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Sample records for clock proteins drosophila

  1. Age-Related Changes in the Expression of the Circadian Clock Protein PERIOD in Drosophila Glial Cells

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    Dani M. Long

    2018-01-01

    Full Text Available Circadian clocks consist of molecular negative feedback loops that coordinate physiological, neurological, and behavioral variables into “circa” 24-h rhythms. Rhythms in behavioral and other circadian outputs tend to weaken during aging, as evident in progressive disruptions of sleep-wake cycles in aging organisms. However, less is known about the molecular changes in the expression of clock genes and proteins that may lead to the weakening of circadian outputs. Western blot studies have demonstrated that the expression of the core clock protein PERIOD (PER declines in the heads of aged Drosophila melanogaster flies. This age-related decline in PER does not occur in the central pacemaker neurons but has been demonstrated so far in retinal photoreceptors. Besides photoreceptors, clock proteins are also expressed in fly glia, which play important roles in neuronal homeostasis and are further categorized into subtypes based on morphology and function. While previous studies of mammalian glial cells have demonstrated the presence of functional clocks in astrocytes and microglia, it is not known which glial cell types in Drosophila express clock proteins and how their expression may change in aged individuals. Here, we conducted immunocytochemistry experiments to identify which glial subtypes express PER protein suggestive of functional circadian clocks. Glial cell subtypes that showed night-time accumulation and day-time absence in PER consistent with oscillations reported in the pacemaker neurons were selected to compare the level of PER protein between young and old flies. Our data demonstrate that some glial subtypes show rhythmic PER expression and the relative PER levels become dampened with advanced age. Identification of glial cell types that display age-related dampening of PER levels may help to understand the cellular changes that contribute to the loss of homeostasis in the aging brain.

  2. QUASIMODO, a novel GPI-anchored zona pellucida protein involved in light input to the Drosophila circadian clock

    Czech Academy of Sciences Publication Activity Database

    Chen, K. F.; Peschel, N.; Závodská, Radka; Sehadová, Hana; Stanewsky, R.

    2011-01-01

    Roč. 21, č. 9 (2011), s. 719-729 ISSN 0960-9822 R&D Projects: GA MŠk LC07032 Institutional research plan: CEZ:AV0Z50070508 Keywords : QUASIMODO * Drosophila * circadian clock Subject RIV: ED - Physiology Impact factor: 9.647, year: 2011

  3. Age-Related Changes in the Expression of the Circadian Clock Protein PERIOD in Drosophila Glial Cells

    OpenAIRE

    Long, Dani M.; Giebultowicz, Jadwiga M.

    2018-01-01

    Circadian clocks consist of molecular negative feedback loops that coordinate physiological, neurological, and behavioral variables into “circa” 24-h rhythms. Rhythms in behavioral and other circadian outputs tend to weaken during aging, as evident in progressive disruptions of sleep-wake cycles in aging organisms. However, less is known about the molecular changes in the expression of clock genes and proteins that may lead to the weakening of circadian outputs. Western blot studies have demo...

  4. clockwork orange encodes a transcriptional repressor important for circadian clock amplitude in Drosophila

    OpenAIRE

    Lim, Chunghun; Chung, Brian Y.; Pitman, Jena L.; McGill, Jermaine J.; Pradhan, Suraj; Lee, Jongbin; Keegan, Kevin P.; Choe, Joonho; Allada, Ravi

    2007-01-01

    Gene transcription is a central timekeeping process in animal clocks. In Drosophila, the basic helix-loop helix (bHLH)-PAS transcription factor heterodimer, CLOCK (CLK)/CYCLE(CYC) transcriptionally activates the clock components period (per), timeless (tim), Par domain protein 1 (Pdp1), and vrille (vri) that feedback and regulate distinct features of CLK/CYC function [1]. Microarray studies have identified numerous rhythmically expressed transcripts [2-7], some of which are potential direct C...

  5. Cryptochrome mediates light-dependent magnetosensitivity of Drosophila's circadian clock.

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    Taishi Yoshii

    2009-04-01

    Full Text Available Since 1960, magnetic fields have been discussed as Zeitgebers for circadian clocks, but the mechanism by which clocks perceive and process magnetic information has remained unknown. Recently, the radical-pair model involving light-activated photoreceptors as magnetic field sensors has gained considerable support, and the blue-light photoreceptor cryptochrome (CRY has been proposed as a suitable molecule to mediate such magnetosensitivity. Since CRY is expressed in the circadian clock neurons and acts as a critical photoreceptor of Drosophila's clock, we aimed to test the role of CRY in magnetosensitivity of the circadian clock. In response to light, CRY causes slowing of the clock, ultimately leading to arrhythmic behavior. We expected that in the presence of applied magnetic fields, the impact of CRY on clock rhythmicity should be altered. Furthermore, according to the radical-pair hypothesis this response should be dependent on wavelength and on the field strength applied. We tested the effect of applied static magnetic fields on the circadian clock and found that flies exposed to these fields indeed showed enhanced slowing of clock rhythms. This effect was maximal at 300 muT, and reduced at both higher and lower field strengths. Clock response to magnetic fields was present in blue light, but absent under red-light illumination, which does not activate CRY. Furthermore, cry(b and cry(OUT mutants did not show any response, and flies overexpressing CRY in the clock neurons exhibited an enhanced response to the field. We conclude that Drosophila's circadian clock is sensitive to magnetic fields and that this sensitivity depends on light activation of CRY and on the applied field strength, consistent with the radical pair mechanism. CRY is widespread throughout biological systems and has been suggested as receptor for magnetic compass orientation in migratory birds. The present data establish the circadian clock of Drosophila as a model system

  6. Neurogenetics of Drosophila circadian clock: expect the unexpected.

    Science.gov (United States)

    Jarabo, Patricia; Martin, Francisco A

    2017-12-01

    Daily biological rhythms (i.e. circadian) are a fundamental part of animal behavior. Numerous reports have shown disruptions of the biological clock in neurodegenerative disorders and cancer. In the latter case, only recently we have gained insight into the molecular mechanisms. After 45 years of intense study of the circadian rhtythms, we find surprising similarities among species on the molecular clock that governs biological rhythms. Indeed, Drosophila is one of the most widely used models in the study of chronobiology. Recent studies in the fruit fly have revealed unpredicted roles for the clock machinery in different aspects of behavior and physiology. Not only the central pacemaker cells do have non-classical circadian functions but also circadian genes work in other cells and tissues different from central clock neurons. In this review, we summarize these new evidences. We also recapitulate the most basic features of Drosophila circadian clock, including recent data about the inputs and outputs that connect the central pacemaker with other regions of the brain. Finally, we discuss the advantages and drawbacks of using natural versus laboratory conditions.

  7. Evaluating the Autonomy of the Drosophila Circadian Clock in Dissociated Neuronal Culture.

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    Sabado, Virginie; Vienne, Ludovic; Nagoshi, Emi

    2017-01-01

    Circadian behavioral rhythms offer an excellent model to study intricate interactions between the molecular and neuronal mechanisms of behavior. In mammals, pacemaker neurons in the suprachiasmatic nucleus (SCN) generate rhythms cell-autonomously, which are synchronized by the network interactions within the circadian circuit to drive behavioral rhythms. However, whether this principle is universal to circadian systems in animals remains unanswered. Here, we examined the autonomy of the Drosophila circadian clock by monitoring transcriptional and post-transcriptional rhythms of individual clock neurons in dispersed culture with time-lapse microscopy. Expression patterns of the transcriptional reporter show that CLOCK/CYCLE (CLK/CYC)-mediated transcription is constantly active in dissociated clock neurons. In contrast, the expression profile of the post-transcriptional reporter indicates that PERIOD (PER) protein levels fluctuate and ~10% of cells display rhythms in PER levels with periods in the circadian range. Nevertheless, PER and TIM are enriched in the cytoplasm and no periodic PER nuclear accumulation was observed. These results suggest that repression of CLK/CYC-mediated transcription by nuclear PER is impaired, and thus the negative feedback loop of the molecular clock is incomplete in isolated clock neurons. We further demonstrate that, by pharmacological assays using the non-amidated form of neuropeptide pigment-dispersing factor (PDF), which could be specifically secreted from larval LNvs and adult s-LNvs, downstream events of the PDF signaling are partly impaired in dissociated larval clock neurons. Although non-amidated PDF is likely to be less active than the amidated one, these results point out the possibility that alteration in PDF downstream signaling may play a role in dampening of molecular rhythms in isolated clock neurons. Taken together, our results suggest that Drosophila clocks are weak oscillators that need to be in the intact circadian

  8. Evaluating the Autonomy of the Drosophila Circadian Clock in Dissociated Neuronal Culture

    Directory of Open Access Journals (Sweden)

    Virginie Sabado

    2017-10-01

    Full Text Available Circadian behavioral rhythms offer an excellent model to study intricate interactions between the molecular and neuronal mechanisms of behavior. In mammals, pacemaker neurons in the suprachiasmatic nucleus (SCN generate rhythms cell-autonomously, which are synchronized by the network interactions within the circadian circuit to drive behavioral rhythms. However, whether this principle is universal to circadian systems in animals remains unanswered. Here, we examined the autonomy of the Drosophila circadian clock by monitoring transcriptional and post-transcriptional rhythms of individual clock neurons in dispersed culture with time-lapse microscopy. Expression patterns of the transcriptional reporter show that CLOCK/CYCLE (CLK/CYC-mediated transcription is constantly active in dissociated clock neurons. In contrast, the expression profile of the post-transcriptional reporter indicates that PERIOD (PER protein levels fluctuate and ~10% of cells display rhythms in PER levels with periods in the circadian range. Nevertheless, PER and TIM are enriched in the cytoplasm and no periodic PER nuclear accumulation was observed. These results suggest that repression of CLK/CYC-mediated transcription by nuclear PER is impaired, and thus the negative feedback loop of the molecular clock is incomplete in isolated clock neurons. We further demonstrate that, by pharmacological assays using the non-amidated form of neuropeptide pigment-dispersing factor (PDF, which could be specifically secreted from larval LNvs and adult s-LNvs, downstream events of the PDF signaling are partly impaired in dissociated larval clock neurons. Although non-amidated PDF is likely to be less active than the amidated one, these results point out the possibility that alteration in PDF downstream signaling may play a role in dampening of molecular rhythms in isolated clock neurons. Taken together, our results suggest that Drosophila clocks are weak oscillators that need to be in the

  9. PDF and cAMP enhance PER stability in Drosophila clock neurons

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    Li, Yue; Guo, Fang; Shen, James; Rosbash, Michael

    2014-01-01

    The neuropeptide PDF is important for Drosophila circadian rhythms: pdf01 (pdf-null) animals are mostly arrhythmic or short period in constant darkness and have an advanced activity peak in light–dark conditions. PDF contributes to the amplitude, synchrony, as well as the pace of circadian rhythms within clock neurons. PDF is known to increase cAMP levels in PDR receptor (PDFR)-containing neurons. However, there is no known connection of PDF or of cAMP with the Drosophila molecular clockworks. We discovered that the mutant period gene perS ameliorates the phenotypes of pdf-null flies. The period protein (PER) is a well-studied repressor of clock gene transcription, and the perS protein (PERS) has a markedly short half-life. The result therefore suggests that the PDF-mediated increase in cAMP might lengthen circadian period by directly enhancing PER stability. Indeed, increasing cAMP levels and cAMP-mediated protein kinase A (PKA) activity stabilizes PER, in S2 tissue culture cells and in fly circadian neurons. Adding PDF to fly brains in vitro has a similar effect. Consistent with these relationships, a light pulse causes more prominent PER degradation in pdf01 circadian neurons than in wild-type neurons. The results indicate that PDF contributes to clock neuron synchrony by increasing cAMP and PKA, which enhance PER stability and decrease clock speed in intrinsically fast-paced PDFR-containing clock neurons. We further suggest that the more rapid degradation of PERS bypasses PKA regulation and makes the pace of clock neurons more uniform, allowing them to avoid much of the asynchrony caused by the absence of PDF. PMID:24707054

  10. Different Levels of Expression of the Clock Protein PER and the Glial Marker REPO in Ensheathing and Astrocyte-Like Glia of the Distal Medulla of Drosophila Optic Lobe.

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    Krzeptowski, Wojciech; Walkowicz, Lucyna; Płonczyńska, Alicja; Górska-Andrzejak, Jolanta

    2018-01-01

    Circadian plasticity of the visual system of Drosophila melanogaster depends on functioning of both the neuronal and glial oscillators. The clock function of the former is already quite well-recognized. The latter, however, is much less known and documented. In this study we focus on the glial oscillators that reside in the distal part of the second visual neuropil, medulla (dMnGl), in vicinity of the PIGMENT-DISPERSING FACTOR (PDF) releasing terminals of the circadian clock ventral Lateral Neurons (LNvs). We reveal the heterogeneity of the dMnGl, which express the clock protein PERIOD (PER) and the pan-glial marker REVERSED POLARITY (REPO) at higher (P1) or lower (P2) levels. We show that the cells with stronger expression of PER display also stronger expression of REPO, and that the number of REPO-P1 cells is bigger during the day than during the night. Using a combination of genetic markers and immunofluorescent labeling with anti PER and REPO Abs, we have established that the P1 and P2 cells can be associated with two different types of the dMnGl, the ensheathing (EnGl), and the astrocyte-like glia (ALGl). Surprisingly, the EnGl belong to the P1 cells, whereas the ALGl, previously reported to play the main role in the circadian rhythms, display the characteristics of the P2 cells (express very low level of PER and low level of REPO). Next to the EnGl and ALGl we have also observed another type of cells in the distal medulla that express PER and REPO, although at very low levels. Based on their morphology we have identified them as the T1 interneurons. Our study reveals the complexity of the distal medulla circadian network, which appears to consist of different types of glial and neuronal peripheral clocks, displaying molecular oscillations of higher (EnGl) and lower (ALGl and T1) amplitudes.

  11. A Circadian Clock Gene, Cry, Affects Heart Morphogenesis and Function in Drosophila as Revealed by Optical Coherence Microscopy.

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    Aneesh Alex

    Full Text Available Circadian rhythms are endogenous, entrainable oscillations of physical, mental and behavioural processes in response to local environmental cues such as daylight, which are present in the living beings, including humans. Circadian rhythms have been related to cardiovascular function and pathology. However, the role that circadian clock genes play in heart development and function in a whole animal in vivo are poorly understood. The Drosophila cryptochrome (dCry is a circadian clock gene that encodes a major component of the circadian clock negative feedback loop. Compared to the embryonic stage, the relative expression levels of dCry showed a significant increase (>100-fold in Drosophila during the pupa and adult stages. In this study, we utilized an ultrahigh resolution optical coherence microscopy (OCM system to perform non-invasive and longitudinal analysis of functional and morphological changes in the Drosophila heart throughout its post-embryonic lifecycle for the first time. The Drosophila heart exhibited major morphological and functional alterations during its development. Notably, heart rate (HR and cardiac activity period (CAP of Drosophila showed significant variations during the pupa stage, when heart remodeling took place. From the M-mode (2D + time OCM images, cardiac structural and functional parameters of Drosophila at different developmental stages were quantitatively determined. In order to study the functional role of dCry on Drosophila heart development, we silenced dCry by RNAi in the Drosophila heart and mesoderm, and quantitatively measured heart morphology and function in those flies throughout its development. Silencing of dCry resulted in slower HR, reduced CAP, smaller heart chamber size, pupal lethality and disrupted posterior segmentation that was related to increased expression of a posterior compartment protein, wingless. Collectively, our studies provided novel evidence that the circadian clock gene, dCry, plays

  12. A Circadian Clock Gene, Cry, Affects Heart Morphogenesis and Function in Drosophila as Revealed by Optical Coherence Microscopy

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    Zeng, Xianxu; Tate, Rebecca E.; McKee, Mary L.; Capen, Diane E.; Zhang, Zhan; Tanzi, Rudolph E.; Zhou, Chao

    2015-01-01

    Circadian rhythms are endogenous, entrainable oscillations of physical, mental and behavioural processes in response to local environmental cues such as daylight, which are present in the living beings, including humans. Circadian rhythms have been related to cardiovascular function and pathology. However, the role that circadian clock genes play in heart development and function in a whole animal in vivo are poorly understood. The Drosophila cryptochrome (dCry) is a circadian clock gene that encodes a major component of the circadian clock negative feedback loop. Compared to the embryonic stage, the relative expression levels of dCry showed a significant increase (>100-fold) in Drosophila during the pupa and adult stages. In this study, we utilized an ultrahigh resolution optical coherence microscopy (OCM) system to perform non-invasive and longitudinal analysis of functional and morphological changes in the Drosophila heart throughout its post-embryonic lifecycle for the first time. The Drosophila heart exhibited major morphological and functional alterations during its development. Notably, heart rate (HR) and cardiac activity period (CAP) of Drosophila showed significant variations during the pupa stage, when heart remodeling took place. From the M-mode (2D + time) OCM images, cardiac structural and functional parameters of Drosophila at different developmental stages were quantitatively determined. In order to study the functional role of dCry on Drosophila heart development, we silenced dCry by RNAi in the Drosophila heart and mesoderm, and quantitatively measured heart morphology and function in those flies throughout its development. Silencing of dCry resulted in slower HR, reduced CAP, smaller heart chamber size, pupal lethality and disrupted posterior segmentation that was related to increased expression of a posterior compartment protein, wingless. Collectively, our studies provided novel evidence that the circadian clock gene, dCry, plays an essential

  13. Investigation of Seasonal and Latitudinal Effects on the Expression of Clock Genes in Drosophila

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    Hosseini, Seyede Sanaz; Nazarimehr, Fahimeh; Jafari, Sajad

    The primary goal in this work is to develop a dynamical model capturing the influence of seasonal and latitudinal variations on the expression of Drosophila clock genes. To this end, we study a specific dynamical system with strange attractors that exhibit changes of Drosophila activity in a range of latitudes and across different seasons. Bifurcations of this system are analyzed to peruse the effect of season and latitude on the behavior of clock genes. Existing experimental data collected from the activity of Drosophila melanogaster corroborate the dynamical model.

  14. Dynamics of the Drosophila circadian clock: theoretical anti-jitter network and controlled chaos.

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    Hassan M Fathallah-Shaykh

    Full Text Available BACKGROUND: Electronic clocks exhibit undesirable jitter or time variations in periodic signals. The circadian clocks of humans, some animals, and plants consist of oscillating molecular networks with peak-to-peak time of approximately 24 hours. Clockwork orange (CWO is a transcriptional repressor of Drosophila direct target genes. METHODOLOGY/PRINCIPAL FINDINGS: Theory and data from a model of the Drosophila circadian clock support the idea that CWO controls anti-jitter negative circuits that stabilize peak-to-peak time in light-dark cycles (LD. The orbit is confined to chaotic attractors in both LD and dark cycles and is almost periodic in LD; furthermore, CWO diminishes the Euclidean dimension of the chaotic attractor in LD. Light resets the clock each day by restricting each molecular peak to the proximity of a prescribed time. CONCLUSIONS/SIGNIFICANCE: The theoretical results suggest that chaos plays a central role in the dynamics of the Drosophila circadian clock and that a single molecule, CWO, may sense jitter and repress it by its negative loops.

  15. Molecular clock in neutral protein evolution

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    Wilke Claus O

    2004-08-01

    Full Text Available Abstract Background A frequent observation in molecular evolution is that amino-acid substitution rates show an index of dispersion (that is, ratio of variance to mean substantially larger than one. This observation has been termed the overdispersed molecular clock. On the basis of in silico protein-evolution experiments, Bastolla and coworkers recently proposed an explanation for this observation: Proteins drift in neutral space, and can temporarily get trapped in regions of substantially reduced neutrality. In these regions, substitution rates are suppressed, which results in an overall substitution process that is not Poissonian. However, the simulation method of Bastolla et al. is representative only for cases in which the product of mutation rate μ and population size Ne is small. How the substitution process behaves when μNe is large is not known. Results Here, I study the behavior of the molecular clock in in silico protein evolution as a function of mutation rate and population size. I find that the index of dispersion decays with increasing μNe, and approaches 1 for large μNe . This observation can be explained with the selective pressure for mutational robustness, which is effective when μNe is large. This pressure keeps the population out of low-neutrality traps, and thus steadies the ticking of the molecular clock. Conclusions The molecular clock in neutral protein evolution can fall into two distinct regimes, a strongly overdispersed one for small μNe, and a mostly Poissonian one for large μNe. The former is relevant for the majority of organisms in the plant and animal kingdom, and the latter may be relevant for RNA viruses.

  16. Association between circadian clock genes and diapause incidence in Drosophila triauraria.

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    Hirokazu Yamada

    Full Text Available Diapause is an adaptive response triggered by seasonal photoperiodicity to overcome unfavorable seasons. The photoperiodic clock is a system that controls seasonal physiological processes, but our knowledge about its physiological mechanisms and genetic architecture remains incomplete. The circadian clock is another system that controls daily rhythmic physiological phenomena. It has been argued that there is a connection between the two clocks. To examine the genetic connection between them, we analyzed the associations of five circadian clock genes (period, timeless, Clock, cycle and cryptochrome with the occurrence of diapause in Drosophila triauraria, which shows a robust reproductive diapause with clear photoperiodicity. Non-diapause strains found in low latitudes were compared in genetic crosses with the diapause strain, in which the diapause trait is clearly dominant. Single nucleotide polymorphism and deletion analyses of the five circadian clock genes in backcross progeny revealed that allelic differences in timeless and cryptochrome between the strains were additively associated with the differences in the incidence of diapause. This suggests that there is a molecular link between certain circadian clock genes and the occurrence of diapause.

  17. Drosophila spaghetti and doubletime link the circadian clock and light to caspases, apoptosis and tauopathy.

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    John C Means

    2015-05-01

    Full Text Available While circadian dysfunction and neurodegeneration are correlated, the mechanism for this is not understood. It is not known if age-dependent circadian dysfunction leads to neurodegeneration or vice-versa, and the proteins that mediate the effect remain unidentified. Here, we show that the knock-down of a regulator (spag of the circadian kinase Dbt in circadian cells lowers Dbt levels abnormally, lengthens circadian rhythms and causes expression of activated initiator caspase (Dronc in the optic lobes during the middle of the day or after light pulses at night. Likewise, reduced Dbt activity lengthens circadian period and causes expression of activated Dronc, and a loss-of-function mutation in Clk also leads to expression of activated Dronc in a light-dependent manner. Genetic epistasis experiments place Dbt downstream of Spag in the pathway, and Spag-dependent reductions of Dbt are shown to require the proteasome. Importantly, activated Dronc expression due to reduced Spag or Dbt activity occurs in cells that do not express the spag RNAi or dominant negative Dbt and requires PDF neuropeptide signaling from the same neurons that support behavioral rhythms. Furthermore, reduction of Dbt or Spag activity leads to Dronc-dependent Drosophila Tau cleavage and enhanced neurodegeneration produced by human Tau in a fly eye model for tauopathy. Aging flies with lowered Dbt or Spag function show markers of cell death as well as behavioral deficits and shortened lifespans, and even old wild type flies exhibit Dbt modification and activated caspase at particular times of day. These results suggest that Dbt suppresses expression of activated Dronc to prevent Tau cleavage, and that the circadian clock defects confer sensitivity to expression of activated Dronc in response to prolonged light. They establish a link between the circadian clock factors, light, cell death pathways and Tau toxicity, potentially via dysregulation of circadian neuronal remodeling in

  18. Drosophila: An Emergent Model for Delineating Interactions between the Circadian Clock and Drugs of Abuse

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    Aliza K. De Nobrega

    2017-01-01

    Full Text Available Endogenous circadian oscillators orchestrate rhythms at the cellular, physiological, and behavioral levels across species to coordinate activity, for example, sleep/wake cycles, metabolism, and learning and memory, with predictable environmental cycles. The 21st century has seen a dramatic rise in the incidence of circadian and sleep disorders with globalization, technological advances, and the use of personal electronics. The circadian clock modulates alcohol- and drug-induced behaviors with circadian misalignment contributing to increased substance use and abuse. Invertebrate models, such as Drosophila melanogaster, have proven invaluable for the identification of genetic and molecular mechanisms underlying highly conserved processes including the circadian clock, drug tolerance, and reward systems. In this review, we highlight the contributions of Drosophila as a model system for understanding the bidirectional interactions between the circadian system and the drugs of abuse, alcohol and cocaine, and illustrate the highly conserved nature of these interactions between Drosophila and mammalian systems. Research in Drosophila provides mechanistic insights into the corresponding behaviors in higher organisms and can be used as a guide for targeted inquiries in mammals.

  19. Modelling of intercellular synchronization in the Drosophila circadian clock

    International Nuclear Information System (INIS)

    Jun-Wei, Wang; Ai-Min, Chen; Jia-Jun, Zhang; Zhan-Jiang, Yuan; Tian-Shou, Zhou

    2009-01-01

    In circadian rhythm generation, intercellular signaling factors are shown to play a crucial role in both sustaining intrinsic cellular rhythmicity and acquiring collective behaviours across a population of circadian neurons. However, the physical mechanism behind their role remains to be fully understood. In this paper, we propose an indirectly coupled multicellular model for the synchronization of Drosophila circadian oscillators combining both intracellular and intercellular dynamics. By simulating different experimental conditions, we find that such an indirect coupling way can synchronize both heterogeneous self-sustained circadian neurons and heterogeneous mutational damped circadian neurons. Moreover, they can also be entrained to ambient light-dark (LD) cycles depending on intercellular signaling. (cross-disciplinary physics and related areas of science and technology)

  20. Cell-permeable Circadian Clock Proteins

    National Research Council Canada - National Science Library

    Johnson, Carl

    2002-01-01

    .... These 'biological clocks' are important to human physiology. For example, psychiatric and medical studies have shown that circadian rhythmicity is involved in some forms of depressive illness, 'jet lag', drug tolerance/efficacy, memory, and insomnia...

  1. A computational model clarifies the roles of positive and negative feedback loops in the Drosophila circadian clock

    International Nuclear Information System (INIS)

    Wang Junwei; Zhou Tianshou

    2010-01-01

    Previous studies showed that a single negative feedback structure should be sufficient for robust circadian oscillations. It is thus pertinent to ask why current cellular clock models almost universally have interlocked negative feedback loop (NFL) and positive feedback loop (PFL). Here, we propose a molecular model that reflects the essential features of the Drosophila circadian clock to clarify the different roles of negative and positive feedback loops. In agreement with experimental observations, the model can simulate circadian oscillations in constant darkness, entrainment by light-dark cycles, as well as phenotypes of per 01 and clk Jrk mutants. Moreover, sustained oscillations persist when the PFL is removed, implying the crucial role of NFL for rhythm generation. Through parameter sensitivity analysis, it is revealed that incorporation of PFL increases the robustness of the system to regulatory processes in PFL itself. Such reduced models can aid understanding of the design principles of circadian clocks in Drosophila and other organisms with complex transcriptional feedback structures.

  2. The central molecular clock is robust in the face of behavioural arrhythmia in a Drosophila model of Alzheimer's disease.

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    Chen, Ko-Fan; Possidente, Bernard; Lomas, David A; Crowther, Damian C

    2014-04-01

    Circadian behavioural deficits, including sleep irregularity and restlessness in the evening, are a distressing early feature of Alzheimer's disease (AD). We have investigated these phenomena by studying the circadian behaviour of transgenic Drosophila expressing the amyloid beta peptide (Aβ). We find that Aβ expression results in an age-related loss of circadian behavioural rhythms despite ongoing normal molecular oscillations in the central clock neurons. Even in the absence of any behavioural correlate, the synchronised activity of the central clock remains protective, prolonging lifespan, in Aβ flies just as it does in control flies. Confocal microscopy and bioluminescence measurements point to processes downstream of the molecular clock as the main site of Aβ toxicity. In addition, there seems to be significant non-cell-autonomous Aβ toxicity resulting in morphological and probably functional signalling deficits in central clock neurons.

  3. Regulation of behavioral circadian rhythms and clock protein PER1 by the deubiquitinating enzyme USP2

    DEFF Research Database (Denmark)

    Yang, Yaoming; Duguay, David; Bédard, Nathalie

    2012-01-01

    Endogenous 24-hour rhythms are generated by circadian clocks located in most tissues. The molecular clock mechanism is based on feedback loops involving clock genes and their protein products. Post-translational modifications, including ubiquitination, are important for regulating the clock...

  4. Biases in Drosophila melanogaster protein trap screens

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    Müller Ilka

    2009-05-01

    Full Text Available Abstract Background The ability to localise or follow endogenous proteins in real time in vivo is of tremendous utility for cell biology or systems biology studies. Protein trap screens utilise the random genomic insertion of a transposon-borne artificial reporter exon (e.g. encoding the green fluorescent protein, GFP into an intron of an endogenous gene to generate a fluorescent fusion protein. Despite recent efforts aimed at achieving comprehensive coverage of the genes encoded in the Drosophila genome, the repertoire of genes that yield protein traps is still small. Results We analysed the collection of available protein trap lines in Drosophila melanogaster and identified potential biases that are likely to restrict genome coverage in protein trap screens. The protein trap screens investigated here primarily used P-element vectors and thus exhibit some of the same positional biases associated with this transposon that are evident from the comprehensive Drosophila Gene Disruption Project. We further found that protein trap target genes usually exhibit broad and persistent expression during embryonic development, which is likely to facilitate better detection. In addition, we investigated the likely influence of the GFP exon on host protein structure and found that protein trap insertions have a significant bias for exon-exon boundaries that encode disordered protein regions. 38.8% of GFP insertions land in disordered protein regions compared with only 23.4% in the case of non-trapping P-element insertions landing in coding sequence introns (p -4. Interestingly, even in cases where protein domains are predicted, protein trap insertions frequently occur in regions encoding surface exposed areas that are likely to be functionally neutral. Considering the various biases observed, we predict that less than one third of intron-containing genes are likely to be amenable to trapping by the existing methods. Conclusion Our analyses suggest that the

  5. Circadian Activators Are Expressed Days before They Initiate Clock Function in Late Pacemaker Neurons from Drosophila.

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    Liu, Tianxin; Mahesh, Guruswamy; Houl, Jerry H; Hardin, Paul E

    2015-06-03

    Circadian pacemaker neurons in the Drosophila brain control daily rhythms in locomotor activity. These pacemaker neurons can be subdivided into early or late groups depending on whether rhythms in period (per) and timeless (tim) expression are initiated at the first instar (L1) larval stage or during metamorphosis, respectively. Because CLOCK-CYCLE (CLK-CYC) heterodimers initiate circadian oscillator function by activating per and tim transcription, a Clk-GFP transgene was used to mark when late pacemaker neurons begin to develop. We were surprised to see that CLK-GFP was already expressed in four of five clusters of late pacemaker neurons during the third instar (L3) larval stage. CLK-GFP is only detected in postmitotic neurons from L3 larvae, suggesting that these four late pacemaker neuron clusters are formed before the L3 larval stage. A GFP-cyc transgene was used to show that CYC, like CLK, is also expressed exclusively in pacemaker neurons from L3 larval brains, demonstrating that CLK-CYC is not sufficient to activate per and tim in late pacemaker neurons at the L3 larval stage. These results suggest that most late pacemaker neurons develop days before novel factors activate circadian oscillator function during metamorphosis. Copyright © 2015 the authors 0270-6474/15/358662-10$15.00/0.

  6. Drosophila Protein Kinase CK2: Genetics, Regulatory Complexity and Emerging Roles during Development

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    Mohna Bandyopadhyay

    2016-12-01

    Full Text Available CK2 is a Ser/Thr protein kinase that is highly conserved amongst all eukaryotes. It is a well-known oncogenic kinase that regulates vital cell autonomous functions and animal development. Genetic studies in the fruit fly Drosophila are providing unique insights into the roles of CK2 in cell signaling, embryogenesis, organogenesis, neurogenesis, and the circadian clock, and are revealing hitherto unknown complexities in CK2 functions and regulation. Here, we review Drosophila CK2 with respect to its structure, subunit diversity, potential mechanisms of regulation, developmental abnormalities linked to mutations in the gene encoding CK2 subunits, and emerging roles in multiple aspects of eye development. We examine the Drosophila CK2 “interaction map” and the eye-specific “transcriptome” databases, which raise the prospect that this protein kinase has many additional targets in the developing eye. We discuss the possibility that CK2 functions during early retinal neurogenesis in Drosophila and mammals bear greater similarity than has been recognized, and that this conservation may extend to other developmental programs. Together, these studies underscore the immense power of the Drosophila model organism to provide new insights and avenues to further investigate developmentally relevant targets of this protein kinase.

  7. The Pyrexia transient receptor potential channel mediates circadian clock synchronization to low temperature cycles in Drosophila melanogaster.

    Science.gov (United States)

    Wolfgang, Werner; Simoni, Alekos; Gentile, Carla; Stanewsky, Ralf

    2013-10-07

    Circadian clocks are endogenous approximately 24 h oscillators that temporally regulate many physiological and behavioural processes. In order to be beneficial for the organism, these clocks must be synchronized with the environmental cycles on a daily basis. Both light : dark and the concomitant daily temperature cycles (TCs) function as Zeitgeber ('time giver') and efficiently entrain circadian clocks. The temperature receptors mediating this synchronization have not been identified. Transient receptor potential (TRP) channels function as thermo-receptors in animals, and here we show that the Pyrexia (Pyx) TRP channel mediates temperature synchronization in Drosophila melanogaster. Pyx is expressed in peripheral sensory organs (chordotonal organs), which previously have been implicated in temperature synchronization. Flies deficient for Pyx function fail to synchronize their behaviour to TCs in the lower range (16-20°C), and this deficit can be partially rescued by introducing a wild-type copy of the pyx gene. Synchronization to higher TCs is not affected, demonstrating a specific role for Pyx at lower temperatures. In addition, pyx mutants speed up their clock after being exposed to TCs. Our results identify the first TRP channel involved in temperature synchronization of circadian clocks.

  8. Oscillating perceptions: the ups and downs of the CLOCK protein in ...

    Indian Academy of Sciences (India)

    2008-12-31

    Dec 31, 2008 ... A functional mouse CLOCK protein has long been thought to be essential for mammalian circadian ... ordinated actions of the Ror families of transcriptional acti- ..... CLOCK-deficient mice expressing the mPER2::LUC fusion.

  9. Tet protein function during Drosophila development.

    Directory of Open Access Journals (Sweden)

    Fei Wang

    Full Text Available The TET (Ten-eleven translocation 1, 2 and 3 proteins have been shown to function as DNA hydroxymethylases in vertebrates and their requirements have been documented extensively. Recently, the Tet proteins have been shown to also hydroxylate 5-methylcytosine in RNA. 5-hydroxymethylcytosine (5hmrC is enriched in messenger RNA but the function of this modification has yet to be elucidated. Because Cytosine methylation in DNA is barely detectable in Drosophila, it serves as an ideal model to study the biological function of 5hmrC. Here, we characterized the temporal and spatial expression and requirement of Tet throughout Drosophila development. We show that Tet is essential for viability as Tet complete loss-of-function animals die at the late pupal stage. Tet is highly expressed in neuronal tissues and at more moderate levels in somatic muscle precursors in embryos and larvae. Depletion of Tet in muscle precursors at early embryonic stages leads to defects in larval locomotion and late pupal lethality. Although Tet knock-down in neuronal tissue does not cause lethality, it is essential for neuronal function during development through its affects upon locomotion in larvae and the circadian rhythm of adult flies. Further, we report the function of Tet in ovarian morphogenesis. Together, our findings provide basic insights into the biological function of Tet in Drosophila, and may illuminate observed neuronal and muscle phenotypes observed in vertebrates.

  10. A computational model clarifies the roles of positive and negative feedback loops in the Drosophila circadian clock

    Energy Technology Data Exchange (ETDEWEB)

    Wang Junwei, E-mail: wangjunweilj@yahoo.com.c [Cisco School of Informatics, Guangdong University of Foreign Studies, Guangzhou 510006 (China); Zhou Tianshou [School of Mathematics and Computational Science, Sun Yat-Sen University, Guangzhou 510275 (China)

    2010-06-14

    Previous studies showed that a single negative feedback structure should be sufficient for robust circadian oscillations. It is thus pertinent to ask why current cellular clock models almost universally have interlocked negative feedback loop (NFL) and positive feedback loop (PFL). Here, we propose a molecular model that reflects the essential features of the Drosophila circadian clock to clarify the different roles of negative and positive feedback loops. In agreement with experimental observations, the model can simulate circadian oscillations in constant darkness, entrainment by light-dark cycles, as well as phenotypes of per{sup 01} and clk{sup Jrk} mutants. Moreover, sustained oscillations persist when the PFL is removed, implying the crucial role of NFL for rhythm generation. Through parameter sensitivity analysis, it is revealed that incorporation of PFL increases the robustness of the system to regulatory processes in PFL itself. Such reduced models can aid understanding of the design principles of circadian clocks in Drosophila and other organisms with complex transcriptional feedback structures.

  11. Drosophila Clock Is Required in Brain Pacemaker Neurons to Prevent Premature Locomotor Aging Independently of Its Circadian Function.

    Directory of Open Access Journals (Sweden)

    Alexandra Vaccaro

    2017-01-01

    Full Text Available Circadian clocks control many self-sustained rhythms in physiology and behavior with approximately 24-hour periodicity. In many organisms, oxidative stress and aging negatively impact the circadian system and sleep. Conversely, loss of the clock decreases resistance to oxidative stress, and may reduce lifespan and speed up brain aging and neurodegeneration. Here we examined the effects of clock disruptions on locomotor aging and longevity in Drosophila. We found that lifespan was similarly reduced in three arrhythmic mutants (ClkAR, cyc0 and tim0 and in wild-type flies under constant light, which stops the clock. In contrast, ClkAR mutants showed significantly faster age-related locomotor deficits (as monitored by startle-induced climbing than cyc0 and tim0, or than control flies under constant light. Reactive oxygen species accumulated more with age in ClkAR mutant brains, but this did not appear to contribute to the accelerated locomotor decline of the mutant. Clk, but not Cyc, inactivation by RNA interference in the pigment-dispersing factor (PDF-expressing central pacemaker neurons led to similar loss of climbing performance as ClkAR. Conversely, restoring Clk function in these cells was sufficient to rescue the ClkAR locomotor phenotype, independently of behavioral rhythmicity. Accelerated locomotor decline of the ClkAR mutant required expression of the PDF receptor and correlated to an apparent loss of dopaminergic neurons in the posterior protocerebral lateral 1 (PPL1 clusters. This neuronal loss was rescued when the ClkAR mutation was placed in an apoptosis-deficient background. Impairing dopamine synthesis in a single pair of PPL1 neurons that innervate the mushroom bodies accelerated locomotor decline in otherwise wild-type flies. Our results therefore reveal a novel circadian-independent requirement for Clk in brain circadian neurons to maintain a subset of dopaminergic cells and avoid premature locomotor aging in Drosophila.

  12. On Variations in the Level of PER in Glial Clocks of Drosophila Optic Lobe and Its Negative Regulation by PDF Signaling.

    Science.gov (United States)

    Górska-Andrzejak, Jolanta; Chwastek, Elżbieta M; Walkowicz, Lucyna; Witek, Kacper

    2018-01-01

    We show that the level of the core protein of the circadian clock Period (PER) expressed by glial peripheral oscillators depends on their location in the Drosophila optic lobe. It appears to be controlled by the ventral lateral neurons (LNvs) that release the circadian neurotransmitter Pigment Dispersing Factor (PDF). We demonstrate that glial cells of the distal medulla neuropil (dMnGl) that lie in the vicinity of the PDF-releasing terminals of the LNvs possess receptors for PDF (PDFRs) and express PER at significantly higher level than other types of glia. Surprisingly, the amplitude of PER molecular oscillations in dMnGl is increased twofold in PDF-free environment, that is in Pdf 0 mutants. The Pdf 0 mutants also reveal an increased level of glia-specific protein REPO in dMnGl. The photoreceptors of the compound eye (R-cells) of the PDF-null flies, on the other hand, exhibit de-synchrony of PER molecular oscillations, which manifests itself as increased variability of PER-specific immunofluorescence among the R-cells. Moreover, the daily pattern of expression of the presynaptic protein Bruchpilot (BRP) in the lamina terminals of the R-cells is changed in Pdf 0 mutant. Considering that PDFRs are also expressed by the marginal glia of the lamina that surround the R-cell terminals, the LNv pacemakers appear to be the likely modulators of molecular cycling in the peripheral clocks of both the glial cells and the photoreceptors of the compound eye. Consequently, some form of PDF-based coupling of the glial clocks and the photoreceptors of the eye with the central LNv pacemakers must be operational.

  13. Temporal requirements of the fragile X mental retardation protein in modulating circadian clock circuit synaptic architecture

    Directory of Open Access Journals (Sweden)

    Cheryl L Gatto

    2009-08-01

    Full Text Available Loss of fragile X mental retardation 1 (FMR1 gene function is the most common cause of inherited mental retardation and autism spectrum disorders, characterized by attention disorder, hyperactivity and disruption of circadian activity cycles. Pursuit of effective intervention strategies requires determining when the FMR1 product (FMRP is required in the regulation of neuronal circuitry controlling these behaviors. In the well-characterized Drosophila disease model, loss of the highly conserved dFMRP causes circadian arrhythmicity and conspicuous abnormalities in the circadian clock circuitry. Here, a novel Sholl Analysis was used to quantify over-elaborated synaptic architecture in dfmr1-null small ventrolateral neurons (sLNvs, a key subset of clock neurons. The transgenic Gene-Switch system was employed to drive conditional neuronal dFMRP expression in the dfmr1-null mutant background in order to dissect temporal requirements within the clock circuit. Introduction of dFMRP during early brain development, including the stages of neurogenesis, neuronal fate specification and early pathfinding, provided no rescue of dfmr1 mutant phenotypes. Similarly, restoring normal dFMRP expression in the adult failed to restore circadian circuit architecture. In sharp contrast, supplying dFMRP during a transient window of very late brain development, wherein synaptogenesis and substantial subsequent synaptic reorganization (e.g. use-dependent pruning occur, provided strong morphological rescue to reestablish normal sLNvs synaptic arbors. We conclude that dFMRP plays a developmentally restricted role in sculpting synaptic architecture in these neurons that cannot be compensated for by later reintroduction of the protein at maturity.

  14. Regulation of behavioral circadian rhythms and clock protein PER1 by the deubiquitinating enzyme USP2

    Directory of Open Access Journals (Sweden)

    Yaoming Yang

    2012-06-01

    Endogenous 24-hour rhythms are generated by circadian clocks located in most tissues. The molecular clock mechanism is based on feedback loops involving clock genes and their protein products. Post-translational modifications, including ubiquitination, are important for regulating the clock feedback mechanism. Previous work has focused on the role of ubiquitin ligases in the clock mechanism. Here we show a role for the rhythmically-expressed deubiquitinating enzyme ubiquitin specific peptidase 2 (USP2 in clock function. Mice with a deletion of the Usp2 gene (Usp2 KO display a longer free-running period of locomotor activity rhythms and altered responses of the clock to light. This was associated with altered expression of clock genes in synchronized Usp2 KO mouse embryonic fibroblasts and increased levels of clock protein PERIOD1 (PER1. USP2 can be coimmunoprecipitated with several clock proteins but directly interacts specifically with PER1 and deubiquitinates it. Interestingly, this deubiquitination does not alter PER1 stability. Taken together, our results identify USP2 as a new core component of the clock machinery and demonstrate a role for deubiquitination in the regulation of the circadian clock, both at the level of the core pacemaker and its response to external cues.

  15. Reciprocal cholinergic and GABAergic modulation of the small ventrolateral pacemaker neurons of Drosophila's circadian clock neuron network.

    Science.gov (United States)

    Lelito, Katherine R; Shafer, Orie T

    2012-04-01

    The relatively simple clock neuron network of Drosophila is a valuable model system for the neuronal basis of circadian timekeeping. Unfortunately, many key neuronal classes of this network are inaccessible to electrophysiological analysis. We have therefore adopted the use of genetically encoded sensors to address the physiology of the fly's circadian clock network. Using genetically encoded Ca(2+) and cAMP sensors, we have investigated the physiological responses of two specific classes of clock neuron, the large and small ventrolateral neurons (l- and s-LN(v)s), to two neurotransmitters implicated in their modulation: acetylcholine (ACh) and γ-aminobutyric acid (GABA). Live imaging of l-LN(v) cAMP and Ca(2+) dynamics in response to cholinergic agonist and GABA application were well aligned with published electrophysiological data, indicating that our sensors were capable of faithfully reporting acute physiological responses to these transmitters within single adult clock neuron soma. We extended these live imaging methods to s-LN(v)s, critical neuronal pacemakers whose physiological properties in the adult brain are largely unknown. Our s-LN(v) experiments revealed the predicted excitatory responses to bath-applied cholinergic agonists and the predicted inhibitory effects of GABA and established that the antagonism of ACh and GABA extends to their effects on cAMP signaling. These data support recently published but physiologically untested models of s-LN(v) modulation and lead to the prediction that cholinergic and GABAergic inputs to s-LN(v)s will have opposing effects on the phase and/or period of the molecular clock within these critical pacemaker neurons.

  16. Molecular cogs of the insect circadian clock.

    Science.gov (United States)

    Shirasu, Naoto; Shimohigashi, Yasuyuki; Tominaga, Yoshiya; Shimohigashi, Miki

    2003-08-01

    During the last five years, enormous progress has been made in understanding the molecular basis of circadian systems, mainly by molecular genetic studies using the mouse and fly. Extensive evidence has revealed that the core clock machinery involves "clock genes" and "clock proteins" functioning as molecular cogs. These participate in transcriptional/translational feedback loops and many homologous clock-components in the fruit fly Drosophila are also expressed in mammalian clock tissues with circadian rhythms. Thus, the mechanisms of the central clock seem to be conserved across animal kingdom. However, some recent studies imply that the present widely accepted molecular models of circadian clocks may not always be supported by the experimental evidence.

  17. Pigment-dispersing factor modulates pheromone production in clock cells that influence mating in Drosophila

    NARCIS (Netherlands)

    Krupp, Joshua J.; Billeter, Jean-Christophe; Wong, Amy; Choi, Charles; Nitabach, Michael N.; Levine, Joel D.

    2013-01-01

    Social cues contribute to the circadian entrainment of physiological and behavioral rhythms. These cues supplement the influence of daily and seasonal cycles in light and temperature. In Drosophila, the social environment modulates circadian mechanisms that regulate sex pheromone production and

  18. A central role for ubiquitination within a circadian clock protein modification code

    Directory of Open Access Journals (Sweden)

    Katarina eStojkovic

    2014-08-01

    Full Text Available Circadian rhythms, endogenous cycles of about 24 h in physiology, are generated by a master clock located in the suprachiasmatic nucleus of the hypothalamus and other clocks located in the brain and peripheral tissues. Circadian disruption is known to increase the incidence of various illnesses, such as mental disorders, metabolic syndrome and cancer. At the molecular level, periodicity is established by a set of clock genes via autoregulatory translation-transcription feedback loops. This clock mechanism is regulated by post-translational modifications such as phosphorylation and ubiquitination, which set the pace of the clock. Ubiquitination in particular has been found to regulate the stability of core clock components, but also other clock protein functions. Mutation of genes encoding ubiquitin ligases can cause either elongation or shortening of the endogenous circadian period. Recent research has also started to uncover roles for deubiquitination in the molecular clockwork. Here we review the role of the ubiquitin pathway in regulating the circadian clock and we propose that ubiquitination is a key element in a clock protein modification code that orchestrates clock mechanisms and circadian behavior over the daily cycle.

  19. Distribution of DNA replication proteins in Drosophila cells

    Science.gov (United States)

    Easwaran, Hariharan P; Leonhardt, Heinrich; Cardoso, M Cristina

    2007-01-01

    Background DNA replication in higher eukaryotic cells is organized in discrete subnuclear sites called replication foci (RF). During the S phase, most replication proteins assemble at the RF by interacting with PCNA via a PCNA binding domain (PBD). This has been shown to occur for many mammalian replication proteins, but it is not known whether this mechanism is conserved in evolution. Results Fluorescent fusions of mammalian replication proteins, Dnmt1, HsDNA Lig I and HsPCNA were analyzed for their ability to target to RF in Drosophila cells. Except for HsPCNA, none of the other proteins and their deletions showed any accumulation at RF in Drosophila cells. We hypothesized that in Drosophila cells there might be some other peptide sequence responsible for targeting proteins to RF. To test this, we identified the DmDNA Lig I and compared the protein sequence with HsDNA Lig I. The two orthologs shared the PBD suggesting a functionally conserved role for this domain in the Drosophila counterpart. A series of deletions of DmDNA Lig I were analyzed for their ability to accumulate at RF in Drosophila and mammalian cells. Surprisingly, no accumulation at RF was observed in Drosophila cells, while in mammalian cells DmDNA Lig I accumulated at RF via its PBD. Further, GFP fusions with the PBD domains from Dnmt1, HsDNA Lig I and DmDNA Lig I, were able to target to RF only in mammalian cells but not in Drosophila cells. Conclusion We show that S phase in Drosophila cells is characterized by formation of RF marked by PCNA like in mammalian cells. However, other than PCNA none of the replication proteins and their deletions tested here showed accumulation at RF in Drosophila cells while the same proteins and deletions are capable of accumulating at RF in mammalian cells. We hypothesize that unlike mammalian cells, in Drosophila cells, replication proteins do not form long-lasting interactions with the replication machinery, and rather perform their functions via very

  20. Evolution of robust circadian clocks in Drosophila melanogaster populations reared in constant dark for over 330 generations

    Science.gov (United States)

    Shindey, Radhika; Varma, Vishwanath; Nikhil, K. L.; Sharma, Vijay Kumar

    2016-10-01

    Robustness is considered to be an important feature of biological systems which may evolve when the functionality of a trait is associated with higher fitness across multiple environmental conditions. Thus, the ability to maintain stable biological phenotypes across environments is thought to be of adaptive value. Previously, we have reported higher intrinsic activity levels (activity levels of free-running rhythm in constant darkness) and power of rhythm (as assessed by amplitude of the periodogram) in Drosophila melanogaster populations (stocks) reared in constant darkness (DD stocks) as compared to those reared in constant light (LL stocks) and 12:12-h light-dark cycles (LD stocks) for over 19 years (˜330 generations). In the current study, we intended to examine whether the enhanced levels of activity observed in DD stocks persist under various environments such as photoperiods, ambient temperatures, non-24-h light-dark (LD) cycles, and semi-natural conditions (SN). We found that DD stocks largely retain their phenotype of enhanced activity levels across most of the above-mentioned environments suggesting the evolution of robust circadian clocks in DD stocks. Furthermore, we compared the peak activity levels of the three stocks across different environmental conditions relative to their peaks in constant darkness and found that the change in peak activity levels upon entrainment was not significantly different across the three stocks for any of the examined environmental conditions. This suggests that the enhancement of activity levels in DD stocks is not due to differential sensitivity to environment. Thus, these results suggest that rearing in constant darkness (DD) leads to evolution of robust circadian clocks suggesting a possible adaptive value of possessing such rhythms under constant dark environments.

  1. Drosophila DNA-Binding Proteins in Polycomb Repression

    Directory of Open Access Journals (Sweden)

    Maksim Erokhin

    2018-01-01

    Full Text Available The formation of individual gene expression patterns in different cell types is required during differentiation and development of multicellular organisms. Polycomb group (PcG proteins are key epigenetic regulators responsible for gene repression, and dysregulation of their activities leads to developmental abnormalities and diseases. PcG proteins were first identified in Drosophila, which still remains the most convenient system for studying PcG-dependent repression. In the Drosophila genome, these proteins bind to DNA regions called Polycomb response elements (PREs. A major role in the recruitment of PcG proteins to PREs is played by DNA-binding factors, several of which have been characterized in detail. However, current knowledge is insufficient for comprehensively describing the mechanism of this process. In this review, we summarize and discuss the available data on the role of DNA-binding proteins in PcG recruitment to chromatin.

  2. Genetic analysis of a Drosophila microtubule-associated protein

    OpenAIRE

    1992-01-01

    The 205-kD microtubule-associated protein (205K MAP) is one of the principal MAPs in Drosophila. 205K MAP is similar to the HeLa 210K/MAP4 family of MAPs since it shares the following biochemical properties: it is present in several isoforms, has a molecular mass of approximately 200 kD, and is thermostable. Furthermore, immuno-crossreactivity has been observed between mouse MAP4, HeLa 210K, and Drosophila 205K MAP. Currently, there is little information concerning the biological function of ...

  3. A Drosophila gene encoding a protein resembling the human β-amyloid protein precursor

    International Nuclear Information System (INIS)

    Rosen, D.R.; Martin-Morris, L.; Luo, L.; White, K.

    1989-01-01

    The authors have isolated genomic and cDNA clones for a Drosophila gene resembling the human β-amyloid precursor protein (APP). This gene produces a nervous system-enriched 6.5-kilobase transcript. Sequencing of cDNAs derived from the 6.5-kilobase transcript predicts an 886-amino acid polypeptide. This polypeptide contains a putative transmembrane domain and exhibits strong sequence similarity to cytoplasmic and extracellular regions of the human β-amyloid precursor protein. There is a high probability that this Drosophila gene corresponds to the essential Drosophila locus vnd, a gene required for embryonic nervous system development

  4. Drosophila Protein interaction Map (DPiM)

    OpenAIRE

    Guruharsha, K.G.; Obar, Robert A.; Mintseris, Julian; Aishwarya, K.; Krishnan, R.T.; VijayRaghavan, K.; Artavanis-Tsakonas, Spyros

    2012-01-01

    Proteins perform essential cellular functions as part of protein complexes, often in conjunction with RNA, DNA, metabolites and other small molecules. The genome encodes thousands of proteins but not all of them are expressed in every cell type; and expressed proteins are not active at all times. Such diversity of protein expression and function accounts for the level of biological intricacy seen in nature. Defining protein-protein interactions in protein complexes, and establishing the when,...

  5. Adaptive evolution of relish, a Drosophila NF-kappaB/IkappaB protein.

    OpenAIRE

    Begun, D J; Whitley, P

    2000-01-01

    NF-kappaB and IkappaB proteins have central roles in regulation of inflammation and innate immunity in mammals. Homologues of these proteins also play an important role in regulation of the Drosophila immune response. Here we present a molecular population genetic analysis of Relish, a Drosophila NF-kappaB/IkappaB protein, in Drosophila simulans and D. melanogaster. We find strong evidence for adaptive protein evolution in D. simulans, but not in D. melanogaster. The adaptive evolution appear...

  6. Developmental expression of Drosophila Wiskott-Aldrich Syndrome family proteins

    Science.gov (United States)

    Rodriguez-Mesa, Evelyn; Abreu-Blanco, Maria Teresa; Rosales-Nieves, Alicia E.; Parkhurst, Susan M.

