Feng D, Liu T, Sun Z, Bugge A, Mullican SE, Alenghat T, Liu XS, Lazar MA. A circadian rhythm orchestrated by histone deacetylase 3 controls hepatic lipid metabolism. Science 2011;331:1315-1319. (Reprinted with permission.)
Disruption of the circadian clock exacerbates metabolic diseases, including obesity and diabetes. We show that histone deacetylase 3 (HDAC3) recruitment to the genome displays a circadian rhythm in mouse liver. Histone acetylation is inversely related to HDAC3 binding, and this rhythm is lost when HDAC3 is absent. Although amounts of HDAC3 are constant, its genomic recruitment in liver corresponds to the expression pattern of the circadian nuclear receptor Rev-erbα. Rev-erbα colocalizes with HDAC3 near genes regulating lipid metabolism, and deletion of HDAC3 or Rev-erbα in mouse liver causes hepatic steatosis. Thus, genomic recruitment of HDAC3 by Rev-erbα directs a circadian rhythm of histone acetylation and gene expression required for normal hepatic lipid homeostasis.
Circadian rhythms are responsible for daily variations in organ-specific functions and are essential in coordinating the timing of various physiological processes. Also, the gastrointestinal tract including the liver is subject to circadian rhythms, and a large number of genes involved in the maintenances of metabolic homeostasis is rhythmically expressed in the liver,1 suggesting that circadian and metabolic regulatory networks are tightly connected. Circadian misalignment causes metabolic dysfunction, and mice with genetic disruption of circadian clock components develop hyperlipidemia, hyperglycemia, hypoinsulinemia, as well as hepatic steatosis.2, 3
The nuclear receptor Rev-erbα is a key regulator of the circadian rhythm and is expressed in a circadian manner.4 Rev-erbα is a transcriptional repressor of critical regulators of the circadian rhythms, and it is supposed that the circadian clock regulates metabolism mostly by regulating the expression of liver enzymes at the transcriptional level. Epigenetic alterations, such as hyperacetylation of the chromatin-associated histones, which is responsible for gene silencing, are critical regulators of gene transcription, involving multiple histone acetyltransferases and deacetylases (HDACs).
A recent report in Science demonstrates the existence of circadian changes in histone acetylation in mice, the dysregulation of which potentially causes major perturbations in normal metabolic functions and may also significant affect the development and progression of nonalcoholic fatty liver disease (NAFLD) in men.5
Feng et al. discovered diurnal recruitment of HDAC3 to the liver genome of mice. In the light period, when the mice are inactive, HDAC bound to over 14,000 sites, whereas in the dark period when mice are active and feeding the binding markedly reduced to only 120 sites. This HDAC3 recruitment pattern oscillated in a 24-hour cycle. Deletion of hepatic HDAC3 expression led to similar acetylation levels of histone H3 lysine 9 (H3K9) during the inactive time as observed in control mice during their activity period, indicating that the circadian clock is the pacemaker for the genomic HDAC3 recruitment. Associated with the observed decrease in H3K9 acetylation in mice with hepatic HDAC3 deletion, the authors found a decrease in polymerase II at the transcription start site of genes with HDAC3 binding sites and a reduced expression of these genes, respectively. Thus, diurnal recruitment of HDAC3 orchestrates a rhythm of epigenomic modification, polymerase II recruitment, and gene expression. Although the HDAC3 recruitment to the genome is diurnal, the abundance of HDAC3 was constant throughout the light/dark cycle. HDAC3 enzymatic activity requires interaction with nuclear receptor corepressors, and Feng et al. discovered that Rev-erbα protein oscillated in phase with HDAC3 recruitment (Fig. 1A), and remarkably, Rev-erbα bound to the majority of HDAC3 binding sites during the inactive period but not during the active period of the mice (Fig. 1B). The extent of HDAC3 with Rev-erbα binding was surprising because other nuclear receptors can also interact with corepressors and HDAC3. However, HDAC3 binding was reduced at many sites in Rev-erbα-deficient mice, consistent with a critical role of Rev-erbα. Still, residual HDAC3 binding sites in Rev-erbα-deficient mice reveal that other factors also contribute to HDAC3 recruitment. Of note, the set of genes bound by Rev-erbα and HDAC3 was enriched for genes encoding for proteins that function in lipid metabolic processes, and indeed, livers in which HDAC3 was deleted revealed a significant increase of neutral lipid content. In accord, chow fed Rev-erbα-deficient mice also developed liver steatosis, and the majority of genes up-regulated in livers depleted of Rev-erbα were bound by both Rev-erbα and HDAC3 during the sleeping period of the mice. At that time HDAC3 and Rev-erbα colocalized at more than 100 lipid biosynthetic genes and