Frans J. M. Maathuis

Physiological functions of mineral macronutrients.

Plants require calcium, magnesium, nitrogen, phosphorous, potassium and sulfur in relatively large amounts (>0.1% of dry mass) and each of these so-called macronutrients is essential for a plant to complete its life cycle. Normally, these minerals are taken up by plant roots from the soil solution in ionic form with the metals Ca(2+), Mg(2+) and K(+) present as free cations, P and S as their oxyanions phosphate (PO(4)(3-)) and sulfate (SO(4)(2-)) and N as anionic nitrate (NO(3)(-)) or cation ammonium (NH(4)(+)). Recently, important progress has been made in identifying transport and regulatory mechanisms for macronutrients and the mechanisms of uptake and distribution. These and the main physiological roles of each nutrient will be discussed.

The vacuolar Ca2+-activated channel TPC1 regulates germination and stomatal movement

Cytosolic free calcium ([Ca2+]cyt) is a ubiquitous signalling component in plant cells. Numerous stimuli trigger sustained or transient elevations of [Ca2+]cyt that evoke downstream stimulus-specific responses. Generation of [Ca2+]cyt signals is effected through stimulus-induced opening of Ca2+-permeable ion channels that catalyse a flux of Ca2+ into the cytosol from extracellular or intracellular stores. Many classes of Ca2+ current have been characterized electrophysiologically in plant membranes. However, the identity of the ion channels that underlie these currents has until now remained obscure. Here we show that the TPC1 (‘two-pore channel 1’) gene of Arabidopsis thaliana encodes a class of Ca2+-dependent Ca2+-release channel that is known from numerous electrophysiological studies as the slow vacuolar channel. Slow vacuolar channels are ubiquitous in plant vacuoles, where they form the dominant conductance at micromolar [Ca2+]cyt. We show that a tpc1 knockout mutant lacks functional slow vacuolar channel activity and is defective in both abscisic acid-induced repression of germination and in the response of stomata to extracellular calcium. These studies unequivocally demonstrate a critical role of intracellular Ca2+-release channels in the physiological processes of plants.

Roles of Higher Plant K+ Channels

Living organisms maintain a cellular solute composition very different from that of the external environment. This implicitly requires the transport of solutes across the cell membrane, and ion channels are integral membrane proteins that play indispensable roles in such transport. In the past dozen years, radical advances have aided in our understanding of ion channel function and regulation in higher plants. Nowhere are these advances more striking than with respect to K+ channels, where the synergistic application of electrophysiological, cell biological, physiological, and molecular techniques has demonstrated an array of channel types playing diverse but defined roles in plant physiology. The major function of K+ channels in animal cells is that of membrane voltage control and short-term repolarization of the membrane. Although K+ channels in plants share similar roles in the regulation of the membrane voltage, early research on guard cells led to the model that shows that plant K+ channels in addition provide important pathways for long-term physiological K+ uptake and release. An extensive range of recent studies suggests diverse longterm transport functions of plant K+ channels, including participation in osmotically driven movements, solute loading into the xylem, cation nutrition, and, by virtue of the presence of K+ channels at endomembranes, intracel

CNGCs: prime targets of plant cyclic nucleotide signalling?

Cyclic nucleotide-gated channels (CNGCs) are a recently identified family of plant ion channels. They show a high degree of similarity to Shaker-type voltage-gated channels and contain a C-terminal cyclic nucleotide-binding domain with an overlapping calmodulin-binding domain. Heterologously expressed plant CNGCs show activation by cyclic nucleotides and permeability to monovalent and divalent cations. In plants, downstream effectors of cyclic nucleotide signals have so far remained obscure, and CNGCs might be their prime targets. The unique position of CNGCs as ligand-gated Ca(2+)-permeable channels suggests that they function at key sites where cyclic nucleotide and Ca(2+) signalling pathways interact. Such processes include plant defence responses, and two recently characterized Arabidopsis mutants in CNGC genes indeed show altered pathogen responses.

