Transit of alpha-mannosidase during its maturation in Dictyostelium discoideum
| Autor: | Arnold Kaplan, Ladonna Wood |
|---|---|
| Rok vydání: | 1985 |
| Předmět: |
Mannosidase
Exocytosis Dictyostelium discoideum chemistry.chemical_compound alpha-Mannosidase Mannosidases Organelle Extracellular Dictyostelium Secretion Monensin Sodium dodecyl sulfate Polyacrylamide gel electrophoresis Enzyme Precursors Methionine biology Biological Transport Articles Cell Biology biology.organism_classification Cell Compartmentation Kinetics chemistry Biochemistry Starvation Lysosomes Protein Processing Post-Translational Subcellular Fractions |
| Zdroj: | The Journal of Cell Biology |
| ISSN: | 1540-8140 0021-9525 |
| Popis: | We proposed that Dictyostelium discoideum contains two linked pools of mature alpha-mannosidase (Wood, L., R. N. Pannell, and A. Kaplan, 1983, J. Biol. Chem., 258:9426-9430). To obtain physical evidence for these pools, cells were pulse-labeled with [35S]methionine, homogenized, and subjected to Percoll gradient centrifugation. After immune precipitation of alpha-mannosidase, its polypeptides were separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis and detected by fluorography. After a 30-min pulse with [35S]methionine, the precursor and small amounts of cleaved enzyme were detected in a low density fraction (1.04 g/ml). Subsequently, cleaved enzyme was transferred to higher density fractions (1.05 and 1.07 g/ml) that were enriched in lysosomal enzymes. The half time for formation of the 1.07 g/ml pool was approximately 45 min, whereas formation of the 1.05 g/ml pool was not detected until 1.5 h after the pulse. The transfer of mature forms out of the 1.04 g/ml pool was inhibited by monensin (3.5 microM). Thus, alpha-mannosidase precursor appears to be cleaved in a prelysosomal organelle. The data also indicate that starving cells secrete precursor directly from this organelle to the extracellular space, whereas cleaved forms are first transferred into lysosomes before they are secreted. Furthermore, 2 h after starvation, the secretion of mature forms ceases even though both transit of mature forms between the two pools and secretion of precursor continues. From this we inferred that the cessation of secretion of mature forms is due to a halt in fusion of lysosomes with the plasma membrane and that precursor follows a different route to the plasma membrane. |
| Databáze: | OpenAIRE |
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