PtdCho synthesis in CHO58 cells expressing CCT-GFP mutants. CHO58 cells were transfected with CCT-GFP or the indicated mutants as described in the legend to Fig. 2. After 2 h at 42°C, cells were incubated with choline-free DMEM containing [3H]choline (2 μCi/ml) for 1 h, followed by treatment without (grey bars) or with (black bars) 300 μM oleate/complexed with BSA for 1 h. [3H]PtdCho was measured and normalized to expression of individual CCT-GFP proteins by immunoblotting of whole-cell lysates, and expressed relative to synthesis in unstimulated CHO58 cells transfected with CCT-GFP. Background [3H]PtdCho synthesis in cells transfected with pEGFP was subtracted. Total PtdCho synthesis was 1,045 and 910 dpm/dish in control and oleate-treated pEGFP-transfected cells, respectively. Total PtdCho synthesis was 13,655 and 36,550 dpm/dish in control and oleate-treated CCT-GFP-expressing cells, respectively. *P < 0.05 compared with corresponding unstimulated control. Results are the mean and SEM of three separate experiments.