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1.
Microbiome ; 7(1): 113, 2019 Aug 09.
Artigo em Inglês | MEDLINE | ID: mdl-31399081

RESUMO

BACKGROUND: Space environment imposes a range of challenges to mammalian physiology and the gut microbiota, and interactions between the two are thought to be important in mammalian health in space. While previous findings have demonstrated a change in the gut microbial community structure during spaceflight, specific environmental factors that alter the gut microbiome and the functional relevance of the microbiome changes during spaceflight remain elusive. METHODS: We profiled the microbiome using 16S rRNA gene amplicon sequencing in fecal samples collected from mice after a 37-day spaceflight onboard the International Space Station. We developed an analytical tool, named STARMAPs (Similarity Test for Accordant and Reproducible Microbiome Abundance Patterns), to compare microbiome changes reported here to other relevant datasets. We also integrated the gut microbiome data with the publically available transcriptomic data in the liver of the same animals for a systems-level analysis. RESULTS: We report an elevated microbiome alpha diversity and an altered microbial community structure that were associated with spaceflight environment. Using STARMAPs, we found the observed microbiome changes shared similarity with data reported in mice flown in a previous space shuttle mission, suggesting reproducibility of the effects of spaceflight on the gut microbiome. However, such changes were not comparable with those induced by space-type radiation in Earth-based studies. We found spaceflight led to significantly altered taxon abundance in one order, one family, five genera, and six species of microbes. This was accompanied by a change in the inferred microbial gene abundance that suggests an altered capacity in energy metabolism. Finally, we identified host genes whose expression in the liver were concordantly altered with the inferred gut microbial gene content, particularly highlighting a relationship between host genes involved in protein metabolism and microbial genes involved in putrescine degradation. CONCLUSIONS: These observations shed light on the specific environmental factors that contributed to a robust effect on the gut microbiome during spaceflight with important implications for mammalian metabolism. Our findings represent a key step toward a better understanding the role of the gut microbiome in mammalian health during spaceflight and provide a basis for future efforts to develop microbiota-based countermeasures that mitigate risks to crew health during long-term human space expeditions.

2.
Neurol Clin ; 37(3): 487-504, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31256785

RESUMO

In mammals, genetic influences of circadian rhythms occur at many levels. A set of core "clock genes" have been identified that form a feedback loop of gene transcription and translation. The core genetic clockwork generates circadian rhythms in cells throughout the body. Polymorphisms in both core clock genes and interacting genes contribute to individual differences in the expression and properties of circadian rhythms. The circadian clock profoundly influences the patterns of gene expression and cellular functions, providing a mechanistic basis for the impact of the genetic circadian system on normal physiological processes as well as the development of diseases.

3.
Science ; 364(6436)2019 04 12.
Artigo em Inglês | MEDLINE | ID: mdl-30975860

RESUMO

To understand the health impact of long-duration spaceflight, one identical twin astronaut was monitored before, during, and after a 1-year mission onboard the International Space Station; his twin served as a genetically matched ground control. Longitudinal assessments identified spaceflight-specific changes, including decreased body mass, telomere elongation, genome instability, carotid artery distension and increased intima-media thickness, altered ocular structure, transcriptional and metabolic changes, DNA methylation changes in immune and oxidative stress-related pathways, gastrointestinal microbiota alterations, and some cognitive decline postflight. Although average telomere length, global gene expression, and microbiome changes returned to near preflight levels within 6 months after return to Earth, increased numbers of short telomeres were observed and expression of some genes was still disrupted. These multiomic, molecular, physiological, and behavioral datasets provide a valuable roadmap of the putative health risks for future human spaceflight.


