title: C-terminal truncation impairs glycosylation of transmembrane collagen XVII and leads to intracellular accumulation.
abstract: Collagen XVII, a type II transmembrane protein in hemidesmosomes, is involved in the anchorage of stratified epithelia to the underlying mesenchyme. Its functions are regulated by ectodomain shedding, and its genetic defects lead to epidermal detachment in junctional epidermolysis bullosa (JEB), a heritable skin fragility syndrome, but the molecular disease mechanisms remain elusive. Here we used a spontaneously occurring homozygous COL17A1 deletion mutant in JEB to discern glycosylation of collagen XVII. The mutation truncated the distal ectodomain and positioned the only N-glycosylation site 34 amino acids from the newly formed C terminus, which impaired efficient N-glycosylation. Immunofluorescence staining of authentic JEB keratinocytes and of COS-7 cells transfected with the mutant indicated intracellular accumulation of collagen XVII precursor molecules. Cell surface biotinylation and quantification of ectodomain shedding demonstrated that only about 15% of the truncated collagen XVII reached the cell surface. The cell surface-associated molecules were N-glycosylated in a normal manner, in contrast to the molecules retained within the cells, indicating that N-glycosylation of the ectodomain is required for targeting of collagen XVII to the plasma membrane and that reduced accessibility of the N-glycosylation site negatively regulates this process. Functional consequences of the strong reduction of collagen XVII on the cell surface included scattered deposition of cell adhesion molecule laminin 5 into the extracellular environment and, as a consequence of faulty collagen XVII-laminin ligand interactions, aberrant motility of the mutant cells.
authors.affiliation: Department of Dermatology, University Medical Center Freiburg, 79104 Freiburg, Germany.
authors.forename: Claus-Werner
authors.initials: CW
authors.lastname: Franzke
authors.forename: Cristina
authors.initials: C
authors.lastname: Has
authors.forename: Carsten
authors.initials: C
authors.lastname: Schulte
authors.forename: Laura
authors.initials: L
authors.lastname: Huilaja
authors.forename: Kaisa
authors.initials: K
authors.lastname: Tasanen
authors.forename: Monique
authors.initials: M
authors.lastname: Aumailley
authors.forename: Leena
authors.initials: L
authors.lastname: Bruckner-Tuderman
mesh_terms: D000595:Amino Acid Sequence; D001324:Autoantigens / Q000235:genetics* / Q000378:metabolism* / Q000502:physiology; D001485:Basement Membrane / Q000378:metabolism; D002478:Cells, Cultured; D006031:Glycosylation; D006801:Humans; D007223:Infant; D007231:Infant, Newborn; D007424:Intracellular Fluid / Q000378:metabolism*; D015603:Keratinocytes / Q000378:metabolism; D008969:Molecular Sequence Data; D024041:Non-Fibrillar Collagens / Q000235:genetics* / Q000378:metabolism* / Q000502:physiology; D010446:Peptide Fragments / Q000235:genetics* / Q000378:metabolism* / Q000502:physiology; D017434:Protein Structure, Tertiary / Q000235:genetics; D017384:Sequence Deletion / Q000235:genetics*; D000097805:Collagen Type XVII
chemical_list: D001324:Autoantigens; D024041:Non-Fibrillar Collagens; D010446:Peptide Fragments; D000097805:Collagen Type XVII