    2012-01-01

    Background Wiskott-Aldrich Syndrome (WASP) family proteins participate in many cellular processes involving rearrangements of the actin cytoskeleton. To the date, four WASP subfamily members have been described in Drosophila: Wash, WASp, SCAR, and Whamy. Wash, WASp, and SCAR are essential during early Drosophila development where they function in orchestrating cytoplasmic events including membrane-cytoskeleton interactions. A mutant for Whamy has not yet been reported. Results We generated monoclonal antibodies that are specific to Drosophila Wash, WASp, SCAR, and Whamy, and use these to describe their spatial and temporal localization patterns. Consistent with the importance of WASP family proteins in flies, we find that Wash, WASp, SCAR, and Whamy are dynamically expressed throughout oogenesis and embryogenesis. For example, we find that Wash accumulates at the oocyte cortex. WASp is highly expressed in the PNS, while SCAR is the most abundantly expressed in the CNS. Whamy exhibits an asymmetric subcellular localization that overlaps with mitochondria and is highly expressed in muscle. Conclusion All four WASP family members show specific expression patterns, some of which reflect their previously known roles and others revealing new potential functions. The monoclonal antibodies developed offer valuable new tools to investigate how WASP family proteins regulate actin cytoskeleton dynamics. PMID:22275148

  7. Light at night alters daily patterns of cortisol and clock proteins in female Siberian hamsters.

    Science.gov (United States)

    Bedrosian, T A; Galan, A; Vaughn, C A; Weil, Z M; Nelson, R J

    2013-06-01

    Humans and other organisms have adapted to a 24-h solar cycle in response to life on Earth. The rotation of the planet on its axis and its revolution around the sun cause predictable daily and seasonal patterns in day length. To successfully anticipate and adapt to these patterns in the environment, a variety of biological processes oscillate with a daily rhythm of approximately 24 h in length. These rhythms arise from hierarchally-coupled cellular clocks generated by positive and negative transcription factors of core circadian clock gene expression. From these endogenous cellular clocks, overt rhythms in activity and patterns in hormone secretion and other homeostatic processes emerge. These circadian rhythms in physiology and behaviour can be organised by a variety of cues, although they are most potently entrained by light. In recent history, there has been a major change from naturally-occurring light cycles set by the sun, to artificial and sometimes erratic light cycles determined by the use of electric lighting. Virtually every individual living in an industrialised country experiences light at night (LAN) but, despite its prevalence, the biological effects of such unnatural lighting have not been fully considered. Using female Siberian hamsters (Phodopus sungorus), we investigated the effects of chronic nightly exposure to dim light on daily rhythms in locomotor activity, serum cortisol concentrations and brain expression of circadian clock proteins (i.e. PER1, PER2, BMAL1). Although locomotor activity remained entrained to the light cycle, the diurnal fluctuation of cortisol concentrations was blunted and the expression patterns of clock proteins in the suprachiasmatic nucleus and hippocampus were altered. These results demonstrate that chronic exposure to dim LAN can dramatically affect fundamental cellular function and emergent physiology. © 2013 British Society for Neuroendocrinology.

  8. Photoperiodic plasticity in circadian clock neurons in insects

    Directory of Open Access Journals (Sweden)

    Sakiko eShiga

    2013-08-01

    Full Text Available Since Bünning’s observation of circadian rhythms and photoperiodism in the runner bean Phaseolus multiflorus in 1936, many studies have shown that photoperiodism is based on the circadian clock system. In insects, involvement of circadian clock genes or neurons has been recently shown in the photoperiodic control of developmental arrests, diapause. Based on molecular and neuronal studies in Drosophila melanogaster, photoperiodic changes have been reported for expression patterns of the circadian clock genes, subcellular distribution of clock proteins, fiber distribution, or the number of plausible clock neurons in different species. Photoperiod sets peaks of per or tim mRNA abundance at lights-off in Sarcophaga crassipalpis, Chymomyza costata and Protophormia terraenovae. Abundance of per and Clock mRNA changes by photoperiod in Pyrrhocoris apterus. Subcellular Per distribution in circadian clock neurons changes with photoperiod in P. terraenovae. Although photoperiodism is not known in Leucophaea maderae, under longer day length, more stomata and longer commissural fibers of circadian clock neurons have been found. These plastic changes in the circadian clock neurons could be an important constituent for photoperiodic clock mechanisms to integrate repetitive photoperiodic information and produce different outputs based on day length.

  9. Lipoic acid entrains the hepatic circadian clock and lipid metabolic proteins that have been desynchronized with advanced age

    International Nuclear Information System (INIS)

    Keith, Dove; Finlay, Liam; Butler, Judy; Gómez, Luis; Smith, Eric; Moreau, Régis; Hagen, Tory

    2014-01-01

    Highlights: • 24 month old rats were supplemented with 0.2% lipoic acid in the diet for 2 weeks. • Lipoic acid shifts phase of core circadian clock proteins. • Lipoic acid corrects age-induced desynchronized lipid metabolism rhythms. - Abstract: It is well established that lipid metabolism is controlled, in part, by circadian clocks. However, circadian clocks lose temporal precision with age and correlates with elevated incidence in dyslipidemia and metabolic syndrome in older adults. Because our lab has shown that lipoic acid (LA) improves lipid homeostasis in aged animals, we hypothesized that LA affects the circadian clock to achieve these results. We fed 24 month old male F344 rats a diet supplemented with 0.2% (w/w) LA for 2 weeks prior to sacrifice and quantified hepatic circadian clock protein levels and clock-controlled lipid metabolic enzymes. LA treatment caused a significant phase-shift in the expression patterns of the circadian clock proteins Period (Per) 2, Brain and Muscle Arnt-Like1 (BMAL1), and Reverse Erythroblastosis virus (Rev-erb) β without altering the amplitude of protein levels during the light phase of the day. LA also significantly altered the oscillatory patterns of clock-controlled proteins associated with lipid metabolism. The level of peroxisome proliferator-activated receptor (PPAR) α was significantly increased and acetyl-CoA carboxylase (ACC) and fatty acid synthase (FAS) were both significantly reduced, suggesting that the LA-supplemented aged animals are in a catabolic state. We conclude that LA remediates some of the dyslipidemic processes associated with advanced age, and this mechanism may be at least partially through entrainment of circadian clocks

  10. Lipoic acid entrains the hepatic circadian clock and lipid metabolic proteins that have been desynchronized with advanced age

    Energy Technology Data Exchange (ETDEWEB)

    Keith, Dove; Finlay, Liam; Butler, Judy [Linus Pauling Institute, Oregon State University (United States); Gómez, Luis; Smith, Eric [Linus Pauling Institute, Oregon State University (United States); Biochemistry Biophysics Department, Oregon State University (United States); Moreau, Régis [Linus Pauling Institute, Oregon State University (United States); Hagen, Tory [Linus Pauling Institute, Oregon State University (United States); Biochemistry Biophysics Department, Oregon State University (United States)

    2014-07-18

    Highlights: • 24 month old rats were supplemented with 0.2% lipoic acid in the diet for 2 weeks. • Lipoic acid shifts phase of core circadian clock proteins. • Lipoic acid corrects age-induced desynchronized lipid metabolism rhythms. - Abstract: It is well established that lipid metabolism is controlled, in part, by circadian clocks. However, circadian clocks lose temporal precision with age and correlates with elevated incidence in dyslipidemia and metabolic syndrome in older adults. Because our lab has shown that lipoic acid (LA) improves lipid homeostasis in aged animals, we hypothesized that LA affects the circadian clock to achieve these results. We fed 24 month old male F344 rats a diet supplemented with 0.2% (w/w) LA for 2 weeks prior to sacrifice and quantified hepatic circadian clock protein levels and clock-controlled lipid metabolic enzymes. LA treatment caused a significant phase-shift in the expression patterns of the circadian clock proteins Period (Per) 2, Brain and Muscle Arnt-Like1 (BMAL1), and Reverse Erythroblastosis virus (Rev-erb) β without altering the amplitude of protein levels during the light phase of the day. LA also significantly altered the oscillatory patterns of clock-controlled proteins associated with lipid metabolism. The level of peroxisome proliferator-activated receptor (PPAR) α was significantly increased and acetyl-CoA carboxylase (ACC) and fatty acid synthase (FAS) were both significantly reduced, suggesting that the LA-supplemented aged animals are in a catabolic state. We conclude that LA remediates some of the dyslipidemic processes associated with advanced age, and this mechanism may be at least partially through entrainment of circadian clocks.

  11. Adaptive response in Drosophila melanogaster heat shock proteins mutant strains

    International Nuclear Information System (INIS)

    Shaposhnikov, M.V.; Moskalev, A.A.; Turysheva, E.V.

    2007-01-01

    Complete text of publication follows. The members of the heat shock proteins (Hsp) family function as molecular chaperones and assist intracellular folding of newly synthesized proteins. Also it is possible that molecular chaperones are induced during adaptive response to oxidative stress and radiation. The aim of our research was to exam the role of heat shock proteins in adaptive response to oxidative stress after low dose rate gamma-irradiation in Drosophila melanogaster. Drosophilamelanogaster strains were kindly provided by Bloomington Drosophila Stock Center (University of state of Indiana, Bloomington, USA). We used wild type strain (CS), heat shock protein mutant strains (Hsp22, Hsp70, Hsp83), and heat shock factor mutant strain (Hsf). Strains were chronically exposured to adaptive dose of gamma-irradiation in dose rate of 0.17 cGy/h during all stages of life history (from the embrional stage to the stage of matured imago). The rate of absorbed dose was 60 cGy. For oxidative-stress challenge twodays old flies were starved in empty vials for 6 h and then transferred to vials containing only filter paper soaked with 20 mM paraquat in 5% sucrose solution. Survival data were collected after 26 h of treatment. Dead flies were counted daily. The obtained data were subjected to survival analysis by Kaplan and Meier method and presented as survival curves. Statistical analysis was held by non-parametric methods. To test the significance of the difference between the two age distributions Kolmogorov-Smirnov test was applied. Gehan-Braslow- Wilcoxon and Cox-Mantel tests were used for estimation of median life span differences. In addition the minimal and maximal life span, time of 90% death, and mortality rate doubling time (MRDT) were estimated. The obtained results will be discussed in presentation.

  12. Identification and characterization of proteins involved in nuclear organization using Drosophila GFP protein trap lines.

    Directory of Open Access Journals (Sweden)

    Margaret Rohrbaugh

    Full Text Available Strains from a collection of Drosophila GFP protein trap lines express GFP in the normal tissues where the endogenous protein is present. This collection can be used to screen for proteins distributed in the nucleus in a non-uniform pattern.We analyzed four lines that show peripheral or punctate nuclear staining. One of these lines affects an uncharacterized gene named CG11138. The CG11138 protein shows a punctate distribution in the nuclear periphery similar to that of Drosophila insulator proteins but does not co-localize with known insulators. Interestingly, mutations in Lamin proteins result in alterations in CG11138 localization, suggesting that this protein may be a novel component of the nuclear lamina. A second line affects the Decondensation factor 31 (Df31 gene, which encodes a protein with a unique nuclear distribution that appears to segment the nucleus into four different compartments. The X-chromosome of males is confined to one of these compartments. We also find that Drosophila Nucleoplasmin (dNlp is present in regions of active transcription. Heat shock leads to loss of dNlp from previously transcribed regions of polytene chromosome without redistribution to the heat shock genes. Analysis of Stonewall (Stwl, a protein previously found to be necessary for the maintenance of germline stem cells, shows that Stwl is present in a punctate pattern in the nucleus that partially overlaps with that of known insulator proteins. Finally we show that Stwl, dNlp, and Df31 form part of a highly interactive network. The properties of other components of this network may help understand the role of these proteins in nuclear biology.These results establish screening of GFP protein trap alleles as a strategy to identify factors with novel cellular functions. Information gained from the analysis of CG11138 Stwl, dNlp, and Df31 sets the stage for future studies of these proteins.

  13. A Drosophila protein-tyrosine phosphatase associates with an adapter protein required for axonal guidance.

    Science.gov (United States)

    Clemens, J C; Ursuliak, Z; Clemens, K K; Price, J V; Dixon, J E

    1996-07-19

    We have used the yeast two-hybrid system to isolate a novel Drosophila adapter protein, which interacts with the Drosophila protein-tyrosine phosphatase (PTP) dPTP61F. Absence of this protein in Drosophila causes the mutant photoreceptor axon phenotype dreadlocks (dock) (Garrity, P. A., Rao, Y., Salecker, I., and Zipursky, S. L.(1996) Cell 85, 639-650). Dock is similar to the mammalian oncoprotein Nck and contains three Src homology 3 (SH3) domains and one Src homology 2 (SH2) domain. The interaction of dPTP61F with Dock was confirmed in vivo by immune precipitation experiments. A sequence containing five PXXP motifs from the non-catalytic domain of the PTP is sufficient for interaction with Dock. This suggests that binding to the PTP is mediated by one or more of the SH3 domains of Dock. Immune precipitations of Dock also co-precipitate two tyrosine-phosphorylated proteins having molecular masses of 190 and 145 kDa. Interactions between Dock and these tyrosine-phosphorylated proteins are likely mediated by the Dock SH2 domain. These findings identify potential signal-transducing partners of Dock and propose a role for dPTP61F and the unidentified phosphoproteins in axonal guidance.

  14. Drosophila Vps13 Is Required for Protein Homeostasis in the Brain.

    Directory of Open Access Journals (Sweden)

    Jan J Vonk

    Full Text Available Chorea-Acanthocytosis is a rare, neurodegenerative disorder characterized by progressive loss of locomotor and cognitive function. It is caused by loss of function mutations in the Vacuolar Protein Sorting 13A (VPS13A gene, which is conserved from yeast to human. The consequences of VPS13A dysfunction in the nervous system are still largely unspecified. In order to study the consequences of VPS13A protein dysfunction in the ageing central nervous system we characterized a Drosophila melanogaster Vps13 mutant line. The Drosophila Vps13 gene encoded a protein of similar size as human VPS13A. Our data suggest that Vps13 is a peripheral membrane protein located to endosomal membranes and enriched in the fly head. Vps13 mutant flies showed a shortened life span and age associated neurodegeneration. Vps13 mutant flies were sensitive to proteotoxic stress and accumulated ubiquitylated proteins. Levels of Ref(2P, the Drosophila orthologue of p62, were increased and protein aggregates accumulated in the central nervous system. Overexpression of the human Vps13A protein in the mutant flies partly rescued apparent phenotypes. This suggests a functional conservation of human VPS13A and Drosophila Vps13. Our results demonstrate that Vps13 is essential to maintain protein homeostasis in the larval and adult Drosophila brain. Drosophila Vps13 mutants are suitable to investigate the function of Vps13 in the brain, to identify genetic enhancers and suppressors and to screen for potential therapeutic targets for Chorea-Acanthocytosis.

  15. Immunoreactivities to three circadian clock proteins in two ground crickets suggest interspecific diversity of the circadian clock structure

    Czech Academy of Sciences Publication Activity Database

    Shao, Q. M.; Sehadová, H.; Ichihara, N.; Sehnal, František; Takeda, M.

    2006-01-01

    Roč. 21, č. 2 (2006), s. 118-131 ISSN 0748-7304 Grant - others:Japan Society for the Promotion of Science(JP) JSPS 99L01205; Japan Society for the Promotion of Science(JP) ID No. P 04197 Institutional research plan: CEZ:AV0Z50070508 Keywords : circadian rhythm * photoperiodic clock * cryptochrome (CRY) Subject RIV: ED - Physiology Impact factor: 4.633, year: 2006

  16. [PIWI protein as a nucleolus visitor in Drosophila melanogaster].

    Science.gov (United States)

    Mikhaleva, E A; Iakushev, E Iu; Stoliarenko, A D; Klenov, M S; Pozovskiĭ, Ia M; Gvozdev, V A

    2015-01-01

    The evolutionarily conserved nuclear Piwi protein of Drosophila melanogaster is a representative of the Argonaute small RNA binding protein family. Guided by small piRNAs, Piwi functions in transposon silencing in somatic and germ cells of the gonad. We found that in ovarian somatic and germ cells, as well as in the established ovarian somatic cell line, Piwi is concentrated predominantly in the nucleolus--the main nuclear compartment, participating not only in rRNA synthesis, but also in various cell stress responses. We demonstrated the colocalization of Piwi with nucleolar marker proteins--fibrillarin and Nopp140. A mutation preventing Piwi transport to the nucleus and disturbing transposon silencing (piwi(Nt)) leads to 6-8-fold upregulation of rRNA genes expression, as evaluated by the level of transcripts of transposon insertions in 28S rRNA genes. RNase treatment of live cultured ovarian somatic cells depletes Piwi from the nucleolus. The same effect is observed upon inhibiting RNA polymerase I which transcribes rRNA, but not RNA polymerase II. In contrast, upon heat shock Piwi is concentrated in the nucleolus and is depleted from the nucleoplasm. These results implicate Piwi in RNA polymerase activity modulation and stress response in the nucleolus. We discuss possible noncanonical Piwi functions along with its canonical role in transposon silencing by piRNAs.

  17. Circadian clock protein KaiC forms ATP-dependent hexameric rings and binds DNA.

    Science.gov (United States)

    Mori, Tetsuya; Saveliev, Sergei V; Xu, Yao; Stafford, Walter F; Cox, Michael M; Inman, Ross B; Johnson, Carl H

    2002-12-24

    KaiC from Synechococcus elongatus PCC 7942 (KaiC) is an essential circadian clock protein in cyanobacteria. Previous sequence analyses suggested its inclusion in the RecADnaB superfamily. A characteristic of the proteins of this superfamily is that they form homohexameric complexes that bind DNA. We show here that KaiC also forms ring complexes with a central pore that can be visualized by electron microscopy. A combination of analytical ultracentrifugation and chromatographic analyses demonstrates that these complexes are hexameric. The association of KaiC molecules into hexamers depends on the presence of ATP. The KaiC sequence does not include the obvious DNA-binding motifs found in RecA or DnaB. Nevertheless, KaiC binds forked DNA substrates. These data support the inclusion of KaiC into the RecADnaB superfamily and have important implications for enzymatic activity of KaiC in the circadian clock mechanism that regulates global changes in gene expression patterns.

  18. Social Isolation Modulates CLOCK Protein and Beta-Catenin Expression Pattern in Gonadotropin-Inhibitory Hormone Neurons in Male Rats

    Directory of Open Access Journals (Sweden)

    Chuin Hau Teo

    2017-09-01

    Full Text Available Postweaning social isolation reduces the amplitude of the daily variation of CLOCK protein in the brain and induces lower reproductive activity. Gonadotropin-inhibitory hormone (GnIH acts as an inhibitor in the reproductive system and has been linked to stress. Social isolation has been shown to lower neuronal activity of GnIH-expressing neurons in the dorsomedial hypothalamus (DMH. The exact mechanism by which social isolation may affect GnIH is still unclear. We investigated the impact of social isolation on regulatory cellular mechanisms in GnIH neurons. We examined via immunohistochemistry the expression of CLOCK protein at four different times throughout the day in GnIH cells tagged with enhanced fluorescent green protein (EGFP-GnIH in 9-week-old adult male rats that have been raised for 6 weeks under postweaning social isolation and compared them with group-raised control rats of the same age. We also studied the expression of β-catenin—which has been shown to be affected by circadian proteins such as Bmal1—in EGFP-GnIH neurons to determine whether it could play a role in linking CLOCK in GnIH neurons. We found that social isolation modifies the pattern of CLOCK expression in GnIH neurons in the DMH. Socially isolated rats displayed greater CLOCK expression in the dark phase, while control rats displayed increased CLOCK expression in the light phase. Furthermore, β-catenin expression pattern in GnIH cells was disrupted by social isolation. This suggests that social isolation triggers changes in CLOCK and GnIH expression, which may be associated with an increase in nuclear β-catenin during the dark phase.

  19. Social Isolation Modulates CLOCK Protein and Beta-Catenin Expression Pattern in Gonadotropin-Inhibitory Hormone Neurons in Male Rats.

    Science.gov (United States)

    Teo, Chuin Hau; Soga, Tomoko; Parhar, Ishwar S

    2017-01-01

    Postweaning social isolation reduces the amplitude of the daily variation of CLOCK protein in the brain and induces lower reproductive activity. Gonadotropin-inhibitory hormone (GnIH) acts as an inhibitor in the reproductive system and has been linked to stress. Social isolation has been shown to lower neuronal activity of GnIH-expressing neurons in the dorsomedial hypothalamus (DMH). The exact mechanism by which social isolation may affect GnIH is still unclear. We investigated the impact of social isolation on regulatory cellular mechanisms in GnIH neurons. We examined via immunohistochemistry the expression of CLOCK protein at four different times throughout the day in GnIH cells tagged with enhanced fluorescent green protein (EGFP-GnIH) in 9-week-old adult male rats that have been raised for 6 weeks under postweaning social isolation and compared them with group-raised control rats of the same age. We also studied the expression of β-catenin-which has been shown to be affected by circadian proteins such as Bmal1-in EGFP-GnIH neurons to determine whether it could play a role in linking CLOCK in GnIH neurons. We found that social isolation modifies the pattern of CLOCK expression in GnIH neurons in the DMH. Socially isolated rats displayed greater CLOCK expression in the dark phase, while control rats displayed increased CLOCK expression in the light phase. Furthermore, β-catenin expression pattern in GnIH cells was disrupted by social isolation. This suggests that social isolation triggers changes in CLOCK and GnIH expression, which may be associated with an increase in nuclear β-catenin during the dark phase.

  20. Genetic Variation and Its Reflection on Posttranslational Modifications in Frequency Clock and Mating Type a-1 Proteins in Sordaria fimicola

    OpenAIRE

    Arif, Rabia; Akram, Faiza; Jamil, Tazeen; Mukhtar, Hamid; Lee, Siu Fai; Saleem, Muhammad

    2017-01-01

    Posttranslational modifications (PTMs) occur in all essential proteins taking command of their functions. There are many domains inside proteins where modifications take place on side-chains of amino acids through various enzymes to generate different species of proteins. In this manuscript we have, for the first time, predicted posttranslational modifications of frequency clock and mating type a-1 proteins in Sordaria fimicola collected from different sites to see the effect of environment o...

  1. Expression of circadian clock genes and proteins in urothelial cancer is related to cancer-associated genes

    International Nuclear Information System (INIS)

    Litlekalsoy, Jorunn; Rostad, Kari; Kalland, Karl-Henning; Hostmark, Jens G.; Laerum, Ole Didrik

    2016-01-01

    The purpose of this study was to evaluate invasive and metastatic potential of urothelial cancer by investigating differential expression of various clock genes/proteins participating in the 24 h circadian rhythms and to compare these gene expressions with transcription of other cancer-associated genes. Twenty seven paired samples of tumour and benign tissue collected from patients who underwent cystectomy were analysed and compared to 15 samples of normal bladder tissue taken from patients who underwent cystoscopy for benign prostate hyperplasia (unrelated donors). Immunohistochemical analyses were made for clock and clock-related proteins. In addition, the gene-expression levels of 22 genes (clock genes, casein kinases, oncogenes, tumour suppressor genes and cytokeratins) were analysed by real-time quantitative PCR (qPCR). Considerable up- or down-regulation and altered cellular distribution of different clock proteins, a reduction of casein kinase1A1 (CSNK1A1) and increase of casein kinase alpha 1 E (CSNK1E) were found. The pattern was significantly correlated with simultaneous up-regulation of stimulatory tumour markers, and a down-regulation of several suppressor genes. The pattern was mainly seen in aneuploid high-grade cancers. Considerable alterations were also found in the neighbouring bladder mucosa. The close correlation between altered expression of various clock genes and common tumour markers in urothelial cancer indicates that disturbed function in the cellular clock work may be an important additional mechanism contributing to cancer progression and malignant behaviour. The online version of this article (doi:10.1186/s12885-016-2580-y) contains supplementary material, which is available to authorized users

  2. Circadian Clock Proteins and Melatonin Receptors in Neurons and Glia of the Sapajus apella Cerebellum

    Directory of Open Access Journals (Sweden)

    Leila M. Guissoni Campos

    2018-02-01

    Full Text Available Oscillations of brain proteins in circadian rhythms are important for determining several cellular and physiological processes in anticipation of daily and seasonal environmental rhythms. In addition to the suprachiasmatic nucleus, the primary central oscillator, the cerebellum shows oscillations in gene and protein expression. The variety of local circuit rhythms that the cerebellar cortex contains influences functions such as motivational processes, regulation of feeding, food anticipation, language, and working memory. The molecular basis of the cerebellar oscillator has been demonstrated by “clock gene” expression within cells of the cerebellar layers. Genetic and epidemiological evidence suggests that disruption of circadian rhythms in humans can lead to many pathological conditions. Despite this importance, data about clock gene and protein expression in the cerebellum of diurnal (day-active species, specifically primates, is currently poorly explored, mainly in regard to cellular identity, as well as the relationship with other molecules also involved in cerebellar functions. These studies could contribute to clarification of the possible mechanisms behind cerebellar rhythmicity. Considering that calcium binding proteins (CaBPs play crucial roles in preserving and modulating cerebellar functions and that clock gene expression can be controlled by afferent projections or paracrine circadian signals such as the hormone melatonin, the present study aimed to describe cellular identities, distribution patterns and day/night expression changes in PER1, PER2, CaBPs, and MT1 and MT2 melatonin receptors in the cerebellar cortex of a diurnal primate using conventional fluorescence and peroxidase-antiperoxidase immunocytochemical techniques. PER1 and PER2 immunoreactive (IR cells were observed in the Purkinje cells of the cerebellum, and MT1 and MT2 receptors were localized around Purkinje cells in the Pj layer in Bergmann cells. This identity

  3. Diurnal rhythms in neurexins transcripts and inhibitory/excitatory synapse scaffold proteins in the biological clock.

    Directory of Open Access Journals (Sweden)

    Mika Shapiro-Reznik

    Full Text Available The neurexin genes (NRXN1/2/3 encode two families (α and β of highly polymorphic presynaptic proteins that are involved in excitatory/inhibitory synaptic balance. Recent studies indicate that neuronal activation and memory formation affect NRXN1/2/3α expression and alternative splicing at splice sites 3 and 4 (SS#3/SS#4. Neurons in the biological clock residing in the suprachiasmatic nuclei of the hypothalamus (SCN act as self-sustained oscillators, generating rhythms in gene expression and electrical activity, to entrain circadian bodily rhythms to the 24 hours day/night cycles. Cell autonomous oscillations in NRXN1/2/3α expression and SS#3/SS#4 exons splicing and their links to rhythms in excitatory/inhibitory synaptic balance in the circadian clock were explored. NRXN1/2/3α expression and SS#3/SS#4 splicing, levels of neurexin-2α and the synaptic scaffolding proteins PSD-95 and gephyrin (representing excitatory and inhibitory synapses, respectively were studied in mRNA and protein extracts obtained from SCN of C3H/J mice at different times of the 24 hours day/night cycle. Further studies explored the circadian oscillations in these components and causality relationships in immortalized rat SCN2.2 cells. Diurnal rhythms in mNRXN1α and mNRXN2α transcription, SS#3/SS#4 exon-inclusion and PSD-95 gephyrin and neurexin-2α levels were found in the SCN in vivo. No such rhythms were found with mNRXN3α. SCN2.2 cells also exhibited autonomous circadian rhythms in rNRXN1/2 expression SS#3/SS#4 exon inclusion and PSD-95, gephyrin and neurexin-2α levels. rNRXN3α and rNRXN1/2β were not expressed. Causal relationships were demonstrated, by use of specific siRNAs, between rNRXN2α SS#3 exon included transcripts and gephyrin levels in the SCN2.2 cells. These results show for the first time dynamic, cell autonomous, diurnal rhythms in expression and splicing of NRXN1/2 and subsequent effects on the expression of neurexin-2α and postsynaptic

  4. [Drosophila melanogaster as a model for studying the function of animal viral proteins].

    Science.gov (United States)

    Omelianchuk, L V; Iudina, O S

    2011-07-01

    Studies in which Drosophila melanogaster individuals carrying transgenes of animal viruses were used to analyze the action of animal viral proteins on the cell are reviewed. The data presented suggest that host specificity of viruses is determined by their proteins responsible for the penetration of the virus into the cell, while viral proteins responsible for interactions with the host cell are much less host-specific. Due to this, the model of Drosophila with its developed system of searching for genetic interactions can be used to find intracellular targets for the action of viral proteins of the second group.

  5. Drosophila protein interaction map (DPiM): a paradigm for metazoan protein complex interactions.

    Science.gov (United States)

    Guruharsha, K G; Obar, Robert A; Mintseris, Julian; Aishwarya, K; Krishnan, R T; Vijayraghavan, K; Artavanis-Tsakonas, Spyros

    2012-01-01

    Proteins perform essential cellular functions as part of protein complexes, often in conjunction with RNA, DNA, metabolites and other small molecules. The genome encodes thousands of proteins but not all of them are expressed in every cell type; and expressed proteins are not active at all times. Such diversity of protein expression and function accounts for the level of biological intricacy seen in nature. Defining protein-protein interactions in protein complexes, and establishing the when, what and where of potential interactions, is therefore crucial to understanding the cellular function of any protein-especially those that have not been well studied by traditional molecular genetic approaches. We generated a large-scale resource of affinity-tagged expression-ready clones and used co-affinity purification combined with tandem mass-spectrometry to identify protein partners of nearly 5,000 Drosophila melanogaster proteins. The resulting protein complex "map" provided a blueprint of metazoan protein complex organization. Here we describe how the map has provided valuable insights into protein function in addition to generating hundreds of testable hypotheses. We also discuss recent technological advancements that will be critical in addressing the next generation of questions arising from the map.

  6. Lego clocks: building a clock from parts.

    Science.gov (United States)

    Brunner, Michael; Simons, Mirre J P; Merrow, Martha

    2008-06-01

    A new finding opens up speculation that the molecular mechanism of circadian clocks in Synechococcus elongatus is composed of multiple oscillator systems (Kitayama and colleagues, this issue, pp. 1513-1521), as has been described in many eukaryotic clock model systems. However, an alternative intepretation is that the pacemaker mechanism-as previously suggested-lies primarily in the rate of ATP hydrolysis by the clock protein KaiC.

  7. A low protein diet increases the hypoxic tolerance in Drosophila.

    Directory of Open Access Journals (Sweden)

    Paul Vigne

    2006-12-01

    Full Text Available Dietary restriction is well known to increase the life span of a variety of organisms from yeast to mammals, but the relationships between nutrition and the hypoxic tolerance have not yet been considered. Hypoxia is a major cause of cell death in myocardial infarction and stroke. Here we forced hypoxia-related death by exposing one-day-old male Drosophila to chronic hypoxia (5% O(2 and analysed their survival. Chronic hypoxia reduced the average life span from 33.6 days to 6.3 days when flies were fed on a rich diet. A demographic analysis indicated that chronic hypoxia increased the slope of the mortality trajectory and not the short-term risk of death. Dietary restriction produced by food dilution, by yeast restriction, or by amino acid restriction partially reversed the deleterious action of hypoxia. It increased the life span of hypoxic flies up to seven days, which represented about 25% of the life time of an hypoxic fly. Maximum survival of hypoxic flies required only dietary sucrose, and it was insensitive to drugs such as rapamycin and resveratrol, which increase longevity of normoxic animals. The results thus uncover a new link between protein nutrition, nutrient signalling, and resistance to hypoxic stresses.

  8. Structure of Drosophila Oskar reveals a novel RNA binding protein

    Science.gov (United States)

    Yang, Na; Yu, Zhenyu; Hu, Menglong; Wang, Mingzhu; Lehmann, Ruth; Xu, Rui-Ming

    2015-01-01

    Oskar (Osk) protein plays critical roles during Drosophila germ cell development, yet its functions in germ-line formation and body patterning remain poorly understood. This situation contrasts sharply with the vast knowledge about the function and mechanism of osk mRNA localization. Osk is predicted to have an N-terminal LOTUS domain (Osk-N), which has been suggested to bind RNA, and a C-terminal hydrolase-like domain (Osk-C) of unknown function. Here, we report the crystal structures of Osk-N and Osk-C. Osk-N shows a homodimer of winged-helix–fold modules, but without detectable RNA-binding activity. Osk-C has a lipase-fold structure but lacks critical catalytic residues at the putative active site. Surprisingly, we found that Osk-C binds the 3′UTRs of osk and nanos mRNA in vitro. Mutational studies identified a region of Osk-C important for mRNA binding. These results suggest possible functions of Osk in the regulation of stability, regulation of translation, and localization of relevant mRNAs through direct interaction with their 3′UTRs, and provide structural insights into a novel protein–RNA interaction motif involving a hydrolase-related domain. PMID:26324911

  9. Developmental environment mediates male seminal protein investment in Drosophila melanogaster.

    Science.gov (United States)

    Wigby, Stuart; Perry, Jennifer C; Kim, Yon-Hee; Sirot, Laura K

    2016-03-01

    Males of many species fine-tune their ejaculates in response to sperm competition risk. Resource availability and the number of competitors during development can also strongly influence sperm production. However, despite the key role of seminal proteins in mediating reproductive processes, it is unclear whether seminal protein investment is dependent on the developmental environment.We manipulated the developmental environment of Drosophila melanogaster by rearing flies at low and high density. As expected, this resulted in large and small (i.e. high and low condition) adult phenotypes, respectively.As predicted, large males produced more of two key seminal proteins, sex peptide (SP) and ovulin, and were more successful at obtaining matings with both virgin and previously mated females. However, there was only a weak and non-significant trend for large males to transfer more absolute quantities of SP at mating, and thus, small males ejaculated proportionally more of their stored accessory gland SP resources.Males transferred more receptivity-inhibiting SP to large females. Despite this, large females remated more quickly than small females and thus responded to their developmental environment over and above the quantity of SP they received.The results are consistent with two non-mutually exclusive hypotheses. First, flies might respond to condition-dependent reproductive opportunities, with (i) small males investing heavily in ejaculates when mating opportunities arise and large males strategically partitioning SP resources and (ii) small females remating at reduced rates because they have higher mating costs or need to replenish sperm less often.Second, flies may be primed by their larval environment to deal with similar adult population densities, with (i) males perceiving high density as signalling increased competition, leading small males to invest proportionally more SP resources at mating and (ii) females perceiving high density as signalling abundant

  10. Molecular identification of a Drosophila G protein-coupled receptor specific for crustacean cardioactive peptide

    DEFF Research Database (Denmark)

    Cazzamali, Giuseppe; Hauser, Frank; Kobberup, Sune

    2003-01-01

    The Drosophila Genome Project website (www.flybase.org) contains the sequence of an annotated gene (CG6111) expected to code for a G protein-coupled receptor. We have cloned this receptor and found that its gene was not correctly predicted, because an annotated neighbouring gene (CG14547) was also...... part of the receptor gene. DNA corresponding to the corrected gene CG6111 was expressed in Chinese hamster ovary cells, where it was found to code for a receptor that could be activated by low concentrations of crustacean cardioactive peptide, which is a neuropeptide also known to occur in Drosophila...... and other insects (EC(50), 5.4 x 10(-10)M). Other known Drosophila neuropeptides, such as adipokinetic hormone, did not activate the receptor. The receptor is expressed in all developmental stages from Drosophila, but only very weakly in larvae. In adult flies, the receptor is mainly expressed in the head...

  11. Identifying neuropeptide and protein hormone receptors in Drosophila melanogaster by exploiting genomic data

    DEFF Research Database (Denmark)

    Hauser, Frank; Williamson, Michael; Cazzamali, Giuseppe

    2006-01-01

    insect genome, that of the fruitfly Drosophila melanogaster, was sequenced in 2000, and about 200 GPCRs have been annnotated in this model insect. About 50 of these receptors were predicted to have neuropeptides or protein hormones as their ligands. Since 2000, the cDNAs of most of these candidate...... receptors have been cloned and for many receptors the endogenous ligand has been identified. In this review, we will give an update about the current knowledge of all Drosophila neuropeptide and protein hormone receptors, and discuss their phylogenetic relationships. Udgivelsesdato: 2006-Feb...

  12. Diel pattern of circadian clock and storage protein gene expression in leaves and during seed filling in cowpea (Vigna unguiculata).

    Science.gov (United States)

    Weiss, Julia; Terry, Marta I; Martos-Fuentes, Marina; Letourneux, Lisa; Ruiz-Hernández, Victoria; Fernández, Juan A; Egea-Cortines, Marcos

    2018-02-14

    Cowpea (Vigna unguiculata) is an important source of protein supply for animal and human nutrition. The major storage globulins VICILIN and LEGUMIN (LEG) are synthesized from several genes including LEGA, LEGB, LEGJ and CVC (CONVICILIN). The current hypothesis is that the plant circadian core clock genes are conserved in a wide array of species and that primary metabolism is to a large extent controlled by the plant circadian clock. Our aim was to investigate a possible link between gene expression of storage proteins and the circadian clock. We identified cowpea orthologues of the core clock genes VunLHY, VunTOC1, VunGI and VunELF3, the protein storage genes VunLEG, VunLEGJ, and VunCVC as well as nine candidate reference genes used in RT-PCR. ELONGATION FACTOR 1-A (ELF1A) resulted the most suitable reference gene. The clock genes VunELF3, VunGI, VunTOC1 and VunLHY showed a rhythmic expression profile in leaves with a typical evening/night and morning/midday phased expression. The diel patterns were not completely robust and only VungGI and VungELF3 retained a rhythmic pattern under free running conditions of darkness. Under field conditions, rhythmicity and phasing apparently faded during early pod and seed development and was regained in ripening pods for VunTOC1 and VunLHY. Mature seeds showed a rhythmic expression of VunGI resembling leaf tissue under controlled growth chamber conditions. Comparing time windows during developmental stages we found that VunCVC and VunLEG were significantly down regulated during the night in mature pods as compared to intermediate ripe pods, while changes in seeds were non-significant due to high variance. The rhythmic expression under field conditions was lost under growth chamber conditions. The core clock gene network is conserved in cowpea leaves showing a robust diel expression pattern except VunELF3 under growth chamber conditions. There appears to be a clock transcriptional reprogramming in pods and seeds compared to

  13. Drosophila SMN complex proteins Gemin2, Gemin3, and Gemin5 are components of U bodies

    International Nuclear Information System (INIS)

    Cauchi, Ruben J.; Sanchez-Pulido, Luis; Liu, Ji-Long

    2010-01-01

    Uridine-rich small nuclear ribonucleoproteins (U snRNPs) play key roles in pre-mRNA processing in the nucleus. The assembly of most U snRNPs takes place in the cytoplasm and is facilitated by the survival motor neuron (SMN) complex. Discrete cytoplasmic RNA granules called U bodies have been proposed to be specific sites for snRNP assembly because they contain U snRNPs and SMN. U bodies invariably associate with P bodies, which are involved in mRNA decay and translational control. However, it remains unknown whether other SMN complex proteins also localise to U bodies. In Drosophila there are four SMN complex proteins, namely SMN, Gemin2/CG10419, Gemin3 and Gemin5/Rigor mortis. Drosophila Gemin3 was originally identified as the Drosophila orthologue of human and yeast Dhh1, a component of P bodies. Through an in silico analysis of the DEAD-box RNA helicases we confirmed that Gemin3 is the bona fide Drosophila orthologue of vertebrate Gemin3 whereas the Drosophila orthologue of Dhh1 is Me31B. We then made use of the Drosophila egg chamber as a model system to study the subcellular distribution of the Gemin proteins as well as Me31B. Our cytological investigations show that Gemin2, Gemin3 and Gemin5 colocalise with SMN in U bodies. Although they are excluded from P bodies, as components of U bodies, Gemin2, Gemin3 and Gemin5 are consistently found associated with P bodies, wherein Me31B resides. In addition to a role in snRNP biogenesis, SMN complexes residing in U bodies may also be involved in mRNP assembly and/or transport.

  14. Drosophila SMN complex proteins Gemin2, Gemin3, and Gemin5 are components of U bodies

    Energy Technology Data Exchange (ETDEWEB)

    Cauchi, Ruben J.; Sanchez-Pulido, Luis [MRC Functional Genomics Unit, Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford OX1 3QX (United Kingdom); Liu, Ji-Long, E-mail: jilong.liu@dpag.ox.ac.uk [MRC Functional Genomics Unit, Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford OX1 3QX (United Kingdom)

    2010-08-15

    Uridine-rich small nuclear ribonucleoproteins (U snRNPs) play key roles in pre-mRNA processing in the nucleus. The assembly of most U snRNPs takes place in the cytoplasm and is facilitated by the survival motor neuron (SMN) complex. Discrete cytoplasmic RNA granules called U bodies have been proposed to be specific sites for snRNP assembly because they contain U snRNPs and SMN. U bodies invariably associate with P bodies, which are involved in mRNA decay and translational control. However, it remains unknown whether other SMN complex proteins also localise to U bodies. In Drosophila there are four SMN complex proteins, namely SMN, Gemin2/CG10419, Gemin3 and Gemin5/Rigor mortis. Drosophila Gemin3 was originally identified as the Drosophila orthologue of human and yeast Dhh1, a component of P bodies. Through an in silico analysis of the DEAD-box RNA helicases we confirmed that Gemin3 is the bona fide Drosophila orthologue of vertebrate Gemin3 whereas the Drosophila orthologue of Dhh1 is Me31B. We then made use of the Drosophila egg chamber as a model system to study the subcellular distribution of the Gemin proteins as well as Me31B. Our cytological investigations show that Gemin2, Gemin3 and Gemin5 colocalise with SMN in U bodies. Although they are excluded from P bodies, as components of U bodies, Gemin2, Gemin3 and Gemin5 are consistently found associated with P bodies, wherein Me31B resides. In addition to a role in snRNP biogenesis, SMN complexes residing in U bodies may also be involved in mRNP assembly and/or transport.

  15. A nanobody-based toolset to investigate the role of protein localization and dispersal in Drosophila.

    Science.gov (United States)

    Harmansa, Stefan; Alborelli, Ilaria; Bieli, Dimitri; Caussinus, Emmanuel; Affolter, Markus

    2017-04-11

    The role of protein localization along the apical-basal axis of polarized cells is difficult to investigate in vivo, partially due to lack of suitable tools. Here, we present the GrabFP system, a collection of four nanobody-based GFP-traps that localize to defined positions along the apical-basal axis. We show that the localization preference of the GrabFP traps can impose a novel localization on GFP-tagged target proteins and results in their controlled mislocalization. These new tools were used to mislocalize transmembrane and cytoplasmic GFP fusion proteins in the Drosophila wing disc epithelium and to investigate the effect of protein mislocalization. Furthermore, we used the GrabFP system as a tool to study the extracellular dispersal of the Decapentaplegic (Dpp) protein and show that the Dpp gradient forming in the lateral plane of the Drosophila wing disc epithelium is essential for patterning of the wing imaginal disc.

  16. SOLO: a meiotic protein required for centromere cohesion, coorientation, and SMC1 localization in Drosophila melanogaster.

    Science.gov (United States)

    Yan, Rihui; Thomas, Sharon E; Tsai, Jui-He; Yamada, Yukihiro; McKee, Bruce D

    2010-02-08

    Sister chromatid cohesion is essential to maintain stable connections between homologues and sister chromatids during meiosis and to establish correct centromere orientation patterns on the meiosis I and II spindles. However, the meiotic cohesion apparatus in Drosophila melanogaster remains largely uncharacterized. We describe a novel protein, sisters on the loose (SOLO), which is essential for meiotic cohesion in Drosophila. In solo mutants, sister centromeres separate before prometaphase I, disrupting meiosis I centromere orientation and causing nondisjunction of both homologous and sister chromatids. Centromeric foci of the cohesin protein SMC1 are absent in solo mutants at all meiotic stages. SOLO and SMC1 colocalize to meiotic centromeres from early prophase I until anaphase II in wild-type males, but both proteins disappear prematurely at anaphase I in mutants for mei-S332, which encodes the Drosophila homologue of the cohesin protector protein shugoshin. The solo mutant phenotypes and the localization patterns of SOLO and SMC1 indicate that they function together to maintain sister chromatid cohesion in Drosophila meiosis.

  17. cGMP-dependent protein kinase I, the circadian clock, sleep and learning.

    Science.gov (United States)

    Feil, Robert; Hölter, Sabine M; Weindl, Karin; Wurst, Wolfgang; Langmesser, Sonja; Gerling, Andrea; Feil, Susanne; Albrecht, Urs

    2009-07-01

    The second messenger cGMP controls cardiovascular and gastrointestinal homeostasis in mammals. However, its physiological relevance in the nervous system is poorly understood.1 Now, we have reported that the cGMP-dependent protein kinase type I (PRKG1) is implicated in the regulation of the timing and quality of sleep and wakefulness.2Prkg1 mutant mice showed altered distribution of sleep and wakefulness as well as reduction in rapid-eye-movement sleep (REMS) duration and in non-REMS consolidation. Furthermore, the ability to sustain waking episodes was compromised. These observations were also reflected in wheel-running and drinking activity. A decrease in electroencephalogram power in the delta frequency range (1-4 Hz) under baseline conditions was observed, which was normalized after sleep deprivation. Together with the finding that circadian clock amplitude is reduced in Prkg1 mutants these results indicate a decrease of the wake-promoting output of the circadian system affecting sleep. Because quality of sleep might affect learning we tested Prkg1 mutants in several learning tasks and find normal spatial learning but impaired object recognition memory in these animals. Our findings indicate that Prkg1 impinges on circadian rhythms, sleep and distinct aspects of learning.

  18. The carnegie protein trap library: a versatile tool for Drosophila developmental studies.

    Science.gov (United States)

    Buszczak, Michael; Paterno, Shelley; Lighthouse, Daniel; Bachman, Julia; Planck, Jamie; Owen, Stephenie; Skora, Andrew D; Nystul, Todd G; Ohlstein, Benjamin; Allen, Anna; Wilhelm, James E; Murphy, Terence D; Levis, Robert W; Matunis, Erika; Srivali, Nahathai; Hoskins, Roger A; Spradling, Allan C

    2007-03-01

    Metazoan physiology depends on intricate patterns of gene expression that remain poorly known. Using transposon mutagenesis in Drosophila, we constructed a library of 7404 protein trap and enhancer trap lines, the Carnegie collection, to facilitate gene expression mapping at single-cell resolution. By sequencing the genomic insertion sites, determining splicing patterns downstream of the enhanced green fluorescent protein (EGFP) exon, and analyzing expression patterns in the ovary and salivary gland, we found that 600-900 different genes are trapped in our collection. A core set of 244 lines trapped different identifiable protein isoforms, while insertions likely to act as GFP-enhancer traps were found in 256 additional genes. At least 8 novel genes were also identified. Our results demonstrate that the Carnegie collection will be useful as a discovery tool in diverse areas of cell and developmental biology and suggest new strategies for greatly increasing the coverage of the Drosophila proteome with protein trap insertions.

  19. Regulation of lifespan, metabolism, and stress responses by the Drosophila SH2B protein, Lnk.

    Directory of Open Access Journals (Sweden)

    Cathy Slack

    2010-03-01

    Full Text Available Drosophila Lnk is the single ancestral orthologue of a highly conserved family of structurally-related intracellular adaptor proteins, the SH2B proteins. As adaptors, they lack catalytic activity but contain several protein-protein interaction domains, thus playing a critical role in signal transduction from receptor tyrosine kinases to form protein networks. Physiological studies of SH2B function in mammals have produced conflicting data. However, a recent study in Drosophila has shown that Lnk is an important regulator of the insulin/insulin-like growth factor (IGF-1 signaling (IIS pathway during growth, functioning in parallel to the insulin receptor substrate, Chico. As this pathway also has an evolutionary conserved role in the determination of organism lifespan, we investigated whether Lnk is required for normal lifespan in Drosophila. Phenotypic analysis of mutants for Lnk revealed that loss of Lnk function results in increased lifespan and improved survival under conditions of oxidative stress and starvation. Starvation resistance was found to be associated with increased metabolic stores of carbohydrates and lipids indicative of impaired metabolism. Biochemical and genetic data suggest that Lnk functions in both the IIS and Ras/Mitogen activated protein Kinase (MapK signaling pathways. Microarray studies support this model, showing transcriptional feedback onto genes in both pathways as well as indicating global changes in both lipid and carbohydrate metabolism. Finally, our data also suggest that Lnk itself may be a direct target of the IIS responsive transcription factor, dFoxo, and that dFoxo may repress Lnk expression. We therefore describe novel functions for a member of the SH2B protein family and provide the first evidence for potential mechanisms of SH2B regulation. Our findings suggest that IIS signaling in Drosophila may require the activity of a second intracellular adaptor, thereby yielding fundamental new insights into the

  20. Analysis of Thioester-Containing Proteins during the Innate Immune Response of Drosophila melanogaster

    Science.gov (United States)

    Bou Aoun, Richard; Hetru, Charles; Troxler, Laurent; Doucet, Daniel; Ferrandon, Dominique; Matt, Nicolas

    2010-01-01

    Thioester-containing proteins (TEPs) are conserved proteins among insects that are thought to be involved in innate immunity. In Drosophila, the Tep family is composed of 6 genes named Tep1–Tep6. In this study, we investigated the phylogeny, expression pattern and roles of these genes in the host defense of Drosophila. Protostomian Tep genes are clustered in 3 distinct branches, 1 of which is specific to mosquitoes. Most D. melanogaster Tep genes are expressed in hemocytes, can be induced in the fat body, and are expressed in specific regions of the hypodermis. This expression pattern is consistent with a role in innate immunity. However, we find that TEP1, TEP2, and TEP4 are not strictly required in the body cavity to fight several bacterial and fungal infections. One possibility is that Drosophila TEPs act redundantly or that their absence can be compensated by other components of the immune response. TEPs may thus provide a subtle selective advantage during evolution. Alternatively, they may be required in host defense against specific as yet unidentified natural pathogens of Drosophila. PMID:21063077

  1. Rampant adaptive evolution in regions of proteins with unknown function in Drosophila simulans.

    Directory of Open Access Journals (Sweden)

    Alisha K Holloway

    2007-10-01

    Full Text Available Adaptive protein evolution is pervasive in Drosophila. Genomic studies, thus far, have analyzed each protein as a single entity. However, the targets of adaptive events may be localized to particular parts of proteins, such as protein domains or regions involved in protein folding. We compared the population genetic mechanisms driving sequence polymorphism and divergence in defined protein domains and non-domain regions. Interestingly, we find that non-domain regions of proteins are more frequent targets of directional selection. Protein domains are also evolving under directional selection, but appear to be under stronger purifying selection than non-domain regions. Non-domain regions of proteins clearly play a major role in adaptive protein evolution on a genomic scale and merit future investigations of their functional properties.

  2. Klp10A modulates the localization of centriole-associated proteins during Drosophila male gametogenesis.

    Science.gov (United States)

    Gottardo, Marco; Callaini, Giuliano; Riparbelli, Maria Giovanna

    2016-12-16

    Mutations in Klp10A, a microtubule-depolymerising Kinesin-13, lead to overly long centrioles in Drosophila male germ cells. We demonstrated that the loss of Klp10A modifies the distribution of typical proteins involved in centriole assembly and function. In the absence of Klp10A the distribution of Drosophila pericentrin-like protein (Dplp), Sas-4 and Sak/Plk4 that are restricted in control testes to the proximal end of the centriole increase along the centriole length. Remarkably, the cartwheel is lacking or it appears abnormal in mutant centrioles, suggesting that this structure may spatially delimit protein localization. Moreover, the parent centrioles that in control cells have the same dimensions grow at different rates in mutant testes with the mother centrioles longer than the daughters. Daughter centrioles have often an ectopic position with respect to the proximal end of the mothers and failed to recruit Dplp.

  3. The HIV-1 Vpu protein induces apoptosis in Drosophila via activation of JNK signaling.

    Directory of Open Access Journals (Sweden)

    Christelle Marchal

    Full Text Available The genome of the human immunodeficiency virus type 1 (HIV-1 encodes the canonical retroviral proteins, as well as additional accessory proteins that enhance the expression of viral genes, the infectivity of the virus and the production of virions. The accessory Viral Protein U (Vpu, in particular, enhances viral particle production, while also promoting apoptosis of HIV-infected human T lymphocytes. Some Vpu effects rely on its interaction with the ubiquitin-proteasome protein degradation system, but the mechanisms responsible for its pro-apoptotic effects in vivo are complex and remain largely to be elucidated.We took advantage of the Drosophila model to study the effects of Vpu activity in vivo. Expression of Vpu in the developing Drosophila wing provoked tissue loss due to caspase-dependent apoptosis. Moreover, Vpu induced expression of the pro-apoptotic gene reaper, known to down-regulate Inhibitor of Apoptosis Proteins (IAPs which are caspase-antagonizing E3 ubiquitin ligases. Indeed, Vpu also reduced accumulation of Drosophila IAP1 (DIAP1. Though our results demonstrate a physical interaction between Vpu and the proteasome-addressing SLIMB/β-TrCP protein, as in mammals, both SLIMB/βTrCP-dependent and -independent Vpu effects were observed in the Drosophila wing. Lastly, the pro-apoptotic effect of Vpu in this tissue was abrogated upon inactivation of the c-Jun N-terminal Kinase (JNK pathway. Our results in the fly thus provide the first functional evidence linking Vpu pro-apoptotic effects to activation of the conserved JNK pathway.

  4. Expression and purification of sea raven type II antifreeze protein from Drosophila melanogaster S2 cells.

    Science.gov (United States)

    Scotter, Andrew J; Kuntz, Douglas A; Saul, Michelle; Graham, Laurie A; Davies, Peter L; Rose, David R

    2006-06-01

    We present a system for the expression and purification of recombinant sea raven type II antifreeze protein, a cysteine-rich, C-type lectin-like globular protein that has proved to be a difficult target for recombinant expression and purification. The cDNAs encoding the pro- and mature forms of the sea raven protein were cloned into a modified pMT Drosophila expression vector. These constructs produced N-terminally His(6)-tagged pro- and mature forms of the type II antifreeze protein under the control of a metallothionein promoter when transfected into Drosophila melanogaster S2 cells. Upon induction of stable cell lines the two proteins were expressed at high levels and secreted into the medium. The proteins were then purified from the cell medium in a simple and rapid protocol using immobilized metal affinity chromatography and specific protease cleavage by tobacco etch virus protease. The proteins demonstrated antifreeze activity indistinguishable from that of wild-type sea raven antifreeze protein purified from serum as illustrated by ice affinity purification, ice crystal morphology, and their ability to inhibit ice crystal growth. This expression and purification system gave yields of 95 mg/L of fully active mature sea raven type II AFP and 9.6 mg/L of the proprotein. This surpasses all previous attempts to express this protein in Escherichia coli, baculovirus-infected fall armyworm cells and Pichia pastoris and will provide sufficient protein for structural analysis.

