The regulation of amino acid metabolism during hyperosmotic stress in Acanthamoeba castellanii

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Abstract
The effects of hyperosmotic stress on amino acid metabolism have been examined in Acanthamoeba castellanii incubated in a defined growth medium. Following hyperosmotic shock, the rapid appearance in the medium of previously ingested proline indicated a rapid release of pinosome contents induced by osmotic dehydration. Initially endocytotic uptake and protein synthesis were both markedly inhibited, but they recovered over the following hours. Total cell proteins decreased by 20% within 30 minutes after hyperosmotic stress, and this decrease was concomitant with a rapid accumulation of free ninhydrin‐positive substances.Following a hyperosmotic shock, free alanine, glutamate, and proline transiently accumulated, in this sequence. Aminooxyacetate prevented the accumulation of alanine while greatly stimulating that of glutamate, but its effect on proline accumulation was small. In cells incubated with radioactive alanine, the accumulation of free alanine proceeded mostly from unlabeled precursors and coincided with a high rate of radioactive glutamate synthesis. This newly synthesized glutamate was extensively converted into proline, while proline synthesis from ornithine was barely stimulated. During acclimation to hyperosmotic conditions, osmoregulatory mechanisms in A. castellanii appear to rely only transiently on free amino acids, particularly on those whose access to main metabolic pathways is not allosterically regulated.