TRANSLOCATION OF NEWLY SYNTHESIZED GANGLIOSIDES TO THE CELL-SURFACE
TRANSLOCATION OF NEWLY SYNTHESIZED GANGLIOSIDES TO THE CELL-SURFACE
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DOI：
10.1021/bi00257a003
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发表时间：
1982-01-01
期刊：
影响因子：
2.9
通讯作者：
FISHMAN, PH
中科院分区：
文献类型：
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作者：
MILLERPODRAZA, H;FISHMAN, PH;FISHMAN, PH
Steven Dower， 8 Halina Miller-Podraza和Peter H. Fishman摘要：一种新的方法被开发出来跟踪神经节苷脂从细胞内的合成位点到质膜的转运。培养的小鼠神经母细胞瘤N18和大鼠神经胶质瘤C6细胞用D-[l-3H]半乳糖标记，然后用NaI04轻度氧化。在选择的条件下，氧化基本上局限于细胞表面唾液酸残基，这些残基转化为具有醛功能的衍生物。将标记的神经节苷脂从细胞中分离出来，与二硝基苯肼反应生成氧化神经节苷脂的二硝基苯（DNP）衍生物。然后用薄层色谱法将dnp -神经节苷与未修饰的对应物分离。因此，将表面神经节苷脂的标记率与总神经节苷脂的标记率区分开来。我们的研究结果表明，神经节苷脂从合成位点转移到细胞表面大约需要20分钟，并且新合成的神经节苷脂似乎以恒定的速率运输到质膜。不同神经节苷类在N18细胞内的易位率以及N18和C6细胞间的易位率均无本质差异。许多神经节苷脂的生物合成途径已被描述（Fishman & Brady, 1976）。一般认为，节脂苷的碳水化合物链是由膜结合的多酶复合物形成的，该复合物催化活性单糖逐步加入糖脂受体（Roseman, 1970; Fishman, 1974; Caputto et al., 1976）。也有人认为神经节苷基化发生在高尔基体和/或细胞内质网中（Keenan et al., 1974; Fleischer, 1977; Pacuszka et al., 1978; Eppler et al., 1980），从那里新合成的化合物移动到它们的最终目的地，即质膜。目前对……知之甚少
Steven Dower, 8 Halina Miller-Podraza, and Peter H. Fishman abstract: A new method was developed to follow the translocation of gangliosides from their site of synthesis within the cell to the plasma membrane. Cultured mouse neuro-blastoma N18 and rat glioma C6 cells were labeled for in-creasing times with D-[l-3H] galactose and then subjected to mild oxidation with NaI04. Underthe conditions chosen, oxidation was essentially restricted to cell-surface sialic acid residues, which were converted to derivatives with an aldehyde function. The labeled gangliosides were isolated from thecells and reacted with dinitrophenylhydrazine to form dinitrophenyl (DNP) derivatives of the oxidized gangliosides. The DNP-gangliosides then were separated from their unmodified counterparts by thin-layer chromatography. Thus, the rate of labeling of surface gangliosides was distinguished from the rate of labeling of total gangliosides. Our results indicated that the transfer of gangliosides from thesite of synthesis to the cell surface required approximately 20 min and that newly synthesized gangliosides appeared to be transported to the plasma membrane at a constant rate. No essential differences were found in the rates of translocation of different ganglioside species by N18 cells or between gangliosides of N18 and C6 cells. e biosynthetic pathways of many gangliosides have been described (Fishman & Brady, 1976). It is generally accepted that the carbohydrate chains of gangliosides are formed by membrane-bound multienzyme complexes which catalyze the stepwise addition of activated monosaccharides to glycolipid acceptors (Roseman, 1970; Fishman, 1974; Caputto et al., 1976). It is also believed that glycosylation of gangliosides takes place in the Golgi apparatus and/or in the endoplasmic reticulum of cells (Keenan et al., 1974; Fleischer, 1977; Pacuszka et al., 1978; Eppler et al., 1980) from where the newly synthesized compounds move to their final destination, the plasma membrane. Very little is known at present about the