ABSTRACT
In plants, γ-aminobutyric acid (GABA) accumulates in the cytosol in response to a variety of stresses. GABA is transported into mitochondria, where it is catabolized into TCA cycle or other intermediates. Although there is circumstantial evidence for mitochondrial GABA transporters in eukaryotes, none have yet been identified. Described here is an Arabidopsis protein similar in sequence and topology to unicellular GABA transporters. The expression of this protein complements a GABA-transport-deficient yeast mutant. Thus the protein was termed AtGABP to indicate GABA-permease activity. In vivo localization of GABP fused to GFP and immunobloting of subcellular fractions demonstrate its mitochondrial localization. Direct [(3) H]GABA uptake measurements into isolated mitochondria revealed impaired uptake into mitochondria of a gabp mutant compared with wild-type (WT) mitochondria, implicating AtGABP as a major mitochondrial GABA carrier. Measurements of CO(2) release, derived from radiolabeled substrates in whole seedlings and in isolated mitochondria, demonstrate impaired GABA-derived input into the TCA cycle, and a compensatory increase in TCA cycle activity in gabp mutants. Finally, growth abnormalities of gabp mutants under limited carbon availability on artificial media, and in soil under low light intensity, combined with their metabolite profiles, suggest an important role for AtGABP in primary carbon metabolism and plant growth. Thus, AtGABP-mediated transport of GABA from the cytosol into mitochondria is important to ensure proper GABA-mediated respiration and carbon metabolism. This function is particularly essential for plant growth under conditions of limited carbon.
Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/enzymology , Citric Acid Cycle , GABA Plasma Membrane Transport Proteins/metabolism , Mitochondria/enzymology , gamma-Aminobutyric Acid/metabolism , Amino Acid Sequence , Analysis of Variance , Arabidopsis/genetics , Arabidopsis/growth & development , Arabidopsis Proteins/genetics , Blotting, Southern , Carbon/metabolism , Carbon Dioxide/metabolism , Cytosol/metabolism , GABA Plasma Membrane Transport Proteins/genetics , Genetic Complementation Test , Genetic Vectors , Genotype , Green Fluorescent Proteins/metabolism , Immunoblotting/methods , Light , Microscopy, Confocal , Mutagenesis, Insertional , Open Reading Frames , Plant Leaves/growth & development , Plant Leaves/metabolism , Plant Roots/growth & development , Plant Roots/metabolism , Proline/metabolism , Protoplasts/metabolism , Recombinant Fusion Proteins , Saccharomyces cerevisiae/enzymology , Saccharomyces cerevisiae/genetics , Seedlings/growth & development , Seedlings/metabolism , Nicotiana/genetics , Nicotiana/metabolismABSTRACT
Calmodulin CaM is the most prominent Ca2+ transducer in eukaryotic cells, regulating the activity of numerous proteins with diverse cellular functions. Many features of CaM and its downstream targets are similar in plants and other eukaryotes. However, plants possess a unique set of CaM-related proteins, and several unique CaM target proteins. This review discusses recent progress in identifying plant-specific CaM-binding proteins and their roles in response to biotic and abiotic stresses and development. The review also addresses aspects emerging from recent structural studies of CaM interactions with target proteins relevant to plants.
Subject(s)
Calmodulin-Binding Proteins/metabolism , Plant Proteins/metabolism , Plants/metabolism , Calcium/metabolism , Signal TransductionABSTRACT
In plants, succinic semialdehyde dehydrogenase (SSADH)-deficiency results in the accumulation of reactive oxygen intermediates (ROI), necrotic lesions, dwarfism, and hypersensitivity to environmental stresses. We report that Arabidopsis ssadh knockout mutants contain five times the normal level of gamma-hydroxybutyrate (GHB), which in SSADH-deficient mammals accounts for phenotypic abnormalities. Moreover, the level of GHB in Arabidopsis is light dependent. Treatment with gamma-vinyl-gamma-aminobutyrate, a specific gamma-aminobutyrate (GABA)-transaminase inhibitor, prevents the accumulation of ROI and GHB in ssadh mutants, inhibits cell death, and improves growth. These results provide novel evidence for the relationship between the GABA shunt and ROI, which may, in part, explain the phenotype of SSADH-deficient plants and animals.
