Silencing of Barkor/ATG14 sensitizes osteosarcoma cells to cisplatin‑induced apoptosis

Zhao,Zhifang; Tao,Lijiang; Shen,Chengchun; Liu,Bing; Yang,Zhengming; Tao,Huimin

doi:10.3892/ijmm.2013.1578

Silencing of Barkor/ATG14 sensitizes osteosarcoma cells to cisplatin‑induced apoptosis

Authors:
- Zhifang Zhao
- Lijiang Tao
- Chengchun Shen
- Bing Liu
- Zhengming Yang
- Huimin Tao
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Affiliations: Department of Orthopedic Surgery, The 117th Hospital of People's Liberation Army, Hangzhou, Zhejiang, P.R. China, Department of Orthopedic Surgery, The Second Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou, Zhejiang, P.R. China
Published online on: December 9, 2013 https://doi.org/10.3892/ijmm.2013.1578
Pages: 271-276
Copyright: © Zhao et al. This is an open access article distributed under the terms of Creative Commons Attribution License [CC BY_NC 3.0].

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Abstract

Although surgical excision following neoadjuvant chemotherapy has contributed to the long-term survival of osteosarcoma patients, patients that do not respond to commonly used drugs including cisplatin, have a poor prognosis. Autophagy is important in the inhibition of chemotherapeutic apoptosis. Therefore, we investigated whether knockdown of Beclin1-associated autophagy-related key regulator (Barkor/ATG14) promoted cisplatin-induced apoptosis in a drug-resistant osteosarcoma cell line in vitro. Saos-2 cells were transfected with Barkor siRNA. Sensitivity of the Barkor siRNA-transfected cell line to cisplatin was evaluated. Silencing of Barkor did not directly inhibit the growth rate of the transfected cells, but it significantly increased their sensitivity to cisplatin. The results of flow cytometry and 4',6-diamidino-2-phenylindole (DAPI) staining revealed that Barkor siRNA-transfected Saos-2 cells treated with cisplatin exhibited much higher rates of apoptosis than the control and control siRNA-transfected cells. Additionally, the combination of silencing of Barkor with cisplatin treatment promoted the expression of caspase-12 and calpain. The increase of cisplatin cytotoxicity may therefore be involved in endoplasmic reticulum (ER) stress-associated apoptosis. Bcl-2 was markedly downregulated in dose‑dependent cisplatin‑treated Barkor-transfected-Saos-2 cells. Findings of the present study suggest that the combination of silencing of Barkor and cisplatin enhanced the antitumor efficacy through the Barkor‑related ER- and mitochondrial-mediated apoptotic pathway.

