Processing, transport, and secretion of the lysosomal enzyme acid phosphatase in Dictyostelium discoideum

• Published in: The Journal of biological chemistry 1989-05, Vol.264 (13), p.7630-7636
• Main Authors: Bush, J M, Cardelli, J A
• Format: Article
• Language: English
• Subjects: ACID PHOSPHATASE; Acid Phosphatase - metabolism; Amines - pharmacology; Analytical, structural and metabolic biochemistry; Biological and medical sciences; Biological Transport - drug effects; Cell Compartmentation - drug effects; CHAMPIGNON; Dictyostelium - enzymology; Dictyostelium discoideum; Endoplasmic Reticulum - enzymology; Enzymes and enzyme inhibitors; Fluorescent Antibody Technique; FOSFATASA ACIDA; Fundamental and applied biological sciences. Psychology; FUNGI; Glycoside Hydrolases - metabolism; Glycosylation; Golgi Apparatus - enzymology; Hydrogen-Ion Concentration; Hydrolases; IMMUNOFLUORESCENCE; INMUNOFLUORESCENCIA; Lysosomes - enzymology; Molecular Weight; PHOSPHATASE ACIDE; Protease Inhibitors - pharmacology; Protein Precursors - metabolism; Protein Processing, Post-Translational; SECRECION; SECRETION
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/S0021-9258(18)83281-1

To explain the different secretion kinetics of lysosomal enzymes in Dictyostelium discoideum, previous investigators have hypothesized the existence of a heterogeneous population of lysosomes containing either the enzyme acid phosphatase or other hydrolase enzymes. This proposal predicts that at least two targeting mechanisms exist for lysosomal enzymes in this organism. To begin to investigate this possibility, the transport, processing, and targeting of acid phosphatase was studied by using a combination of radiolabel pulse-chase procedures, subcellular fractionations, and indirect immunofluorescence microscopy. Acid phosphatase was initially synthesized in axenically growing cells as a 56-kDa precursor polypeptide that was proteolytically processed after 20 min to a 55-kDa mature protein. This enzyme was rapidly transported from the endoplasmic reticulum to Golgi complex (halftime of 3 min) as measured by the acquisition of resistance to the enzyme endoglycosidase H. Furthermore, Percoll gradient fractionations indicated that radiolabeled forms of acid phosphatase reached dense lysosomal vesicles at about the same time as final processing was occurring. Proper sorting of acid phosphatase in D. discoideum apparently was not critically dependent on low intravacuolar pH since the addition of ammonium chloride did not stimulate the missorting and secretion of acid phosphatase. These results are very similar to previous observations concerning other Dictyostelium lysosomal enzymes. Consistent with the existence of a heterogeneus population of lysosomes, the percentage of radiolabeled acid phosphatase secreted 4 h into a chase period was 15-fold lower as compared with another lysosomal enzyme, β-glucosidase. However, acid phosphatase, α-mannosidase, and β-glucosidase were all predominantly colocalized as determined by indirect immunofluorescence, which for the first time demonstrates the homogeneous nature of the lysosomal system in D. discoideum. Taken together these results suggest that the processing and transport of acid phosphatase may be similar in nature to the glycosidases. However, the different kinetics of secretion of acid phosphatase versus the colocalized glycosidase enzymes suggests that an undefined mechanism operates to distinguish these classes of enzymes at a step after localization to lysosomes but prior to secretion.