Targeted quantitative profiling of metabolites and gene transcripts associated with 4-aminobutyrate (GABA) in apple fruit stored under multiple abiotic stresses.

4-Aminobutyrate accumulates in plants under abiotic stress. Here, targeted quantitative profiling of metabolites and transcripts was conducted to monitor glutamate- and polyamine-derived 4-aminobutyrate production and its subsequent catabolism to succinate or 4-hydroxybutyrate in apple (Malus x domestica Borkh.) fruit stored at 0 °C with 2.5 kPa O2 and 0.03 or 5 kPa CO2 for 16 weeks. Low-temperature-induced protein hydrolysis appeared to be responsible for the enhanced availability of amino acids during early storage, and the resulting higher glutamate level stimulated 4-aminobutyrate levels more than polyamines. Elevated CO2 increased the levels of polyamines, as well as succinate and 4-hydroxybutyrate, during early storage, and 4-aminobutyrate and 4-hydroxybutyrate over the longer term. Expression of all of the genes likely involved in 4-aminobutyrate metabolism from glutamate/polyamines to succinate/4-hydroxybutyrate was induced in a co-ordinated manner. CO2-regulated expression of apple GLUTAMATE DECARBOXYLASE 2, AMINE OXIDASE 1, ALDEHYDE DEHYDROGENASE 10A8 and POLYAMINE OXIDASE 2 was evident with longer term storage. Evidence suggested that respiratory activities were restricted by the elevated CO2/O2 environment, and that decreasing NAD+ availability and increasing NADPH and NADPH/NADP+, respectively, played key roles in the regulation of succinate and 4-hydroxybutyate accumulation. Together, these findings suggest that both transcriptional and biochemical mechanisms are associated with 4-aminobutyrate and 4-hydroxybutyrate metabolism in apple fruit stored under multiple abiotic stresses.

Plant Glyoxylate/Succinic Semialdehyde Reductases: Comparative Biochemical Properties, Function during Chilling Stress, and Subcellular Localization.

Plant NADPH-dependent glyoxylate/succinic semialdehyde reductases 1 and 2 (cytosolic GLYR1 and plastidial/mitochondrial GLYR2) are considered to be of particular importance under abiotic stress conditions. Here, the apple (Malus × domestica Borkh.) and rice (Oryza sativa L.) GLYR1s and GLYR2s were characterized and their kinetic properties were compared to those of previously characterized GLYRs from Arabidopsis thaliana [L.] Heynh. The purified recombinant GLYRs had an affinity for glyoxylate and succinic semialdehyde, respectively, in the low micromolar and millimolar ranges, and were inhibited by NADP+. Comparison of the GLYR activity in cell-free extracts from wild-type Arabidopsis and a glyr1 knockout mutant revealed that approximately 85 and 15% of the cellular GLYR activity is cytosolic and plastidial/mitochondrial, respectively. Recovery of GLYR activity in purified mitochondria from the Arabidopsis glyr1 mutant, free from cytosolic GLYR1 or plastidial GLYR2 contamination, provided additional support for the targeting of GLYR2 to mitochondria, as well as plastids. The growth of plantlets or roots of various Arabidopsis lines with altered GLYR activity responded differentially to succinic semialdehyde or glyoxylate under chilling conditions. Taken together, these findings highlight the potential regulation of highly conserved plant GLYRs by NADPH/NADP+ ratios in planta, and their roles in the reduction of toxic aldehydes in plants subjected to chilling stress.