mTOR regulation of metabolism limits LPS-induced monocyte inflammatory and procoagulant responses.

Translocated lipopolysaccharide (LPS) activates monocytes via TLR4 and is hypothesized to increase cardiovascular disease risk in persons living with HIV. We tested whether mTOR activity supports LPS-stimulated monocyte production of pro-inflammatory cytokines and tissue factor (TF), as it propels the inflammatory response in several immune cell types besides monocytes. However, multi-omics analyses here demonstrate that mTOR activates a metabolic pathway that limits abundance of these gene products in monocytes.

NADH inhibition of SIRT1 links energy state to transcription during time-restricted feeding.

In mammals, circadian rhythms are entrained to the light cycle and drive daily oscillations in levels of NAD, a cosubstrate of the class III histone deacetylase sirtuin 1 (SIRT1) that associates with clock transcription factors. Although NAD also participates in redox reactions, the extent to which NAD(H) couples nutrient state with circadian transcriptional cycles remains unknown. Here we show that nocturnal animals subjected to time-restricted feeding of a calorie-restricted diet (TRF-CR) only during night-time display reduced body temperature and elevated hepatic NADH during daytime.

Phosphatase of Regenerating Liver-1 Selectively Times Circadian Behavior in Darkness via Function in PDF Neurons and Dephosphorylation of TIMELESS.

The timing of behavior under natural light-dark conditions is a function of circadian clocks and photic input pathways, but a mechanistic understanding of how these pathways collaborate in animals is lacking. Here we demonstrate in Drosophila that the Phosphatase of Regenerating Liver-1 (PRL-1) sets period length and behavioral phase gated by photic signals. PRL-1 knockdown in PDF clock neurons dramatically lengthens circadian period.

A novel mouse model overexpressing Nocturnin results in decreased fat mass in male mice.

Nocturnin (NOCT) belongs to the Mg dependent Exonucleases, Endonucleases, Phosphatase (EEP) family of enzymes that exhibit various functions in vitro and in vivo. NOCT is known to function as a deadenylase, cleaving poly-A tails from mRNA (messenger RNA) transcripts. Previously, we reported a role for NOCT in regulating bone marrow stromal cell differentiation through its interactions with PPARγ.

Requirement for NF-κB in maintenance of molecular and behavioral circadian rhythms in mice.

The mammalian circadian clock is encoded by an autoregulatory transcription feedback loop that drives rhythmic behavior and gene expression in the brain and peripheral tissues. Transcriptomic analyses indicate cell type-specific effects of circadian cycles on rhythmic physiology, although how clock cycles respond to environmental stimuli remains incompletely understood. Here, we show that activation of the inducible transcription factor NF-κB in response to inflammatory stimuli leads to marked inhibition of clock repressors, including the , , and genes, within the negative limb.

Circadian measurements of sirtuin biology.

Many of our behavioral and physiological processes display daily oscillations that are under the control of the circadian clock. The core molecular clock network is present in both the brain and peripheral tissues and is composed of a complex series of interlocking transcriptional/translational feedback loops that oscillate with a periodicity of ~24 h. Recent evidence has implicated NAD(+) biosynthesis and the sirtuin family of NAD(+)-dependent protein deacetylases as part of a novel feedback loop within the core clock network, findings which underscore the importance of taking circadian timing into consideration when designing and interpreting metabolic studies, particularly in regard to sirtuin biology.

Usf1, a suppressor of the circadian Clock mutant, reveals the nature of the DNA-binding of the CLOCK:BMAL1 complex in mice.

Genetic and molecular approaches have been critical for elucidating the mechanism of the mammalian circadian clock. Here, we demonstrate that the ClockΔ19 mutant behavioral phenotype is significantly modified by mouse strain genetic background. We map a suppressor of the ClockΔ19 mutation to a ∼900 kb interval on mouse chromosome 1 and identify the transcription factor, Usf1, as the responsible gene.

Competing E3 ubiquitin ligases govern circadian periodicity by degradation of CRY in nucleus and cytoplasm.

Period determination in the mammalian circadian clock involves the turnover rate of the repressors CRY and PER. We show that CRY ubiquitination engages two competing E3 ligase complexes that either lengthen or shorten circadian period in mice. Cloning of a short-period circadian mutant, Past-time, revealed a glycine to glutamate missense mutation in Fbxl21, an F-box protein gene that is a paralog of Fbxl3 that targets the CRY proteins for degradation.

Block of hERG K+ channel and prolongation of action potential duration by fluphenazine at submicromolar concentration.

Fluphenazine is a potent antipsychotic drug that can increase action potential duration and induce QT prolongation in several animal models and in humans. As the block of cardiac human ether-a-go-go-related gene (hERG) channels is one of the leading causes of acquired long QT syndrome, we investigated the acute effects of fluphenazine on hERG channels to determine the electrophysiological basis for its proarrhythmic potential. Fluphenazine at concentrations of 0.

