Isolation and characterization of a Chinese hamster ovary cell mutant with altered regulation of phosphatidylserine biosynthesis.
Isolation and characterization of a Chinese hamster ovary cell mutant with altered regulation of phosphatidylserine biosynthesis.
复制标题
磷脂酰丝氨酸生物合成调节发生改变的中国仓鼠卵巢细胞突变体的分离和表征。
DOI：
10.1016/s0021-9258(19)47193-7
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发表时间：
1989
期刊：
影响因子：
--
通讯作者：
Y. Akamatsu
中科院分区：
文献类型：
--
作者：
K. Hasegawa;O. Kuge;M. Nishijima;Y. Akamatsu
我们筛选了约10，000个固定在聚酯布上的中国仓鼠卵巢（CHO）细胞菌落，以寻找[14 C]乙醇胺掺入三氯乙酸可沉淀磷脂中的缺陷突变体。在以这种方式发现的突变体29中，参与CDP-乙醇胺途径的酶的活性是正常的;然而，磷酸乙醇胺的细胞内池升高，是亲本CHO-K1细胞的10倍以上。这些结果表明，在突变体29中，[14 C]乙醇胺掺入磷脂酰乙醇胺的减少是由于磷酸-[14 C]乙醇胺随着细胞磷酸乙醇胺量的增加而稀释。有趣的是，与亲本细胞相比，突变体29细胞中丝氨酸掺入磷脂酰丝氨酸的速率和磷脂酰丝氨酸的含量分别增加了3倍和1.5倍。突变体29细胞中磷酸乙醇胺的过量产生归因于磷脂酰丝氨酸生物合成水平的升高，因为乙醇胺是作为磷脂酰乙醇胺转化为磷脂酰丝氨酸的反应产物产生的，磷脂酰丝氨酸的转化是由磷脂-丝氨酸碱基交换酶催化的。使用完整的细胞和细胞提取物的颗粒级分，在CHO-K1细胞中的磷脂酰丝氨酸生物合成被磷脂酰丝氨酸本身所抑制，而在突变体29细胞中的磷脂酰丝氨酸生物合成与亲本细胞相比对抑制具有很大的抗性。作为结论，可以假设突变体29细胞具有通过丝氨酸交换酶活性调节磷脂酰丝氨酸生物合成的损伤，这也导致磷脂酰丝氨酸和磷酸乙醇胺的过量产生。
We have screened approximately 10,000 colonies of Chinese hamster ovary (CHO) cells immobilized on polyester cloth for mutants defective in [14C]ethanolamine incorporation into trichloroacetic acid-precipitable phospholipids. In mutant 29, discovered in this way, the activities of enzymes involved in the CDP-ethanolamine pathway were normal; however, the intracellular pool of phosphorylethanolamine was elevated, being more than 10-fold that in the parental CHO-K1 cells. These results suggested that the reduced incorporation of [14C]ethanolamine into phosphatidylethanolamine in mutant 29 was due to dilution of phosphoryl-[14C]ethanolamine with the increased amount of cellular phosphorylethanolamine. Interestingly, the rate of incorporation of serine into phosphatidylserine and the content of phosphatidylserine in mutant 29 cells were increased 3-fold and 1.5-fold, respectively, compared with the parent cells. The overproduction of phosphorylethanolamine in mutant 29 cells was ascribed to the elevated level of phosphatidylserine biosynthesis, because ethanolamine is produced as a reaction product on the conversion of phosphatidylethanolamine to phosphatidylserine, which is catalyzed by phospholipid-serine base-exchange enzymes. Using both intact cells and the particulate fraction of a cell extract, phosphatidylserine biosynthesis in CHO-K1 cells was shown to be inhibited by phosphatidylserine itself, whereas that in mutant 29 cells was greatly resistant to the inhibition, compared with the parental cells. As a conclusion, it may be assumed that mutant 29 cells have a lesion in the regulation of phosphatidylserine biosynthesis by serine-exchange enzyme activity, which results in the overproduction of phosphatidylserine and phosphorylethanolamine as well.