Assuntos
Adaptação Fisiológica , Astronautas , Voo Espacial , Imunidade Adaptativa , Peso Corporal , Artérias Carótidas/diagnóstico por imagem , Espessura Intima-Media Carotídea , Dano ao DNA , Metilação de DNA , Microbioma Gastrointestinal , Instabilidade Genômica , Humanos , Masculino , Homeostase do Telômero , Fatores de Tempo , Estados Unidos , United States National Aeronautics and Space Administration
4.
Sci Rep ; 9(1): 4808, 2019 Mar 18.
Artigo em Inglês | MEDLINE | ID: mdl-30886221

RESUMO

In addition to the characteristic motor symptoms, Parkinson's disease (PD) often involves a constellation of sleep and mood symptoms. However, the mechanisms underlying these comorbidities are largely unknown. We have previously reconstructed gene networks in the striatum of a population of (C57BL/6J x A/J) F2 mice and associated the networks to sleep and affective phenotypes, providing a resource for integrated analyses to investigate perturbed sleep and affective functions at the gene network level. Combining this resource with PD-relevant transcriptomic datasets from humans and mice, we identified four networks that showed elevated gene expression in PD patients, including a circadian clock and mitotic network that was altered similarly in mouse models of PD. We then utilized multiple types of omics data from public databases and linked this gene network to postsynaptic dopamine signaling in the striatum, CDK1-modulated transcriptional regulation, and the genetic susceptibility of PD. These findings suggest that dopamine deficiency, a key aspect of PD pathology, perturbs a circadian/mitotic gene network in striatal neurons. Since the normal functions of this network were relevant to sleep and affective behaviors, these findings implicate that dysregulation of functional gene networks may be involved in the emergence of non-motor symptoms in PD. Our analyses present a framework for integrating multi-omics data from diverse sources in mice and humans to reveal insights into comorbid symptoms of complex diseases.

5.
Int J Mol Sci ; 17(12)2016 Dec 02.
Artigo em Inglês | MEDLINE | ID: mdl-27918452

RESUMO

BACKGROUND: Colorectal cancer (CRC) is associated with the modern lifestyle. Chronic alcohol consumption-a frequent habit of majority of modern societies-increases the risk of CRC. Our group showed that chronic alcohol consumption increases polyposis in a mouse mode of CRC. Here we assess the effect of circadian disruption-another modern life style habit-in promoting alcohol-associated CRC. METHOD: TS4Cre × adenomatous polyposis coli (APC)lox468 mice underwent (a) an alcohol-containing diet while maintained on a normal 12 h light:12 h dark cycle; or (b) an alcohol-containing diet in conjunction with circadian disruption by once-weekly 12 h phase reversals of the light:dark (LD) cycle. Mice were sacrificed after eight weeks of full alcohol and/or LD shift to collect intestine samples. Tumor number, size, and histologic grades were compared between animal groups. Mast cell protease 2 (MCP2) and 6 (MCP6) histology score were analyzed and compared. Stool collected at baseline and after four weeks of experimental manipulations was used for microbiota analysis. RESULTS: The combination of alcohol and LD shifting accelerated intestinal polyposis, with a significant increase in polyp size, and caused advanced neoplasia. Consistent with a pathogenic role of stromal tryptase-positive mast cells in colon carcinogenesis, the ratio of mMCP6 (stromal)/mMCP2 (intraepithelial) mast cells increased upon LD shifting. Baseline microbiota was similar between groups, and experimental manipulations resulted in a significant difference in the microbiota composition between groups. CONCLUSIONS: Circadian disruption by Light:dark shifting exacerbates alcohol-induced polyposis and CRC. Effect of circadian disruption could, at least partly, be mediated by promoting a pro-tumorigenic inflammatory milieu via changes in microbiota.


Assuntos
Alcoolismo/complicações , Carcinogênese/patologia , Neoplasias Colorretais/etiologia , Inflamação/patologia , Intestinos/microbiologia , Intestinos/patologia , Microbiota , Fotoperíodo , Animais , Neoplasias Colorretais/microbiologia , Neoplasias Colorretais/patologia , Disbiose/complicações , Disbiose/microbiologia , Disbiose/patologia , Células Epiteliais/patologia , Comportamento Alimentar , Mastócitos/patologia , Camundongos
6.
Am J Physiol Gastrointest Liver Physiol ; 311(1): G192-201, 2016 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-27198191

