Role of Malate Synthase in Citric Acid Synthesis by Maturing Cotton Embryos: A Proposal

Cotton embryos from 34 to 54 days after anthesis were analyzed for organic acids, and enzymes associated with organic acid metabolism. During this developmental period, embryos accumulated citrate. Malate synthase activity appeared at 46 days after anthesis and increased rapidly to 54 days. Of other enzymes examined, only citrate synthase activity increased during this period. As isocitrate lyase activity was absent from cotton embryos during maturation, an alternative source of glyoxylate would be required for in vivo malate synthase activity. Of several metabolic sources tested, glycine was converted to glyoxylate via a transamination reaction.Halves of 50-day (mature) cotton embryos incorporated radioactivity from [1-(14)C]acetate, [1-(14)C]glyoxylate, and [1-(14)C]glycine into organic acids. Embryo halves incubated with [(14)C]glyoxylate plus [(3)H]acetate synthesized double-labeled malate and citrate. Radioactive citrate isolated from 50-day cotton embryos incubated with [1-(14)C]acetate was degraded; label was distributed as follows: 55% in C(1), 33% in C(5), and 12% in C(6). Taken together, these data strongly suggest participation of malate synthase in citrate production in vivo.Separation of organelles by sucrose density gradient sedimentation revealed that malate synthase, malate dehydrogenase, and citrate synthase were compartmentalized together only in the peroxisome fraction (1.24 grams per milliliter). Peroxisomes isolated from 50-day embryos, when incubated with glyoxylate and [(3)H]acetyl-CoA, synthesized labeled malate and citrate, but only radioactive citrate accumulated. Incubations with glycine plus alpha-ketoglutarate, in place of glyoxylate, also resulted in synthesis of radioactive citrate.A metabolic scheme illustrating the participation of cotton embryo peroxisomes in citrate synthesis is proposed. This scheme suggests a function for plant peroxisomes not previously elucidated. The ontogenetic and metabolic relationship between these organelles and glyoxysomes active in gluconeogenesis during postgerminative growth remains to be examined.