  5. G-rich, a Drosophila selenoprotein, is a Golgi-resident type III membrane protein

    International Nuclear Information System (INIS)

    Chen, Chang Lan; Shim, Myoung Sup; Chung, Jiyeol; Yoo, Hyun-Seung; Ha, Ji Min; Kim, Jin Young; Choi, Jinmi; Zang, Shu Liang; Hou, Xiao; Carlson, Bradley A.; Hatfield, Dolph L.; Lee, Byeong Jae

    2006-01-01

    G-rich is a Drosophila melanogaster selenoprotein, which is a homologue of human and mouse SelK. Subcellular localization analysis using GFP-tagged G-rich showed that G-rich was localized in the Golgi apparatus. The fusion protein was co-localized with the Golgi marker proteins but not with an endoplasmic reticulum (ER) marker protein in Drosophila SL2 cells. Bioinformatic analysis of G-rich suggests that this protein is either type II or type III transmembrane protein. To determine the type of transmembrane protein experimentally, GFP-G-rich in which GFP was tagged at the N-terminus of G-rich, or G-rich-GFP in which GFP was tagged at the C-terminus of G-rich, were expressed in SL2 cells. The tagged proteins were then digested with trypsin, and analyzed by Western blot analysis. The results showed that the C-terminus of the G-rich protein was exposed to the cytoplasm indicating it is a type III microsomal membrane protein. G-rich is First selenoprotein identified in the Golgi apparatus

  6. The functional interplay between protein kinase CK2 and CCA1 transcriptional activity is essential for clock temperature compensation in Arabidopsis.

    Directory of Open Access Journals (Sweden)

    Sergi Portolés

    2010-11-01

    Full Text Available Circadian rhythms are daily biological oscillations driven by an endogenous mechanism known as circadian clock. The protein kinase CK2 is one of the few clock components that is evolutionary conserved among different taxonomic groups. CK2 regulates the stability and nuclear localization of essential clock proteins in mammals, fungi, and insects. Two CK2 regulatory subunits, CKB3 and CKB4, have been also linked with the Arabidopsis thaliana circadian system. However, the biological relevance and the precise mechanisms of CK2 function within the plant clockwork are not known. By using ChIP and Double-ChIP experiments together with in vivo luminescence assays at different temperatures, we were able to identify a temperature-dependent function for CK2 modulating circadian period length. Our study uncovers a previously unpredicted mechanism for CK2 antagonizing the key clock regulator CIRCADIAN CLOCK-ASSOCIATED 1 (CCA1. CK2 activity does not alter protein accumulation or subcellular localization but interferes with CCA1 binding affinity to the promoters of the oscillator genes. High temperatures enhance the CCA1 binding activity, which is precisely counterbalanced by the CK2 opposing function. Altering this balance by over-expression, mutation, or pharmacological inhibition affects the temperature compensation profile, providing a mechanism by which plants regulate circadian period at changing temperatures. Therefore, our study establishes a new model demonstrating that two opposing and temperature-dependent activities (CCA1-CK2 are essential for clock temperature compensation in Arabidopsis.

  7. Functional dissection of the Hox protein Abdominal-B in Drosophila cell culture

    Energy Technology Data Exchange (ETDEWEB)

    Zhai, Zongzhao [Key Laboratory of the Zoological Systematics and Evolution, Institute of Zoology, Chinese Academy of Sciences, Beichen West Road, Chaoyang, Beijing 100101 (China); CellNetworks - Cluster of Excellence, Centre for Organismal Studies (COS) Heidelberg, University of Heidelberg, D-69120 Heidelberg (Germany); Graduate School of Chinese Academy of Sciences, Beijing 100039 (China); Yang, Xingke, E-mail: yangxk@ioz.ac.cn [Key Laboratory of the Zoological Systematics and Evolution, Institute of Zoology, Chinese Academy of Sciences, Beichen West Road, Chaoyang, Beijing 100101 (China); Lohmann, Ingrid, E-mail: ilohmann@flydev.org [CellNetworks - Cluster of Excellence, Centre for Organismal Studies (COS) Heidelberg, University of Heidelberg, D-69120 Heidelberg (Germany)

    2011-11-04

    Highlights: Black-Right-Pointing-Pointer ct340 CRM was identified to be the posterior spiracle enhancer of gene cut. Black-Right-Pointing-Pointer ct340 is under the direct transcriptional control of Hox protein Abd-B. Black-Right-Pointing-Pointer An efficient cloning system was developed to assay protein-DNA interaction. Black-Right-Pointing-Pointer New features of Abd-B dependent target gene regulation were detected. -- Abstract: Hox transcription factors regulate the morphogenesis along the anterior-posterior (A/P) body axis through the interaction with small cis-regulatory modules (CRMs) of their target gene, however so far very few Hox CRMs are known and have been analyzed in detail. In this study we have identified a new Hox CRM, ct340, which guides the expression of the cell type specification gene cut (ct) in the posterior spiracle under the direct control of the Hox protein Abdominal-B (Abd-B). Using the ct340 enhancer activity as readout, an efficient cloning system to generate VP16 activation domain fusion protein was developed to unambiguously test protein-DNA interaction in Drosophila cell culture. By functionally dissecting the Abd-B protein, new features of Abd-B dependent target gene regulation were detected. Due to its easy adaptability, this system can be generally used to map functional domains within sequence-specific transcriptional factors in Drosophila cell culture, and thus provide preliminary knowledge of the protein functional domain structure for further in vivo analysis.

  8. Drosophila CTCF tandemly aligns with other insulator proteins at the borders of H3K27me3 domains.

    Science.gov (United States)

    Van Bortle, Kevin; Ramos, Edward; Takenaka, Naomi; Yang, Jingping; Wahi, Jessica E; Corces, Victor G

    2012-11-01

    Several multiprotein DNA complexes capable of insulator activity have been identified in Drosophila melanogaster, yet only CTCF, a highly conserved zinc finger protein, and the transcription factor TFIIIC have been shown to function in mammals. CTCF is involved in diverse nuclear activities, and recent studies suggest that the proteins with which it associates and the DNA sequences that it targets may underlie these various roles. Here we show that the Drosophila homolog of CTCF (dCTCF) aligns in the genome with other Drosophila insulator proteins such as Suppressor of Hairy wing [SU(HW)] and Boundary Element Associated Factor of 32 kDa (BEAF-32) at the borders of H3K27me3 domains, which are also enriched for associated insulator proteins and additional cofactors. RNAi depletion of dCTCF and combinatorial knockdown of gene expression for other Drosophila insulator proteins leads to a reduction in H3K27me3 levels within repressed domains, suggesting that insulators are important for the maintenance of appropriate repressive chromatin structure in Polycomb (Pc) domains. These results shed new insights into the roles of insulators in chromatin domain organization and support recent models suggesting that insulators underlie interactions important for Pc-mediated repression. We reveal an important relationship between dCTCF and other Drosophila insulator proteins and speculate that vertebrate CTCF may also align with other nuclear proteins to accomplish similar functions.

  9. Toxic effect of visible light on Drosophila lifespan depending upon diet protein content.

    Science.gov (United States)

    Shen, Jie; Zhu, Xiang; Gu, Yitian; Zhang, Chiqian; Huang, Jiahong; Qing, Xiao

    2018-03-01

    We investigated the toxic effect of visible light on Drosophila lifespan in both sexes. The toxic effect of ultraviolet (UV) light on organisms is well known. However, the effects of illumination with visible light remain unclear. Here, we found that visible light could be toxic to Drosophila survival, depending on the protein content in diet. In addition, further analysis revealed significant interaction between light and sex, and showed that strong light shortened life span by causing opposite direction changes in mortality rate parameters in females versus males. Our findings suggest that photoageing may be a general phenomenon, and support the theory of sexual antagonistic pleiotropy in aging intervention. The results caution that exposure to visible light could be hazardous to life span and suggest that identification of the underlying mechanism would allow better understanding of aging intervention.

  10. Involvement of adenosine monophosphate-activated protein kinase in the influence of timed high-fat evening diet on the hepatic clock and lipogenic gene expression in mice.

    Science.gov (United States)

    Huang, Yan; Zhu, Zengyan; Xie, Meilin; Xue, Jie

    2015-09-01

    A high-fat diet may result in changes in hepatic clock gene expression, but potential mechanisms are not yet elucidated. Adenosine monophosphate-activated protein kinase (AMPK) is a serine/threonine protein kinase that is recognized as a key regulator of energy metabolism and certain clock genes. Therefore, we hypothesized that AMPK may be involved in the alteration of hepatic clock gene expression under a high-fat environment. This study aimed to examine the effects of timed high-fat evening diet on the activity of hepatic AMPK, clock genes, and lipogenic genes. Mice with hyperlipidemic fatty livers were induced by orally administering high-fat milk via gavage every evening (19:00-20:00) for 6 weeks. Results showed that timed high-fat diet in the evening not only decreased the hepatic AMPK protein expression and activity but also disturbed its circadian rhythm. Accordingly, the hepatic clock genes, including clock, brain-muscle-Arnt-like 1, cryptochrome 2, and period 2, exhibited prominent changes in their expression rhythms and/or amplitudes. The diurnal rhythms of the messenger RNA expression of peroxisome proliferator-activated receptorα, acetyl-CoA carboxylase 1α, and carnitine palmitoyltransferase 1 were also disrupted; the amplitude of peroxisome proliferator-activated receptorγcoactivator 1α was significantly decreased at 3 time points, and fatty liver was observed. These findings demonstrate that timed high-fat diet at night can change hepatic AMPK protein levels, activity, and circadian rhythm, which may subsequently alter the circadian expression of several hepatic clock genes and finally result in the disorder of hepatic lipogenic gene expression and the formation of fatty liver. Copyright © 2015 Elsevier Inc. All rights reserved.

  11. Functional dissection of the Hox protein Abdominal-B in Drosophila cell culture

    International Nuclear Information System (INIS)

    Zhai, Zongzhao; Yang, Xingke; Lohmann, Ingrid

    2011-01-01

    Highlights: ► ct340 CRM was identified to be the posterior spiracle enhancer of gene cut. ► ct340 is under the direct transcriptional control of Hox protein Abd-B. ► An efficient cloning system was developed to assay protein–DNA interaction. ► New features of Abd-B dependent target gene regulation were detected. -- Abstract: Hox transcription factors regulate the morphogenesis along the anterior–posterior (A/P) body axis through the interaction with small cis-regulatory modules (CRMs) of their target gene, however so far very few Hox CRMs are known and have been analyzed in detail. In this study we have identified a new Hox CRM, ct340, which guides the expression of the cell type specification gene cut (ct) in the posterior spiracle under the direct control of the Hox protein Abdominal-B (Abd-B). Using the ct340 enhancer activity as readout, an efficient cloning system to generate VP16 activation domain fusion protein was developed to unambiguously test protein–DNA interaction in Drosophila cell culture. By functionally dissecting the Abd-B protein, new features of Abd-B dependent target gene regulation were detected. Due to its easy adaptability, this system can be generally used to map functional domains within sequence-specific transcriptional factors in Drosophila cell culture, and thus provide preliminary knowledge of the protein functional domain structure for further in vivo analysis.

  12. Overlapping functions of argonaute proteins in patterning and morphogenesis of Drosophila embryos.

    Directory of Open Access Journals (Sweden)

    Wibke J Meyer

    2006-08-01

    Full Text Available Argonaute proteins are essential components of the molecular machinery that drives RNA silencing. In Drosophila, different members of the Argonaute family of proteins have been assigned to distinct RNA silencing pathways. While Ago1 is required for microRNA function, Ago2 is a crucial component of the RNA-induced silencing complex in siRNA-triggered RNA interference. Drosophila Ago2 contains an unusual amino-terminus with two types of imperfect glutamine-rich repeats (GRRs of unknown function. Here we show that the GRRs of Ago2 are essential for the normal function of the protein. Alleles with reduced numbers of GRRs cause specific disruptions in two morphogenetic processes associated with the midblastula transition: membrane growth and microtubule-based organelle transport. These defects do not appear to result from disruption of siRNA-dependent processes but rather suggest an interference of the mutant Ago2 proteins in an Ago1-dependent pathway. Using loss-of-function alleles, we further demonstrate that Ago1 and Ago2 act in a partially redundant manner to control the expression of the segment-polarity gene wingless in the early embryo. Our findings argue against a strict separation of Ago1 and Ago2 functions and suggest that these proteins act in concert to control key steps of the midblastula transition and of segmental patterning.

  13. Dietary protein content affects evolution for body size, body fat and viability in Drosophila melanogaster

    DEFF Research Database (Denmark)

    Kristensen, Torsten N; Overgaard, Johannes; Loeschcke, Volker

    2011-01-01

    The ability to use different food sources is likely to be under strong selection if organisms are faced with natural variation in macro-nutrient (protein, carbohydrate and lipid) availabilities. Here, we use experimental evolution to study how variable dietary protein content affects adult body...... composition and developmental success in Drosophila melanogaster. We reared flies on either a standard diet or a protein-enriched diet for 17 generations before testing them on both diet types. Flies from lines selected on protein-rich diet produced phenotypes with higher total body mass and relative lipid...... content when compared with those selected on a standard diet, irrespective of which of the two diets they were tested on. However, selection on protein-rich diet incurred a cost as flies reared on this diet had markedly lower developmental success in terms of egg-to-adult viability on both medium types...

  14. INHIBITION OF THE DNA-BINDING ACTIVITY OF DROSOPHILA SUPPRESSOR OF HAIRLESS AND OF ITS HUMAN HOMOLOG, KBF2/RBP-J-KAPPA, BY DIRECT PROTEIN-PROTEIN INTERACTION WITH DROSOPHILA HAIRLESS

    NARCIS (Netherlands)

    BROU, C; LOGEAT, F; LECOURTOIS, M; VANDEKERCKHOVE, Joël; KOURILSKY, P; SCHWEISGUTH, F; ISRAEL, A

    1994-01-01

    We have purified the sequence-specific DNA-binding protein KBF2 and cloned the corresponding cDNA, which is derived from the previously described RBP-J kappa gene, the human homolog of the Drosophila Suppressor of Hairless [Su(H)] gene. Deletion studies of the RBP-J kappa and Su(H) proteins allowed

  15. Naltrexone Reverses Ethanol Preference and Protein Kinase C Activation in Drosophila melanogaster

    Directory of Open Access Journals (Sweden)

    Rajeswari Koyyada

    2018-03-01

    Full Text Available Alcohol use disorder (AUD is a major health, social and economic problem for which there are few effective treatments. The opiate antagonist naltrexone is currently prescribed clinically with mixed success. We have used naltrexone in an established behavioral assay (CAFE in Drosophila melanogaster that measures the flies' preference for ethanol-containing food. We have confirmed that Drosophila exposed to ethanol develop a preference toward this drug and we demonstrate that naltrexone, in a dose dependant manner, reverses the ethanol-induced ethanol preference. This effect is not permanent, as preference for alcohol returns after discontinuing naltrexone. Additionally, naltrexone reduced the alcohol-induced increase in protein kinase C activity. These findings are of interest because they confirm that Drosophila is a useful model for studying human responses to addictive drugs. Additionally because of the lack of a closely conserved opiate system in insects, our results could either indicate that a functionally related system does exist in insects or that in insects, and potentially also in mammals, naltrexone binds to alternative sites. Identifying such sites could lead to improved treatment strategies for AUD.

  16. Naltrexone Reverses Ethanol Preference and Protein Kinase C Activation in Drosophila melanogaster

    Science.gov (United States)

    Koyyada, Rajeswari; Latchooman, Nilesh; Jonaitis, Julius; Ayoub, Samir S.; Corcoran, Olivia; Casalotti, Stefano O.

    2018-01-01

    Alcohol use disorder (AUD) is a major health, social and economic problem for which there are few effective treatments. The opiate antagonist naltrexone is currently prescribed clinically with mixed success. We have used naltrexone in an established behavioral assay (CAFE) in Drosophila melanogaster that measures the flies' preference for ethanol-containing food. We have confirmed that Drosophila exposed to ethanol develop a preference toward this drug and we demonstrate that naltrexone, in a dose dependant manner, reverses the ethanol-induced ethanol preference. This effect is not permanent, as preference for alcohol returns after discontinuing naltrexone. Additionally, naltrexone reduced the alcohol-induced increase in protein kinase C activity. These findings are of interest because they confirm that Drosophila is a useful model for studying human responses to addictive drugs. Additionally because of the lack of a closely conserved opiate system in insects, our results could either indicate that a functionally related system does exist in insects or that in insects, and potentially also in mammals, naltrexone binds to alternative sites. Identifying such sites could lead to improved treatment strategies for AUD. PMID:29593550

  17. Genetic Variation and Its Reflection on Posttranslational Modifications in Frequency Clock and Mating Type a-1 Proteins in Sordaria fimicola

    Directory of Open Access Journals (Sweden)

    Rabia Arif

    2017-01-01

    Full Text Available Posttranslational modifications (PTMs occur in all essential proteins taking command of their functions. There are many domains inside proteins where modifications take place on side-chains of amino acids through various enzymes to generate different species of proteins. In this manuscript we have, for the first time, predicted posttranslational modifications of frequency clock and mating type a-1 proteins in Sordaria fimicola collected from different sites to see the effect of environment on proteins or various amino acids pickings and their ultimate impact on consensus sequences present in mating type proteins using bioinformatics tools. Furthermore, we have also measured and walked through genomic DNA of various Sordaria strains to determine genetic diversity by genotyping the short sequence repeats (SSRs of wild strains of S. fimicola collected from contrasting environments of two opposing slopes (harsh and xeric south facing slope and mild north facing slope of Evolution Canyon (EC, Israel. Based on the whole genome sequence of S. macrospora, we targeted 20 genomic regions in S. fimicola which contain short sequence repeats (SSRs. Our data revealed genetic variations in strains from south facing slope and these findings assist in the hypothesis that genetic variations caused by stressful environments lead to evolution.

  18. Genetic Variation and Its Reflection on Posttranslational Modifications in Frequency Clock and Mating Type a-1 Proteins in Sordaria fimicola.

    Science.gov (United States)

    Arif, Rabia; Akram, Faiza; Jamil, Tazeen; Mukhtar, Hamid; Lee, Siu Fai; Saleem, Muhammad

    2017-01-01

    Posttranslational modifications (PTMs) occur in all essential proteins taking command of their functions. There are many domains inside proteins where modifications take place on side-chains of amino acids through various enzymes to generate different species of proteins. In this manuscript we have, for the first time, predicted posttranslational modifications of frequency clock and mating type a-1 proteins in Sordaria fimicola collected from different sites to see the effect of environment on proteins or various amino acids pickings and their ultimate impact on consensus sequences present in mating type proteins using bioinformatics tools. Furthermore, we have also measured and walked through genomic DNA of various Sordaria strains to determine genetic diversity by genotyping the short sequence repeats (SSRs) of wild strains of S. fimicola collected from contrasting environments of two opposing slopes (harsh and xeric south facing slope and mild north facing slope) of Evolution Canyon (EC), Israel. Based on the whole genome sequence of S. macrospora , we targeted 20 genomic regions in S. fimicola which contain short sequence repeats (SSRs). Our data revealed genetic variations in strains from south facing slope and these findings assist in the hypothesis that genetic variations caused by stressful environments lead to evolution.

  19. Loss of the clock protein PER2 shortens the erythrocyte life span in mice.

    Science.gov (United States)

    Sun, Qi; Zhao, Yue; Yang, Yunxia; Yang, Xiao; Li, Minghui; Xu, Xi; Wen, Dan; Wang, Junsong; Zhang, Jianfa

    2017-07-28

    Cell proliferation and release from the bone marrow have been demonstrated to be controlled by circadian rhythms in both humans and mice. However, it is unclear whether local circadian clocks in the bone marrow influence physiological functions and life span of erythrocytes. Here, we report that loss of the clock gene Per2 significantly decreased erythrocyte life span. Mice deficient in Per2 were more susceptible to acute stresses in the erythrocytes, becoming severely anemic upon phenylhydrazine, osmotic, and H 2 O 2 challenges. 1 H NMR-based metabolomics analysis revealed that the Per2 depletion causes significant changes in metabolic profiles of erythrocytes, including increased lactate and decreased ATP levels compared with wild-type mice. The lower ATP levels were associated with hyperfunction of Na + /K + -ATPase activity in Per2 -null erythrocytes, and inhibition of Na + /K + -ATPase activity by ouabain efficiently rescued ATP levels. Per2 -null mice displayed increased levels of Na + /K + -ATPase α1 (ATP1A1) in the erythrocyte membrane, and transfection of Per2 cDNA into the erythroleukemic cell line TF-1 inhibited Atp1a1 expression. Furthermore, we observed that PER2 regulates Atp1a1 transcription through interacting with trans-acting transcription factor 1 (SP1). Our findings reveal that Per2 function in the bone marrow is required for the regulation of life span in circulating erythrocytes. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  20. Growth inhibition and differences in protein profiles in azadirachtin-treated Drosophila melanogaster larvae.

    Science.gov (United States)

    Wang, Hao; Lai, Duo; Yuan, Mei; Xu, Hanhong

    2014-04-01

    Azadirachtin A is a very effective biopesticide widely used in insect pest control. It has strong antifeeding and growth inhibitory activity against most insects, however, its mode of action is still unclear. Proteomic experiments using 2DE indicate significant effects of Azadirachtin A on the amount of proteins related to growth inhibition in Drosophila melanogaster larvae. Twenty-one spots with different intensity in azadirachtin-treated larvae were identified. These proteins are involved in cytoskeletal organization, transcription and translation, hormonal regulation, and energy metabolism. Protein network analysis reveals heat shock protein 23 to be a potential target of azadirachtin. These results provide new insights into understanding the mechanism of growth inhibition in insects in response to azadirachtin. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  1. Biochemical and genetic analysis of the Drk SH2/SH3 adaptor protein of Drosophila.

    OpenAIRE

    Raabe, T; Olivier, J P; Dickson, B J; Liu, X; Gish, G D; Pawson, T; Hafen, E

    1995-01-01

    The Drk SH3-SH2-SH3 adaptor protein has been genetically identified in a screen for rate-limiting components acting downstream of the Sevenless (Sev) receptor tyrosine kinase in the developing eye of Drosophila. It provides a link between the activated Sev receptor and Sos, a guanine nucleotide release factor that activates Ras1. We have used a combined biochemical and genetic approach to study the interactions between Sev, Drk and Sos. We show that Tyr2546 in the cytoplasmic tail of Sev is r...

  2. Drosophila photoreceptor axon guidance and targeting requires the dreadlocks SH2/SH3 adapter protein.

    Science.gov (United States)

    Garrity, P A; Rao, Y; Salecker, I; McGlade, J; Pawson, T; Zipursky, S L

    1996-05-31

    Mutations in the Drosophila gene dreadlocks (dock) disrupt photoreceptor cell (R cell) axon guidance and targeting. Genetic mosaic analysis and cell-type-specific expression of dock transgenes demonstrate dock is required in R cells for proper innervation. Dock protein contains one SH2 and three SH3 domains, implicating it in tyrosine kinase signaling, and is highly related to the human proto-oncogene Nck. Dock expression is detected in R cell growth cones in the target region. We propose Dock transmits signals in the growth cone in response to guidance and targeting cues. These findings provide an important step for dissection of signaling pathways regulating growth cone motility.

  3. A family of related proteins is encoded by the major Drosophila heat shock gene family

    International Nuclear Information System (INIS)

    Wadsworth, S.C.

    1982-01-01

    At least four proteins of 70,000 to 75,000 molecular weight (70-75K) were synthesized from mRNA which hybridized with a cloned heat shock gene previously shown to be localized to the 87A and 87C heat shock puff sites. These in vitro-synthesized proteins were indistinguishable from in vivo-synthesized heat shock-induced proteins when analyzed on sodium dodecyl sulfate-polyacrylamide gels. A comparison of the pattern of this group of proteins synthesized in vivo during a 5-min pulse or during continuous labeling indicates that the 72-75K proteins are probably not kinetic precursors to the major 70K heat shock protein. Partial digestion products generated with V8 protease indicated that the 70-75K heat shock proteins are closely related, but that there are clear differences between them. The partial digestion patterns obtained from heat shock proteins from the Kc cell line and from the Oregon R strain of Drosophila melanogaster are very similar. Genetic analysis of the patterns of 70-75K heat shock protein synthesis indicated that the genes encoding at least two of the three 72-75K heat shock proteins are located outside of the major 87A and 87C puff sites

  4. The Heat Shock Protein 26 Gene is Required for Ethanol Tolerance in Drosophila

    Directory of Open Access Journals (Sweden)

    Awoyemi A. Awofala

    2011-01-01

    Full Text Available Stress plays an important role in drug- and addiction-related behaviours. However, the mechanisms underlying these behavioural responses are still poorly understood. In the light of recent reports that show consistent regulation of many genes encoding stress proteins including heat shock proteins following ethanol exposure in Drosophila , it was hypothesised that transition to alcohol dependence may involve the dysregulation of the circuits that mediate behavioural responses to stressors. Thus, behavioural genetic methodologies were used to investigate the role of the Drosophila hsp26 gene, a small heat shock protein coding gene which is induced in response to various stresses, in the development of rapid tolerance to ethanol sedation. Rapid tolerance was quantified as the percentage difference in the mean sedation times between the second and first ethanol exposure. Two independently isolated P-element mutations near the hsp26 gene eliminated the capacity for tolerance. In addition, RNAi-mediated functional knockdown of hsp26 expression in the glial cells and the whole nervous system also caused a defect in tolerance development. The rapid tolerance phenotype of the hsp26 mutants was rescued by the expression of the wild-type hsp26 gene in the nervous system. None of these manipulations of the hsp26 gene caused changes in the rate of ethanol absorption. Hsp26 genes are evolutionary conserved, thus the role of hsp26 in ethanol tolerance may present a new direction for research into alcohol dependency.

  5. Circadian Clock Protein Content and Daily Rhythm of Locomotor Activity Are Altered after Chronic Exposure to Lead in Rat

    Science.gov (United States)

    Sabbar, Mariam; Dkhissi-Benyahya, Ouria; Benazzouz, Abdelhamid; Lakhdar-Ghazal, Nouria

    2017-01-01

    Lead exposure has been reported to produce many clinical features, including parkinsonism. However, its consequences on the circadian rhythms are still unknown. Here we aimed to examine the circadian rhythms of locomotor activity following lead intoxication and investigate the mechanisms by which lead may induce alterations of circadian rhythms in rats. Male Wistar rats were injected with lead or sodium acetate (10 mg/kg/day, i.p.) during 4 weeks. Both groups were tested in the “open field” to quantify the exploratory activity and in the rotarod to evaluate motor coordination. Then, animals were submitted to continuous 24 h recordings of locomotor activity under 14/10 Light/dark (14/10 LD) cycle and in complete darkness (DD). At the end of experiments, the clock proteins BMAL1, PER1-2, and CRY1-2 were assayed in the suprachiasmatic nucleus (SCN) using immunohistochemistry. We showed that lead significantly reduced the number of crossing in the open field, impaired motor coordination and altered the daily locomotor activity rhythm. When the LD cycle was advanced by 6 h, both groups adjusted their daily locomotor activity to the new LD cycle with high onset variability in lead-intoxicated rats compared to controls. Lead also led to a decrease in the number of immunoreactive cells (ir-) of BMAL1, PER1, and PER2 without affecting the number of ir-CRY1 and ir-CRY2 cells in the SCN. Our data provide strong evidence that lead intoxication disturbs the rhythm of locomotor activity and alters clock proteins expression in the SCN. They contribute to the understanding of the mechanism by which lead induce circadian rhythms disturbances. PMID:28970786

  6. Codon usage regulates protein structure and function by affecting translation elongation speed in Drosophila cells.

    Science.gov (United States)

    Zhao, Fangzhou; Yu, Chien-Hung; Liu, Yi

    2017-08-21

    Codon usage biases are found in all eukaryotic and prokaryotic genomes and have been proposed to regulate different aspects of translation process. Codon optimality has been shown to regulate translation elongation speed in fungal systems, but its effect on translation elongation speed in animal systems is not clear. In this study, we used a Drosophila cell-free translation system to directly compare the velocity of mRNA translation elongation. Our results demonstrate that optimal synonymous codons speed up translation elongation while non-optimal codons slow down translation. In addition, codon usage regulates ribosome movement and stalling on mRNA during translation. Finally, we show that codon usage affects protein structure and function in vitro and in Drosophila cells. Together, these results suggest that the effect of codon usage on translation elongation speed is a conserved mechanism from fungi to animals that can affect protein folding in eukaryotic organisms. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

  7. The insulator protein SU(HW fine-tunes nuclear lamina interactions of the Drosophila genome.

    Directory of Open Access Journals (Sweden)

    Joke G van Bemmel

    Full Text Available Specific interactions of the genome with the nuclear lamina (NL are thought to assist chromosome folding inside the nucleus and to contribute to the regulation of gene expression. High-resolution mapping has recently identified hundreds of large, sharply defined lamina-associated domains (LADs in the human genome, and suggested that the insulator protein CTCF may help to demarcate these domains. Here, we report the detailed structure of LADs in Drosophila cells, and investigate the putative roles of five insulator proteins in LAD organization. We found that the Drosophila genome is also organized in discrete LADs, which are about five times smaller than human LADs but contain on average a similar number of genes. Systematic comparison to new and published insulator binding maps shows that only SU(HW binds preferentially at LAD borders and at specific positions inside LADs, while GAF, CTCF, BEAF-32 and DWG are mostly absent from these regions. By knockdown and overexpression studies we demonstrate that SU(HW weakens genome - NL interactions through a local antagonistic effect, but we did not obtain evidence that it is essential for border formation. Our results provide insights into the evolution of LAD organization and identify SU(HW as a fine-tuner of genome - NL interactions.

  8. Fatty-acid binding proteins modulate sleep and enhance long-term memory consolidation in Drosophila.

    Directory of Open Access Journals (Sweden)

    Jason R Gerstner

    2011-01-01

    Full Text Available Sleep is thought to be important for memory consolidation, since sleep deprivation has been shown to interfere with memory processing. However, the effects of augmenting sleep on memory formation are not well known, and testing the role of sleep in memory enhancement has been limited to pharmacological and behavioral approaches. Here we test the effect of overexpressing the brain-type fatty acid binding protein (Fabp7 on sleep and long-term memory (LTM formation in Drosophila melanogaster. Transgenic flies carrying the murine Fabp7 or the Drosophila homologue dFabp had reduced baseline sleep but normal LTM, while Fabp induction produced increases in both net sleep and LTM. We also define a post-training consolidation "window" that is sufficient for the observed Fabp-mediated memory enhancement. Since Fabp overexpression increases consolidated daytime sleep bouts, these data support a role for longer naps in improving memory and provide a novel role for lipid-binding proteins in regulating memory consolidation concurrently with changes in behavioral state.

  9. Ribosomal S6 Kinase Cooperates with Casein Kinase 2 to Modulate the Drosophila Circadian Molecular Oscillator

    Science.gov (United States)

    Akten, Bikem; Tangredi, Michelle M.; Jauch, Eike; Roberts, Mary A.; Ng, Fanny; Raabe, Thomas; Jackson, F. Rob

    2009-01-01

    There is a universal requirement for post-translational regulatory mechanisms in circadian clock systems. Previous work in Drosophila has identified several kinases, phosphatases and an E3 ligase that are critical for determining the nuclear translocation and/or stability of clock proteins. The present study evaluated the function of p90 ribosomal S6 kinase (RSK) in the Drosophila circadian system. In mammals, RSK1 is a light- and clock-regulated kinase known to be activated by the MAPK pathway, but there is no direct evidence that it functions as a component of the circadian system. Here, we show that Drosophila S6KII RNA displays rhythms in abundance, indicative of circadian control. Importantly, an S6KII null mutant exhibits a short-period circadian phenotype that can be rescued by expression of the wild-type gene in clock neurons, indicating a role for S6KII in the molecular oscillator. Peak PER clock protein expression is elevated in the mutant, indicative of enhanced stability, whereas per mRNA level is decreased, consistent with enhanced feedback repression. Gene reporter assays show that decreased S6KII is associated with increased PER repression. Surprisingly, we demonstrate a physical interaction between S6KII and the Casein Kinase 2 regulatory subunit (CK2β), suggesting a functional relationship between the two kinases. In support of such a relationship, there are genetic interactions between S6KII and CK2 mutations, in vivo, which indicate that CK2 activity is required for S6KII action. We propose that the two kinases cooperate within clock neurons to fine-tune circadian period, improving the precision of the clock mechanism. PMID:19144847

  10. Reproductive hacking. A male seminal protein acts through intact reproductive pathways in female Drosophila.

    Science.gov (United States)

    Rubinstein, C Dustin; Wolfner, Mariana F

    2014-01-01

    Seminal proteins are critical for reproductive success in all animals that have been studied. Although seminal proteins have been identified in many taxa, and female reproductive responses to receipt of these proteins have been documented in several, little is understood about the mechanisms by which seminal proteins affect female reproductive physiology. To explore this topic, we investigated how a Drosophila seminal protein, ovulin, increases ovulation rate in mated females. Ovulation is a relatively simple physiological process, with known female regulators: previous studies have shown that ovulation rate is promoted by the neuromodulator octopamine (OA) in D. melanogaster and other insects. We found that ovulin stimulates ovulation by increasing OA signaling in the female. This finding supports a model in which a male seminal protein acts through "hacking" a well-conserved, regulatory system females use to adjust reproductive output, rather than acting downstream of female mechanisms of control or in parallel pathways altogether. We also discuss similarities between 2 forms of intersexual control of behavior through chemical communication: seminal proteins and pheromones.

  11. Widespread Positive Selection Drives Differentiation of Centromeric Proteins in the Drosophila melanogaster subgroup.

    Science.gov (United States)

    Beck, Emily A; Llopart, Ana

    2015-11-25

    Rapid evolution of centromeric satellite repeats is thought to cause compensatory amino acid evolution in interacting centromere-associated kinetochore proteins. Cid, a protein that mediates kinetochore/centromere interactions, displays particularly high amino acid turnover. Rapid evolution of both Cid and centromeric satellite repeats led us to hypothesize that the apparent compensatory evolution may extend to interacting partners in the Condensin I complex (i.e., SMC2, SMC4, Cap-H, Cap-D2, and Cap-G) and HP1s. Missense mutations in these proteins often result in improper centromere formation and aberrant chromosome segregation, thus selection for maintained function and coevolution among proteins of the complex is likely strong. Here, we report evidence of rapid evolution and recurrent positive selection in seven centromere-associated proteins in species of the Drosophila melanogaster subgroup, and further postulate that positive selection on these proteins could be a result of centromere drive and compensatory changes, with kinetochore proteins competing for optimal spindle attachment.

  12. Characterization of the Drosophila ortholog of the human Usher Syndrome type 1G protein sans.

    Directory of Open Access Journals (Sweden)

    Fabio Demontis

    Full Text Available BACKGROUND: The Usher syndrome (USH is the most frequent deaf-blindness hereditary disease in humans. Deafness is attributed to the disorganization of stereocilia in the inner ear. USH1, the most severe subtype, is associated with mutations in genes encoding myosin VIIa, harmonin, cadherin 23, protocadherin 15, and sans. Myosin VIIa, harmonin, cadherin 23, and protocadherin 15 physically interact in vitro and localize to stereocilia tips in vivo, indicating that they form functional complexes. Sans, in contrast, localizes to vesicle-like structures beneath the apical membrane of stereocilia-displaying hair cells. How mutations in sans result in deafness and blindness is not well understood. Orthologs of myosin VIIa and protocadherin 15 have been identified in Drosophila melanogaster and their genetic analysis has identified essential roles in auditory perception and microvilli morphogenesis, respectively. PRINCIPAL FINDINGS: Here, we have identified and characterized the Drosophila ortholog of human sans. Drosophila Sans is expressed in tubular organs of the embryo, in lens-secreting cone cells of the adult eye, and in microvilli-displaying follicle cells during oogenesis. Sans mutants are viable, fertile, and mutant follicle cells appear to form microvilli, indicating that Sans is dispensable for fly development and microvilli morphogenesis in the follicle epithelium. In follicle cells, Sans protein localizes, similar to its vertebrate ortholog, to intracellular punctate structures, which we have identified as early endosomes associated with the syntaxin Avalanche. CONCLUSIONS: Our work is consistent with an evolutionary conserved function of Sans in vesicle trafficking. Furthermore it provides a significant basis for further understanding of the role of this Usher syndrome ortholog in development and disease.

  13. The role of the Drosophila LAMMER protein kinase DOA in somatic ...

    Indian Academy of Sciences (India)

    2010-09-06

    Sep 6, 2010 ... Somatic sexual identity in Drosophila melanogaster is under the control of a ... between stages 12 and 14, in response to an X to autosome .... MER kinases used were Drosophila DOA, mouse CLK1, human CLK2 and.

  14. Time of Day Influences Memory Formation and dCREB2 Proteins in Drosophila

    Directory of Open Access Journals (Sweden)

    Robin eFropf

    2014-03-01

    Full Text Available Many biological phenomena oscillate under the control of the circadian system, exhibiting peaks and troughs of activity across the day/night cycle. In most animal models, memory formation also exhibits this property, but the underlying neuronal and molecular mechanisms remain unclear. The dCREB2 transcription factor shows circadian regulated oscillations in its activity, and has been shown to be important for both circadian biology and memory formation. We show that the time-of-day (TOD of behavioral training affects Drosophila memory formation. dCREB2 exhibits complex changes in protein levels across the daytime and nighttime, and these changes in protein abundance are likely to contribute to oscillations in dCREB2 activity and TOD effects on memory formation.

  15. A Drosophila protein family implicated in pheromone perception is related to Tay-Sachs GM2-activator protein.

    Science.gov (United States)

    Starostina, Elena; Xu, Aiguo; Lin, Heping; Pikielny, Claudio W

    2009-01-02

    Low volatility, lipid-like cuticular hydrocarbon pheromones produced by Drosophila melanogaster females play an essential role in triggering and modulating mating behavior, but the chemosensory mechanisms involved remain poorly understood. Recently, we showed that the CheB42a protein, which is expressed in only 10 pheromone-sensing taste hairs on the front legs of males, modulates progression to late stages of male courtship behavior in response to female-specific cuticular hydrocarbons. Here we report that expression of all 12 genes in the CheB gene family is predominantly or exclusively gustatory-specific, and occurs in many different, often non-overlapping patterns. Only the Gr family of gustatory receptor genes displays a comparable variety of gustatory-specific expression patterns. Unlike Grs, however, expression of all but one CheB gene is sexually dimorphic. Like CheB42a, other CheBs may therefore function specifically in gustatory perception of pheromones. We also show that CheBs belong to the ML superfamily of lipid-binding proteins, and are most similar to human GM2-activator protein (GM2-AP). In particular, GM2-AP residues involved in ligand binding are conserved in CheBs but not in other ML proteins. Finally, CheB42a is specifically secreted into the inner lumen of pheromone-sensing taste hairs, where pheromones interact with membrane-bound receptors. We propose that CheB proteins interact directly with lipid-like Drosophila pheromones and modulate their detection by the gustatory signal transduction machinery. Furthermore, as loss of GM2-AP in Tay-Sachs disease prevents degradation of GM2 gangliosides and results in neurodegeneration, the function of CheBs in pheromone response may involve biochemical mechanisms critical for lipid metabolism in human neurons.

  16. A Drosophila Protein Family Implicated in Pheromone Perception Is Related to Tay-Sachs GM2-Activator Protein*

    Science.gov (United States)

    Starostina, Elena; Xu, Aiguo; Lin, Heping; Pikielny, Claudio W.

    2009-01-01

    Low volatility, lipid-like cuticular hydrocarbon pheromones produced by Drosophila melanogaster females play an essential role in triggering and modulating mating behavior, but the chemosensory mechanisms involved remain poorly understood. Recently, we showed that the CheB42a protein, which is expressed in only 10 pheromone-sensing taste hairs on the front legs of males, modulates progression to late stages of male courtship behavior in response to female-specific cuticular hydrocarbons. Here we report that expression of all 12 genes in the CheB gene family is predominantly or exclusively gustatory-specific, and occurs in many different, often non-overlapping patterns. Only the Gr family of gustatory receptor genes displays a comparable variety of gustatory-specific expression patterns. Unlike Grs, however, expression of all but one CheB gene is sexually dimorphic. Like CheB42a, other CheBs may therefore function specifically in gustatory perception of pheromones. We also show that CheBs belong to the ML superfamily of lipid-binding proteins, and are most similar to human GM2-activator protein (GM2-AP). In particular, GM2-AP residues involved in ligand binding are conserved in CheBs but not in other ML proteins. Finally, CheB42a is specifically secreted into the inner lumen of pheromone-sensing taste hairs, where pheromones interact with membrane-bound receptors. We propose that CheB proteins interact directly with lipid-like Drosophila pheromones and modulate their detection by the gustatory signal transduction machinery. Furthermore, as loss of GM2-AP in Tay-Sachs disease prevents degradation of GM2 gangliosides and results in neurodegeneration, the function of CheBs in pheromone response may involve biochemical mechanisms critical for lipid metabolism in human neurons. PMID:18952610

  17. A novel Drosophila Girdin-like protein is involved in Akt pathway control of cell size

    Energy Technology Data Exchange (ETDEWEB)

    Puseenam, Aekkachai [Department of Applied Biology, Kyoto Institute of Technology, Sakyo-ku, Kyoto 606-8585 (Japan); Insect Biomedical Research Center, Kyoto Institute of Technology, Sakyo-ku, Kyoto 606-8585 (Japan); Yoshioka, Yasuhide [Department of Applied Biology, Kyoto Institute of Technology, Sakyo-ku, Kyoto 606-8585 (Japan); Venture Laboratory, Kyoto Institute of Technology, Sakyo-ku, Kyoto 606-8585 (Japan); Nagai, Rika [Department of Applied Biology, Kyoto Institute of Technology, Sakyo-ku, Kyoto 606-8585 (Japan); Insect Biomedical Research Center, Kyoto Institute of Technology, Sakyo-ku, Kyoto 606-8585 (Japan); Hashimoto, Reina [Department of Applied Biology, Kyoto Institute of Technology, Sakyo-ku, Kyoto 606-8585 (Japan); Venture Laboratory, Kyoto Institute of Technology, Sakyo-ku, Kyoto 606-8585 (Japan); Suyari, Osamu [Insect Biomedical Research Center, Kyoto Institute of Technology, Sakyo-ku, Kyoto 606-8585 (Japan); Itoh, Masanobu [Department of Applied Biology, Kyoto Institute of Technology, Sakyo-ku, Kyoto 606-8585 (Japan); Insect Biomedical Research Center, Kyoto Institute of Technology, Sakyo-ku, Kyoto 606-8585 (Japan); Enomoto, Atsushi [Department of Pathology, Center for Neurological Disease and Cancer, Nagoya University Graduate School of Medicine, Showa-ku, Nagoya, Aichi 466-8550 (Japan); Takahashi, Masahide [Insect Biomedical Research Center, Kyoto Institute of Technology, Sakyo-ku, Kyoto 606-8585 (Japan); Department of Pathology, Center for Neurological Disease and Cancer, Nagoya University Graduate School of Medicine, Showa-ku, Nagoya, Aichi 466-8550 (Japan); Yamaguchi, Masamitsu, E-mail: myamaguc@kit.ac.jp [Department of Applied Biology, Kyoto Institute of Technology, Sakyo-ku, Kyoto 606-8585 (Japan); Insect Biomedical Research Center, Kyoto Institute of Technology, Sakyo-ku, Kyoto 606-8585 (Japan)

    2009-11-15

    The Akt signaling pathway is well known to regulate cell proliferation and growth. Girdin, a novel substrate of Akt, plays a crucial role in organization of the actin cytoskeleton and cell motility under the control of Akt. We here identified a novel Girdin-like protein in Drosophila (dGirdin), which has two isoforms, dGirdin PA and dGirdin PB. dGirdin shows high homology with human Girdin in the N-terminal and coiled-coil domains, while diverging at the C-terminal domain. On establishment of transgenic fly lines, featuring knockdown or overexpression of dGirdin in vivo, overexpression in the wing disc cells induced ectopic apoptosis, implying a role in directing apoptosis. Knockdown of dGirdin in the Drosophila wing imaginal disc cells resulted in reduction of cell size. Furthermore, this was enhanced by half reduction of the Akt gene dose, suggesting that Akt positively regulates dGirdin. In the wing disc, cells in which dGirdin was knocked down exhibited disruption of actin filaments. From these in vivo analyses, we conclude that dGirdin is required for actin organization and regulation of appropriate cell size under control of the Akt signaling pathway.

  18. The ERM protein Moesin is essential for neuronal morphogenesis and long-term memory in Drosophila.

    Science.gov (United States)

    Freymuth, Patrick S; Fitzsimons, Helen L

    2017-08-29

    Moesin is a cytoskeletal adaptor protein that plays an important role in modification of the actin cytoskeleton. Rearrangement of the actin cytoskeleton drives both neuronal morphogenesis and the structural changes in neurons that are required for long-term memory formation. Moesin has been identified as a candidate memory gene in Drosophila, however, whether it is required for memory formation has not been evaluated. Here, we investigate the role of Moesin in neuronal morphogenesis and in short- and long-term memory formation in the courtship suppression assay, a model of associative memory. We found that both knockdown and overexpression of Moesin led to defects in axon growth and guidance as well as dendritic arborization. Moreover, reduction of Moesin expression or expression of a constitutively active phosphomimetic in the adult Drosophila brain had no effect on short term memory, but prevented long-term memory formation, an effect that was independent of its role in development. These results indicate a critical role for Moesin in both neuronal morphogenesis and long-term memory formation.

  19. M6 membrane protein plays an essential role in Drosophila oogenesis.

    Science.gov (United States)

    Zappia, María Paula; Brocco, Marcela Adriana; Billi, Silvia C; Frasch, Alberto C; Ceriani, María Fernanda

    2011-01-01

    We had previously shown that the transmembrane glycoprotein M6a, a member of the proteolipid protein (PLP) family, regulates neurite/filopodium outgrowth, hence, M6a might be involved in neuronal remodeling and differentiation. In this work we focused on M6, the only PLP family member present in Drosophila, and ortholog to M6a. Unexpectedly, we found that decreased expression of M6 leads to female sterility. M6 is expressed in the membrane of the follicular epithelium in ovarioles throughout oogenesis. Phenotypes triggered by M6 downregulation in hypomorphic mutants included egg collapse and egg permeability, thus suggesting M6 involvement in eggshell biosynthesis. In addition, RNAi-mediated M6 knockdown targeted specifically to follicle cells induced an arrest of egg chamber development, revealing that M6 is essential in oogenesis. Interestingly, M6-associated phenotypes evidenced abnormal changes of the follicle cell shape and disrupted follicular epithelium in mid- and late-stage egg chambers. Therefore, we propose that M6 plays a role in follicular epithelium maintenance involving membrane cell remodeling during oogenesis in Drosophila.

  20. A novel Drosophila Girdin-like protein is involved in Akt pathway control of cell size

    International Nuclear Information System (INIS)

    Puseenam, Aekkachai; Yoshioka, Yasuhide; Nagai, Rika; Hashimoto, Reina; Suyari, Osamu; Itoh, Masanobu; Enomoto, Atsushi; Takahashi, Masahide; Yamaguchi, Masamitsu

    2009-01-01

    The Akt signaling pathway is well known to regulate cell proliferation and growth. Girdin, a novel substrate of Akt, plays a crucial role in organization of the actin cytoskeleton and cell motility under the control of Akt. We here identified a novel Girdin-like protein in Drosophila (dGirdin), which has two isoforms, dGirdin PA and dGirdin PB. dGirdin shows high homology with human Girdin in the N-terminal and coiled-coil domains, while diverging at the C-terminal domain. On establishment of transgenic fly lines, featuring knockdown or overexpression of dGirdin in vivo, overexpression in the wing disc cells induced ectopic apoptosis, implying a role in directing apoptosis. Knockdown of dGirdin in the Drosophila wing imaginal disc cells resulted in reduction of cell size. Furthermore, this was enhanced by half reduction of the Akt gene dose, suggesting that Akt positively regulates dGirdin. In the wing disc, cells in which dGirdin was knocked down exhibited disruption of actin filaments. From these in vivo analyses, we conclude that dGirdin is required for actin organization and regulation of appropriate cell size under control of the Akt signaling pathway.

  1. The PP2C Alphabet is a negative regulator of stress-activated protein kinase signaling in Drosophila.

    Science.gov (United States)

    Baril, Caroline; Sahmi, Malha; Ashton-Beaucage, Dariel; Stronach, Beth; Therrien, Marc

    2009-02-01

    The Jun N-terminal kinase and p38 pathways, also known as stress-activated protein kinase (SAPK) pathways, are signaling conduits reiteratively used throughout the development and adult life of metazoans where they play central roles in the control of apoptosis, immune function, and environmental stress responses. We recently identified a Drosophila Ser/Thr phosphatase of the PP2C family, named Alphabet (Alph), which acts as a negative regulator of the Ras/ERK pathway. Here we show that Alph also plays an inhibitory role with respect to Drosophila SAPK signaling during development as well as under stress conditions such as oxidative or genotoxic stresses. Epistasis experiments suggest that Alph acts at a step upstream of the MAPKKs Hep and Lic. Consistent with this interpretation, biochemical experiments identify the upstream MAPKKKs Slpr, Tak1, and Wnd as putative substrates. Together with previous findings, this work identifies Alph as a general attenuator of MAPK signaling in Drosophila.

  2. Positive Selection Drives the Evolution of rhino, a Member of the Heterochromatin Protein 1 Family in Drosophila.

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    2005-07-01

    Full Text Available Heterochromatin comprises a significant component of many eukaryotic genomes. In comparison to euchromatin, heterochromatin is gene poor, transposon rich, and late replicating. It serves many important biological roles, from gene silencing to accurate chromosome segregation, yet little is known about the evolutionary constraints that shape heterochromatin. A complementary approach to the traditional one of directly studying heterochromatic DNA sequence is to study the evolution of proteins that bind and define heterochromatin. One of the best markers for heterochromatin is the heterochromatin protein 1 (HP1, which is an essential, nonhistone chromosomal protein. Here we investigate the molecular evolution of five HP1 paralogs present in Drosophila melanogaster. Three of these paralogs have ubiquitous expression patterns in adult Drosophila tissues, whereas HP1D/rhino and HP1E are expressed predominantly in ovaries and testes respectively. The HP1 paralogs also have distinct localization preferences in Drosophila cells. Thus, Rhino localizes to the heterochromatic compartment in Drosophila tissue culture cells, but in a pattern distinct from HP1A and lysine-9 dimethylated H3. Using molecular evolution and population genetic analyses, we find that rhino has been subject to positive selection in all three domains of the protein: the N-terminal chromo domain, the C-terminal chromo-shadow domain, and the hinge region that connects these two modules. Maximum likelihood analysis of rhino sequences from 20 species of Drosophila reveals that a small number of residues of the chromo and shadow domains have been subject to repeated positive selection. The rapid and positive selection of rhino is highly unusual for a gene encoding a chromosomal protein and suggests that rhino is involved in a genetic conflict that affects the germline, belying the notion that heterochromatin is simply a passive recipient of "junk DNA" in eukaryotic genomes.

  3. Positive selection drives the evolution of rhino, a member of the heterochromatin protein 1 family in Drosophila.

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    Danielle Vermaak

    2005-07-01

    Full Text Available Heterochromatin comprises a significant component of many eukaryotic genomes. In comparison to euchromatin, heterochromatin is gene poor, transposon rich, and late replicating. It serves many important biological roles, from gene silencing to accurate chromosome segregation, yet little is known about the evolutionary constraints that shape heterochromatin. A complementary approach to the traditional one of directly studying heterochromatic DNA sequence is to study the evolution of proteins that bind and define heterochromatin. One of the best markers for heterochromatin is the heterochromatin protein 1 (HP1, which is an essential, nonhistone chromosomal protein. Here we investigate the molecular evolution of five HP1 paralogs present in Drosophila melanogaster. Three of these paralogs have ubiquitous expression patterns in adult Drosophila tissues, whereas HP1D/rhino and HP1E are expressed predominantly in ovaries and testes respectively. The HP1 paralogs also have distinct localization preferences in Drosophila cells. Thus, Rhino localizes to the heterochromatic compartment in Drosophila tissue culture cells, but in a pattern distinct from HP1A and lysine-9 dimethylated H3. Using molecular evolution and population genetic analyses, we find that rhino has been subject to positive selection in all three domains of the protein: the N-terminal chromo domain, the C-terminal chromo-shadow domain, and the hinge region that connects these two modules. Maximum likelihood analysis of rhino sequences from 20 species of Drosophila reveals that a small number of residues of the chromo and shadow domains have been subject to repeated positive selection. The rapid and positive selection of rhino is highly unusual for a gene encoding a chromosomal protein and suggests that rhino is involved in a genetic conflict that affects the germline, belying the notion that heterochromatin is simply a passive recipient of "junk DNA" in eukaryotic genomes.

  4. Time-of-day- and light-dependent expression of ubiquitin protein ligase E3 component N-recognin 4 (UBR4 in the suprachiasmatic nucleus circadian clock.

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    Harrod H Ling

    Full Text Available Circadian rhythms of behavior and physiology are driven by the biological clock that operates endogenously but can also be entrained to the light-dark cycle of the environment. In mammals, the master circadian pacemaker is located in the suprachiasmatic nucleus (SCN, which is composed of individual cellular oscillators that are driven by a set of core clock genes interacting in transcriptional/translational feedback loops. Light signals can trigger molecular events in the SCN that ultimately impact on the phase of expression of core clock genes to reset the master pacemaker. While transcriptional regulation has received much attention in the field of circadian biology in the past, other mechanisms including targeted protein degradation likely contribute to the clock timing and entrainment process. In the present study, proteome-wide screens of the murine SCN led to the identification of ubiquitin protein ligase E3 component N-recognin 4 (UBR4, a novel E3 ubiquitin ligase component of the N-end rule pathway, as a time-of-day-dependent and light-inducible protein. The spatial and temporal expression pattern of UBR4 in the SCN was subsequently characterized by immunofluorescence microscopy. UBR4 is expressed across the entire rostrocaudal extent of the SCN in a time-of-day-dependent fashion. UBR4 is localized exclusively to arginine vasopressin (AVP-expressing neurons of the SCN shell. Upon photic stimulation in the early subjective night, the number of UBR4-expressing cells within the SCN increases. This study is the first to identify a novel E3 ubiquitin ligase component, UBR4, in the murine SCN and to implicate the N-end rule degradation pathway as a potential player in regulating core clock mechanisms and photic entrainment.

  5. The spliceosome-associated protein Mfap1 binds to VCP in Drosophila.

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    Sandra Rode

    Full Text Available Posttranscriptional regulation of gene expression contributes to many developmental transitions. Previously, we found that the AAA chaperone Valosin-Containing Protein (VCP regulates ecdysone-dependent dendrite pruning of Drosophila class IV dendritic arborization (c4da neurons via an effect on RNA metabolism. In a search for RNA binding proteins associated with VCP, we identified the spliceosome-associated protein Mfap1, a component of the tri-snRNP complex. Mfap1 is a nucleolar protein in neurons and its levels are regulated by VCP. Mfap1 binds to VCP and TDP-43, a disease-associated RNA-binding protein. via distinct regions in its N- and C-terminal halfs. Similar to vcp mutations, Mfap1 overexpression causes c4da neuron dendrite pruning defects and mislocalization of TDP-43 in these cells, but genetic analyses show that Mfap1 is not a crucial VCP target during dendrite pruning. Finally, rescue experiments with a lethal mfap1 mutant show that the VCP binding region is not essential for Mfap1 function, but may act to increase its stability or activity.

