CHARACTERIZATION OF LYSOSOMES ISOLATED FROM DICTYOSTELIUM-DISCOIDEUM BY MAGNETIC FRACTIONATION

Superparamagnetic particles were prepared with iron oxide cores of con gruent-to 8 nm diameter and dextran coats. After feeding the probe to the amoeba, Dictyostelium discoideum, for 15 min and chasing for 15 mi n, a lysosome fraction was isolated magnetically. Isolates contained 7 6% of ingested iron, 82% of ingested fluorescent dextran, 1.3% of cell protein, 4% of the lipid, 28% of acid phosphatase, and 5% of the vacu olar H+-ATPase. Enrichment in endocytic markers was congruent-to 60-fo ld; markers for other organelles were <0.5%. The lysosomes were homoge neous, round (0.4-1.1 mum in diameter), and frequently adherent to one another through zones of intimate apposition. Cells were also fed the iron probe continuously for 3 h to fill their entire endocytic pathwa y; in this case, isolates contained 3.3% of cell protein, 11% of lipid , and 49% of cell acid phosphatase. Bis(monoacylglycerol)phosphate (BM P), a lipid characteristic of lysosomes in animal cells, comprised con gruent-to 6% of biosynthetically labelled cell lipids and up to half o f the lipid in the endocytic pathway. Essentially all of the cellular BMP was recovered in isolates prepared after 3 h of feeding. The speci ficity and abundance of BMP in the endocytic organelles of this early diverged protist suggests that this phospholipid serves a universal an d essential function in endocytosis.