Research Paper Volume 13, Issue 8 pp 10989—11009

Figure 2. Phosphorylation of eIF2α plays a key role in protein translation. (A) Metabolic labelling by ³⁵S-methionine/cysteine incorporation in WT and eIF2α.S52A cells grown in YES medium. Cells were cultivated to exponential growth phase (time 0) or later time points, as indicated, and labelled for 10 min prior to harvesting. Labelled proteins present in cell extracts were visualized by autoradiography after SDS-PAGE separation (top panel). Coomassie staining shows the total protein levels loaded in each lane (bottom panel). (B) Polysome profile analysis of WT and eIF2α.S52A cells. Cell extracts were separated in 7-50 % sucrose gradients and fractioned as described in the Materials and Methods. Polysome/monosome ratios (P/M) were quantified and are shown for each profile. Values are expressed as %, considering the 100% value for WT cells at time = 0, and represent the mean value of three independent experiments. Data information: (A, B) Representative results from at least three independent experiments.