Figure 11.
Examination of a-factor biogenesis after α-factor induction. Strain SM1227 (MFA1 mfa2::URA3) was preincubated with 25 μM α-factor (except for the 0-min sample). At the indicated times, which represent minutes after the addition of pheromone, cells were pulse labeled with [35S]cysteine for 5 min, and the label was chased for 45 min. (For the 0-min control sample, the pulse chase was carried out in the absence of any α-factor.) The intracellular (I) fraction was prepared from a portion of the 5-min pulse-labeled cells to examine the total amount of a-factor synthesized, and the extracellular (E) fraction was prepared from an equivalent portion of the culture that had completed the 45-min chase to examine the portion of the a-factor synthesized in the pulse that ultimately underwent maturation and export. Fractions were immunoprecipitated with a-factor antiserum and subjected to SDS-PAGE analysis.