Figure 2.

Growth and precursor processing defect of SSC1-I485T is partly alleviated by D107A. (A) pRS314-SSC1 plasmids carrying either a wt or mutant copy of SSC1 (as indicated) were transformed into DG252 cells; transformants were streaked on plates containing 5-fluoroorotic acid to select for loss of the URA3-marked plasmid containing wt SSC1. 5-Fluoroorotic acid-resistant cells were grown in YPD overnight, 1:10 serial dilutions were made, and cells were plated on YPD and incubated at the indicated temperatures. (B and C) In vivo pulse–chase analysis of mitochondrial precursor processing. Strains containing wt or mutant copies of SSC1 were pulse labeled with [³⁵S]methionine and cysteine at 37°C for 2 min, at which time cold methionine and cysteine were added, and incubation continued for the indicated time in minutes before harvest and immunoprecipitation with antibodies to Hsp60 (B) or F₁β (C). Samples were analyzed by SDS/PAGE and visualized by using a PhosphorImager.