title: Similarities and differences in the biogenesis, processing and lysosomal targeting between zebrafish and human pro-Cathepsin D: functional implications.
abstract: The lysosomal protease Cathepsin D (CD) plays a role in neurodegenerative diseases, cancer, and embryo-fetus abnormalities. It is therefore of interest to know how this protein is synthesized in animal species used for modeling human diseases. Zebrafish (Danio rerio) is emerging as a valuable 'in vivo' vertebrate model for several human diseases. We have characterized the biogenetic pathways of zebrafish and human CD transgenically expressed in both human SH-SY5Y cells and zebrafish PAC2 cells. Differently from human CD, zebrafish CD was synthesized as a mono-glycosylated precursor (pro-CD) that was eventually processed into a single-chain mature polypeptide. In PAC2 cells, ammonium chloride and chloroquine impaired the N-glycosylation, and greatly stimulated the secretion, of pro-CD; still, a portion of un-glycosylated pro-CD reached the lysosomes and was processed to mature CD. The treatment with tunicamycin, which abrogates N-glycosylation, resulted in a similar effect. Zebrafish pro-CD was correctly processed when expressed in human cells, and its glycosylation, transport and maturation were not impaired by ammonium chloride. On the contrary, the transport and processing of human pro-CD expressed in zebrafish cells were profoundly altered: while the intermediate single-chain was not detectable, a small amount of double-chain mature CD still formed. This fact indicates that the enzyme machinery for single- to double-chain processing of mammal CD is present in zebrafish. Our data highlight the respective impact of the information imparted by the primary sequence and of the cellular transport and processing machineries in the biogenesis of lysosomal CD.
authors.affiliation: Department of Health Sciences, Università del Piemonte Orientale Amedeo Avogadro, Via Solaroli 17, 28100, Novara, Italy. carlo.follo@med.unipmn.it
authors.forename: Carlo
authors.initials: C
authors.lastname: Follo
authors.forename: Matteo
authors.initials: M
authors.lastname: Ozzano
authors.forename: Claudia
authors.initials: C
authors.lastname: Montalenti
authors.forename: Maneerat
authors.initials: M
authors.lastname: Ekkapongpisit
authors.forename: Ciro
authors.initials: C
authors.lastname: Isidoro
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002402:Cathepsin D / Q000737:chemistry / Q000378:metabolism*; D002460:Cell Line; D004792:Enzyme Precursors / Q000737:chemistry / Q000378:metabolism*; D006031:Glycosylation; D006801:Humans; D008247:Lysosomes / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011499:Protein Processing, Post-Translational; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D017386:Sequence Homology, Amino Acid; D015027:Zebrafish; D029961:Zebrafish Proteins / Q000737:chemistry / Q000378:metabolism*
chemical_list: D004792:Enzyme Precursors; D029961:Zebrafish Proteins; C533922:CTSD protein, human; D002402:Cathepsin D