The physiological role and mechanism of nutrient storage within vacuoles of specific cell types is poorly understood. Transcript profiles from Arabidopsis thaliana leaf cells differing in calcium concentration ([Ca], epidermis <10 mM versus mesophyll >60 mM) were compared using a microarray screen and single-cell quantitative PCR. Three tonoplast-localized Ca 2+ transporters, CAX1 (Ca 2+ /H + -antiporter), ACA4, and ACA11 (Ca 2+ -ATPases), were identified as preferentially expressed in Ca-rich mesophyll. Analysis of respective loss-of-function mutants demonstrated that only a mutant that lacked expression of both CAX1 and CAX3, a gene ectopically expressed in leaves upon knockout of CAX1, had reduced mesophyll [Ca]. Reduced capacity for mesophyll Ca accumulation resulted in reduced cell wall extensibility, stomatal aperture, transpiration, CO 2 assimilation, and leaf growth rate; increased transcript abundance of other Ca 2+ transporter genes; altered expression of cell wall-modifying proteins, including members of the pectinmethylesterase, expansin, cellulose synthase, and polygalacturonase families; and higher pectin concentrations and thicker cell walls. We demonstrate that these phenotypes result from altered apoplastic free [Ca 2+ ], which is threefold greater in cax1/cax3 than in wild-type plants. We establish CAX1 as a key regulator of apoplastic [Ca 2+ ] through compartmentation into mesophyll vacuoles, a mechanism essential for optimal plant function and productivity.

Cation levels within the cytosol are coordinated by a network of transporters. Here, we examine the functional roles of calcium exchanger 1 (CAX1), a vacuolar H 1 /Ca 21 transporter, and the closely related transporter CAX3. We demonstrate that like CAX1, CAX3 is also localized to the tonoplast. We show that CAX1 is predominately expressed in leaves, while CAX3 is highly expressed in roots. Previously, using a yeast assay, we demonstrated that an N-terminal truncation of CAX1 functions as an H At the whole-plant level, it has been well documented that there is a complex interplay among various ions (Marschner, 1995). For example, supplemental Ca 21 is known to mitigate the adverse effects of salinity on plant growth (Epstein, 1972). Recently, it has become possible to measure the sum total of the plant's mineral nutrient and trace element composition, termed the ionome (Lahner et al., 2003). The ionome phenotypes now allow investigators to assess how alterations in specific transporters affect these ionic relationships.Ca 21 and other cations can accumulate to millimolar levels in the vacuole, whereas the concentrations of these cations are maintained in the micromolar range in the cytosol (Taiz et al., 1990; Marty, 1999 (Sze et al., 2000). The driving force for cation antiport activity is the pH gradient generated by two electrogenic proton pumps located on the membrane, an ATPase and a pyrophosphatase (PPase; Sze et al., 1999). In principle, the proton pumps and the H 1 / cation exchangers can both dramatically alter the cation content of the vacuoles.Plant H Article, publication date, and citation information can be found at www.plantphysiol.org/cgi

The Arabidopsis Ca 2 ؉ /H ؉ transporter CAX1 (Cation Exchanger1) may be an important regulator of intracellular Ca 2 ؉ levels. Here, we describe the preliminary localization of CAX1 to the tonoplast and the molecular and biochemical characterization of cax1 mutants. We show that these mutants exhibit a 50% reduction in tonoplast Ca 2 ؉ /H ؉ antiport activity, a 40% reduction in tonoplast V-type H ؉ -translocating ATPase activity, a 36% increase in tonoplast Ca 2 ؉ -ATPase activity, and increased expression of the putative vacuolar Ca 2 ؉ /H ؉ antiporters CAX3 and CAX4 . Enhanced growth was displayed by the cax1 lines under Mn 2 ؉ and Mg 2 ؉ stress conditions. The mutants exhibited altered plant development, perturbed hormone sensitivities, and altered expression of an auxin-regulated promoter-reporter gene fusion. We propose that CAX1 regulates myriad plant processes and discuss the observed phenotypes with regard to the compensatory alterations in other transporters.