  6. Fragile X mental retardation protein regulates trans-synaptic signaling in Drosophila

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    Samuel H. Friedman

    2013-11-01

    Fragile X syndrome (FXS, the most common inherited determinant of intellectual disability and autism spectrum disorders, is caused by loss of the fragile X mental retardation 1 (FMR1 gene product (FMRP, an mRNA-binding translational repressor. A number of conserved FMRP targets have been identified in the well-characterized Drosophila FXS disease model, but FMRP is highly pleiotropic in function and the full spectrum of FMRP targets has yet to be revealed. In this study, screens for upregulated neural proteins in Drosophila fmr1 (dfmr1 null mutants reveal strong elevation of two synaptic heparan sulfate proteoglycans (HSPGs: GPI-anchored glypican Dally-like protein (Dlp and transmembrane Syndecan (Sdc. Our recent work has shown that Dlp and Sdc act as co-receptors regulating extracellular ligands upstream of intracellular signal transduction in multiple trans-synaptic pathways that drive synaptogenesis. Consistently, dfmr1 null synapses exhibit altered WNT signaling, with changes in both Wingless (Wg ligand abundance and downstream Frizzled-2 (Fz2 receptor C-terminal nuclear import. Similarly, a parallel anterograde signaling ligand, Jelly belly (Jeb, and downstream ERK phosphorylation (dpERK are depressed at dfmr1 null synapses. In contrast, the retrograde BMP ligand Glass bottom boat (Gbb and downstream signaling via phosphorylation of the transcription factor MAD (pMAD seem not to be affected. To determine whether HSPG upregulation is causative for synaptogenic defects, HSPGs were genetically reduced to control levels in the dfmr1 null background. HSPG correction restored both (1 Wg and Jeb trans-synaptic signaling, and (2 synaptic architecture and transmission strength back to wild-type levels. Taken together, these data suggest that FMRP negatively regulates HSPG co-receptors controlling trans-synaptic signaling during synaptogenesis, and that loss of this regulation causes synaptic structure and function defects characterizing the FXS disease state.

  7. Clueless, a protein required for mitochondrial function, interacts with the PINK1-Parkin complex in Drosophila

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    Aditya Sen

    2015-06-01

    Full Text Available Loss of mitochondrial function often leads to neurodegeneration and is thought to be one of the underlying causes of neurodegenerative diseases such as Parkinson's disease (PD. However, the precise events linking mitochondrial dysfunction to neuronal death remain elusive. PTEN-induced putative kinase 1 (PINK1 and Parkin (Park, either of which, when mutated, are responsible for early-onset PD, mark individual mitochondria for destruction at the mitochondrial outer membrane. The specific molecular pathways that regulate signaling between the nucleus and mitochondria to sense mitochondrial dysfunction under normal physiological conditions are not well understood. Here, we show that Drosophila Clueless (Clu, a highly conserved protein required for normal mitochondrial function, can associate with Translocase of the outer membrane (TOM 20, Porin and PINK1, and is thus located at the mitochondrial outer membrane. Previously, we found that clu genetically interacts with park in Drosophila female germ cells. Here, we show that clu also genetically interacts with PINK1, and our epistasis analysis places clu downstream of PINK1 and upstream of park. In addition, Clu forms a complex with PINK1 and Park, further supporting that Clu links mitochondrial function with the PINK1-Park pathway. Lack of Clu causes PINK1 and Park to interact with each other, and clu mutants have decreased mitochondrial protein levels, suggesting that Clu can act as a negative regulator of the PINK1-Park pathway. Taken together, these results suggest that Clu directly modulates mitochondrial function, and that Clu's function contributes to the PINK1-Park pathway of mitochondrial quality control.

  8. Neighboring genes for DNA-binding proteins rescue male sterility in Drosophila hybrids.

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    Liénard, Marjorie A; Araripe, Luciana O; Hartl, Daniel L

    2016-07-19

    Crosses between closely related animal species often result in male hybrids that are sterile, and the molecular and functional basis of genetic factors for hybrid male sterility is of great interest. Here, we report a molecular and functional analysis of HMS1, a region of 9.2 kb in chromosome 3 of Drosophila mauritiana, which results in virtually complete hybrid male sterility when homozygous in the genetic background of sibling species Drosophila simulans. The HMS1 region contains two strong candidate genes for the genetic incompatibility, agt and Taf1 Both encode unrelated DNA-binding proteins, agt for an alkyl-cysteine-S-alkyltransferase and Taf1 for a subunit of transcription factor TFIID that serves as a multifunctional transcriptional regulator. The contribution of each gene to hybrid male sterility was assessed by means of germ-line transformation, with constructs containing complete agt and Taf1 genomic sequences as well as various chimeric constructs. Both agt and Taf1 contribute about equally to HMS1 hybrid male sterility. Transgenes containing either locus rescue sterility in about one-half of the males, and among fertile males the number of offspring is in the normal range. This finding suggests compensatory proliferation of the rescued, nondysfunctional germ cells. Results with chimeric transgenes imply that the hybrid incompatibilities result from interactions among nucleotide differences residing along both agt and Taf1 Our results challenge a number of preliminary generalizations about the molecular and functional basis of hybrid male sterility, and strongly reinforce the role of DNA-binding proteins as a class of genes contributing to the maintenance of postzygotic reproductive isolation.

  9. Cryptochromes define a novel circadian clock mechanism in monarch butterflies that may underlie sun compass navigation.

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    Haisun Zhu

    2008-01-01

    Full Text Available The circadian clock plays a vital role in monarch butterfly (Danaus plexippus migration by providing the timing component of time-compensated sun compass orientation, a process that is important for successful navigation. We therefore evaluated the monarch clockwork by focusing on the functions of a Drosophila-like cryptochrome (cry, designated cry1, and a vertebrate-like cry, designated cry2, that are both expressed in the butterfly and by placing these genes in the context of other relevant clock genes in vivo. We found that similar temporal patterns of clock gene expression and protein levels occur in the heads, as occur in DpN1 cells, of a monarch cell line that contains a light-driven clock. CRY1 mediates TIMELESS degradation by light in DpN1 cells, and a light-induced TIMELESS decrease occurs in putative clock cells in the pars lateralis (PL in the brain. Moreover, monarch cry1 transgenes partially rescue both biochemical and behavioral light-input defects in cry(b mutant Drosophila. CRY2 is the major transcriptional repressor of CLOCK:CYCLE-mediated transcription in DpN1 cells, and endogenous CRY2 potently inhibits transcription without involvement of PERIOD. CRY2 is co-localized with clock proteins in the PL, and there it translocates to the nucleus at the appropriate time for transcriptional repression. We also discovered CRY2-positive neural projections that oscillate in the central complex. The results define a novel, CRY-centric clock mechanism in the monarch in which CRY1 likely functions as a blue-light photoreceptor for entrainment, whereas CRY2 functions within the clockwork as the transcriptional repressor of a negative transcriptional feedback loop. Our data further suggest that CRY2 may have a dual role in the monarch butterfly's brain-as a core clock element and as an output that regulates circadian activity in the central complex, the likely site of the sun compass.

  10. Dietary live yeast alters metabolic profiles, protein biosynthesis and thermal stress tolerance of Drosophila melanogaster.

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    Colinet, Hervé; Renault, David

    2014-04-01

    The impact of nutritional factors on insect's life-history traits such as reproduction and lifespan has been excessively examined; however, nutritional determinant of insect's thermal tolerance has not received a lot of attention. Dietary live yeast represents a prominent source of proteins and amino acids for laboratory-reared drosophilids. In this study, Drosophila melanogaster adults were fed on diets supplemented or not with live yeast. We hypothesized that manipulating nutritional conditions through live yeast supplementation would translate into altered physiology and stress tolerance. We verified how live yeast supplementation affected body mass characteristics, total lipids and proteins, metabolic profiles and cold tolerance (acute and chronic stress). Females fed with live yeast had increased body mass and contained more lipids and proteins. Using GC/MS profiling, we found distinct metabolic fingerprints according to nutritional conditions. Metabolite pathway enrichment analysis corroborated that live yeast supplementation was associated with amino acid and protein biosyntheses. The cold assays revealed that the presence of dietary live yeast greatly promoted cold tolerance. Hence, this study conclusively demonstrates a significant interaction between nutritional conditions and thermal tolerance. Copyright © 2014 Elsevier Inc. All rights reserved.

  11. Drosophila Nora virus capsid proteins differ from those of other picorna-like viruses.

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    Ekström, Jens-Ola; Habayeb, Mazen S; Srivastava, Vaibhav; Kieselbach, Thomas; Wingsle, Gunnar; Hultmark, Dan

    2011-09-01

    The recently discovered Nora virus from Drosophila melanogaster is a single-stranded RNA virus. Its published genomic sequence encodes a typical picorna-like cassette of replicative enzymes, but no capsid proteins similar to those in other picorna-like viruses. We have now done additional sequencing at the termini of the viral genome, extending it by 455 nucleotides at the 5' end, but no more coding sequence was found. The completeness of the final 12,333-nucleotide sequence was verified by the production of infectious virus from the cloned genome. To identify the capsid proteins, we purified Nora virus particles and analyzed their proteins by mass spectrometry. Our results show that the capsid is built from three major proteins, VP4A, B and C, encoded in the fourth open reading frame of the viral genome. The viral particles also contain traces of a protein from the third open reading frame, VP3. VP4A and B are not closely related to other picorna-like virus capsid proteins in sequence, but may form similar jelly roll folds. VP4C differs from the others and is predicted to have an essentially α-helical conformation. In a related virus, identified from EST database sequences from Nasonia parasitoid wasps, VP4C is encoded in a separate open reading frame, separated from VP4A and B by a frame-shift. This opens a possibility that VP4C is produced in non-equimolar quantities. Altogether, our results suggest that the Nora virus capsid has a different protein organization compared to the order Picornavirales. Copyright © 2011 Elsevier B.V. All rights reserved.

  12. Inverse regulation of two classic Hippo pathway target genes in Drosophila by the dimerization hub protein Ctp.

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    Barron, Daniel A; Moberg, Kenneth

    2016-03-14

    The LC8 family of small ~8 kD proteins are highly conserved and interact with multiple protein partners in eukaryotic cells. LC8-binding modulates target protein activity, often through induced dimerization via LC8:LC8 homodimers. Although many LC8-interactors have roles in signaling cascades, LC8's role in developing epithelia is poorly understood. Using the Drosophila wing as a developmental model, we find that the LC8 family member Cut up (Ctp) is primarily required to promote epithelial growth, which correlates with effects on the pro-growth factor dMyc and two genes, diap1 and bantam, that are classic targets of the Hippo pathway coactivator Yorkie. Genetic tests confirm that Ctp supports Yorkie-driven tissue overgrowth and indicate that Ctp acts through Yorkie to control bantam (ban) and diap1 transcription. Quite unexpectedly however, Ctp loss has inverse effects on ban and diap1: it elevates ban expression but reduces diap1 expression. In both cases these transcriptional changes map to small segments of these promoters that recruit Yorkie. Although LC8 complexes with Yap1, a Yorkie homolog, in human cells, an orthologous interaction was not detected in Drosophila cells. Collectively these findings reveal that that Drosophila Ctp is a required regulator of Yorkie-target genes in vivo and suggest that Ctp may interact with a Hippo pathway protein(s) to exert inverse transcriptional effects on Yorkie-target genes.

  13. Inverse European Latitudinal Cline at the timeless Locus of Drosophila melanogaster Reveals Selection on a Clock Gene: Population Genetics of ls-tim.

    Science.gov (United States)

    Zonato, Valeria; Vanin, Stefano; Costa, Rodolfo; Tauber, Eran; Kyriacou, Charalambos P

    2018-02-01

    The spread of adaptive genetic variants in populations is a cornerstone of evolutionary theory but with relatively few biologically well-understood examples. Previous work on the ls-tim variant of timeless, which encodes the light-sensitive circadian regulator in Drosophila melanogaster, suggests that it may have originated in southeastern Italy. Flies characterized by the new allele show photoperiod-related phenotypes likely to be adaptive in seasonal environments. ls-tim may be spreading from its point of origin in Italy by directional selection, but there are alternative explanations for its observed clinal geographical distribution, including balancing selection and demography. From population analyses of ls-tim frequencies collected on the eastern side of the Iberian Peninsula, we show that ls-tim frequencies are inverted compared with those in Italy. This pattern is consistent with a scenario of directional selection rather than latitude-associated balancing selection. Neutrality tests further reveal the signature of directional selection at the ls-tim site, which is reduced a few kb pairs either side of ls-tim. A reanalysis of allele frequencies from a large number of microsatellite loci do not demonstrate any frequent ls-tim-like spatial patterns, so a general demographic effect or population expansion from southeastern Italy cannot readily explain current ls-tim frequencies. Finally, a revised estimate of the age of ls-tim allele using linkage disequilibrium and coalescent-based approaches reveals that it may be only 300 to 3000 years old, perhaps explaining why it has not yet gone to fixation. ls-tim thus provides a rare temporal snapshot of a new allele that has come under selection before it reaches equilibrium.

  14. Altered protein glycosylation predicts Alzheimer's disease and modulates its pathology in disease model Drosophila.

    Science.gov (United States)

    Frenkel-Pinter, Moran; Stempler, Shiri; Tal-Mazaki, Sharon; Losev, Yelena; Singh-Anand, Avnika; Escobar-Álvarez, Daniela; Lezmy, Jonathan; Gazit, Ehud; Ruppin, Eytan; Segal, Daniel

    2017-08-01

    The pathological hallmarks of Alzheimer's disease (AD) are pathogenic oligomers and fibrils of misfolded amyloidogenic proteins (e.g., β-amyloid and hyper-phosphorylated tau in AD), which cause progressive loss of neurons in the brain and nervous system. Although deviations from normal protein glycosylation have been documented in AD, their role in disease pathology has been barely explored. Here our analysis of available expression data sets indicates that many glycosylation-related genes are differentially expressed in brains of AD patients compared with healthy controls. The robust differences found enabled us to predict the occurrence of AD with remarkable accuracy in a test cohort and identify a set of key genes whose expression determines this classification. We then studied in vivo the effect of reducing expression of homologs of 6 of these genes in transgenic Drosophila overexpressing human tau, a well-established invertebrate AD model. These experiments have led to the identification of glycosylation genes that may augment or ameliorate tauopathy phenotypes. Our results indicate that OstDelta, l(2)not and beta4GalT7 are tauopathy suppressors, whereas pgnat5 and CG33303 are enhancers, of tauopathy. These results suggest that specific alterations in protein glycosylation may play a causal role in AD etiology. Copyright © 2017 Elsevier Inc. All rights reserved.

  15. A role for the membrane protein M6 in the Drosophila visual system.

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    Zappia, María Paula; Bernabo, Guillermo; Billi, Silvia C; Frasch, Alberto C; Ceriani, María Fernanda; Brocco, Marcela Adriana

    2012-07-04

    Members of the proteolipid protein family, including the four-transmembrane glycoprotein M6a, are involved in neuronal plasticity in mammals. Results from our group previously demonstrated that M6, the only proteolipid protein expressed in Drosophila, localizes to the cell membrane in follicle cells. M6 loss triggers female sterility, which suggests a role for M6 in follicular cell remodeling. These results were the basis of the present study, which focused on the function and requirements of M6 in the fly nervous system. The present study identified two novel, tissue-regulated M6 isoforms with variable N- and C- termini, and showed that M6 is the functional fly ortholog of Gpm6a. In the adult brain, the protein was localized to several neuropils, such as the optic lobe, the central complex, and the mushroom bodies. Interestingly, although reduced M6 levels triggered a mild rough-eye phenotype, hypomorphic M6 mutants exhibited a defective response to light. Based on its ability to induce filopodium formation we propose that M6 is key in cell remodeling processes underlying visual system function. These results bring further insight into the role of M6/M6a in biological processes involving neuronal plasticity and behavior in flies and mammals.

  16. cGMP-dependent protein kinase I, the circadian clock, sleep and learning

    OpenAIRE

    Feil, Robert; Hölter, Sabine M; Weindl, Karin; Wurst, Wolfgang; Langmesser, Sonja; Gerling, Andrea; Feil, Susanne; Albrecht, Urs

    2009-01-01

    The second messenger cGMP controls cardiovascular and gastrointestinal homeostasis in mammals. However, its physiological relevance in the nervous system is poorly understood.1 Now, we have reported that the cGMP-dependent protein kinase type I (PRKG1) is implicated in the regulation of the timing and quality of sleep and wakefulness.2 Prkg1 mutant mice showed altered distribution of sleep and wakefulness as well as reduction in rapid-eye-movement sleep (REMS) duration and in non-REMS consoli...

  17. Cooperation of the BTB-Zinc finger protein, Abrupt, with cytoskeletal regulators in Drosophila epithelial tumorigenesis

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    Nezaket Turkel

    2015-08-01

    Full Text Available The deregulation of cell polarity or cytoskeletal regulators is a common occurrence in human epithelial cancers. Moreover, there is accumulating evidence in human epithelial cancer that BTB-ZF genes, such as Bcl6 and ZBTB7A, are oncogenic. From our previous studies in the vinegar fly, Drosophila melanogaster, we have identified a cooperative interaction between a mutation in the apico-basal cell polarity regulator Scribble (Scrib and overexpression of the BTB-ZF protein Abrupt (Ab. Herein, we show that co-expression of ab with actin cytoskeletal regulators, RhoGEF2 or Src64B, in the developing eye-antennal epithelial tissue results in the formation of overgrown amorphous tumours, whereas ab and DRac1 co-expression leads to non-cell autonomous overgrowth. Together with ab, these genes affect the expression of differentiation genes, resulting in tumours locked in a progenitor cell fate. Finally, we show that the expression of two mammalian genes related to ab, Bcl6 and ZBTB7A, which are oncogenes in mammalian epithelial cancers, significantly correlate with the upregulation of cytoskeletal genes or downregulation of apico-basal cell polarity neoplastic tumour suppressor genes in colorectal, lung and other human epithelial cancers. Altogether, this analysis has revealed that upregulation of cytoskeletal regulators cooperate with Abrupt in Drosophila epithelial tumorigenesis, and that high expression of human BTB-ZF genes, Bcl6 and ZBTB7A, shows significant correlations with cytoskeletal and cell polarity gene expression in specific epithelial tumour types. This highlights the need for further investigation of the cooperation between these genes in mammalian systems.

  18. Mushroom body miscellanea: transgenic Drosophila strains expressing anatomical and physiological sensor proteins in Kenyon cells

    Science.gov (United States)

    Pech, Ulrike; Dipt, Shubham; Barth, Jonas; Singh, Priyanka; Jauch, Mandy; Thum, Andreas S.; Fiala, André; Riemensperger, Thomas

    2013-01-01

    The fruit fly Drosophila melanogaster represents a key model organism for analyzing how neuronal circuits regulate behavior. The mushroom body in the central brain is a particularly prominent brain region that has been intensely studied in several insect species and been implicated in a variety of behaviors, e.g., associative learning, locomotor activity, and sleep. Drosophila melanogaster offers the advantage that transgenes can be easily expressed in neuronal subpopulations, e.g., in intrinsic mushroom body neurons (Kenyon cells). A number of transgenes has been described and engineered to visualize the anatomy of neurons, to monitor physiological parameters of neuronal activity, and to manipulate neuronal function artificially. To target the expression of these transgenes selectively to specific neurons several sophisticated bi- or even multipartite transcription systems have been invented. However, the number of transgenes that can be combined in the genome of an individual fly is limited in practice. To facilitate the analysis of the mushroom body we provide a compilation of transgenic fruit flies that express transgenes under direct control of the Kenyon-cell specific promoter, mb247. The transgenes expressed are fluorescence reporters to analyze neuroanatomical aspects of the mushroom body, proteins to restrict ectopic gene expression to mushroom bodies, or fluorescent sensors to monitor physiological parameters of neuronal activity of Kenyon cells. Some of the transgenic animals compiled here have been published already, whereas others are novel and characterized here for the first time. Overall, the collection of transgenic flies expressing sensor and reporter genes in Kenyon cells facilitates combinations with binary transcription systems and might, ultimately, advance the physiological analysis of mushroom body function. PMID:24065891

  19. A transgenic Drosophila model demonstrates that the Helicobacter pylori CagA protein functions as a eukaryotic Gab adaptor.

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    Crystal M Botham

    2008-05-01

    Full Text Available Infection with the human gastric pathogen Helicobacter pylori is associated with a spectrum of diseases including gastritis, peptic ulcers, gastric adenocarcinoma, and gastric mucosa-associated lymphoid tissue lymphoma. The cytotoxin-associated gene A (CagA protein of H. pylori, which is translocated into host cells via a type IV secretion system, is a major risk factor for disease development. Experiments in gastric tissue culture cells have shown that once translocated, CagA activates the phosphatase SHP-2, which is a component of receptor tyrosine kinase (RTK pathways whose over-activation is associated with cancer formation. Based on CagA's ability to activate SHP-2, it has been proposed that CagA functions as a prokaryotic mimic of the eukaryotic Grb2-associated binder (Gab adaptor protein, which normally activates SHP-2. We have developed a transgenic Drosophila model to test this hypothesis by investigating whether CagA can function in a well-characterized Gab-dependent process: the specification of photoreceptors cells in the Drosophila eye. We demonstrate that CagA expression is sufficient to rescue photoreceptor development in the absence of the Drosophila Gab homologue, Daughter of Sevenless (DOS. Furthermore, CagA's ability to promote photoreceptor development requires the SHP-2 phosphatase Corkscrew (CSW. These results provide the first demonstration that CagA functions as a Gab protein within the tissue of an organism and provide insight into CagA's oncogenic potential. Since many translocated bacterial proteins target highly conserved eukaryotic cellular processes, such as the RTK signaling pathway, the transgenic Drosophila model should be of general use for testing the in vivo function of bacterial effector proteins and for identifying the host genes through which they function.

  20. Drosophila DJ-1 decreases neural sensitivity to stress by negatively regulating Daxx-like protein through dFOXO.

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    Soojin Hwang

    2013-04-01

    Full Text Available DJ-1, a Parkinson's disease (PD-associated gene, has been shown to protect against oxidative stress in Drosophila. However, the molecular mechanism underlying oxidative stress-induced phenotypes, including apoptosis, locomotive defects, and lethality, in DJ-1-deficient flies is not fully understood. Here we showed that Daxx-like protein (DLP, a Drosophila homologue of the mammalian Death domain-associated protein (Daxx, was upregulated under oxidative stress conditions in the loss-of-function mutants of Drosophila DJ-1β, a Drosophila homologue of DJ-1. DLP overexpression induced apoptosis via the c-Jun N-terminal kinase (JNK/Drosophila forkhead box subgroup O (dFOXO pathway, whereas loss of DLP increased resistance to oxidative stress and UV irradiation. Moreover, the oxidative stress-induced phenotypes of DJ-1β mutants were dramatically rescued by DLP deficiency, suggesting that enhanced expression of DLP contributes to the DJ-1β mutant phenotypes. Interestingly, we found that dFOXO was required for the increase in DLP expression in DJ-1β mutants and that dFOXO activity was increased in the heads of DJ-1β mutants. In addition, subcellular localization of DLP appeared to be influenced by DJ-1 expression so that cytosolic DLP was increased in DJ-1β mutants. Similarly, in mammalian cells, Daxx translocation from the nucleus to the cytosol was suppressed by overexpressed DJ-1β under oxidative stress conditions; and, furthermore, targeted expression of DJ-1β to mitochondria efficiently inhibited the Daxx translocation. Taken together, our findings demonstrate that DJ-1β protects flies against oxidative stress- and UV-induced apoptosis by regulating the subcellular localization and gene expression of DLP, thus implying that Daxx-induced apoptosis is involved in the pathogenesis of DJ-1-associated PD.

  1. Analysis of temporal transcription expression profiles reveal links between protein function and developmental stages of Drosophila melanogaster.

    Science.gov (United States)

    Wan, Cen; Lees, Jonathan G; Minneci, Federico; Orengo, Christine A; Jones, David T

    2017-10-01

    Accurate gene or protein function prediction is a key challenge in the post-genome era. Most current methods perform well on molecular function prediction, but struggle to provide useful annotations relating to biological process functions due to the limited power of sequence-based features in that functional domain. In this work, we systematically evaluate the predictive power of temporal transcription expression profiles for protein function prediction in Drosophila melanogaster. Our results show significantly better performance on predicting protein function when transcription expression profile-based features are integrated with sequence-derived features, compared with the sequence-derived features alone. We also observe that the combination of expression-based and sequence-based features leads to further improvement of accuracy on predicting all three domains of gene function. Based on the optimal feature combinations, we then propose a novel multi-classifier-based function prediction method for Drosophila melanogaster proteins, FFPred-fly+. Interpreting our machine learning models also allows us to identify some of the underlying links between biological processes and developmental stages of Drosophila melanogaster.

  2. Analysis of temporal transcription expression profiles reveal links between protein function and developmental stages of Drosophila melanogaster.

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    Cen Wan

    2017-10-01

    Full Text Available Accurate gene or protein function prediction is a key challenge in the post-genome era. Most current methods perform well on molecular function prediction, but struggle to provide useful annotations relating to biological process functions due to the limited power of sequence-based features in that functional domain. In this work, we systematically evaluate the predictive power of temporal transcription expression profiles for protein function prediction in Drosophila melanogaster. Our results show significantly better performance on predicting protein function when transcription expression profile-based features are integrated with sequence-derived features, compared with the sequence-derived features alone. We also observe that the combination of expression-based and sequence-based features leads to further improvement of accuracy on predicting all three domains of gene function. Based on the optimal feature combinations, we then propose a novel multi-classifier-based function prediction method for Drosophila melanogaster proteins, FFPred-fly+. Interpreting our machine learning models also allows us to identify some of the underlying links between biological processes and developmental stages of Drosophila melanogaster.

  3. The Drosophila BTB domain protein Jim Lovell has roles in multiple larval and adult behaviors.

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    Sonia M Bjorum

    Full Text Available Innate behaviors have their origins in the specification of neural fates during development. Within Drosophila, BTB (Bric-a-brac,Tramtrack, Broad domain proteins such as Fruitless are known to play key roles in the neural differentiation underlying such responses. We previously identified a gene, which we have termed jim lovell (lov, encoding a BTB protein with a role in gravity responses. To understand more fully the behavioral roles of this gene we have investigated its function through several approaches. Transcript and protein expression patterns have been examined and behavioral phenotypes of new lov mutations have been characterized. Lov is a nuclear protein, suggesting a role as a transcriptional regulator, as for other BTB proteins. In late embryogenesis, Lov is expressed in many CNS and PNS neurons. An examination of the PNS expression indicates that lov functions in the late specification of several classes of sensory neurons. In particular, only two of the five abdominal lateral chordotonal neurons express Lov, predicting functional variation within this highly similar group. Surprisingly, Lov is also expressed very early in embryogenesis in ways that suggests roles in morphogenetic movements, amnioserosa function and head neurogenesis. The phenotypes of two new lov mutations that delete adjacent non-coding DNA regions are strikingly different suggesting removal of different regulatory elements. In lov(47 , Lov expression is lost in many embryonic neurons including the two lateral chordotonal neurons. lov(47 mutant larvae show feeding and locomotor defects including spontaneous backward movement. Adult lov(47 males perform aberrant courtship behavior distinguished by courtship displays that are not directed at the female. lov(47 adults also show more defective negative gravitaxis than the previously isolated lov(91Y mutant. In contrast, lov(66 produces largely normal behavior but severe female sterility associated with ectopic lov

  4. Drosophila melanogaster cellular repressor of E1A-stimulated genes is a lysosomal protein essential for fly development

    OpenAIRE

    Kowalewski-Nimmerfall, Elisabeth; Sch?hs, Philipp; Maresch, Daniel; Rendic, Dubravko; Kr?mer, Helmut; Mach, Lukas

    2014-01-01

    Mammalian cellular repressor of E1A-stimulated genes is a lysosomal glycoprotein implicated in cellular growth and differentiation. The genome of the fruit fly Drosophila melanogaster encodes a putative orthologue (dCREG), suggesting evolutionarily conserved physiological functions of this protein. In D. melanogaster S2 cells, dCREG was found to localize in lysosomes. Further studies revealed that intracellular dCREG is subject of proteolytic maturation. Processing and turnover could be subst...

  5. Biochemical and genetic analysis of the Drk SH2/SH3 adaptor protein of Drosophila.

    Science.gov (United States)

    Raabe, T; Olivier, J P; Dickson, B; Liu, X; Gish, G D; Pawson, T; Hafen, E

    1995-06-01

    The Drk SH3-SH2-SH3 adaptor protein has been genetically identified in a screen for rate-limiting components acting downstream of the Sevenless (Sev) receptor tyrosine kinase in the developing eye of Drosophila. It provides a link between the activated Sev receptor and Sos, a guanine nucleotide release factor that activates Ras1. We have used a combined biochemical and genetic approach to study the interactions between Sev, Drk and Sos. We show that Tyr2546 in the cytoplasmic tail of Sev is required for Drk binding, probably because it provides a recognition site for the Drk SH2 domain. Interestingly, a mutation at this site does not completely block Sev function in vivo. This may suggest that Sev can signal in a Drk-independent, parallel pathway or that Drk can also bind to an intermediate docking protein. Analysis of the Drk-Sos interaction has identified a high affinity binding site for Drk SH3 domains in the Sos tail. We show that the N-terminal Drk SH3 domain is primarily responsible for binding to the tail of Sos in vitro, and for signalling to Ras in vivo.

  6. The Ly6 protein coiled is required for septate junction and blood brain barrier organisation in Drosophila.

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    Assia Hijazi

    Full Text Available BACKGROUND: Genetic analysis of the Drosophila septate junctions has greatly contributed to our understanding of the mechanisms controlling the assembly of these adhesion structures, which bear strong similarities with the vertebrate tight junctions and the paranodal septate junctions. These adhesion complexes share conserved molecular components and have a common function: the formation of paracellular barriers restraining the diffusion of solutes through epithelial and glial envelopes. METHODOLOGY/PRINCIPAL FINDINGS: In this work we characterise the function of the Drosophila cold gene, that codes for a protein belonging to the Ly6 superfamily of extracellular ligands. Analysis of cold mutants shows that this gene is specifically required for the organisation of the septate junctions in epithelial tissues and in the nervous system, where its contribution is essential for the maintenance of the blood-brain barrier. We show that cold acts in a cell autonomous way, and we present evidence indicating that this protein could act as a septate junction component. CONCLUSION/SIGNIFICANCE: We discuss the specific roles of cold and three other Drosophila members of the Ly6 superfamily that have been shown to participate in a non-redundant way in the process of septate junction assembly. We propose that vertebrate Ly6 proteins could fulfill analogous roles in tight junctions and/or paranodal septate junctions.

  7. The Ly6 protein coiled is required for septate junction and blood brain barrier organisation in Drosophila.

    Science.gov (United States)

    Hijazi, Assia; Haenlin, Marc; Waltzer, Lucas; Roch, Fernando

    2011-03-15

    Genetic analysis of the Drosophila septate junctions has greatly contributed to our understanding of the mechanisms controlling the assembly of these adhesion structures, which bear strong similarities with the vertebrate tight junctions and the paranodal septate junctions. These adhesion complexes share conserved molecular components and have a common function: the formation of paracellular barriers restraining the diffusion of solutes through epithelial and glial envelopes. In this work we characterise the function of the Drosophila cold gene, that codes for a protein belonging to the Ly6 superfamily of extracellular ligands. Analysis of cold mutants shows that this gene is specifically required for the organisation of the septate junctions in epithelial tissues and in the nervous system, where its contribution is essential for the maintenance of the blood-brain barrier. We show that cold acts in a cell autonomous way, and we present evidence indicating that this protein could act as a septate junction component. We discuss the specific roles of cold and three other Drosophila members of the Ly6 superfamily that have been shown to participate in a non-redundant way in the process of septate junction assembly. We propose that vertebrate Ly6 proteins could fulfill analogous roles in tight junctions and/or paranodal septate junctions.

  8. The ALS-associated proteins FUS and TDP-43 function together to affect Drosophila locomotion and life span

    Science.gov (United States)

    Wang, Ji-Wu; Brent, Jonathan R.; Tomlinson, Andrew; Shneider, Neil A.; McCabe, Brian D.

    2011-01-01

    The fatal adult motor neuron disease amyotrophic lateral sclerosis (ALS) shares some clinical and pathological overlap with frontotemporal dementia (FTD), an early-onset neurodegenerative disorder. The RNA/DNA-binding proteins fused in sarcoma (FUS; also known as TLS) and TAR DNA binding protein-43 (TDP-43) have recently been shown to be genetically and pathologically associated with familial forms of ALS and FTD. It is currently unknown whether perturbation of these proteins results in disease through mechanisms that are independent of normal protein function or via the pathophysiological disruption of molecular processes in which they are both critical. Here, we report that Drosophila mutants in which the homolog of FUS is disrupted exhibit decreased adult viability, diminished locomotor speed, and reduced life span compared with controls. These phenotypes were fully rescued by wild-type human FUS, but not ALS-associated mutant FUS proteins. A mutant of the Drosophila homolog of TDP-43 had similar, but more severe, deficits. Through cross-rescue analysis, we demonstrated that FUS acted together with and downstream of TDP-43 in a common genetic pathway in neurons. Furthermore, we found that these proteins associated with each other in an RNA-dependent complex. Our results establish that FUS and TDP-43 function together in vivo and suggest that molecular pathways requiring the combined activities of both of these proteins may be disrupted in ALS and FTD. PMID:21881207

  9. Effects of different per translational kinetics on the dynamics of a core circadian clock model.

    Science.gov (United States)

    Nieto, Paula S; Revelli, Jorge A; Garbarino-Pico, Eduardo; Condat, Carlos A; Guido, Mario E; Tamarit, Francisco A

    2015-01-01

    Living beings display self-sustained daily rhythms in multiple biological processes, which persist in the absence of external cues since they are generated by endogenous circadian clocks. The period (per) gene is a central player within the core molecular mechanism for keeping circadian time in most animals. Recently, the modulation PER translation has been reported, both in mammals and flies, suggesting that translational regulation of clock components is important for the proper clock gene expression and molecular clock performance. Because translational regulation ultimately implies changes in the kinetics of translation and, therefore, in the circadian clock dynamics, we sought to study how and to what extent the molecular clock dynamics is affected by the kinetics of PER translation. With this objective, we used a minimal mathematical model of the molecular circadian clock to qualitatively characterize the dynamical changes derived from kinetically different PER translational mechanisms. We found that the emergence of self-sustained oscillations with characteristic period, amplitude, and phase lag (time delays) between per mRNA and protein expression depends on the kinetic parameters related to PER translation. Interestingly, under certain conditions, a PER translation mechanism with saturable kinetics introduces longer time delays than a mechanism ruled by a first-order kinetics. In addition, the kinetic laws of PER translation significantly changed the sensitivity of our model to parameters related to the synthesis and degradation of per mRNA and PER degradation. Lastly, we found a set of parameters, with realistic values, for which our model reproduces some experimental results reported recently for Drosophila melanogaster and we present some predictions derived from our analysis.

  10. Rapid DNA Synthesis During Early Drosophila Embryogenesis Is Sensitive to Maternal Humpty Dumpty Protein Function.

    Science.gov (United States)

    Lesly, Shera; Bandura, Jennifer L; Calvi, Brian R

    2017-11-01

    Problems with DNA replication cause cancer and developmental malformations. It is not fully understood how DNA replication is coordinated with development and perturbed in disease. We had previously identified the Drosophila gene humpty dumpty ( hd ), and showed that null alleles cause incomplete DNA replication, tissue undergrowth, and lethality. Animals homozygous for the missense allele, hd 272-9 , were viable, but adult females had impaired amplification of eggshell protein genes in the ovary, resulting in the maternal effects of thin eggshells and embryonic lethality. Here, we show that expression of an hd transgene in somatic cells of the ovary rescues amplification and eggshell synthesis but not embryo viability. The germline of these mothers remain mutant for the hd 272-9 allele, resulting in reduced maternal Hd protein and embryonic arrest during mitosis of the first few S/M nuclear cleavage cycles with chromosome instability and chromosome bridges. Epistasis analysis of hd with the rereplication mutation plutonium indicates that the chromosome bridges of hd embryos are the result of a failed attempt to segregate incompletely replicated sister chromatids. This study reveals that maternally encoded Humpty dumpty protein is essential for DNA replication and genome integrity during the little-understood embryonic S/M cycles. Moreover, the two hd 272-9 maternal-effect phenotypes suggest that ovarian gene amplification and embryonic cleavage are two time periods in development that are particularly sensitive to mild deficits in DNA replication function. This last observation has broader relevance for interpreting why mild mutations in the human ortholog of humpty dumpty and other DNA replication genes cause tissue-specific malformations of microcephalic dwarfisms. Copyright © 2017 by the Genetics Society of America.

  11. The Drosophila DOCK family protein Sponge is required for development of the air sac primordium

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    Morishita, Kazushge; Anh Suong, Dang Ngoc; Yoshida, Hideki; Yamaguchi, Masamitsu, E-mail: myamaguc@kit.ac.jp

    2017-05-15

    Dedicator of cytokinesis (DOCK) family genes are known as DOCK1-DOCK11 in mammals. DOCK family proteins mainly regulate actin filament polymerization and/or depolymerization and are GEF proteins, which contribute to cellular signaling events by activating small G proteins. Sponge (Spg) is a Drosophila counterpart to mammalian DOCK3/DOCK4, and plays a role in embryonic central nervous system development, R7 photoreceptor cell differentiation, and adult thorax development. In order to conduct further functional analyses on Spg in vivo, we examined its localization in third instar larval wing imaginal discs. Immunostaining with purified anti-Spg IgG revealed that Spg mainly localized in the air sac primordium (ASP) in wing imaginal discs. Spg is therefore predicted to play an important role in the ASP. The specific knockdown of Spg by the breathless-GAL4 driver in tracheal cells induced lethality accompanied with a defect in ASP development and the induction of apoptosis. The monitoring of ERK signaling activity in wing imaginal discs by immunostaining with anti-diphospho-ERK IgG revealed reductions in the ERK signal cascade in Spg knockdown clones. Furthermore, the overexpression of D-raf suppressed defects in survival and the proliferation of cells in the ASP induced by the knockdown of Spg. Collectively, these results indicate that Spg plays a critical role in ASP development and tracheal cell viability that is mediated by the ERK signaling pathway. - Highlights: • Spg mainly localizes in the air sac primordium in wing imaginal discs. • Spg plays a critical role in air sac primordium development. • Spg positively regulates the ERK signal cascade.

  12. Circadian clock, cell cycle and cancer

    Directory of Open Access Journals (Sweden)

    Cansu Özbayer

    2011-12-01

    Full Text Available There are a few rhythms of our daily lives that we are under the influence. One of them is characterized by predictable changes over a 24-hour timescale called circadian clock. This cellular clock is coordinated by the suprachiasmatic nucleus in the anterior hypothalamus. The clock consist of an autoregulatory transcription-translation feedback loop compose of four genes/proteins; BMAL1, Clock, Cyrptochrome, and Period. BMAL 1 and Clock are transcriptional factors and Period and Cyrptochrome are their targets. Period and Cyrptochrome dimerize in the cytoplasm to enter the nucleus where they inhibit Clock/BMAL activity.It has been demonstrate that circadian clock plays an important role cellular proliferation, DNA damage and repair mechanisms, checkpoints, apoptosis and cancer.

  13. Tre1, a G protein-coupled receptor, directs transepithelial migration of Drosophila germ cells.

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    Prabhat S Kunwar

    2003-12-01

    Full Text Available In most organisms, germ cells are formed distant from the somatic part of the gonad and thus have to migrate along and through a variety of tissues to reach the gonad. Transepithelial migration through the posterior midgut (PMG is the first active step during Drosophila germ cell migration. Here we report the identification of a novel G protein-coupled receptor (GPCR, Tre1, that is essential for this migration step. Maternal tre1 RNA is localized to germ cells, and tre1 is required cell autonomously in germ cells. In tre1 mutant embryos, most germ cells do not exit the PMG. The few germ cells that do leave the midgut early migrate normally to the gonad, suggesting that this gene is specifically required for transepithelial migration and that mutant germ cells are still able to recognize other guidance cues. Additionally, inhibiting small Rho GTPases in germ cells affects transepithelial migration, suggesting that Tre1 signals through Rho1. We propose that Tre1 acts in a manner similar to chemokine receptors required during transepithelial migration of leukocytes, implying an evolutionarily conserved mechanism of transepithelial migration. Recently, the chemokine receptor CXCR4 was shown to direct migration in vertebrate germ cells. Thus, germ cells may more generally use GPCR signaling to navigate the embryo toward their target.

  14. Antioxidant proteins TSA and PAG interact synergistically with Presenilin to modulate Notch signaling in Drosophila.

    Science.gov (United States)

    Wangler, Michael F; Reiter, Lawrence T; Zimm, Georgianna; Trimble-Morgan, Jennifer; Wu, Jane; Bier, Ethan

    2011-07-01

    Alzheimer's disease (AD) pathogenesis is characterized by senile plaques in the brain and evidence of oxidative damage. Oxidative stress may precede plaque formation in AD; however, the link between oxidative damage and plaque formation remains unknown. Presenilins are transmembrane proteins in which mutations lead to accelerated plaque formation and early-onset familial Alzheimer's disease. Presenilins physically interact with two antioxidant enzymes thiol-specific antioxidant (TSA) and proliferation-associated gene (PAG) of the peroxiredoxin family. The functional consequences of these interactions are unclear. In the current study we expressed a presenilin transgene in Drosophila wing and sensory organ precursors of the fly. This caused phenotypes typical of Notch signaling loss-of-function mutations. We found that while expression of TSA or PAG alone produced no phenotype, co-expression of TSA and PAG with presenilin led to an enhanced Notch loss-of-function phenotype. This phenotype was more severe and more penetrant than that caused by the expression of Psn alone. In order to determine whether these phenotypes were indeed affecting Notch signaling, this experiment was performed in a genetic background carrying an activated Notch (Abruptex) allele. The phenotypes were almost completely rescued by this activated Notch allele. These results link peroxiredoxins with the in vivo function of Presenilin, which ultimately connects two key pathogenetic mechanisms in AD, namely, antioxidant activity and plaque formation, and raises the possibility of a role for peroxiredoxin family members in Alzheimer's pathogenesis.

  15. DISCO Interacting Protein 2 regulates axonal bifurcation and guidance of Drosophila mushroom body neurons.

    Science.gov (United States)

    Nitta, Yohei; Yamazaki, Daisuke; Sugie, Atsushi; Hiroi, Makoto; Tabata, Tetsuya

    2017-01-15

    Axonal branching is one of the key processes within the enormous complexity of the nervous system to enable a single neuron to send information to multiple targets. However, the molecular mechanisms that control branch formation are poorly understood. In particular, previous studies have rarely addressed the mechanisms underlying axonal bifurcation, in which axons form new branches via splitting of the growth cone. We demonstrate that DISCO Interacting Protein 2 (DIP2) is required for precise axonal bifurcation in Drosophila mushroom body (MB) neurons by suppressing ectopic bifurcation and regulating the guidance of sister axons. We also found that DIP2 localize to the plasma membrane. Domain function analysis revealed that the AMP-synthetase domains of DIP2 are essential for its function, which may involve exerting a catalytic activity that modifies fatty acids. Genetic analysis and subsequent biochemical analysis suggested that DIP2 is involved in the fatty acid metabolization of acyl-CoA. Taken together, our results reveal a function of DIP2 in the developing nervous system and provide a potential functional relationship between fatty acid metabolism and axon morphogenesis. Copyright © 2016 Elsevier Inc. All rights reserved.

  16. Silver nanoparticles induced heat shock protein 70, oxidative stress and apoptosis in Drosophila melanogaster.

    Science.gov (United States)

    Ahamed, Maqusood; Posgai, Ryan; Gorey, Timothy J; Nielsen, Mark; Hussain, Saber M; Rowe, John J

    2010-02-01

    Due to the intensive commercial application of silver nanoparticles (Ag NPs), risk assessment of this nanoparticle is of great importance. Our previous in vitro study demonstrated that Ag NPs caused DNA damage and apoptosis in mouse embryonic stem cells and fibroblasts. However, toxicity of Ag NPs in vivo is largely lacking. This study was undertaken to examine the toxic effects of well-characterized polysaccharide coated 10 nm Ag NPs on heat shock stress, oxidative stress, DNA damage and apoptosis in Drosophila melanogaster. Third instar larvae of D. melanogaster were fed a diet of standard cornmeal media mixed with Ag NPs at the concentrations of 50 and 100 microg/ml for 24 and 48 h. Ag NPs up-regulated the expression of heat shock protein 70 and induced oxidative stress in D. melanogaster. Malondialdehyde level, an end product of lipid peroxidation was significantly higher while antioxidant glutathione content was significantly lower in Ag NPs exposed organisms. Activities of antioxidant enzyme superoxide dismutase and catalase were also significantly higher in the organisms exposed to Ag NPs. Furthermore, Ag NPs up-regulated the cell cycle checkpoint p53 and cell signaling protein p38 that are involved in the DNA damage repair pathway. Moreover, activities of caspase-3 and caspase-9, markers of apoptosis were significantly higher in Ag NPs exposed organisms. The results indicate that Ag NPs in D. melanogaster induce heat shock stress, oxidative stress, DNA damage and apoptosis. This study suggests that the organism is stressed and thus warrants more careful assessment of Ag NPs using in vivo models to determine if chronic exposure presents developmental and reproductive toxicity. Copyright 2009 Elsevier Inc. All rights reserved.

  17. Silver nanoparticles induced heat shock protein 70, oxidative stress and apoptosis in Drosophila melanogaster

    International Nuclear Information System (INIS)

    Ahamed, Maqusood; Posgai, Ryan; Gorey, Timothy J.; Nielsen, Mark; Hussain, Saber M.; Rowe, John J.

    2010-01-01

    Due to the intensive commercial application of silver nanoparticles (Ag NPs), risk assessment of this nanoparticle is of great importance. Our previous in vitro study demonstrated that Ag NPs caused DNA damage and apoptosis in mouse embryonic stem cells and fibroblasts. However, toxicity of Ag NPs in vivo is largely lacking. This study was undertaken to examine the toxic effects of well-characterized polysaccharide coated 10 nm Ag NPs on heat shock stress, oxidative stress, DNA damage and apoptosis in Drosophila melanogaster. Third instar larvae of D. melanogaster were fed a diet of standard cornmeal media mixed with Ag NPs at the concentrations of 50 and 100 μg/ml for 24 and 48 h. Ag NPs up-regulated the expression of heat shock protein 70 and induced oxidative stress in D. melanogaster. Malondialdehyde level, an end product of lipid peroxidation was significantly higher while antioxidant glutathione content was significantly lower in Ag NPs exposed organisms. Activities of antioxidant enzyme superoxide dismutase and catalase were also significantly higher in the organisms exposed to Ag NPs. Furthermore, Ag NPs up-regulated the cell cycle checkpoint p53 and cell signaling protein p38 that are involved in the DNA damage repair pathway. Moreover, activities of caspase-3 and caspase-9, markers of apoptosis were significantly higher in Ag NPs exposed organisms. The results indicate that Ag NPs in D. melanogaster induce heat shock stress, oxidative stress, DNA damage and apoptosis. This study suggests that the organism is stressed and thus warrants more careful assessment of Ag NPs using in vivo models to determine if chronic exposure presents developmental and reproductive toxicity.

  18. Odorant binding protein 69a connects social interaction to modulation of social responsiveness in Drosophila.

    Science.gov (United States)

    Bentzur, Assa; Shmueli, Anat; Omesi, Liora; Ryvkin, Julia; Knapp, Jon-Michael; Parnas, Moshe; Davis, Fred P; Shohat-Ophir, Galit

    2018-04-01

    Living in a social environment requires the ability to respond to specific social stimuli and to incorporate information obtained from prior interactions into future ones. One of the mechanisms that facilitates social interaction is pheromone-based communication. In Drosophila melanogaster, the male-specific pheromone cis-vaccenyl acetate (cVA) elicits different responses in male and female flies, and functions to modulate behavior in a context and experience-dependent manner. Although it is the most studied pheromone in flies, the mechanisms that determine the complexity of the response, its intensity and final output with respect to social context, sex and prior interaction, are still not well understood. Here we explored the functional link between social interaction and pheromone-based communication and discovered an odorant binding protein that links social interaction to sex specific changes in cVA related responses. Odorant binding protein 69a (Obp69a) is expressed in auxiliary cells and secreted into the olfactory sensilla. Its expression is inversely regulated in male and female flies by social interactions: cVA exposure reduces its levels in male flies and increases its levels in female flies. Increasing or decreasing Obp69a levels by genetic means establishes a functional link between Obp69a levels and the extent of male aggression and female receptivity. We show that activation of cVA-sensing neurons is sufficeint to regulate Obp69a levels in the absence of cVA, and requires active neurotransmission between the sensory neuron to the second order olfactory neuron. The cross-talk between sensory neurons and non-neuronal auxiliary cells at the olfactory sensilla, represents an additional component in the machinery that promotes behavioral plasticity to the same sensory stimuli in male and female flies.

  19. The Putative HORMA Domain Protein Atg101 Dimerizes and Is Required for Starvation-Induced and Selective Autophagy in Drosophila

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    Krisztina Hegedűs

    2014-01-01

    Full Text Available The large-scale turnover of intracellular material including organelles is achieved by autophagy-mediated degradation in lysosomes. Initiation of autophagy is controlled by a protein kinase complex consisting of an Atg1-family kinase, Atg13, FIP200/Atg17, and the metazoan-specific subunit Atg101. Here we show that loss of Atg101 impairs both starvation-induced and basal autophagy in Drosophila. This leads to accumulation of protein aggregates containing the selective autophagy cargo ref(2P/p62. Mapping experiments suggest that Atg101 binds to the N-terminal HORMA domain of Atg13 and may also interact with two unstructured regions of Atg1. Another HORMA domain-containing protein, Mad2, forms a conformational homodimer. We show that Drosophila Atg101 also dimerizes, and it is predicted to fold into a HORMA domain. Atg101 interacts with ref(2P as well, similar to Atg13, Atg8a, Atg16, Atg18, Keap1, and RagC, a known regulator of Tor kinase which coordinates cell growth and autophagy. These results raise the possibility that the interactions and dimerization of the putative HORMA domain protein Atg101 play critical roles in starvation-induced autophagy and proteostasis, by promoting the formation of protein aggregate-containing autophagosomes.

  20. Responses of antioxidant enzymes and heat shock proteins in drosophila to treatment with a pesticide mixture

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    Doganlar Oguzhan

    2015-01-01

    Full Text Available The effects of a mixture of seven pesticides were examined on the expression of antioxidant enzymes, Mn superoxide dismutase (Mn-SOD, catalase (CAT, glutathione synthetase (GS, and heat shock proteins (HSP 26, 60, 70 and 83 in adult fruit flies (Drosophila melanogaster Oregon R. The flies were reared under controlled conditions on artificial diets and treated with a mixture of seven pesticides (molinate, thiobencarb, linuron, phorate, primiphos-methyl, fenvalerate and lambda-cyhalothrin commonly found in water, at concentrations of 0.1, 0.5 and 1 parts per billion (ppb for 1 and 5 days. Quantitative real-time PCR (qRT-PCR analysis of Mn-SOD, CAT and GS expression revealed that the analyzed markers responded significantly to pesticide-induced oxidative stress, in particular on the 5th day of treatment. On the 1st day of treatment, the relative expression of HSP26 and HSP60 genes increased only after exposure to the highest concentrations of pesticides, whereas HSP70 and HSP83 expression increased after exposure to 0.5 and 1 ppb. After five days of treatment, the expression of all HSP genes was increased after exposure to all pesticide concentrations. A positive correlation was determined between the relative expression levels of some HSPs (except HSP60, and antioxidant genes. The observed changes in antioxidant enzyme and HSP mRNA levels in D. melanogaster suggest that the permissible limits of pesticide concentrations for clean drinking water outlined in the regulations of several countries are potentially cytotoxic. The presented findings lend support for reevaluation of these limits.

  1. PH Domain-Arf G Protein Interactions Localize the Arf-GEF Steppke for Cleavage Furrow Regulation in Drosophila.

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    Donghoon M Lee

    Full Text Available The recruitment of GDP/GTP exchange factors (GEFs to specific subcellular sites dictates where they activate small G proteins for the regulation of various cellular processes. Cytohesins are a conserved family of plasma membrane GEFs for Arf small G proteins that regulate endocytosis. Analyses of mammalian cytohesins have identified a number of recruitment mechanisms for these multi-domain proteins, but the conservation and developmental roles for these mechanisms are unclear. Here, we report how the pleckstrin homology (PH domain of the Drosophila cytohesin Steppke affects its localization and activity at cleavage furrows of the early embryo. We found that the PH domain is necessary for Steppke furrow localization, and for it to regulate furrow structure. However, the PH domain was not sufficient for the localization. Next, we examined the role of conserved PH domain amino acid residues that are required for mammalian cytohesins to bind PIP3 or GTP-bound Arf G proteins. We confirmed that the Steppke PH domain preferentially binds PIP3 in vitro through a conserved mechanism. However, disruption of residues for PIP3 binding had no apparent effect on GFP-Steppke localization and effects. Rather, residues for binding to GTP-bound Arf G proteins made major contributions to this Steppke localization and activity. By analyzing GFP-tagged Arf and Arf-like small G proteins, we found that Arf1-GFP, Arf6-GFP and Arl4-GFP, but not Arf4-GFP, localized to furrows. However, analyses of embryos depleted of Arf1, Arf6 or Arl4 revealed either earlier defects than occur in embryos depleted of Steppke, or no detectable furrow defects, possibly because of redundancies, and thus it was difficult to assess how individual Arf small G proteins affect Steppke. Nonetheless, our data show that the Steppke PH domain and its conserved residues for binding to GTP-bound Arf G proteins have substantial effects on Steppke localization and activity in early Drosophila embryos.

