A Genetic Defect in Phosphatidylcholine Biosynthesis Triggers Apoptosis in Chinese Hamster Ovary Cells

Abstract

We have investigated the cell death of a Chinese hamster ovary mutant (MT-58) with a thermo-sensitive CTP:phosphocholine cytidylyltransferase, the rate-limiting enzyme of the CDP-choline pathway for phosphatidylcholine biosynthesis (Esko, J. D., Wermuth, M. M., and Raetz, C. R. H. (1981) J. Biol. Chem. 256, 7388-7393). After MT-58 cells were shifted to the restrictive temperature of 40 degrees C, the cytidylyltransferase was inactivated immediately leading to a decrease in phosphatidylcholine biosynthesis and cell death. DNA content and number of cells in the S phase decreased significantly in the dying MT-58 cells according to flow cytometrical analyses. The fragmentation of genomic DNA was detected by DNA ladders in agarose gel and release of the prelabeled genomic DNA into cytosolic fractions 14 h after the temperature shift. The dying cells underwent a dramatic reduction of cellular volume while maintaining the membrane containment of cellular contents. These events indicated that the inactivation of cytidylyltransferase triggered apoptosis in Chinese hamster ovary cells. This is the first report that apoptosis was induced in cultured cells, not by an added agent, but by a mutation in phospholipid biosynthesis.

The follicular microenviroment as a predictor of pregnancy: MALDI-TOF MS lipid profile in cumulus cells

“…Therefore, PC is important for maintaining cellular integrity and cell growth. Furthermore, several studies have clearly demonstrated that PC deficiency alone may induce apoptosis [8]. Although we identified PC only in the pregnant group, as shown in Table 2, this result does not mean that it is absent in the non-pregnant group.…”

Choline kinase inhibition induces the increase in ceramides resulting in a highly specific and selective cytotoxic antitumoral strategy as a potential mechanism of action

“…An alternative pathway to generate PC implies the methylation of PE by phosphatidylethanolamine-N-methyl-transferase (PEMT) (Vance et al, 1997). However, this pathway is not relevant in mammalian cells as demonstrated by restoration of PC synthesis by PEMT transfection experiments in cells rendered temperature-sensitive mutant for CT (MT58 cell line) (Esko et al, 1981(Esko et al, , 1982Houweling et al, 1995;Cui et al, 1996). The inability of PEMT to rescue MT58 cells under restrictive temperature from apoptosis suggests that the PC species generated from the Kennedy pathway and the PEMT pathway are functionally different (Cui and Houweling, 2002).…”

Lipid metabolic changes caused by short-chain ceramides and the connection with apoptosis

“…Indeed, recent work shows that defective synthesis of PtdCho is by itself sufficient to cause apoptosis ; thus mutant Chinesehamster ovary (' CHO ') cells possessing a temperature-sensitive cytidylyltransferase undergo apoptosis when exposed to temperatures which inactivate this enzyme [28], and HeLa cells or HL60 cells are rescued from apoptosis caused by inhibitors of PtdCho synthesis if cytidylyltransferase is up-regulated [26] or if lysoPtdCho is added as an alternative source of PtdCho [27]. There is even a possibility that C # -ceramide as a natural metabolite could affect normal lipid synthesis, since at least in HL60 cells, endogenous C # -ceramide can be formed from platelet-activating factor [29].…”