Membranous intermediates in endocytosis are labile, as shown in a temperature-sensitive mutant

Membranous tubules, especially prevalent in mammalian absorptive epithelia and insect oocytes, are one of the pleomorphic endocytic compartments that have a role in receptor-mediated endocytosis. To determine whether these tubules are evanescent, and to investigate their temporal relationships with other endocytic intermediates, we studied these tubules during oocyte vitellogenesis in the temperature-sensitive mutant Drosophila melanogaster, shibiretsl. Raising the temperature of shibire oocytes for 1 min from 19 degrees C to 29 degrees C caused a loss of these membranous tubules. The percentage of membrane in tubules decreased from 36% at 19 degrees C to 1.5% after 5 min at 29 degrees C. Concomitantly, the amount of surface membrane increased from 64% at 19 degrees C to 98% after 5 min at 29 degrees C, causing surface membrane invaginations to extend deeper into the cortex. At 29 degrees C the cytoplasmic face of the plasma membrane was studded with coated pits, and the extracellular face was coated with electron-dense material. Return from 29 degrees C to either 19 degrees C or 26 degrees C for 1-2 min produced a rapid reappearance of tubules containing extracellular horseradish peroxidase in the cortex. These data suggest that tubular intermediates are evanescent structures, and that temperature shock (i) rapidly blocks their formation from the plasma membrane, (ii) causes existing tubules to rapidly recycle to the plasma membrane, and (iii) is rapidly reversed, as newly formed tubules derive their membrane and content from the cell surface.