Cytotoxic T Cells Use Mechanical Force to Potentiate Target Cell Killing

Basu, Roshni; Whitlock, Benjamin M.; Husson, Julien; Floc’h, Audrey Le; Jin, Weiyang; Oyler-Yaniv, Alon; Dotiwala, Farokh; Giannone, Grégory; Hivroz, Claire; Biais, Nicolas; Lieberman, Judy; Kam, Lance C.; Huse, Morgan

doi:10.1016/j.cell.2016.01.021

Cited by 333 publications

(374 citation statements)

References 56 publications

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“…The microindenter's bending stiffness was evaluated against standard microindenters that had been previously calibrated. One method to calibrate these standard microindenters was to measure their deflection under the gravitational force exerted on their tip by a piece of ultrathin paper of known mass (25). We confirmed this initial method by using a commercial force probe (model 406A, with a force range of 0-500 nN, Aurora Scientific, Aurora, Ontario, Canada).…”

Section: Microindenter Fabrication and Calibrationmentioning

confidence: 75%

Mechanical Criterion for the Rupture of a Cell Membrane under Compression

Gonzalez‐Rodriguez

Guillou

Cornat

et al. 2016

Biophysical Journal

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We investigate the mechanical conditions leading to the rupture of the plasma membrane of an endothelial cell subjected to a local, compressive force. Membrane rupture is induced by tilted microindentation, a technique used to perform mechanical measurements on adherent cells. In this technique, the applied force can be deduced from the measured horizontal displacement of a microindenter's tip, as imaged with an inverted microscope and without the need for optical sensors to measure the microindenter's deflection. We show that plasma membrane rupture of endothelial cells occurs at a well-defined value of the applied compressive stress. As a point of reference, we use numerical simulations to estimate the magnitude of the compressive stresses exerted on endothelial cells during the deployment of a stent.

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Section: Microindenter Fabrication and Calibrationmentioning

confidence: 75%

Mechanical Criterion for the Rupture of a Cell Membrane under Compression

Gonzalez‐Rodriguez

Guillou

Cornat

et al. 2016

Biophysical Journal

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“…This paradoxical extension of αβ TCR-pMHC-bond lifetime under force, so-called catch bond behavior, was then observed experimentally (19,33). The importance of force on TCR triggering has been confirmed in other work (34)(35)(36)(37)(38)(39)(40). However, during the initial T-cell surface contact, external (scanning) and internal (retrograde flow) force appear to operate in opposing directions (41).…”

mentioning

confidence: 68%

Mechanosensing drives acuity of αβ T-cell recognition

Feng

Brazin

Kobayashi

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

151

197

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T lymphocytes use surface αβ T-cell receptors (TCRs) to recognize peptides bound to MHC molecules (pMHCs) on antigen-presenting cells (APCs). How the exquisite specificity of high-avidity T cells is achieved is unknown but essential, given the paucity of foreign pMHC ligands relative to the ubiquitous self-pMHC array on an APC. Using optical traps, we determine physicochemical triggering thresholds based on load and force direction. Strikingly, chemical thresholds in the absence of external load require orders of magnitude higher pMHC numbers than observed physiologically. In contrast, force applied in the shear direction (∼10 pN per TCR molecule) triggers T-cell Ca 2+ flux with as few as two pMHC molecules at the interacting surface interface with rapid positional relaxation associated with similarly directed motor-dependent transport via ∼8-nm steps, behaviors inconsistent with serial engagement during initial TCR triggering. These synergistic directional forces generated during cell motility are essential for adaptive T-cell immunity against infectious pathogens and cancers.mechanosensor | T-cell receptor | T-cell activation | optical tweezers | cellular force relaxation T he T-cell receptor (TCR) expressed on T lymphocytes of the adaptive immune system is a stout and squat (12-nm wide × 8-nm tall) multisubunit surface complex with a ligand binding moiety that is an αβ disulfide-linked heterodimer buttressed by the associated invariant CD3 subunits (1-3). The αβ chains are each encoded by V and J gene segments and in the case of the β, a D segment as well (4). The clone-specific TCR unique to each T lymphocyte endows mammals with the capacity to detect perturbations in host cellular function resulting from myriad infectious pathogens, physical damage (thermal, irradiation, etc.), or premalignant or malignant cellular transformations while averting strong self-reactivities that could induce autoimmunity (5).Signaling is initiated through ligation of the clonotype by its cognate antigenic peptide bound to an MHC molecule (pMHC) and displayed on the surface of antigen-presenting cells (APCs) (6, 7). Ligation impacts disposition and function of the associated CD3 dimers (CD3 γ, CD3 δ, and CD3ζζ) as well as the transmembrane domains of the TCR heterodimer and CD3 subunits that interdigitate in the membrane to signal into the cytoplasm (8, 9). A cascade of intracellular events involving phosphorylation of immunoreceptor tyrosine-based activation motifs (ITAMs) on the CD3 cytoplasmic domains with subsequent ZAP70 activation and downstream signaling follows (10, 11). Membrane-associated CD8 and CD4 coreceptors, marking cytotoxic T lymphocytes (CTLs) and helper T cells, respectively, function to bring the membrane-anchored Src family tyrosine kinase Lck to the TCR-pMHC complex for the initiation of ITAM phosphorylation. Signaling, in turn, leads to a transient rise of cytosolic Ca 2+ and other biochemical events resulting in transcriptional activation, ultimately resulting in developmental decisions or effector functi...

