl.Acad. Sci. U.S.A. 97, 9801–9806.

20. Holdaway-Clarke, T.L. Feijo, J.F., Hackett, G.R., Kunkel, J.G. andHepler, P.K. (1997). Pollen tube growth and the intracellular cytoso-lic calcium gradient oscillates in phase while extracellular calciuminflux is delayed. Plant Cell 9, 1999–2010.