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Ultrarapid lytic granule release from CTLs activates Ca²⁺-dependent synaptic resistance pathways in melanoma cells.

ABSTRACT: Human cytotoxic T lymphocytes (CTLs) exhibit ultrarapid lytic granule secretion, but whether melanoma cells mobilize defense mechanisms with commensurate rapidity remains unknown. We used single-cell time-lapse microscopy to offer high spatiotemporal resolution analyses of subcellular events in melanoma cells upon CTL attack. Target cell perforation initiated an intracellular Ca²⁺ wave that propagated outward from the synapse within milliseconds and triggered lysosomal mobilization to the synapse, facilitating membrane repair and conferring resistance to CTL induced cytotoxicity. Inhibition of Ca²⁺ flux and silencing of synaptotagmin VII limited synaptic lysosomal exposure and enhanced cytotoxicity. Multiplexed immunohistochemistry of patient melanoma nodules combined with automated image analysis showed that melanoma cells facing CD8⁺ CTLs in the tumor periphery or peritumoral area exhibited significant lysosomal enrichment. Our results identified synaptic Ca²⁺ entry as the definitive trigger for lysosomal deployment to the synapse upon CTL attack and highlighted an unpredicted defensive topology of lysosome distribution in melanoma nodules.

SUBMITTER: Filali L

PROVIDER: S-EPMC8849291 | biostudies-literature | 2022 Feb

REPOSITORIES: biostudies-literature

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Ultrarapid lytic granule release from CTLs activates Ca<sup>2+</sup>-dependent synaptic resistance pathways in melanoma cells.

Filali Liza L Puissegur Marie-Pierre MP Cortacero Kevin K Cussat-Blanc Sylvain S Khazen Roxana R Van Acker Nathalie N Frenois François-Xavier FX Abreu Arnaud A Lamant Laurence L Meyer Nicolas N Vergier Béatrice B Müller Sabina S McKenzie Brienne B Valitutti Salvatore S

Science advances 20220216 7

Human cytotoxic T lymphocytes (CTLs) exhibit ultrarapid lytic granule secretion, but whether melanoma cells mobilize defense mechanisms with commensurate rapidity remains unknown. We used single-cell time-lapse microscopy to offer high spatiotemporal resolution analyses of subcellular events in melanoma cells upon CTL attack. Target cell perforation initiated an intracellular Ca<sup>2+</sup> wave that propagated outward from the synapse within milliseconds and triggered lysosomal mobilization to ...[more]

PMID: 35171665

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Project description:Amyloid β peptide oligomers (AβOs), toxic aggregates with pivotal roles in Alzheimer's disease, trigger persistent and low magnitude Ca2+ signals in neurons. We reported previously that these Ca2+ signals, which arise from Ca2+ entry and subsequent amplification by Ca2+ release through ryanodine receptor (RyR) channels, promote mitochondrial network fragmentation and reduce RyR2 expression. Here, we examined if AβOs, by inducing redox sensitive RyR-mediated Ca2+ release, stimulate mitochondrial Ca2+-uptake, ROS generation and mitochondrial fragmentation, and also investigated the effects of the antioxidant N-acetyl cysteine (NAC) and the mitochondrial antioxidant EUK-134 on AβOs-induced mitochondrial dysfunction. In addition, we studied the contribution of the RyR2 isoform to AβOs-induced Ca2+ release, mitochondrial Ca2+ uptake and fragmentation. We show here that inhibition of NADPH oxidase type-2 prevented the emergence of RyR-mediated cytoplasmic Ca2+ signals induced by AβOs in primary hippocampal neurons. Treatment with AβOs promoted mitochondrial Ca2+ uptake and increased mitochondrial superoxide and hydrogen peroxide levels; ryanodine, at concentrations that suppress RyR activity, prevented these responses. The antioxidants NAC and EUK-134 impeded the mitochondrial ROS increase induced by AβOs. Additionally, EUK-134 prevented the mitochondrial fragmentation induced by AβOs, as previously reported for NAC and ryanodine. These findings show that both antioxidants, NAC and EUK-134, prevented the Ca2+-mediated noxious effects of AβOs on mitochondrial function. Our results also indicate that Ca2+ release mediated by the RyR2 isoform causes the deleterious effects of AβOs on mitochondrial function. Knockdown of RyR2 with antisense oligonucleotides reduced by about 50% RyR2 mRNA and protein levels in primary hippocampal neurons, decreased by 40% Ca2+ release induced by the RyR agonist 4-chloro-m-cresol, and significantly reduced the cytoplasmic and mitochondrial Ca2+ signals and the mitochondrial fragmentation induced by AβOs. Based on our results, we propose that AβOs-induced Ca2+ entry and ROS generation jointly stimulate RyR2 activity, causing mitochondrial Ca2+ overload and fragmentation in a feed forward injurious cycle. The present novel findings highlight the specific participation of RyR2-mediated Ca2+ release on AβOs-induced mitochondrial malfunction.

Dataset Information

Ultrarapid lytic granule release from CTLs activates Ca²⁺-dependent synaptic resistance pathways in melanoma cells.

Publications

Ultrarapid lytic granule release from CTLs activates Ca<sup>2+</sup>-dependent synaptic resistance pathways in melanoma cells.

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Ultrarapid lytic granule release from CTLs activates Ca2+-dependent synaptic resistance pathways in melanoma cells.

Publications

Ultrarapid lytic granule release from CTLs activates Ca&lt;sup&gt;2+&lt;/sup&gt;-dependent synaptic resistance pathways in melanoma cells.

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OmicsDI is part of the ELIXIR infrastructure

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Ultrarapid lytic granule release from CTLs activates Ca²⁺-dependent synaptic resistance pathways in melanoma cells.

Ultrarapid lytic granule release from CTLs activates Ca<sup>2+</sup>-dependent synaptic resistance pathways in melanoma cells.