Membrane transport systems active in cellular inorganic phosphate (P(i)) acquisition play a key role in maintaining cellular P(i) homeostasis, independent of whether the cell is a unicellular microorganism or is contained in the tissue of a higher eukaryotic organism. Since unicellular eukaryotes such as yeast interact directly with the nutritious environment, regulation of P(i) transport is maintained solely by transduction of nutrient signals across the plasma membrane. The individual yeast cell thus recognizes nutrients that can act as both signals and sustenance. The present review provides an overview of P(i) acquisition via the plasma membrane P(i) transporters of Saccharomyces cerevisiae and the regulation of internal P(i) stores under the prevailing P(i) status.

In this study, the putative anion transporter 1 (ANTR1) from Arabidopsis thaliana was shown to be localized to the chloroplast thylakoid membrane by Western blotting with two different peptide-specific antibodies. ANTR1 is homologous to the type I of mammalian Na ؉ -dependent inorganic phosphate (P i ) transporters. The function of ANTR1 as a Na ؉ -dependent P i transporter was demonstrated by heterologous expression and uptake of radioactive P i into Escherichia coli cells. The expression of ANTR1 conferred increased growth rates to the transformed cells and stimulated P i uptake in a pH-and Na ؉ -dependent manner as compared with the control cells. Among various tested effectors, P i was the preferred substrate. Although it competed with the uptake of P i , glutamate was not transported by ANTR1 into E. coli. In relation to its function as a P i transporter, several physiological roles for ANTR1 in the thylakoid membrane are proposed, such as export of P i produced during nucleotide metabolism in the thylakoid lumen back to the chloroplast stroma and balance of the trans-thylakoid H ؉ electrochemical gradient storage.Solute and metabolite transporters play essential roles in physiological processes, including nutrient uptake, cell homeostasis, signal transduction, growth, and stress responses in every living organism. In the plant chloroplast, the photosynthetic organelle, most transporters have been identified and biochemically characterized from the envelope membrane (1, 2). Among them, there are several translocators for inorganic phosphate (P i ), all functioning as antiport systems using P i or phosphorylated C3 and C6 compounds as counter substrates (3, 4). Much less information is available for transport processes across the chloroplast thylakoid membrane, which is mostly studied as the site of light-driven photosynthetic reactions coupled to ATP synthesis. Only a few thylakoid transporters have been identified and functionally characterized. Examples are ATP transport across spinach thylakoid membrane into the lumenal space and a thylakoid ATP/ADP carrier identified and characterized in Arabidopsis thaliana (5,6). An active nucleotide metabolism in the thylakoid lumen (5) implies the existence of additional, yet unidentified, transporters, such as those recycling P i to the soluble stroma.A few Na ϩ -coupled P i transporters (NaP i ) 6 have in recent years been reported in green algae and vascular plants (7,8). NaP i systems are known to be mostly active in mammals, whereas H ϩ -coupled P i transport is dominant in plants (4). Three NaP i types have been described in eukaryotes, NaP i -II and III being the main ones in mammals (9 -11). NaP i -II type plays the role of an intracellular P i accumulation system, whereas NaP i -III type has the characteristics of a housekeeping system (9). The molecular mechanisms controlling the NaP i -II and III uptake systems have recently been reviewed (11). NaP i -I represents a group of proteins for which the endogenous substrate, ionic coupling, and physiological fu...

In Saccharomyces cerevisiae, phosphate uptake is mainly dependent on the proton-coupled Pho84 permease under phosphate-limited growth conditions. Phosphate addition causes Pho84-mediated activation of the protein kinase A (PKA) pathway as well as rapid internalization and vacuolar breakdown of Pho84. We show that Pho84 undergoes phosphate-induced phosphorylation and subsequent ubiquitination on amino acids located in the large middle intracellular loop prior to endocytosis. The attachment of ubiquitin is dependent on the ubiquitin conjugating enzymes Ubc2 and Ubc4. In addition, we show that the Pho84 endocytotic process is delayed in strains with reduced PKA activity. Our results suggest that Pho84-mediated activation of the PKA pathway is responsible for its own downregulation by phosphorylation, ubiquination, internalization, and vacuolar breakdown.

The Na(+)-coupled, high-affinity Pho89 plasma membrane phosphate transporter in Saccharomyces cerevisiae has so far been difficult to study because of its low activity and special properties. In this study, we have used a pho84Deltapho87Deltapho90Deltapho91Delta quadruple deletion strain of S. cerevisiae devoid of all transporter genes specific for inorganic phosphate, except for PHO89, to functionally characterize Pho89 under conditions where its expression is hyperstimulated. Under these conditions, the Pho89 protein is strongly upregulated and is the sole high-capacity phosphate transporter sustaining cellular acquisition of inorganic phosphate. Even if Pho89 is synthesized in cells grown at pH 4.5-8.0, the transporter is functionally active under alkaline conditions only, with a K(m) value reflecting high-affinity properties of the transporter and with a transport rate about 100-fold higher than that of the protein in a wild-type strain. Even under these hyperexpressive conditions, Pho89 is unable to sense and signal extracellular phosphate levels. In cells grown at pH 8.0, Pho89-mediated phosphate uptake at alkaline pH is cation-dependent with a strong activation by Na(+) ions and sensitivity to carbonyl cyanide m-chlorophenylhydrazone. The contribution of H(+)- and Na(+)-coupled phosphate transport systems in wild-type cells grown at different pH values was quantified. The contribution of the Na(+)-coupled transport system to the total cellular phosphate uptake activity increases progressively with increasing pH.

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