Instruct: Given a scientific claim, retrieve documents that support or refute the claim.
Query: Rhythmic expression of Cry1 translates directly into a circadian regulation of cAMP signaling in gluconeogenesis.
During fasting, mammals maintain normal glucose homeostasis by stimulating hepatic gluconeogenesis. Elevations in circulating glucagon and epinephrine, two hormones that activate hepatic gluconeogenesis, trigger the cAMP-mediated phosphorylation of cAMP response element-binding protein (Creb) and dephosphorylation of the Creb-regulated transcription coactivator-2 (Crtc2)--two key transcriptional regulators of this process. Although the underlying mechanism is unclear, hepatic gluconeogenesis is also regulated by the circadian clock, which coordinates glucose metabolism with changes in the external environment. Circadian control of gene expression is achieved by two transcriptional activators, Clock and Bmal1, which stimulate cryptochrome (Cry1 and Cry2) and Period (Per1, Per2 and Per3) repressors that feed back on Clock-Bmal1 activity. Here we show that Creb activity during fasting is modulated by Cry1 and Cry2, which are rhythmically expressed in the liver. Cry1 expression was elevated during the night-day transition, when it reduced fasting gluconeogenic gene expression by blocking glucagon-mediated increases in intracellular cAMP concentrations and in the protein kinase A-mediated phosphorylation of Creb. In biochemical reconstitution studies, we found that Cry1 inhibited accumulation of cAMP in response to G protein-coupled receptor (GPCR) activation but not to forskolin, a direct activator of adenyl cyclase. Cry proteins seemed to modulate GPCR activity directly through interaction with G(s)α. As hepatic overexpression of Cry1 lowered blood glucose concentrations and improved insulin sensitivity in insulin-resistant db/db mice, our results suggest that compounds that enhance cryptochrome activity may provide therapeutic benefit to individuals with type 2 diabetes.
Transcriptional factors binding to cAMP-responsive elements (CREs) in the promoters of various genes belong to the basic domain-leucine zipper superfamily and are composed of three genes in mammals, CREB, CREM, and ATF-1. A large number of CREB, CREM, and ATF-1 proteins are generated by posttranscriptional events, mostly alternative splicing, and regulate gene expression by acting as activators or repressors. Activation is classically brought about by signaling-dependent phosphorylation of a key acceptor site (Ser133 in CREB) by a number of possible kinases, including PKA, CamKIV, and Rsk-2. Phosphorylation is the prerequisite for the interaction of CBP (CREB-binding protein), a co-activator that has also histone acetyltransferase activity. Repression may involve dynamic dephosphorylation of the activators and thus decreased association with CBP. Another pathway of transcriptional repression on CRE sites implicates the inducible repressor ICER (inducible cAMP early repressor), a product of the CREM gene. Being an inducible repressor, ICER is involved in autoregulatory feedback loops of transcription that govern the down-regulation of early response genes, such as the proto-oncogene c-fos. The liver represents a remarkable physiological setting where cAMP-responsive signaling plays a major role. Indeed, a finely tuned program of gene expression is triggered by partial hepatectomy, so that through specific checkpoints a coordinated regeneration of the tissue is obtained. Temporal kinetics of transcriptional activation after hepatectomy reveals a pattern of early induction for several genes, some of them controlled by the CREB/CREM transcription factors. An important role of CREM in liver physiology was suggested by the robust induction of ICER after partial hepatectomy. The delay in tissue regeneration in CREM-deficient mice confirmed the important function of this factor in regulating hepatocyte proliferation. As gene induction is accompanied by critical changes in chromatin organization, the deciphering of the specific modification codes that histones display during liver regeneration and physiology will provide exciting new insights into the dynamics of chromatin architecture.
