title: Granule-mediated killing by granzyme B and perforin requires a mannose 6-phosphate receptor and is augmented by cell surface heparan sulfate.
abstract: During granule-mediated killing by cytotoxic T lymphocytes or natural killer cells, the serine protease granzyme B enters the target cell by endocytosis and induces apoptosis. Previous studies suggested a role for the mannose 6-phosphate receptor, but further experiments with purified granzyme B indicated this was not essential. Additionally, it is now clear that grB is exocytosed from killer cells in a high-molecular-weight complex with the proteoglycan serglycin. Here granzyme B was delivered as a purified monomer, or in complex with either glycosaminoglycans or serglycin, and killing was evaluated. When granzyme B was a monomer, soluble mannose 6-phosphate had a limited impact, whereas apoptosis induced by the complexed grB was effectively inhibited by mannose 6-phosphate. Most importantly, when granzyme B and perforin were delivered together from granules, inhibition by mannose 6-phosphate was also observed. In pulldown assays mediated by the cation-independent mannose 6-phosphate receptor, granzyme B bound to the receptor more intensely in the presence of immobilized heparan sulfate. We therefore propose the model that under physiological conditions serglycin-bound granzyme B is critically endocytosed by a mannose 6-phosphate receptor, and receptor binding is enhanced by cell surface heparan sulfate.
authors.affiliation: Department of Biochemistry, University of Alberta, Edmonton, Alberta, Canada T6G 2H7.
authors.forename: Kirstin
authors.initials: K
authors.lastname: Veugelers
authors.forename: Bruce
authors.initials: B
authors.lastname: Motyka
authors.forename: Ing Swie
authors.initials: IS
authors.lastname: Goping
authors.forename: Irene
authors.initials: I
authors.lastname: Shostak
authors.forename: Tracy
authors.initials: T
authors.lastname: Sawchuk
authors.forename: R Chris
authors.initials: RC
authors.lastname: Bleackley
mesh_terms: D000818:Animals; D017209:Apoptosis; D002460:Cell Line; D006025:Glycosaminoglycans / Q000378:metabolism; D053804:Granzymes; D006497:Heparan Sulfate / Q000737:chemistry / Q000502:physiology*; D006801:Humans; D019169:Jurkat Cells; D008562:Membrane Glycoproteins / Q000502:physiology*; D008565:Membrane Proteins / Q000502:physiology*; D051379:Mice; D008954:Models, Biological; D054353:Perforin; D052899:Pore Forming Cytotoxic Proteins; D011509:Proteoglycans / Q000502:physiology; D017527:Receptor, IGF Type 2 / Q000502:physiology*; D022142:Secretory Vesicles / Q000201:enzymology / Q000502:physiology*; D012697:Serine Endopeptidases / Q000502:physiology*; D013602:T-Lymphocytes, Cytotoxic / Q000201:enzymology / Q000276:immunology*; D033921:Vesicular Transport Proteins / Q000502:physiology
chemical_list: D006025:Glycosaminoglycans; D053804:Granzymes; D006497:Heparan Sulfate; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; D054353:Perforin; D052899:Pore Forming Cytotoxic Proteins; D011509:Proteoglycans; D017527:Receptor, IGF Type 2; D012697:Serine Endopeptidases; D033921:Vesicular Transport Proteins; C075430:serglycin; C509213:GZMB protein, human; C509214:Gzmb protein, mouse