RESUMO

Alcohol-induced intestinal hyperpermeability (AIHP) is a known risk factor for alcoholic liver disease (ALD), but only 20-30% of heavy alcoholics develop AIHP and ALD. The hypothesis of this study is that circadian misalignment would promote AIHP. We studied two groups of healthy subjects on a stable work schedule for 3 mo [day workers (DW) and night workers (NW)]. Subjects underwent two circadian phase assessments with sugar challenge to access intestinal permeability between which they drank 0.5 g/kg alcohol daily for 7 days. Sleep architecture by actigraphy did not differ at baseline or after alcohol between either group. After alcohol, the dim light melatonin onset (DLMO) in the DW group did not change significantly, but in the NW group there was a significant 2-h phase delay. Both the NW and DW groups had no change in small bowel permeability with alcohol, but only in the NW group was there an increase in colonic and whole gut permeability. A lower area under the curve of melatonin inversely correlated with increased colonic permeability. Alcohol also altered peripheral clock gene amplitude of peripheral blood mononuclear cells in CLOCK, BMAL, PER1, CRY1, and CRY2 in both groups, and inflammatory markers lipopolysaccharide-binding protein, LPS, and IL-6 had an elevated mesor at baseline in NW vs. DW and became arrhythmic with alcohol consumption. Together, our data suggest that central circadian misalignment is a previously unappreciated risk factor for AIHP and that night workers may be at increased risk for developing liver injury with alcohol consumption.


Assuntos
Consumo de Bebidas Alcoólicas/efeitos adversos , Ritmo Circadiano , Colo/efeitos dos fármacos , Intestino Delgado/efeitos dos fármacos , Admissão e Escalonamento de Pessoal , Transtornos do Sono do Ritmo Circadiano/complicações , Sono , Tolerância ao Trabalho Programado , Adulto , Biomarcadores/sangue , Peptídeos e Proteínas de Sinalização do Ritmo Circadiano/sangue , Peptídeos e Proteínas de Sinalização do Ritmo Circadiano/genética , Colo/metabolismo , Colo/fisiopatologia , Regulação da Expressão Gênica , Humanos , Mediadores da Inflamação/sangue , Intestino Delgado/metabolismo , Intestino Delgado/fisiopatologia , Melatonina/sangue , Pessoa de Meia-Idade , Permeabilidade , Transtornos do Sono do Ritmo Circadiano/sangue , Transtornos do Sono do Ritmo Circadiano/diagnóstico , Transtornos do Sono do Ritmo Circadiano/fisiopatologia , Fatores de Tempo , Adulto Jovem
7.
Sci Rep ; 5: 16896, 2015 Nov 20.
Artigo em Inglês | MEDLINE | ID: mdl-26584570

RESUMO

A variety of environmental factors contribute to progressive development of osteoarthritis (OA). Environmental factors that upset circadian rhythms have been linked to various diseases. Our recent work establishes chronic environmental circadian disruption - analogous to rotating shiftwork-associated disruption of circadian rhythms in humans - as a novel risk factor for the development of OA. Evidence suggests shift workers are prone to obesity and also show altered eating habits (i.e., increased preference for high-fat containing food). In the present study, we investigated the impact of chronic circadian rhythm disruption in combination with a high-fat diet (HFD) on progression of OA in a mouse model. Our study demonstrates that when mice with chronically circadian rhythms were fed a HFD, there was a significant proteoglycan (PG) loss and fibrillation in knee joint as well as increased activation of the expression of the catabolic mediators involved in cartilage homeostasis. Our results, for the first time, provide the evidence that environmental disruption of circadian rhythms plus HFD potentiate OA-like pathological changes in the mouse joints. Thus, our findings may open new perspectives on the interactions of chronic circadian rhythms disruption with diet in the development of OA and may have potential clinical implications.


