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. 2010 Jun 29;29(15):2501–2514. doi: 10.1038/emboj.2010.141

Figure 5.

Figure 5

CHIP is required for the inhibition of sorbitol-induced sustained activation of ERK. (A) Function of MEKK2 and CHIP in sorbitol-induced activation of ERK pathway. After transfection with the indicated combination of control, MEKK2 and CHIP siRNAs for 24 h, HEK293 cells were transfected with pFA-Elk1, pFR-Luc and pSV7d-βGal for 24 h. Cells were transiently treated with 500 mM sorbitol for 1 h. After 7 h, the Elk1-promoter luciferase activities were determined as described in Materials and methods. The luciferase activities relative to the β-galactosidase activities are shown as fold increase compared with control cells (n=3, mean±s.e., *P<0.05, **P<0.01). (B, C) Requirement of CHIP for the inhibition of sorbitol-induced ERK activation at a late phase. HEK293 cells transfected with siRNAs against control, CHIP#1 or CHIP#2 for 48 h were treated with 500 mM sorbitol for 1 h. Cell lysates were subjected to immunoblotting with indicated antibodies. The bottom schema represents the experimental time schedules (B). Sorbitol-induced ERK, JNK and p38 phosphorylation levels relative to the amount of total proteins are shown as fold increase compared with non-treated cells (0 h) (n=3, mean±s.e., **P<0.01) (C).