RESUMEN

BACKGROUND: While the daily rhythm of allergic rhinitis (AR) has long been recognized, the molecular mechanism underlying this phenomenon remains enigmatic. OBJECTIVE: We aimed to investigate the role of circadian clock in AR development and to clarify the mechanism by which the daily rhythm of AR is generated. METHODS: AR was induced in mice with ovalbumin. Toluidine blue staining, liquid chromatography-tandem mass spectrometry analysis, real-time quantitative PCR, and immunoblotting were performed with AR and control mice. RESULTS: Ovalbumin-induced AR is diurnally rhythmic and associated with clock gene disruption in nasal mucosa. In particular, Rev-erbα is generally downregulated and its rhythm retained, but with a near-12-hour phase shift. Furthermore, global knockout of core clock gene Bmal1 or Rev-erbα increases the susceptibility of mice to AR and blunts AR rhythmicity. Importantly, nasal solitary chemosensory cells (SCCs) are rhythmically activated, and inhibition of the SCC pathway leads to attenuated AR and a loss of its rhythm. Moreover, rhythmic activation of SCCs is accounted for by diurnal expression of ChAT (an enzyme responsible for the synthesis of acetylcholine) and temporal generation of the neurotransmitter acetylcholine. Mechanistically, Rev-erbα trans-represses Chat through direct binding to a specific response element, generating a diurnal oscillation in this target gene. CONCLUSION: SCCs, under the control of Rev-erbα, are a driver of AR rhythmicity; targeting SCCs should be considered as a new avenue for AR management.

Asunto(s)

Ritmo Circadiano , Modelos Animales de Enfermedad , Mucosa Nasal , Rinitis Alérgica , Animales , Rinitis Alérgica/inmunología , Rinitis Alérgica/metabolismo , Ratones , Mucosa Nasal/metabolismo , Mucosa Nasal/inmunología , Mucosa Nasal/patología , Miembro 1 del Grupo D de la Subfamilia 1 de Receptores Nucleares/genética , Miembro 1 del Grupo D de la Subfamilia 1 de Receptores Nucleares/metabolismo , Ratones Noqueados , Ovalbúmina/inmunología , Factores de Transcripción ARNTL/genética , Factores de Transcripción ARNTL/metabolismo , Relojes Circadianos/genética , Ratones Endogámicos BALB C

RESUMEN

With the continuous development of the field of building optimization, more and more optimization methods have sprung up, among which there are many kinds of intelligent optimization algorithms. This kind of intelligent optimization algorithm usually relies on the traditional building performance simulation method to obtain the building performance index for optimization. However, intelligent optimization algorithms generally require large-scale calculations. At the same time, the time required for building performance simulation is often limited by the complexity of building models and the configuration of computers, which leads to a long time for performance optimization, which cannot give efficient and accurate feedback to designers in engineering. Building performance optimization methods based on intelligent optimization algorithms are mainly used in scientific research and are difficult to put into practical projects. Therefore, this paper builds an accurate and efficient platform for building performance prediction and optimization to help designers make decisions combined with BP neural network and the SPEA-II multiobjective optimization algorithm. Besides, the optimization results of the case are quantitatively and qualitatively analyzed and presented in visual form based on the BP neural network prediction model. Quantitative analysis includes the evolution process of solution set, convergence process, and comprehensive quality evaluation of solution set. Qualitative analysis includes Pareto frontier and optimal architectural scheme analysis. Finally, the conclusion shows that the platform prediction and optimization can give accurate and reliable optimal solution, and the optimal building scheme is reasonable and has high engineering application value.

