Fig. 4. Secretion of Hsp150p to the cell surface was unaffected by OSH temperature-sensitive mutants. Autoradiograph of Hsp150 purified using Con A-affinity chromatography from the extracellular medium and cell extracts of wild-type (Osh⁺; SEY6210), osh osh4-1 (CBY926), osh OSH4 (CBY924), wild-type (W.T.; RSY255) and sec1-1 (RSY782) strains. Cultures were incubated at 37°C for 60 minutes and, after a 10 minute pulse with [³⁵S]-label followed by the addition of the chase solution, samples were removed at 0, 10, 30 and 60 minutes. In all samples from OSH mutants, Hsp150p was secreted into the medium with equivalent kinetics to its wild-type control. Moreover, 60 minutes after pulse labeling and addition of chase solution no intracellular Hsp150p was detectable. In the exocytosis-defective sec1-1 strain scarcely any Hsp150p was secreted into the medium during the 60 minutes following pulse labeling and addition of chase solution, and Hsp150p was detected within cells (internalized Hsp150p appeared to degrade 60 minutes after the addition of chase solution but was nonetheless detectable). The autoradiograph of intracellular glycoproteins represented a longer exposure than that showing secreted extracellular glycoproteins.