Department of Biochemistry and Cell Biology, Faculty of Veterinary Medicine, and Institute of Biomembranes, University of Utrecht, 3584 CM, The Netherlands.
J Lipid Res. 2009 Nov;50(11):2182-92. doi: 10.1194/jlr.M800660-JLR200. Epub 2009 May 20.
The mutant Chinese hamster ovary cell line MT58 contains a thermosensitive mutation in CTP:phosphocholine cytidylyltransferase, the regulatory enzyme in the CDP-choline pathway. As a result, MT58 cells have a 50% decrease in their phosphatidylcholine (PC) level within 24 h when cultured at the nonpermissive temperature (40 degrees C). This is due to a relative rapid breakdown of PC that is not compensated for by the inhibition of de novo PC synthesis. Despite this drastic decrease in cellular PC content, cells are viable and can proliferate by addition of lysophosphatidylcholine. By [(3)H]oleate labeling, we found that the FA moiety of the degraded PC is recovered in triacylglycerol. In accordance with this finding, an accumulation of lipid droplets is seen in MT58 cells. Analysis of PC-depleted MT58 cells by electron and fluorescence microscopy revealed a partial dilation of the rough endoplasmic reticulum, resulting in spherical structures on both sites of the nucleus, whereas the morphology of the plasma membrane, mitochondria, and Golgi complex was unaffected. In contrast to these morphological observations, protein transport from the ER remains intact. Surprisingly, protein transport at the level of the Golgi complex is impaired. Our data suggest that the transport processes at the Golgi complex are regulated by distal changes in lipid metabolism.
突变的中国仓鼠卵巢细胞系 MT58 在 CTP:磷酸胆碱胞苷转移酶中有一个热敏突变，该酶是 CDP-胆碱途径中的调节酶。结果，当在非许可温度（40 摄氏度）下培养时，MT58 细胞的磷脂酰胆碱（PC）水平在 24 小时内降低了 50%。这是由于 PC 的相对快速分解，而新合成的 PC 合成被抑制，无法得到补偿。尽管细胞内 PC 含量急剧下降，但细胞仍具有活力，并可以通过添加溶血磷脂酰胆碱进行增殖。通过 [(3)H]油酸标记，我们发现降解的 PC 的 FA 部分在三酰基甘油中被回收。根据这一发现，在 MT58 细胞中可以看到脂质滴的积累。用电子和荧光显微镜对 PC 耗尽的 MT58 细胞进行分析显示，粗面内质网部分扩张，导致核两侧出现球形结构，而质膜、线粒体和高尔基体的形态不受影响。与这些形态学观察结果相反，内质网的蛋白质运输仍然完整。令人惊讶的是，高尔基体水平的蛋白质运输受到损害。我们的数据表明，高尔基体的运输过程受到远端脂质代谢变化的调节。