Asunto(s)

Algoritmos , Redes Neurales de la Computación , Simulación por Computador , Ingeniería , Inteligencia

RESUMEN

The role of intestine clock in energy homeostasis remains elusive. Here we show that mice with Bmal1 specifically deleted in the intestine (Bmal1iKO mice) have a normal phenotype on a chow diet. However, on a high-fat diet (HFD), Bmal1iKO mice are protected against development of obesity and related abnormalities such as hyperlipidemia and fatty livers. These metabolic phenotypes are attributed to impaired lipid resynthesis in the intestine and reduced fat secretion. Consistently, wild-type mice fed a HFD during nighttime (with a lower BMAL1 expression) show alleviated obesity compared to mice fed ad libitum. Mechanistic studies uncover that BMAL1 transactivates the Dgat2 gene (encoding the triacylglycerol synthesis enzyme DGAT2) via direct binding to an E-box in the promoter, thereby promoting dietary fat absorption. Supporting these findings, intestinal deficiency of Rev-erbα, a known BMAL1 repressor, enhances dietary fat absorption and exacerbates HFD-induced obesity and comorbidities. Moreover, small-molecule targeting of REV-ERBα/BMAL1 by SR9009 ameliorates HFD-induced obesity in mice. Altogether, intestine clock functions as an accelerator in dietary fat absorption and targeting intestinal BMAL1 may be a promising approach for management of metabolic diseases induced by excess fat intake.

Asunto(s)

Factores de Transcripción ARNTL/genética , Ritmo Circadiano/genética , Diacilglicerol O-Acetiltransferasa/genética , Hígado Graso/genética , Hiperlipidemias/genética , Miembro 1 del Grupo D de la Subfamilia 1 de Receptores Nucleares/genética , Obesidad/genética , Factores de Transcripción ARNTL/deficiencia , Animales , Diacilglicerol O-Acetiltransferasa/metabolismo , Dieta Alta en Grasa/efectos adversos , Grasas de la Dieta/administración & dosificación , Grasas de la Dieta/metabolismo , Hígado Graso/etiología , Hígado Graso/metabolismo , Hígado Graso/prevención & control , Regulación de la Expresión Génica , Homeostasis/efectos de los fármacos , Homeostasis/genética , Hiperlipidemias/etiología , Hiperlipidemias/metabolismo , Hiperlipidemias/prevención & control , Mucosa Intestinal/efectos de los fármacos , Mucosa Intestinal/metabolismo , Metabolismo de los Lípidos/efectos de los fármacos , Metabolismo de los Lípidos/genética , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Miembro 1 del Grupo D de la Subfamilia 1 de Receptores Nucleares/antagonistas & inhibidores , Miembro 1 del Grupo D de la Subfamilia 1 de Receptores Nucleares/metabolismo , Obesidad/etiología , Obesidad/metabolismo , Obesidad/prevención & control , Regiones Promotoras Genéticas , Unión Proteica , Pirrolidinas/farmacología , Transducción de Señal , Tiofenos/farmacología , Triglicéridos/biosíntesis

RESUMEN

Identifying drugs with dosing time-dependent effects (chronoeffects) and understanding the underlying mechanisms would help to improve drug treatment outcome. Here, we aimed to determine chronoeffects of the herbal medicines Puerariae radix (PR) and Coptidis rhizoma (CR), and investigate a potential role of REV-ERBα as a drug target in generating chronoeffects. The pharmacological effect of PR on hyperhomocysteinemia in mice was evaluated by measuring total homocysteine, triglyceride levels and lipid accumulation. PR dosed at ZT10 generated a stronger effect on hyperhomocysteinemia than drug dosed at ZT2. Furthermore, PR increased the expression levels of REV-ERBα target genes Bhmt, Cbs and Cth (encoding three key enzymes responsible for homocysteine catabolism), thereby alleviating hyperhomocysteinemia in mice. Moreover, CR attenuated chronic colitis in mice in a dosing time-dependent manner based on measurements of disease activity index, colon length, malondialdehyde/myeloperoxidase activities and IL-1ß/IL-6 levels. ZT10 dosing generated a stronger anti-colitis effect as compared to ZT2 dosing. This was accompanied by lower production of colonic inflammatory cytokines (i.e., Nlrp3, IL-1ß, IL-6, Tnf-α and Ccl2, REV-ERBα target genes) in colitis mice dosed at ZT10. The diurnal patterns of PR and CR effects were respectively consistent with those of puerarin (a main active constituent of PR, a REV-ERBα antagonist) and berberine (a main active constituent of CR, a REV-ERBα agonist). In addition, loss of Rev-erbα in mice abolished the dosing time-dependency in PR and CR effects. In conclusion, the therapeutic effects of PR and CR depend on dosing time in mice, which are probably attributed to diurnal expression of REV-ERBα as the drug target. Our findings have implications for improving therapeutic outcomes of herbal medicines with a chronotherapeutic approach.