MOTIVATION Circadian oscillations have been observed in animals, plants, fungi and cyanobacteria and play a fundamental role in coordinating the homeostasis and behavior of biological systems. Genetically encoded molecular clocks found in nearly every cell, based on negative transcription/translation feedback loops and involving only a dozen genes, play a central role in maintaining these oscillations. However, high-throughput gene expression experiments reveal that in a typical tissue, a much larger fraction ([Formula: see text]) of all transcripts oscillate with the day-night cycle and the oscillating species vary with tissue type suggesting that perhaps a much larger fraction of all transcripts, and perhaps also other molecular species, may bear the potential for circadian oscillations. RESULTS To better quantify the pervasiveness and plasticity of circadian oscillations, we conduct the first large-scale analysis aggregating the results of 18 circadian transcriptomic studies and 10 circadian metabolomic studies conducted in mice using different tissues and under different conditions. We find that over half of protein coding genes in the cell can produce transcripts that are circadian in at least one set of conditions and similarly for measured metabolites. Genetic or environmental perturbations can disrupt existing oscillations by changing their amplitudes and phases, suppressing them or giving rise to novel circadian oscillations. The oscillating species and their oscillations provide a characteristic signature of the physiological state of the corresponding cell/tissue. Molecular networks comprise many oscillator loops that have been sculpted by evolution over two trillion day-night cycles to have intrinsic circadian frequency. These oscillating loops are coupled by shared nodes in a large network of coupled circadian oscillators where the clock genes form a major hub. Cells can program and re-program their circadian repertoire through epigenetic and other mechanisms. AVAILABILITY AND IMPLEMENTATION High-resolution and tissue/condition specific circadian data and networks available at http://circadiomics.igb.uci.edu. CONTACT pfbaldi@ics.uci.edu   SUPPLEMENTARY INFORMATION Supplementary data are available at Bioinformatics online.
The MYC oncogene encodes MYC, a transcription factor that binds the genome through sites termed E-boxes (5'-CACGTG-3'), which are identical to the binding sites of the heterodimeric CLOCK-BMAL1 master circadian transcription factor. Hence, we hypothesized that ectopic MYC expression perturbs the clock by deregulating E-box-driven components of the circadian network in cancer cells. We report here that deregulated expression of MYC or N-MYC disrupts the molecular clock in vitro by directly inducing REV-ERBα to dampen expression and oscillation of BMAL1, and this could be rescued by knockdown of REV-ERB. REV-ERBα expression predicts poor clinical outcome for N-MYC-driven human neuroblastomas that have diminished BMAL1 expression, and re-expression of ectopic BMAL1 in neuroblastoma cell lines suppresses their clonogenicity. Further, ectopic MYC profoundly alters oscillation of glucose metabolism and perturbs glutaminolysis. Our results demonstrate an unsuspected link between oncogenic transformation and circadian and metabolic dysrhythmia, which we surmise to be advantageous for cancer.
Cryptochromes are flavin-containing blue light photoreceptors related to photolyases-they are found in both plants and animals and have recently been described for bacteria. In plants, cryptochromes perform a variety of functions including the entrainment of circadian rhythms. They serve a similar role in Drosophila and mammals, where the cryptochromes also perform an additional function as an essential component of the circadian clock.
Glucose is the primary source of energy for the budding yeast Saccharomyces cerevisiae. Although yeast cells can utilize a wide range of carbon sources, presence of glucose suppresses molecular activities involved in the use of alternate carbon sources as well as it represses respiration and gluconeogenesis. This dominant effect of glucose on yeast carbon metabolism is coordinated by several signaling and metabolic interactions that mainly regulate transcriptional activity but are also effective at post-transcriptional and post-translational levels. This review describes effects of glucose repression on yeast carbon metabolism with a focus on roles of the Snf3/Rgt2 glucose-sensing pathway and Snf1 signal transduction in establishment and relief of glucose repression.
