It is generally accepted that K ؉ uptake into guard cells via inwardrectifying K ؉ channels is required for stomatal opening. To test whether the guard cell K ؉ channel KAT1 is essential for stomatal opening, a knockout mutant, KAT1::En-1, was isolated from an En-1 mutagenized Arabidopsis thaliana population. Stomatal action and K ؉ uptake, however, were not impaired in KAT1-deficient plants. Reverse transcription-PCR experiments with isolated guard cell protoplasts showed that in addition to KAT1, the K ؉ channels AKT1, AKT2͞3, AtKC1, and KAT2 were expressed in this cell type. In impalement measurements, intact guard cells exhibited inwardrectifying K ؉ currents across the plasma membrane of both wildtype and KAT1::En-1 plants. This study demonstrates that multiple K ؉ channel transcripts exist in guard cells and that KAT1 is not essential for stomatal action.

The potassium-channel gene, AKT3, has recently been isolated from an Arabidopsis thaliana cDNA library. By using the whole-mount and in situ hybridization techniques, we found AKT3 predominantly expressed in the phloem. To study the physiological role of this channel type, AKT3 was heterologously expressed in Xenopus oocytes, and the electrical properties were examined with voltage-clamp techniques. Unlike the plant inward-rectifying guard cell K ؉ channels KAT1 and KST1, the AKT3 channels were only weakly regulated by the membrane potential. Furthermore, AKT3 was blocked by physiological concentrations of external Ca 2؉ and showed an inverted pH regulation. Extracellular acidification decreased the macroscopic AKT3 currents by reducing the single-channel conductance. Because assimilate transport in the vascular tissue coincides with both H ؉ and K ؉ f luxes, AKT3 K ؉ channels may be involved in K ؉ transport accompanying phloem loading and unloading processes.The plant vascular system, which consists of xylem and phloem, is specialized for long-distance solute and water transport. In both tissues, potassium represents one of the major mineral nutrients and is likely to assist in osmotic homeostasis. After uptake from the soil, K ϩ ions circulate between roots and shoots through the xylem and phloem to adopt the specific demands for this cation in the various tissues (1, 2). Recently, in vivo and in vitro analysis demonstrated the presence of K ϩ uptake and release channels in xylem parenchyma cells (3-6). In comparison, information about K ϩ transport across the plasma membrane of phloem cells, the underlying mechanisms, and the physiological role in long-distance solute transport is still limited. Phloem loading with assimilates is accompanied by ionic movements (7). Protons are pumped into the apoplast by a H ϩ -ATPase, generating transmembrane gradients in electropotential and pH that in turn enable the uphill transport of assimilates into the phloem through assimilate/ H ϩ -cotransporters (8-10). The phloem loading coincides with an increase in the symplastic K ϩ concentration likely to maintain electrical neutrality that is required for creating the pH gradient (2, 7). In addition, the K ϩ concentration in the sieve tube may affect the volume flow rate in the phloem (11). Furthermore, the membrane potential that transiently changes during phloem-propagating action potentials is possibly reestablished on K ϩ release from the sieve tube (12). Evidence for the expression of K ϩ channels in the phloem was recently provided (P.A., K. Philippar, and R.H., unpublished data) to support the idea that K ϩ channels may also mediate the transmembrane K ϩ fluxes in the phloem.In the present paper, we localized the cloned Arabidopsis K ϩ channel AKT3 to the phloem and revealed its unique dependence on voltage, Ca 2ϩ , and pH properties, which are well suited for meeting its supposed role in processes associated with the phloem. MATERIALS AND METHODSRNA Extraction and Northern Blot Analysis. Total RNA was isolate...

Agrobacterium tumefaciens causes crown gall disease by transferring and integrating bacterial DNA (T-DNA) into the plant genome. To examine the physiological changes and adaptations during Agrobacterium-induced tumor development, we compared the profiles of salicylic acid (SA), ethylene (ET), jasmonic acid (JA), and auxin (indole-3-acetic acid [IAA]) with changes in the Arabidopsis thaliana transcriptome. Our data indicate that host responses were much stronger toward the oncogenic strain C58 than to the disarmed strain GV3101 and that auxin acts as a key modulator of the ArabidopsisAgrobacterium interaction. At initiation of infection, elevated levels of IAA and ET were associated with the induction of host genes involved in IAA, but not ET signaling. After T-DNA integration, SA as well as IAA and ET accumulated, but JA did not. This did not correlate with SA-controlled pathogenesis-related gene expression in the host, although high SA levels in mutant plants prevented tumor development, while low levels promoted it. Our data are consistent with a scenario in which ET and later on SA control virulence of agrobacteria, whereas ET and auxin stimulate neovascularization during tumor formation. We suggest that crosstalk among IAA, ET, and SA balances pathogen defense launched by the host and tumor growth initiated by agrobacteria.

Members of the AKT2/3 family have been identified as photosynthate-induced phloem K(+) channels. Here we describe the isolation and characterisation of an AKT2/3 loss-of-function mutant (akt2/3-1) from Arabidopsis thaliana (L.) Heynh. Microautoradiography following (14)CO(2) incubation in the light revealed that a major fraction of (14)CO(2)-derived photosynthates leaking out of sieve tubes appears not to be effectively reloaded (retrieval) into the phloem of the mutant. Using the aphid stylectomy technique we showed that the phloem sap of the mutant, lacking the phloem channels of the AKT2/3 type, contained only half the sucrose content of the wild type. Furthermore, the akt2/3-1 mutant exhibited a reduced K(+) dependence of the phloem potential. Xenopus oocytes expressing the phloem sucrose/proton symporter depolarise upon sucrose application. When, however, the phloem channel was co-expressed - mimicking the situation in the sieve tube/companion cell complex - depolarisation was prevented. From our studies we thus conclude that AKT2/3 regulates the sucrose/H(+) symporters via the phloem potential.

Distribution of K, Ca, Cl, S, and P in freeze-dried sections of Arabidopsis flower stalk was analyzed by energy dispersive x-ray imaging. Concentrations of these elements in different cell types were quantified by microanalysis of single-cell samples and phloem exudates. Results showed a differential pattern of distribution for all five elements. K concentration was found to be highest in the parenchymatous tissue around vascular bundles. Ca and Cl were present mainly in the central part of the flower stalk. P was largely located in the bundles and in the parenchyma surrounding them. S signal was extraordinary high in groups of cells (S-cells) situated between the phloem of every vascular bundle and the endodermis. Enzymatic hydrolysis by thioglucosidase of cell sap collected from S-cells using a glass microcapillary resulted in the release of glucose, indicating that these cells contain glucosinolates at high (> 100 mm) concentration, which is consistent with the concentration of S (> 200 mm) estimated by x-ray analysis of cell sap samples. Since their position outside of the phloem is ideally suited for protecting the long-distance transport system from feeding insects, the possible roles of these cells as components of a plant defense system are discussed.