  2. USP2-45 Is a Circadian Clock Output Effector Regulating Calcium Absorption at the Post-Translational Level.

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    Daniel Pouly

    Full Text Available The mammalian circadian clock influences most aspects of physiology and behavior through the transcriptional control of a wide variety of genes, mostly in a tissue-specific manner. About 20 clock-controlled genes (CCGs oscillate in virtually all mammalian tissues and are generally considered as core clock components. One of them is Ubiquitin-Specific Protease 2 (Usp2, whose status remains controversial, as it may be a cogwheel regulating the stability or activity of core cogwheels or an output effector. We report here that Usp2 is a clock output effector related to bodily Ca2+ homeostasis, a feature that is conserved across evolution. Drosophila with a whole-body knockdown of the orthologue of Usp2, CG14619 (dUsp2-kd, predominantly die during pupation but are rescued by dietary Ca2+ supplementation. Usp2-KO mice show hyperabsorption of dietary Ca2+ in small intestine, likely due to strong overexpression of the membrane scaffold protein NHERF4, a regulator of the Ca2+ channel TRPV6 mediating dietary Ca2+ uptake. In this tissue, USP2-45 is found in membrane fractions and negatively regulates NHERF4 protein abundance in a rhythmic manner at the protein level. In clock mutant animals (Cry1/Cry2-dKO, rhythmic USP2-45 expression is lost, as well as the one of NHERF4, confirming the inverse relationship between USP2-45 and NHERF4 protein levels. Finally, USP2-45 interacts in vitro with NHERF4 and endogenous Clathrin Heavy Chain. Taken together these data prompt us to define USP2-45 as the first clock output effector acting at the post-translational level at cell membranes and possibly regulating membrane permeability of Ca2+.

  3. The Drosophila Pericentrin-like-protein (PLP cooperates with Cnn to maintain the integrity of the outer PCM

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    Jennifer H. Richens

    2015-08-01

    Full Text Available Centrosomes comprise a pair of centrioles surrounded by a matrix of pericentriolar material (PCM. In vertebrate cells, Pericentrin plays an important part in mitotic PCM assembly, but the Drosophila Pericentrin-like protein (PLP appears to have a more minor role in mitotic fly cells. Here we investigate the function of PLP during the rapid mitotic cycles of the early Drosophila embryo. Unexpectedly, we find that PLP is specifically enriched in the outer-most regions of the PCM, where it largely co-localizes with the PCM scaffold protein Cnn. In the absence of PLP the outer PCM appears to be structurally weakened, and it rapidly disperses along the centrosomal microtubules (MTs. As a result, centrosomal MTs are subtly disorganized in embryos lacking PLP, although mitosis is largely unperturbed and these embryos develop and hatch at near-normal rates. Y2H analysis reveals that PLP can potentially form multiple interactions with itself and with the PCM recruiting proteins Asl, Spd-2 and Cnn. A deletion analysis suggests that PLP participates in a complex network of interactions that ultimately help to strengthen the PCM.

  4. Drosophila MOF controls Checkpoint protein2 and regulates genomic stability during early embryogenesis.

    Science.gov (United States)

    Pushpavalli, Sreerangam N C V L; Sarkar, Arpita; Ramaiah, M Janaki; Chowdhury, Debabani Roy; Bhadra, Utpal; Pal-Bhadra, Manika

    2013-01-24

    In Drosophila embryos, checkpoints maintain genome stability by delaying cell cycle progression that allows time for damage repair or to complete DNA synthesis. Drosophila MOF, a member of MYST histone acetyl transferase is an essential component of male X hyperactivation process. Until recently its involvement in G2/M cell cycle arrest and defects in ionizing radiation induced DNA damage pathways was not well established. Drosophila MOF is highly expressed during early embryogenesis. In the present study we show that haplo-insufficiency of maternal MOF leads to spontaneous mitotic defects like mitotic asynchrony, mitotic catastrophe and chromatid bridges in the syncytial embryos. Such abnormal nuclei are eliminated and digested in the yolk tissues by nuclear fall out mechanism. MOF negatively regulates Drosophila checkpoint kinase 2 tumor suppressor homologue. In response to DNA damage the checkpoint gene Chk2 (Drosophila mnk) is activated in the mof mutants, there by causing centrosomal inactivation suggesting its role in response to genotoxic stress. A drastic decrease in the fall out nuclei in the syncytial embryos derived from mof¹/+; mnkp⁶/+ females further confirms the role of DNA damage response gene Chk2 to ensure the removal of abnormal nuclei from the embryonic precursor pool and maintain genome stability. The fact that mof mutants undergo DNA damage has been further elucidated by the increased number of single and double stranded DNA breaks. mof mutants exhibited genomic instability as evidenced by the occurance of frequent mitotic bridges in anaphase, asynchronous nuclear divisions, disruption of cytoskeleton, inactivation of centrosomes finally leading to DNA damage. Our findings are consistent to what has been reported earlier in mammals that; reduced levels of MOF resulted in increased genomic instability while total loss resulted in lethality. The study can be further extended using Drosophila as model system and carry out the interaction of MOF

  5. Drosophila MOF controls Checkpoint protein2 and regulates genomic stability during early embryogenesis

    Directory of Open Access Journals (Sweden)

    Pushpavalli Sreerangam NCVL

    2013-01-01

    Full Text Available Abstract Background In Drosophila embryos, checkpoints maintain genome stability by delaying cell cycle progression that allows time for damage repair or to complete DNA synthesis. Drosophila MOF, a member of MYST histone acetyl transferase is an essential component of male X hyperactivation process. Until recently its involvement in G2/M cell cycle arrest and defects in ionizing radiation induced DNA damage pathways was not well established. Results Drosophila MOF is highly expressed during early embryogenesis. In the present study we show that haplo-insufficiency of maternal MOF leads to spontaneous mitotic defects like mitotic asynchrony, mitotic catastrophe and chromatid bridges in the syncytial embryos. Such abnormal nuclei are eliminated and digested in the yolk tissues by nuclear fall out mechanism. MOF negatively regulates Drosophila checkpoint kinase 2 tumor suppressor homologue. In response to DNA damage the checkpoint gene Chk2 (Drosophila mnk is activated in the mof mutants, there by causing centrosomal inactivation suggesting its role in response to genotoxic stress. A drastic decrease in the fall out nuclei in the syncytial embryos derived from mof1/+; mnkp6/+ females further confirms the role of DNA damage response gene Chk2 to ensure the removal of abnormal nuclei from the embryonic precursor pool and maintain genome stability. The fact that mof mutants undergo DNA damage has been further elucidated by the increased number of single and double stranded DNA breaks. Conclusion mof mutants exhibited genomic instability as evidenced by the occurance of frequent mitotic bridges in anaphase, asynchronous nuclear divisions, disruption of cytoskeleton, inactivation of centrosomes finally leading to DNA damage. Our findings are consistent to what has been reported earlier in mammals that; reduced levels of MOF resulted in increased genomic instability while total loss resulted in lethality. The study can be further extended using

  6. Variation in sperm displacement and its association with accessory gland protein loci in Drosophila melanogaster.

    Science.gov (United States)

    Clark, A G; Aguadé, M; Prout, T; Harshman, L G; Langley, C H

    1995-01-01

    Genes that influence mating and/or fertilization success may be targets for strong natural selection. If females remate frequently relative to the duration of sperm storage and rate of sperm use, sperm displacement may be an important component of male reproductive success. Although it has long been known that mutant laboratory stocks of Drosophila differ in sperm displacement, the magnitude of the naturally occurring genetic variation in this character has not been systematically quantified. Here we report the results of a screen for variation in sperm displacement among 152 lines of Drosophilia melanogaster that were made homozygous for second and/or third chromosomes recovered from natural populations. Sperm displacement was assayed by scoring the progeny of cn;bw females that had been mated sequentially to cn;bw and tested males in either order. Highly significant differences were seen in both the ability to displace sperm that is resident in the female's reproductive tract and in the ability to resist displacement by subsequent sperm. Most lines exhibited nearly complete displacement, having nearly all progeny sired by the second male, but several lines had as few as half the progeny fathered by the second male. Lines that were identified in the screen for naturally occurring variation in sperm displacement were also characterized for single-strand conformation polymorphisms (SSCP) at seven accessory gland protein (Acp) genes, Glucose dehydrogenase (Gld), and Esterase-6 (Est-6). Acp genes encode proteins that are in some cases known to be transmitted to the female in the seminal fluid and are likely candidates for genes that might mediate the phenomenon of sperm displacement. Significant associations were found between particular Acp alleles at four different loci (Acp26Aa/Ab, Acp29B, Acp36DE and Acp53E) and the ability of males to resist displacement by subsequent sperm. There was no correlation between the ability to displace resident sperm and the ability

  7. Relativistic Ideal Clock

    OpenAIRE

    Bratek, Łukasz

    2015-01-01

    Two particularly simple ideal clocks exhibiting intrinsic circular motion with the speed of light and opposite spin alignment are described. The clocks are singled out by singularities of an inverse Legendre transformation for relativistic rotators of which mass and spin are fixed parameters. Such clocks work always the same way, no matter how they move. When subject to high accelerations or falling in strong gravitational fields of black holes, the clocks could be used to test the clock hypo...

  8. Lie Group Analysis of the Photo-Induced Fluorescence of Drosophila Oogenesis with the Asymmetrically Localized Gurken Protein.

    Directory of Open Access Journals (Sweden)

    Jen-Cheng Wang

    Full Text Available Lie group analysis of the photo-induced fluorescence of Drosophila oogenesis with the asymmetrically localized Gurken protein has been performed systematically to assess the roles of ligand-receptor complexes in follicle cells. The (2×2 matrix representations resulting from the polarized tissue spectra were employed to characterize the asymmetrical Gurken distributions. It was found that the fluorescence of the wild-type egg shows the Lie point symmetry X 23 at early stages of oogenesis. However, due to the morphogen regulation by intracellular proteins and extracellular proteins, the fluorescence of the embryogenesis with asymmetrically localized Gurken expansions exhibits specific symmetry features: Lie point symmetry Z 1 and Lie point symmetry X 1. The novel approach developed herein was successfully used to validate that the invariant-theoretical characterizations are consonant with the observed asymmetric fluctuations during early embryological development.

  9. The phospho-occupancy of an atypical SLIMB-binding site on PERIOD that is phosphorylated by DOUBLETIME controls the pace of the clock.

    Science.gov (United States)

    Chiu, Joanna C; Vanselow, Jens T; Kramer, Achim; Edery, Isaac

    2008-07-01

    A common feature of animal circadian clocks is the progressive phosphorylation of PERIOD (PER) proteins, which is highly dependent on casein kinase Idelta/epsilon (CKIdelta/epsilon; termed DOUBLETIME [DBT] in Drosophila) and ultimately leads to the rapid degradation of hyperphosphorylated isoforms via a mechanism involving the F-box protein, beta-TrCP (SLIMB in Drosophila). Here we use the Drosophila melanogaster model system, and show that a key step in controlling the speed of the clock is phosphorylation of an N-terminal Ser (S47) by DBT, which collaborates with other nearby phosphorylated residues to generate a high-affinity atypical SLIMB-binding site on PER. DBT-dependent increases in the phospho-occupancy of S47 are temporally gated, dependent on the centrally located DBT docking site on PER and partially counterbalanced by protein phosphatase activity. We propose that the gradual DBT-mediated phosphorylation of a nonconsensus SLIMB-binding site establishes a temporal threshold for when in a daily cycle the majority of PER proteins are tagged for rapid degradation. Surprisingly, most of the hyperphosphorylation is unrelated to direct effects on PER stability. We also use mass spectrometry to map phosphorylation sites on PER, leading to the identification of a number of "phospho-clusters" that explain several of the classic per mutants.

  10. Basic leucine zipper protein Cnc-C is a substrate and transcriptional regulator of the Drosophila 26S proteasome.

    Science.gov (United States)

    Grimberg, Kristian Björk; Beskow, Anne; Lundin, Daniel; Davis, Monica M; Young, Patrick

    2011-02-01

    While the 26S proteasome is a key proteolytic complex, little is known about how proteasome levels are maintained in higher eukaryotic cells. Here we describe an RNA interference (RNAi) screen of Drosophila melanogaster that was used to identify transcription factors that may play a role in maintaining levels of the 26S proteasome. We used an RNAi library against 993 Drosophila transcription factor genes to identify genes whose suppression in Schneider 2 cells stabilized a ubiquitin-green fluorescent protein reporter protein. This screen identified Cnc (cap 'n' collar [CNC]; basic region leucine zipper) as a candidate transcriptional regulator of proteasome component expression. In fact, 20S proteasome activity was reduced in cells depleted of cnc. Immunoblot assays against proteasome components revealed a general decline in both 19S regulatory complex and 20S proteasome subunits after RNAi depletion of this transcription factor. Transcript-specific silencing revealed that the longest of the seven transcripts for the cnc gene, cnc-C, was needed for proteasome and p97 ATPase production. Quantitative reverse transcription-PCR confirmed the role of Cnc-C in activation of transcription of genes encoding proteasome components. Expression of a V5-His-tagged form of Cnc-C revealed that the transcription factor is itself a proteasome substrate that is stabilized when the proteasome is inhibited. We propose that this single cnc gene in Drosophila resembles the ancestral gene family of mammalian nuclear factor erythroid-derived 2-related transcription factors, which are essential in regulating oxidative stress and proteolysis.

  11. Loss of circadian clock accelerates aging in neurodegeneration-prone mutants.

    Science.gov (United States)

    Krishnan, Natraj; Rakshit, Kuntol; Chow, Eileen S; Wentzell, Jill S; Kretzschmar, Doris; Giebultowicz, Jadwiga M

    2012-03-01

    Circadian clocks generate rhythms in molecular, cellular, physiological, and behavioral processes. Recent studies suggest that disruption of the clock mechanism accelerates organismal senescence and age-related pathologies in mammals. Impaired circadian rhythms are observed in many neurological diseases; however, it is not clear whether loss of rhythms is the cause or result of neurodegeneration, or both. To address this important question, we examined the effects of circadian disruption in Drosophila melanogaster mutants that display clock-unrelated neurodegenerative phenotypes. We combined a null mutation in the clock gene period (per(01)) that abolishes circadian rhythms, with a hypomorphic mutation in the carbonyl reductase gene sniffer (sni(1)), which displays oxidative stress induced neurodegeneration. We report that disruption of circadian rhythms in sni(1) mutants significantly reduces their lifespan compared to single mutants. Shortened lifespan in double mutants was coupled with accelerated neuronal degeneration evidenced by vacuolization in the adult brain. In addition, per(01)sni(1) flies showed drastically impaired vertical mobility and increased accumulation of carbonylated proteins compared to age-matched single mutant flies. Loss of per function does not affect sni mRNA expression, suggesting that these genes act via independent pathways producing additive effects. Finally, we show that per(01) mutation accelerates the onset of brain pathologies when combined with neurodegeneration-prone mutation in another gene, swiss cheese (sws(1)), which does not operate through the oxidative stress pathway. Taken together, our data suggest that the period gene may be causally involved in neuroprotective pathways in aging Drosophila. Copyright © 2011 Elsevier Inc. All rights reserved.

  12. Interspecific studies of circadian genes period and timeless in Drosophila.

    Science.gov (United States)

    Noreen, Shumaila; Pegoraro, Mirko; Nouroz, Faisal; Tauber, Eran; Kyriacou, Charalambos P

    2018-03-30

    The level of rescue of clock function in genetically arrhythmic Drosophila melanogaster hosts using interspecific clock gene transformation was used to study the putative intermolecular coevolution between interacting clock proteins. Among them PER and TIM are the two important negative regulators of the circadian clock feedback loop. We transformed either the D. pseudoobscura per or tim transgenes into the corresponding arrhythmic D. melanogaster mutant (per01 or tim01) and observed >50% rhythmicity but the period of activity rhythm was either longer (D. pseudoobscura-per) or shorter than 24 h (D. pseudoobscura-tim) compared to controls. By introducing both transgenes simultaneously into double mutants, we observed that the period of the activity rhythm was rescued by the pair of hemizygous transgenes (~24 h). These flies also showed a more optimal level of temperature compensation for the period. Under LD 12:12 these flies have a D. pseudoobscura like activity profile with the absence of morning anticipation as well as a very prominent earlier evening peak of activity rhythm. These observation are consistent with the view that TIM and PER form a heterospecific coevolved module at least for the circadian period of activity rhythms. However the strength of rhythmicity was reduced by having both transgenes present, so while evidence for a coevolution between PER and TIM is observed for some characters it is not for others. Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.

  13. The intracellular domain of the Drosophila cholinesterase-like neural adhesion protein, gliotactin, is natively unfolded

    NARCIS (Netherlands)

    Zeev-Ben-Mordehai, T; Rydberg, EH; Solomon, A.; Toker, L; Auld, VJ; Silman, I.; Botti, S; Sussman, J.L.

    2003-01-01

    Drosophila gliotactin (Gli) is a 109-kDa transmembrane, cholinesterase-like adhesion molecule (CLAM), expressed in peripheral glia, that is crucial for formation of the blood-nerve barrier. The intracellular portion (Gli-cyt) was cloned and expressed in the cytosolic fraction of Escherichia coli

  14. Unfolded Protein Response-regulated Drosophila Fic (dFic) Protein Reversibly AMPylates BiP Chaperone during Endoplasmic Reticulum Homeostasis*

    Science.gov (United States)

    Ham, Hyeilin; Woolery, Andrew R.; Tracy, Charles; Stenesen, Drew; Krämer, Helmut; Orth, Kim

    2014-01-01

    Drosophila Fic (dFic) mediates AMPylation, a covalent attachment of adenosine monophosphate (AMP) from ATP to hydroxyl side chains of protein substrates. Here, we identified the endoplasmic reticulum (ER) chaperone BiP as a substrate for dFic and mapped the modification site to Thr-366 within the ATPase domain. The level of AMPylated BiP in Drosophila S2 cells is high during homeostasis, whereas the level of AMPylated BiP decreases upon the accumulation of misfolded proteins in the ER. Both dFic and BiP are transcriptionally activated upon ER stress, supporting the role of dFic in the unfolded protein response pathway. The inactive conformation of BiP is the preferred substrate for dFic, thus endorsing a model whereby AMPylation regulates the function of BiP as a chaperone, allowing acute activation of BiP by deAMPylation during an ER stress response. These findings not only present the first substrate of eukaryotic AMPylator but also provide a target for regulating the unfolded protein response, an emerging avenue for cancer therapy. PMID:25395623

  15. The lysosomal enzyme receptor protein (LERP is not essential, but is implicated in lysosomal function in Drosophila melanogaster

    Directory of Open Access Journals (Sweden)

    Medina Hasanagic

    2015-10-01

    Full Text Available The lysosomal enzyme receptor protein (LERP of Drosophila melanogaster is the ortholog of the mammalian cation-independent mannose 6-phosphate (Man 6-P receptor, which mediates trafficking of newly synthesized lysosomal acid hydrolases to lysosomes. However, flies lack the enzymes necessary to make the Man 6-P mark, and the amino acids implicated in Man 6-P binding by the mammalian receptor are not conserved in LERP. Thus, the function of LERP in sorting of lysosomal enzymes to lysosomes in Drosophila is unclear. Here, we analyze the consequence of LERP depletion in S2 cells and intact flies. RNAi-mediated knockdown of LERP in S2 cells had little or no effect on the cellular content or secretion of several lysosomal hydrolases. We generated a novel Lerp null mutation, LerpF6, which abolishes LERP protein expression. Lerp mutants have normal viability and fertility and display no overt phenotypes other than reduced body weight. Lerp mutant flies exhibit a 30–40% decrease in the level of several lysosomal hydrolases, and are hypersensitive to dietary chloroquine and starvation, consistent with impaired lysosome function. Loss of LERP also enhances an eye phenotype associated with defective autophagy. Our findings implicate Lerp in lysosome function and autophagy.

  16. Circadian Clock Dysfunction and Psychiatric Disease: Could Fruit Flies have a Say?

    Science.gov (United States)

    Zordan, Mauro Agostino; Sandrelli, Federica

    2015-01-01

    There is evidence of a link between the circadian system and psychiatric diseases. Studies in humans and mammals suggest that environmental and/or genetic disruption of the circadian system leads to an increased liability to psychiatric disease. Disruption of clock genes and/or the clock network might be related to the etiology of these pathologies; also, some genes, known for their circadian clock functions, might be associated to mental illnesses through clock-independent pleiotropy. Here, we examine the features which we believe make Drosophila melanogaster a model apt to study the role of the circadian clock in psychiatric disease. Despite differences in the organization of the clock system, the molecular architecture of the Drosophila and mammalian circadian oscillators are comparable and many components are evolutionarily related. In addition, Drosophila has a rather complex nervous system, which shares much at the cell and neurobiological level with humans, i.e., a tripartite brain, the main neurotransmitter systems, and behavioral traits: circadian behavior, learning and memory, motivation, addiction, social behavior. There is evidence that the Drosophila brain shares some homologies with the vertebrate cerebellum, basal ganglia, and hypothalamus-pituitary-adrenal axis, the dysfunctions of which have been tied to mental illness. We discuss Drosophila in comparison to mammals with reference to the: organization of the brain and neurotransmitter systems; architecture of the circadian clock; clock-controlled behaviors. We sum up current knowledge on behavioral endophenotypes, which are amenable to modeling in flies, such as defects involving sleep, cognition, or social interactions, and discuss the relationship of the circadian system to these traits. Finally, we consider if Drosophila could be a valuable asset to understand the relationship between circadian clock malfunction and psychiatric disease.

  17. Circadian clock dysfunction and psychiatric disease: could fruit flies have a say?

    Directory of Open Access Journals (Sweden)

    Mauro Agostino Zordan

    2015-04-01

    Full Text Available There is evidence of a link between the circadian system and psychiatric diseases. Studies in humans and mammals suggest that environmental and/or genetic disruption of the circadian system lead to an increased liability to psychiatric disease. Disruption of clock genes and/or the clock network might be related to the etiology of these pathologies; also, some genes, known for their circadian clock functions, might be associated to mental illnesses through clock-independent pleiotropy. Here we examine the features which we believe make Drosophila melanogaster a model apt to study the role of the circadian clock in psychiatric disease. Despite differences in the organization of the clock system, the molecular architecture of the Drosophila and mammalian circadian oscillators are comparable and many components are evolutionarily related. In addition, Drosophila has a rather complex nervous system, which shares much at the cell and neurobiological level with humans, i.e. a tripartite brain, the main neurotransmitter systems, and behavioral traits: circadian behavior, learning and memory, motivation, addiction, social behavior. There is evidence that the Drosophila brain shares some homologies with the vertebrate cerebellum, basal ganglia and hypothalamus-pituitary-adrenal axis, the dysfunctions of which have been tied to mental illness. We discuss Drosophila in comparison to mammals with reference to the: organization of the brain and neurotransmitter systems; architecture of the circadian clock; clock-controlled behaviors. We sum up current knowledge on behavioral endophenotypes which are amenable to modeling in flies, such as defects involving sleep, cognition, or social interactions and discuss the relationship of the circadian system to these traits. Finally, we consider if Drosophila could be a valuable asset to understand the relationship between circadian clock malfunction and psychiatric disease.

  18. dyschronic, a Drosophila homolog of a deaf-blindness gene, regulates circadian output and Slowpoke channels.

    Directory of Open Access Journals (Sweden)

    James E C Jepson

    Full Text Available Many aspects of behavior and physiology are under circadian control. In Drosophila, the molecular clock that regulates rhythmic patterns of behavior has been extensively characterized. In contrast, genetic loci involved in linking the clock to alterations in motor activity have remained elusive. In a forward-genetic screen, we uncovered a new component of the circadian output pathway, which we have termed dyschronic (dysc. dysc mutants exhibit arrhythmic locomotor behavior, yet their eclosion rhythms are normal and clock protein cycling remains intact. Intriguingly, dysc is the closest Drosophila homolog of whirlin, a gene linked to type II Usher syndrome, the leading cause of deaf-blindness in humans. Whirlin and other Usher proteins are expressed in the mammalian central nervous system, yet their function in the CNS has not been investigated. We show that DYSC is expressed in major neuronal tracts and regulates expression of the calcium-activated potassium channel SLOWPOKE (SLO, an ion channel also required in the circadian output pathway. SLO and DYSC are co-localized in the brain and control each other's expression post-transcriptionally. Co-immunoprecipitation experiments demonstrate they form a complex, suggesting they regulate each other through protein-protein interaction. Furthermore, electrophysiological recordings of neurons in the adult brain show that SLO-dependent currents are greatly reduced in dysc mutants. Our work identifies a Drosophila homolog of a deaf-blindness gene as a new component of the circadian output pathway and an important regulator of ion channel expression, and suggests novel roles for Usher proteins in the mammalian nervous system.

  19. The Drosophila melanogaster DmCK2beta transcription unit encodes for functionally non-redundant protein isoforms.

    Science.gov (United States)

    Jauch, Eike; Wecklein, Heike; Stark, Felix; Jauch, Mandy; Raabe, Thomas

    2006-06-07

    Genes encoding for the two evolutionary highly conserved subunits of a heterotetrameric protein kinase CK2 holoenzyme are present in all examined eukaryotic genomes. Depending on the organism, multiple transcription units encoding for a catalytically active CK2alpha subunit and/or a regulatory CK2beta subunit may exist. The phosphotransferase activity of members of the protein kinase CK2alpha family is thought to be independent of second messengers but is modulated by interaction with CK2beta-like proteins. In the genome of Drosophila melanogaster, one gene encoding for a CK2alpha subunit and three genes encoding for CK2beta-like proteins are present. The X-linked DmCK2beta transcription unit encodes for several CK2beta protein isoforms due to alternative splicing of its primary transcript. We addressed the question whether CK2beta-like proteins are redundant in function. Our in vivo experiments show that variations of the very C-terminal tail of CK2beta isoforms encoded by the X-linked DmCK2beta transcription unit influence their functional properties. In addition, we find that CK2beta-like proteins encoded by the autosomal D. melanogaster genes CK2betates and CK2beta' cannot fully substitute for a loss of CK2beta isoforms encoded by DmCK2beta.

  20. Isolation of a novel protein, P12-from adult Drosophila melanogaster that inhibits deoxyribonucleoside and protein kinase activities and activates 3'-5'-exonuclease activity

    DEFF Research Database (Denmark)

    Christiansen, Louise Slot; Zanten, Gabriella van; Berenstein, Dvora

    2016-01-01

    We have previously found that Drosophila melanogaster only has one deoxyribonucleoside kinase, Dm-dNK, however, capable to phosphorylate all four natural deoxyribonucleosides. Dm-dNK was originally isolated from an embryonic cell line. We wanted to study the expression of Dm-dNK during development......-dNK, also inhibited the two protein kinases to the same degree. Furthermore, testing P12 in a DNA polymerase based assay we found that the 3'-5'-exonuclease part of the DNA polymerase (Klenow polymerase) was activated....

  1. Protein and carbohydrate composition of larval food affects tolerance tothermal stress and desiccation in adult Drosophila melanogaster

    DEFF Research Database (Denmark)

    Andersen, Laila H; Kristensen, Torsten N; Loeschcke, Volker

    2010-01-01

    stress compared to males. Egg production was highest in females that had developed on the protein-enriched medium. However, there was a sex-specific effect of nutrition on egg-to-adult viability, with higher viability for males developing on the sucrose-enriched medium, while female survival was highest......Larval nutrition may affect a range of different life history traits as well as responses to environmental stress in adult insects. Here we test whether raising larvae of fruit flies, Drosophila melanogaster, on two different nutritional regimes affects resistance to cold, heat and desiccation....... In contrast, flies developed on the carbohydrate-enriched growth medium recovered faster from chill coma stress compared to flies developed on a protein-enriched medium. We also found gender differences in stress tolerance, with female flies being more tolerant to chill coma, heat knockdown and desiccation...

  2. High-resolution two-dimensional gel analysis of proteins in wing imaginal discs: A data base of Drosophila

    International Nuclear Information System (INIS)

    Santaren, J.F.; Garcia-Bellido, A.

    1990-01-01

    An improved method of high-resolution two-dimensional gel electrophoresis has been used to study the patterns of protein synthesis in wing imaginal discs of late instar larvae of Drosophila melanogaster. A small number of discs were radiolabeled with a mixture of 14 C-labeled amino acids or with [ 35 S]methionine and the pattern of labeled proteins was analyzed. One thousand and twenty-five polypeptides (787 acidic (IEF) and 238 basic (NEPHGE)) from wing discs of several wild-type strains have so far been separated and cataloged. All these polypeptides have been numbered and presented in a reference map for further studies. When comparing patterns of label we have found small quantitative differences in rate of synthesis between individuals of the same strain, not due to sexual differences, and very few quantitative and qualitative differences between groups of individuals of different strains

  3. The Microtubule Regulatory Protein Stathmin Is Required to Maintain the Integrity of Axonal Microtubules in Drosophila

    Science.gov (United States)

    Duncan, Jason E.; Lytle, Nikki K.; Zuniga, Alfredo; Goldstein, Lawrence S. B.

    2013-01-01

    Axonal transport, a form of long-distance, bi-directional intracellular transport that occurs between the cell body and synaptic terminal, is critical in maintaining the function and viability of neurons. We have identified a requirement for the stathmin (stai) gene in the maintenance of axonal microtubules and regulation of axonal transport in Drosophila . The stai gene encodes a cytosolic phosphoprotein that regulates microtubule dynamics by partitioning tubulin dimers between pools of soluble tubulin and polymerized microtubules, and by directly binding to microtubules and promoting depolymerization. Analysis of stai function in Drosophila , which has a single stai gene, circumvents potential complications with studies performed in vertebrate systems in which mutant phenotypes may be compensated by genetic redundancy of other members of the stai gene family. This has allowed us to identify an essential function for stai in the maintenance of the integrity of axonal microtubules. In addition to the severe disruption in the abundance and architecture of microtubules in the axons of stai mutant Drosophila , we also observe additional neurological phenotypes associated with loss of stai function including a posterior paralysis and tail-flip phenotype in third instar larvae, aberrant accumulation of transported membranous organelles in stai deficient axons, a progressive bang-sensitive response to mechanical stimulation reminiscent of the class of Drosophila mutants used to model human epileptic seizures, and a reduced adult lifespan. Reductions in the levels of Kinesin-1, the primary anterograde motor in axonal transport, enhance these phenotypes. Collectively, our results indicate that stai has an important role in neuronal function, likely through the maintenance of microtubule integrity in the axons of nerves of the peripheral nervous system necessary to support and sustain long-distance axonal transport. PMID:23840848

  4. Variation in Sperm Displacement and Its Association with Accessory Gland Protein Loci in Drosophila Melanogaster

    OpenAIRE

    Clark, A. G.; Aguade, M.; Prout, T.; Harshman, L. G.; Langley, C. H.

    1995-01-01

    Genes that influence mating and/or fertilization success may be targets for strong natural selection. If females remate frequently relative to the duration of sperm storage and rate of sperm use, sperm displacement may be an important component of male reproductive success. Although it has long been known that mutant laboratory stocks of Drosophila differ in sperm displacement, the magnitude of the naturally occurring genetic variation in this character has not been systematically quantified....

  5. CREB Binding Protein Functions During Successive Stages of Eye Development in Drosophila

    OpenAIRE

    Kumar, Justin P.; Jamal, Tazeen; Doetsch, Alex; Turner, F. Rudolf; Duffy, Joseph B.

    2004-01-01

    During the development of the compound eye of Drosophila several signaling pathways exert both positive and inhibitory influences upon an array of nuclear transcription factors to produce a near-perfect lattice of unit eyes or ommatidia. Individual cells within the eye are exposed to many extracellular signals, express multiple surface receptors, and make use of a large complement of cell-subtype-specific DNA-binding transcription factors. Despite this enormous complexity, each cell will make...

  6. Localization and activation of the Drosophila protease easter require the ER-resident saposin-like protein seele.

    Science.gov (United States)

    Stein, David; Charatsi, Iphigenie; Cho, Yong Suk; Zhang, Zhenyu; Nguyen, Jesse; DeLotto, Robert; Luschnig, Stefan; Moussian, Bernard

    2010-11-09

    Drosophila embryonic dorsal-ventral polarity is generated by a series of serine protease processing events in the egg perivitelline space. Gastrulation Defective processes Snake, which then cleaves Easter, which then processes Spätzle into the activating ligand for the Toll receptor. seele was identified in a screen for mutations that, when homozygous in ovarian germline clones, lead to the formation of progeny embryos with altered embryonic patterning; maternal loss of seele function leads to the production of moderately dorsalized embryos. By combining constitutively active versions of Gastrulation Defective, Snake, Easter, and Spätzle with loss-of-function alleles of seele, we find that Seele activity is dispensable for Spätzle-mediated activation of Toll but is required for Easter, Snake, and Gastrulation Defective to exert their effects on dorsal-ventral patterning. Moreover, Seele function is required specifically for secretion of Easter from the developing embryo into the perivitelline space and for Easter processing. Seele protein resides in the endoplasmic reticulum of blastoderm embryos, suggesting a role in the trafficking of Easter to the perivitelline space, prerequisite to its processing and function. Easter transport to the perivitelline space represents a previously unappreciated control point in the signal transduction pathway that controls Drosophila embryonic dorsal-ventral polarity. Copyright © 2010 Elsevier Ltd. All rights reserved.

  7. Organ-specific gene expression: the bHLH protein Sage provides tissue specificity to Drosophila FoxA.

    Science.gov (United States)

    Fox, Rebecca M; Vaishnavi, Aria; Maruyama, Rika; Andrew, Deborah J

    2013-05-01

    FoxA transcription factors play major roles in organ-specific gene expression, regulating, for example, glucagon expression in the pancreas, GLUT2 expression in the liver, and tyrosine hydroxylase expression in dopaminergic neurons. Organ-specific gene regulation by FoxA proteins is achieved through cooperative regulation with a broad array of transcription factors with more limited expression domains. Fork head (Fkh), the sole Drosophila FoxA family member, is required for the development of multiple distinct organs, yet little is known regarding how Fkh regulates tissue-specific gene expression. Here, we characterize Sage, a bHLH transcription factor expressed exclusively in the Drosophila salivary gland (SG). We show that Sage is required for late SG survival and normal tube morphology. We find that many Sage targets, identified by microarray analysis, encode SG-specific secreted cargo, transmembrane proteins, and the enzymes that modify these proteins. We show that both Sage and Fkh are required for the expression of Sage target genes, and that co-expression of Sage and Fkh is sufficient to drive target gene expression in multiple cell types. Sage and Fkh drive expression of the bZip transcription factor Senseless (Sens), which boosts expression of Sage-Fkh targets, and Sage, Fkh and Sens colocalize on SG chromosomes. Importantly, expression of Sage-Fkh target genes appears to simply add to the tissue-specific gene expression programs already established in other cell types, and Sage and Fkh cannot alter the fate of most embryonic cell types even when expressed early and continuously.

  8. Functional analysis of the glycogen binding subunit CG9238/Gbs-70E of protein phosphatase 1 in Drosophila melanogaster.

    Science.gov (United States)

    Kerekes, Éva; Kókai, Endre; Páldy, Ferenc Sándor; Dombrádi, Viktor

    2014-06-01

    The product of the CG9238 gene that we termed glycogen binding subunit 70E (Gbs-70E) was characterized by biochemical and molecular genetics methods. The interaction between Gbs-70E and all catalytic subunits of protein phosphatase 1 (Pp1-87B, Pp1-9C, Pp1-96A and Pp1-13C) of Drosophila melanogaster was confirmed by pairwise yeast two-hybrid tests, co-immunoprecipitation and pull down experiments. The binding of Gbs-70E to glycogen was demonstrated by sedimentation analysis. With RT-PCR we found that the mRNAs coding for the longer Gbs-70E PB/PC protein were expressed in all developmental stages of the fruit flies while the mRNA for the shorter Gbs-70E PA was restricted to the eggs and the ovaries of the adult females. The development specific expression of the shorter splice variant was not conserved in different Drosophila species. The expression level of the gene was manipulated by P-element insertions and gene deletion to analyze the functions of the gene product. A small or moderate reduction in the gene expression resulted in no significant changes, however, a deletion mutant expressing very low level of the transcript lived shorter and exhibited reduced glycogen content in the imagos. In addition, the gene deletion decreased the fertility of the fruit flies. Our results prove that Gbs-70E functions as the glycogen binding subunit of protein phosphatase 1 that regulates glycogen content and plays a role in the development of eggs in D. melanogaster. Copyright © 2014 Elsevier Ltd. All rights reserved.

  9. Protein O-Mannosyltransferases Affect Sensory Axon Wiring and Dynamic Chirality of Body Posture in the Drosophila Embryo.

    Science.gov (United States)

    Baker, Ryan; Nakamura, Naosuke; Chandel, Ishita; Howell, Brooke; Lyalin, Dmitry; Panin, Vladislav M

    2018-02-14

    Genetic defects in protein O-mannosyltransferase 1 (POMT1) and POMT2 underlie severe muscular dystrophies. POMT genes are evolutionarily conserved in metazoan organisms. In Drosophila , both male and female POMT mutants show a clockwise rotation of adult abdominal segments, suggesting a chirality of underlying pathogenic mechanisms. Here we described and analyzed a similar phenotype in POMT mutant embryos that shows left-handed body torsion. Our experiments demonstrated that coordinated muscle contraction waves are associated with asymmetric embryo rolling, unveiling a new chirality marker in Drosophila development. Using genetic and live-imaging approaches, we revealed that the torsion phenotype results from differential rolling and aberrant patterning of peristaltic waves of muscle contractions. Our results demonstrated that peripheral sensory neurons are required for normal contractions that prevent the accumulation of torsion. We found that POMT mutants show abnormal axonal connections of sensory neurons. POMT transgenic expression limited to sensory neurons significantly rescued the torsion phenotype, axonal connectivity defects, and abnormal contractions in POMT mutant embryos. Together, our data suggested that protein O-mannosylation is required for normal sensory feedback to control coordinated muscle contractions and body posture. This mechanism may shed light on analogous functions of POMT genes in mammals and help to elucidate the etiology of neurological defects in muscular dystrophies. SIGNIFICANCE STATEMENT Protein O-mannosyltransferases (POMTs) are evolutionarily conserved in metazoans. Mutations in POMTs cause severe muscular dystrophies associated with pronounced neurological defects. However, neurological functions of POMTs remain poorly understood. We demonstrated that POMT mutations in Drosophila result in abnormal muscle contractions and cause embryo torsion. Our experiments uncovered a chirality of embryo movements and a unique POMT -dependent

  10. Drosophila Pumilio protein contains multiple autonomous repression domains that regulate mRNAs independently of Nanos and brain tumor.

    Science.gov (United States)

    Weidmann, Chase A; Goldstrohm, Aaron C

    2012-01-01

    Drosophila melanogaster Pumilio is an RNA-binding protein that potently represses specific mRNAs. In developing embryos, Pumilio regulates a key morphogen, Hunchback, in collaboration with the cofactor Nanos. To investigate repression by Pumilio and Nanos, we created cell-based assays and found that Pumilio inhibits translation and enhances mRNA decay independent of Nanos. Nanos robustly stimulates repression through interactions with the Pumilio RNA-binding domain. We programmed Pumilio to recognize a new binding site, which garners repression of new target mRNAs. We show that cofactors Brain Tumor and eIF4E Homologous Protein are not obligatory for Pumilio and Nanos activity. The conserved RNA-binding domain of Pumilio was thought to be sufficient for its function. Instead, we demonstrate that three unique domains in the N terminus of Pumilio possess the major repressive activity and can function autonomously. The N termini of insect and vertebrate Pumilio and Fem-3 binding factors (PUFs) are related, and we show that corresponding regions of human PUM1 and PUM2 have repressive activity. Other PUF proteins lack these repression domains. Our findings suggest that PUF proteins have evolved new regulatory functions through protein sequences appended to their conserved PUF repeat RNA-binding domains.

  11. rigor mortis encodes a novel nuclear receptor interacting protein required for ecdysone signaling during Drosophila larval development.

    Science.gov (United States)

    Gates, Julie; Lam, Geanette; Ortiz, José A; Losson, Régine; Thummel, Carl S

    2004-01-01

    Pulses of the steroid hormone ecdysone trigger the major developmental transitions in Drosophila, including molting and puparium formation. The ecdysone signal is transduced by the EcR/USP nuclear receptor heterodimer that binds to specific response elements in the genome and directly regulates target gene transcription. We describe a novel nuclear receptor interacting protein encoded by rigor mortis (rig) that is required for ecdysone responses during larval development. rig mutants display defects in molting, delayed larval development, larval lethality, duplicated mouth parts, and defects in puparium formation--phenotypes that resemble those seen in EcR, usp, E75A and betaFTZ-F1 mutants. Although the expression of these nuclear receptor genes is essentially normal in rig mutant larvae, the ecdysone-triggered switch in E74 isoform expression is defective. rig encodes a protein with multiple WD-40 repeats and an LXXLL motif, sequences that act as specific protein-protein interaction domains. Consistent with the presence of these elements and the lethal phenotypes of rig mutants, Rig protein interacts with several Drosophila nuclear receptors in GST pull-down experiments, including EcR, USP, DHR3, SVP and betaFTZ-F1. The ligand binding domain of betaFTZ-F1 is sufficient for this interaction, which can occur in an AF-2-independent manner. Antibody stains reveal that Rig protein is present in the brain and imaginal discs of second and third instar larvae, where it is restricted to the cytoplasm. In larval salivary gland and midgut cells, however, Rig shuttles between the cytoplasm and nucleus in a spatially and temporally regulated manner, at times that correlate with the major lethal phase of rig mutants and major switches in ecdysone-regulated gene expression. Taken together, these data indicate that rig exerts essential functions during larval development through gene-specific effects on ecdysone-regulated transcription, most likely as a cofactor for one or more

  12. The Drosophila Microtubule-Associated Protein Mars Stabilizes Mitotic Spindles by Crosslinking Microtubules through Its N-Terminal Region

    Science.gov (United States)

    Zhang, Gang; Beati, Hamze; Nilsson, Jakob; Wodarz, Andreas

    2013-01-01

    Correct segregation of genetic material relies on proper assembly and maintenance of the mitotic spindle. How the highly dynamic microtubules (MTs) are maintained in stable mitotic spindles is a key question to be answered. Motor and non-motor microtubule associated proteins (MAPs) have been reported to stabilize the dynamic spindle through crosslinking adjacent MTs. Mars, a novel MAP, is essential for the early development of Drosophila embryos. Previous studies showed that Mars is required for maintaining an intact mitotic spindle but did not provide a molecular mechanism for this function. Here we show that Mars is able to stabilize the mitotic spindle in vivo. Both in vivo and in vitro data reveal that the N-terminal region of Mars functions in the stabilization of the mitotic spindle by crosslinking adjacent MTs. PMID:23593258

  13. The Drosophila microtubule-associated protein mars stabilizes mitotic spindles by crosslinking microtubules through its N-terminal region.

    Directory of Open Access Journals (Sweden)

    Gang Zhang

    Full Text Available Correct segregation of genetic material relies on proper assembly and maintenance of the mitotic spindle. How the highly dynamic microtubules (MTs are maintained in stable mitotic spindles is a key question to be answered. Motor and non-motor microtubule associated proteins (MAPs have been reported to stabilize the dynamic spindle through crosslinking adjacent MTs. Mars, a novel MAP, is essential for the early development of Drosophila embryos. Previous studies showed that Mars is required for maintaining an intact mitotic spindle but did not provide a molecular mechanism for this function. Here we show that Mars is able to stabilize the mitotic spindle in vivo. Both in vivo and in vitro data reveal that the N-terminal region of Mars functions in the stabilization of the mitotic spindle by crosslinking adjacent MTs.

  14. The Drosophila microtubule-associated protein mars stabilizes mitotic spindles by crosslinking microtubules through its N-terminal region.

    Science.gov (United States)

    Zhang, Gang; Beati, Hamze; Nilsson, Jakob; Wodarz, Andreas

    2013-01-01

    Correct segregation of genetic material relies on proper assembly and maintenance of the mitotic spindle. How the highly dynamic microtubules (MTs) are maintained in stable mitotic spindles is a key question to be answered. Motor and non-motor microtubule associated proteins (MAPs) have been reported to stabilize the dynamic spindle through crosslinking adjacent MTs. Mars, a novel MAP, is essential for the early development of Drosophila embryos. Previous studies showed that Mars is required for maintaining an intact mitotic spindle but did not provide a molecular mechanism for this function. Here we show that Mars is able to stabilize the mitotic spindle in vivo. Both in vivo and in vitro data reveal that the N-terminal region of Mars functions in the stabilization of the mitotic spindle by crosslinking adjacent MTs.

  15. A novel Drosophila model of TDP-43 proteinopathies: N-terminal sequences combined with the Q/N domain induce protein functional loss and locomotion defects

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    Simona Langellotti

    2016-06-01

    Full Text Available Transactive response DNA-binding protein 43 kDa (TDP-43, also known as TBPH in Drosophila melanogaster and TARDBP in mammals is the main protein component of the pathological inclusions observed in neurons of patients affected by different neurodegenerative disorders, including amyotrophic lateral sclerosis (ALS and fronto-temporal lobar degeneration (FTLD. The number of studies investigating the molecular mechanisms underlying neurodegeneration is constantly growing; however, the role played by TDP-43 in disease onset and progression is still unclear. A fundamental shortcoming that hampers progress is the lack of animal models showing aggregation of TDP-43 without overexpression. In this manuscript, we have extended our cellular model of aggregation to a transgenic Drosophila line. Our fly model is not based on the overexpression of a wild-type TDP-43 transgene. By contrast, we engineered a construct that includes only the specific TDP-43 amino acid sequences necessary to trigger aggregate formation and capable of trapping endogenous Drosophila TDP-43 into a non-functional insoluble form. Importantly, the resulting recombinant product lacks functional RNA recognition motifs (RRMs and, thus, does not have specific TDP-43-physiological functions (i.e. splicing regulation ability that might affect the animal phenotype per se. This novel Drosophila model exhibits an evident degenerative phenotype with reduced lifespan and early locomotion defects. Additionally, we show that important proteins involved in neuromuscular junction function, such as syntaxin (SYX, decrease their levels as a consequence of TDP-43 loss of function implying that the degenerative phenotype is a consequence of TDP-43 sequestration into the aggregates. Our data lend further support to the role of TDP-43 loss-of-function in the pathogenesis of neurodegenerative disorders. The novel transgenic Drosophila model presented in this study will help to gain further insight into the

  16. Similar rates of protein adaptation in Drosophila miranda and D. melanogaster, two species with different current effective population sizes

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    Bachtrog Doris

    2008-12-01

    Full Text Available Abstract Background Adaptive protein evolution is common in several Drosophila species investigated. Some studies point to very weak selection operating on amino-acid mutations, with average selection intensities on the order of Nes ~ 5 in D. melanogaster and D. simulans. Species with lower effective population sizes should undergo less adaptation since they generate fewer mutations and selection is ineffective on a greater proportion of beneficial mutations. Results Here I study patterns of polymorphism and divergence at 91 X-linked loci in D. miranda, a species with a roughly 5-fold smaller effective population size than D. melanogaster. Surprisingly, I find a similar fraction of amino-acid mutations being driven to fixation by positive selection in D. miranda and D. melanogaster. Genes with higher rates of amino-acid evolution show lower levels of neutral diversity, a pattern predicted by recurrent adaptive protein evolution. I fit a hitchhiking model to patterns of polymorphism in D. miranda and D. melanogaster and estimate an order of magnitude higher selection coefficients for beneficial mutations in D. miranda. Conclusion This analysis suggests that effective population size may not be a major determinant in rates of protein adaptation. Instead, adaptation may not be mutation-limited, or the distribution of fitness effects for beneficial mutations might differ vastly between different species or populations. Alternative explanation such as biases in estimating the fraction of beneficial mutations or slightly deleterious mutation models are also discussed.

  17. A database and tool, IM Browser, for exploring and integrating emerging gene and protein interaction data for Drosophila

    Directory of Open Access Journals (Sweden)

    Parrish Jodi R

    2006-04-01

    Full Text Available Abstract Background Biological processes are mediated by networks of interacting genes and proteins. Efforts to map and understand these networks are resulting in the proliferation of interaction data derived from both experimental and computational techniques for a number of organisms. The volume of this data combined with the variety of specific forms it can take has created a need for comprehensive databases that include all of the available data sets, and for exploration tools to facilitate data integration and analysis. One powerful paradigm for the navigation and analysis of interaction data is an interaction graph or map that represents proteins or genes as nodes linked by interactions. Several programs have been developed for graphical representation and analysis of interaction data, yet there remains a need for alternative programs that can provide casual users with rapid easy access to many existing and emerging data sets. Description Here we describe a comprehensive database of Drosophila gene and protein interactions collected from a variety of sources, including low and high throughput screens, genetic interactions, and computational predictions. We also present a program for exploring multiple interaction data sets and for combining data from different sources. The program, referred to as the Interaction Map (IM Browser, is a web-based application for searching and visualizing interaction data stored in a relational database system. Use of the application requires no downloads and minimal user configuration or training, thereby enabling rapid initial access to interaction data. IM Browser was designed to readily accommodate and integrate new types of interaction data as it becomes available. Moreover, all information associated with interaction measurements or predictions and the genes or proteins involved are accessible to the user. This allows combined searches and analyses based on either common or technique-specific attributes

  18. Developmental shaping of dendritic arbors in Drosophila relies on tightly regulated intra-neuronal activity of protein kinase A (PKA).

    Science.gov (United States)

    Copf, Tijana

    2014-09-15

    Dendrites develop morphologies characterized by multiple levels of complexity that involve neuron type specific dendritic length and particular spatial distribution. How this is developmentally regulated and in particular which signaling molecules are crucial in the process is still not understood. Using Drosophila class IV dendritic arborization (da) neurons we test in vivo the effects of cell-autonomous dose-dependent changes in the activity levels of the cAMP-dependent Protein Kinase A (PKA) on the formation of complex dendritic arbors. We find that genetic manipulations of the PKA activity levels affect profoundly the arbor complexity with strongest impact on distal branches. Both decreasing and increasing PKA activity result in a reduced complexity of the arbors, as reflected in decreased dendritic length and number of branching points, suggesting an inverted U-shape response to PKA. The phenotypes are accompanied by changes in organelle distribution: Golgi outposts and early endosomes in distal dendritic branches are reduced in PKA mutants. By using Rab5 dominant negative we find that PKA interacts genetically with the early endosomal pathway. We test if the possible relationship between PKA and organelles may be the result of phosphorylation of the microtubule motor dynein components or Rab5. We find that Drosophila cytoplasmic dynein components are direct PKA phosphorylation targets in vitro, but not in vivo, thus pointing to a different putative in vivo target. Our data argue that tightly controlled dose-dependent intra-neuronal PKA activity levels are critical in determining the dendritic arbor complexity, one of the possible ways being through the regulation of organelle distribution. Copyright © 2014 Elsevier Inc. All rights reserved.

  19. The RNA-binding proteins FMR1, rasputin and caprin act together with the UBA protein lingerer to restrict tissue growth in Drosophila melanogaster.

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    Roland Baumgartner

    Full Text Available Appropriate expression of growth-regulatory genes is essential to ensure normal animal development and to prevent diseases like cancer. Gene regulation at the levels of transcription and translational initiation mediated by the Hippo and Insulin signaling pathways and by the TORC1 complex, respectively, has been well documented. Whether translational control mediated by RNA-binding proteins contributes to the regulation of cellular growth is less clear. Here, we identify Lingerer (Lig, an UBA domain-containing protein, as growth suppressor that associates with the RNA-binding proteins Fragile X mental retardation protein 1 (FMR1 and Caprin (Capr and directly interacts with and regulates the RNA-binding protein Rasputin (Rin in Drosophila melanogaster. lig mutant organs overgrow due to increased proliferation, and a reporter for the JAK/STAT signaling pathway is upregulated in a lig mutant situation. rin, Capr or FMR1 in combination as double mutants, but not the respective single mutants, display lig like phenotypes, implicating a redundant function of Rin, Capr and FMR1 in growth control in epithelial tissues. Thus, Lig regulates cell proliferation during development in concert with Rin, Capr and FMR1.

  20. The RNA-binding proteins FMR1, rasputin and caprin act together with the UBA protein lingerer to restrict tissue growth in Drosophila melanogaster.

    Science.gov (United States)

    Baumgartner, Roland; Stocker, Hugo; Hafen, Ernst

    2013-01-01

    Appropriate expression of growth-regulatory genes is essential to ensure normal animal development and to prevent diseases like cancer. Gene regulation at the levels of transcription and translational initiation mediated by the Hippo and Insulin signaling pathways and by the TORC1 complex, respectively, has been well documented. Whether translational control mediated by RNA-binding proteins contributes to the regulation of cellular growth is less clear. Here, we identify Lingerer (Lig), an UBA domain-containing protein, as growth suppressor that associates with the RNA-binding proteins Fragile X mental retardation protein 1 (FMR1) and Caprin (Capr) and directly interacts with and regulates the RNA-binding protein Rasputin (Rin) in Drosophila melanogaster. lig mutant organs overgrow due to increased proliferation, and a reporter for the JAK/STAT signaling pathway is upregulated in a lig mutant situation. rin, Capr or FMR1 in combination as double mutants, but not the respective single mutants, display lig like phenotypes, implicating a redundant function of Rin, Capr and FMR1 in growth control in epithelial tissues. Thus, Lig regulates cell proliferation during development in concert with Rin, Capr and FMR1.

  1. The full-length form of the Drosophila amyloid precursor protein is involved in memory formation.

    Science.gov (United States)

    Bourdet, Isabelle; Preat, Thomas; Goguel, Valérie

    2015-01-21

    The APP plays a central role in AD, a pathology that first manifests as a memory decline. Understanding the role of APP in normal cognition is fundamental in understanding the progression of AD, and mammalian studies have pointed to a role of secreted APPα in memory. In Drosophila, we recently showed that APPL, the fly APP ortholog, is required for associative memory. In the present study, we aimed to characterize which form of APPL is involved in this process. We show that expression of a secreted-APPL form in the mushroom bodies, the center for olfactory memory, is able to rescue the memory deficit caused by APPL partial loss of function. We next assessed the impact on memory of the Drosophila α-secretase kuzbanian (KUZ), the enzyme initiating the nonamyloidogenic pathway that produces secreted APPLα. Strikingly, KUZ overexpression not only failed to rescue the memory deficit caused by APPL loss of function, it exacerbated this deficit. We further show that in addition to an increase in secreted-APPL forms, KUZ overexpression caused a decrease of membrane-bound full-length species that could explain the memory deficit. Indeed, we observed that transient expression of a constitutive membrane-bound mutant APPL form is sufficient to rescue the memory deficit caused by APPL reduction, revealing for the first time a role of full-length APPL in memory formation. Our data demonstrate that, in addition to secreted APPL, the noncleaved form is involved in memory, raising the possibility that secreted and full-length APPL act together in memory processes. Copyright © 2015 the authors 0270-6474/15/351043-09$15.00/0.

  2. Pervasive adaptive protein evolution apparent in diversity patterns around amino acid substitutions in Drosophila simulans.