The circadian clock and the cell cycle are major cellular systems that organize global physiology in temporal fashion. It seems conceivable that the potentially conflicting programs are coordinated. We show here that overexpression of MYC in U2OS cells attenuates the clock and conversely promotes cell proliferation while downregulation of MYC strengthens the clock and reduces proliferation. Inhibition of the circadian clock is crucially dependent on the formation of repressive complexes of MYC with MIZ1 and subsequent downregulation of the core clock genes BMAL1 (ARNTL), CLOCK and NPAS2. We show furthermore that BMAL1 expression levels correlate inversely with MYC levels in 102 human lymphomas. Our data suggest that MYC acts as a master coordinator that inversely modulates the impact of cell cycle and circadian clock on gene expression.
Many physiological, biochemical and behavioral processes operate under the circadian rhythm, which is generated by an internal time-keeping mechanism commonly referred to as the biological clock, in almost all organisms from bacteria to mammals. The core circadian oscillator is composed of an autoregulatory transcription-translation feedback loop, in which CLOCK and BMAL1 are positive regulators. A cell has two mechanisms, "cell cycle" and "cell rhythm", the relationship between which remains controversial. Therefore, the aim of this study was to explore the effect of Clock and Bmal1 on cell cycle, especially on the G1 phase, using vectors with the tetracycline operator-repressor system. The present study revealed that simultaneous induction of Bmal1 and Clock had an influential effect on the cell cycle in SW480/T-REx/Clock/Bmal1 cells, in which both Clock and Bmal1 could be induced by tetracycline. The observation that induction of both Clock and Bmal1 inhibited cell growth and the significant increase of the G1 phase proportion of in SW480/T-REx/Clock/Bmal1 cells indicated that entry from the G1 to S phase was inhibited by the induction of Clock and Bmal1. Furthermore, overexpression of Clock and Bmal1 prevented the cells from entering into the G2/M phase induced by Paclitaxel, and made the cells more resistant to the agent. In conclusion, we found that overexpression of both Clock and Bmal1 suppressed cell growth. In addition, the present study raised the possibility that Clock and Bmal1 may in part play a role in preventing the cells from entering G1 to S phase of cell cycle via suppression of CyclinD1 expression, and thus acquiring resistance to Paclitaxel.
Circadian disruption accelerates cancer progression, whereas circadian reinforcement could halt it. Mice with P03 pancreatic adenocarcinoma (n = 77) were synchronized and fed ad libitum (AL) or with meal timing (MT) from Zeitgeber time (ZT) 2 to ZT6 with normal or fat diet. Tumor gene expression profiling was determined with DNA microarrays at endogenous circadian time (CT) 4 and CT16. Circadian mRNA expression patterns were determined for clock genes Rev-erbalpha, Per2, and Bmal1, cellular stress genes Hspa8 and Cirbp, and cyclin A2 gene Ccna2 in liver and tumor. The 24-hour patterns in telemetered rest-activity and body temperature and plasma corticosterone and insulin-like growth factor-I (IGF-I) were assessed. We showed that MT inhibited cancer growth by approximately 40% as compared with AL (P = 0.011) irrespective of calorie intake. Clock gene transcription remained arrhythmic in tumors irrespective of feeding schedule or diet. Yet, MT upregulated or downregulated the expression of 423 tumor genes, according to CT. Moreover, 36 genes involved in cellular stress, cell cycle, and metabolism were upregulated at one CT and downregulated 12 h apart. MT induced >10-fold circadian expression of Hspa8, Cirbp, and Ccna2 in tumors. Corticosterone or IGF-I patterns played no role in tumor growth inhibition. In contrast, MT consistently doubled the circadian amplitude of body temperature. Peak and trough respectively corresponded to peak expressions of Hspa8 and Cirbp in tumors. The reinforcement of the host circadian timing system with MT induced 24-hour rhythmic expression of critical genes in clock-deficient tumors, which translated into cancer growth inhibition. Targeting circadian clocks represents a novel potential challenge for cancer therapeutics.
Mammalian cells depend on growth factor signaling to take up nutrients; however, coordination of glucose and glutamine uptake has been a mystery. In this issue of Genes & Development, Wellen and colleagues (pp. 2784-2799) show that glucose flux through the hexosamine biosynthesis pathway regulates growth factor receptor glycosylation and enables glutamine consumption. This mechanism ensures that cells do not engage in anabolic metabolism when nutrients are limiting, and highlights how substrate availability for protein modifications can modulate cell signaling.
