巨自噬与分子伴侣介导的自噬在饥饿诱导的Raji细胞中相继顺序被激活
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摘要
目的研究巨自噬与分子伴侣介导的自噬在饥饿诱导的Raji细胞中出现的先后顺序及其机制。方法用激光共聚焦显微镜和荧光显微镜观察MDC荧光染色后,Raji细胞内自噬体的形成情况; Western blot检测细胞LC3Ⅰ/Ⅱ、Beclin-1、Hsc70及Lamp-2a等蛋白的表达水平。结果在饥饿诱导的Raji细胞中,巨自噬与分子伴侣介导的自噬是相继顺序,而非同步被激活,巨自噬活性在饥饿诱导的最初几小时快速升高并达到峰值,然后随着饥饿时间的延长逐渐下降,而与巨自噬活性下降相伴随的是分子伴侣介导的自噬活性逐渐增加。结论肿瘤细胞在饥饿诱导的不同阶段通过不同的自噬方式应对压力刺激,巨自噬与分子伴侣介导的自噬之间此消彼长、协调补充的关系更有助于肿瘤细胞适应内外环境的变化,维持自身的生长与增殖。
Aim To research the cross-talk and conversion between macroautophagy and chaperone-mediated autophagy( CMA) in cultured Burkitt lymphoma Raji cells induced by starvation. Methods The autophagic vacuoles were observed by fluorescence microscopy and confocal laser-scanning microscopy with monodansylcadaverine staining. The expression of autophagy associated-proteins were determined by Western blot. Results Both macroautophagy and CMA were activated sequentially instead of simultaneously in starvation-induced Raji cells,and macroautophagy was quickly activated and peaked during the first hours of nutrition deprivation,and then gradually decreased tonear baseline. With starvation persisted,CMA progressively increased along with the decline of macroautophagy. Conclusions Macroautophagy and CMA are maximally activated during different stages of starvation.Activation of these two pathways is often sequential.The sequential switch from macroautophagy to CMA might be conducive to the adaption of cancer cells to miscellaneous intracellular or extracellular changes,maintaining their own growth and proliferation.
Aim To research the cross-talk and conversion between macroautophagy and chaperone-mediated autophagy( CMA) in cultured Burkitt lymphoma Raji cells induced by starvation. Methods The autophagic vacuoles were observed by fluorescence microscopy and confocal laser-scanning microscopy with monodansylcadaverine staining. The expression of autophagy associated-proteins were determined by Western blot. Results Both macroautophagy and CMA were activated sequentially instead of simultaneously in starvation-induced Raji cells,and macroautophagy was quickly activated and peaked during the first hours of nutrition deprivation,and then gradually decreased tonear baseline. With starvation persisted,CMA progressively increased along with the decline of macroautophagy. Conclusions Macroautophagy and CMA are maximally activated during different stages of starvation.Activation of these two pathways is often sequential.The sequential switch from macroautophagy to CMA might be conducive to the adaption of cancer cells to miscellaneous intracellular or extracellular changes,maintaining their own growth and proliferation.
引文
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[2]Li C L,Wei H L,Chen J,et al.Arsenic trioxide induces autophagy and antitumor effects in Burkitt's lymphoma Raji cells[J].Oncol Rep,2014,4:1557-63.
[3]林宇宁,王声善,吕贝贝,等.白花丹醌对肝癌细胞SMMC-7721凋亡和自噬的影响[J].中国药理学通报,2018,34(10):1419-24.[3]Lin Y N,Wang S S,Lyu B B,et al.Effect of plumbagin on apoptosis and autophagy of hepatocellular carcinoma cell line SMMC-7721[J].Chin Pharmacol Bull,2018,34(10):1419-24.
[4]Du P,Cao H,Wu H R,et al.Blocking Bcl-2 leads to autophagy activation and cell death of the HepG2 liver cancer cell line[J].Asian Pac J Cancer Prev,2013,14:5849-53.
[5]Kon M,Kiffin R,Koga H,et al.Chaperone-mediated autophagy is required for tumor growth[J].Cancer,2011,109:1-13.
[6]Cuervo A M,Dice J F.Regulation of lamp2a levels in the lysosomal membrane[J].Traffic,2000,1:570-83.
[7]Cuervo A M,Mann L,Bonten E J,et al.Cathepsin A regulates chaperone-mediated autophagy through cleavage of the lysosomal receptor[J].EMBO J,2003,22,47-59.
[8]Park C,Cuervo A M.Selective autophagy:talking with the UPS[J].Cell Biochem Biophys,2013,67:3-13.
[9]Saha T.LAMP2A overexpression in breast tumors promotes cancer cell survival via chaperone-mediated autophagy[J].Autophagy,2012,8:1643-56.
[10]Massey A C,Zhang C,Cuervo A M.Chaperone-mediated autophagy in aging and disease[J].Curr Top Dev Biol,2006,73:205-35.
[11]Rodriguez M N,Koga H,Garcia L L,et al.Balance between autophagic pathways preserves retinal homeostasis[J].Aging Cell,2013,12:478-88.
[12]Mark S K,Mc Cormark A L,Manning-Bog A B,et al.Lysosomal degradation ofα-synuclein in vivo[J].J Biol Chem,2010,285(18):13621-9.
[13]Wang Y,Martinez-Vicente M,Krger U,et al.Tau fragmentation,aggregation and clearance:the dual role of lysosomal processing[J].Hum Mol Genet,2009,18(21):4153-70.
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