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    Shmuel Sattath

    2011-02-01

    Full Text Available In Drosophila, multiple lines of evidence converge in suggesting that beneficial substitutions to the genome may be common. All suffer from confounding factors, however, such that the interpretation of the evidence-in particular, conclusions about the rate and strength of beneficial substitutions-remains tentative. Here, we use genome-wide polymorphism data in D. simulans and sequenced genomes of its close relatives to construct a readily interpretable characterization of the effects of positive selection: the shape of average neutral diversity around amino acid substitutions. As expected under recurrent selective sweeps, we find a trough in diversity levels around amino acid but not around synonymous substitutions, a distinctive pattern that is not expected under alternative models. This characterization is richer than previous approaches, which relied on limited summaries of the data (e.g., the slope of a scatter plot, and relates to underlying selection parameters in a straightforward way, allowing us to make more reliable inferences about the prevalence and strength of adaptation. Specifically, we develop a coalescent-based model for the shape of the entire curve and use it to infer adaptive parameters by maximum likelihood. Our inference suggests that ∼13% of amino acid substitutions cause selective sweeps. Interestingly, it reveals two classes of beneficial fixations: a minority (approximately 3% that appears to have had large selective effects and accounts for most of the reduction in diversity, and the remaining 10%, which seem to have had very weak selective effects. These estimates therefore help to reconcile the apparent conflict among previously published estimates of the strength of selection. More generally, our findings provide unequivocal evidence for strongly beneficial substitutions in Drosophila and illustrate how the rapidly accumulating genome-wide data can be leveraged to address enduring questions about the genetic basis

  3. Pervasive adaptive protein evolution apparent in diversity patterns around amino acid substitutions in Drosophila simulans.

    Science.gov (United States)

    Sattath, Shmuel; Elyashiv, Eyal; Kolodny, Oren; Rinott, Yosef; Sella, Guy

    2011-02-10

    In Drosophila, multiple lines of evidence converge in suggesting that beneficial substitutions to the genome may be common. All suffer from confounding factors, however, such that the interpretation of the evidence-in particular, conclusions about the rate and strength of beneficial substitutions-remains tentative. Here, we use genome-wide polymorphism data in D. simulans and sequenced genomes of its close relatives to construct a readily interpretable characterization of the effects of positive selection: the shape of average neutral diversity around amino acid substitutions. As expected under recurrent selective sweeps, we find a trough in diversity levels around amino acid but not around synonymous substitutions, a distinctive pattern that is not expected under alternative models. This characterization is richer than previous approaches, which relied on limited summaries of the data (e.g., the slope of a scatter plot), and relates to underlying selection parameters in a straightforward way, allowing us to make more reliable inferences about the prevalence and strength of adaptation. Specifically, we develop a coalescent-based model for the shape of the entire curve and use it to infer adaptive parameters by maximum likelihood. Our inference suggests that ∼13% of amino acid substitutions cause selective sweeps. Interestingly, it reveals two classes of beneficial fixations: a minority (approximately 3%) that appears to have had large selective effects and accounts for most of the reduction in diversity, and the remaining 10%, which seem to have had very weak selective effects. These estimates therefore help to reconcile the apparent conflict among previously published estimates of the strength of selection. More generally, our findings provide unequivocal evidence for strongly beneficial substitutions in Drosophila and illustrate how the rapidly accumulating genome-wide data can be leveraged to address enduring questions about the genetic basis of adaptation.

  4. GPS Composite Clock Analysis

    OpenAIRE

    Wright, James R.

    2008-01-01

    The GPS composite clock defines GPS time, the timescale used today in GPS operations. GPS time is illuminated by examination of its role in the complete estimation and control problem relative to UTC/TAI. The phase of each GPS clock is unobservable from GPS pseudorange measurements, and the mean phase of the GPS clock ensemble (GPS time) is unobservable. A new and useful observability definition is presented, together with new observability theorems, to demonstrate explicitly that GPS time is...

  5. DISCO interacting protein 2 determines direction of axon projection under the regulation of c-Jun N-terminal kinase in the Drosophila mushroom body

    International Nuclear Information System (INIS)

    Nitta, Yohei; Sugie, Atsushi

    2017-01-01

    Precisely controlled axon guidance for complex neuronal wiring is essential for appropriate neuronal function. c-Jun N-terminal kinase (JNK) was found to play a role in axon guidance recently as well as in cell proliferation, protection and apoptosis. In spite of many genetic and molecular studies on these biological processes regulated by JNK, how JNK regulates axon guidance accurately has not been fully explained thus far. To address this question, we use the Drosophila mushroom body (MB) as a model since the α/β axons project in two distinct directions. Here we show that DISCO interacting protein 2 (DIP2) is required for the accurate direction of axonal guidance. DIP2 expression is under the regulation of Basket (Bsk), the Drosophila homologue of JNK. We additionally found that the Bsk/DIP2 pathway is independent from the AP-1 transcriptional factor complex pathway, which is directly activated by Bsk. In conclusion, our findings revealed DIP2 as a novel effector downstream of Bsk modulating the direction of axon projection. - Highlights: • DIP2 is required for accurate direction of axon guidance in Drosophila mushroom body. • DIP2 is a downstream of JNK in the axon guidance of Drosophila mushroom body neuron. • JNK/DIP2 pathway is independent from JNK/AP-1 transcriptional factor complex pathway.

  6. Loss of PTB or negative regulation of Notch mRNA reveals distinct zones of Notch and actin protein accumulation in Drosophila embryo.

    Directory of Open Access Journals (Sweden)

    Cedric S Wesley

    Full Text Available Polypyrimidine Tract Binding (PTB protein is a regulator of mRNA processing and translation. Genetic screens and studies of wing and bristle development during the post-embryonic stages of Drosophila suggest that it is a negative regulator of the Notch pathway. How PTB regulates the Notch pathway is unknown. Our studies of Drosophila embryogenesis indicate that (1 the Notch mRNA is a potential target of PTB, (2 PTB and Notch functions in the dorso-lateral regions of the Drosophila embryo are linked to actin regulation but not their functions in the ventral region, and (3 the actin-related Notch activity in the dorso-lateral regions might require a Notch activity at or near the cell surface that is different from the nuclear Notch activity involved in cell fate specification in the ventral region. These data raise the possibility that the Drosophila embryo is divided into zones of different PTB and Notch activities based on whether or not they are linked to actin regulation. They also provide clues to the almost forgotten role of Notch in cell adhesion and reveal a role for the Notch pathway in cell fusions.

  7. Loss of PTB or Negative Regulation of Notch mRNA Reveals Distinct Zones of Notch and Actin Protein Accumulation in Drosophila Embryo

    Science.gov (United States)

    Wesley, Cedric S.; Guo, Heng; Chaudhry, Kanita A.; Thali, Markus J.; Yin, Jerry C.; Clason, Todd; Wesley, Umadevi V.

    2011-01-01

    Polypyrimidine Tract Binding (PTB) protein is a regulator of mRNA processing and translation. Genetic screens and studies of wing and bristle development during the post-embryonic stages of Drosophila suggest that it is a negative regulator of the Notch pathway. How PTB regulates the Notch pathway is unknown. Our studies of Drosophila embryogenesis indicate that (1) the Notch mRNA is a potential target of PTB, (2) PTB and Notch functions in the dorso-lateral regions of the Drosophila embryo are linked to actin regulation but not their functions in the ventral region, and (3) the actin-related Notch activity in the dorso-lateral regions might require a Notch activity at or near the cell surface that is different from the nuclear Notch activity involved in cell fate specification in the ventral region. These data raise the possibility that the Drosophila embryo is divided into zones of different PTB and Notch activities based on whether or not they are linked to actin regulation. They also provide clues to the almost forgotten role of Notch in cell adhesion and reveal a role for the Notch pathway in cell fusions. PMID:21750738

  8. cGMP-Dependent Protein Kinase Inhibition Extends the Upper Temperature Limit of Stimulus-Evoked Calcium Responses in Motoneuronal Boutons of Drosophila melanogaster Larvae.

    Science.gov (United States)

    Krill, Jennifer L; Dawson-Scully, Ken

    2016-01-01

    While the mammalian brain functions within a very narrow range of oxygen concentrations and temperatures, the fruit fly, Drosophila melanogaster, has employed strategies to deal with a much wider range of acute environmental stressors. The foraging (for) gene encodes the cGMP-dependent protein kinase (PKG), has been shown to regulate thermotolerance in many stress-adapted species, including Drosophila, and could be a potential therapeutic target in the treatment of hyperthermia in mammals. Whereas previous thermotolerance studies have looked at the effects of PKG variation on Drosophila behavior or excitatory postsynaptic potentials at the neuromuscular junction (NMJ), little is known about PKG effects on presynaptic mechanisms. In this study, we characterize presynaptic calcium ([Ca2+]i) dynamics at the Drosophila larval NMJ to determine the effects of high temperature stress on synaptic transmission. We investigated the neuroprotective role of PKG modulation both genetically using RNA interference (RNAi), and pharmacologically, to determine if and how PKG affects presynaptic [Ca2+]i dynamics during hyperthermia. We found that PKG activity modulates presynaptic neuronal Ca2+ responses during acute hyperthermia, where PKG activation makes neurons more sensitive to temperature-induced failure of Ca2+ flux and PKG inhibition confers thermotolerance and maintains normal Ca2+ dynamics under the same conditions. Targeted motoneuronal knockdown of PKG using RNAi demonstrated that decreased PKG expression was sufficient to confer thermoprotection. These results demonstrate that the PKG pathway regulates presynaptic motoneuronal Ca2+ signaling to influence thermotolerance of presynaptic function during acute hyperthermia.

  9. Precision Clock Evaluation Facility

    Data.gov (United States)

    Federal Laboratory Consortium — FUNCTION: Tests and evaluates high-precision atomic clocks for spacecraft, ground, and mobile applications. Supports performance evaluation, environmental testing,...

  10. Drosophila TDP-43 RNA-Binding Protein Facilitates Association of Sister Chromatid Cohesion Proteins with Genes, Enhancers and Polycomb Response Elements.

    Directory of Open Access Journals (Sweden)

    Amanda Swain

    2016-09-01

    Full Text Available The cohesin protein complex mediates sister chromatid cohesion and participates in transcriptional control of genes that regulate growth and development. Substantial reduction of cohesin activity alters transcription of many genes without disrupting chromosome segregation. Drosophila Nipped-B protein loads cohesin onto chromosomes, and together Nipped-B and cohesin occupy essentially all active transcriptional enhancers and a large fraction of active genes. It is unknown why some active genes bind high levels of cohesin and some do not. Here we show that the TBPH and Lark RNA-binding proteins influence association of Nipped-B and cohesin with genes and gene regulatory sequences. In vitro, TBPH and Lark proteins specifically bind RNAs produced by genes occupied by Nipped-B and cohesin. By genomic chromatin immunoprecipitation these RNA-binding proteins also bind to chromosomes at cohesin-binding genes, enhancers, and Polycomb response elements (PREs. RNAi depletion reveals that TBPH facilitates association of Nipped-B and cohesin with genes and regulatory sequences. Lark reduces binding of Nipped-B and cohesin at many promoters and aids their association with several large enhancers. Conversely, Nipped-B facilitates TBPH and Lark association with genes and regulatory sequences, and interacts with TBPH and Lark in affinity chromatography and immunoprecipitation experiments. Blocking transcription does not ablate binding of Nipped-B and the RNA-binding proteins to chromosomes, indicating transcription is not required to maintain binding once established. These findings demonstrate that RNA-binding proteins help govern association of sister chromatid cohesion proteins with genes and enhancers.

  11. Characterization of the Drosophila group ortholog to the amino-terminus of the alpha-thalassemia and mental retardation X-Linked (ATRX vertebrate protein.

    Directory of Open Access Journals (Sweden)

    Brenda López-Falcón

    Full Text Available The human ATRX gene encodes hATRX, a chromatin-remodeling protein harboring an helicase/ATPase and ADD domains. The ADD domain has two zinc fingers that bind to histone tails and mediate hATRX binding to chromatin. dAtrx, the putative ATRX homolog in Drosophila melanogaster, has a conserved helicase/ATPase domain but lacks the ADD domain. A bioinformatic search of the Drosophila genome using the human ADD sequence allowed us to identify the CG8290 annotated gene, which encodes three ADD harboring- isoforms generated by alternative splicing. This Drosophila ADD domain is highly similar in structure and in the amino acids which mediate the histone tail contacts to the ADD domain of hATRX as shown by 3D modeling. Very recently the CG8290 annotated gene has been named dadd1. We show through pull-down and CoIP assays that the products of the dadd1 gene interact physically with dAtrxL and HP1a and all of them mainly co-localize in the chromocenter, although euchromatic localization can also be observed through the chromosome arms. We confirm through ChIP analyses that these proteins are present in vivo in the same heterochromatic regions. The three isoforms are expressed throughout development. Flies carrying transheterozygous combinations of the dadd1 and atrx alleles are semi-viable and have different phenotypes including the appearance of melanotic masses. Interestingly, the dAdd1-b and c isoforms have extra domains, such as MADF, which suggest newly acquired functions of these proteins. These results strongly support that, in Drosophila, the atrx gene diverged and that the dadd1-encoded proteins participate with dAtrx in some cellular functions such as heterochromatin maintenance.

  12. Preliminary small-angle X-ray scattering and X-ray diffraction studies of the BTB domain of lola protein from Drosophila melanogaster

    Science.gov (United States)

    Boyko, K. M.; Nikolaeva, A. Yu.; Kachalova, G. S.; Bonchuk, A. N.; Dorovatovskii, P. V.; Popov, V. O.

    2017-11-01

    The Drosophila genome has several dozens of transcription factors (TTK group) containing BTB domains assembled into octamers. The LOLA protein belongs to this family. The purification, crystallization, and preliminary X-ray diffraction and small-angle X-ray scattering (SAXS) studies of the BTB domain of this protein are reported. The crystallization conditions were found by the vapor-diffusion technique. A very low diffraction resolution (8.7 Å resolution) of the crystals was insufficient for the determination of the threedimensional structure of the BTB domain. The SAXS study demonstrated that the BTB domain of the LOLA protein exists as an octamer in solution.

  13. The conserved, disease-associated RNA binding protein dNab2 interacts with the Fragile-X protein ortholog in Drosophila neurons

    Science.gov (United States)

    Bienkowski, Rick S.; Banerjee, Ayan; Rounds, J. Christopher; Rha, Jennifer; Omotade, Omotola F.; Gross, Christina; Morris, Kevin J.; Leung, Sara W.; Pak, ChangHui; Jones, Stephanie K.; Santoro, Michael R.; Warren, Stephen T.; Zheng, James Q.; Bassell, Gary J.; Corbett, Anita H.; Moberg, Kenneth H.

    2017-01-01

    Summary The Drosophila dNab2 protein is an ortholog of human ZC3H14, a poly(A) RNA-binding protein required for intellectual function. dNab2 supports memory and axon projection, but its molecular role in neurons is undefined. Here we present a network of interactions that links dNab2 to cytoplasmic control of neuronal mRNAs in conjunction with and the Fragile-X protein ortholog dFMRP. dNab2 and dfmr1 interact genetically in control of neurodevelopment and olfactory memory and their encoded proteins co-localize in puncta within neuronal processes. dNab2 regulates CaMKII but not futsch mRNA, implying a selective role in control of dFMRP-bound transcripts. Reciprocally, dFMRP and vertebrate FMRP restrict mRNA poly(A)-tail length similar to dNab2/ZC3H14. Parallel studies of murine hippocampal neurons indicate that ZC3H14 is also a cytoplasmic regulator of neuronal mRNAs. In sum these findings suggest that dNab2 represses expression of a subset of dFMRP-target mRNAs, which could underlie brain-specific defects in patients lacking ZC3H14. PMID:28793261

  14. The Conserved, Disease-Associated RNA Binding Protein dNab2 Interacts with the Fragile X Protein Ortholog in Drosophila Neurons

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    Rick S. Bienkowski

    2017-08-01

    Full Text Available The Drosophila dNab2 protein is an ortholog of human ZC3H14, a poly(A RNA binding protein required for intellectual function. dNab2 supports memory and axon projection, but its molecular role in neurons is undefined. Here, we present a network of interactions that links dNab2 to cytoplasmic control of neuronal mRNAs in conjunction with the fragile X protein ortholog dFMRP. dNab2 and dfmr1 interact genetically in control of neurodevelopment and olfactory memory, and their encoded proteins co-localize in puncta within neuronal processes. dNab2 regulates CaMKII, but not futsch, implying a selective role in control of dFMRP-bound transcripts. Reciprocally, dFMRP and vertebrate FMRP restrict mRNA poly(A tail length, similar to dNab2/ZC3H14. Parallel studies of murine hippocampal neurons indicate that ZC3H14 is also a cytoplasmic regulator of neuronal mRNAs. Altogether, these findings suggest that dNab2 represses expression of a subset of dFMRP-target mRNAs, which could underlie brain-specific defects in patients lacking ZC3H14.

  15. Targeted mutagenesis of the Sap47 gene of Drosophila: Flies lacking the synapse associated protein of 47 kDa are viable and fertile

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    Huber Saskia

    2004-04-01

    Full Text Available Abstract Background Conserved proteins preferentially expressed in synaptic terminals of the nervous system are likely to play a significant role in brain function. We have previously identified and molecularly characterized the Sap47 gene which codes for a novel synapse associated protein of 47 kDa in Drosophila. Sequence comparison identifies homologous proteins in numerous species including C. elegans, fish, mouse and human. First hints as to the function of this novel protein family can be obtained by generating mutants for the Sap47 gene in Drosophila. Results Attempts to eliminate the Sap47 gene through targeted mutagenesis by homologous recombination were unsuccessful. However, several mutants were generated by transposon remobilization after an appropriate insertion line had become available from the Drosophila P-element screen of the Bellen/Hoskins/Rubin/Spradling labs. Characterization of various deletions in the Sap47 gene due to imprecise excision of the P-element identified three null mutants and three hypomorphic mutants. Null mutants are viable and fertile and show no gross structural or obvious behavioural deficits. For cell-specific over-expression and "rescue" of the knock-out flies a transgenic line was generated which expresses the most abundant transcript under the control of the yeast enhancer UAS. In addition, knock-down of the Sap47 gene was achieved by generating 31 transgenic lines expressing Sap47 RNAi constructs, again under UAS control. When driven by a ubiquitously expressed yeast transcription factor (GAL4, Sap47 gene suppression in several of these lines is highly efficient resulting in residual SAP47 protein concentrations in heads as low as 6% of wild type levels. Conclusion The conserved synaptic protein SAP47 of Drosophila is not essential for basic synaptic function. The Sap47 gene region may be refractory to targeted mutagenesis by homologous recombination. RNAi using a construct linking genomic DNA to anti

  16. Polycomb group (PcG) proteins and Pax6 cooperate to inhibit in vivo reprogramming of the developing Drosophila eye.

    Science.gov (United States)

    Zhu, Jinjin; Ordway, Alison J; Weber, Lena; Buddika, Kasun; Kumar, Justin P

    2018-04-04

    How different cells and tissues commit to and determine their fates has been a central question in developmental biology since the seminal embryological experiments conducted by Wilhelm Roux and Hans Driesch in sea urchins and frogs. Here, we demonstrate that Polycomb group (PcG) proteins maintain Drosophila eye specification by suppressing the activation of alternative fate choices. The loss of PcG in the developing eye results in a cellular reprogramming event in which the eye is redirected to a wing fate. This fate transformation occurs with either the individual loss of Polycomb proteins or the simultaneous reduction of the Pleiohomeotic repressive complex and Pax6. Interestingly, the requirement for retinal selector genes is limited to Pax6, as the removal of more downstream members does not lead to the eye-wing transformation. We also show that distinct PcG complexes are required during different developmental windows throughout eye formation. These findings build on earlier observations that the eye can be reprogrammed to initiate head epidermis, antennal and leg development. © 2018. Published by The Company of Biologists Ltd.

  17. Regulation of the Drosophila Enhancer of split and invected-engrailed gene complexes by sister chromatid cohesion proteins.

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    Cheri A Schaaf

    2009-07-01

    Full Text Available The cohesin protein complex was first recognized for holding sister chromatids together and ensuring proper chromosome segregation. Cohesin also regulates gene expression, but the mechanisms are unknown. Cohesin associates preferentially with active genes, and is generally absent from regions in which histone H3 is methylated by the Enhancer of zeste [E(z] Polycomb group silencing protein. Here we show that transcription is hypersensitive to cohesin levels in two exceptional cases where cohesin and the E(z-mediated histone methylation simultaneously coat the entire Enhancer of split and invected-engrailed gene complexes in cells derived from Drosophila central nervous system. These gene complexes are modestly transcribed, and produce seven of the twelve transcripts that increase the most with cohesin knockdown genome-wide. Cohesin mutations alter eye development in the same manner as increased Enhancer of split activity, suggesting that similar regulation occurs in vivo. We propose that cohesin helps restrain transcription of these gene complexes, and that deregulation of similarly cohesin-hypersensitive genes may underlie developmental deficits in Cornelia de Lange syndrome.

  18. Drosophila melanogaster cellular repressor of E1A-stimulated genes is a lysosomal protein essential for fly development.

    Science.gov (United States)

    Kowalewski-Nimmerfall, Elisabeth; Schähs, Philipp; Maresch, Daniel; Rendic, Dubravko; Krämer, Helmut; Mach, Lukas

    2014-12-01

    Mammalian cellular repressor of E1A-stimulated genes is a lysosomal glycoprotein implicated in cellular growth and differentiation. The genome of the fruit fly Drosophila melanogaster encodes a putative orthologue (dCREG), suggesting evolutionarily conserved physiological functions of this protein. In D. melanogaster S2 cells, dCREG was found to localize in lysosomes. Further studies revealed that intracellular dCREG is subject of proteolytic maturation. Processing and turnover could be substantially reduced by RNAi-mediated silencing of cathepsin L. In contrast to mammalian cells, lysosomal delivery of dCREG does not depend on its carbohydrate moiety. Furthermore, depletion of the putative D. melanogaster lysosomal sorting receptor lysosomal enzyme receptor protein did not compromise cellular retention of dCREG. We also investigated the developmental consequences of dCREG ablation in whole D. melanogaster flies. Ubiquitous depletion of dCREG proved lethal at the late pupal stage once a knock-down efficiency of >95% was achieved. These results demonstrate that dCREG is essential for proper completion of fly development. Copyright © 2014. Published by Elsevier B.V.

  19. Yolk proteins in the male reproductive system of the fruit fly Drosophila melanogaster: spatial and temporal patterns of expression.

    Science.gov (United States)

    Majewska, Magdalena M; Suszczynska, Agnieszka; Kotwica-Rolinska, Joanna; Czerwik, Tomasz; Paterczyk, Bohdan; Polanska, Marta A; Bernatowicz, Piotr; Bebas, Piotr

    2014-04-01

    In insects, spermatozoa develop in the testes as clones of single spermatogonia covered by specialized somatic cyst cells (cc). Upon completion of spermatogenesis, spermatozoa are released to the vas deferens, while the cc remain in the testes and die. In the fruit fly Drosophila melanogaster, the released spermatozoa first reach the seminal vesicles (SV), the organ where post-testicular maturation begins. Here, we demonstrate the temporal (restricted to the evening and early night hours) accumulation of membranous vesicles containing proteins in the SV lumen of D. melanogaster. When SV vesicles were isolated from the semen and co-incubated with testis-derived spermatozoa in vitro, their contents bound to the spermatozoa along their tails. The proteins of the SV vesicles were then characterized using 2-D electrophoresis. We identified a prominent protein spot of around 45-47 kDa, which disappears from the SV vesicles in the night, i.e. shortly after they appear in the SV lumen. Sequencing of peptides derived from this spot by mass spectrometry revealed identity with three yolk proteins (YP1-3). This unexpected result was confirmed by western blotting, which demonstrated that SV vesicles contain proteins that are immunoreactive with an antibody against D. melanogaster YP1-3. The expression of all yp genes was shown to be a unique feature of testis tissues. Using RNA probes we found that their transcripts localize exclusively to the cc that cover fully developed spermatozoa in the distal part of each testis. Temporally, the expression of yp genes was found to be restricted to a short period during the day and is followed by the evening accumulation of YP proteins in the cc. Immunohistochemical staining confirmed that cc are the source of SV vesicles containing YPs that are released into the SV lumen. These vesicles interact with spermatozoa and as a result, YPs become extrinsic proteins of the sperm membrane. Thus, we describe for the first time the expression of

  20. Dephosphorylation of the Core Clock Protein KaiC in the Cyanobacterial KaiABC Circadian Oscillator Proceeds via an ATP Synthase Mechanism

    Energy Technology Data Exchange (ETDEWEB)

    Egli, Martin; Mori, Tetsuya; Pattanayek, Rekha; Xu, Yao; Qin, Ximing; Johnson, Carl H. (Vanderbilt)

    2014-10-02

    The circadian clock of the cyanobacterium Synechococcus elongatus can be reconstituted in vitro from three proteins, KaiA, KaiB, and KaiC in the presence of ATP, to tick in a temperature-compensated manner. KaiC, the central cog of this oscillator, forms a homohexamer with 12 ATP molecules bound between its N- and C-terminal domains and exhibits unusual properties. Both the N-terminal (CI) and C-terminal (CII) domains harbor ATPase activity, and the subunit interfaces between CII domains are the sites of autokinase and autophosphatase activities. Hydrolysis of ATP correlates with phosphorylation at threonine and serine sites across subunits in an orchestrated manner, such that first T432 and then S431 are phosphorylated, followed by dephosphorylation of these residues in the same order. Although structural work has provided insight into the mechanisms of ATPase and kinase, the location and mechanism of the phosphatase have remained enigmatic. From the available experimental data based on a range of approaches, including KaiC crystal structures and small-angle X-ray scattering models, metal ion dependence, site-directed mutagenesis (i.e., E318, the general base), and measurements of the associated clock periods, phosphorylation patterns, and dephosphorylation courses as well as a lack of sequence motifs in KaiC that are typically associated with known phosphatases, we hypothesized that KaiCII makes use of the same active site for phosphorylation and dephosphorlyation. We observed that wild-type KaiC (wt-KaiC) exhibits an ATP synthase activity that is significantly reduced in the T432A/S431A mutant. We interpret the first observation as evidence that KaiCII is a phosphotransferase instead of a phosphatase and the second that the enzyme is capable of generating ATP, both from ADP and P{sub i} (in a reversal of the ATPase reaction) and from ADP and P-T432/P-S431 (dephosphorylation). This new concept regarding the mechanism of dephosphorylation is also supported by the

  1. Mining for novel candidate clock genes in the circadian regulatory network

    OpenAIRE

    Bhargava, Anuprabha; Herzel, Hanspeter; Ananthasubramaniam, Bharath

    2015-01-01

    Background Most physiological processes in mammals are temporally regulated by means of a master circadian clock in the brain and peripheral oscillators in most other tissues. A transcriptional-translation feedback network of clock genes produces near 24 h oscillations in clock gene and protein expression. Here, we aim to identify novel additions to the clock network using a meta-analysis of public chromatin immunoprecipitation sequencing (ChIP-seq), proteomics and protein-protein interaction...

  2. The GTP- and Phospholipid-Binding Protein TTD14 Regulates Trafficking of the TRPL Ion Channel in Drosophila Photoreceptor Cells

    Science.gov (United States)

    Cerny, Alexander C.; Altendorfer, André; Schopf, Krystina; Baltner, Karla; Maag, Nathalie; Sehn, Elisabeth; Wolfrum, Uwe; Huber, Armin

    2015-01-01

    Recycling of signaling proteins is a common phenomenon in diverse signaling pathways. In photoreceptors of Drosophila, light absorption by rhodopsin triggers a phospholipase Cβ-mediated opening of the ion channels transient receptor potential (TRP) and TRP-like (TRPL) and generates the visual response. The signaling proteins are located in a plasma membrane compartment called rhabdomere. The major rhodopsin (Rh1) and TRP are predominantly localized in the rhabdomere in light and darkness. In contrast, TRPL translocates between the rhabdomeral plasma membrane in the dark and a storage compartment in the cell body in the light, from where it can be recycled to the plasma membrane upon subsequent dark adaptation. Here, we identified the gene mutated in trpl translocation defective 14 (ttd14), which is required for both TRPL internalization from the rhabdomere in the light and recycling of TRPL back to the rhabdomere in the dark. TTD14 is highly conserved in invertebrates and binds GTP in vitro. The ttd14 mutation alters a conserved proline residue (P75L) in the GTP-binding domain and abolishes binding to GTP. This indicates that GTP binding is essential for TTD14 function. TTD14 is a cytosolic protein and binds to PtdIns(3)P, a lipid enriched in early endosome membranes, and to phosphatidic acid. In contrast to TRPL, rhabdomeral localization of the membrane proteins Rh1 and TRP is not affected in the ttd14 P75L mutant. The ttd14 P75L mutation results in Rh1-independent photoreceptor degeneration and larval lethality suggesting that other processes are also affected by the ttd14 P75L mutation. In conclusion, TTD14 is a novel regulator of TRPL trafficking, involved in internalization and subsequent sorting of TRPL into the recycling pathway that enables this ion channel to return to the plasma membrane. PMID:26509977

  3. The GTP- and Phospholipid-Binding Protein TTD14 Regulates Trafficking of the TRPL Ion Channel in Drosophila Photoreceptor Cells.

    Directory of Open Access Journals (Sweden)

    Alexander C Cerny

    2015-10-01

    Full Text Available Recycling of signaling proteins is a common phenomenon in diverse signaling pathways. In photoreceptors of Drosophila, light absorption by rhodopsin triggers a phospholipase Cβ-mediated opening of the ion channels transient receptor potential (TRP and TRP-like (TRPL and generates the visual response. The signaling proteins are located in a plasma membrane compartment called rhabdomere. The major rhodopsin (Rh1 and TRP are predominantly localized in the rhabdomere in light and darkness. In contrast, TRPL translocates between the rhabdomeral plasma membrane in the dark and a storage compartment in the cell body in the light, from where it can be recycled to the plasma membrane upon subsequent dark adaptation. Here, we identified the gene mutated in trpl translocation defective 14 (ttd14, which is required for both TRPL internalization from the rhabdomere in the light and recycling of TRPL back to the rhabdomere in the dark. TTD14 is highly conserved in invertebrates and binds GTP in vitro. The ttd14 mutation alters a conserved proline residue (P75L in the GTP-binding domain and abolishes binding to GTP. This indicates that GTP binding is essential for TTD14 function. TTD14 is a cytosolic protein and binds to PtdIns(3P, a lipid enriched in early endosome membranes, and to phosphatidic acid. In contrast to TRPL, rhabdomeral localization of the membrane proteins Rh1 and TRP is not affected in the ttd14P75L mutant. The ttd14P75L mutation results in Rh1-independent photoreceptor degeneration and larval lethality suggesting that other processes are also affected by the ttd14P75L mutation. In conclusion, TTD14 is a novel regulator of TRPL trafficking, involved in internalization and subsequent sorting of TRPL into the recycling pathway that enables this ion channel to return to the plasma membrane.

  4. Sex-lethal enables germline stem cell differentiation by down-regulating Nanos protein levels during Drosophila oogenesis.

    Science.gov (United States)

    Chau, Johnnie; Kulnane, Laura Shapiro; Salz, Helen K

    2012-06-12

    Drosophila ovarian germ cells require Sex-lethal (Sxl) to exit from the stem cell state and to enter the differentiation pathway. Sxl encodes a female-specific RNA binding protein and in somatic cells serves as the developmental switch gene for somatic sex determination and X-chromosome dosage compensation. None of the known Sxl target genes are required for germline differentiation, leaving open the question of how Sxl promotes the transition from stem cell to committed daughter cell. We address the mechanism by which Sxl regulates this transition through the identification of nanos as one of its target genes. Previous studies have shown that Nanos protein is necessary for GSC self-renewal and is rapidly down-regulated in the daughter cells fated to differentiate in the adult ovary. We find that this dynamic expression pattern is limited to female germ cells and is under Sxl control. In the absence of Sxl, or in male germ cells, Nanos protein is continuously expressed. Furthermore, this female-specific expression pattern is dependent on the presence of canonical Sxl binding sites located in the nanos 3' untranslated region. These results, combined with the observation that nanos RNA associates with the Sxl protein in ovarian extracts and loss and gain of function studies, suggest that Sxl enables the switch from germline stem cell to committed daughter cell by posttranscriptional down-regulation of nanos expression. These findings connect sexual identity to the stem cell self-renewal/differentiation decision and highlight the importance of posttranscriptional gene regulatory networks in controlling stem cell behavior.

  5. Crystallization and preliminary X-ray diffraction studies of Drosophila melanogaster Gαo-subunit of heterotrimeric G protein in complex with the RGS domain of CG5036

    International Nuclear Information System (INIS)

    Tishchenko, Svetlana; Gabdulkhakov, Azat; Tin, Uliana; Kostareva, Olga; Lin, Chen; Katanaev, Vladimir L.

    2012-01-01

    D. melanogaster Gαo-subunit and the RGS domain of its interacting partner CG5036 have been overproduced and purified; the crystallization and preliminary X-ray crystallographic analysis of the complex of the two proteins are reported. Regulator of G-protein signalling (RGS) proteins negatively regulate heterotrimeric G-protein signalling through their conserved RGS domains. RGS domains act as GTPase-activating proteins, accelerating the GTP hydrolysis rate of the activated form of Gα-subunits. Although omnipresent in eukaryotes, RGS proteins have not been adequately analysed in non-mammalian organisms. The Drosophila melanogaster Gαo-subunit and the RGS domain of its interacting partner CG5036 have been overproduced and purified; the crystallization of the complex of the two proteins using PEG 4000 as a crystallizing agent and preliminary X-ray crystallographic analysis are reported. Diffraction data were collected to 2.0 Å resolution using a synchrotron-radiation source

  6. Egyptian "Star Clocks"

    Science.gov (United States)

    Symons, Sarah

    Diagonal, transit, and Ramesside star clocks are tables of astronomical information occasionally found in ancient Egyptian temples, tombs, and papyri. The tables represent the motions of selected stars (decans and hour stars) throughout the Egyptian civil year. Analysis of star clocks leads to greater understanding of ancient Egyptian constellations, ritual astronomical activities, observational practices, and pharaonic chronology.

  7. Biological Clocks & Circadian Rhythms

    Science.gov (United States)

    Robertson, Laura; Jones, M. Gail

    2009-01-01

    The study of biological clocks and circadian rhythms is an excellent way to address the inquiry strand in the National Science Education Standards (NSES) (NRC 1996). Students can study these everyday phenomena by designing experiments, gathering and analyzing data, and generating new experiments. As students explore biological clocks and circadian…

  8. Identification of ER Proteins Involved in the Functional Organisation of the Early Secretory Pathway in Drosophila Cells by a Targeted RNAi Screen

    Science.gov (United States)

    Kondylis, Vangelis; Tang, Yang; Fuchs, Florian; Boutros, Michael; Rabouille, Catherine

    2011-01-01

    Background In Drosophila, the early secretory apparatus comprises discrete paired Golgi stacks in close proximity to exit sites from the endoplasmic reticulum (tER sites), thus forming tER-Golgi units. Although many components involved in secretion have been identified, the structural components sustaining its organisation are less known. Here we set out to identify novel ER resident proteins involved in the of tER-Golgi unit organisation. Results To do so, we designed a novel screening strategy combining a bioinformatics pre-selection with an RNAi screen. We first selected 156 proteins exhibiting known or related ER retention/retrieval signals from a list of proteins predicted to have a signal sequence. We then performed a microscopy-based primary and confirmation RNAi screen in Drosophila S2 cells directly scoring the organisation of the tER-Golgi units. We identified 49 hits, most of which leading to an increased number of smaller tER-Golgi units (MG for “more and smaller Golgi”) upon depletion. 16 of them were validated and characterised, showing that this phenotype was not due to an inhibition in secretion, a block in G2, or ER stress. Interestingly, the MG phenotype was often accompanied by an increase in the cell volume. Out of 6 proteins, 4 were localised to the ER. Conclusions This work has identified novel proteins involved in the organisation of the Drosophila early secretory pathway. It contributes to the effort of assigning protein functions to gene annotation in the secretory pathway, and analysis of the MG hits revealed an enrichment of ER proteins. These results suggest a link between ER localisation, aspects of cell metabolism and tER-Golgi structural organisation. PMID:21383842

  9. Regulation of axonal development by the nuclear protein hindsight (pebbled) in the Drosophila visual system.

    Science.gov (United States)

    Oliva, Carlos; Sierralta, Jimena

    2010-08-15

    The molecules and networks involved in the process of acquisition and maintenance of the form of a mature neuron are not completely known. Using a misexpression screen we identified the gene hindsight as a gene involved in the process of acquisition of the neuronal morphogenesis in the Drosophila adult nervous system. hindsight encodes a transcription factor known for its role in early developmental processes such as embryonic germ band retraction and dorsal closure, as well as in the establishment of cell morphology, planar cell polarity, and epithelial integrity during retinal development. We describe here a novel function for HNT by showing that both loss and gain of function of HNT affects the pathfinding of the photoreceptors axons. By manipulating the timing and level of HNT expression, together with the number of cells manipulated we show here that the function of HNT in axonal guidance is independent of the HNT functions previously reported in retinal cells. Based on genetic interaction experiments we show that part of HNT function in axonal development is exerted through the regulation of genes involved in the dynamics of the actin cytoskeleton. Copyright 2010 Elsevier Inc. All rights reserved.

  10. Changes in protein synthetic activity in early Drosophila embryos mutant for the segmentation gene Krueppel

    International Nuclear Information System (INIS)

    Bedian, V.; Summers, M.C.; Kauffman, S.A.

    1988-01-01

    We have identified early embryo proteins related to the segmentation gene Krueppel by [35S]methionine pulse labelling and two-dimensional gel electrophoresis. Protein synthesis differences shared by homozygous embryos of two Krueppel alleles when compared to heterozygous and wild-type embryos are reported. The study was extended to syncytial blastoderm stages by pulse labelling and gel analysis of single embryos, using Krueppel-specific proteins from gastrula stages as molecular markers for identifying homozygous Krueppel embryos. Localized expression of interesting proteins was examined in embryo fragments. The earliest differences detected at nuclear migration stages showed unregulated synthesis in mutant embryos of two proteins that have stage specific synthesis in normal embryos. At the cellular blastoderm stage one protein was not synthesized and two proteins showed apparent shifts in isoelectric point in mutant embryos. Differences observed in older embryos included additional proteins with shifted isoelectric points and a number of qualitative and quantitative changes in protein synthesis. Five of the proteins with altered rates of synthesis in mutant embryos showed localized synthesis in normal embryos. The early effects observed are consistent with the hypothesis that the Krueppel product can be a negative or positive regulator of expression of other loci, while blastoderm and gastrula stage shifts in isoelectric point indicate that a secondary effect of Krueppel function may involve post-translational modification of proteins

  11. Mechanical Control of Whole Body Shape by a Single Cuticular Protein Obstructor-E in Drosophila melanogaster.

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    Reiko Tajiri

    2017-01-01

    Full Text Available Body shapes are much more variable than body plans. One way to alter body shapes independently of body plans would be to mechanically deform bodies. To what extent body shapes are regulated physically, or molecules involved in physical control of morphogenesis, remain elusive. During fly metamorphosis, the cuticle (exoskeleton covering the larval body contracts longitudinally and expands laterally to become the ellipsoidal pupal case (puparium. Here we show that Drosophila melanogaster Obstructor-E (Obst-E is a protein constituent of the larval cuticle that confers the oriented contractility/expandability. In the absence of obst-E function, the larval cuticle fails to undergo metamorphic shape change and finally becomes a twiggy puparium. We present results indicating that Obst-E regulates the arrangement of chitin, a long-chain polysaccharide and a central component of the insect cuticle, and directs the formation of supracellular ridges on the larval cuticle. We further show that Obst-E is locally required for the oriented shape change of the cuticle during metamorphosis, which is associated with changes in the morphology of those ridges. Thus, Obst-E dramatically affects the body shape in a direct, physical manner by controlling the mechanical property of the exoskeleton.

  12. Fasting, but Not Aging, Dramatically Alters the Redox Status of Cysteine Residues on Proteins in Drosophila melanogaster

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    Katja E. Menger

    2015-06-01

    Full Text Available Altering the redox state of cysteine residues on protein surfaces is an important response to environmental challenges. Although aging and fasting alter many redox processes, the role of cysteine residues is uncertain. To address this, we used a redox proteomic technique, oxidative isotope-coded affinity tags (OxICAT, to assess cysteine-residue redox changes in Drosophila melanogaster during aging and fasting. This approach enabled us to simultaneously identify and quantify the redox state of several hundred cysteine residues in vivo. Cysteine residues within young flies had a bimodal distribution with peaks at ∼10% and ∼85% reversibly oxidized. Surprisingly, these cysteine residues did not become more oxidized with age. In contrast, 24 hr of fasting dramatically oxidized cysteine residues that were reduced under fed conditions while also reducing cysteine residues that were initially oxidized. We conclude that fasting, but not aging, dramatically alters cysteine-residue redox status in D. melanogaster.

  13. Glue protein production can be triggered by steroid hormone signaling independent of the developmental program in Drosophila melanogaster.

    Science.gov (United States)

    Kaieda, Yuya; Masuda, Ryota; Nishida, Ritsuo; Shimell, MaryJane; O'Connor, Michael B; Ono, Hajime

    2017-10-01

    Steroid hormones regulate life stage transitions, allowing animals to appropriately follow a developmental timeline. During insect development, the steroid hormone ecdysone is synthesized and released in a regulated manner by the prothoracic gland (PG) and then hydroxylated to the active molting hormone, 20-hydroxyecdysone (20E), in peripheral tissues. We manipulated ecdysteroid titers, through temporally controlled over-expression of the ecdysteroid-inactivating enzyme, CYP18A1, in the PG using the GeneSwitch-GAL4 system in the fruit fly Drosophila melanogaster. We monitored expression of a 20E-inducible glue protein gene, Salivary gland secretion 3 (Sgs3), using a Sgs3:GFP fusion transgene. In wild type larvae, Sgs3-GFP expression is activated at the midpoint of the third larval instar stage in response to the rising endogenous level of 20E. By first knocking down endogenous 20E levels during larval development and then feeding 20E to these larvae at various stages, we found that Sgs3-GFP expression could be triggered at an inappropriate developmental stage after a certain time lag. This stage-precocious activation of Sgs3 required expression of the Broad-complex, similar to normal Sgs3 developmental regulation, and a small level of nutritional input. We suggest that these studies provide evidence for a tissue-autonomic regulatory system for a metamorphic event independent from the primary 20E driven developmental progression. Copyright © 2017 Elsevier Inc. All rights reserved.

  14. Dystroglycan and mitochondrial ribosomal protein L34 regulate differentiation in the Drosophila eye.

    Directory of Open Access Journals (Sweden)

    Yougen Zhan

    2010-05-01

    Full Text Available Mutations that diminish the function of the extracellular matrix receptor Dystroglycan (DG result in muscular dystrophies, with associated neuronal migration defects in the brain and mental retardation e.g. Muscle Eye Brain Disease. To gain insight into the function of DG in the nervous system we initiated a study to examine its contribution to development of the eye of Drosophila melanogaster. Immuno-histochemistry showed that DG is concentrated on the apical surface of photoreceptors (R cells during specification of cell-fate in the third instar larva and is maintained at this location through early pupal stages. In point mutations that are null for DG we see abortive R cell elongation during differentiation that first appears in the pupa and results in stunted R cells in the adult. Overexpression of DG in R cells results in a small but significant increase in their size. R cell differentiation defects appear at the same stage in a deficiency line Df(2RDg(248 that affects Dg and the neighboring mitochondrial ribosomal gene, mRpL34. In the adult, these flies have severely disrupted R cells as well as defects in the lens and ommatidia. Expression of an mRpL34 transgene rescues much of this phenotype. We conclude that DG does not affect neuronal commitment but functions R cell autonomously to regulate neuronal elongation during differentiation in the pupa. We discuss these findings in view of recent work implicating DG as a regulator of cell metabolism and its genetic interaction with mRpL34, a member of a class of mitochondrial genes essential for normal metabolic function.

  15. Drosophila TIM binds importin α1, and acts as an adapter to transport PER to the nucleus.

    Science.gov (United States)

    Jang, A Reum; Moravcevic, Katarina; Saez, Lino; Young, Michael W; Sehgal, Amita

    2015-02-01

    Regulated nuclear entry of clock proteins is a conserved feature of eukaryotic circadian clocks and serves to separate the phase of mRNA activation from mRNA repression in the molecular feedback loop. In Drosophila, nuclear entry of the clock proteins, PERIOD (PER) and TIMELESS (TIM), is tightly controlled, and impairments of this process produce profound behavioral phenotypes. We report here that nuclear entry of PER-TIM in clock cells, and consequently behavioral rhythms, require a specific member of a classic nuclear import pathway, Importin α1 (IMPα1). In addition to IMPα1, rhythmic behavior and nuclear expression of PER-TIM require a specific nuclear pore protein, Nup153, and Ran-GTPase. IMPα1 can also drive rapid and efficient nuclear expression of TIM and PER in cultured cells, although the effect on PER is mediated by TIM. Mapping of interaction domains between IMPα1 and TIM/PER suggests that TIM is the primary cargo for the importin machinery. This is supported by attenuated interaction of IMPα1 with TIM carrying a mutation previously shown to prevent nuclear entry of TIM and PER. TIM is detected at the nuclear envelope, and computational modeling suggests that it contains HEAT-ARM repeats typically found in karyopherins, consistent with its role as a co-transporter for PER. These findings suggest that although PER is the major timekeeper of the clock, TIM is the primary target of nuclear import mechanisms. Thus, the circadian clock uses specific components of the importin pathway with a novel twist in that TIM serves a karyopherin-like role for PER.

  16. BMAA neurotoxicity in Drosophila.

    Science.gov (United States)

    Zhou, Xianchong; Escala, Wilfredo; Papapetropoulos, Spyridon; Bradley, Walter G; Zhai, R Grace

    2009-01-01

    We report the establishment of an in vivo model using the fruit fly Drosophila melanogaster to investigate the toxic effects of L-BMAA. We found that dietary intake of BMAA reduced the lifespan as well as the neurological functions of flies. Furthermore, we have developed an HPLC method to reliably detect both free and protein-bound BMAA in fly tissue extracts.

  17. KPNB1 mediates PER/CRY nuclear translocation and circadian clock function.

    Science.gov (United States)

    Lee, Yool; Jang, A Reum; Francey, Lauren J; Sehgal, Amita; Hogenesch, John B

    2015-08-29

    Regulated nuclear translocation of the PER/CRY repressor complex is critical for negative feedback regulation of the circadian clock of mammals. However, the precise molecular mechanism is not fully understood. Here, we report that KPNB1, an importin β component of the ncRNA repressor of nuclear factor of activated T cells (NRON) ribonucleoprotein complex, mediates nuclear translocation and repressor function of the PER/CRY complex. RNAi depletion of KPNB1 traps the PER/CRY complex in the cytoplasm by blocking nuclear entry of PER proteins in human cells. KPNB1 interacts mainly with PER proteins and directs PER/CRY nuclear transport in a circadian fashion. Interestingly, KPNB1 regulates the PER/CRY nuclear entry and repressor function, independently of importin α, its classical partner. Moreover, inducible inhibition of the conserved Drosophila importin β in lateral neurons abolishes behavioral rhythms in flies. Collectively, these data show that KPNB1 is required for timely nuclear import of PER/CRY in the negative feedback regulation of the circadian clock.

  18. Proteomic identification of Drosophila melanogaster male accessory gland proteins, including a pro-cathepsin and a soluble γ-glutamyl transpeptidase

    Directory of Open Access Journals (Sweden)

    Sajid Mohammed

    2006-05-01

    Full Text Available Background In Drosophila melanogaster, the male seminal fluid contains proteins that are important for reproductive success. Many of these proteins are synthesised by the male accessory glands and are secreted into the accessory gland lumen, where they are stored until required. Previous studies on the identification of Drosophila accessory gland products have largely focused on characterisation of male-specific accessory gland cDNAs from D. melanogaster and, more recently, Drosophila simulans. In the present study, we have used a proteomics approach without any sex bias to identify proteins in D. melanogaster accessory gland secretions. Results Thirteen secreted accessory gland proteins, including seven new accessory gland proteins, were identified by 2D-gel electrophoresis combined with mass spectrometry of tryptic fragments. They included protein-folding and stress-response proteins, a hormone, a lipase, a serpin, a cysteine-rich protein and two peptidases, a pro-enzyme form of a cathepsin K-like cysteine peptidase and a γ-glutamyl transpeptidase. Enzymatic studies established that accessory gland secretions contain a cysteine peptidase zymogen that can be activated at low pH. This peptidase may have a role in the processing of female and other male-derived proteins, but is unlikely to be involved in the processing of the sex peptide. γ-Glutamyl transpeptidases are type II integral membrane proteins; however, the identified AG γ-glutamyl transpeptidase (GGT-1 is unusual in that it is predicted to be a soluble secreted protein, a prediction that is supported by biochemical evidence. GGT-1 is possibly involved in maintaining a protective redox environment for sperm. The strong γ-glutamyl transpeptidase activity found in the secretions provides an explanation for the observation that glutamic acid is the most abundant free amino acid in accessory gland secretions of D. melanogaster. Conclusion We have applied biochemical approaches, not used

  19. Proteomic identification of Drosophila melanogaster male accessory gland proteins, including a pro-cathepsin and a soluble gamma-glutamyl transpeptidase.

    Science.gov (United States)

    Walker, Michael J; Rylett, Caroline M; Keen, Jeff N; Audsley, Neil; Sajid, Mohammed; Shirras, Alan D; Isaac, R Elwyn

    2006-05-02

    In Drosophila melanogaster, the male seminal fluid contains proteins that are important for reproductive success. Many of these proteins are synthesised by the male accessory glands and are secreted into the accessory gland lumen, where they are stored until required. Previous studies on the identification of Drosophila accessory gland products have largely focused on characterisation of male-specific accessory gland cDNAs from D. melanogaster and, more recently, Drosophila simulans. In the present study, we have used a proteomics approach without any sex bias to identify proteins in D. melanogaster accessory gland secretions. Thirteen secreted accessory gland proteins, including seven new accessory gland proteins, were identified by 2D-gel electrophoresis combined with mass spectrometry of tryptic fragments. They included protein-folding and stress-response proteins, a hormone, a lipase, a serpin, a cysteine-rich protein and two peptidases, a pro-enzyme form of a cathepsin K-like cysteine peptidase and a gamma-glutamyl transpeptidase. Enzymatic studies established that accessory gland secretions contain a cysteine peptidase zymogen that can be activated at low pH. This peptidase may have a role in the processing of female and other male-derived proteins, but is unlikely to be involved in the processing of the sex peptide. gamma-Glutamyl transpeptidases are type II integral membrane proteins; however, the identified AG gamma-glutamyl transpeptidase (GGT-1) is unusual in that it is predicted to be a soluble secreted protein, a prediction that is supported by biochemical evidence. GGT-1 is possibly involved in maintaining a protective redox environment for sperm. The strong gamma-glutamyl transpeptidase activity found in the secretions provides an explanation for the observation that glutamic acid is the most abundant free amino acid in accessory gland secretions of D. melanogaster. We have applied biochemical approaches, not used previously, to characterise

  20. Atomic and gravitational clocks

    International Nuclear Information System (INIS)

    Canuto, V.M.; City Coll., New York; Goldman, I.

    1982-01-01

    Atomic and gravitational clocks are governed by the laws of electrodynamics and gravity respectively. While the strong equivalence principle (SEP) assumes that the two clocks have been synchronous at all times, recent planetary data seem to suggest a possible violation of the SEP. Past analysis of the implications of an SEP violation on different physical phenomena revealed no disagreement. However, these studies assumed that the two different clocks can be consistently constructed within the framework. The concept of scale invariance, and the physical meaning of different systems of units, are now reviewed and the construction of two clocks that do not remain synchronous-whose rates are related by a non-constant function βsub(a)-is demonstrated. The cosmological character of βsub(a) is also discussed. (author)

  1. Clock synchronization and dispersion

    International Nuclear Information System (INIS)

    Giovannetti, Vittorio; Lloyd, Seth; Maccone, Lorenzo; Wong, Franco N C

    2002-01-01

    We present a method to defeat effects of dispersion of timing signals when synchronizing clocks. It is based on the recently proposed 'conveyor belt synchronization' scheme and on the quantum dispersion cancellation effect

  2. A homeostatic clock sets daughter centriole size in flies

    Science.gov (United States)

    Aydogan, Mustafa G.; Steinacker, Thomas L.; Novak, Zsofia A.; Baumbach, Janina; Muschalik, Nadine

    2018-01-01

    Centrioles are highly structured organelles whose size is remarkably consistent within any given cell type. New centrioles are born when Polo-like kinase 4 (Plk4) recruits Ana2/STIL and Sas-6 to the side of an existing “mother” centriole. These two proteins then assemble into a cartwheel, which grows outwards to form the structural core of a new daughter. Here, we show that in early Drosophila melanogaster embryos, daughter centrioles grow at a linear rate during early S-phase and abruptly stop growing when they reach their correct size in mid- to late S-phase. Unexpectedly, the cartwheel grows from its proximal end, and Plk4 determines both the rate and period of centriole growth: the more active the centriolar Plk4, the faster centrioles grow, but the faster centriolar Plk4 is inactivated and growth ceases. Thus, Plk4 functions as a homeostatic clock, establishing an inverse relationship between growth rate and period to ensure that daughter centrioles grow to the correct size. PMID:29500190

  3. Cell type-specific recruitment of Drosophila Lin-7 to distinct MAGUK-based protein complexes defines novel roles for Sdt and Dlg-S97.

    Science.gov (United States)

    Bachmann, André; Timmer, Marco; Sierralta, Jimena; Pietrini, Grazia; Gundelfinger, Eckart D; Knust, Elisabeth; Thomas, Ulrich

    2004-04-15

    Stardust (Sdt) and Discs-Large (Dlg) are membrane-associated guanylate kinases (MAGUKs) involved in the organization of supramolecular protein complexes at distinct epithelial membrane compartments in Drosophila. Loss of either Sdt or Dlg affects epithelial development with severe effects on apico-basal polarity. Moreover, Dlg is required for the structural and functional integrity of synaptic junctions. Recent biochemical and cell culture studies have revealed that various mammalian MAGUKs can interact with mLin-7/Veli/MALS, a small PDZ-domain protein. To substantiate these findings for their in vivo significance with regard to Sdt- and Dlg-based protein complexes, we analyzed the subcellular distribution of Drosophila Lin-7 (DLin-7) and performed genetic and biochemical assays to characterize its interaction with either of the two MAGUKs. In epithelia, Sdt mediates the recruitment of DLin-7 to the subapical region, while at larval neuromuscular junctions, a particular isoform of Dlg, Dlg-S97, is required for postsynaptic localization of DLin-7. Ectopic expression of Dlg-S97 in epithelia, however, was not sufficient to induce a redistribution of DLin-7. These results imply that the recruitment of DLin-7 to MAGUK-based protein complexes is defined by cell-type specific mechanisms and that DLin-7 acts downstream of Sdt in epithelia and downstream of Dlg at synapses.