Assuntos
Ritmo Circadiano , Dieta Hiperlipídica/efeitos adversos , Articulação do Joelho/patologia , Osteoartrite do Joelho/etiologia , Animais , Peso Corporal , Modelos Animais de Doenças , Progressão da Doença , Meio Ambiente , Humanos , Articulação do Joelho/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Obesidade/etiologia , Obesidade/fisiopatologia , Osteoartrite do Joelho/fisiopatologia , Proteoglicanas/metabolismo
8.
J Cell Physiol ; 230(9): 2174-2183, 2015 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-25655021

RESUMO

Circadian rhythm dysfunction is linked to many diseases, yet pathophysiological roles in articular cartilage homeostasis and degenerative joint disease including osteoarthritis (OA) remains to be investigated in vivo. Here, we tested whether environmental or genetic disruption of circadian homeostasis predisposes to OA-like pathological changes. Male mice were examined for circadian locomotor activity upon changes in the light:dark (LD) cycle or genetic disruption of circadian rhythms. Wild-type (WT) mice were maintained on a constant 12 h:12 h LD cycle (12:12 LD) or exposed to weekly 12 h phase shifts. Alternatively, male circadian mutant mice (Clock(Δ19) or Csnk1e(tau) mutants) were compared with age-matched WT littermates that were maintained on a constant 12:12 LD cycle. Disruption of circadian rhythms promoted osteoarthritic changes by suppressing proteoglycan accumulation, upregulating matrix-degrading enzymes and downregulating anabolic mediators in the mouse knee joint. Mechanistically, these effects involved activation of the PKCδ-ERK-RUNX2/NFκB and ß-catenin signaling pathways, stimulation of MMP-13 and ADAMTS-5, as well as suppression of the anabolic mediators SOX9 and TIMP-3 in articular chondrocytes of phase-shifted mice. Genetic disruption of circadian homeostasis does not predispose to OA-like pathological changes in joints. Our results, for the first time, provide compelling in vivo evidence that environmental disruption of circadian rhythms is a risk factor for the development of OA-like pathological changes in the mouse knee joint.


Assuntos
Proteínas CLOCK/genética , Cartilagem Articular/metabolismo , Ritmo Circadiano/genética , Osteoartrite do Joelho/genética , Animais , Cartilagem Articular/patologia , Ritmo Circadiano/fisiologia , Subunidade alfa 1 de Fator de Ligação ao Core/biossíntese , Suscetibilidade a Doenças , Meio Ambiente , Homeostase/genética , Humanos , Articulação do Joelho/metabolismo , Articulação do Joelho/fisiopatologia , Sistema de Sinalização das MAP Quinases/genética , Metaloproteinase 13 da Matriz/biossíntese , Camundongos , Atividade Motora/genética , Atividade Motora/fisiologia , Osteoartrite do Joelho/fisiopatologia
9.
J Neurosci Methods ; 235: 59-64, 2014 Sep 30.
Artigo em Inglês | MEDLINE | ID: mdl-24992574

RESUMO

BACKGROUND: The forced swim test (FST) is used to predict the effectiveness of novel antidepressant treatments. In this test, a mouse or rat is placed in a beaker of water for several minutes, and the amount of time spent passively floating is measured; antidepressants reduce the amount of such immobility. Though the FST is commonly used, manually scoring the test is time-consuming and involves considerable subjectivity. NEW METHOD: We developed a simple MATLAB-based motion-detection method to quantify mice's activity in videos of FST. FST trials are video-recorded from a side view. Each pixel of the video is compared between subsequent video frames; if the pixel's color difference surpasses a threshold, a motion count is recorded. RESULTS: Human-scored immobility time correlates well with total motion detected by the computer (r=-0.80) and immobility time determined by the computer (r=0.83). Our computer method successfully detects group differences in activity between genotypes and different days of testing. Furthermore, we observe heterosis for this behavior, in which (C57BL/6J×A/J) F1 hybrid mice are more active in the FST than the parental strains. COMPARISON WITH EXISTING METHODS: This computer-scoring method is much faster and more objective than human scoring. Other automatic scoring methods exist, but they require the purchase of expensive hardware and/or software. CONCLUSION: This computer-scoring method is an effective, fast, and low-cost method of quantifying the FST. It is validated by replicating statistical differences observed in traditional visual scoring. We also demonstrate a case of heterosis in the FST.


