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. Author manuscript; available in PMC: 2016 Aug 4.
Published in final edited form as: Cell Rep. 2015 Feb 7;10(5):684–693. doi: 10.1016/j.celrep.2015.01.014

Figure 3. Rheb induces P- eIF2α independent of mTORC1 and promotes P-eIF2α predominantly through PERK.

Figure 3

(A) Western blotting of indicated proteins of HEK293 expressing cDNAs of myc (control) or myc-Rheb WT were grown in DMEM with serum (+) or without serum (–) or pretreated with the inhibitors rapamycin (250 nM) or DMSO (0.5%, control). (B) Western blotting of indicated proteins in HEK293 cells expressing myc or myc-Rheb and pretreated with rapamycin or DMSO as in A (C). Western blotting of indicated proteins in HEK293 cells expressing myc or myc-Rheb and pretreated with inhibitors of MAPK [PD98059, 50 µM] or PI3K [wortmannin, 100 nM], rapamycin or DMSO. (D) Western blotting of indicated proteins in GCN2−/− cells and their isogeneic wild-type (+/+) MEFs were infected with adenoviral-null or adenoviral-Rheb. (E) Autoradiography of [35S]-Met incorporation and total proteins (ponceu) in GCN2 MEF cells infected with adenoviral-null or adenoviral-Rheb (F) Western blotting of indicated proteins in PERK−/− cells and their isogeneic wild-type (+/+) MEFs were infected with adenoviral-null or adenoviral-Rheb. GTPase Rheb fails to induce P-eIF2α in PERK−/− MEF cells. (G) Autoradiography of [35S]-Met incorporation and total proteins (ponceu) in PERK MEF cells infected with adenoviral-null or adenoviral-Rheb (***p<0.001: **p<0.01: *p<0.05, Student t test). Data are means ± SEM from 3 experiments.