FIGURE 2.

Multiple sites of Raptor are phosphorylated in response to osmotic stress. A, the MS/MS spectrum of the peptide with phospho-Ser-696 and phospho-Thr-706. B, the MS/MS spectrum of the peptide with phospho-Ser-863. C, a schematic representation of the Raptor domains and phosphorylated sites (P) induced by osmotic stress. D, FLAG-Raptor WT or S696A/T706A/S863A mutant (3A) was expressed and exposed to 0.5 m sorbitol for 60 min. The level of phosphorylation was determined by the mobility shift in SDS-PAGE or phospho-specific antibodies. p-Raptor, phospho-Raptor. E, lysates from Fig. 1B were subjected to immunoblotting with the indicated phospho-Raptor antibodies.