Antigen-receptor interaction requirement for conjugate formation and lethal-hit triggering by cytotoxic T lymphocytes can be bypassed by protein kinase C activators and Ca2+ ionophores.

Berrebi, Gabriel; Takayama, Hajime; Sitkovsky, Michail V.

doi:10.1073/pnas.84.5.1364

Cited by 38 publications

(25 citation statements)

References 36 publications

(30 reference statements)

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“…TCR-induced cell activation can be simulated by PMA and ionomycin. 11,12 Interestingly, we have seen that PMA alone is sufficient to induce degranulation of preformed FasL. This was unexpected as stimulation with both PMA and ionomycin is required for inducing the release of lytic granule markers.…”

Section: Discussionmentioning

confidence: 91%

“…(d) Human T cell blasts were pre-incubated with or without ML-7 and CD107a and cell surface expression was measured. Typical experiments out of three are shown with previous reports that described the need for both PMA and ionomycin for the activation-induced release of CD63, CD107a, perforin and granzymes 11,12,24 (and own results). Our findings are further supported by the observation that weak TCR signals, that do not elicit a detectable increase in intracellular calcium, induce FasL-mediated killing, whereas strong TCR activation also promote perforin/granzymemediated cytotoxicity.…”

Section: Discussionmentioning

confidence: 98%

“…T cell receptor (TCR)-induced T lymphocyte activation can be simulated by phorbol ester and ionomycin 11,12 activating the protein kinase pathways and the release of intracellular calcium, respectively. Accordingly, we observed that phorbol myristate acetate (PMA) plus ionomycin efficiently induced functional FasL cell surface expression (Figure 2a).…”

Section: Resultsmentioning

confidence: 99%

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Distinct requirements for activation-induced cell surface expression of preformed Fas/CD95 ligand and cytolytic granule markers in T cells

et al. 2008

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Fas (CD95/Apo-1) ligand is a potent inducer of apoptosis and one of the major killing effector mechanisms of cytotoxic T cells. Thus, Fas ligand activity has to be tightly regulated, involving various transcriptional and post-transcriptional processes. For example, preformed Fas ligand is stored in secretory lysosomes of activated T cells, and rapidly released by degranulation upon reactivation. In this study, we analyzed the minimal requirements for activation-induced degranulation of Fas ligand. T cell receptor activation can be mimicked by calcium ionophore and phorbol ester. Unexpectedly, we found that stimulation with phorbol ester alone is sufficient to trigger Fas ligand release, whereas calcium ionophore is neither sufficient nor necessary. The relevance of this process was confirmed in primary CD4 þ and CD8 þ T cells and NK cells. Although the activation of protein kinase(s) was absolutely required for Fas ligand degranulation, protein kinase C or A were not involved. Previous reports have shown that preformed Fas ligand co-localizes with other markers of cytolytic granules. We found, however, that the activationinduced degranulation of Fas ligand has distinct requirements and involves different mechanisms than those of the granule markers CD63 and CD107a/Lamp-1. We conclude that activation-induced degranulation of Fas ligand in cytotoxic lymphocytes is differently regulated than other classical cytotoxic granule proteins.

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Section: Discussionmentioning

confidence: 91%

Section: Discussionmentioning

confidence: 98%

Section: Resultsmentioning

confidence: 99%

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Distinct requirements for activation-induced cell surface expression of preformed Fas/CD95 ligand and cytolytic granule markers in T cells

et al. 2008

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“…13 Control, RMA-exposed NK cells showed a robust [Ca 2ϩ ]i response upon NKG2D cross-linking. In contrast, [Ca 2ϩ ]i mobilization was essentially absent in RMA-H60-exposed NK cells (Figure 4), suggesting that NKG2D on these NK cells is signaling deficient.…”

Section: Basis For Defective Nkg2d Functionmentioning

confidence: 99%

Altered NKG2D function in NK cells induced by chronic exposure to NKG2D ligand–expressing tumor cells

