Fig. 3. Effect of NH₄Cl on the biosynthesis of cathepsin B, D and L in NIH/3T3 cells and M6P/IGF2R-negative mouse embryonic fibroblasts. (A) Igf2r^-/- MEFs and receptor-positive NIH/3T3 cells were metabolically labeled with [³⁵S]methionine and chased for 5 hours in the absence (-) or continuous presence (+) of 10 mM NH₄Cl as described in Fig. 1. Cathepsin B was then immunoprecipitated from equivalent amounts of cell and medium extracts and analyzed by SDS-PAGE followed by fluorography. Note that the nonspecific band described in Fig. 1 was not observed in this experiment. proCB, procathepsin B; scCB, mature cathepsin B (single-chain form). (B) Igf2r^-/- MEFs were metabolically labeled with [³⁵S]methionine and chased for 5 hours in the absence (-) or continuous presence (+) of 10 mM NH₄Cl as described in Fig. 1. Cathepsin D and L were then sequentially immunoprecipitated from equivalent amounts of cell and medium extracts and analyzed by SDS-PAGE followed by fluorography. proCL, procathepsin L; scCL, mature cathepsin L (single-chain form); tcCL, heavy-chain of mature cathepsin L (two-chain form); proCD, procathepsin D; scCD, mature cathepsin D (single-chain form).