ABSTRACT
The regulation of intracellular ion concentrations is a fundamental property of living cells. Although many ion transporters have been identified, the systems that modulate their activity remain largely unknown. We have characterized two partially redundant genes from Saccharomyces cerevisiae, HAL4/SAT4 and HAL5, that encode homologous protein kinases implicated in the regulation of cation uptake. Overexpression of these genes increases the tolerance of yeast cells to sodium and lithium, whereas gene disruptions result in greater cation sensitivity. These phenotypic effects of the mutations correlate with changes in cation uptake and are dependent on a functional Trk1-Trk2 potassium transport system. In addition, hal4 hal5 and trk1 trk2 mutants exhibit similar phenotypes: (i) they are deficient in potassium uptake; (ii) their growth is sensitive to a variety of toxic cations, including lithium, sodium, calcium, tetramethylammonium, hygromycin B, and low pH; and (iii) they exhibit increased uptake of methylammonium, an indicator of membrane potential. These results suggest that the Hal4 and Hal5 protein kinases activate the Trk1-Trk2 potassium transporter, increasing the influx of potassium and decreasing the membrane potential. The resulting loss in electrical driving force reduces the uptake of toxic cations and improves salt tolerance. Our data support a role for regulation of membrane potential in adaptation to salt stress that is mediated by the Hal4 and Hal5 kinases.
Subject(s)
Carrier Proteins/metabolism , Cation Transport Proteins , Fungal Proteins/metabolism , Membrane Proteins/metabolism , Potassium/metabolism , Protein Kinases/metabolism , Saccharomyces cerevisiae Proteins , Saccharomyces cerevisiae/metabolism , Salts/metabolism , Biological Transport , Cations/pharmacology , Cloning, Molecular , Gene Expression Regulation, Fungal , Homeostasis , Membrane Potentials , Methylamines , Mutation , PhenotypeABSTRACT
We report the cloning of both the cDNA and the corresponding genomic sequence of a new PP2C from Arabidopsis thaliana, named AtP2C-HA (for homology to ABI1/ABI2). The AtP2C-HA cDNA contains an open reading frame of 1536 bp and encodes a putative protein of 511 amino acids with a predicted molecular mass of 55.7 kDa. The AtP2C-HA protein is composed of two domains, a C-terminal PP2C catalytic domain and a N-terminal extension of ca. 180 amino acid residues. The deduced amino acid sequence is 55% and 54% identical to ABI1 and ABI2, respectively. Comparison of the genomic structure of the ABI1, ABI2 and AtP2C-HA genes suggests that they belong to a multigene family. The expression of the AtP2C-HA gene is up-regulated by abscisic acid (ABA) treatment.
Subject(s)
Arabidopsis Proteins , Arabidopsis/genetics , Phosphoprotein Phosphatases/genetics , Saccharomyces cerevisiae Proteins , Abscisic Acid/pharmacology , Amino Acid Sequence , Arabidopsis/chemistry , Arabidopsis/enzymology , Base Sequence , Blotting, Northern , Blotting, Southern , Cloning, Molecular , DNA, Complementary/chemistry , DNA, Complementary/genetics , DNA, Complementary/isolation & purification , DNA, Plant/chemistry , DNA, Plant/genetics , Exons , Gene Expression Regulation, Enzymologic/drug effects , Gene Expression Regulation, Plant/drug effects , Genes, Plant/genetics , Introns , Molecular Sequence Data , Protein Phosphatase 2 , Protein Phosphatase 2C , RNA, Messenger/drug effects , RNA, Messenger/genetics , RNA, Messenger/metabolism , Sequence Alignment , Sequence Analysis, DNA , Sequence Homology, Amino Acid , Tissue DistributionABSTRACT
The plant hormone abscisic acid (ABA) mediates various responses such as stomatal closure, maintenance of seed dormancy, and inhibition of plant growth. All three responses are regulated by the ABI1 gene product. The ABI1 protein (ABI1p) has been characterized as a protein serine/threonine phosphatase of type 2C that is highly affected in its activity by changes in the proton and magnesium ion concentrations. In the ABA-insensitive mutant abi1 of Arabidopsis thaliana a single amino acid exchange in the primary structure results in both a dominant insensitive phenotype and a strongly reduced protein phosphatase activity in vitro by possibly impairing metal ion coordination.
Subject(s)
Arabidopsis Proteins , Phosphoprotein Phosphatases/drug effects , Phosphoprotein Phosphatases/isolation & purification , Plant Proteins/isolation & purification , Arabidopsis/enzymology , Dose-Response Relationship, Drug , Hydrogen-Ion Concentration , Magnesium/pharmacology , Phosphoprotein Phosphatases/metabolism , Plant Proteins/metabolism , Protons , Serine/metabolism , Threonine/metabolismABSTRACT
The plant hormone abscisic acid (ABA) mediates various responses such as stomatal closure, the maintenance of seed dormancy, and the inhibition of plant growth. All three responses are affected in the ABA-insensitive mutant abi1 of Arabidopsis thaliana, suggesting that an early step in the signaling of ABA is controlled by the ABI1 locus. The ABI1 gene was cloned by chromosome walking, and a missense mutation was identified in the structural gene of the abi1 mutant. The ABI1 gene encodes a protein with high similarity to protein serine or threonine phosphatases of type 2C with the novel feature of a putative Ca2+ binding site. Thus, the control of the phosphorylation state of cell signaling components by the ABI1 product could mediate pleiotropic hormone responses.