  4. FUNCTIONAL IMPLICATIONS OF THE CLOCK 3111T/C SINGLE-NUCLEOTIDE POLYMORPHISM

    Directory of Open Access Journals (Sweden)

    Angela Renee Ozburn

    2016-04-01

    Full Text Available Circadian rhythm disruptions are prominently associated with Bipolar Disorder (BD. Circadian rhythms are regulated by the molecular clock, a family of proteins that function together in a transcriptional-translational feedback loop. The CLOCK protein is a key transcription factor of this feedback loop, and previous studies have found that manipulations of the Clock gene are sufficient to produce manic-like behavior in mice (Roybal et al., 2007. The Clock 3111T/C single-nucleotide polymorphism (SNP; rs1801260 is a genetic variation of the human Clock gene that is significantly associated with increased frequency of manic episodes in BD patients (Benedetti et al., 2003. The 3111T/C SNP is located in the 3’ untranslated region of the Clock gene. In this study, we sought to examine the functional implications of the human Clock 3111T/C SNP by transfecting a mammalian cell line (mouse embryonic fibroblasts isolated from Clock -/- knockout mice with pcDNA plasmids containing the human Clock gene with either the T or C SNP at position 3111. We then measured circadian gene expression over a 24 hour time period. We found that the Clock3111C SNP resulted in higher mRNA levels than the Clock 3111T SNP. Further, we found that Per2, a transcriptional target of CLOCK, was also more highly expressed with Clock 3111C expression, indicating the 3’UTR SNP affects the expression, function and stability of Clock mRNA.

  5. The WW domain protein Kibra acts upstream of Hippo in Drosophila

    DEFF Research Database (Denmark)

    Baumgartner, Roland; Poernbacher, Ingrid; Buser, Nathalie

    2010-01-01

    inactivating the transcriptional coactivator Yorkie is well established, much less is known about the upstream events that regulate Hippo signaling activity. The FERM domain proteins Expanded and Merlin appear to represent two different signaling branches that feed into the Hippo pathway. Signaling...... by the atypical cadherin Fat may act via Expanded, but how Merlin is regulated has remained elusive. Here, we show that the WW domain protein Kibra is a Hippo signaling component upstream of Hippo and Merlin. Kibra acts synergistically with Expanded, and it physically interacts with Merlin. Thus, Kibra...

  6. Drosophila Vps13 Is Required for Protein Homeostasis in the Brain

    NARCIS (Netherlands)

    Vonk, Jan J.; Yeshaw, Wondwossen M.; Pinto, Francesco; Faber, Anita I. E.; Lahaye, Liza L.; Kanon, Bart; van der Zwaag, Marianne; Velayos-Baeza, Antonio; Freire, Raimundo; van IJzendoorn, Sven C.; Grzeschik, Nicola A.; Sibon, Ody C. M.

    2017-01-01

    Chorea-Acanthocytosis is a rare, neurodegenerative disorder characterized by progressive loss of locomotor and cognitive function. It is caused by loss of function mutations in the Vacuolar Protein Sorting 13A (VPS13A) gene, which is conserved from yeast to human. The consequences of VPS13A

  7. Protein expression profiling of the drosophila fragile X mutant brain reveals up-regulation of monoamine synthesis.

    Science.gov (United States)

    Zhang, Yong Q; Friedman, David B; Wang, Zhe; Woodruff, Elvin; Pan, Luyuan; O'donnell, Janis; Broadie, Kendal

    2005-03-01

    Fragile X syndrome is the most common form of inherited mental retardation, associated with both cognitive and behavioral anomalies. The disease is caused by silencing of the fragile X mental retardation 1 (fmr1) gene, which encodes the mRNA-binding, translational regulator FMRP. Previously we established a disease model through mutation of Drosophila fmr1 (dfmr1) and showed that loss of dFMRP causes defects in neuronal structure, function, and behavioral output similar to the human disease state. To uncover molecular targets of dFMRP in the brain, we use here a proteomic approach involving two-dimensional difference gel electrophoresis analyses followed by mass spectrometry identification of proteins with significantly altered expression in dfmr1 null mutants. We then focus on two misregulated enzymes, phenylalanine hydroxylase (Henna) and GTP cyclohydrolase (Punch), both of which mediate in concert the synthetic pathways of two key monoamine neuromodulators, dopamine and serotonin. Brain enzymatic assays show a nearly 2-fold elevation of Punch activity in dfmr1 null mutants. Consistently brain neurochemical assays show that both dopamine and serotonin are significantly increased in dfmr1 null mutants. At a cellular level, dfmr1 null mutant neurons display a highly significant elevation of the dense core vesicles that package these monoamine neuromodulators for secretion. Taken together, these data indicate that dFMRP normally down-regulates the monoamine pathway, which is consequently up-regulated in the mutant condition. Elevated brain levels of dopamine and serotonin provide a plausible mechanistic explanation for aspects of cognitive and behavioral deficits in human patients.

  8. The Human dsRNA binding protein PACT is unable to functionally substitute for the Drosophila dsRNA binding protein R2D2 [v1; ref status: indexed, http://f1000r.es/201

    Directory of Open Access Journals (Sweden)

    Benjamin K Dickerman

    2013-10-01

    Full Text Available The primary function of the dsRNA binding protein (dsRBP PACT/RAX is to activate the dsRNA dependent protein kinase PKR in response to stress signals.  Additionally, it has been identified as a component of the small RNA processing pathway.  A role for PACT/RAX in this pathway represents an important interplay between two modes of post-transcriptional gene regulation.  The function of PACT/RAX in this context is poorly understood.  Thus, additional models are required to clarify the mechanism by which PACT/RAX functions.  In this study, Drosophila melanogaster was employed to identify functionally orthologous dsRNA-binding proteins.  Transgenic Drosophila expressing human PACT were generated to determine whether PACT is capable of functionally substituting for the Drosophila dsRBP R2D2, which has a well-defined role in small RNA biogenesis.  Results presented here indicate that PACT is unable to substitute for R2D2 at the whole organism level.

  9. Exploring some of the physico-chemical properties of the LAMMER protein kinase DOA of Drosophila

    Czech Academy of Sciences Publication Activity Database

    Farkaš, R.; Kováčiková, M.; Liszeková, D.; Beňo, M.; Daniš, P.; Rabinow, L.; Chase, B. A.; Raška, Ivan

    2009-01-01

    Roč. 3, č. 2 (2009), s. 130-142 ISSN 1933-6934 Grant - others:GA MŠk(CZ) LC535; GA ČR(CZ) GA304/02/0342; GA ČR(CZ) GA304/04/0692 Program:LC; GA Institutional research plan: CEZ:AV0Z50110509 Keywords : protein kinase structure * function * genetics Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.156, year: 2009

  10. Electrostatic Interactions between Elongated Monomers Drive Filamentation of Drosophila Shrub, a Metazoan ESCRT-III Protein

    Directory of Open Access Journals (Sweden)

    Brian J. McMillan

    2016-08-01

    Full Text Available The endosomal sorting complex required for transport (ESCRT is a conserved protein complex that facilitates budding and fission of membranes. It executes a key step in many cellular events, including cytokinesis and multi-vesicular body formation. The ESCRT-III protein Shrub in flies, or its homologs in yeast (Snf7 or humans (CHMP4B, is a critical polymerizing component of ESCRT-III needed to effect membrane fission. We report the structural basis for polymerization of Shrub and define a minimal region required for filament formation. The X-ray structure of the Shrub core shows that individual monomers in the lattice interact in a staggered arrangement using complementary electrostatic surfaces. Mutations that disrupt interface salt bridges interfere with Shrub polymerization and function. Despite substantial sequence divergence and differences in packing interactions, the arrangement of Shrub subunits in the polymer resembles that of Snf7 and other family homologs, suggesting that this intermolecular packing mechanism is shared among ESCRT-III proteins.

  11. Atomic clocks for geodesy

    Science.gov (United States)

    Mehlstäubler, Tanja E.; Grosche, Gesine; Lisdat, Christian; Schmidt, Piet O.; Denker, Heiner

    2018-06-01

    We review experimental progress on optical atomic clocks and frequency transfer, and consider the prospects of using these technologies for geodetic measurements. Today, optical atomic frequency standards have reached relative frequency inaccuracies below 10‑17, opening new fields of fundamental and applied research. The dependence of atomic frequencies on the gravitational potential makes atomic clocks ideal candidates for the search for deviations in the predictions of Einstein’s general relativity, tests of modern unifying theories and the development of new gravity field sensors. In this review, we introduce the concepts of optical atomic clocks and present the status of international clock development and comparison. Besides further improvement in stability and accuracy of today’s best clocks, a large effort is put into increasing the reliability and technological readiness for applications outside of specialized laboratories with compact, portable devices. With relative frequency uncertainties of 10‑18, comparisons of optical frequency standards are foreseen to contribute together with satellite and terrestrial data to the precise determination of fundamental height reference systems in geodesy with a resolution at the cm-level. The long-term stability of atomic standards will deliver excellent long-term height references for geodetic measurements and for the modelling and understanding of our Earth.

  12. Use of a transfected and amplified Drosophila heat shock promoter construction for inducible production of toxic mouse c-myc proteins in CHO cells

    International Nuclear Information System (INIS)

    Wurm, F.M.; Gwinn, K.A.; Papoulas, O.; Pallavicini, M.; Kingston, R.E.

    1987-01-01

    After transfection and selection with methotrexate, CHO cell lines were established which contained up to 2000 copies of an expression vector for c-myc protein. The vector contained the Drosophila heat shock protein 70 (hsp70) promoter fused with the coding region of the mouse c-myc gene. Incubation of cells for up to 3 hours at 43 0 C resulted in at least a 100-fold induction of recombinant c-myc mRNA. When cells were shifted back to 37 0 C, within 1 to 4 hours, this RNA was translated into protein to yield about 250 μg per 10 9 cells. Cells died a few hours later, suggesting that high concentrations of intracellular c-myc are cytotoxic. 47 refs., 5 figs

  13. Intact interval timing in circadian CLOCK mutants.

    Science.gov (United States)

    Cordes, Sara; Gallistel, C R

    2008-08-28

    While progress has been made in determining the molecular basis for the circadian clock, the mechanism by which mammalian brains time intervals measured in seconds to minutes remains a mystery. An obvious question is whether the interval-timing mechanism shares molecular machinery with the circadian timing mechanism. In the current study, we trained circadian CLOCK +/- and -/- mutant male mice in a peak-interval procedure with 10 and 20-s criteria. The mutant mice were more active than their wild-type littermates, but there were no reliable deficits in the accuracy or precision of their timing as compared with wild-type littermates. This suggests that expression of the CLOCK protein is not necessary for normal interval timing.

  14. The Drosophila inner-membrane protein PMI controls crista biogenesis and mitochondrial diameter.

    Science.gov (United States)

    Macchi, Marc; El Fissi, Najla; Tufi, Roberta; Bentobji, Mélanie; Liévens, Jean-Charles; Martins, L Miguel; Royet, Julien; Rival, Thomas

    2013-02-01

    Cristae are mitochondrial inner-membrane structures that concentrate respiratory chain complexes and hence regulate ATP production. Mechanisms controlling crista morphogenesis are poorly understood and few crista determinants have been identified. Among them are the Mitofilins that are required to establish crista junctions and ATP-synthase subunits that bend the membrane at the tips of the cristae. We report here the phenotypic consequences associated with the in vivo inactivation of the inner-membrane protein Pantagruelian Mitochondrion I (PMI) both at the scale of the whole organism, and at the level of mitochondrial ultrastructure and function. We show that flies in which PMI is genetically inactivated experience synaptic defects and have a reduced life span. Electron microscopy analysis of the inner-membrane morphology demonstrates that loss of PMI function increases the average length of mitochondrial cristae in embryonic cells. This phenotype is exacerbated in adult neurons in which cristae form a dense tangle of elongated membranes. Conversely, we show that PMI overexpression is sufficient to reduce crista length in vivo. Finally, these crista defects are associated with impaired respiratory chain activity and increases in the level of reactive oxygen species. Since PMI and its human orthologue TMEM11 are regulators of mitochondrial morphology, our data suggest that, by controlling crista length, PMI influences mitochondrial diameter and tubular shape.

  15. SH3 domain-mediated binding of the Drk protein to Dos is an important step in signaling of Drosophila receptor tyrosine kinases.

    Science.gov (United States)

    Feller, Stephan M; Wecklein, Heike; Lewitzky, Marc; Kibler, Eike; Raabe, Thomas

    2002-08-01

    Activation of the Sevenless (Sev) receptor tyrosine kinase (RTK) in the developing Drosophila eye is required for the specification of the R7 photoreceptor cell fate. Daughter of Sevenless (Dos), a putative multi-site adaptor protein, is a substrate of the Sev kinase and is known to associate with the tyrosine phosphatase Corkscrew (Csw). Binding of Csw to Dos depends on the Csw Src homology 2 (SH2) domains and is an essential step for signaling by the Sev RTK. Dos, however, lacks a recognizable phosphotyrosine interaction domain and it was previously unclear how it is recruited to the Sev receptor. Here it is shown that the SH2/SH3 domain adaptor protein Drk can provide this link. Drk binds with its SH2 domain to the autophosphorylated Sev receptor while the C-terminal SH3 domain is able to associate with Dos. The Drk SH3 domain binding motifs on Dos were mapped to two sites which do not conform the known Drk SH3 domain binding motif (PxxPxR) but instead have the consensus PxxxRxxKP. Mutational analysis in vitro and in vivo provided evidence that both Drk binding sites fulfil an important function in the context of Sev and Drosophila epidermal growth factor receptor mediated signaling processes.

  16. Drosophila C-terminal binding protein, dCtBP is required for sensory organ prepattern and sharpens proneural transcriptional activity of the GATA factor Pnr.

    Science.gov (United States)

    Biryukova, Inna; Heitzler, Pascal

    2008-11-01

    The peripheral nervous system is required for animals to detect and to relay environmental stimuli to central nervous system for the information processing. In Drosophila, the precise spatial and temporal expression of two proneural genes achaete (ac) and scute (sc), is necessary for development of the sensory organs. Here we present an evidence that the transcription co-repressor, dCtBP acts as a negative regulator of sensory organ prepattern. Loss of dCtBP function mutant exhibits ectopic sensory organs, while overexpression of dCtBP results in a dramatic loss of sensory organs. These phenotypes are correlated with mis-emerging of sensory organ precursors and perturbated expression of proneural transcription activator Ac. Mammalian CtBP-1 was identified via interaction with the consensus motif PXDLSX(K/R) of adenovirus E1A oncoprotein. We demonstrated that dCtBP binds directly to PLDLS motif of Drosophila Friend of GATA-1 protein, U-shaped and sharpens the adult sensory organ development. Moreover, we found that dCtBP mediates multivalent interaction with the GATA transcriptional activator Pannier and acts as a direct co-repressor of the Pannier-mediated activation of proneural genes. We demonstrated that Pannier genetically interacts with dCtBP-interacting protein HDAC1, suggesting that the dCtBP-dependent regulation of Pannier activity could utilize a repressive mechanism involving alteration of local chromatine structure.

  17. Decamp Clock Board Firmware

    International Nuclear Information System (INIS)

    Vicente, J. de; Castilla, J.; Martinez, G.

    2007-01-01

    Decamp (Dark Energy Survey Camera) is a new instrument designed to explore the universe aiming to reveal the nature of Dark Energy. The camera consists of 72 CCDs and 520 Mpixels. The readout electronics of DECam is based on the Monsoon system. Monsoon is a new image acquisition system developed by the NOAO (National Optical Astronomical Observatory) for the new generation of astronomical cameras. The Monsoon system uses three types of boards inserted in a Eurocard format based crate: master control board, acquisition board and clock board. The direct use of the Monsoon system for DECam readout electronics requires nine crates mainly due to the high number of clock boards needed. Unfortunately, the available space for DECam electronics is constrained to four crates at maximum. The major drawback to achieve such desired compaction degree resides in the clock board signal density. This document describes the changes performed at CIEMAT on the programmable logic of the Monsoon clock board aiming to meet such restricted space constraints. (Author) 5 refs

  18. Decamp Clock Board Firmware

    Energy Technology Data Exchange (ETDEWEB)

    Vicente, J. de; Castilla, J.; Martinez, G.

    2007-09-27

    Decamp (Dark Energy Survey Camera) is a new instrument designed to explore the universe aiming to reveal the nature of Dark Energy. The camera consists of 72 CCDs and 520 Mpixels. The readout electronics of DECam is based on the Monsoon system. Monsoon is a new image acquisition system developed by the NOAO (National Optical Astronomical Observatory) for the new generation of astronomical cameras. The Monsoon system uses three types of boards inserted in a Eurocard format based crate: master control board, acquisition board and clock board. The direct use of the Monsoon system for DECam readout electronics requires nine crates mainly due to the high number of clock boards needed. Unfortunately, the available space for DECam electronics is constrained to four crates at maximum. The major drawback to achieve such desired compaction degree resides in the clock board signal density. This document describes the changes performed at CIEMAT on the programmable logic of the Monsoon clock board aiming to meet such restricted space constraints. (Author) 5 refs.

  19. A cGMP-dependent protein kinase gene, foraging, modifies habituation-like response decrement of the giant fiber escape circuit in Drosophila.

    Science.gov (United States)

    Engel, J E; Xie, X J; Sokolowski, M B; Wu, C F

    2000-01-01

    The Drosophila giant fiber jump-and-flight escape response is a model for genetic analysis of both the physiology and the plasticity of a sensorimotor behavioral pathway. We previously established the electrically induced giant fiber response in intact tethered flies as a model for habituation, a form of nonassociative learning. Here, we show that the rate of stimulus-dependent response decrement of this neural pathway in a habituation protocol is correlated with PKG (cGMP-Dependent Protein Kinase) activity and foraging behavior. We assayed response decrement for natural and mutant rover and sitter alleles of the foraging (for) gene that encodes a Drosophila PKG. Rover larvae and adults, which have higher PKG activities, travel significantly farther while foraging than sitters with lower PKG activities. Response decrement was most rapid in genotypes previously shown to have low PKG activities and sitter-like foraging behavior. We also found differences in spontaneous recovery (the reversal of response decrement during a rest from stimulation) and a dishabituation-like phenomenon (the reversal of response decrement evoked by a novel stimulus). This electrophysiological study in an intact animal preparation provides one of the first direct demonstrations that PKG can affect plasticity in a simple learning paradigm. It increases our understanding of the complex interplay of factors that can modulate the sensitivity of the giant fiber escape response, and it defines a new adult-stage phenotype of the foraging locus. Finally, these results show that behaviorally relevant neural plasticity in an identified circuit can be influenced by a single-locus genetic polymorphism existing in a natural population of Drosophila.

  20. The Drosophila IKK-related kinase (Ik2 and Spindle-F proteins are part of a complex that regulates cytoskeleton organization during oogenesis

    Directory of Open Access Journals (Sweden)

    Shaanan Boaz

    2008-09-01

    Full Text Available Abstract Background IkappaB kinases (IKKs regulate the activity of Rel/NF-kappaB transcription factors by targeting their inhibitory partner proteins, IkappaBs, for degradation. The Drosophila genome encodes two members of the IKK family. Whereas the first is a kinase essential for activation of the NF-kappaB pathway, the latter does not act as IkappaB kinase. Instead, recent findings indicate that Ik2 regulates F-actin assembly by mediating the function of nonapoptotic caspases via degradation of DIAP1. Also, it has been suggested that ik2 regulates interactions between the minus ends of the microtubules and the actin-rich cortex in the oocyte. Since spn-F mutants display oocyte defects similar to those of ik2 mutant, we decided to investigate whether Spn-F could be a direct regulatory target of Ik2. Results We found that Ik2 binds physically to Spn-F, biomolecular interaction analysis of Spn-F and Ik2 demonstrating that both proteins bind directly and form a complex. We showed that Ik2 phosphorylates Spn-F and demonstrated that this phosphorylation does not lead to Spn-F degradation. Ik2 is localized to the anterior ring of the oocyte and to punctate structures in the nurse cells together with Spn-F protein, and both proteins are mutually required for their localization. Conclusion We conclude that Ik2 and Spn-F form a complex, which regulates cytoskeleton organization during Drosophila oogenesis and in which Spn-F is the direct regulatory target for Ik2. Interestingly, Ik2 in this complex does not function as a typical IKK in that it does not direct SpnF for degradation following phosphorylation.

  1. The cohesion protein SOLO associates with SMC1 and is required for synapsis, recombination, homolog bias and cohesion and pairing of centromeres in Drosophila Meiosis.

    Science.gov (United States)

    Yan, Rihui; McKee, Bruce D

    2013-01-01

    Cohesion between sister chromatids is mediated by cohesin and is essential for proper meiotic segregation of both sister chromatids and homologs. solo encodes a Drosophila meiosis-specific cohesion protein with no apparent sequence homology to cohesins that is required in male meiosis for centromere cohesion, proper orientation of sister centromeres and centromere enrichment of the cohesin subunit SMC1. In this study, we show that solo is involved in multiple aspects of meiosis in female Drosophila. Null mutations in solo caused the following phenotypes: 1) high frequencies of homolog and sister chromatid nondisjunction (NDJ) and sharply reduced frequencies of homolog exchange; 2) reduced transmission of a ring-X chromosome, an indicator of elevated frequencies of sister chromatid exchange (SCE); 3) premature loss of centromere pairing and cohesion during prophase I, as indicated by elevated foci counts of the centromere protein CID; 4) instability of the lateral elements (LE)s and central regions of synaptonemal complexes (SCs), as indicated by fragmented and spotty staining of the chromosome core/LE component SMC1 and the transverse filament protein C(3)G, respectively, at all stages of pachytene. SOLO and SMC1 are both enriched on centromeres throughout prophase I, co-align along the lateral elements of SCs and reciprocally co-immunoprecipitate from ovarian protein extracts. Our studies demonstrate that SOLO is closely associated with meiotic cohesin and required both for enrichment of cohesin on centromeres and stable assembly of cohesin into chromosome cores. These events underlie and are required for stable cohesion of centromeres, synapsis of homologous chromosomes, and a recombination mechanism that suppresses SCE to preferentially generate homolog crossovers (homolog bias). We propose that SOLO is a subunit of a specialized meiotic cohesin complex that mediates both centromeric and axial arm cohesion and promotes homolog bias as a component of chromosome

  2. Molecular cloning, genomic organization, developmental regulation, and a knock-out mutant of a novel leu-rich repeats-containing G protein-coupled receptor (DLGR-2) from Drosophila melanogaster

    DEFF Research Database (Denmark)

    Eriksen, Kathrine Krageskov; Hauser, Frank; Schiøtt, Morten

    2000-01-01

    After screening the Berkeley Drosophila Genome Project database with sequences from a recently characterized Leu-rich repeats-containing G protein-coupled receptor (LGR) fromDrosophila (DLGR-1), we identified a second gene for a different LGR (DLGR-2) and cloned its cDNA. DLGR-2 is 1360 amino aci...... knock-out mutants, where the DLGR-2 gene is interrupted by a P element insertion, die around the time of hatching. This finding, together with the expression data, strongly suggests that DLGR-2 is exclusively involved in development....

  3. bHLH-O proteins balance the self-renewal and differentiation of Drosophila neural stem cells by regulating Earmuff expression.

    Science.gov (United States)

    Li, Xiaosu; Chen, Rui; Zhu, Sijun

    2017-11-15

    Balancing self-renewal and differentiation of stem cells requires differential expression of self-renewing factors in two daughter cells generated from the asymmetric division of the stem cells. In Drosophila type II neural stem cell (or neuroblast, NB) lineages, the expression of the basic helix-loop-helix-Orange (bHLH-O) family proteins, including Deadpan (Dpn) and E(spl) proteins, is required for maintaining the self-renewal and identity of type II NBs, whereas the absence of these self-renewing factors is essential for the differentiation of intermediate neural progenitors (INPs) generated from type II NBs. Here, we demonstrate that Dpn maintains type II NBs by suppressing the expression of Earmuff (Erm). We provide evidence that Dpn and E(spl) proteins suppress Erm by directly binding to C-sites and N-boxes in the cis-regulatory region of erm. Conversely, the absence of bHLH-O proteins in INPs allows activation of erm and Erm-mediated maturation of INPs. Our results further suggest that Pointed P1 (PntP1) mediates the dedifferentiation of INPs resulting from the loss of Erm or overexpression of Dpn or E(spl) proteins. Taken together, these findings reveal mechanisms underlying the regulation of the maintenance of type II NBs and differentiation of INPs through the differential expression of bHLH-O family proteins. Copyright © 2017 Elsevier Inc. All rights reserved.

  4. Wolbachia Protein TomO Targets nanos mRNA and Restores Germ Stem Cells in Drosophila Sex-lethal Mutants.

    Science.gov (United States)

    Ote, Manabu; Ueyama, Morio; Yamamoto, Daisuke

    2016-09-12

    Wolbachia, endosymbiotic bacteria prevalent in invertebrates, manipulate their hosts in a variety of ways: they induce cytoplasmic incompatibility, male lethality, male-to-female transformation, and parthenogenesis. However, little is known about the molecular basis for host manipulation by these bacteria. In Drosophila melanogaster, Wolbachia infection makes otherwise sterile Sex-lethal (Sxl) mutant females capable of producing mature eggs. Through a functional genomic screen for Wolbachia genes with growth-inhibitory effects when expressed in cultured Drosophila cells, we identified the gene WD1278 encoding a novel protein we call toxic manipulator of oogenesis (TomO), which phenocopies some of the Wolbachia effects in Sxl mutant D. melanogaster females. We demonstrate that TomO enhances the maintenance of germ stem cells (GSCs) by elevating Nanos (Nos) expression via its interaction with nos mRNA, ultimately leading to the restoration of germ cell production in Sxl mutant females that are otherwise without GSCs. Copyright © 2016 Elsevier Ltd. All rights reserved.

  5. The role of biological clock in glucose homeostasis 

    Directory of Open Access Journals (Sweden)

    Piotr Chrościcki

    2013-06-01

    Full Text Available The mechanism of the biological clock is based on a rhythmic expression of clock genes and clock-controlled genes. As a result of their transcripto-translational associations, endogenous rhythms in the synthesis of key proteins of various physiological and metabolic processes are created. The major timekeeping mechanism for these rhythms exists in the central nervous system. The master circadian clock, localized in suprachiasmatic nucleus (SCN, regulates multiple metabolic pathways, while feeding behavior and metabolite availability can in turn regulate the circadian clock. It is also suggested that in the brain there is a food entrainable oscillator (FEO or oscillators, resulting in activation of both food anticipatory activity and hormone secretion that control digestion processes. Moreover, most cells and tissues express autonomous clocks. Maintenance of the glucose homeostasis is particularly important for the proper function of the body, as this sugar is the main source of energy for the brain, retina, erythrocytes and skeletal muscles. Thus, glucose production and utilization are synchronized in time. The hypothalamic excited orexin neurons control energy balance of organism and modulate the glucose production and utilization. Deficiency of orexin action results in narcolepsy and weight gain, whereas glucose and amino acids can affect activity of the orexin cells. Large-scale genetic studies in rodents and humans provide evidence for the involvement of disrupted clock gene expression rhythms in the pathogenesis of obesity and type 2 diabetes. In general, the current lifestyle of the developed modern societies disturbs the action of biological clock

  6. UNC79 and UNC80, putative auxiliary subunits of the NARROW ABDOMEN ion channel, are indispensable for robust circadian locomotor rhythms in Drosophila.

    Directory of Open Access Journals (Sweden)

    Bridget C Lear

    Full Text Available In the fruit fly Drosophila melanogaster, a network of circadian pacemaker neurons drives daily rhythms in rest and activity. The ion channel NARROW ABDOMEN (NA, orthologous to the mammalian sodium leak channel NALCN, functions downstream of the molecular circadian clock in pacemaker neurons to promote behavioral rhythmicity. To better understand the function and regulation of the NA channel, we have characterized two putative auxiliary channel subunits in Drosophila, unc79 (aka dunc79 and unc80 (aka CG18437. We have generated novel unc79 and unc80 mutations that represent strong or complete loss-of-function alleles. These mutants display severe defects in circadian locomotor rhythmicity that are indistinguishable from na mutant phenotypes. Tissue-specific RNA interference and rescue analyses indicate that UNC79 and UNC80 likely function within pacemaker neurons, with similar anatomical requirements to NA. We observe an interdependent, post-transcriptional regulatory relationship among the three gene products, as loss of na, unc79, or unc80 gene function leads to decreased expression of all three proteins, with minimal effect on transcript levels. Yet despite this relationship, we find that the requirement for unc79 and unc80 in circadian rhythmicity cannot be bypassed by increasing NA protein expression, nor can these putative auxiliary subunits substitute for each other. These data indicate functional requirements for UNC79 and UNC80 beyond promoting channel subunit expression. Immunoprecipitation experiments also confirm that UNC79 and UNC80 form a complex with NA in the Drosophila brain. Taken together, these data suggest that Drosophila NA, UNC79, and UNC80 function together in circadian clock neurons to promote rhythmic behavior.

  7. Comparisons of mental clocks.

    Science.gov (United States)

    Paivio, A

    1978-02-01

    Subjects in three experiments were presented with pairs of clock times and were required to choose the one in which the hour and minute hand formed the smaller angle. In Experiments 1 and 2, the times were presented digitally, necessitating a transformation into symbolic representations from which the angular size difference could be inferred. The results revealed orderly symbolic distance effects so that comparison reaction time increased as the angular size difference decreased. Moreover, subjects generally reported using imagery to make the judgment, and subjects scoring high on test of imagery ability were faster than those scoring low on such tests. Experiment 3 added a direct perceptual condition in which subjects compared angles between pairs of hands on two drawn (analog) clocks, as well as a mixed condition involving one digital and one analog clock time. The results showed comparable distance effects for all conditions. In addition, reaction time increased from the perceptual, to the mixed, to the pure-digital condition. These results are consistent with predictions from an image-based dual-coding theory.

  8. Circadian expression of clock genes and clock-controlled genes in the rat retina

    NARCIS (Netherlands)

    Kamphuis, Willem; Cailotto, Cathy; Dijk, Frederike; Bergen, Arthur; Buijs, Ruud M.

    2005-01-01

    The circadian expression patterns of genes encoding for proteins that make up the core of the circadian clock were measured in rat retina using real-time quantitative PCR (qPCR). Transcript levels of several genes previously used for normalization of qPCR assays were determined and the effect of

  9. SUMOylation in Drosophila Development

    Directory of Open Access Journals (Sweden)

    Albert J. Courey

    2012-07-01

    Full Text Available Small ubiquitin-related modifier (SUMO, an ~90 amino acid ubiquitin-like protein, is highly conserved throughout the eukaryotic domain. Like ubiquitin, SUMO is covalently attached to lysine side chains in a large number of target proteins. In contrast to ubiquitin, SUMO does not have a direct role in targeting proteins for proteasomal degradation. However, like ubiquitin, SUMO does modulate protein function in a variety of other ways. This includes effects on protein conformation, subcellular localization, and protein–protein interactions. Significant insight into the in vivo role of SUMOylation has been provided by studies in Drosophila that combine genetic manipulation, proteomic, and biochemical analysis. Such studies have revealed that the SUMO conjugation pathway regulates a wide variety of critical cellular and developmental processes, including chromatin/chromosome function, eggshell patterning, embryonic pattern formation, metamorphosis, larval and pupal development, neurogenesis, development of the innate immune system, and apoptosis. This review discusses our current understanding of the diverse roles for SUMO in Drosophila development.

  10. Combining the auxin-inducible degradation system with CRISPR/Cas9-based genome editing for the conditional depletion of endogenous Drosophila melanogaster proteins.

    Science.gov (United States)

    Bence, Melinda; Jankovics, Ferenc; Lukácsovich, Tamás; Erdélyi, Miklós

    2017-04-01

    Inducible protein degradation techniques have considerable advantages over classical genetic approaches, which generate loss-of-function phenotypes at the gene or mRNA level. The plant-derived auxin-inducible degradation system (AID) is a promising technique which enables the degradation of target proteins tagged with the AID motif in nonplant cells. Here, we present a detailed characterization of this method employed during the adult oogenesis of Drosophila. Furthermore, with the help of CRISPR/Cas9-based genome editing, we improve the utility of the AID system in the conditional elimination of endogenously expressed proteins. We demonstrate that the AID system induces efficient and reversible protein depletion of maternally provided proteins both in the ovary and the early embryo. Moreover, the AID system provides a fine spatiotemporal control of protein degradation and allows for the generation of different levels of protein knockdown in a well-regulated manner. These features of the AID system enable the unraveling of the discrete phenotypes of genes with highly complex functions. We utilized this system to generate a conditional loss-of-function allele which allows for the specific degradation of the Vasa protein without affecting its alternative splice variant (solo) and the vasa intronic gene (vig). With the help of this special allele, we demonstrate that dramatic decrease of Vasa protein in the vitellarium does not influence the completion of oogenesis as well as the establishment of proper anteroposterior and dorsoventral polarity in the developing oocyte. Our study suggests that both the localization and the translation of gurken mRNA in the vitellarium is independent from Vasa. © 2017 Federation of European Biochemical Societies.

  11. Global regulation of mRNA translation and stability in the early Drosophila embryo by the Smaug RNA-binding protein.

    Science.gov (United States)

    Chen, Linan; Dumelie, Jason G; Li, Xiao; Cheng, Matthew Hk; Yang, Zhiyong; Laver, John D; Siddiqui, Najeeb U; Westwood, J Timothy; Morris, Quaid; Lipshitz, Howard D; Smibert, Craig A

    2014-01-07

    Smaug is an RNA-binding protein that induces the degradation and represses the translation of mRNAs in the early Drosophila embryo. Smaug has two identified direct target mRNAs that it differentially regulates: nanos and Hsp83. Smaug represses the translation of nanos mRNA but has only a modest effect on its stability, whereas it destabilizes Hsp83 mRNA but has no detectable effect on Hsp83 translation. Smaug is required to destabilize more than one thousand mRNAs in the early embryo, but whether these transcripts represent direct targets of Smaug is unclear and the extent of Smaug-mediated translational repression is unknown. To gain a panoramic view of Smaug function in the early embryo, we identified mRNAs that are bound to Smaug using RNA co-immunoprecipitation followed by hybridization to DNA microarrays. We also identified mRNAs that are translationally repressed by Smaug using polysome gradients and microarrays. Comparison of the bound mRNAs to those that are translationally repressed by Smaug and those that require Smaug for their degradation suggests that a large fraction of Smaug's target mRNAs are both translationally repressed and degraded by Smaug. Smaug directly regulates components of the TRiC/CCT chaperonin, the proteasome regulatory particle and lipid droplets, as well as many metabolic enzymes, including several glycolytic enzymes. Smaug plays a direct and global role in regulating the translation and stability of a large fraction of the mRNAs in the early Drosophila embryo, and has unanticipated functions in control of protein folding and degradation, lipid droplet function and metabolism.

  12. Olfactory receptor signaling is regulated by the post-synaptic density 95, Drosophila discs large, zona-occludens 1 (PDZ) scaffold multi-PDZ domain protein 1.

    LENUS (Irish Health Repository)

    Dooley, Ruth

    2009-12-01

    The unique ability of mammals to detect and discriminate between thousands of different odorant molecules is governed by the diverse array of olfactory receptors expressed by olfactory sensory neurons in the nasal epithelium. Olfactory receptors consist of seven transmembrane domain G protein-coupled receptors and comprise the largest gene superfamily in the mammalian genome. We found that approximately 30% of olfactory receptors possess a classical post-synaptic density 95, Drosophila discs large, zona-occludens 1 (PDZ) domain binding motif in their C-termini. PDZ domains have been established as sites for protein-protein interaction and play a central role in organizing diverse cell signaling assemblies. In the present study, we show that multi-PDZ domain protein 1 (MUPP1) is expressed in the apical compartment of olfactory sensory neurons. Furthermore, on heterologous co-expression with olfactory sensory neurons, MUPP1 was shown to translocate to the plasma membrane. We found direct interaction of PDZ domains 1 + 2 of MUPP1 with the C-terminus of olfactory receptors in vitro. Moreover, the odorant-elicited calcium response of OR2AG1 showed a prolonged decay in MUPP1 small interfering RNA-treated cells. We have therefore elucidated the first building blocks of the putative \\'olfactosome\\

  13. Crystallization and preliminary crystallographic studies of the W2 domain of Drosophila melanogaster eukaryotic translation initiation factor 5C domain-containing protein

    International Nuclear Information System (INIS)

    Zhao, Hui; Wang, Hong; Liu, Huihui; Teng, Maikun; Li, Xu

    2012-01-01

    The crystallization and preliminary crystallographic studies of the carboxy-terminal domain of D. melanogaster eukaryotic translation initiation factor 5C domain-containing protein are reported. The Drosophila melanogaster eukaryotic translation initiation factor 5C domain-containing protein (ECP) is composed of two independently folded domains which belong to the basic leucine-zipper and W2 domain-containing protein (BZW) family. Based on the sequence similarity between the C-terminal W2 domain of ECP and some eukaryotic translation initiation factors (such as eIF2B∊, eIF4γ, eIF5 etc.), ECP has been speculated to participate in the translation initiation process. Structural information on the C-terminal W2 domain of ECP would be helpful in understanding the specific cellular function of this protein. Here, the W2 domain of ECP was expressed and crystallized. Crystals grown by the hanging-drop vapour-diffusion method diffracted to 2.70 Å resolution and belonged to space group I4, with unit-cell parameters a = b = 81.05, c = 57.44 Å. The Matthews coefficient suggested that there was one molecule per asymmetric unit in the crystal

  14. Characterization of big bang, a novel gene encoding for PDZ domain-containing proteins that are dynamically expressed throughout Drosophila development.

    Science.gov (United States)

    Kim, Sabrina Y; Renihan, Maia K; Boulianne, Gabrielle L

    2006-06-01

    PDZ (PSD-95, Discs-large, ZO-1) domain proteins often function as scaffolding proteins and have been shown to play important roles in diverse cellular processes such as the establishment and maintenance of cell polarity, and signal transduction. Here, we report the identification and cloning of a novel Drosophila melanogaster gene that is predicted to produce several different PDZ domain-containing proteins through alternative promoter usage and alternative splicing. This gene, that we have named big bang (bbg), was first identified as C96-GAL4, a GAL4 enhancer trap line that was generated in our lab. To further characterize bbg, its expression pattern was examined in ovaries, embryos, and late third instar larvae using UAS reporter gene constructs, in situ hybridization, or immunocytochemistry. In addition, the expression of alternatively spliced transcripts was examined in more detail using in situ hybridization. We find that during embryogenesis bbg is predominantly expressed in the developing gut, but it is also expressed in external sensory organs found in the epidermis. In the late third instar larva, bbg is expressed along the presumptive wing margin in the wing disc, broadly in the eye disc, and in other imaginal discs as well as in the brain. The expression patterns observed are dynamic and specific during development, suggesting that like other genes that encode for several different PDZ domain protein isoforms, bbg likely plays important roles in multiple developmental processes.

  15. The MAP kinase ERK and its scaffold protein MP1 interact with the chromatin regulator Corto during Drosophila wing tissue development

    Science.gov (United States)

    2011-01-01

    Background Mitogen-activated protein kinase (MAPK) cascades (p38, JNK, ERK pathways) are involved in cell fate acquisition during development. These kinase modules are associated with scaffold proteins that control their activity. In Drosophila, dMP1, that encodes an ERK scaffold protein, regulates ERK signaling during wing development and contributes to intervein and vein cell differentiation. Functional relationships during wing development between a chromatin regulator, the Enhancer of Trithorax and Polycomb Corto, ERK and its scaffold protein dMP1, are examined here. Results Genetic interactions show that corto and dMP1 act together to antagonize rolled (which encodes ERK) in the future intervein cells, thus promoting intervein fate. Although Corto, ERK and dMP1 are present in both cytoplasmic and nucleus compartments, they interact exclusively in nucleus extracts. Furthermore, Corto, ERK and dMP1 co-localize on several sites on polytene chromosomes, suggesting that they regulate gene expression directly on chromatin. Finally, Corto is phosphorylated. Interestingly, its phosphorylation pattern differs between cytoplasm and nucleus and changes upon ERK activation. Conclusions Our data therefore suggest that the Enhancer of Trithorax and Polycomb Corto could participate in regulating vein and intervein genes during wing tissue development in response to ERK signaling. PMID:21401930

  16. The E3 ligase Ubr3 regulates Usher syndrome and MYH9 disorder proteins in the auditory organs of Drosophila and mammals.

    Science.gov (United States)

    Li, Tongchao; Giagtzoglou, Nikolaos; Eberl, Daniel F; Jaiswal, Sonal Nagarkar; Cai, Tiantian; Godt, Dorothea; Groves, Andrew K; Bellen, Hugo J

    2016-06-22

    Myosins play essential roles in the development and function of auditory organs and multiple myosin genes are associated with hereditary forms of deafness. Using a forward genetic screen in Drosophila, we identified an E3 ligase, Ubr3, as an essential gene for auditory organ development. Ubr3 negatively regulates the mono-ubiquitination of non-muscle Myosin II, a protein associated with hearing loss in humans. The mono-ubiquitination of Myosin II promotes its physical interaction with Myosin VIIa, a protein responsible for Usher syndrome type IB. We show that ubr3 mutants phenocopy pathogenic variants of Myosin II and that Ubr3 interacts genetically and physically with three Usher syndrome proteins. The interactions between Myosin VIIa and Myosin IIa are conserved in the mammalian cochlea and in human retinal pigment epithelium cells. Our work reveals a novel mechanism that regulates protein complexes affected in two forms of syndromic deafness and suggests a molecular function for Myosin IIa in auditory organs.

  17. The MAP kinase ERK and its scaffold protein MP1 interact with the chromatin regulator Corto during Drosophila wing tissue development.

    Science.gov (United States)

    Mouchel-Vielh, Emmanuèle; Rougeot, Julien; Decoville, Martine; Peronnet, Frédérique

    2011-03-14

    Mitogen-activated protein kinase (MAPK) cascades (p38, JNK, ERK pathways) are involved in cell fate acquisition during development. These kinase modules are associated with scaffold proteins that control their activity. In Drosophila, dMP1, that encodes an ERK scaffold protein, regulates ERK signaling during wing development and contributes to intervein and vein cell differentiation. Functional relationships during wing development between a chromatin regulator, the Enhancer of Trithorax and Polycomb Corto, ERK and its scaffold protein dMP1, are examined here. Genetic interactions show that corto and dMP1 act together to antagonize rolled (which encodes ERK) in the future intervein cells, thus promoting intervein fate. Although Corto, ERK and dMP1 are present in both cytoplasmic and nucleus compartments, they interact exclusively in nucleus extracts. Furthermore, Corto, ERK and dMP1 co-localize on several sites on polytene chromosomes, suggesting that they regulate gene expression directly on chromatin. Finally, Corto is phosphorylated. Interestingly, its phosphorylation pattern differs between cytoplasm and nucleus and changes upon ERK activation. Our data therefore suggest that the Enhancer of Trithorax and Polycomb Corto could participate in regulating vein and intervein genes during wing tissue development in response to ERK signaling.

  18. The MAP kinase ERK and its scaffold protein MP1 interact with the chromatin regulator Corto during Drosophila wing tissue development

    Directory of Open Access Journals (Sweden)

    Peronnet Frédérique

    2011-03-01

    Full Text Available Abstract Background Mitogen-activated protein kinase (MAPK cascades (p38, JNK, ERK pathways are involved in cell fate acquisition during development. These kinase modules are associated with scaffold proteins that control their activity. In Drosophila, dMP1, that encodes an ERK scaffold protein, regulates ERK signaling during wing development and contributes to intervein and vein cell differentiation. Functional relationships during wing development between a chromatin regulator, the Enhancer of Trithorax and Polycomb Corto, ERK and its scaffold protein dMP1, are examined here. Results Genetic interactions show that corto and dMP1 act together to antagonize rolled (which encodes ERK in the future intervein cells, thus promoting intervein fate. Although Corto, ERK and dMP1 are present in both cytoplasmic and nucleus compartments, they interact exclusively in nucleus extracts. Furthermore, Corto, ERK and dMP1 co-localize on several sites on polytene chromosomes, suggesting that they regulate gene expression directly on chromatin. Finally, Corto is phosphorylated. Interestingly, its phosphorylation pattern differs between cytoplasm and nucleus and changes upon ERK activation. Conclusions Our data therefore suggest that the Enhancer of Trithorax and Polycomb Corto could participate in regulating vein and intervein genes during wing tissue development in response to ERK signaling.

  19. The E3 ligase Ubr3 regulates Usher syndrome and MYH9 disorder proteins in the auditory organs of Drosophila and mammals

    Science.gov (United States)

    Li, Tongchao; Giagtzoglou, Nikolaos; Eberl, Daniel F; Jaiswal, Sonal Nagarkar; Cai, Tiantian; Godt, Dorothea; Groves, Andrew K; Bellen, Hugo J

    2016-01-01

    Myosins play essential roles in the development and function of auditory organs and multiple myosin genes are associated with hereditary forms of deafness. Using a forward genetic screen in Drosophila, we identified an E3 ligase, Ubr3, as an essential gene for auditory organ development. Ubr3 negatively regulates the mono-ubiquitination of non-muscle Myosin II, a protein associated with hearing loss in humans. The mono-ubiquitination of Myosin II promotes its physical interaction with Myosin VIIa, a protein responsible for Usher syndrome type IB. We show that ubr3 mutants phenocopy pathogenic variants of Myosin II and that Ubr3 interacts genetically and physically with three Usher syndrome proteins. The interactions between Myosin VIIa and Myosin IIa are conserved in the mammalian cochlea and in human retinal pigment epithelium cells. Our work reveals a novel mechanism that regulates protein complexes affected in two forms of syndromic deafness and suggests a molecular function for Myosin IIa in auditory organs. DOI: http://dx.doi.org/10.7554/eLife.15258.001 PMID:27331610

  20. Prediction of GNSS satellite clocks

    International Nuclear Information System (INIS)

    Broederbauer, V.

    2010-01-01

    This thesis deals with the characterisation and prediction of GNSS-satellite-clocks. A prerequisite to develop powerful algorithms for the prediction of clock-corrections is the thorough study of the behaviour of the different clock-types of the satellites. In this context the predicted part of the IGU-clock-corrections provided by the Analysis Centers (ACs) of the IGS was compared to the IGS-Rapid-clock solutions to determine reasonable estimates of the quality of already existing well performing predictions. For the shortest investigated interval (three hours) all ACs obtain almost the same accuracy of 0,1 to 0,4 ns. For longer intervals the individual predictions results start to diverge. Thus, for a 12-hours- interval the differences range from nearly 10 ns (GFZ, CODE) until up to some 'tens of ns'. Based on the estimated clock corrections provided via the IGS Rapid products a simple quadratic polynomial turns out to be sufficient to describe the time series of Rubidium-clocks. On the other hand Cesium-clocks show a periodical behaviour (revolution period) with an amplitude of up to 6 ns. A clear correlation between these amplitudes and the Sun elevation angle above the orbital planes can be demonstrated. The variability of the amplitudes is supposed to be caused by temperature-variations affecting the oscillator. To account for this periodical behaviour a quadratic polynomial with an additional sinus-term was finally chosen as prediction model both for the Cesium as well as for the Rubidium clocks. The three polynomial-parameters as well as amplitude and phase shift of the periodic term are estimated within a least-square-adjustment by means of program GNSS-VC/static. Input-data are time series of the observed part of the IGU clock corrections. With the estimated parameters clock-corrections are predicted for various durations. The mean error of the prediction of Rubidium-clock-corrections for an interval of six hours reaches up to 1,5 ns. For the 12-hours

  1. Radioisotope clocks in archaeology

    Energy Technology Data Exchange (ETDEWEB)

    Hedges, R E.M. [Oxford Univ. (UK). Research Lab. for Archaeology

    1979-09-06

    Methods of absolute dating which use the rate of disintegration of a radioactive nucleus as the clock, are reviewed. The use of the abundant radioisotopes (/sup 40/K, Th and U) and of the rare radioisotopes (/sup 14/C, /sup 10/Be, /sup 26/Al, /sup 32/Si, /sup 36/Cl, /sup 41/Ca, /sup 53/Mn) is discussed and radiation integration techniques (fission track dating, thermoluminescence and related techniques) are considered. Specific fields of use of the various methods and their accuracy are examined.

  2. Methodologies for steering clocks

    Science.gov (United States)

    Chadsey, Harold

    1995-01-01

    One of the concerns of the PTTI community is the coordination of one time scale with another. This is accomplished through steering one clock system to another, with a goal of a zero or constant offset in time and frequency. In order to attain this goal, rate differences are calculated and allowed for by the steering algorithm. This paper will present several of these different methods of determining rate differences. Ideally, any change in rate should not cause the offset to change sign (overshoot) by any amount, but certainly not by as much as its previous absolute value. The advantages and disadvantages of each depend on the user's situation.

  3. Clocks and special relativity

    International Nuclear Information System (INIS)

    MacRoberts, D.T.

    1980-01-01

    A kinematic theory without precise definitions of the 'space' and 'time' used is an uninterpreted calculus. The definition of 'time' in special relativity is based on light propagation and the 'constant velocity of light' is a tautological consequence of the definition. When this definition is reified in a 'clock' the phenomenon of 'time dilation' occurs, in terms of the defined time, but is not reciprocal between moving systems; the postulate of relativity is not observed. The new definition of time is compatible with an ether theory without the relativity principle. The derivation of the Lorentz transformations, which requires both postulates, is purely formalistic and is not ontologically sound. (Auth.)

  4. A Light Clock Satisfying the Clock Hypothesis of Special Relativity

    Science.gov (United States)

    West, Joseph

    2007-01-01

    The design of the FMEL, a floor-mirrored Einstein-Langevin "light clock", is introduced. The clock provides a physically intuitive manner to calculate and visualize the time dilation effects for a spatially extended set of observers (an accelerated "frame") undergoing unidirectional acceleration or observers on a rotating cylinder of constant…

  5. Insulator protein Su(Hw) recruits SAGA and Brahma complexes and constitutes part of Origin Recognition Complex-binding sites in the Drosophila genome

    Science.gov (United States)

    Vorobyeva, Nadezhda E.; Mazina, Marina U.; Golovnin, Anton K.; Kopytova, Daria V.; Gurskiy, Dmitriy Y.; Nabirochkina, Elena N.; Georgieva, Sofia G.; Georgiev, Pavel G.; Krasnov, Aleksey N.

    2013-01-01

    Despite increasing data on the properties of replication origins, molecular mechanisms underlying origin recognition complex (ORC) positioning in the genome are still poorly understood. The Su(Hw) protein accounts for the activity of best-studied Drosophila insulators. Here, we show that Su(Hw) recruits the histone acetyltransferase complex SAGA and chromatin remodeler Brahma to Su(Hw)-dependent insulators, which gives rise to regions with low nucleosome density and creates conditions for ORC binding. Depletion in Su(Hw) leads to a dramatic drop in the levels of SAGA, Brahma and ORC subunits and a significant increase in nucleosome density on Su(Hw)-dependent insulators, whereas artificial Su(Hw) recruitment itself is sufficient for subsequent SAGA, Brahma and ORC binding. In contrast to the majority of replication origins that associate with promoters of active genes, Su(Hw)-binding sites constitute a small proportion (6%) of ORC-binding sites that are localized preferentially in transcriptionally inactive chromatin regions termed BLACK and BLUE chromatin. We suggest that the key determinants of ORC positioning in the genome are DNA-binding proteins that constitute different DNA regulatory elements, including insulators, promoters and enhancers. Su(Hw) is the first example of such a protein. PMID:23609538

  6. The PTK7-related transmembrane proteins off-track and off-track 2 are co-receptors for Drosophila Wnt2 required for male fertility.

    Science.gov (United States)

    Linnemannstöns, Karen; Ripp, Caroline; Honemann-Capito, Mona; Brechtel-Curth, Katja; Hedderich, Marie; Wodarz, Andreas

    2014-07-01

    Wnt proteins regulate many developmental processes and are required for tissue homeostasis in adult animals. The cellular responses to Wnts are manifold and are determined by the respective Wnt ligand and its specific receptor complex in the plasma membrane. Wnt receptor complexes contain a member of the Frizzled family of serpentine receptors and a co-receptor, which commonly is a single-pass transmembrane protein. Vertebrate protein tyrosine kinase 7 (PTK7) was identified as a Wnt co-receptor required for control of planar cell polarity (PCP) in frogs and mice. We found that flies homozygous for a complete knock-out of the Drosophila PTK7 homolog off track (otk) are viable and fertile and do not show PCP phenotypes. We discovered an otk paralog (otk2, CG8964), which is co-expressed with otk throughout embryonic and larval development. Otk and Otk2 bind to each other and form complexes with Frizzled, Frizzled2 and Wnt2, pointing to a function as Wnt co-receptors. Flies lacking both otk and otk2 are viable but male sterile due to defective morphogenesis of the ejaculatory duct. Overexpression of Otk causes female sterility due to malformation of the oviduct, indicating that Otk and Otk2 are specifically involved in the sexually dimorphic development of the genital tract.

  7. The PTK7-related transmembrane proteins off-track and off-track 2 are co-receptors for Drosophila Wnt2 required for male fertility.

    Directory of Open Access Journals (Sweden)

    Karen Linnemannstöns

    2014-07-01

    Full Text Available Wnt proteins regulate many developmental processes and are required for tissue homeostasis in adult animals. The cellular responses to Wnts are manifold and are determined by the respective Wnt ligand and its specific receptor complex in the plasma membrane. Wnt receptor complexes contain a member of the Frizzled family of serpentine receptors and a co-receptor, which commonly is a single-pass transmembrane protein. Vertebrate protein tyrosine kinase 7 (PTK7 was identified as a Wnt co-receptor required for control of planar cell polarity (PCP in frogs and mice. We found that flies homozygous for a complete knock-out of the Drosophila PTK7 homolog off track (otk are viable and fertile and do not show PCP phenotypes. We discovered an otk paralog (otk2, CG8964, which is co-expressed with otk throughout embryonic and larval development. Otk and Otk2 bind to each other and form complexes with Frizzled, Frizzled2 and Wnt2, pointing to a function as Wnt co-receptors. Flies lacking both otk and otk2 are viable but male sterile due to defective morphogenesis of the ejaculatory duct. Overexpression of Otk causes female sterility due to malformation of the oviduct, indicating that Otk and Otk2 are specifically involved in the sexually dimorphic development of the genital tract.

  8. Sumoylation of the Plant Clock Transcription Factor CCA1 Suppresses DNA Binding

    NARCIS (Netherlands)

    Hansen, L.L.; Imrie, L.; Le Bihan, T.; van den Burg, H.A.; van Ooijen, G.

