DIFFERENTIAL TARGETING AND PROCESSING OF PROCATHEPSIN-D IN NORMAL ANDTRANSFORMED MURINE 3T3 FIBROBLASTS

The kinetics of transport and the processing of procathepsin D (proCD) , the precursor of a lysosomal aspartyl protease involved in tumor-cel l proliferation and metastasis, were compared in normal and SV-40- or benzo[a]pyrene-transformed 3T3 mouse fibroblasts. Sorting of newly syn thesized proCD in normal cells was almost complete within 3 hr, while in transformed cells a fraction of the precursor survives a long time. In both normal and transformed 3T3 cultures, secretion of proCD start ed at 3 hr of chase. However, in normal cells secretion of proCD remai ned constant between 3 and 24 hr of chase, while in transformed cells it increased along with the chase incubation. The efficiency of format ion of the mannose-6-phosphate group on proCD varied among the 3 cell types, being minimal in benzo[a]pyrene-transformed 3T3 cells. Ammonium chloride, a drug known to disrupt the segregation and to enhance the secretion of lysosomal proenzymes, was 2-fold more effective in normal than in transformed 3T3 cells. Despite vacuolar alkalinization, about one third of proCD was segregated into the endosomal-lysosomal pathwa y in normal and in transformed 3T3 fibroblasts, indicating the existen ce in these cells of alternative, mannose-6-phosphate receptor-indepen dent mechanisms for targeting proCD. Thus, while hypersecretion of pro CD and reduced sensitivity to vacuolar alkalinization are common featu res of both transformed cell types, the mechanisms responsible for ine fficient segregation of proCD may differ between virally and chemicall y transformed 3T3 cells. (C) 1997 Wiley-Liss, Inc.