title: Expression of human cathepsin D in Xenopus oocytes: phosphorylation and intracellular targeting.
abstract: We have obtained expression of a cDNA clone for human cathepsin D in Xenopus laevis oocytes. Biosynthetic studies with [35S]methionine labeling demonstrated that most of the cathepsin D remained intracellular and underwent proteolytic cleavage, converting a precursor of Mr 47,000 D to a mature form of Mr 39,000 D with processing intermediates of Mr 43,000-41,000 D. greater than 90% of the cathepsin D synthesized by oocytes bound to a mannose 6-phosphate (Man-6-P) receptor affinity column, indicating the presence of phosphomannosyl residues. An analysis of [2-3H]mannose-labeled oligosaccharides directly demonstrated phosphomannosyl residues on cathepsin D. Sucrose-gradient fractionation, performed to define the membranous compartments that cathepsin D traversed during its biosynthesis, demonstrated that cathepsin D is targeted to a subpopulation of yolk platelets, the oocyte equivalent of a lysosome. Xenopus oocytes were able to endocytose lysosomal enzymes from the medium and this uptake was inhibited by Man-6-P, thus demonstrating the presence of Man-6-P receptors in these cells. Therefore, the entire Man-6-P dependent pathway for targeting of lysosomal enzymes is present in the oocytes. Xenopus oocytes should be a useful system for examining signals responsible for the specific targeting of lysosomal enzymes to lysosomes.
authors: Department of Medicine, Washington University School of Medicine, St. Louis, Missouri 63110.
authors: P L
authors: PL
authors: Faust
authors: D A
authors: DA
authors: Wall
authors: E
authors: E
authors: Perara
authors: V R
authors: VR
authors: Lingappa
authors: S
authors: S
authors: Kornfeld
mesh_terms: D000818:Animals; D002402:Cathepsin D / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification; D003001:Cloning, Molecular; D004247:DNA / Q000378:metabolism; D005260:Female; D006801:Humans; D007700:Kinetics; D008970:Molecular Weight; D009865:Oocytes / Q000378:metabolism*; D010766:Phosphorylation; D010957:Plasmids; D011499:Protein Processing, Post-Translational; D014982:Xenopus laevis
chemical_list: D004247:DNA; D002402:Cathepsin D