The two-pore channel TPK1 gene encodes the vacuolar K+ conductance and plays a role in K+ homeostasis

The Arabidopsis thaliana genome contains five genes that encode two pore K+ (TPK) channels. The most abundantly expressed isoform of this family, TPK1, is expressed at the tonoplast where it mediates K+-selective currents between cytoplasmic and vacuolar compartments. TPK1 open probability depends on both cytoplasmic Ca2+ and cytoplasmic pH but not on the tonoplast membrane voltage. The channel shows intrinsic rectification and can be blocked by Ba2+, tetraethylammonium, and quinine. TPK1 current was found in all shoot cell types and shows all of the hallmarks of the previously described vacuolar K (VK) tonoplast channel characterized in guard cells. Characterization of TPK1 loss-of-function mutants and TPK1-overexpressing plants shows that TPK1 has a role in intracellular K+ homeostasis affecting seedling growth at high and low ambient K+ levels. In stomata, TPK1 function is consistent with vacuolar K+ release, and removal of this channel leads to slower stomatal closure kinetics. During germination, TPK1 contributes to the radicle development through vacuolar K+ deposition to provide expansion growth or in the redistribution of essential minerals.

Energization of potassium uptake in Arabidopsis thaliana

Plant roots accumulate K+ from micromolar external concentrations. However, the absence of a firm determination of the trans-plasma-membrane electrochemical gradient for K+ in these conditions has precluded an assessment of whether K+-accumulation requires energization in addition to the driving force provided by the inside-negative membrane electrical potential (Em). To address this question unequivocally, we measured Em, and the cytosolic and external K+-activities in root cells of Arabidopsis thaliana (L.) Heynh. cv. Columbia in conditions in which net K+-accumulation occurs at low external K+ (10 μM). In these conditions, net K+-uptake was about 0.1 μmol · (g FW)-1 · h-1, Em varied between-153 and -129 mV and the cytosolic K+-activity, determined with K+-selective electrodes, was 83 ± 4 mM. These values yield an outwardly-directed driving force on K+ of at least 6.5 kJ · mol-1. Only if external potassium is raised to the region of 1 mM does Em become sufficient to drive net K+-accumulation. It is therefore concluded that at micromolar external K+-activities which prevail in most soils, K+-uptake cannot be solely energized by Em — as exemplified by a channel-mediated mechanism. The nature of the energization mechanism is discussed in relation to processes operating in fungal and algal cells.

The Heat Shock Response in Moss Plants Is Regulated by Specific Calcium-Permeable Channels in the Plasma Membrane

Land plants are prone to strong thermal variations and must therefore sense early moderate temperature increments to induce appropriate cellular defenses, such as molecular chaperones, in anticipation of upcoming noxious temperatures. To investigate how plants perceive mild changes in ambient temperature, we monitored in recombinant lines of the moss Physcomitrella patens the activation of a heat-inducible promoter, the integrity of a thermolabile enzyme, and the fluctuations of cytoplasmic calcium. Mild temperature increments, or isothermal treatments with membrane fluidizers or Hsp90 inhibitors, induced a heat shock response (HSR) that critically depended on a preceding Ca2+ transient through the plasma membrane. Electrophysiological experiments revealed the presence of a Ca2+-permeable channel in the plasma membrane that is transiently activated by mild temperature increments or chemical perturbations of membrane fluidity. The amplitude of the Ca2+ influx during the first minutes of a temperature stress modulated the intensity of the HSR, and Ca2+ channel blockers prevented HSR and the onset of thermotolerance. Our data suggest that early sensing of mild temperature increments occurs at the plasma membrane of plant cells independently from cytosolic protein unfolding. The heat signal is translated into an effective HSR by way of a specific membrane-regulated Ca2+ influx, leading to thermotolerance.