Brain-specific rescue of Clock reveals system-driven transcriptional rhythms in peripheral tissue.

The circadian regulatory network is organized in a hierarchical fashion, with a central oscillator in the suprachiasmatic nuclei (SCN) orchestrating circadian oscillations in peripheral tissues. The nature of the relationship between central and peripheral oscillators, however, is poorly understood. We used the tetOFF expression system to specifically restore Clock function in the brains of Clock(Δ19) mice, which have compromised circadian clocks.

Generation of N-ethyl-N-nitrosourea (ENU) diabetes models in mice demonstrates genotype-specific action of glucokinase activators.

We performed genome-wide mutagenesis in C57BL/6J mice using N-ethyl-N-nitrosourea to identify mutations causing high blood glucose early in life and to produce new animal models of diabetes. Of a total of 13 new lines confirmed by heritability testing, we identified two semi-dominant pedigrees with novel missense mutations (Gck(K140E) and Gck(P417R)) in the gene encoding glucokinase (Gck), the mammalian glucose sensor that is mutated in human maturity onset diabetes of the young type 2 and the target of emerging anti-hyperglycemic agents that function as glucokinase activators (GKAs). Diabetes phenotype corresponded with genotype (mild-to-severe: Gck(+/+) < Gck(P417R/+), Gck(K140E)(/+) < Gck(P417R/P417R), Gck(P417R/K140E), and Gck(K140E/K140E)) and with the level of expression of GCK in liver.

Block of the human ether-a-go-go-related gene (hERG) K+ channel by the antidepressant desipramine.

Desipramine is a tricyclic antidepressant for psychiatric disorders that can induce QT prolongation, which may lead to torsades de pointes. Since blockade of cardiac human ether-a-go-go-related gene (hERG) channels is an important cause of acquired long QT syndrome, we investigated the acute effects of desipramine on hERG channels to determine the electrophysiological basis for its pro-arrhythmic potential. We examined the effects of desipramine on the hERG channels expressed in Xenopus oocytes using two-microelectrode voltage-clamp techniques.

Inhibition of the human ether-a-go-go-related gene (HERG) K+ channels by Lindera erythrocarpa.

Lindera erythrocarpa Makino (Lauraceae) is used as a traditional medicine for analgesic, antidote, and antibacterial purposes and shows anti-tumor activity. We studied the effects of Lindera erythrocarpa on the human ether-a-go-go-related gene (HERG) channel, which appears of importance in favoring cancer progression in vivo and determining cardiac action potential duration. Application of MeOH extract of Lindera erythrocarpa showed a dose-dependent decrease in the amplitudes of the outward currents measured at the end of the pulse (I(HERG)) and the tail currents of HERG (I(tail)).

Block of HERG k channel by classic histamine h(1) receptor antagonist chlorpheniramine.

Chlorpheniramine is a potent first-generation histamine H(1) receptor antagonist that can increase action potential duration and induce QT prolongation in several animal models. Since block of cardiac human ether-a-go-go-related gene (hERG) channels is one of leading causes of acquired long QT syndrome, we investigated the acute effects of chlorpheniramine on hERG channels to determine the electrophysiological basis for its proarrhythmic potential. We examined the effects of chlorpheniramine on the hERG channels expressed in Xenopus oocytes using two-microelectrode voltage-clamp techniques.

H(1) antihistamine drug promethazine directly blocks hERG K(+) channel.

Promethazine is a phenothiazine derivative with antihistaminic (H(1)), sedative, antiemetic, anticholinergic, and antimotion sickness properties that can induce QT prolongation, which may lead to torsades de pointes. Since block of cardiac human ether-a-go-go-related gene (hERG) channels is one of the leading causes of acquired long QT syndrome, we investigated the acute effects of promethazine on hERG channels to determine the electrophysiological basis for its proarrhythmic potential. Promethazine increased the action potential duration at 90% of repolarization (APD(90)) in a concentration-dependent manner, with an IC(50) of 0.

Circadian clock feedback cycle through NAMPT-mediated NAD+ biosynthesis.

The circadian clock is encoded by a transcription-translation feedback loop that synchronizes behavior and metabolism with the light-dark cycle. Here we report that both the rate-limiting enzyme in mammalian nicotinamide adenine dinucleotide (NAD+) biosynthesis, nicotinamide phosphoribosyltransferase (NAMPT), and levels of NAD+ display circadian oscillations that are regulated by the core clock machinery in mice. Inhibition of NAMPT promotes oscillation of the clock gene Per2 by releasing CLOCK:BMAL1 from suppression by SIRT1.

Papaverine, a vasodilator, blocks the pore of the HERG channel at submicromolar concentration.

