Low density lipoprotein receptor (LDLR) plays an important role in the cholesterol homeostasis. We examined the possible circadian regulation of LDLR and mechanism(s) underlying it. In mice, blood glucose and plasma triglyceride, total and high density lipoprotein cholesterol varied distinctively throughout a day. In addition, LDLR mRNA oscillated in the liver in a functional clock-dependent manner. Accordingly, analysis of human LDLR promoter sequence revealed three putative E-boxes, raising the possible regulation of LDLR expression by E-box-binding transcription factors. To test this possibility, human LDLR promoter reporter constructs were transfected into HepG2 cells and the effects of CLOCK/BMAL1, Hes1, and Hes6 expression were analyzed. It was found that positive circadian transcription factor complex CLOCK/BMAL1 upregulated human LDLR promoter activity in a serum-independent manner, while Hes family members Hes1 and Hes6 downregulated it only under serum-depleted conditions. Both effects were mapped to proximal promoter region of human LDLR, where mutation or deletion of well-known sterol regulatory element (SRE) abolished only the repressive effect of Hes1. Interestingly, hes6 and hes1 mRNA oscillated in an anti-phasic manner in the wild-type but not in the per1-/-per2-/- mouse. Comparative analysis of mouse, rat and human hes6 genes revealed that three E-boxes are conserved among three species. Transfection and site-directed mutagenesis studies with hes6 reporter constructs confirmed that the third E-box in the exon IV is functionally induced by CLOCK/BMAL1. Taken together, these results suggest that LDLR expression is under circadian control involving CLOCK/BMAL1 and Hes family members Hes1 and Hes6.

(2013) Adrenal peripheral clock disruption leads to altered circadian behavioral responses to voluntary exercise in middle-aged female mice, Animal Cells and Systems, 17:6, 397-405, DOI: 10.1080/19768354.2013 We recently established an adrenal clock-disrupted transgenic mouse line (BMAS) that exhibits a dampened rhythm of corticosterone secretion and reduced amplitude of day/night activity. Here, we observe that voluntary wheel running increases the robustness and amplitude of both body temperature and home cage activity (HCA) rhythms in wild-type, but not in BMAS mice, but without affecting estrous cycle. Surprisingly, wheel running alters the HCA waveform of BMAS females in a way that preferentially increases the late nighttime (ZT21-ZT24) HCA. These results indicate that adrenal clock disruption causes the animals to respond differently to the voluntary exercise cue in middle-aged female mice.