Fig. 7. Loss of Vps20p or Vta1p causes secretion of Golgi-modified p2 CPY precursor and degradation of Vps10p. Wild-type (SF838-9D), vps20 (SF838-9Dvpl10) and vta1 (PLY3046) cells were converted to PEP4 as described in Materials and Methods. They were pulse-labelled with [³⁵S]methionine/cysteine for 10 minutes at 30°C and then chased with excess unlabelled methionine/cysteine at the same temperature. Samples were taken at 0 or 60 minutes of chase, and further membrane transport was stopped by addition of sodium azide and sodium fluoride to 20 mM. The cells in each sample were converted to spheroplasts and fractionated into intracellular (I) and extracellular (E) fractions. CPY was immunoprecipitated from half the intracellular (I) and extracellular (E) fractions of the 60-minute chase samples (A). Vps10p was immunoprecipitated from the intracellular (I) fractions of the 0- and 60-minute chase samples (B). Indicated are the mature (mCPY) and Golgi-modified (p2CPY) forms of CPY, and both full-length Vps10p (Vps10p) and the lower protease-cleaved band of Vps10p (*).