Among lower eukaryotes, glucose repression is a conserved, widely spread mechanism regulating carbon catabolism. The yeast Snf1 kinase, the Mig1 DNA-binding repressor and the Mig1-interacting co-repressor complex Cyc8(Ssn6)-Tup1 are central components of this pathway. Previous experiments suggested that cytoplasmic translocation of Mig1, upon its phosphorylation by Snf1 in the nucleus, is the key regulatory step for releasing glucose repression. In this report we re-evaluate this model. We establish the coordinated repressive action of Mig1 and Cyc8-Tup1 on GAL1 transcription, but we find that Cyc8-Tup1 is not tethered by Mig1 to the promoter DNA. We demonstrate that both negative regulators occupy GAL1 continuously under either repression or activation conditions, although the majority of the Mig1 is redistributed to the cytoplasm upon activation. We show that Snf1-dependent phosphorylation of Mig1 abolishes interaction with Cyc8-Tup1, and we propose that regulation of this interaction, not the Mig1 cytoplasmic localization, is the molecular switch that controls transcriptional repression/de-repression.
Endoplasmic reticulum (ER) stress causes pancreatic β-cell dysfunction and contributes to β-cell loss and the progression of type 2 diabetes. Wolfram syndrome 1 (WFS1) has been shown to be an important regulator of the ER stress signalling pathway; however, its role in β-cell function remains unclear. Here we provide evidence that WFS1 is essential for glucose- and glucagon-like peptide 1 (GLP-1)-stimulated cyclic AMP production and regulation of insulin biosynthesis and secretion. Stimulation with glucose causes WFS1 translocation from the ER to the plasma membrane, where it forms a complex with adenylyl cyclase 8 (AC8), an essential cAMP-generating enzyme in the β-cell that integrates glucose and GLP-1 signalling. ER stress and mutant WFS1 inhibit complex formation and activation of AC8, reducing cAMP synthesis and insulin secretion. These findings reveal that an ER-stress-related protein has a distinct role outside the ER regulating both insulin biosynthesis and secretion. The reduction of WFS1 protein on the plasma membrane during ER stress is a contributing factor for β-cell dysfunction and progression of type 2 diabetes.
The rapamycin-sensitive TOR signalling pathway in Saccharomyces cerevisiae activates a cell-growth program in response to nutrients such as nitrogen and carbon. The TOR1 and TOR2 kinases (TOR) control cytoplasmic protein synthesis and degradation through the conserved TAP42 protein. Upon phosphorylation by TOR, TAP42 binds and possibly inhibits type 2A and type-2A-related phosphatases; however, the mechanism by which TOR controls nuclear events such as global repression of starvation-specific transcription is unknown. Here we show that TOR prevents transcription of genes expressed upon nitrogen limitation by promoting the association of the GATA transcription factor GLN3 with the cytoplasmic protein URE2. The binding of GLN3 to URE2 requires TOR-dependent phosphorylation of GLN3. Phosphorylation and cytoplasmic retention of GLN3 are also dependent on the TOR effector TAP42, and are antagonized by the type-2A-related phosphatase SIT4. TOR inhibits expression of carbon-source-regulated genes by stimulating the binding of the transcriptional activators MSN2 and MSN4 to the cytoplasmic 14-3-3 protein BMH2. Thus, the TOR signalling pathway broadly controls nutrient metabolism by sequestering several transcription factors in the cytoplasm.