Assuntos
Depressão/diagnóstico , Movimento (Física) , Testes Neuropsicológicos , Reconhecimento Automatizado de Padrão/métodos , Natação , Gravação em Vídeo/métodos , Animais , Camundongos , Camundongos Endogâmicos C57BL , Atividade Motora , Software , Especificidade da Espécie , Fatores de Tempo
10.
Sleep ; 37(4): 785-93, 793A-793C, 2014 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-24744456

RESUMO

STUDY OBJECTIVES: Efforts to identify the genetic basis of mammalian sleep have included quantitative trait locus (QTL) mapping and gene targeting of known core circadian clock genes. We combined three different genetic approaches to identify and test a positional candidate sleep gene - the circadian gene casein kinase 1 epsilon (Csnk1e), which is located in a QTL we identified for rapid eye movement (REM) sleep on chromosome 15. MEASUREMENTS AND RESULTS: Using electroencephalographic (EEG) and electromyographic (EMG) recordings, baseline sleep was examined in a 12-h light:12-h dark (LD 12:12) cycle in mice of seven genotypes, including Csnk1e(tau/tau) and Csnk1e(-/-) mutant mice, Csnk1e (B6.D2) and Csnk1e (D2.B6) congenic mice, and their respective wild-type littermate control mice. Additionally, Csnk1e(tau/tau) and wild-type mice were examined in constant darkness (DD). Csnk1e(tau/tau) mutant mice and both Csnk1e (B6.D2) and Csnk1e (D2.B6) congenic mice showed significantly higher proportion of sleep time spent in REM sleep during the dark period than wild-type controls - the original phenotype for which the QTL on chromosome 15 was identified. This phenotype persisted in Csnk1e(tau/tau) mice while under free-running DD conditions. Other sleep phenotypes observed in Csnk1e(tau/tau) mice and congenics included a decreased number of bouts of nonrapid eye movement (NREM) sleep and an increased average NREM sleep bout duration. CONCLUSIONS: These results demonstrate a role for Csnk1e in regulating not only the timing of sleep, but also the REM sleep amount and NREM sleep architecture, and support Csnk1e as a causal gene in the sleep QTL on chromosome 15.


Assuntos
Caseína Quinase Iépsilon/genética , Relógios Circadianos/genética , Sono REM/genética , Sono REM/fisiologia , Sono/genética , Sono/fisiologia , Animais , Caseína Quinase Iépsilon/metabolismo , Cromossomos de Mamíferos/genética , Relógios Circadianos/fisiologia , Ritmo Circadiano/genética , Ritmo Circadiano/fisiologia , Escuridão , Eletroencefalografia , Eletromiografia , Genótipo , Luz , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos DBA , Mutação/genética , Fenótipo , Locos de Características Quantitativas/genética , Fatores de Tempo
11.
PLoS One ; 8(6): e67102, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-23825629

RESUMO

The circadian clock orchestrates temporal patterns of physiology and behavior relative to the environmental light:dark cycle by generating and organizing transcriptional and biochemical rhythms in cells and tissues throughout the body. Circadian clock genes have been shown to regulate the physiology and function of the gastrointestinal tract. Disruption of the intestinal epithelial barrier enables the translocation of proinflammatory bacterial products, such as endotoxin, across the intestinal wall and into systemic circulation; a process that has been linked to pathologic inflammatory states associated with metabolic, hepatic, cardiovascular and neurodegenerative diseases - many of which are commonly reported in shift workers. Here we report, for the first time, that circadian disorganization, using independent genetic and environmental strategies, increases permeability of the intestinal epithelial barrier (i.e., gut leakiness) in mice. Utilizing chronic alcohol consumption as a well-established model of induced intestinal hyperpermeability, we also found that both genetic and environmental circadian disruption promote alcohol-induced gut leakiness, endotoxemia and steatohepatitis, possibly through a mechanism involving the tight junction protein occludin. Circadian organization thus appears critical for the maintenance of intestinal barrier integrity, especially in the context of injurious agents, such as alcohol. Circadian disruption may therefore represent a previously unrecognized risk factor underlying the susceptibility to or development of alcoholic liver disease, as well as other conditions associated with intestinal hyperpermeability and an endotoxin-triggered inflammatory state.