polymerase II recruitment to the transcription start site of many of these genes increased, when the mice were active and ate. These findings suggest that biosynthesis was actively suppressed, and indeed, Rev-erbα- and HDAC3-deficient mice revealed increased de novo biosynthesis of lipids (Fig. 1C). Thus, this fascinating report provides a molecular mechanism underlying the observation that hepatic lipogenesis in mice follows a diurnal rhythm that is antiphase to Rev-erbα and HDAC3 recruitment to the genome. HDAC3 was already known as a critical regulator of circadian rhythm and glucose metabolism,6 and liver-specific deletion of HDAC3 has been described to cause fatty liver in mice.7 The present report newly connects HDAC3 with the circadian rhythm and impressively demonstrates that not its abundance but its rhythmic recruitment to the genome in concert with Rev-erbα critically affects transcriptional regulation of hepatic lipid metabolisms. The significance of daily variations in hepatic gene expression is still not fully determined but may be related to different requirements of nutrient absorption, energy generation, and energy storage during the feeding and fasting state. In general, the suprachiasmatic nucleus harbors the central pacemaker of the circadian rhythm in mammals, but circadian oscillators exist in most peripheral tissues including the liver. Rats exposed to a light/dark cycle regimen mimicking shift-work during a period of 10 weeks revealed significantly changed hepatic lipid metabolism, including and noteworthy also, Rev-erbα expression.8 In the study by Feng et al. the HDAC3 recruitment pattern to the liver genome was retained in constant darkness, whereas the rhythm of HDAC recruitment to the genome was quickly reversed when food was provided only during the inactive, sleeping period. Because the liver clock is entrained by food intake, these findings indicate that the “hepatic” circadian clock is the pacemaker for the genomic HDAC3 recruitment. It has been shown that temporal feeding restriction under light/dark or dark/dark conditions can change the phase of circadian gene expression in peripheral cell types by up to 12 hours, while leaving the phase of cyclic gene expression in the suprachiasmatic nucleus unaffected.9 Hence, changes in metabolism can lead to an uncoupling of peripheral oscillators from the central pacemaker, and misalignment of fasting/feeding and sleep/wake cycles with endogenous circadian cycles of hepatic fuel utilization or energy storage cause hepatic steatosis. The liver seems to be prone to such a misalignment because food-induced phase resetting proceeds faster in liver than in other organs such as kidney, heart, or pancreas.8 What may be the pathopyhsiological significance of such an imbalance? Feng et al. describe only modestly elevated hepatic transaminases in HDAC-deficient mice, but this was probably due to the short observation time after induced HDAC3 depletion, because a previous study found progressive hepatocellular damage in HDAC-deficient mice with time.7 Also, experimentally induced disruption of the circadian rhythm led to an abolished rhythm in the expression of both central clock as well as hepatic clock genes and caused an altered innate immune response with heightened release of proinflammatory cytokines in response to lipopolysaccharide (LPS) treatment.10 Recent studies revealed the crucial role of innate immunity in the progression of (nonalcoholic) steatosis to (nonalcoholic) steatohepatitis (NASH).11 Together, these studies suggest that disruption of the circadian rhythm affects not only hepatic (lipid) metabolisms but subsequently triggers the progression of NASH. In line with this, genetic variants of molecular clock genes have been identified as risk factors for the development of NAFLD,12 rotating shift work increases the risk for developing the metabolic syndrome,13 and interestingly, this appears to be particularly the case in individuals with elevated alanine aminotransferase serum levels.14 Moreover, circadian disruption was found to accelerate liver carcinogenesis in mice, further suggesting that the tight and proper control of circadian clocks is a prerequisite of hepatic integrity.15
Thus, liver steatosis may be one of the myriad negative health effects of shift work, and, certainly, not only from the hepatologist's perspective should this be avoided. Still, if this is not feasible it seems mandatory to avoid or at least minimize the misalignment of the circadian and the hepatic clock. Of note, balanced diets containing carbohydrates/sugars and proteins were shown to be necessary for proper entrainment of the liver clock in mice.16 Future studies have to show whether these findings may assist in the development of dietary recommendations for shift workers. In addition to the quality and quantity of food, not only for shift-workers and jet-lagged air travelers, the time of food consumption may be a risk factor for fatty liver.