Supplementary Figure 4: Inhibition of eIF4E phosphorylation decreases amplitudes of PER oscillations

(a) and (b) Temporal profiles of PER1(a) and PER2(b) in the SCN over 24 h. Representative microscopic images of SCN sections immunostained for PER1 (a) or PER2 (b) are shown. Quantitation of the staining intensity is shown in Figure 4a,b. For these experiments, animals were entrained for 14 d and dark-adapted for 2 d. On the 3rd day in DD, mice were sacrificed and brains were harvested every 4 h throughout 24 h. SCN sections were stained for PER1 or PER2. Note that the levels of PER1/2 proteins in the SCN exhibited circadian oscillations and reached a peak at around CT14 in both the WT and KI mice. However, PER1/2 levels at CT14 were decreased in the SCN of KI mice as compared to the WT mice. Scale bars: 100 μm. (c) Representative plots of mPER2::Luc bioluminescence patterns of SCN explants. Each plot was from one SCN explant. The period of the plots are quantified in (e). *P<0.05 (d) Representative plots of mPER2::Luc bioluminescence patterns of SCN explants before and after application of the MNK1 inhibitor CGP57380 (10 µM). Note that CGP57380 significantly decreased the amplitude of the mPER2::Luc rhythms in the WT but not in the KI SCN explant. Quantitation of the period is shown in (e) and the amplitude ratios (with CGP/without CGP) are shown in (f). Nine to thirteen animals were used for each group. *P<0.05 vs.WT