Circadian organization changes with age, but we do not know the extent to which age-related changes are the result of alterations in the central pacemakers, the peripheral oscillators, or the coupling mechanisms that hold the system together. By using transgenic rats with a luciferase (luc) reporter, we assessed the effects of aging on the rhythm of expression of the Period 1 (Per1) gene in the suprachiasmatic nucleus (SCN) and in peripheral tissues. Young (2 months) and aged (24-26 months) Per1-luc transgenic rats, entrained to light-dark cycles, were killed, and tissues were removed and cultured. Per1-luc expression was measured from 10 tissues. In the SCN, the central mammalian pacemaker, Per1-luc expression was robustly rhythmic for more than 7 weeks in culture. The only difference between SCN rhythmicity in young and old rats was a small but significant age-related shortening of the free-running period. Circadian rhythmicity in some peripheral tissues was unaffected by aging, whereas rhythmicity in other tissues was either phase advanced relative to the light cycle or absent. Those tissues that were arrhythmic could be induced to oscillate by application of forskolin, suggesting that they retained the capacity to oscillate but were not being appropriately driven in vivo. Overall, the results provide new insights into the effects of aging on the mammalian circadian system. Aging seems to affect rhythms in some but not in all tissues and may act primarily on interactions among circadian oscillators, perhaps attenuating the ability of the SCN to drive damped oscillators in the periphery.
In this study, we aimed to investigate changes in the expression of human Clock (hClock), a gene at the core of the circadian gene family, in colorectal carcinomas (CRCs) and to discuss the possible effects. Previous studies have revealed that the disruption of circadian rhythms is one of the endogenous factors that contribute to the initiation and development of CRCs. However, the underlying molecular changes to the circadian genes associated with CRCs have not been explored. Immunofluorescence and quantitative polymerase chain reaction (qPCR) analysis of the hCLOCK protein and gene expression were performed in 30 cases of CRC. The hCLOCK protein was expressed in all specimens obtained from 30 CRC patients. Higher levels of hCLOCK expression were observed in human CRC tissues compared with the paired non-cancerous tissues. hCLOCK expression was significantly higher in poorly differentiated, or late-stage, Dukes' grade tumors and in 64.3% of tumor cases with lymph node metastasis. The hClock gene was expressed in all specimens. A significantly higher expression of hClock was found in human CRC cases compared with paired non-cancerous tissues. There was a strong positive linear correlation between hClock gene expression and protein expression in human CRCs. A strong positive linear correlation was also found between hClock gene expression and ARNT, HIF-1α and VEGF expression in human CRCs. There was no significant correlation between hClock and Bak, Bax, Bid, tumor necrosis factor receptor I (TNFR I) and TNFR II. The circadian gene hClock was stably expressed in human colorectal mucosa and was important in regulating the expression of downstream clock-controlled genes. hCLOCK may interact with HIF-1α/ARNT and activate VEGF to stimulate tumor angiogenesis and metastasis.
The expression of clock genes in vertebrates is widespread and not restricted to classical clock structures. The expression of the Clock gene in zebrafish shows a strong circadian oscillation in many tissues in vivo and in culture, showing that endogenous oscillators exist in peripheral organs. A defining feature of circadian clocks is that they can be set or entrained to local time, usually by the environmental light-dark cycle. An important question is whether peripheral oscillators are entrained to local time by signals from central pacemakers such as the eyes or are themselves directly light-responsive. Here we show that the peripheral organ clocks of zebrafish are set by light-dark cycles in culture. We also show that a zebrafish-derived cell line contains a circadian oscillator, which is also directly light entrained.
Highly conserved among eukaryotic cells, the AMP-activated kinase (AMPK) is a central regulator of carbon metabolism. To map the complete network of interactions around AMPK in yeast (Snf1) and to evaluate the role of its regulatory subunit Snf4, we measured global mRNA, protein and metabolite levels in wild type, Deltasnf1, Deltasnf4, and Deltasnf1Deltasnf4 knockout strains. Using four newly developed computational tools, including novel DOGMA sub-network analysis, we showed the benefits of three-level ome-data integration to uncover the global Snf1 kinase role in yeast. We for the first time identified Snf1's global regulation on gene and protein expression levels, and showed that yeast Snf1 has a far more extensive function in controlling energy metabolism than reported earlier. Additionally, we identified complementary roles of Snf1 and Snf4. Similar to the function of AMPK in humans, our findings showed that Snf1 is a low-energy checkpoint and that yeast can be used more extensively as a model system for studying the molecular mechanisms underlying the global regulation of AMPK in mammals, failure of which leads to metabolic diseases.