RESUMEN

1. Retrorsine (RTS) is a pyrrolizidine alkaloid (distributed in many medicinal plants) that has significant hepatotoxicity. Here, we aimed to determine the daily variations in RTS hepatotoxicity (chronotoxicity) in mice, and to investigate the role of metabolism in generating RTS chronotoxicity.2. Acute toxicity and pharmacokinetic studies were performed with mice after RTS administration at different times of the day. Hepatotoxicity was assessed by measuring plasma ALT (alanine aminotransferase) and AST (aspartate aminotransferase) levels. mRNA and proteins were determined by qPCR and Western blotting, respectively. Time-dependent in vitro metabolism of RTS was assessed by using mouse liver microsomes.3. We found that RTS toxicity was more severe in the dark phase (zeitgeber time 14 or ZT14 and ZT18) than in the light phase (ZT2 and ZT6). This chronotoxicity was associated with a dosing time difference in the systemic exposures of RTS and a pyrrolic ester metabolite (a cause of hepatotoxicity, measured by the levels of pyrrole-GSH conjugate and pyrrole-protein adducts due to a high chemical reactivity). Moreover, the CYP3A11 (a major enzyme for RTS bioactivation) inhibitor ketoconazole decreased the production of pyrrole-GSH conjugate and abrogated diurnal rhythm in RTS metabolism. In addition, E4bp4 (a circadian regulator of Cyp3a11) ablation abolished the rhythm of CYP3A11 expression and abrogated the dosing time-dependency of RTS toxicity.4. In conclusion, RTS chronotoxicity in mice was attributed to time-varying hepatic metabolism regulated by the circadian clock. Our findings have implications for reducing pyrrolizidine alkaloid-induced toxicity via a chronotherapeutic approach.

Asunto(s)

Relojes Circadianos , Alcaloides de Pirrolicidina , Alanina Transaminasa , Animales , Ritmo Circadiano , Citocromo P-450 CYP3A/genética , Hígado , Proteínas de la Membrana , Ratones , Alcaloides de Pirrolicidina/toxicidad

RESUMEN

To design potentially more effective therapies, we need to further understand the mechanisms underlying epilepsy. Here, we uncover the role of Rev-erbα in circadian regulation of epileptic seizures. We first show up-regulation of REV-ERBα/Rev-erbα in brain tissues from patients with epilepsy and a mouse model. Ablation or pharmacological modulation of Rev-erbα in mice decreases the susceptibility to acute and chronic seizures, and abolishes diurnal rhythmicity in seizure severity, whereas activation of Rev-erbα increases the animal susceptibility. Rev-erbα ablation or antagonism also leads to prolonged spontaneous inhibitory postsynaptic currents and elevated frequency in the mouse hippocampus, indicating enhanced GABAergic signaling. We also identify the transporters Slc6a1 and Slc6a11 as regulators of Rev-erbα-mediated clearance of GABA. Mechanistically, Rev-erbα promotes the expressions of Slc6a1 and Slc6a11 through transcriptional repression of E4bp4. Our findings propose Rev-erbα as a regulator of synaptic function at the crosstalk between pathways regulating the circadian clock and epilepsy.

