Autophagy in tobacco suspension-cultured cells in response to sucrose starvation
Autophagy in tobacco suspension-cultured cells in response to sucrose starvation
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DOI：
10.1104/pp.111.4.1233
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发表时间：
1996-08-01
期刊：
影响因子：
7.4
通讯作者：
Ohsumi, Y
中科院分区：
文献类型：
--
作者：
Moriyasu, Y;Ohsumi, Y
研究了烟草悬浮培养细胞(BY-2)对营养饥饿的反应。将在含蔗糖3%(w/v)的Murashige-Skoog培养液中培养的细胞转移到不含蔗糖的相同培养基上，2d内可降解30%~45%的胞内蛋白质，十二烷基硫酸钠-聚丙烯酰胺凝胶电泳法分析表明，蛋白质被非选择性降解。在相同的处理条件下，以荧光素硫代氨基甲酰酪蛋白为底物在pH值为5.0的条件下测得的细胞内蛋白水解酶活性在1d后提高了3~7倍，当半胱氨酸蛋白酶抑制剂(2S，3S)-trans-epoxysuccinyl-L-leucylamido-3-methyl-butane(10mU M)存在于饥饿介质中时，蛋白质的降解和蛋白水解酶活性的增加都被有效地抑制。光镜分析显示，抑制剂处理8h后，胞浆核周区域聚集了许多小的球形小体。这些小体是直径1-6微米的膜结合囊泡，内含多个颗粒。奎纳克林对这些小泡和中央液泡进行了染色，因此，两个细胞器都是酸性隔室。以1-萘磷酸和β-甘油磷酸为底物的细胞化学酶分析表明，这些囊泡中含有一种酸性磷酸酶(S)。我们认为，这些小泡有助于蔗糖饥饿条件下刺激的细胞蛋白质降解。
The response of tobacco (Nicotiana tabacum) suspension-cultured cells (BY-2) to nutrient starvation was investigated. When the cells that were grown in Murashige-Skoog medium containing 3% (w/v) sucrose were transferred to the same medium without sucrose, 30 to 45% of the intracellular proteins were degraded in 2 d. An analysis with sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that proteins were degraded nonselectively. With the same treatment, protease activity in the cell, which was measured at pH 5.0 using fluorescein thiocarbamoyl-casein as a substrate, increased 3- to 7-fold after 1 d. When the cysteine protease inhibitor (2S,3S)-trans-epoxysuccinyl-L-leucylamido-3-methyl-butane (10 mu M) was present in the starvation medium, both the protein degradation and the increase in the protease activity were effectively inhibited. Light microscopy analysis showed that many small spherical bodies accumulated in the perinuclear region of the cytosol 8 h after the start of the inhibitor treatment. These bodies were shown to be membrane-bound vesicles of 1 to 6 mu m in diameter that contained several particles. Quinacrine stained these vesicles and the central vacuole; thus, both organelles are acidic compartments. Cytochemical enzyme analysis using 1-naphthylphosphate and beta-glycerophosphate as substrates showed that these vesicles contained an acid phosphatase(s). We suggest that these vesicles contribute to cellular protein degradation stimulated under sucrose starvation conditions.