Cardamonin inhibits cell proliferation by caspase-mediated cleavage of Raptor
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ORIGINAL ARTICLE
Cardamonin inhibits cell proliferation by caspase-mediated cleavage of Raptor Yanting Zhu 1 & Jintuo Zhou 1 & Peiguang Niu 1 & Huajiao Chen 1 & Daohua Shi 1 Received: 31 August 2020 / Accepted: 4 October 2020 # Springer-Verlag GmbH Germany, part of Springer Nature 2020
Abstract The antiproliferative effect of cardamonin on mTORC1 is related with downregulation of Raptor. We investigated the mechanism that cardamonin decreases Raptor expression through caspase-mediated protein degradation. SKOV3 cells and HeLa cells were pretreated with caspase inhibitor z-VAD-fmk for 30 min and then exposed to different doses of cardamonin and cisplatin, respectively. We analyzed the gene expression of caspases based on TCGA and GTEx gene expression data in serous cystadenocarcinoma and normal tissues, monitored caspase activity by caspase colorimetric assay kit, detected expression of mTORC1-associated proteins and apoptosis-associated proteins by western blotting, and finally detected cell viability by methyl thiazolyl tetrazolium (MTT) assay. A different expression of caspases except caspase-1 was found between serous cystadenocarcinoma and normal tissues. Raptor was cleaved when caspases were activated by cisplatin and caspase-6/caspase-8 was activated by cardamonin in SKOV3 cells. We further used a monoclonal antibody recognizing the N-terminal part of Raptor to find that Raptor was cleaved into a smaller fragment of about 70 kDa by cardamonin and was rescued by z-VADfmk treatment. As a result of Raptor cleavage, mTORC1 activity was decreased and cell viability was inhibited, while cell apoptosis was induced in SKOV3 cells. Notably, similar results are only observed in HeLa cells with a high dose of cardamonin. We concluded that caspase-mediated cleavage of Raptor might be an important mechanism in that cardamonin regulated Raptor and mTORC1 activity. Keywords Raptor . Caspase . Cardamonin . mTORC1
Introduction Caspases are a family of cysteine proteases, which play an important role in the process of programmed cell death or apoptosis. There are three main groups in human caspase family including the inflammatory caspases 1, 4, 5, and 12; the apoptotic effector caspases 3, 6, and 7; and the human initiator caspases 2, 8, 9, and 10(Van Opdenbosch and Lamkanfi 2019). Caspase-6 has been characterized as an executioner caspase because of its short prodomain and sequence Electronic supplementary material The online version of this article (https://doi.org/10.1007/s00210-020-01986-z) contains supplementary material, which is available to authorized users. * Daohua Shi [email protected]; [email protected] 1
Department of Pharmacy, Fujian Maternity and Child Health Hospital, Affiliated Hospital of Fujian Medical University, 18 Daoshan Road, Fuzhou 350001, Fujian, China
recognition motif, but its own activation is not sufficient to result in apoptosis of all cells (Gray et al. 2010). Upon activation, caspases begin to cleave the specific substrates, and for the most part, it cooperates in the process of cell death t
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