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1	Mirabello L, Troisi RJ and Savage SA: Osteosarcoma incidence and survival rates from 1973 to 2004: data from the Surveillance, Epidemiology, and End Results Program. Cancer. 115:1531–1543. 2009. View Article : Google Scholar : PubMed/NCBI
2	Ottaviani G and Jaffe N: The epidemiology of osteosarcoma. Cancer Treat Res. 152:3–13. 2009. View Article : Google Scholar
3	Wittig JC, Bickels J, Priebat D, et al: Osteosarcoma: a multidisciplinary approach to diagnosis and treatment. Am Fam Physician. 65:1123–1132. 2002.PubMed/NCBI
4	Meyers PA, Schwartz CL, Krailo M, et al: Osteosarcoma: a randomized, prospective trial of the addition of ifosfamide and/or muramyl tripeptide to cisplatin, doxorubicin, and high-dose methotrexate. J Clin Oncol. 23:2004–2011. 2005. View Article : Google Scholar : PubMed/NCBI
5	Reggiori F and Klionsky DJ: Autophagy in the eukaryotic cell. Eukaryot Cell. 1:11–21. 2002. View Article : Google Scholar : PubMed/NCBI
6	Codogno P and Meijer AJ: Autophagy and signaling: their role in cell survival and cell death. Cell Death Differ. 12(Suppl 2): 1509–1518. 2005. View Article : Google Scholar : PubMed/NCBI
7	Levine B and Yuan J: Autophagy in cell death: an innocent convict? J Clin Invest. 115:2679–2688. 2005. View Article : Google Scholar : PubMed/NCBI
8	Sun WL, Chen J, Wang YP and Zheng H: Autophagy protects breast cancer cells from epirubicin-induced apoptosis and facilitates epirubicin-resistance development. Autophagy. 7:1035–1044. 2011. View Article : Google Scholar : PubMed/NCBI
9	O’Donovan TR, O’Sullivan GC and McKenna SL: Induction of autophagy by drug-resistant esophageal cancer cells promotes their survival and recovery following treatment with chemotherapeutics. Autophagy. 7:509–524. 2011.PubMed/NCBI
10	Zhong Y, Wang QJ, Li X, et al: Distinct regulation of autophagic activity by Atg14L and Rubicon associated with Beclin 1-phosphatidylinositol-3-kinase complex. Nat Cell Biol. 11:468–476. 2009. View Article : Google Scholar : PubMed/NCBI
11	Matsunaga K, Saitoh T, Tabata K, et al: Two Beclin 1-binding proteins, Atg14L and Rubicon, reciprocally regulate autophagy at different stages. Nat Cell Biol. 11:385–396. 2009. View Article : Google Scholar : PubMed/NCBI
12	Momoi T: Caspases involved in ER stress-mediated cell death. J Chem Neuroanat. 28:101–105. 2004. View Article : Google Scholar : PubMed/NCBI
13	Nakagawa T and Yuan J: Cross-talk between two cysteine protease families. Activation of caspase-12 by calpain in apoptosis. J Cell Biol. 150:887–894. 2000. View Article : Google Scholar : PubMed/NCBI
14	Tan Y, Dourdin N, Wu C, De Veyra T, Elce JS and Greer PA: Ubiquitous calpains promote caspase-12 and JNK activation during endoplasmic reticulum stress-induced apoptosis. J Biol Chem. 281:16016–16024. 2006. View Article : Google Scholar : PubMed/NCBI
15	Masud A, Mohapatra A, Lakhani SA, Ferrandino A, Hakem R and Flavell RA: Endoplasmic reticulum stress-induced death of mouse embryonic fibroblasts requires the intrinsic pathway of apoptosis. J Biol Chem. 282:14132–14139. 2007. View Article : Google Scholar : PubMed/NCBI
16	Rao RV, Hermel E, Castro-Obregon S, et al: Coupling endoplasmic reticulum stress to the cell death program. Mechanism of caspase activation. J Biol Chem. 276:33869–33874. 2001. View Article : Google Scholar : PubMed/NCBI
17	Martinez JA, Zhang Z, Svetlov SI, Hayes RL, Wang KK and Larner SF: Calpain and caspase processing of caspase-12 contribute to the ER stress-induced cell death pathway in differentiated PC12 cells. Apoptosis. 15:1480–1493. 2010. View Article : Google Scholar : PubMed/NCBI
18	Huang J, Ni J, Liu K, et al: HMGB1 promotes drug resistance in osteosarcoma. Cancer Res. 72:230–238. 2012. View Article : Google Scholar : PubMed/NCBI
19	Picci P: Osteosarcoma (osteogenic sarcoma). Orphanet J Rare Dis. 2:62007. View Article : Google Scholar
20	Yang Z and Klionsky DJ: Eaten alive: a history of macroautophagy. Nat Cell Biol. 12:814–822. 2010. View Article : Google Scholar : PubMed/NCBI
21	Itakura E, Kishi C, Inoue K and Mizushima N: Beclin 1 forms two distinct phosphatidylinositol 3-kinase complexes with mammalian Atg14 and UVRAG. Mol Biol Cell. 19:5360–5372. 2008. View Article : Google Scholar : PubMed/NCBI
22	Sun Q, Fan W, Chen K, Ding X, Chen S and Zhong Q: Identification of Barkor as a mammalian autophagy-specific factor for Beclin 1 and class III phosphatidylinositol 3-kinase. Proc Natl Acad Sci USA. 105:19211–19216. 2008. View Article : Google Scholar : PubMed/NCBI
23	Fan W, Nassiri A and Zhong Q: Autophagosome targeting and membrane curvature sensing by Barkor/Atg14(L). Proc Natl Acad Sci USA. 108:7769–7774. 2011. View Article : Google Scholar : PubMed/NCBI