Papaverine, a vasodilator used as a therapeutic agent for a range of diseases, has been reported to increase the risk of occasional serious ventricular arrhythmias. To examine the mechanism for this effect, we herein tested the effects of papaverine on human ether-a-go-go (HERG) K channels expressed in HEK293 cells and Xenopus oocytes. Our results revealed that papaverine dose-dependently decreased the tail currents of HERG channel expressed in HEK293 cells with the IC50 and the Hill coefficient of 0.

The genetics of mammalian circadian order and disorder: implications for physiology and disease.

Circadian cycles affect a variety of physiological processes, and disruptions of normal circadian biology therefore have the potential to influence a range of disease-related pathways. The genetic basis of circadian rhythms is well studied in model organisms and, more recently, studies of the genetic basis of circadian disorders has confirmed the conservation of key players in circadian biology from invertebrates to humans. In addition, important advances have been made in understanding how these molecules influence physiological functions in tissues throughout the body.

Clomipramine block of the hERG K+ channel: accessibility to F656 and Y652.

Clomipramine is a tricyclic antidepressant for psychiatric disorders that can induce QT prolongation, which may lead to torsades de pointes. Since blockade of cardiac human ether-a-go-go-related gene (hERG) channels is an important cause of acquired long QT syndrome, we investigated the acute effects of clomipramine on hERG channels to determine the electrophysiological basis for its proarrhythmic potential. We examined the effects of clomipramine on the hERG channels expressed in Xenopus oocytes and HEK293 cells using two-microelectrode voltage-clamp and patch-clamp techniques.

Protriptyline block of the human ether-à-go-go-related gene (HERG) K+ channel.

Protriptyline, a tricyclic antidepressant for psychiatric disorders, can induce prolonged QT, torsades de pointes, and sudden death. We studied the effects of protriptyline on human ether-à-go-go-related gene (HERG) channels expressed in Xenopus oocytes and HEK293 cells. Protriptyline induced a concentration-dependent decrease in current amplitudes at the end of the voltage steps and HERG tail currents.

Maprotiline block of the human ether-a-go-go-related gene (HERG) K+ channel.

Maprotiline, an atypical antidepressant, can induce prolonged QT and torsades de pointes. We studied the effects of maprotiline on human ether-a-go-go-related gene (HERG) channels expressed in Xenopus oocytes and HEK293 cells. Maprotiline induced a concentration-dependent decrease in current amplitudes at the end of the voltage steps and tail currents of HERG.

Inducible and reversible Clock gene expression in brain using the tTA system for the study of circadian behavior.

The mechanism of circadian oscillations in mammals is cell autonomous and is generated by a set of genes that form a transcriptional autoregulatory feedback loop. While these "clock genes" are well conserved among animals, their specific functions remain to be fully understood and their roles in central versus peripheral circadian oscillators remain to be defined. We utilized the in vivo inducible tetracycline-controlled transactivator (tTA) system to regulate Clock gene expression conditionally in a tissue-specific and temporally controlled manner.

Dissecting the functions of the mammalian clock protein BMAL1 by tissue-specific rescue in mice.

The basic helix-loop-helix (bHLH)-Per-Arnt-Sim (PAS) domain transcription factor BMAL1 is an essential component of the mammalian circadian pacemaker. Bmal1-/- mice lose circadian rhythmicity but also display tendon calcification and decreased activity, body weight, and longevity. To investigate whether these diverse functions of BMAL1 are tissue-specific, we produced transgenic mice that constitutively express Bmal1 in brain or muscle and examined the effects of rescued gene expression in Bmal1-/- mice.

PERIOD2::LUCIFERASE real-time reporting of circadian dynamics reveals persistent circadian oscillations in mouse peripheral tissues.

Mammalian circadian rhythms are regulated by the suprachiasmatic nucleus (SCN), and current dogma holds that the SCN is required for the expression of circadian rhythms in peripheral tissues. Using a PERIOD2::LUCIFERASE fusion protein as a real-time reporter of circadian dynamics in mice, we report that, contrary to previous work, peripheral tissues are capable of self-sustained circadian oscillations for >20 cycles in isolation. In addition, peripheral organs expressed tissue-specific differences in circadian period and phase.

The gene for soluble N-ethylmaleimide sensitive factor attachment protein alpha is mutated in hydrocephaly with hop gait (hyh) mice.

The spontaneous autosomal recessive mouse mutant for hydrocephaly with hop gait (hyh) exhibits dramatic cystic dilation of the ventricles at birth and invariably develops hopping gait. We show that the gene for soluble N-ethylmaleimide sensitive factor attachment protein alpha, also known as alpha-SNAP, is mutated in hyh mice. alpha-SNAP plays a key role in a wide variety of membrane fusion events in eukaryotic cells, including the regulated exocytosis of neurotransmitters.