Asunto(s)

Neuronas GABAérgicas/metabolismo , Regulación de la Expresión Génica , Miembro 1 del Grupo D de la Subfamilia 1 de Receptores Nucleares/genética , Convulsiones/genética , Enfermedad Aguda , Animales , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/genética , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/metabolismo , Enfermedad Crónica , Modelos Animales de Enfermedad , Epilepsia del Lóbulo Temporal/genética , Epilepsia del Lóbulo Temporal/patología , Epilepsia del Lóbulo Temporal/fisiopatología , Proteínas Transportadoras de GABA en la Membrana Plasmática/genética , Proteínas Transportadoras de GABA en la Membrana Plasmática/metabolismo , Regulación de la Expresión Génica/efectos de los fármacos , Hipocampo/patología , Humanos , Potenciales Postsinápticos Inhibidores/efectos de los fármacos , Isoquinolinas/farmacología , Excitación Neurológica/efectos de los fármacos , Ratones Endogámicos C57BL , Ratones Noqueados , Miembro 1 del Grupo D de la Subfamilia 1 de Receptores Nucleares/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Convulsiones/patología , Convulsiones/fisiopatología , Bibliotecas de Moléculas Pequeñas/farmacología , Tiofenos/farmacología , Ácido gamma-Aminobutírico/metabolismo

RESUMEN

BACKGROUND AND PURPOSE: Acute kidney injury (AKI) is a common and critical illness, resulting in severe morbidity and a high mortality. There is a considerable interest in identifying novel molecular targets for management of AKI. We investigated the potential role of the circadian clock components Rev-erb-α/ß in regulation of ferroptosis and AKI. EXPERIMENTAL APPROACH: AKI model was established by treating mice with folic acid. Regulatory effects of Rev-erb-α/ß on AKI and ferroptosis were determined using single-gene knockout (Rev-erb-α-/- and Rev-erb-ß-/- ) mice, incomplete double-knockout (icDKO, Rev-erb-α+/- Rev-erb-ß-/- ) mice and cells with erastin-induced ferroptosis. Targeted antagonism of Rev-erb-α/ß to alleviate AKI and ferroptosis was assessed using the small-molecule antagonist SR8278. Transcriptional gene regulation was investigated using luciferase reporter, mobility shift and chromatin immunoprecipitation assays. KEY RESULTS: Loss of Rev-erb-α or Rev-erb-ß reduced the sensitivity of mice to folic acid-induced AKI and eliminated the circadian time dependency in disease severity. This coincided with less extensive ferroptosis, a main cause of folic acid-induced AKI. Moreover, icDKO mice were more resistant to folic acid-induced AKI and ferroptosis as compared with single-gene knockout mice. Supporting this, targeting Rev-erb-α/ß by SR8278 attenuated ferroptosis to ameliorate folic acid-induced AKI in mice. Rev-erb-α/ß promoted ferroptosis by repressing the transcription of Slc7a11 and HO1 (two ferroptosis-inhibitory genes) via direct binding to a RORE cis-element. CONCLUSION AND IMPLICATIONS: Targeted inhibition of Rev-erb-α/ß limits ferroptosis to ameliorate folic acid-induced AKI in mice. The findings may have implications for improved understanding of circadian clock-controlled ferroptosis and for formulating new strategies to treat AKI.

Asunto(s)

Lesión Renal Aguda , Ferroptosis , Lesión Renal Aguda/inducido químicamente , Lesión Renal Aguda/tratamiento farmacológico , Animales , Ritmo Circadiano , Ácido Fólico , Ratones , Miembro 1 del Grupo D de la Subfamilia 1 de Receptores Nucleares/genética