Assuntos
Relógios Circadianos , Etanol/farmacologia , Hepatite Alcoólica/etiologia , Fígado/efeitos dos fármacos , Animais , Fígado/patologia , Camundongos , Camundongos Endogâmicos C57BL
12.
PLoS One ; 7(5): e37668, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22649550

RESUMO

BACKGROUND: The circadian clock has been linked to reproduction at many levels in mammals. Epidemiological studies of female shift workers have reported increased rates of reproductive abnormalities and adverse pregnancy outcomes, although whether the cause is circadian disruption or another factor associated with shift work is unknown. Here we test whether environmental disruption of circadian rhythms, using repeated shifts of the light:dark (LD) cycle, adversely affects reproductive success in mice. METHODOLOGY/PRINCIPAL FINDINGS: Young adult female C57BL/6J (B6) mice were paired with B6 males until copulation was verified by visual identification of vaginal plug formation. Females were then randomly assigned to one of three groups: control, phase-delay or phase-advance. Controls remained on a constant 12-hr light:12-hr dark cycle, whereas phase-delayed and phase-advanced mice were subjected to 6-hr delays or advances in the LD cycle every 5-6 days, respectively. The number of copulations resulting in term pregnancies was determined. Control females had a full-term pregnancy success rate of 90% (11/12), which fell to 50% (9/18; p<0.1) in the phase-delay group and 22% (4/18; p<0.01) in the phase-advance group. CONCLUSIONS/SIGNIFICANCE: Repeated shifting of the LD cycle, which disrupts endogenous circadian timekeeping, dramatically reduces pregnancy success in mice. Advances of the LD cycle have a greater negative impact on pregnancy outcomes and, in non-pregnant female mice, require longer for circadian re-entrainment, suggesting that the magnitude or duration of circadian misalignment may be related to the severity of the adverse impact on pregnancy. These results explicitly link disruptions of circadian entrainment to adverse pregnancy outcomes in mammals, which may have important implications for the reproductive health of female shift workers, women with circadian rhythm sleep disorders and/or women with disturbed circadian rhythms for other reasons.


Assuntos
Ritmo Circadiano/fisiologia , Meio Ambiente , Reprodução/fisiologia , Animais , Feminino , Locomoção/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Fotoperíodo , Gravidez , Resultado da Gravidez
13.
PLoS One ; 6(9): e25079, 2011.
Artigo em Inglês | MEDLINE | ID: mdl-21949859

RESUMO

The neuroendocrine and metabolic effects of leptin have been extensively researched since the discovery, and the later identification, of the leptin gene mutated within the ob/ob mouse. Leptin is required for optimal health in a number of physiological systems (e.g. fertility, bone density, body weight regulation). Despite the extensive leptin literature and many observations of leptin's cyclical pattern over the 24-hour day, few studies have specifically examined how the circadian rhythm of leptin may be essential to leptin signaling and health. Here we present data indicating that a rhythmic leptin profile (e.g. 1 peak every 24 hours) leads to excessive weight gain during desynchronized feeding whereas non-rhythmic leptin provided in a continuous manner does not lead to excessive body weight gain under similar feeding conditions. This study suggests that feeding time can interact with leptin's endogenous rhythm to influence metabolic signals, specifically leading to excessive body weight gains during 'wrongly' timed feeding.


