Surface Labeling of Membrane Glycoproteins and Their Drastic Changes during Development of Dictyostelium discoideum

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Abstract
Surface glycoproteins on plasma membranes from D. discoideum were labeled by sodium metaperiodate oxidation and sodium boro[3H]hydride reduction. The amount of incorporation of tritium from NaB3H4 reached a plateau after 10 min at periodate concentration of 20 mM. Density analysis carried out by sucrose density-gradient centrifugation showed that plasma membranes were selectively labeled. About 84% of the radioactivity incorporated was released by hydrolysis with 0.25 M H2SO4 for 3 h at 100.degree. C. Released materials were eluted at a bed volume after Sephadex G-50 chromatography. Paper chromatography of the eluate detected 4 radioactive spots; 2 were glycerol and glyceraldehyde and the other 2 spots seemed to be oligosaccharides. Plasma membranes from aggregation-phase cells were labeled 4 times more than those from growth-phase cells. The labeled plasma membrane fraction during the aggregation phase was separated into 39 distinct glycoprotein bands on a 1-dimensional dodecylsulfate polyacrylamide gel. Of the 39 bands 6 increased in density and 2 new bands appeared. Four bands decreased in density in the aggregation phase. Surface glycoproteins were analyzed directly by gel electrophoresis of the lysate of labeled whole cells without preparing plasma membranes. Changes during development of glycoproteins on outer cell surfaces were confirmed by O''Farrell''s method of 2-dimensional polyacrylamide gel electrophoresis. Glycoproteins on plasma membranes of D. discoideum were separated into 63 individual spots; 45 spots changed during the early developmental course of D. discoideum, in contrast to the slight change in soluble and membrane proteins during this phase. Glycoproteins may have important roles in the process of aggregation.

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