RESUMEN

UDP-glucuronosyltransferases (UGTs) are a family of phase II enzymes that play an important role in metabolism and elimination of numerous endo- and xenobiotics. Here, we aimed to characterize diurnal rhythm of Ugt1a9 in mouse liver and to determine the molecular mechanisms underlying the rhythmicity. Hepatic Ugt1a9 mRNA and protein displayed robust diurnal rhythms in wild-type mice with peak levels at zeitgeber time (ZT) 6. Rhythmicity in Ugt1a9 expression was confirmed using synchronized Hepa-1c1c7 cells. We observed time-varying glucuronidation (ZT6 > ZT18) of propofol, a specific Ugt1a9 substrate, consistent with the diurnal pattern of Ugt1a9 protein. Loss of Rev-erbα (a circadian clock component) downregulated the Ugt1a9 expression and blunted its rhythm in mouse liver. Accordingly, propofol glucuronidation was reduced and its dosing time dependency was lost in Rev-erbα -/- mice. Dec2 (a transcription factor) was screened to be the potential intermediate that mediated Rev-erbα regulation of Ugt1a9. We confirmed Rev-erbα as a negative regulator of Dec2 in mice and in Hepa-1c1c7 cells. Based on promoter analysis and luciferase reporter assays, it was found that Dec2 trans-repressed Ugt1a9 via direct binding to an E-box-like motif in the gene promoter. Additionally, regulation of Ugt1a9 by Rev-erbα was Dec2-dependent. In conclusion, Rev-erbα generates and regulates rhythmic Ugt1a9 through periodical inhibition of Dec2, a transcriptional repressor of Ugt1a9. Our study may have implications for understanding of circadian clock-controlled drug metabolism and of metabolism-based chronotherapeutics. SIGNIFICANCE STATEMENT: Hepatic Ugt1a9 displays diurnal rhythmicities in expression and glucuronidation activity in mice. It is uncovered that Rev-erbα generates and regulates rhythmic Ugt1a9 through periodical inhibition of Dec2, a transcriptional repressor of Ugt1a9. The findings may have implications for understanding of circadian clock-controlled drug metabolism and of metabolism-based chronotherapeutics.

Asunto(s)

Relojes Circadianos/genética , Ritmo Circadiano/genética , Glucurónidos/metabolismo , Glucuronosiltransferasa/genética , Miembro 1 del Grupo D de la Subfamilia 1 de Receptores Nucleares/metabolismo , Factores de Transcripción/metabolismo , Animales , Línea Celular Tumoral , Regulación hacia Abajo , Elementos E-Box/genética , Regulación de la Expresión Génica , Glucuronosiltransferasa/metabolismo , Inyecciones Intraperitoneales , Masculino , Ratones , Ratones Noqueados , Miembro 1 del Grupo D de la Subfamilia 1 de Receptores Nucleares/genética , Fotoperiodo , Regiones Promotoras Genéticas , Propofol/administración & dosificación , Propofol/farmacocinética , UDP Glucuronosiltransferasa 1A9

RESUMEN

Hyperhomocysteinemia is associated with poor health, including cardiovascular and brain diseases. Puerarin, initially isolated from Puerariae radix, has been shown to possess anti-hyperhomocysteinemia effect. However, the mechanism of puerarin action remains unknown. Here, we uncovered that puerarin targeted the circadian clock protein Rev-erbα to alleviate hyperhomocysteinemia in mice in a circadian time-dependent manner. We first identified puerarin as an antagonist of Rev-erbα based on luciferase reporter, Gal4 co-transfection and target gene expression assays. Consistent with an antagonistic effect, puerarin induced mRNA and protein expressions of Bhmt, Cbs and Cth (three enzymes involved in homocysteine catabolism and known targets of Rev-erbα) in Hepa-1c1c7 cells. These induction effects of puerarin were lost in Rev-erbα-deficient cells. Furthermore, puerarin dose-dependently alleviated methionine-induced hyperhomocysteinemia in mice as evidenced by decreased levels of total homocysteine and triglyceride. This was accompanied by increased expressions of Bhmt, Cbs and Cth in the liver. Moreover, puerarin dosed at ZT10 generated stronger pharmacological effects than drug dosed at ZT22 consistent with diurnally rhythmic expression of Rev-erbα (a high expression at ZT10 and a low expression at ZT22). In conclusion, puerarin targets Rev-erbα to alleviate hyperhomocysteinemia in mice in a circadian time-dependent manner. The finding of a circadian gene as drug target encourages chronotherapeutic practices on puerarin and related medications for optimized efficacy.

