Isolation and characterization of cultured mammalian cell mutants defective in phospholipid biosynthesis

(1)克隆了两个补充PSA-3突变的CHO细胞基因，PSA-3是CHO细胞的磷脂酰丝氨酸营养缺陷型突变体。(2)我们检测了膜磷脂修饰对Sindbis病毒在PSA-3细胞中增殖的影响，发现当PSA-3细胞在没有磷脂酰丝氨酸的情况下生长时，病毒的结合和内化正常发生，但病毒粒子的产量和病毒RNA的合成大大减少。这些结果表明，细胞内的磷脂酰丝氨酸和/或磷脂酰乙醇胺参与了Sindbis病毒感染的某个步骤，在病毒颗粒内化之后，病毒核衣壳进入细胞质之前。(3)获得了一株在调节磷脂酰丝氨酸生物合成方面存在缺陷的CHO细胞突变体(#29)。在该突变细胞中，与亲本细胞相比，磷脂酰丝氨酸的双合成增加了约两倍，并且显著地抵抗了外源磷脂酰丝氨酸的抑制。(4)我们发现CHO细胞有两种肌醇转运系统，即钠依赖和非钠依赖系统。我们获得了钠依赖肌醇转运系统缺陷的CHO细胞突变体。(5)分离到葡萄糖转运缺陷的CHO细胞突变体。其中一个突变体(GTS-31)的葡萄糖转运蛋白水平比亲本细胞降低了80%。

(1) We cloned two CHO cell genes that complement the mutation of PSA-3, a phosphatidylserine auxotrophic mutant of CHO cell line. (2) We examined the effects of the modification of membrane phospholipids on the proliferation of the sindbis virus in PSA-3 cells and found that when PSA-3 cells are grown without phosphatidylserine, the binding and internalization of the virus occur normally but the yield of virions and viral RNA synthesis greatly decreased. These results indicate that cellular phosphatidylserine and/or phosphatidylethanolamine participate in a certain step of sindbis virus infection, after the internalization of virus particles, but before penetration of the viral nucleocapsids into the cytoplasm. (3) We obtained a CHO cell mutant (# 29) which is defective in the regulation of phosphatidylserine biosynthesis. In this mutant cells, phosphatidylserine bisynthesis was elevated about two-fold and was remarkably resistant to the inhibition by exogenous phosphatidylserine as compared to the parental cells. (4) We found that CHO cells have two kinds of inositol transport systems, namely, sodium-dependent and sodium-independent systems. We obtained CHO cell mutants defective in sodium-dependent inositol transport system. (5) We isolated CHO cell mutants defective in glucose transport. In one of the mutants (GTS-31), the level of glucose transporter was reduced by 80% as compared to the parental cells.