FIGURE 5.

Gcn2-CTD binds eEF1A independently of ribosomes. A, various amounts of WCE, and of the supernatant of WCE obtained from a high velocity centrifugation (PRS), were subjected to SDS-PAGE and immunoblotting using antibodies against eEF1A and RPS22 to verify that the amount of ribosomes was significantly reduced in the PRS. The top panel shows Ponceau S staining of the immunoblotting membrane. 1 x stands for the amount of total protein used (25 μg) in co-precipitation assays in B. B, GST-Gcn2-CTD, or GST alone as control, were subjected to co-precipitation assays using PRS or WCE (25 μg of total protein) from A, and then subjected to immunoblotting as described in Fig. 3.