Assuntos
Peso Corporal/efeitos dos fármacos , Ritmo Circadiano/efeitos dos fármacos , Leptina/farmacologia , Obesidade/etiologia , Periodicidade , Ganho de Peso/efeitos dos fármacos , Animais , Comportamento Alimentar , Masculino , Camundongos , Camundongos Obesos
15.
Proc Natl Acad Sci U S A ; 107(18): 8399-403, 2010 May 04.
Artigo em Inglês | MEDLINE | ID: mdl-20404168

RESUMO

Most laboratory mouse strains including C57BL/6J do not produce detectable levels of pineal melatonin owing to deficits in enzymatic activity of arylalkylamine N-acetyltransferase (AANAT) and N-acetylserotonin O-methyl transferase (ASMT), two enzymes necessary for melatonin biosynthesis. Here we report that alleles segregating at these two loci in C3H/HeJ mice, an inbred strain producing melatonin, suppress the circadian period-lengthening effect of the Clock mutation. Through a functional mapping approach, we localize mouse Asmt to chromosome X and show that it, and the Aanat locus on chromosome 11, are significantly associated with pineal melatonin levels. Treatment of suprachiasmatic nucleus (SCN) explant cultures from Period2(Luciferase) (Per2(Luc)) Clock/+ reporter mice with melatonin, or the melatonin agonist, ramelteon, phenocopies the genetic suppression of the Clock mutant phenotype observed in living animals. These results demonstrate that melatonin suppresses the Clock/+ mutant phenotype and interacts with Clock to affect the mammalian circadian system.


Assuntos
Proteínas CLOCK/metabolismo , Ritmo Circadiano , Regulação para Baixo , Melatonina/biossíntese , Mutação , Acetilserotonina O-Metiltransferasa/metabolismo , Animais , Arilalquilamina N-Acetiltransferase/metabolismo , Comportamento Animal , Proteínas CLOCK/genética , Cromossomos , Camundongos , Camundongos Endogâmicos C3H , Fenótipo
16.
Proc Natl Acad Sci U S A ; 103(24): 9327-32, 2006 Jun 13.
Artigo em Inglês | MEDLINE | ID: mdl-16754844

RESUMO

The mouse Clock gene encodes a basic helix-loop-helix-PAS transcription factor, CLOCK, that acts in concert with BMAL1 to form the positive elements of the circadian clock mechanism in mammals. The original Clock mutant allele is a dominant negative (antimorphic) mutation that deletes exon 19 and causes an internal deletion of 51 aa in the C-terminal activation domain of the CLOCK protein. Here we report that heterozygous Clock/+ mice exhibit high-amplitude phase-resetting responses to 6-h light pulses (Type 0 resetting) as compared with wild-type mice that have low amplitude (Type 1) phase resetting. The magnitude and time course of acute light induction in the suprachiasmatic nuclei of the only known light-induced core clock genes, Per1 and Per2, are not affected by the Clock/+ mutation. However, the amplitude of the circadian rhythms of Per gene expression are significantly reduced in Clock homozygous and heterozygous mutants. Rhythms of PER2::LUCIFERASE expression in suprachiasmatic nuclei explant cultures also are reduced in amplitude in Clock heterozygotes. The phase-response curves to changes in culture medium are Type 0 in Clock heterozygotes, but Type 1 in wild types, similar to that seen for light in vivo. The increased efficacy of resetting stimuli and decreased PER expression amplitude can be explained in a unified manner by a model in which the Clock mutation reduces circadian pacemaker amplitude in the suprachiasmatic nuclei.


Assuntos
Relógios Biológicos/fisiologia , Ritmo Circadiano/fisiologia , Mutação , Transativadores/metabolismo , Animais , Proteínas CLOCK , Proteínas de Ciclo Celular , Regulação da Expressão Gênica , Luz , Camundongos , Camundongos Endogâmicos , Atividade Motora/fisiologia , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Proteínas Circadianas Period , Fotoperíodo , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Núcleo Supraquiasmático/fisiologia , Transativadores/genética , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo
17.
Trends Neurosci ; 29(4): 233-40, 2006 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-16519954

RESUMO

Significant developments have occurred in our understanding of the mammalian genome thanks to informatics, expression profiling and sequencing of the human and rodent genomes. However, although these facets of genomic analysis are being addressed, analysis of in vivo gene function remains a formidable task. Evaluation of the phenotype of mutants provides powerful access to gene function, and this approach is particularly relevant to the nervous system and behavior. Here, we discuss the complementary mouse genetic approaches of gene-driven, targeted mutagenesis and phenotype-driven, chemical mutagenesis. We highlight an NIH-supported large-scale effort to use phenotype-driven mutagenesis screens to identify mouse mutants with neural and behavioral alterations. Such single-gene mutations can then be used for gene identification using positional candidate gene-cloning methods.