Coudert

Zimmer

Tomasello

et al. 2005

Blood

195

223

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NKG2D is an activation receptor that allows natural killer (NK) cells to detect diseased host cells. The engagement of NKG2D with corresponding ligand results in surface modulation of the receptor and reduced function upon subsequent receptor engagement. However, it is not clear whether in addition to modulation the NKG2D receptor complex and/or its signaling capacity is preserved. We show here that the prolonged encounter with tumor cell-bound, but not soluble, ligand can completely uncouple the NKG2D receptor from the intracellular mobilization of calcium and the exertion of cell-mediated cytolysis. However, cytolytic effector function is intact since NKG2D ligand-exposed NK cells can be activated via the Ly49D receptor. While NKG2D-dependent cytotoxicity is impaired, prolonged ligand exposure results in constitutive interferon ␥ (IFN␥) production, suggesting sustained signal IntroductionNumerous activation receptors can trigger natural killer (NK) cell-mediated cytotoxic and/or cytokine responses. The NKG2D receptor is specific for endogenous ligands (ie, histocompatibility 60 (H60), retinoic acid early inducible gene-1 (RAE-1), and mouse UL16-binding proteinlike transcript 1 (MULT1) in mice), which are usually not expressed on normal resting cells but which are readily induced upon different forms of cellular distress (for a review see Raulet 1 ). Thus, the de novo expression of NKG2D ligands renders cells susceptible to NK cell-mediated lysis. On many tumor cell lines of distinct tissue origins, NKG2D ligands are constitutively expressed. The ectopic expression of NKG2D ligands induces the rejection of several transplantable tumor cell lines. 2,3 Despite NKG2D's prominent role in NK cell activation and T-cell costimulation, it is noteworthy that many established tumor cell lines constitutively express NKG2D ligands, suggesting that tumors can escape NKG2D recognition. Indeed, T and NK cells, which were infiltrating major histocompatibility class I chain-related protein A (MICA)-positive tumors had reduced NKG2D cell surface levels. 4,5 In vitro, soluble MICA or tumor cells expressing MICA induced the down-modulation and degradation of NKG2D expressed by CD8 ϩ T cells, which led to reduced effector functions. 4 In addition, NK cells derived from nonobese diabetic (NOD) mice (a strain of mice prone to autoimmune diabetes) displayed impaired NKG2D function. 6 NKG2D dysfunction and modulation were explained by an interaction of NKG2D with its ligands, which are aberrantly induced on NOD NK cells upon culture. 6 In general, reduced NKG2D function is thought to be the consequence of and directly related to the lower NKG2D cell surface levels. However, it has not been determined whether sustained NKG2D engagement leaves the NKG2D receptor complex and/or its signal transduction capacity intact. To this end we established a culture system in which NK cells are exposed to tumor cells expressing NKG2D ligand. We show that chronic engagement with tumor cell-bound NKG2D ligand can uncouple NKG2D recognition from the ex...

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“…Moreover, we show that the cells exerting the enhanced anti-MOPC-315 lytic activity in the presence of PMA are of the classic CTL phenotype, i. e., are CD8+/CD4 -. Finally, the calcium ionophore ionomycin, which acts synergistically with PMA in some systems to activate CTL lysis against antigenically unrelated tumors [3,10,23], did not lead to lysis of EL4 tumor cells by L-PAM TuB spleen cells exposed to PMA.…”

Section: Introductionmentioning

confidence: 99%

Phorbol ester-induced enhancement in lytic activity of CD8+ splenic T cells from low-dose melphalan-treated MOPC-315-tumor bearers

Weiskirch

Baumgartel

Barker

et al. 1991

Cancer Immunol Immunother

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We have previously shown that while spleen cells from untreated mice bearing a large MOPC-315 tumor are not cytotoxic in vitro for MOPC-315 tumor cells, spleen cells obtained from such mice on day 7 after low-dose melphalan (L-phenylalanine mustard); L-PAM therapy exert a substantial anti-MOPC-315 cytotoxicity [Mokyr et al. (1989) Cancer Res 49: 4597]. Here we show that this anti-MOPC-315 lytic activity is evident by day 5, and peaks on day 7 after the low-dose chemotherapy, at a time when the mice are actively engaged in tumor eradication. Short-term exposure of spleen cells from mice bearing a MOPC-315 tumor and treated with low-dose L-PAM (L-PAM TuB mice) to phorbol 12-myristate 13-acetate (PMA) was found to enhance greatly the ability of these spleen cells to lyse MOPC-315 tumor cells. The highest level of anti-MOPC-315 cytotoxicity was obtained when spleen cells from tumor-bearing mice that had received chemotherapy 7 days earlier were exposed to PMA at a concentration of 1-10 ng/ml. The exertion of the enhanced anti-MOPC-315 lytic activity by L-PAM TuB spleen cells exposed to PMA was found to require CD8+, but not CD4+, T cells. The apparent specificity of the lytic activity exerted by the PMA-stimulated L-PAM TuB spleen cells was illustrated not only by the inability of the spleen cells to lyse an allogeneic, antigenically unrelated thymoma (EL4), but also by their relatively weak lytic activity for two antigenically related syngeneic plasmacytomas. In addition, when EL4 target cells were admixed with MOPC-315 tumor cells, the lytic activity triggered in the L-PAM TuB spleen cells by the MOPC-315 tumor cells plus PMA was not effective in lysing the antigenically unrelated target cells. Moreover, even in the presence of the calcium-specific ionophore, ionomycin, L-PAM TuB spleen cells exposed to PMA were unable to lyse the EL4 target cells. Thus, fresh CD8+ splenic T cells from L-PAM TuB mice that are in the process of eradicating a large MOPC-315 tumor as a consequence of low-dose L-PAM therapy can be triggered with PMA to exert enhanced lytic activity against MOPC-315 tumor cells.(ABSTRACT TRUNCATED AT 400 WORDS)

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Antigen-receptor interaction requirement for conjugate formation and lethal-hit triggering by cytotoxic T lymphocytes can be bypassed by protein kinase C activators and Ca2+ ionophores.

Cited by 38 publications

References 36 publications

Distinct requirements for activation-induced cell surface expression of preformed Fas/CD95 ligand and cytolytic granule markers in T cells

Distinct requirements for activation-induced cell surface expression of preformed Fas/CD95 ligand and cytolytic granule markers in T cells

Altered NKG2D function in NK cells induced by chronic exposure to NKG2D ligand–expressing tumor cells

Phorbol ester-induced enhancement in lytic activity of CD8+ splenic T cells from low-dose melphalan-treated MOPC-315-tumor bearers

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