Asunto(s)

Ritmo Circadiano/genética , Hiperhomocisteinemia/tratamiento farmacológico , Isoflavonas/farmacología , Miembro 1 del Grupo D de la Subfamilia 1 de Receptores Nucleares/genética , Animales , Línea Celular , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Células HEK293 , Humanos , Hiperhomocisteinemia/genética , Isoflavonas/administración & dosificación , Masculino , Ratones , Ratones Endogámicos C57BL , Pueraria/química , ARN Mensajero/metabolismo , Factores de Tiempo

RESUMEN

Flavin-containing monooxygenase 5 (FMO5) is a phase I enzyme that plays an important role in xenobiotic metabolism. Here, we aimed to characterize diurnal rhythms of Fmo5 expression and activity in mouse liver and to investigate the potential roles of clock genes (Bmal1, Rev-erbα, and E4bp4) in the generation of diurnal rhythms. Fmo5 mRNA and protein showed robust diurnal rhythms, with peak values at zeitgeber time (ZT) 10/14 and trough values at ZT2/22 in mouse liver. Consistently, a diurnal rhythm was observed for in vitro microsomal Baeyer-Villiger oxidation of pentoxifylline (PTX), a specific reaction catalyzed by Fmo5. Pharmacokinetic studies revealed a more extensive Baeyer-Villiger oxidation of PTX at dosing time of ZT14 than at ZT2, consistent with the diurnal pattern of Fmo5 protein. Fmo5 expression was downregulated and its rhythm was blunted in Bmal1 -/- and Rev-erbα -/- mice. Positive regulation of Fmo5 by Bmal1 and Rev-erbα was confirmed in primary mouse hepatocytes and/or Hepa1-6 cells. Furthermore, Fmo5 expression was upregulated and its rhythm was attenuated in E4bp4 -/- mice. Negative regulation of Fmo5 by E4bp4 was validated using primary mouse hepatocytes. Combined luciferase reporter and chromatin immunoprecipitation assays demonstrated that Bmal1 (a known Rev-erbα activator) activated Fmo5 transcription via direct binding to an E-box (-1822/-1816 bp) in the promoter, whereas E4bp4 (a known Rev-erbα target gene) inhibited Fmo5 transcription by binding to two D-boxes (-1726/-1718 and -804/-796 bp). In conclusion, circadian clock genes control diurnal expression of Fmo5 through transcriptional actions on E-box and D-box cis-elements. SIGNIFICANCE STATEMENT: Hepatic Fmo5 displayed diurnal rhythmicities in expression and activity in mice. We uncovered the molecular mechanism by which the rhythmic Fmo5 expression was generated. Fmo5 promoter presents E-box and D-box binding elements for transcriptional actions from circadian clock proteins such as Bmal1, E4bp4, and Dbp. These findings have implications for understanding clock-controlled drug metabolism and for facilitating the practice of chronotherapeutics.

Asunto(s)

Ritmo Circadiano/fisiología , Regulación Enzimológica de la Expresión Génica , Hígado/enzimología , Oxigenasas/genética , Animales , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/fisiología , Masculino , Ratones , Ratones Endogámicos C57BL , Miembro 1 del Grupo D de la Subfamilia 1 de Receptores Nucleares/fisiología , Oxigenasas/metabolismo , Regiones Promotoras Genéticas , Transcripción Genética

RESUMEN

The intestinal exporter MRP2 plays an important role in disposition and elimination of a wide range of drugs. Here, we aimed to clarify the impact of circadian clock on intestinal MRP2, and to determine the molecular mechanisms for generation of diurnal MRP2 expression. Methods: The regulatory effects of Bmal1 on intestinal MRP2 expression were assessed using intestine-specific Bmal1 knockout (Bmal1iKO ) mice and colon cancer cells. The relative mRNA and protein levels were determined by qPCR and Western blotting, respectively. Everted gut sac, cell viability and in situ intestinal perfusion experiments were performed to evaluate intestinal efflux of the MRP2 substrate methotrexate (MTX). Toxicity and pharmacokinetic experiments were performed with Bmal1iKO mice and control littermates (Bmal1fl/fl mice) after oral gavage of MTX. Transcriptional gene regulation was investigated using luciferase reporter, mobility shift and chromatin immunoprecipitation (ChIP) assays. Results: Bmal1iKO mice were generated by inter-crossing the mice carrying a Bmal1 exon 8 floxed allele (Bmal1fl/fl ) with Villin-Cre mice. Intestinal MRP2 expression exhibited a diurnal oscillation in Bmal1fl/fl mice with a zenith value at ZT6. Bmal1 ablation caused reductions in Mrp2 mRNA and protein levels [as well as in transport activity (measured by MTX)], and blunted their diurnal rhythms. Intestinal ablation of Bmal1 abrogated circadian time-dependency of MTX pharmacokinetics and toxicity. Bmal1/BMAL1 regulation of rhythmic Mrp2/MRP2 expression was also confirmed in the colon cancer CT26 and Caco-2 cells. Based on a combination of luciferase reporter, mobility shift and ChIP assays, we found that Dbp activated and E4bp4 repressed Mrp2 transcription via specific binding to a same D-box (-100/-89 bp) element in promoter region. Further, Bmal1 directly activated the transcription of Dbp and Rev-erbα through the E-boxes, whereas it negatively regulated E4bp4 via the transcriptional repressor Rev-erbα. Positive regulation of Mrp2 by Rev-erbα was also observed, and attained through modulation of E4bp4. Conclusion: Bmal1 coordinates temporal expressions of DBP (a MRP2 activator), REV-ERBα (an E4BP4 repressor) and E4BP4 (a MRP2 repressor), generating diurnal MRP2 expression.

