Cold atmospheric plasma induces stress granule formation <i>via</i> an eIF2α-dependent pathway

Motaln, Helena; Čerček, Urša; Recek, Nina; Česnik, Ana Bajc; Mozetič, Miran; Rogelj, Boris

doi:10.1039/d0bm00488j

Cited by 11 publications

(10 citation statements)

References 39 publications

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“…SH-SY5Y-TR-FRT cell line was generated as described before. 18 FlpIn SH-SY5Y-TR-FRT cells containing a single FRT insertion site were used for the generation of an inducible SH-SY5Y-TR-FRT-mScarletI-DPR cell line. Gene sequences of PolyGA, polyGP, polyPA and polyPR were cut out of the pEGFP constructs using KpnI and HindIII restriction enzymes (both Thermo Scientific) and polyGR construct were cut out of a pcDNA5-myc plasmid by KpnI and EcoRV (Thermo Scientific) enzymes.…”

Section: Methodsmentioning

confidence: 99%

Interactome screening ofC9orf72dipeptide repeats reveals VCP sequestration and functional impairment by polyGA

Božič

Motaln

Janež

et al. 2021

Brain

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Repeat expansions in the C9orf72 gene are a common cause of amyotrophic lateral sclerosis and frontotemporal lobar degeneration, two devastating neurodegenerative disorders. One of the proposed mechanisms of GGGGCC repeat expansion is their translation into non-canonical dipeptide repeats, which can then accumulate as aggregates and contribute to these pathologies. There are five different dipeptide repeat proteins (polyGA, polyGR, polyPR, polyPA and polyGP), some of which are known to be neurotoxic. In the present study, we used BioID2 proximity labelling to identify the interactomes of all five dipeptide repeat proteins consisting of 125 repeats each. We identified 113 interacting partners for polyGR, 90 for polyGA, 106 for polyPR, 25 for polyPA, and 27 for polyGP. Gene Ontology enrichment analysis of the proteomic data revealed that these target interaction partners are involved in a variety of functions, including protein translation, signal transduction pathways, protein catabolic processes, amide metabolic processes, and RNA-binding. Using autopsy brain tissue from patients with C9orf72 expansion complemented with cell culture analysis, we evaluated the interactions between polyGA and valosin-containing protein (VCP). Functional analysis of this interaction revealed sequestration of VCP with polyGA aggregates, altering levels of soluble VCP protein. VCP also functions in autophagy processes, and consistent with this, we observed altered autophagy in cells expressing polyGA. We also observed altered co-localisation of polyGA aggregates and p62 in cells depleted of the VCP protein. Altogether these data suggest that sequestration of VCP with polyGA aggregates contributes to the loss of VCP function, and consequently to alterations in autophagy processes in C9orf72 expansion disorders.

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Section: Methodsmentioning

confidence: 99%

Interactome screening ofC9orf72dipeptide repeats reveals VCP sequestration and functional impairment by polyGA

Božič

Motaln

Janež

et al. 2021

Brain

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“…CAP treatment promotes cell growth or cell death depending on the cell type, plasma type, and exposure parameters [26,27]. The exposure parameters effects on these processes were presented already by Bauer [28] and Tanaka [29].…”

Section: Plasma's Ability To Differentially Affect Cell Fitnessmentioning

confidence: 83%

“…The cells' early response to this oxidative stress signals very much resembles the initial response of the cells to transient stress, which is reversible and allows cells to resume normal cell functions or even boost their metabolism after stress withdrawal. This way CAP was shown to induce stress granule formation in the exposed cells via eIF2α-signalling, and in dynamic very much resembling stress granule formation in cells exposed to Arsenite-the oxidative stress inducer [26]. Stress granules are transient structures within the cells formed by proteins and RNA, that initiate translation halt, and protect the cellular proteins and RNA during stress.…”

Section: Plasma's Ability To Differentially Affect Cell Fitnessmentioning

confidence: 84%

Intracellular Responses Triggered by Cold Atmospheric Plasma and Plasma-Activated Media in Cancer Cells

2021

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Cold atmospheric plasma (CAP), an ionized gas operating at room temperature, has been increasingly studied with respect to its potential use in medicine, where its beneficial effects on tumor reduction in oncology have been demonstrated. This review discusses the cellular changes appearing in cell membranes, cytoplasm, various organelles, and DNA content upon cells’ direct or indirect exposure to CAP or CAP-activated media/solutions (PAM), respectively. In addition, the CAP/PAM impact on the main cellular processes of proliferation, migration, protein degradation and various forms of cell death is addressed, especially in light of CAP use in the oncology field of plasma medicine.

