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Activation of lysosomal mediated cell death in the course of autophagy by mTORC1 inhibitor.

ABSTRACT: Lysosomal biogenesis plays a vital role in cell fate. Under certain conditions, excessive lysosomal biogenesis leads to susceptibility for lysosomal membrane permeabilization resulting in various pathological conditions including cell death. In cancer cells apoptosis machinery becomes dysregulated during the course of treatment, thus allows cancer cells to escape apoptosis. So it is therefore imperative to identify cytotoxic agents that exploit non-apoptotic mechanisms of cell death. Our study showed that pancreatic cancer cells treated with SDS-203 triggered an incomplete autophagic response and a nuclear translocation of transcriptional factor TFEB. This resulted in abundant biosynthesis and accumulation of autophagosomes and lysosomes into the cells leading to their death. It was observed that the silencing of autophagy genes didn't alter the cell fate, whereas siRNA-mediated silencing of TFEB subdued SDS-203 mediated lysosomal biogenesis and associated cell death. Further mouse tumors treated with SDS-203 showed a significant reduction in tumor burden and increased expression of lysosomal markers. Taken together this study demonstrates that SDS-203 treatment triggers non-apoptotic cell death in pancreatic cancer cells through a mechanism of lysosome over accumulation.

SUBMITTER: Khan SU

PROVIDER: S-EPMC8943151 | biostudies-literature | 2022 Mar

REPOSITORIES: biostudies-literature

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Activation of lysosomal mediated cell death in the course of autophagy by mTORC1 inhibitor.

Khan Sameer Ullah SU Pathania Anup Singh AS Wani Abubakar A Fatima Kaneez K Mintoo Mubashir Javed MJ Hamza Baseerat B Paddar Masroor Ahmad MA Bhumika Wadhwa W Anand Loveleena Kour LK Maqbool Mir Shahid MS Mir Sameer Ahmad SA Kour Jaspreet J Venkateswarlu Vunnam V Mondhe Dilip Manikrao DM Sawant Sanghapal D SD Malik Fayaz F

Scientific reports 20220323 1

Lysosomal biogenesis plays a vital role in cell fate. Under certain conditions, excessive lysosomal biogenesis leads to susceptibility for lysosomal membrane permeabilization resulting in various pathological conditions including cell death. In cancer cells apoptosis machinery becomes dysregulated during the course of treatment, thus allows cancer cells to escape apoptosis. So it is therefore imperative to identify cytotoxic agents that exploit non-apoptotic mechanisms of cell death. Our study s ...[more]

PMID: 35322026

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Project description:Nervous necrosis virus (NNV) is the etiological agent of viral nervous necrosis (VNN), also known as viral encephalopathy and retinopathy (VER), which results in heavy economic losses to the aquaculture industry worldwide. Dramatic cytoplasmic vacuoles were observed during NNV infection both in vitro and in vivo; however, the origin and mechanism of cytoplasmic vacuolization remains unknown. In this report, we found that the cytoplasmic vacuole morphology became fused and enlarged during infection with red spotted grouper nervous necrosis virus (RGNNV), which was accompanied by increased cell death. Notably, Lyso-Tracker, but not Mito-Tracker or ER-Tracker, was accumulated in the vacuoles, and abnormal lysosome swelling was observed in RGNNV-infected cells, suggesting that the cytoplasmic vacuoles originated from lysosomal organelles. Cytoplasmic vacuolization and cell death in RGNNV-infected cells was completely blocked by the vacuolar H+-ATPase inhibitor (bafilomycin A1), and was significantly weakened by chloroquine (CQ), a lysosomotropic agent that induces the acidification of the lysosomes. This suggests that lysosome acidification was essential for vacuole formation. Significant inhibitory effects on vacuolization and cell death were also observed in the RGNNV-infected cells following treatment with nigericin and monensin (ionophores that uncouple the proton gradient present in lysosomes). This indicated that lysosome function was tightly associated with RGNNV infection-induced cell death. In addition, vacuoles were found to be partially co-localized with GFP-LC3II punctate dots during RGNNV infection. Moreover, the severity of vacuolization and cell death were both significantly decreased after treatment with the autophagy inhibitor, 3-MA, suggesting that autophagy was involved in lysosomal vacuolization and cell death evoked by RGNNV infection. Thus, our results demonstrate that autophagy participates in lysosomal vacuolation-mediated cell death during RGNNV infection, and provides new insight into our understanding of the potential mechanisms underlying nodavirus pathogenesis in vitro.

Project description:Increasing evidence suggests that induction of lethal macroautophagy/autophagy carries potential significance for the treatment of glioblastoma (GBM). In continuation of previous work, we demonstrate that pimozide and loperamide trigger an ATG5- and ATG7 (autophagy related 5 and 7)-dependent type of cell death that is significantly reduced with cathepsin inhibitors and the lipid reactive oxygen species (ROS) scavenger α-tocopherol in MZ-54 GBM cells. Global proteomic analysis after treatment with both drugs also revealed an increase of proteins related to lipid and cholesterol metabolic processes. These changes were accompanied by a massive accumulation of cholesterol and other lipids in the lysosomal compartment, indicative of impaired lipid transport/degradation. In line with these observations, pimozide and loperamide treatment were associated with a pronounced increase of bioactive sphingolipids including ceramides, glucosylceramides and sphingoid bases measured by targeted lipidomic analysis. Furthermore, pimozide and loperamide inhibited the activity of SMPD1/ASM (sphingomyelin phosphodiesterase 1) and promoted induction of lysosomal membrane permeabilization (LMP), as well as release of CTSB (cathepsin B) into the cytosol in MZ-54 wild-type (WT) cells. Whereas LMP and cell death were significantly attenuated in ATG5 and ATG7 knockout (KO) cells, both events were enhanced by depletion of the lysophagy receptor VCP (valosin containing protein), supporting a pro-survival function of lysophagy under these conditions. Collectively, our data suggest that pimozide and loperamide-driven autophagy and lipotoxicity synergize to induce LMP and cell death. The results also support the notion that simultaneous overactivation of autophagy and induction of LMP represents a promising approach for the treatment of GBM.Abbreviations: ACD: autophagic cell death; AKT1: AKT serine/threonine kinase 1; ATG5: autophagy related 5; ATG7: autophagy related 7; ATG14: autophagy related 14; CERS1: ceramide synthase 1; CTSB: cathepsin B; CYBB/NOX2: cytochrome b-245 beta chain; ER: endoplasmatic reticulum; FBS: fetal bovine serum; GBM: glioblastoma; GO: gene ontology; HTR7/5-HT7: 5-hydroxytryptamine receptor 7; KD: knockdown; KO: knockout; LAMP1: lysosomal associated membrane protein 1; LAP: LC3-associated phagocytosis; LMP: lysosomal membrane permeabilization; MAP1LC3B: microtubule associated protein 1 light chain 3 beta; MTOR: mechanistic target of rapamycin kinase; RB1CC1: RB1 inducible coiled-coil 1; ROS: reactive oxygen species; RPS6: ribosomal protein S6; SMPD1/ASM: sphingomyelin phosphodiesterase 1; VCP/p97: valosin containing protein; WT: wild-type.

Dataset Information

Activation of lysosomal mediated cell death in the course of autophagy by mTORC1 inhibitor.

Publications

Activation of lysosomal mediated cell death in the course of autophagy by mTORC1 inhibitor.

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