Figure 3: MYC directly represses clock genes via MIZ1. | Nature Communications

Figure 3: MYC directly represses clock genes via MIZ1.

From: MYC/MIZ1-dependent gene repression inversely coordinates the circadian clock with cell cycle and proliferation

Figure 3

(a) Kinetics of MYC-dependent repression of p21-luc, p15-luc and Bmal1-luc and activation of 6xEbox-luc (n=3). Unsynchronized U2OS t-rex tetO-MYC:V5 cells were transiently transfected with the indicated reporter plasmids. At time point 0 MYC:V5 expression was either induced with doxycycline (MYC ox) or not induced with PBS (Ctrl). (b) Left panel: schematic of MIZ1 binding sites in NPAS2, CLOCK and BMAL1 based on data from Walz et al.14. Black triangles indicate regions amplified in ChIP-PCR analysis. Right panel: ChIP-PCR analysis showing recruitment of induced MYC:V5 to MIZ1 binding sites in NPAS2, CLOCK and BMAL1 at 24 and 36 h after synchronization of U2OS t-rex tetO-MYC:V5 cells (n=3). (c) Downregulation of MIZ1 with siRNA reduces expression of NPAS2, CLOCK and BMAL1 in U2OS cells (n=3). Transcript levels of the indicated genes were determined by quantitative PCR (qPCR). (d) MYC-induced attenuation of the circadian clock is rescued by downregulation of MIZ1 (n=3). Synchronized U2OS t-rex tetO-MYC:V5 Bmal1-luc cells were transfected with MIZ1 or control siRNAs. Note that downregulation of MIZ1 attenuates the circadian rhythm of Bmal1-luc and lengthens the period (Supplementary Fig. 3a,b). Data are presented as mean±s.e.m. *P<0.05; Student’s t-test.

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