Liver Clock Protein BMAL1 Promotes de Novo Lipogenesis through Insulin-mTORC2-AKT Signaling

The clock protein BMAL1 (brain and muscle Arnt-like protein 1) participates in circadian regulation of lipid metabolism, but its contribution to insulin AKT-regulated hepatic lipid synthesis is unclear. Here we used both Bmal1^−/− and acute liver-specific Bmal1-depleted mice to study the role of BMAL1 in refeeding-induced de novo lipogenesis in the liver. Both global deficiency and acute hepatic depletion of Bmal1 reduced lipogenic gene expression in the liver upon refeeding. Conversely, Bmal1 overexpression in mouse liver by adenovirus was sufficient to elevate the levels of mRNA of lipogenic enzymes. Bmal1^−/− primary mouse hepatocytes displayed decreased levels of de novo lipogenesis and lipogenic enzymes, supporting the notion that BMAL1 regulates lipid synthesis in hepatocytes in a cell-autonomous manner. Both refed mouse liver and insulin-treated primary mouse hepatocytes showed impaired AKT activation in the case of either Bmal1 deficiency or Bmal1 depletion by adenoviral shRNA. Restoring AKT activity by a constitutively active mutant of AKT nearly normalized de novo lipogenesis in Bmal1^−/− hepatocytes. Finally, Bmal1 deficiency or knockdown decreased the protein abundance of RICTOR, the key component of the mTORC2 complex, without affecting the gene expression of key factors of insulin signaling. Thus, our study uncovered a novel metabolic function of hepatic BMAL1 that promotes de novo lipogenesis via the insulin-mTORC2-AKT signaling during refeeding.     

Circadian clock rhythms in different adipose tissue model systems

ABSTRACT

Circadian clock-controlled 24-h oscillations in adipose tissues play an important role in the regulation of energy homeostasis, thus representing a potential drug target for prevention and therapy of metabolic diseases. For pharmacological screens, scalable adipose model systems are needed that largely recapitulate clock properties observed in vivo. In this study, we compared molecular circadian clock regulation in different ex vivo and in vitro models derived from murine adipose tissues. Explant cultures from three different adipose depots of PER2::LUC circadian reporter mice revealed stable and comparable rhythms of luminescence ex vivo. Likewise, primary pre- and mature adipocytes from these mice displayed stable luminescence rhythms, but with strong damping in mature adipocytes. Stable circadian periods were also observed using Bmal1-luc and Per2-luc reporters after lentiviral transduction of wild-type pre-adipocytes. SV40 immortalized adipocytes of murine brown, subcutaneous and epididymal adipose tissue origin showed rhythmic mRNA expression of the core clock genes Bmal1, Per2, Dbp and REV-erbα in pre- and mature adipocytes, with a maturation-associated increase in overall mRNA levels and amplitudes. A comparison of clock gene mRNA rhythm phases revealed specific changes between in vivo and ex vivo conditions. In summary, our data indicate that adipose culture systems to a large extent mimic in vivo tissue clock regulation. Thus, both explant and cell systems may be useful tools for large-scale screens for adipose clock regulating factors.     

Clock-controlled mir-142-3p can target its activator, Bmal1

ABSTRACT: BACKGROUND: microRNAs (miRNAs) are shown to be involved in the regulation of circadian clock. However, it remains largely unknown whether miRNAs can regulate the core clock genes (Clock and Bmal1). RESULTS: In this study, we found that mir-142-3p directly targeted the 3'UTR of human BMAL1 and mouse Bmal1. The over-expression (in 293ET and NIH3T3 cells) and knockdown (in U87MG cells) of mir-142-3p reduced and up-regulated the Bmal1/BMAL1 mRNA and protein levels, respectively. Moreover, the expression level of mir-142-3p oscillated in serum-shocked NIH3T3 cells and the results of ChIP and luciferase reporter assays suggested that the expression of mir-142-3p was directly controlled by CLOCK/BMAL1 heterodimers in NIH3T3 cells. CONCLUSIONS: Our study demonstrates that mir-142-3p can directly target the 3'UTR of Bmal1. In addition, the expression of mir-142-3p is controlled by CLOCK/BMAL1 heterodimers, suggesting a potential negative feedback loop consisting of the miRNAs and the core clock genes. These findings open new perspective for studying the molecular mechanism of circadian clock.     

High‐fat Diet Followed by Fasting Disrupts Circadian Expression of Adiponectin Signaling Pathway in Muscle and Adipose Tissue

The circadian clock controls energy homeostasis by regulating circadian expression of proteins involved in metabolism. Disruption of circadian rhythms leads to obesity and metabolic disorders. Little is known regarding the control of the biological clock over adiponectin signaling pathway in adipose tissue, the adiponectin producer, and muscle, an adiponectin target tissue under fasting, low‐fat (LF), or high‐fat (HF) diet. Mice were fed LF or HF diet for 7 weeks and fasted on the last day. The circadian mRNA expression of clock genes and components of adiponectin metabolic pathway (mAdipoR1, mAdipoR2, mPparα, mPparγ, mAmpk, and mAcc) in the muscle and adipose tissue were tested. Using average daily levels of multiple time points around the circadian cycle, we assessed mRNA levels of the different adiponectin signaling components. In addition, serum glucose, adiponectin, and insulin were measured. Under LF diet, adiponectin signaling pathway components exhibited circadian rhythmicity at the mRNA levels. Fasting and HF diet followed by fasting disrupted this circadian expression causing a phase advance or delay, respectively. Changes were also found in the expression levels of adiponectin receptor, mAmpk, mAcc, mPparα, and mPparγ reflecting a defect in adiponectin signaling. As both peroxisome proliferator‐activated receptor α (PPARα) and mAMPK are linked to the core clock mechanism, they could mediate the disruptions seen in clock gene expression under HF diet. In turn, the circadian clock affects the daily rhythm of these adiponectin signaling components.