title: Perforin activity at membranes leads to invaginations and vesicle formation.
abstract: The cytotoxic cell granule secretory pathway is essential for immune defence. How the pore-forming protein perforin (PFN) facilitates the cytosolic delivery of granule-associated proteases (granzymes) remains enigmatic. Here we show that PFN is able to induce invaginations and formation of complete internal vesicles in giant unilamellar vesicles. Formation of internal vesicles depends on native PFN and calcium and antibody labeling shows the localization of PFN at the invaginations. This vesiculation is recapitulated in large unilamellar vesicles and in this case PFN oligomers can be seen associated with the necks of the invaginations. Capacitance measurements show PFN is able to increase a planar lipid membrane surface area in the absence of pore formation, in agreement with the ability to induce invaginations. Finally, addition of PFN to Jurkat cells causes the formation of internal vesicles prior to pore formation. PFN is capable of triggering an endocytosis-like event in addition to pore formation, suggesting a new paradigm for its role in delivering apoptosis-inducing granzymes into target cells.
authors.affiliation: Department of Biology, Biotechnical Faculty, University of Ljubljana, Večna pot 111, 1000 Ljubljana, Slovenia.
authors.forename: Tilen
authors.initials: T
authors.lastname: Praper
authors.forename: Andreas F-P
authors.initials: AF
authors.lastname: Sonnen
authors.forename: Ales
authors.initials: A
authors.lastname: Kladnik
authors.forename: Alberto O
authors.initials: AO
authors.lastname: Andrighetti
authors.forename: Gabriella
authors.initials: G
authors.lastname: Viero
authors.forename: Keith J
authors.initials: KJ
authors.lastname: Morris
authors.forename: Emanuela
authors.initials: E
authors.lastname: Volpi
authors.forename: Lorenzo
authors.initials: L
authors.lastname: Lunelli
authors.forename: Mauro
authors.initials: M
authors.lastname: Dalla Serra
authors.forename: Christopher J
authors.initials: CJ
authors.lastname: Froelich
authors.forename: Robert J C
authors.initials: RJ
authors.lastname: Gilbert
authors.forename: Gregor
authors.initials: G
authors.lastname: Anderluh
mesh_terms: D002462:Cell Membrane / Q000378:metabolism*; D020285:Cryoelectron Microscopy; D004705:Endocytosis / Q000502:physiology*; D053804:Granzymes / Q000378:metabolism*; D006801:Humans; D007113:Immunity, Innate / Q000502:physiology*; D019169:Jurkat Cells; D008856:Microscopy, Fluorescence; D054353:Perforin / Q000276:immunology / Q000378:metabolism* / Q000502:physiology; D022142:Secretory Vesicles / Q000378:metabolism*
chemical_list: D054353:Perforin; D053804:Granzymes