A secretory pathway-localized cation diffusion facilitator confers plant manganese tolerance

Manganese toxicity is a major problem for plant growth in acidic soils, but cellular mechanisms that facilitate growth in such conditions have not been clearly delineated. Established mechanisms that counter metal toxicity in plants involve chelation and cytoplasmic export of the metal across the plasma or vacuolar membranes out of the cell or sequestered into a large organelle, respectively. We report here that expression of the Arabidopsis and poplar MTP11 cation diffusion facilitators in a manganese-hypersensitive yeast mutant restores manganese tolerance to wild-type levels. Microsomes from yeast expressing AtMTP11 exhibit enhanced manganese uptake. In accord with a presumed function of MTP11 in manganese tolerance, Arabidopsis mtp11 mutants are hypersensitive to elevated levels of manganese, whereas plants overexpressing MTP11 are hypertolerant. In contrast, sensitivity to manganese deficiency is slightly decreased in mutants and increased in overexpressing lines. Promoter-GUS studies showed that AtMTP11 is most highly expressed in root tips, shoot margins, and hydathodes, but not in epidermal cells and trichomes, which are generally associated with manganese accumulation. Surprisingly, imaging of MTP11–EYFP fusions demonstrated that MTP11 localizes neither to the plasma membrane nor to the vacuole, but to a punctate endomembrane compartment that largely coincides with the distribution of the trans-Golgi marker sialyl transferase. Golgi-based manganese accumulation might therefore result in manganese tolerance through vesicular trafficking and exocytosis. In accord with this proposal, Arabidopsis mtp11 mutants exhibit enhanced manganese concentrations in shoots and roots. We propose that Golgi-mediated exocytosis comprises a conserved mechanism for heavy metal tolerance in plants.

Contrasting roles in ion transport of two K+-channel types in root cells of Arabidopsis thaliana

Plant roots accumulate K+ over a range of external concentrations. Root cells have evolved at least two parallel plasma-membrane K+ transporters which operate at millimolar and micromolar external [K+]: high-affinity K+ uptake is energised by symport with H+, while low-affinity uptake is assumed to occur via ion channels. To determine the role of ion channels in low-affinity K+ uptake, a characterisation of the principal K+-selective ion channels in the plasma membrane of Arabidopsis thaliana (L.) Heynh. cv. Columbia roots was undertaken. Two classes of K+-selective channels were frequently observed: one inward (IRC) and one outward (ORC) rectifying with unitary conductances of 5 pS, 20 pS (IRCs) and 15 pS (ORC), measured in symmetrical 10 mM KCl. The dominant IRC (5 pS) and ORC (15 pS) were highly cation-selective (PCl PK < 0.025) but less selective amongst monovalent cations (PNa∶PK≈0.17–0.3). Both the IRC and the ORC were blocked by Ba2+, Cs+ and tetra-ethyl-ammonium, whereas 4-aminopyridine and quinidine selectively inhibited the ORC. The ORC open probability was steeply voltage-dependent and ORC activation potentials were close to the potassium equilibrium potential (EK+), enabling ORCs to conduct mainly outward, but occasionally inward, K+ current. By contrast, gating of the 5-pS IRC was weakly voltageependent and IRC gating was invariably restricted to membrane potentials more negative than EK+, ensuring K+ transport was always inwardly directed. Studies on channel activity were conducted for a large number of root cells grown at two levels of external [K+], one where K+ uptake is likely to be principally through channels (6 mM K+) and one where it must be energised (100 μM K+). Shifting growth conditions from high to low K+ did not affect single-channel properties such as conductance and selectivity, nor the manifestation of the ORC and 20-pS IRC, but led to enhanced activity of the 5-pS IRC. The enhanced activity of the 5-pS IRC was mirrored by a parallel increase in unidirectional 86Rb+ influx after low-K+ growth, clearly indicating a dominant role of this particular channel in K+ uptake at supra millimolar external [K+].

The Arabidopsis thaliana aquaglyceroporin AtNIP7;1 is a pathway for arsenite uptake

The data suggest that AtNIP7;1 can mediate AsIII transport and contributes to AsIII uptake in plants.