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“…Although mutant YT-KR/AA GB is not efficiently delivered to target cells by perforin in loading experiment, it is efficiently delivered to target cells during killer cell attack. It was recently shown that effector cells exert synaptic force to enhance perforin activity, 57 explaining the discrepancy between the loading experiment and the physiological effector/target interaction. However, GB KR/AA, which cannot enter the mitochondria is significantly impaired in its ability to trigger ROS and cell death both in vitro and in vivo, indicating that GB mitochondrial entry is necessary for efficient cell death induction in agreement with Jacquemin et al 18 Tob55/Sam50 belongs to the bacterial β-barrel-shaped channels of the Omp85 family.…”

Section: Discussionmentioning

confidence: 99%

Granzyme B enters the mitochondria in a Sam50-, Tim22- and mtHsp70-dependent manner to induce apoptosis

et al. 2017

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We have found that granzyme B (GB)-induced apoptosis also requires reactive oxygen species resulting from the alteration of mitochondrial complex I. How GB, which does not possess a mitochondrial targeting sequence, enter this organelle is unknown. We show that GB enters the mitochondria independently of the translocase of the outer mitochondrial membrane complex, but requires instead Sam50, the central subunit of the sorting and assembly machinery that integrates outer membrane β-barrel proteins. Moreover, GB breaches the inner membrane through Tim22, the metabolite carrier translocase pore, in a mitochondrial heat-shock protein 70 (mtHsp70)-dependent manner. Granzyme A (GA) and caspase-3 use a similar route to the mitochondria. Finally, preventing GB from entering the mitochondria either by mutating lysine 243 and arginine 244 or depleting Sam50 renders cells more resistant to GB-mediated reactive oxygen species and cell death. Similarly, Sam50 depletion protects cells from GA-, GM-and caspase-3-mediated cell death. Therefore, cytotoxic molecules enter the mitochondria to induce efficiently cell death through a noncanonical Sam50-, Tim22-and mtHsp70-dependent import pathway.Cytotoxic lymphocytes eradicate pathogen-infected or transformed cells mainly through the cytotoxic granule pathway. [1][2][3][4] Among the five human granzymes (A, B, H, K and M), granzyme B (GB) triggers cell death in both a caspasedependent and caspase-independent manner, although human and mouse GB differ in their requirement for caspase activation. 5-10 GB-induced cell death also involves Bid/Bax/ Bak-mediated mitochondrial outer membrane permeabilization for the release of the apoptogenic factors cytochrome c, Smac/Diablo, Endo G, HtrA2 and AIF. 1,9-17 We have recently reported that mitochondrial reactive oxygen species (ROS) have a critical role in GB pathway by amplifying apoptogenic factor release, oligonucleosomal DNA fragmentation and lysosomal membrane rupture. 18 Mitochondrial ROS resulted from GB-mediated cleavage of NDUFV1, NDUFS1 and NDUFS2, three subunits of mitochondrial complex I. 18 How GB that does not possess a mitochondrial targeting sequence enters the mitochondria is not known. The mitochondrial nucleoid encodes only for 13 structural proteins, while the rest of the mitochondrial proteome is nuclear-encoded and transported to the mitochondria through a sophisticated protein import machinery. [19][20][21][22][23][24][25][26][27][28] The translocase of the outer mitochondrial membrane (TOM), the entry gate to the mitochondria, 20 passes on the cargos to the sorting and assembly machinery (SAM) complex, to the mitochondrial intermembrane space assembly (MIA) complex and to the translocases of the inner mitochondrial membrane TIM22 or TIM23 complexes for delivery to the outer membrane, the intermembrane space, the inner membrane or the matrix, respectively. [19][20][21][22]25,26,28 Unexpectedly, we found that GB mitochondrial entry is independent of the TOM-TIM23 complexes, but requires instead the Sam50 and Tim22 chann...

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Cytotoxic T Cells Use Mechanical Force to Potentiate Target Cell Killing

Cited by 333 publications

References 56 publications

Mechanical Criterion for the Rupture of a Cell Membrane under Compression

Mechanical Criterion for the Rupture of a Cell Membrane under Compression

Mechanosensing drives acuity of αβ T-cell recognition

Granzyme B enters the mitochondria in a Sam50-, Tim22- and mtHsp70-dependent manner to induce apoptosis

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