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“…Neuroblastoma cells SH-SY5Y cell lines (ATCC ® CRL-2266 ™ ), FlpIn G3BP1 SH-SY5Y generated by us, 43 and the NSC-34 motor neuron-like cells obtained from Cedarlane Laboratories were maintained and differentiated as published. 43-45 Along with mouse primary cortical neurons, these differentiated cells were transfected with 1 μg of c-Src, c-Fyn, or c-Abl plasmid, or in combination with 1 μg of GFP-FUS wild-type plasmid using NeuroMag (OZ Biosciences) according to the manufacturer’s instructions. Untransfected differentiated cells/neurons and those transfected with GFP-FUS wild-type only were used as controls.…”

Section: Methodsmentioning

confidence: 99%

Abl kinase-mediated FUS Tyr526 phosphorylation alters nucleocytoplasmic FUS localization in FTLD-FUS

Motaln

Čerček

Yamoah

et al. 2023

Brain

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Nuclear to cytoplasmic mislocalization and aggregation of multiple RNA-binding proteins (RBPs), including Fused in sarcoma (FUS), are the main neuropathological features of the majority of cases of amyotrophic lateral sclerosis (ALS) and frontotemporal lobular degeneration (FTLD). In ALS-FUS these aggregates arise from disease-associated mutations in FUS, whereas in FTLD-FUS the cytoplasmic inclusions do not contain mutant FUS, suggesting different molecular mechanisms of FUS pathogenesis in FTLD that remain to be investigated. We have previously shown that phosphorylation of the C-terminal Tyr526 of FUS results in increased cytoplasmic retention of FUS due to impaired binding to the nuclear import receptor Transportin 1 (TNPO1). Inspired by the above notions, in the current study we developed a novel antibody against the C-terminally phosphorylated Tyr526 FUS (FUSp-Y526) that is specifically capable of recognizing phosphorylated cytoplasmic FUS, which is poorly recognized by other commercially available FUS antibodies. Using this FUSp-Y526 antibody, we demonstrated a FUS phosphorylation-specific effect on the cytoplasmic distribution of soluble and insoluble FUSp-Y526 in various cells and confirmed the involvement of the Src kinase family in Tyr526 FUS phosphorylation. In addition, we found that FUSp-Y526 expression pattern corelates with active pSrc/pAbl kinases in specific brain regions of mice, indicating preferential involvement of cAbl in the cytoplasmic mislocalization of FUSp-Y526 in cortical neurons. Finally, the pattern of immunoreactivity of active cAbl kinase and FUSp-Y526 revealed altered cytoplasmic distribution of FUSp-Y526 in cortical neurons of post-mortem frontal cortex tissue from FTLD patients compared with controls. The overlap of FUSp-Y526 and FUS signals was found preferentially in small diffuse inclusions and was absent in mature aggregates, suggesting possible involvement of FUSp-Y526 in the formation of early toxic FUS aggregates in the cytoplasm that are largely undetected by commercially available FUS antibodies. Given the overlapping patterns of cAbl activity and FUSp-Y526 distribution in cortical neurons, and cAbl induced sequestration of FUSp-Y526 into G3BP1 positive granules in stressed cells, we propose that cAbl kinase is actively involved in mediating cytoplasmic mislocalization and promoting toxic aggregation of wild-type FUS in the brains of FTLD patients, as a novel putative underlying mechanism of FTLD-FUS pathophysiology and progression.

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Cold atmospheric plasma induces stress granule formation via an eIF2α-dependent pathway

Abstract: Cold atmospheric plasma treatment of FlpIn SH-SY5Y cells with an inducible expression of G3BP1 results in stress granule assembly resembling a cellular oxidative stress response that has been shown to be eIF2α-signaling dependent and inhibited by an ISRIB inhibitor.

Cited by 11 publications

References 39 publications

Interactome screening ofC9orf72dipeptide repeats reveals VCP sequestration and functional impairment by polyGA

Interactome screening ofC9orf72dipeptide repeats reveals VCP sequestration and functional impairment by polyGA

Intracellular Responses Triggered by Cold Atmospheric Plasma and Plasma-Activated Media in Cancer Cells

Abl kinase-mediated FUS Tyr526 phosphorylation alters nucleocytoplasmic FUS localization in FTLD-FUS

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