The phloem network is as essential for plants as the vascular system is for humans. This network, assembled by nucleus-and vacuole-free interconnected living cells, represents a long distance transport pathway for nutrients and information. According to the Mü nch hypothesis, osmolytes such as sucrose generate the hydrostatic pressure that drives nutrient and water flow between the source and the sink phloem (Mü nch, E. (1930) Die Stoffbewegungen in der Pflanze, Gustav Fischer, Jena, Germany). Although proton-coupled sucrose carriers have been localized to the sieve tube and the companion cell plasma membrane of both source and sink tissues, knowledge of the molecular representatives and the mechanism of the sucrose phloem efflux is still scant. We expressed ZmSUT1, a maize sucrose/proton symporter, in Xenopus oocytes and studied the transport characteristics of the carrier by electrophysiological methods. Using the patch clamp techniques in the giant inside-out patch mode, we altered the chemical and electrochemical gradient across the sucrose carrier and analyzed the currents generated by the proton flux. Thereby we could show that ZmSUT1 is capable of mediating both the sucrose uptake into the phloem in mature leaves (source) as well as the desorption of sugar from the phloem vessels into heterotrophic tissues (sink). As predicted from a perfect molecular machine, the ZmSUT1-mediated sucrose-coupled proton current was reversible and depended on the direction of the sucrose and pH gradient as well as the membrane potential across the transporter.To ensure adequate partitioning of sucrose throughout the plant body, sucrose has to be translocated from the mesophyll cells to the sieve element-companion cell complex. Because of the energy-dependent sucrose/H ϩ symporter in apoplasmic loading plant species, the transport sugar accumulates at concentrations of several hundred mM to Ͼ1 molar in the conducting vascular cells. The hydrostatic pressure difference between source and sink tissues drives the mass flow of water and nutrients in the phloem vessels (1). In sink tissues, which are dependent on carbon supply via the phloem, a symplasmic unloading of sucrose along its concentration gradient has been shown for many plant species (2). Interestingly, however, sucrose/H ϩ symporter transcripts and proteins have also been localized in sink tissues, suggesting a role in sink loading/ retrieval or unloading of sucrose via these transporters (see Ref. 2 for review). SUT1 from the potato, for example, has been detected in the sieve elements of mature source leaves as well as in developing sink leaves, roots (3), and tubers (4, 5). Using a sink-specific antisense inhibition for SUT1 under the control of a tuber-specific promoter, Kü hn et al. (2003) (4) could demonstrate the involvement of SUT1 in early tuber development and, thus, phloem unloading. Further evidence for a sucrose export system was added by the localization of sucrose/H ϩ symporters expressed in symplasmically isolated tissues such as developing embryos (6...

The Arabidopsis tandem-pore K ؉ (TPK) channels displaying four transmembrane domains and two pore regions share structural homologies with their animal counterparts of the KCNK family. In contrast to the Shaker-like Arabidopsis channels (six transmembrane domains͞one pore region), the functional properties and the biological role of plant TPK channels have not been elucidated yet. Here, we show that AtTPK4 (KCO4) localizes to the plasma membrane and is predominantly expressed in pollen. AtTPK4 (KCO4) resembles the electrical properties of a voltage-independent K ؉ channel after expression in Xenopus oocytes and yeast. Hyperpolarizing as well as depolarizing membrane voltages elicited instantaneous K ؉ currents, which were blocked by extracellular calcium and cytoplasmic protons. Functional complementation assays using a K ؉ transport-deficient yeast confirmed the biophysical and pharmacological properties of the AtTPK4 channel. The features of AtTPK4 point toward a role in potassium homeostasis and membrane voltage control of the growing pollen tube. Thus, AtTPK4 represents a member of plant tandem-pore-K ؉ channels, resembling the characteristics of its animal counterparts as well as plant-specific features with respect to modulation of channel activity by acidosis and calcium.

Sponges (phylum Porifera) are the phylogenetically oldest metazoan animals, their evolution dating back to 600 million years ago. Here we demonstrate that sponges express ADP-ribosyl cyclase activity, which converts NAD ؉ into cyclic ADP-ribose, a potent and universal intracellular Ca 2؉ mobilizer. In Axinella polypoides (Demospongiae, Axinellidae), ADP-ribosyl cyclase was activated by temperature increases by means of an abscisic acid-induced, protein kinase A-dependent mechanism. The thermosensor triggering this signaling cascade was a heat-activated cation channel. Elucidation of the complete thermosensing pathway in sponges highlights a number of features conserved in higher organisms: (i) the cation channel thermoreceptor, sensitive to heat, mechanical stress, phosphorylation, and anesthetics, shares all of the functional characteristics of the mammalian heat-activated background K ؉ channel responsible for central and peripheral thermosensing; (ii) involvement of the phytohormone abscisic acid and cyclic ADP-ribose as its second messenger is reminiscent of the drought stress signaling pathway in plants. These results suggest an ancient evolutionary origin of this stress-signaling cascade in a common precursor of modern Metazoa and Metaphyta.A DP-ribosyl cyclase activity is expressed along the phylogenetic tree from unicellular protists (1) to mammals (2). It converts NAD ϩ to cyclic ADP-ribose (cADPR) (3), an intracellular calcium mobilizer that is being increasingly recognized as a pivotal signaling molecule, involved in such diverse functions as cell cycle regulation (1) (protists), oocyte fertilization (4) (invertebrates), insulin secretion (5), and cell proliferation (6) (mammals). The peculiar position of Porifera in the phylogenetic tree [they are the oldest known Metazoa, sharing a common ancestor with all multicellular animals (7)] and the presence of signaling pathways in these organisms (8) prompted us to investigate the presence, and the functional role, of the ADPribosyl cyclase͞cADPR system in marine sponges. Materials and MethodsSponges. Sponges were collected in the Ligurian Sea (Mediterranean Sea), at Gallinara Island (Genova, Italy) at a depth between 35 and 45 m. The temperature was maintained at 14°C during transfer of the animals to the lab. They were kept in an aquarium in natural seawater (SW) at 14°C for 5-7 days before utilization. All procedures involving sponge tissue were performed at 14°C. As the most ancient and simple metazoan animals, sponges lack a defined tissue organization. Cells are embedded in a collagenous matrix, impregnated with siliceous or calcareous spicules, surrounding a complex network of internal canals, through which water circulates. Intact sponge cells (Ϸ8 m diameter) could be easily obtained by gentle squeezing of cleanly cut Axinella polypoides fragments. Cell viability, as checked microscopically, was always Ն95% after mechanical dissociation and after exposure of the cells to heat stress or abscisic acid (ABA).HPLC and Fluorimetric Assays of Enzymatic A...

Highlights d HKT1-type channels mediate a one-way sodium load into quinoa bladder cells d ClC transporters operate in the Cl À sequestration into vacuoles of bladder cells d The bladder cytoplasm is osmotically balanced by potassium and proline import d On the transcript level, bladders are ''constitutively active'' for salt sequestration