Human skin copes with harmful environmental factors that are circadian in nature, yet how circadian rhythms modulate the function of human epidermal stem cells is mostly unknown. Here we show that in human epidermal stem cells and their differentiated counterparts, core clock genes peak in a successive and phased manner, establishing distinct temporal intervals during the 24 hr day period. Each of these successive clock waves is associated with a peak in the expression of subsets of transcripts that temporally segregate the predisposition of epidermal stem cells to respond to cues that regulate their proliferation or differentiation, such as TGFβ and calcium. Accordingly, circadian arrhythmia profoundly affects stem cell function in culture and in vivo. We hypothesize that this intricate mechanism ensures homeostasis by providing epidermal stem cells with environmentally relevant temporal functional cues during the course of the day and that its perturbation may contribute to aging and carcinogenesis.
The signals and molecular mechanisms that regulate the replication of terminally differentiated beta cells are unknown. Here, we report the identification and characterization of transmembrane protein 27 (Tmem27, collectrin) in pancreatic beta cells. Expression of Tmem27 is reduced in Tcf1(-/-) mice and is increased in islets of mouse models with hypertrophy of the endocrine pancreas. Tmem27 forms dimers and its extracellular domain is glycosylated, cleaved and shed from the plasma membrane of beta cells. This cleavage process is beta cell specific and does not occur in other cell types. Overexpression of full-length Tmem27, but not the truncated or soluble protein, leads to increased thymidine incorporation, whereas silencing of Tmem27 using RNAi results in a reduction of cell replication. Furthermore, transgenic mice with increased expression of Tmem27 in pancreatic beta cells exhibit increased beta cell mass. Our results identify a pancreatic beta cell transmembrane protein that regulates cell growth of pancreatic islets.
Circadian rhythms are oscillations of physiology, behavior, and metabolism that have period lengths near 24 hours. In several model organisms and humans, circadian clock genes have been characterized and found to be transcription factors. Because of this, researchers have used microarrays to characterize global regulation of gene expression and algorithmic approaches to detect cycling. This article presents a new algorithm, JTK_CYCLE, designed to efficiently identify and characterize cycling variables in large data sets. Compared with COSOPT and the Fisher's G test, two commonly used methods for detecting cycling transcripts, JTK_CYCLE distinguishes between rhythmic and nonrhythmic transcripts more reliably and efficiently. JTK_CYCLE's increased resistance to outliers results in considerably greater sensitivity and specificity. Moreover, JTK_CYCLE accurately measures the period, phase, and amplitude of cycling transcripts, facilitating downstream analyses. Finally, JTK_CYCLE is several orders of magnitude faster than COSOPT, making it ideal for large-scale data sets. JTK_CYCLE was used to analyze legacy data sets including NIH3T3 cells, which have comparatively low amplitude oscillations. JTK_CYCLE's improved power led to the identification of a novel cluster of RNA-interacting genes whose abundance is under clear circadian regulation. These data suggest that JTK_CYCLE is an ideal tool for identifying and characterizing oscillations in genome-scale data sets.
The regulated release of anorexigenic α-melanocyte stimulating hormone (α-MSH) and orexigenic Agouti-related protein (AgRP) from discrete hypothalamic arcuate neurons onto common target sites in the central nervous system has a fundamental role in the regulation of energy homeostasis. Both peptides bind with high affinity to the melanocortin-4 receptor (MC4R); existing data show that α-MSH is an agonist that couples the receptor to the Gαs signalling pathway, while AgRP binds competitively to block α-MSH binding and blocks the constitutive activity mediated by the ligand-mimetic amino-terminal domain of the receptor. Here we show that, in mice, regulation of firing activity of neurons from the paraventricular nucleus of the hypothalamus (PVN) by α-MSH and AgRP can be mediated independently of Gαs signalling by ligand-induced coupling of MC4R to closure of inwardly rectifying potassium channel, Kir7.1. Furthermore, AgRP is a biased agonist that hyperpolarizes neurons by binding to MC4R and opening Kir7.1, independently of its inhibition of α-MSH binding. Consequently, Kir7.1 signalling appears to be central to melanocortin-mediated regulation of energy homeostasis within the PVN. Coupling of MC4R to Kir7.1 may explain unusual aspects of the control of energy homeostasis by melanocortin signalling, including the gene dosage effect of MC4R and the sustained effects of AgRP on food intake.
