title: Perforin forms transient pores on the target cell plasma membrane to facilitate rapid access of granzymes during killer cell attack.
abstract: Cytotoxic lymphocytes serve a key role in immune homeostasis by eliminating virus-infected and transformed target cells through the perforin-dependent delivery of proapoptotic granzymes. However, the mechanism of granzyme entry into cells remains unresolved. Using biochemical approaches combined with time-lapse microscopy of human primary cytotoxic lymphocytes engaging their respective targets, we defined the time course of perforin pore formation in the context of the physiological immune synapse. We show that, on recognition of targets, calcium influx into the lymphocyte led to perforin exocytosis and target cell permeabilization in as little as 30 seconds. Within the synaptic cleft, target cell permeabilization by perforin resulted in the rapid diffusion of extracellular milieu-derived granzymes. Repair of these pores was initiated within 20 seconds and was completed within 80 seconds, thus limiting granzyme diffusion. Remarkably, even such a short time frame was sufficient for the delivery of lethal amounts of granzymes into the target cell. Rapid initiation of apoptosis was evident from caspase-dependent target cell rounding within 2 minutes of perforin permeabilization. This study defines the final sequence of events controlling cytotoxic lymphocyte immune defense, in which perforin pores assemble on the target cell plasma membrane, ensuring efficient delivery of lethal granzymes.
authors: Cancer Immunology Program, Peter MacCallum Cancer Centre, East Melbourne, VIC, Australia.
authors: Jamie A
authors: JA
authors: Lopez
authors: Olivia
authors: O
authors: Susanto
authors: Misty R
authors: MR
authors: Jenkins
authors: Natalya
authors: N
authors: Lukoyanova
authors: Vivien R
authors: VR
authors: Sutton
authors: Ruby H P
authors: RH
authors: Law
authors: Angus
authors: A
authors: Johnston
authors: Catherina H
authors: CH
authors: Bird
authors: Phillip I
authors: PI
authors: Bird
authors: James C
authors: JC
authors: Whisstock
authors: Joseph A
authors: JA
authors: Trapani
authors: Helen R
authors: HR
authors: Saibil
authors: Ilia
authors: I
authors: Voskoboinik
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000276:immunology*; D002462:Cell Membrane / Q000276:immunology* / Q000378:metabolism; D015938:Complement Membrane Attack Complex / Q000276:immunology / Q000378:metabolism; D004705:Endocytosis / Q000276:immunology; D005089:Exocytosis / Q000276:immunology; D053804:Granzymes / Q000276:immunology* / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D019169:Jurkat Cells; D007694:Killer Cells, Natural / Q000166:cytology / Q000276:immunology* / Q000378:metabolism; D051379:Mice; D054353:Perforin; D052899:Pore Forming Cytotoxic Proteins / Q000276:immunology* / Q000378:metabolism; D013602:T-Lymphocytes, Cytotoxic / Q000166:cytology / Q000276:immunology* / Q000378:metabolism; D013997:Time Factors
chemical_list: D015938:Complement Membrane Attack Complex; D052899:Pore Forming Cytotoxic Proteins; C510467:perforin 1, mouse; D054353:Perforin; D053804:Granzymes