Fig. 3.

eIF2α phosphorylation represses adipogenesis. (a) Protein expression profile in a stable 3T3-L1 cell line that expresses chimeric Fv2E-PERK. Cell lysates were collected at the indicated times after AP20187 treatment (5 nmol/l) for western blot analysis (* indicates a non-specific band). (b) Oil red O staining. Adipogenesis was stimulated in 3T3-L1 cells expressing Fv2E-PERK in the absence (vehicle) or presence of AP20187. On day 10, cells were fixed and stained with Oil red O. Representative images from indicated cell lines are shown. Scale bar indicates 100 μm. (c) Quantification of Oil red O staining in (b). (d–g) Gene expression profiles for markers of mature adipocytes in Fv2E-PERK 3T3-L1 cells in the absence or presence of AP20187 (5 nmol/l). (h) Oil red O staining of Eif2a^S/S and Eif2a^A/A MEFs after adipogenesis. Differentiation was induced in primary MEFs from wild-type (Eif2a^S/S) or homozygous mutant (Eif2a^A/A) for 14 days. Cells were then fixed and stained with Oil red O. Representative images are shown and scale bar indicates 100 μm. (i) Quantification of Oil red O staining in (h). (j–n) Gene expression profiles during adipocyte differentiation of primary MEFs. Total RNA was isolated from cells at the indicated times during adipogenesis, and mRNA levels were measured by quantitative real-time RT-PCR. Circles, Eif2a^S/S; squares, Eif2a^A/A. D0, D2, D4, D7, D10 indicate day 0, day 2, day 4, day 7 and day 10 after initiation of adipocyte differentiation. Data are presented as means±SEM of three independent experiments with triplicates. **p<0.01, *p<0.05 vs Eif2a^A/A. AU, arbitrary units