Assuntos
Comportamento Animal/fisiologia , Genética Comportamental/métodos , Genômica/métodos , Camundongos Mutantes/genética , Mutagênese/genética , Fenótipo , Animais , Mapeamento Cromossômico , Relações Interinstitucionais , Camundongos , National Institutes of Health (U.S.) , Locos de Características Quantitativas/fisiologia , Projetos de Pesquisa , Estados Unidos
18.
Genome Res ; 16(3): 436-40, 2006 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-16461637

RESUMO

Phenotype-driven genetics can be used to create mouse models of human disease and birth defects. However, the utility of these mutant models is limited without identification of the causal gene. To facilitate genetic mapping, we developed a fixed single nucleotide polymorphism (SNP) panel of 394 SNPs as an alternative to analyses using simple sequence length polymorphism (SSLP) marker mapping. With the SNP panel, chromosomal locations for 22 monogenic mutants were identified. The average number of affected progeny genotyped for mapped monogenic mutations is nine. Map locations for several mutants have been obtained with as few as four affected progeny. The average size of genetic intervals obtained for these mutants is 43 Mb, with a range of 17-83 Mb. Thus, our SNP panel allows for identification of moderate resolution map position with small numbers of mice in a high-throughput manner. Importantly, the panel is suitable for mapping crosses from many inbred and wild-derived inbred strain combinations. The chromosomal localizations obtained with the SNP panel allow one to quickly distinguish between potentially novel loci or remutations in known genes, and facilitates fine mapping and positional cloning. By using this approach, we identified DNA sequence changes in two ethylnitrosourea-induced mutants.


Assuntos
Mapeamento Cromossômico/métodos , Genoma , Camundongos/genética , Polimorfismo de Nucleotídeo Único , Animais , Clonagem de Organismos , Genótipo , Humanos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos , Mutação
19.
Genetica ; 122(1): 51-64, 2004 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-15619961

RESUMO

A step towards annotating the mouse genome is to use forward genetics in phenotype-driven screens to saturate the genome with mutations. The purpose of this article is to highlight the new projects in North America that are focused on isolating mouse mutations after ENU mutagenesis and phenotype screening.


Assuntos
Mapeamento Cromossômico , Camundongos/genética , Mutação , Animais , Modelos Animais de Doenças , Etilnitrosoureia , Feminino , Masculino , National Institutes of Health (U.S.) , Doenças do Sistema Nervoso/genética , Fenótipo , Estados Unidos
20.
Vision Res ; 44(28): 3335-45, 2004 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-15536001

RESUMO

Random mutagenesis combined with phenotypic screening using carefully crafted functional tests has successfully led to the discovery of genes that are essential for a number of functions. This approach does not require prior knowledge of the identity of the genes that are involved and is a way to ascribe function to the nearly 6000 genes for which knowledge of the DNA sequence has been inadequate to determine the function of the gene product. In an effort to identify genes involved in the visual system via this approach, we have tested over 9000 first and third generation offspring of mice treated with the mutagen N-ethyl-N-nitrosourea (ENU) for visual defects, as evidenced by abnormalities in the electroretinogram and appearance of the fundus. We identified 61 putative mutations with this procedure and outline the steps needed to identify the affected genes.


Assuntos
Transtornos da Visão/genética , Seleção Visual/métodos , Animais , Eletrorretinografia/métodos , Etilnitrosoureia , Fundo de Olho , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Mutantes , Mutagênese , Fenótipo , Retina/fisiopatologia , Transtornos da Visão/diagnóstico , Transtornos da Visão/fisiopatologia , Visão Ocular/genética
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