Patterns of daily human activity are controlled by an intrinsic circadian clock that promotes ∼24 hr rhythms in many behavioral and physiological processes. This system is altered in delayed sleep phase disorder (DSPD), a common form of insomnia in which sleep episodes are shifted to later times misaligned with the societal norm. Here, we report a hereditary form of DSPD associated with a dominant coding variation in the core circadian clock gene CRY1, which creates a transcriptional inhibitor with enhanced affinity for circadian activator proteins Clock and Bmal1. This gain-of-function CRY1 variant causes reduced expression of key transcriptional targets and lengthens the period of circadian molecular rhythms, providing a mechanistic link to DSPD symptoms. The allele has a frequency of up to 0.6%, and reverse phenotyping of unrelated families corroborates late and/or fragmented sleep patterns in carriers, suggesting that it affects sleep behavior in a sizeable portion of the human population.
The presence of DNA in the cytoplasm of mammalian cells is a danger signal that triggers host immune responses such as the production of type I interferons. Cytosolic DNA induces interferons through the production of cyclic guanosine monophosphate-adenosine monophosphate (cyclic GMP-AMP, or cGAMP), which binds to and activates the adaptor protein STING. Through biochemical fractionation and quantitative mass spectrometry, we identified a cGAMP synthase (cGAS), which belongs to the nucleotidyltransferase family. Overexpression of cGAS activated the transcription factor IRF3 and induced interferon-β in a STING-dependent manner. Knockdown of cGAS inhibited IRF3 activation and interferon-β induction by DNA transfection or DNA virus infection. cGAS bound to DNA in the cytoplasm and catalyzed cGAMP synthesis. These results indicate that cGAS is a cytosolic DNA sensor that induces interferons by producing the second messenger cGAMP.
Budding yeast grown under continuous, nutrient-limited conditions exhibit robust, highly periodic cycles in the form of respiratory bursts. Microarray studies reveal that over half of the yeast genome is expressed periodically during these metabolic cycles. Genes encoding proteins having a common function exhibit similar temporal expression patterns, and genes specifying functions associated with energy and metabolism tend to be expressed with exceptionally robust periodicity. Essential cellular and metabolic events occur in synchrony with the metabolic cycle, demonstrating that key processes in a simple eukaryotic cell are compartmentalized in time.
Circadian (∼24 hour) clocks are fundamentally important for coordinated physiology in organisms as diverse as cyanobacteria and humans. All current models of the molecular circadian clockwork in eukaryotic cells are based on transcription-translation feedback loops. Non-transcriptional mechanisms in the clockwork have been difficult to study in mammalian systems. We circumvented these problems by developing novel assays using human red blood cells, which have no nucleus (or DNA) and therefore cannot perform transcription. Our results show that transcription is not required for circadian oscillations in humans, and that non-transcriptional events seem to be sufficient to sustain cellular circadian rhythms. Using red blood cells, we found that peroxiredoxins, highly conserved antioxidant proteins, undergo ∼24-hour redox cycles, which persist for many days under constant conditions (that is, in the absence of external cues). Moreover, these rhythms are entrainable (that is, tunable by environmental stimuli) and temperature-compensated, both key features of circadian rhythms. We anticipate that our findings will facilitate more sophisticated cellular clock models, highlighting the interdependency of transcriptional and non-transcriptional oscillations in potentially all eukaryotic cells.