Asunto(s)

Factores de Transcripción ARNTL/metabolismo , Antineoplásicos/metabolismo , Relojes Circadianos , Inactivación Metabólica , Metotrexato/metabolismo , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/metabolismo , Factores de Transcripción ARNTL/deficiencia , Animales , Antineoplásicos/administración & dosificación , Antineoplásicos/farmacocinética , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/metabolismo , Células CACO-2 , Perfilación de la Expresión Génica , Regulación de la Expresión Génica , Humanos , Metotrexato/administración & dosificación , Metotrexato/farmacocinética , Ratones , Ratones Noqueados , Proteína 2 Asociada a Resistencia a Múltiples Medicamentos , Miembro 1 del Grupo D de la Subfamilia 1 de Receptores Nucleares/metabolismo

RESUMEN

Metabolic homeostasis of amino acids is essential for human health. Here, we aimed to investigate a potential role for the clock component reverse erythroblastosis virus α (Rev-erbα) in circadian regulation of amino acid metabolism. RNA-seq with Rev-erbα-/- mice showed expression changes in genes involved in amino acid metabolism, particularly, the urea cycle and methionine metabolism. Rev-erbα ablation increased hepatic mRNA, protein, and enzymatic activity of betaine homocysteine methyltransferase (Bhmt), cystathionine ß-synthase (Cbs), and cystathionine γ-lyase (Cth) and decreased the levels of plasma and liver homocysteine in mice. Cell-based assays confirmed negative regulation of these three genes by Rev-erbα. Combined luciferase reporter, mobility-shift, and chromatin immunoprecipitation assays identified Rev-erbα as a transcriptional repressor of Bhmt, Cbs, and Cth. Rev-erbα ablation or antagonism alleviated chemical-induced hyperhomocysteinemia in mice. This was accompanied by elevated expressions of Bhmt, Cbs, and Cth. Moreover, Rev-erbα ablation or antagonism promoted urea production and ammonia clearance. Of urea cycle-related genes, arginase 1 (Arg1), ornithine transcarbamylase (Otc), and carbamoyl-phosphate synthase 1 (Cps1) expressions were up-regulated in Rev-erbα-/- mice. Negative regulation of these urea cycle genes by Rev-erbα was validated using cell-based experiments. Mechanistic studies revealed that Rev-erbα inhibited CCAAT-enhancer-binding protein α transactivation to repress the transcription of Arg1, Cps1, and Otc. Conclusion: Rev-erbα antagonism alleviates hyperhomocysteinemia and promotes ammonia clearance. Targeting Rev-erbα represents an approach for the management of homocysteine- and ammonia-related diseases.

Asunto(s)

Amoníaco/metabolismo , Ritmo Circadiano/fisiología , Homocisteína/metabolismo , Miembro 1 del Grupo D de la Subfamilia 1 de Receptores Nucleares/antagonistas & inhibidores , Animales , Masculino , Ratones