    2017-01-01

    In plants, the circadian clock regulates the expression of one-third of all transcripts and is crucial to virtually every aspect of metabolism and growth. We now establish sumoylation, a posttranslational protein modification, as a novel regulator of the key clock protein CCA1 in the model plant

  9. Minimal tool set for a prokaryotic circadian clock.

    Science.gov (United States)

    Schmelling, Nicolas M; Lehmann, Robert; Chaudhury, Paushali; Beck, Christian; Albers, Sonja-Verena; Axmann, Ilka M; Wiegard, Anika

    2017-07-21

    Circadian clocks are found in organisms of almost all domains including photosynthetic Cyanobacteria, whereby large diversity exists within the protein components involved. In the model cyanobacterium Synechococcus elongatus PCC 7942 circadian rhythms are driven by a unique KaiABC protein clock, which is embedded in a network of input and output factors. Homologous proteins to the KaiABC clock have been observed in Bacteria and Archaea, where evidence for circadian behavior in these domains is accumulating. However, interaction and function of non-cyanobacterial Kai-proteins as well as homologous input and output components remain mainly unclear. Using a universal BLAST analyses, we identified putative KaiC-based timing systems in organisms outside as well as variations within Cyanobacteria. A systematic analyses of publicly available microarray data elucidated interesting variations in circadian gene expression between different cyanobacterial strains, which might be correlated to the diversity of genome encoded clock components. Based on statistical analyses of co-occurrences of the clock components homologous to Synechococcus elongatus PCC 7942, we propose putative networks of reduced and fully functional clock systems. Further, we studied KaiC sequence conservation to determine functionally important regions of diverged KaiC homologs. Biochemical characterization of exemplary cyanobacterial KaiC proteins as well as homologs from two thermophilic Archaea demonstrated that kinase activity is always present. However, a KaiA-mediated phosphorylation is only detectable in KaiC1 orthologs. Our analysis of 11,264 genomes clearly demonstrates that components of the Synechococcus elongatus PCC 7942 circadian clock are present in Bacteria and Archaea. However, all components are less abundant in other organisms than Cyanobacteria and KaiA, Pex, LdpA, and CdpA are only present in the latter. Thus, only reduced KaiBC-based or even simpler, solely KaiC-based timing systems

  10. An optical clock to go

    Science.gov (United States)

    Ludlow, Andrew D.

    2018-05-01

    Bringing next-generation atomic clocks out of the lab is not an easy task, but doing so will unlock many new possibilities. As a crucial first step, a portable atomic clock has now been deployed for relativistic geodesy measurements in the Alps.

  11. Ultradian rhythm unmasked in the Pdf clock mutant of Drosophila

    Indian Academy of Sciences (India)

    2014-07-20

    Ohtomo et al. .... stand the influence of these two phases, short sampling segments were ... per-long (perL) mutants also showed ultradian rhythms un- der L:L ..... would like to dedicate this paper to the memory of Dr Obaid. Siddiqi.

  12. Small heat shock proteins mediate cell-autonomous and -nonautonomous protection in a Drosophila model for environmental-stress-induced degeneration.

    Science.gov (United States)

    Kawasaki, Fumiko; Koonce, Noelle L; Guo, Linda; Fatima, Shahroz; Qiu, Catherine; Moon, Mackenzie T; Zheng, Yunzhen; Ordway, Richard W

    2016-09-01

    Cell and tissue degeneration, and the development of degenerative diseases, are influenced by genetic and environmental factors that affect protein misfolding and proteotoxicity. To better understand the role of the environment in degeneration, we developed a genetic model for heat shock (HS)-stress-induced degeneration in Drosophila This model exhibits a unique combination of features that enhance genetic analysis of degeneration and protection mechanisms involving environmental stress. These include cell-type-specific failure of proteostasis and degeneration in response to global stress, cell-nonautonomous interactions within a simple and accessible network of susceptible cell types, and precise temporal control over the induction of degeneration. In wild-type flies, HS stress causes selective loss of the flight ability and degeneration of three susceptible cell types comprising the flight motor: muscle, motor neurons and associated glia. Other motor behaviors persist and, accordingly, the corresponding cell types controlling leg motor function are resistant to degeneration. Flight motor degeneration was preceded by a failure of muscle proteostasis characterized by diffuse ubiquitinated protein aggregates. Moreover, muscle-specific overexpression of a small heat shock protein (HSP), HSP23, promoted proteostasis and protected muscle from HS stress. Notably, neurons and glia were protected as well, indicating that a small HSP can mediate cell-nonautonomous protection. Cell-autonomous protection of muscle was characterized by a distinct distribution of ubiquitinated proteins, including perinuclear localization and clearance of protein aggregates associated with the perinuclear microtubule network. This network was severely disrupted in wild-type preparations prior to degeneration, suggesting that it serves an important role in muscle proteostasis and protection. Finally, studies of resistant leg muscles revealed that they sustain proteostasis and the microtubule

  13. echinus, required for interommatidial cell sorting and cell death in the Drosophila pupal retina, encodes a protein with homology to ubiquitin-specific proteases

    Directory of Open Access Journals (Sweden)

    Gorski Sharon M

    2007-07-01

    Full Text Available Abstract Background Programmed cell death is used to remove excess cells between ommatidia in the Drosophila pupal retina. This death is required to establish the crystalline, hexagonal packing of ommatidia that characterizes the adult fly eye. In previously described echinus mutants, interommatidial cell sorting, which precedes cell death, occurred relatively normally. Interommatidial cell death was partially suppressed, resulting in adult eyes that contained excess pigment cells, and in which ommatidia were mildly disordered. These results have suggested that echinus functions in the pupal retina primarily to promote interommatidial cell death. Results We generated a number of new echinus alleles, some likely null mutants. Analysis of these alleles provides evidence that echinus has roles in cell sorting as well as cell death. echinus encodes a protein with homology to ubiquitin-specific proteases. These proteins cleave ubiquitin-conjugated proteins at the ubiquitin C-terminus. The echinus locus encodes multiple splice forms, including two proteins that lack residues thought to be critical for deubiquitination activity. Surprisingly, ubiquitous expression in the eye of versions of Echinus that lack residues critical for ubiquitin specific protease activity, as well as a version predicted to be functional, rescue the echinus loss-of-function phenotype. Finally, genetic interactions were not detected between echinus loss and gain-of-function and a number of known apoptotic regulators. These include Notch, EGFR, the caspases Dronc, Drice, Dcp-1, Dream, the caspase activators, Rpr, Hid, and Grim, the caspase inhibitor DIAP1, and Lozenge or Klumpfuss. Conclusion The echinus locus encodes multiple splice forms of a protein with homology to ubiquitin-specific proteases, but protease activity is unlikely to be required for echinus function, at least when echinus is overexpressed. Characterization of likely echinus null alleles and genetic interactions

  14. Shining a light on the Arabidopsis circadian clock.

    Science.gov (United States)

    Oakenfull, Rachael J; Davis, Seth J

    2017-11-01

    The circadian clock provides essential timing information to ensure optimal growth to prevailing external environmental conditions. A major time-setting mechanism (zeitgeber) in clock synchronization is light. Differing light wavelengths, intensities, and photoperiodic duration are processed for the clock-setting mechanism. Many studies on light-input pathways to the clock have focused on Arabidopsis thaliana. Photoreceptors are specific chromic proteins that detect light signals and transmit this information to the central circadian oscillator through a number of different signalling mechanisms. The most well-characterized clock-mediating photoreceptors are cryptochromes and phytochromes, detecting blue, red, and far-red wavelengths of light. Ultraviolet and shaded light are also processed signals to the oscillator. Notably, the clock reciprocally generates rhythms of photoreceptor action leading to so-called gating of light responses. Intermediate proteins, such as Phytochrome interacting factors (PIFs), constitutive photomorphogenic 1 (COP1) and EARLY FLOWERING 3 (ELF3), have been established in signalling pathways downstream of photoreceptor activation. However, the precise details for these signalling mechanisms are not fully established. This review highlights both historical and recent efforts made to understand overall light input to the oscillator, first looking at how each wavelength of light is detected, this is then related to known input mechanisms and their interactions. © 2017 John Wiley & Sons Ltd.

  15. High Precision Clock Bias Prediction Model in Clock Synchronization System

    Directory of Open Access Journals (Sweden)

    Zan Liu

    2016-01-01

    Full Text Available Time synchronization is a fundamental requirement for many services provided by a distributed system. Clock calibration through the time signal is the usual way to realize the synchronization among the clocks used in the distributed system. The interference to time signal transmission or equipment failures may bring about failure to synchronize the time. To solve this problem, a clock bias prediction module is paralleled in the clock calibration system. And for improving the precision of clock bias prediction, the first-order grey model with one variable (GM(1,1 model is proposed. In the traditional GM(1,1 model, the combination of parameters determined by least squares criterion is not optimal; therefore, the particle swarm optimization (PSO is used to optimize GM(1,1 model. At the same time, in order to avoid PSO getting stuck at local optimization and improve its efficiency, the mechanisms that double subgroups and nonlinear decreasing inertia weight are proposed. In order to test the precision of the improved model, we design clock calibration experiments, where time signal is transferred via radio and wired channel, respectively. The improved model is built on the basis of clock bias acquired in the experiments. The results show that the improved model is superior to other models both in precision and in stability. The precision of improved model increased by 66.4%~76.7%.

  16. Aspergillus nidulans Synthesize Insect Juvenile Hormones upon Expression of a Heterologous Regulatory Protein and in Response to Grazing by Drosophila melanogaster Larvae

    DEFF Research Database (Denmark)

    Nielsen, Morten Thrane; Klejnstrup, Marie Louise; Rohlfs, Marko

    2013-01-01

    , indicating that fungal secondary metabolites remain an underexplored resource of bioactive molecules. In this study, we combine heterologous expression of regulatory proteins in Aspergillus nidulans with systematic variation of growth conditions and observe induced synthesis of insect juvenile hormone......-III and methyl farnesoate. Both compounds are sesquiterpenes belonging to the juvenile hormone class. Juvenile hormones regulate developmental and metabolic processes in insects and crustaceans, but have not previously been reported as fungal metabolites. We found that feeding by Drosophila melanogaster larvae...

  17. Clocks around Sgr A*

    Science.gov (United States)

    Angélil, Raymond; Saha, Prasenjit

    2014-11-01

    The S stars near the Galactic Centre and any pulsars that may be on similar orbits can be modelled in a unified way as clocks orbiting a black hole, and hence are potential probes of relativistic effects, including black hole spin. The high eccentricities of many S stars mean that relativistic effects peak strongly around pericentre; for example, orbit precession is not a smooth effect but almost a kick at pericentre. We argue that concentration around pericentre will be an advantage when analysing redshift or pulse-arrival data to measure relativistic effects, because cumulative precession will be drowned out by Newtonian perturbations from other mass in the Galactic Centre region. Wavelet decomposition may be a way to disentangle relativistic effects from Newton perturbations. Assuming a plausible model for Newtonian perturbations on S2, relativity appears to be strongest in a two-year interval around pericentre, in wavelet modes of time-scale ≈6 months.

  18. Rasputin, the Drosophila homologue of the RasGAP SH3 binding protein, functions in ras- and Rho-mediated signaling.

    Science.gov (United States)

    Pazman, C; Mayes, C A; Fanto, M; Haynes, S R; Mlodzik, M

    2000-04-01

    The small GTPase Ras plays an important role in many cellular signaling processes. Ras activity is negatively regulated by GTPase activating proteins (GAPs). It has been proposed that RasGAP may also function as an effector of Ras activity. We have identified and characterized the Drosophila homologue of the RasGAP-binding protein G3BP encoded by rasputin (rin). rin mutants are viable and display defects in photoreceptor recruitment and ommatidial polarity in the eye. Mutations in rin/G3BP genetically interact with components of the Ras signaling pathway that function at the level of Ras and above, but not with Raf/MAPK pathway components. These interactions suggest that Rin is required as an effector in Ras signaling during eye development, supporting an effector role for RasGAP. The ommatidial polarity phenotypes of rin are similar to those of RhoA and the polarity genes, e.g. fz and dsh. Although rin/G3BP interacts genetically with RhoA, affecting both photoreceptor differentiation and polarity, it does not interact with the gain-of-function genotypes of fz and dsh. These data suggest that Rin is not a general component of polarity generation, but serves a function specific to Ras and RhoA signaling pathways.

  19. Oligomeric structure and chaperone-like activity of Drosophila melanogaster mitochondrial small heat shock protein Hsp22 and arginine mutants in the alpha-crystallin domain.

    Science.gov (United States)

    Dabbaghizadeh, Afrooz; Finet, Stéphanie; Morrow, Genevieve; Moutaoufik, Mohamed Taha; Tanguay, Robert M

    2017-07-01

    The structure and chaperone function of DmHsp22WT, a small Hsp of Drosophila melanogaster localized within mitochondria were examined. Mutations of conserved arginine mutants within the alpha-crystallin domain (ACD) domain (R105G, R109G, and R110G) were introduced, and their effects on oligomerization and chaperone function were assessed. Arginine to glycine mutations do not induce significant changes in tryptophan fluorescence, and the mutated proteins form oligomers that are of equal or smaller size than the wild-type protein. They all form oligomer with one single peak as determined by size exclusion chromatography. While all mutants demonstrate the same efficiency as the DmHsp22WT in a DTT-induced insulin aggregation assay, all are more efficient chaperones to prevent aggregation of malate dehydrogenase. Arginine mutants of DmHsp22 are efficient chaperones to retard aggregation of CS and Luc. In summary, this study shows that mutations of arginine to glycine in DmHsp22 ACD induce a number of structural changes, some of which differ from those described in mammalian sHsps. Interestingly, only the R110G-DmHsp22 mutant, and not the expected R109G equivalent to human R140-HspB1, R116-HspB4, and R120-HspB5, showed different structural properties compared with the DmHsp22WT.

  20. Oligomerization and chaperone-like activity of Drosophila melanogaster small heat shock protein DmHsp27 and three arginine mutants in the alpha-crystallin domain.

    Science.gov (United States)

    Moutaoufik, Mohamed Taha; Morrow, Geneviève; Maaroufi, Halim; Férard, Céline; Finet, Stéphanie; Tanguay, Robert M

    2017-07-01

    The small Hsp DmHsp27 from Drosophila melanogaster is one of the few small heat shock proteins (sHsps) found within the nucleus. We report that its dimerization is independent of disulfide bond formation and seems to rely on salt bridges. Unlike metazoan sHsps, DmHsp27 forms two populations of oligomers not in equilibrium. Mutations at highly conserved arginine residues in mammalian sHsps have been reported to be associated with protein conformational defects and intracellular aggregation. Independent mutation of three highly conserved arginines (R122, R131, and R135) to glycine in DmHsp27 results in only one population of higher molecular weight form. In vitro, the chaperone-like activity of wild-type DmHsp27 was comparable with that of its two isolated populations and to the single population of the R122G, R131G, and R135G using luciferase as substrate. However, using insulin, the chaperone-like activity of wild-type DmHsp27 was lower than that of R122G and R131G mutants. Altogether, the results characterize wild-type DmHsp27 and its alpha-crystallin domain (ACD) arginine mutants and may give insight into protection mechanism of sHsps.

  1. Identification of Atg2 and ArfGAP1 as Candidate Genetic Modifiers of the Eye Pigmentation Phenotype of Adaptor Protein-3 (AP-3) Mutants in Drosophila melanogaster.

    Science.gov (United States)

    Rodriguez-Fernandez, Imilce A; Dell'Angelica, Esteban C

    2015-01-01

    The Adaptor Protein (AP)-3 complex is an evolutionary conserved, molecular sorting device that mediates the intracellular trafficking of proteins to lysosomes and related organelles. Genetic defects in AP-3 subunits lead to impaired biogenesis of lysosome-related organelles (LROs) such as mammalian melanosomes and insect eye pigment granules. In this work, we have performed a forward screening for genetic modifiers of AP-3 function in the fruit fly, Drosophila melanogaster. Specifically, we have tested collections of large multi-gene deletions--which together covered most of the autosomal chromosomes-to identify chromosomal regions that, when deleted in single copy, enhanced or ameliorated the eye pigmentation phenotype of two independent AP-3 subunit mutants. Fine-mapping led us to define two non-overlapping, relatively small critical regions within fly chromosome 3. The first critical region included the Atg2 gene, which encodes a conserved protein involved in autophagy. Loss of one functional copy of Atg2 ameliorated the pigmentation defects of mutants in AP-3 subunits as well as in two other genes previously implicated in LRO biogenesis, namely Blos1 and lightoid, and even increased the eye pigment content of wild-type flies. The second critical region included the ArfGAP1 gene, which encodes a conserved GTPase-activating protein with specificity towards GTPases of the Arf family. Loss of a single functional copy of the ArfGAP1 gene ameliorated the pigmentation phenotype of AP-3 mutants but did not to modify the eye pigmentation of wild-type flies or mutants in Blos1 or lightoid. Strikingly, loss of the second functional copy of the gene did not modify the phenotype of AP-3 mutants any further but elicited early lethality in males and abnormal eye morphology when combined with mutations in Blos1 and lightoid, respectively. These results provide genetic evidence for new functional links connecting the machinery for biogenesis of LROs with molecules implicated in

  2. Physiological links of circadian clock and biological clock of aging.

    Science.gov (United States)

    Liu, Fang; Chang, Hung-Chun

    2017-07-01

    Circadian rhythms orchestrate biochemical and physiological processes in living organisms to respond the day/night cycle. In mammals, nearly all cells hold self-sustained circadian clocks meanwhile couple the intrinsic rhythms to systemic changes in a hierarchical manner. The suprachiasmatic nucleus (SCN) of the hypothalamus functions as the master pacemaker to initiate daily synchronization according to the photoperiod, in turn determines the phase of peripheral cellular clocks through a variety of signaling relays, including endocrine rhythms and metabolic cycles. With aging, circadian desynchrony occurs at the expense of peripheral metabolic pathologies and central neurodegenerative disorders with sleep symptoms, and genetic ablation of circadian genes in model organisms resembled the aging-related features. Notably, a number of studies have linked longevity nutrient sensing pathways in modulating circadian clocks. Therapeutic strategies that bridge the nutrient sensing pathways and circadian clock might be rational designs to defy aging.

  3. Identification of four Drosophila allatostatins as the cognate ligands for the Drosophila orphan receptor DAR-2

    DEFF Research Database (Denmark)

    Lenz, C; Williamson, M; Hansen, G N

    2001-01-01

    The allatostatins are generally inhibitory insect neuropeptides. The Drosophila orphan receptor DAR-2 is a G-protein-coupled receptor, having 47% amino acid residue identity with another Drosophila receptor, DAR-1 (which is also called dros. GPCR, or DGR) that was previously shown...... to be the receptor for an intrinsic Drosophila A-type (cockroach-type) allatostatin. Here, we have permanently expressed DAR-2 in CHO cells and found that it is the cognate receptor for four Drosophila A-type allatostatins, the drostatins-A1 to -A4. Of all the drostatins, drostatin-A4 (Thr...... weakly in the brain. The Drosophila larval gut also contains about 20-30 endocrine cells, expressing the gene for the drostatins-A1 to -A4. We suggest, therefore, that DAR-2 mediates an allatostatin (drostatin)-induced inhibition of gut motility. This is the first report on the permanent and functional...

  4. Heterochromatin protein 1 (HP1a positively regulates euchromatic gene expression through RNA transcript association and interaction with hnRNPs in Drosophila.

    Directory of Open Access Journals (Sweden)

    Lucia Piacentini

    2009-10-01

    Full Text Available Heterochromatin Protein 1 (HP1a is a well-known conserved protein involved in heterochromatin formation and gene silencing in different species including humans. A general model has been proposed for heterochromatin formation and epigenetic gene silencing in different species that implies an essential role for HP1a. According to the model, histone methyltransferase enzymes (HMTases methylate the histone H3 at lysine 9 (H3K9me, creating selective binding sites for itself and the chromodomain of HP1a. This complex is thought to form a higher order chromatin state that represses gene activity. It has also been found that HP1a plays a role in telomere capping. Surprisingly, recent studies have shown that HP1a is present at many euchromatic sites along polytene chromosomes of Drosophila melanogaster, including the developmental and heat-shock-induced puffs, and that this protein can be removed from these sites by in vivo RNase treatment, thus suggesting an association of HP1a with the transcripts of many active genes. To test this suggestion, we performed an extensive screening by RIP-chip assay (RNA-immunoprecipitation on microarrays, and we found that HP1a is associated with transcripts of more than one hundred euchromatic genes. An expression analysis in HP1a mutants shows that HP1a is required for positive regulation of these genes. Cytogenetic and molecular assays show that HP1a also interacts with the well known proteins DDP1, HRB87F, and PEP, which belong to different classes of heterogeneous nuclear ribonucleoproteins (hnRNPs involved in RNA processing. Surprisingly, we found that all these hnRNP proteins also bind heterochromatin and are dominant suppressors of position effect variegation. Together, our data show novel and unexpected functions for HP1a and hnRNPs proteins. All these proteins are in fact involved both in RNA transcript processing and in heterochromatin formation. This suggests that, in general, similar epigenetic mechanisms

  5. Micro Mercury Ion Clock (MMIC)

    Data.gov (United States)

    National Aeronautics and Space Administration — Demonstrate micro clock based on trapped Hg ions with more than 10x size reduction and power; Fractional frequency stability at parts per 1014 level, adequate for...

  6. History of early atomic clocks

    International Nuclear Information System (INIS)

    Ramsey, N.F.

    2005-01-01

    This review of the history of early atomic clocks includes early atomic beam magnetic resonance, methods of separated and successive oscillatory fields, microwave absorption, optical pumping and atomic masers. (author)

  7. Controllable clock circuit design in PEM system

    International Nuclear Information System (INIS)

    Sun Yunhua; Wang Peihua; Hu Tingting; Feng Baotong; Shuai Lei; Huang Huan; Wei Shujun; Li Ke; Zhao Jingwei; Wei Long

    2011-01-01

    A high-precision synchronized clock circuit design will be presented, which can supply steady, reliable and anti-jamming clock signal for the data acquirement (DAQ) system of Positron Emission Mammography (PEM). This circuit design is based on the Single-Chip Microcomputer and high-precision clock chip, and can achieve multiple controllable clock signals. The jamming between the clock signals can be reduced greatly with the differential transmission. Meanwhile, the adoption of CAN bus control in the clock circuit can prompt the clock signals to be transmitted or masked simultaneously when needed. (authors)

  8. Controllable clock circuit design in PEM system

    International Nuclear Information System (INIS)

    Sun Yunhua; Wang Peilin; Hu Tingting; Feng Baotong; Shuai Lei; Huang Huan; Wei Shujun; Li Ke; Zhao Jingwei; Wei Long

    2010-01-01

    A high-precision synchronized clock circuit design will be presented, which can supply steady, reliable and anti-jamming clock signal for the data acquirement (DAQ) system of Positron Emission Mammography (PEM). This circuit design is based on the Single-Chip Microcomputer and high-precision clock chip, and can achieve multiple controllable clock signals. The jamming between the clock signals can be reduced greatly with the differential transmission. Meanwhile, the adoption of CAN bus control in the clock circuit can prompt the clock signals to be transmitted or masked simultaneously when needed. (authors)

  9. Time without clocks - an attempt

    International Nuclear Information System (INIS)

    Karpman, G.

    1978-01-01

    A definition of time intervals separating two states of systems of elementary particles and observers is attempted. The definition is founded on the notion of instant state of the system and uses no information connected with the use of a clock. Applying the definition to a classical clock and to a sample of unstable particles, results are obtained in agreement with experiment. However, if the system contains 'few' elementary particles, the properties of the time interval present some different features. (author)

  10. Physical Layer Ethernet Clock Synchronization

    Science.gov (United States)

    2010-11-01

    42 nd Annual Precise Time and Time Interval (PTTI) Meeting 77 PHYSICAL LAYER ETHERNET CLOCK SYNCHRONIZATION Reinhard Exel, Georg...oeaw.ac.at Nikolaus Kerö Oregano Systems, Mohsgasse 1, 1030 Wien, Austria E-mail: nikolaus.keroe@oregano.at Abstract Clock synchronization ...is a service widely used in distributed networks to coordinate data acquisition and actions. As the requirement to achieve tighter synchronization

  11. Glial-Specific Functions of Microcephaly Protein WDR62 and Interaction with the Mitotic Kinase AURKA Are Essential for Drosophila Brain Growth.

    Science.gov (United States)

    Lim, Nicholas R; Shohayeb, Belal; Zaytseva, Olga; Mitchell, Naomi; Millard, S Sean; Ng, Dominic C H; Quinn, Leonie M

    2017-07-11

    The second most commonly mutated gene in primary microcephaly (MCPH) patients is wd40-repeat protein 62 (wdr62), but the relative contribution of WDR62 function to the growth of major brain lineages is unknown. Here, we use Drosophila models to dissect lineage-specific WDR62 function(s). Interestingly, although neural stem cell (neuroblast)-specific depletion of WDR62 significantly decreased neuroblast number, brain size was unchanged. In contrast, glial lineage-specific WDR62 depletion significantly decreased brain volume. Moreover, loss of function in glia not only decreased the glial population but also non-autonomously caused neuroblast loss. We further demonstrated that WDR62 controls brain growth through lineage-specific interactions with master mitotic signaling kinase, AURKA. Depletion of AURKA in neuroblasts drives brain overgrowth, which was suppressed by WDR62 co-depletion. In contrast, glial-specific depletion of AURKA significantly decreased brain volume, which was further decreased by WDR62 co-depletion. Thus, dissecting relative contributions of MCPH factors to individual neural lineages will be critical for understanding complex diseases such as microcephaly. Crown Copyright © 2017. Published by Elsevier Inc. All rights reserved.

  12. The light gene of Drosophila melanogaster encodes a homologue of VPS41, a yeast gene involved in cellular-protein trafficking.

    Science.gov (United States)

    Warner, T S; Sinclair, D A; Fitzpatrick, K A; Singh, M; Devlin, R H; Honda, B M

    1998-04-01

    Mutations in a number of genes affect eye colour in Drosophila melanogaster; some of these "eye-colour" genes have been shown to be involved in various aspects of cellular transport processes. In addition, combinations of viable mutant alleles of some of these genes, such as carnation (car) combined with either light (lt) or deep-orange (dor) mutants, show lethal interactions. Recently, dor was shown to be homologous to the yeast gene PEP3 (VPS18), which is known to be involved in intracellular trafficking. We have undertaken to extend our earlier work on the lt gene, in order to examine in more detail its expression pattern and to characterize its gene product via sequencing of a cloned cDNA. The gene appears to be expressed at relatively high levels in all stages and tissues examined, and shows strong homology to VPS41, a gene involved in cellular-protein trafficking in yeast and higher eukaryotes. Further genetic experiments also point to a role for lt in transport processes: we describe lethal interactions between viable alleles of lt and dor, as well as phenotypic interactions (reductions in eye pigment) between allels of lt and another eye-colour gene, garnet (g), whose gene product has close homology to a subunit of the human adaptor complex, AP-3.

  13. Identification and characterization of the fibrinogen-like domain of fibrinogen-related proteins in the mosquito, Anopheles gambiae, and the fruitfly, Drosophila melanogaster, genomes

    Directory of Open Access Journals (Sweden)

    Zhao Qin

    2005-09-01

    Full Text Available Abstract Background The fibrinogen-like (FBG domain, which consists of approximately 200 amino acid residues, has high sequence similarity to the C-terminal halves of fibrinogen β and γ chains. Fibrinogen-related proteins (FREPs, which contain FBG domains in their C-terminal region, are found universally in vertebrates and invertebrates. In invertebrates, FREPs are involved in immune responses and other aspects of physiology. To understand the complexity of this family in insects, we analyzed FREPs in the mosquito genome and made comparisons to FREPs in the fruitfly genome. Results By using the genome data of the mosquito, Anopheles gambiae, 53 FREPs were identified, whereas only 20 members were found in the Drosophila melanogaster genome. Using sequence profile analysis, we found that FBG domains have high sequence similarity and are highly conserved throughout the FBG domain region. By secondary structure analysis and comparison, the FBG domains of FREPs are predicted to function in recognition of carbohydrates and their derivatives on the surface of microorganisms in innate immunity. Conclusion Detailed sequence and structural analysis discloses that the FREP family contains FBG domains that have high sequence similarity in the A. gambiae genome. Expansion of the FREP family in mosquitoes during evolutionary history is mainly accounted for by a major expansion of the FBG domain architecture. The characterization of the FBG domains in the FREP family is likely to aid in the experimental analysis of the ability of mosquitoes to recognize parasites in innate immunity and physiologies associated with blood feeding.

  14. Transfection of Chinese hamster ovary DHFR/sup -/ cells with the gene coding for heat shock protein 70 from drosophila melanogaster

    International Nuclear Information System (INIS)

    Duffy, J.J.; Carper, S.W.; Gerner, E.W.

    1987-01-01

    Chinese hamster ovary DHFR/sup -/ cells (CHO-DHFR/sup -/) were transfected with the plasmid pSV2-dhfr expressing the mouse gene coding for dhfr or with the same plasmid containing the gene coding for the Drosophila melanogaster heat shock protein 70 (hsp70), pSVd-hsp70. Three subcloned cell lines selected for expression of the dhfr gene were shown to contain either the vector sequence (G cells) or varying copies of pSVd-hsp70 (H cells). One line of H cells was shown to contain > 30 copies of the D. melanogaster hsp70 gene and to express the hsp70 RNA at significant levels. No difference between G and H cells was observed in the rate of growth, in the development of thermotolerance, or in the sensitivity of actin microfilament bundles to heat shock. However, H cells containing the transfected hsp70 gene had an altered morphology when compared to the G cells and the parental CHO-DHFR/sup -/ cells being more fibroblastic. The adhesion properties of the H cells was also decreased when compared to the G cells. These results show that insertion of the D. melanogaster gene into CHO cells does not effect growth rates or heat shock responses but may alter cell morphology and adhesion

  15. Ubiquitous overexpression of a transgene encoding the extracellular portion of the Drosophila roughest-irregular chiasm C protein induces early embryonic lethality.

    Science.gov (United States)

    Moda, L; Machado, R C; Ramos, R G

    2000-09-01

    The cell adhesion molecule Rst-irreC is a transmembrane glycoprotein of the immunoglobulin superfamily involved in several important developmental processes in Drosophila, including axonal pathfinding in the optic lobe and programmed cell death and pigment cell differentiation in the pupal retina. As an initial step towards the "in vivo" functional analysis of this protein we have generated transgenic fly stocks carrying a truncated cDNA construct encoding only the extracellular domain of Rst-IrreC under the transcriptional control of the heat shock inducible promoter hsp70. We show that heat-shocking embryos bearing the transgene during the first 8hs of development lead to a 3-4 fold reduction in their viability compared to wild type controls. The embryonic lethality can already be produced by applying the heat pulse in the first 3hs of embryonic development, does not seem to be suppressed in the absence of wildtype product and is progressively reduced as the heat treatment is applied later in embryogenesis. These results are compatible with the hypothesis of the lethal phenotype being primarily due to heterophilic interactions between Rst-IrreC extracellular domain and an yet unknown ligand.

  16. Rhythmic Behavior Is Controlled by the SRm160 Splicing Factor in Drosophila melanogaster.

    Science.gov (United States)

    Beckwith, Esteban J; Hernando, Carlos E; Polcowñuk, Sofía; Bertolin, Agustina P; Mancini, Estefania; Ceriani, M Fernanda; Yanovsky, Marcelo J

    2017-10-01

    Circadian clocks organize the metabolism, physiology, and behavior of organisms throughout the day-night cycle by controlling daily rhythms in gene expression at the transcriptional and post-transcriptional levels. While many transcription factors underlying circadian oscillations are known, the splicing factors that modulate these rhythms remain largely unexplored. A genome-wide assessment of the alterations of gene expression in a null mutant of the alternative splicing regulator SR-related matrix protein of 160 kDa (SRm160) revealed the extent to which alternative splicing impacts on behavior-related genes. We show that SRm160 affects gene expression in pacemaker neurons of the Drosophila brain to ensure proper oscillations of the molecular clock. A reduced level of SRm160 in adult pacemaker neurons impairs circadian rhythms in locomotor behavior, and this phenotype is caused, at least in part, by a marked reduction in period ( per ) levels. Moreover, rhythmic accumulation of the neuropeptide PIGMENT DISPERSING FACTOR in the dorsal projections of these neurons is abolished after SRm160 depletion. The lack of rhythmicity in SRm160-downregulated flies is reversed by a fully spliced per construct, but not by an extra copy of the endogenous locus, showing that SRm160 positively regulates per levels in a splicing-dependent manner. Our findings highlight the significant effect of alternative splicing on the nervous system and particularly on brain function in an in vivo model. Copyright © 2017 by the Genetics Society of America.

  17. Apoptosis in Drosophila: which role for mitochondria?

    Science.gov (United States)

    Clavier, Amandine; Rincheval-Arnold, Aurore; Colin, Jessie; Mignotte, Bernard; Guénal, Isabelle

    2016-03-01

    It is now well established that the mitochondrion is a central regulator of mammalian cell apoptosis. However, the importance of this organelle in non-mammalian apoptosis has long been regarded as minor, mainly because of the absence of a crucial role for cytochrome c in caspase activation. Recent results indicate that the control of caspase activation and cell death in Drosophila occurs at the mitochondrial level. Numerous proteins, including RHG proteins and proteins of the Bcl-2 family that are key regulators of Drosophila apoptosis, constitutively or transiently localize in mitochondria. These proteins participate in the cell death process at different levels such as degradation of Diap1, a Drosophila IAP, production of mitochondrial reactive oxygen species or stimulation of the mitochondrial fission machinery. Here, we review these mitochondrial events that might have their counterpart in human.

  18. Identification of the Drosophila Ortholog of HSPB8 IMPLICATION OF HSPB8 LOSS OF FUNCTION IN PROTEIN FOLDING DISEASES

    NARCIS (Netherlands)

    Carra, Serena; Boncoraglio, Alessandra; Kanon, Bart; Brunsting, Jeanette F.; Minoia, Melania; Rana, Anil; Vos, Michel J.; Seidel, Kay; Sibon, Ody C. M.; Kampinga, Harm H.

    2010-01-01

    Protein aggregation is a hallmark of many neuronal disorders, including the polyglutamine disorder spinocerebellar ataxia 3 and peripheral neuropathies associated with the K141E and K141N mutations in the small heat shock protein HSPB8. In cells, HSPB8 cooperates with BAG3 to stimulate autophagy in

  19. Organization and evolution of Drosophila terminin: similarities and differences between Drosophila and human telomeres

    Directory of Open Access Journals (Sweden)

    Grazia Daniela Raffa

    2013-05-01

    Full Text Available Drosophila lacks telomerase and fly telomeres are elongated by occasional transposition of three specialized retroelements. Drosophila telomeres do not terminate with GC-rich repeats and are assembled independently of the sequence of chromosome ends. Recent work has shown that Drosophila telomeres are capped by the terminin complex, which includes the fast-evolving proteins HOAP, HipHop, Moi and Ver. These proteins are not conserves outside Drosophilidae and localize and function exclusively at telomeres, protecting them from fusion events. Other proteins required to prevent end-to-end fusion in flies include HP1, Eff/UbcD1, ATM, the components of the Mre11-Rad50-Nbs (MRN complex, and the Woc transcription factor. These proteins do not share the terminin properties; they are evolutionarily conserved non-fast-evolving proteins that do not accumulate only telomeres and do not serve telomere-specific functions. We propose that following telomerase loss, Drosophila rapidly evolved terminin to bind chromosome ends in a sequence-independent manner. This hypothesis suggests that terminin is the functional analog of the shelterin complex that protects human telomeres. The non-terminin proteins are instead likely to correspond to ancestral telomere-associated proteins that did not evolve as rapidly as terminin because of the functional constraints imposed by their involvement in diverse cellular processes. Thus, it appears that the main difference between Drosophila and human telomeres is in the protective complexes that specifically associate with the DNA termini. We believe that Drosophila telomeres offer excellent opportunities for investigations on human telomere biology. The identification of additional Drosophila genes encoding non-terminin proteins involved in telomere protection might lead to the discovery of novel components of human telomeres.

  20. Pitfalls of Insulin Pump Clocks

    Science.gov (United States)

    Reed, Amy J.

    2014-01-01

    The objective was to raise awareness about the importance of ensuring that insulin pumps internal clocks are set up correctly at all times. This is a very important safety issue because all commercially available insulin pumps are not GPS-enabled (though this is controversial), nor equipped with automatically adjusting internal clocks. Special attention is paid to how basal and bolus dose errors can be introduced by daylight savings time changes, travel across time zones, and am-pm clock errors. Correct setting of insulin pump internal clock is crucial for appropriate insulin delivery. A comprehensive literature review is provided, as are illustrative cases. Incorrect setting can potentially result in incorrect insulin delivery, with potential harmful consequences, if too much or too little insulin is delivered. Daylight saving time changes may not significantly affect basal insulin delivery, given the triviality of the time difference. However, bolus insulin doses can be dramatically affected. Such problems may occur when pump wearers have large variations in their insulin to carb ratio, especially if they forget to change their pump clock in the spring. More worrisome than daylight saving time change is the am-pm clock setting. If this setting is set up incorrectly, both basal rates and bolus doses will be affected. Appropriate insulin delivery through insulin pumps requires correct correlation between dose settings and internal clock time settings. Because insulin pumps are not GPS-enabled or automatically time-adjusting, extra caution should be practiced by patients to ensure correct time settings at all times. Clinicians and diabetes educators should verify the date/time of insulin pumps during patients’ visits, and should remind their patients to always verify these settings. PMID:25355713

  1. Relationships between the circadian system and Alzheimer's disease-like symptoms in Drosophila.

    Directory of Open Access Journals (Sweden)

    Dani M Long

    Full Text Available Circadian clocks coordinate physiological, neurological, and behavioral functions into circa 24 hour rhythms, and the molecular mechanisms underlying circadian clock oscillations are conserved from Drosophila to humans. Clock oscillations and clock-controlled rhythms are known to dampen during aging; additionally, genetic or environmental clock disruption leads to accelerated aging and increased susceptibility to age-related pathologies. Neurodegenerative diseases, such as Alzheimer's disease (AD, are associated with a decay of circadian rhythms, but it is not clear whether circadian disruption accelerates neuronal and motor decline associated with these diseases. To address this question, we utilized transgenic Drosophila expressing various Amyloid-β (Aβ peptides, which are prone to form aggregates characteristic of AD pathology in humans. We compared development of AD-like symptoms in adult flies expressing Aβ peptides in the wild type background and in flies with clocks disrupted via a null mutation in the clock gene period (per01. No significant differences were observed in longevity, climbing ability and brain neurodegeneration levels between control and clock-deficient flies, suggesting that loss of clock function does not exacerbate pathogenicity caused by human-derived Aβ peptides in flies. However, AD-like pathologies affected the circadian system in aging flies. We report that rest/activity rhythms were impaired in an age-dependent manner. Flies expressing the highly pathogenic arctic Aβ peptide showed a dramatic degradation of these rhythms in tune with their reduced longevity and impaired climbing ability. At the same time, the central pacemaker remained intact in these flies providing evidence that expression of Aβ peptides causes rhythm degradation downstream from the central clock mechanism.

  2. Molecular cloning, functional expression, and gene silencing of two Drosophila receptors for the Drosophila neuropeptide pyrokinin-2

    DEFF Research Database (Denmark)

    Rosenkilde, Carina; Cazzamali, Giuseppe; Williamson, Michael

    2003-01-01

    The database of the Drosophila Genome Project contains the sequences of two genes, CG8784 and CG8795, predicted to code for two structurally related G protein-coupled receptors. We have cloned these genes and expressed their coding parts in Chinese hamster ovary cells. We found that both receptors...... can be activated by low concentrations of the Drosophila neuropeptide pyrokinin-2 (CG8784, EC(50) for pyrokinin-2, 1x10(-9)M; CG8795, EC(50) for pyrokinin-2, 5 x 10(-10)M). The precise role of Drosophila pyrokinin-2 (SVPFKPRLamide) in Drosophila is unknown, but in other insects, pyrokinins have...... embryos and first instar larvae. In addition to the two Drosophila receptors, we also identified two probable pyrokinin receptors in the genomic database from the malaria mosquito Anopheles gambiae. The two Drosophila pyrokinin receptors are, to our knowledge, the first invertebrate pyrokinin receptors...

  3. Stress-induced changes in the expression of the clock protein PERIOD1 in the rat limbic forebrain and hypothalamus: role of stress type, time of day, and predictability.

    Directory of Open Access Journals (Sweden)

    Sherin Al-Safadi

    Full Text Available Stressful events can disrupt circadian rhythms in mammals but mechanisms underlying this disruption remain largely unknown. One hypothesis is that stress alters circadian protein expression in the forebrain, leading to functional dysregulation of the brain circadian network and consequent disruption of circadian physiological and behavioral rhythms. Here we characterized the effects of several different stressors on the expression of the core clock protein, PER1 and the activity marker, FOS in select forebrain and hypothalamic nuclei in rats. We found that acute exposure to processive stressors, restraint and forced swim, elevated PER1 and FOS expression in the paraventricular and dorsomedial hypothalamic nuclei and piriform cortex but suppressed PER1 and FOS levels exclusively in the central nucleus of the amygdala (CEAl and oval nucleus of the bed nucleus of the stria terminalis (BNSTov. Conversely, systemic stressors, interleukin-1β and 2-Deoxy-D-glucose, increased PER1 and FOS levels in all regions studied, including the CEAl and BNSTov. PER1 levels in the suprachiasmatic nucleus (SCN, the master pacemaker, were unaffected by any of the stress manipulations. The effect of stress on PER1 and FOS was modulated by time of day and, in the case of daily restraint, by predictability. These results demonstrate that the expression of PER1 in the forebrain is modulated by stress, consistent with the hypothesis that PER1 serves as a link between stress and the brain circadian network. Furthermore, the results show that the mechanisms that control PER1 and FOS expression in CEAl and BNSTov are uniquely sensitive to differences in the type of stressor. Finally, the finding that the effect of stress on PER1 parallels its effect on FOS supports the idea that Per1 functions as an immediate-early gene. Our observations point to a novel role for PER1 as a key player in the interface between stress and circadian rhythms.

  4. Identification of Atg2 and ArfGAP1 as Candidate Genetic Modifiers of the Eye Pigmentation Phenotype of Adaptor Protein-3 (AP-3 Mutants in Drosophila melanogaster.

    Directory of Open Access Journals (Sweden)

    Imilce A Rodriguez-Fernandez

    Full Text Available The Adaptor Protein (AP-3 complex is an evolutionary conserved, molecular sorting device that mediates the intracellular trafficking of proteins to lysosomes and related organelles. Genetic defects in AP-3 subunits lead to impaired biogenesis of lysosome-related organelles (LROs such as mammalian melanosomes and insect eye pigment granules. In this work, we have performed a forward screening for genetic modifiers of AP-3 function in the fruit fly, Drosophila melanogaster. Specifically, we have tested collections of large multi-gene deletions--which together covered most of the autosomal chromosomes-to identify chromosomal regions that, when deleted in single copy, enhanced or ameliorated the eye pigmentation phenotype of two independent AP-3 subunit mutants. Fine-mapping led us to define two non-overlapping, relatively small critical regions within fly chromosome 3. The first critical region included the Atg2 gene, which encodes a conserved protein involved in autophagy. Loss of one functional copy of Atg2 ameliorated the pigmentation defects of mutants in AP-3 subunits as well as in two other genes previously implicated in LRO biogenesis, namely Blos1 and lightoid, and even increased the eye pigment content of wild-type flies. The second critical region included the ArfGAP1 gene, which encodes a conserved GTPase-activating protein with specificity towards GTPases of the Arf family. Loss of a single functional copy of the ArfGAP1 gene ameliorated the pigmentation phenotype of AP-3 mutants but did not to modify the eye pigmentation of wild-type flies or mutants in Blos1 or lightoid. Strikingly, loss of the second functional copy of the gene did not modify the phenotype of AP-3 mutants any further but elicited early lethality in males and abnormal eye morphology when combined with mutations in Blos1 and lightoid, respectively. These results provide genetic evidence for new functional links connecting the machinery for biogenesis of LROs with

  5. Sound Clocks and Sonic Relativity

    Science.gov (United States)

    Todd, Scott L.; Menicucci, Nicolas C.

    2017-10-01

    Sound propagation within certain non-relativistic condensed matter models obeys a relativistic wave equation despite such systems admitting entirely non-relativistic descriptions. A natural question that arises upon consideration of this is, "do devices exist that will experience the relativity in these systems?" We describe a thought experiment in which `acoustic observers' possess devices called sound clocks that can be connected to form chains. Careful investigation shows that appropriately constructed chains of stationary and moving sound clocks are perceived by observers on the other chain as undergoing the relativistic phenomena of length contraction and time dilation by the Lorentz factor, γ , with c the speed of sound. Sound clocks within moving chains actually tick less frequently than stationary ones and must be separated by a shorter distance than when stationary to satisfy simultaneity conditions. Stationary sound clocks appear to be length contracted and time dilated to moving observers due to their misunderstanding of their own state of motion with respect to the laboratory. Observers restricted to using sound clocks describe a universe kinematically consistent with the theory of special relativity, despite the preferred frame of their universe in the laboratory. Such devices show promise in further probing analogue relativity models, for example in investigating phenomena that require careful consideration of the proper time elapsed for observers.

  6. Circadian clocks, epigenetics, and cancer

    KAUST Repository

    Masri, Selma; Kinouchi, Kenichiro; Sassone-Corsi, Paolo

    2015-01-01

    The interplay between circadian rhythm and cancer has been suggested for more than a decade based on the observations that shift work and cancer incidence are linked. Accumulating evidence implicates the circadian clock in cancer survival and proliferation pathways. At the molecular level, multiple control mechanisms have been proposed to link circadian transcription and cell-cycle control to tumorigenesis.The circadian gating of the cell cycle and subsequent control of cell proliferation is an area of active investigation. Moreover, the circadian clock is a transcriptional system that is intricately regulated at the epigenetic level. Interestingly, the epigenetic landscape at the level of histone modifications, DNA methylation, and small regulatory RNAs are differentially controlled in cancer cells. This concept raises the possibility that epigenetic control is a common thread linking the clock with cancer, though little scientific evidence is known to date.This review focuses on the link between circadian clock and cancer, and speculates on the possible connections at the epigenetic level that could further link the circadian clock to tumor initiation or progression.

  7. Titan's methane clock

    Science.gov (United States)

    Nixon, C. A.; Jennings, D. E.; Romani, P. N.; Teanby, N. A.; Irwin, P. G. J.; Flasar, F. M.

    2010-04-01

    Measurements of the 12C/13C and D/H isotopic ratios in Titan's methane show intriguing differences from the values recorded in the giant planets. This implies that either (1) the atmosphere was differently endowed with material at the time of formation, or (2) evolutionary processes are at work in the moon's atmosphere - or some combination of the two. The Huygens Gas Chromatograph Mass Spectrometer Instrument (GCMS) found 12CH4/13CH4 = 82 +/- 1 (Niemann et al. 2005), some 7% lower than the giant planets' value of 88 +/- 7 (Sada et al. 1996), which closely matches the terrestrial inorganic standard of 89. The Cassini Composite Infrared Spectrometer (CIRS) has previously reported 12CH4/13CH4 of 77 +/-3 based on nadir sounding, which we now revise upwards to 80 +/- 4 based on more accurate limb sounding. The CIRS and GCMS results are therefore in agreement about an overall enrichment in 13CH4 of ~10%. The value of D/H in Titan's CH4 has long been controversial: historical measurements have ranged from about 8-15 x 10-5 (e.g. Coustenis et al. 1989, Coustenis et al. 2003). A recent measurement based on CIRS limb data by Bezard et al. (2007) puts the D/H in CH4 at (13 +/- 1) x 10-5, very much greater than in Jupiter and Saturn, ~2 x 10-5 (Mahaffy et al. 1998, Fletcher et al. 2009). To add complexity, the 12C/13C and D/H vary among molecules in Titan atmosphere, typically showing enhancement in D but depletion in 13C in the daughter species (H2, C2H2, C2H6), relative to the photochemical progenitor, methane. Jennings et al. (2009) have sought to interpret the variance in carbon isotopes as a Kinetic Isotope Effect (KIE), whilst an explanation for the D/H in all molecules remains elusive (Cordier et al. 2008). In this presentation we argue that evolution of isotopic ratios in Titan's methane over time forms a ticking 'clock', somewhat analogous to isotopic ratios in geochronology. Under plausible assumptions about the initial values and subsequent replenishment, various

  8. Low-resolution structure of Drosophila translin

    Science.gov (United States)

    Kumar, Vinay; Gupta, Gagan D.

    2012-01-01

    Crystals of native Drosophila melanogaster translin diffracted to 7 Å resolution. Reductive methylation of the protein improved crystal quality. The native and methylated proteins showed similar profiles in size-exclusion chromatography analyses but the methylated protein displayed reduced DNA-binding activity. Crystals of the methylated protein diffracted to 4.2 Å resolution at BM14 of the ESRF synchrotron. Crystals with 49% solvent content belonged to monoclinic space group P21 with eight protomers in the asymmetric unit. Only 2% of low-resolution structures with similar low percentage solvent content were found in the PDB. The crystal structure, solved by molecular replacement method, refined to Rwork (Rfree) of 0.24 (0.29) with excellent stereochemistry. The crystal structure clearly shows that drosophila protein exists as an octamer, and not as a decamer as expected from gel-filtration elution profiles. The similar octameric quaternary fold in translin orthologs and in translin–TRAX complexes suggests an up-down dimer as the basic structural subunit of translin-like proteins. The drosophila oligomer displays asymmetric assembly and increased radius of gyration that accounts for the observed differences between the elution profiles of human and drosophila proteins on gel-filtration columns. This study demonstrates clearly that low-resolution X-ray structure can be useful in understanding complex biological oligomers. PMID:23650579

  9. The Drosophila Microtubule-Associated Protein Mars Stabilizes Mitotic Spindles by Crosslinking Microtubules through Its N-Terminal Region

    DEFF Research Database (Denmark)

    Zhang, Gang; Beati, Hamze; Nilsson, Jakob

    2013-01-01

    Correct segregation of genetic material relies on proper assembly and maintenance of the mitotic spindle. How the highly dynamic microtubules (MTs) are maintained in stable mitotic spindles is a key question to be answered. Motor and non-motor microtubule associated proteins (MAPs) have been...

  10. A library of MiMICs allows tagging of genes and reversible, spatial and temporal knockdown of proteins in Drosophila

    Science.gov (United States)

    Nagarkar-Jaiswal, Sonal; Lee, Pei-Tseng; Campbell, Megan E; Chen, Kuchuan; Anguiano-Zarate, Stephanie; Cantu Gutierrez, Manuel; Busby, Theodore; Lin, Wen-Wen; He, Yuchun; Schulze, Karen L; Booth, Benjamin W; Evans-Holm, Martha; Venken, Koen JT; Levis, Robert W; Spradling, Allan C; Hoskins, Roger A; Bellen, Hugo J

    2015-01-01

    Here, we document a collection of ∼7434 MiMIC (Minos Mediated Integration Cassette) insertions of which 2854 are inserted in coding introns. They allowed us to create a library of 400 GFP-tagged genes. We show that 72% of internally tagged proteins are functional, and that more than 90% can be imaged in unfixed tissues. Moreover, the tagged mRNAs can be knocked down by RNAi against GFP (iGFPi), and the tagged proteins can be efficiently knocked down by deGradFP technology. The phenotypes associated with RNA and protein knockdown typically correspond to severe loss of function or null mutant phenotypes. Finally, we demonstrate reversible, spatial, and temporal knockdown of tagged proteins in larvae and adult flies. This new strategy and collection of strains allows unprecedented in vivo manipulations in flies for many genes. These strategies will likely extend to vertebrates. DOI: http://dx.doi.org/10.7554/eLife.05338.001 PMID:25824290

  11. Planar Cell Polarity Breaks the Symmetry of PAR Protein Distribution prior to Mitosis in Drosophila Sensory Organ Precursor Cells.

    Science.gov (United States)

    Besson, Charlotte; Bernard, Fred; Corson, Francis; Rouault, Hervé; Reynaud, Elodie; Keder, Alyona; Mazouni, Khalil; Schweisguth, François

    2015-04-20

    During development, cell-fate diversity can result from the unequal segregation of fate determinants at mitosis. Polarization of the mother cell is essential for asymmetric cell division (ACD). It often involves the formation of a cortical domain containing the PAR complex proteins Par3, Par6, and atypical protein kinase C (aPKC). In the fly notum, sensory organ precursor cells (SOPs) divide asymmetrically within the plane of the epithelium and along the body axis to generate two distinct cells. Fate asymmetry depends on the asymmetric localization of the PAR complex. In the absence of planar cell polarity (PCP), SOPs divide with a random planar orientation but still asymmetrically, showing that PCP is dispensable for PAR asymmetry at mitosis. To study when and how the PAR complex localizes asymmetrically, we have used a quantitative imaging approach to measure the planar polarization of the proteins Bazooka (Baz, fly Par3), Par6, and aPKC in living pupae. By using imaging of functional GFP-tagged proteins with image processing and computational modeling, we find that Baz, Par6, and aPKC become planar polarized prior to mitosis in a manner independent of the AuroraA kinase and that PCP is required for the planar polarization of Baz, Par6, and aPKC during interphase. This indicates that a "mitosis rescue" mechanism establishes asymmetry at mitosis in PCP mutants. This study therefore identifies PCP as the initial symmetry-breaking signal for the planar polarization of PAR proteins in asymmetrically dividing SOPs. Copyright © 2015 Elsevier Ltd. All rights reserved.

  12. The Circadian Clock-controlled Transcriptome of Developing Soybean Seeds

    Directory of Open Access Journals (Sweden)

    Karen A. Hudson

    2010-07-01

    Full Text Available A number of metabolic and physiological processes in plants are controlled by the circadian clock, which enables a plant to anticipate daily changes in the environment. Relatively little is known about circadian rhythms in developing seeds, which may be important for determining the extent and timing of nutrient storage in grain. Microarray expression profiling was used to identify genes expressed in developing soybean ( seeds that are controlled by the circadian clock. Genes with predicted functions in protein synthesis, fatty acid metabolism, and photosynthesis totaling 1.8% of the mRNAs detected in seed were found to be expressed in a circadian rhythm. Known circadian and light-controlled promoter elements were identified as over-represented in the promoters of clock-controlled seed genes, with the over-represented elements varying according to the phase of circadian expression. A subset of circadian-regulated genes were found to be expressed in different phases in developing seeds with respect to leaves from the same plants, many of which have roles in photosynthesis and carbon metabolism. These results help to characterize the genes and processes in seeds that may be regulated by the circadian clock, and provide some insight into organ-specific phasing of clock controlled gene expression.

  13. Naming analog clocks conceptually facilitates naming digital clocks

    NARCIS (Netherlands)

    Meeuwissen, M.H.W.; Roelofs, A.P.A.; Levelt, W.J.M.

    2004-01-01

    Naming digital clocks (e.g., 2:45, say "quarter to three") requires conceptual operations on the minute and hour information displayed in the input for producing the correct relative time expression. The interplay of these conceptual operations was investigated using a repetition priming paradigm.

  14. The Drosophila chitinase-like protein IDGF3 is involved in protection against nematodes and woung healing

    Czech Academy of Sciences Publication Activity Database

    Kučerová, Lucie; Brož, Václav; Arefin, B.; Maaroufi, H. O.; Hurychová, J.; Strnad, Hynek; Žurovec, Michal; Theopold, U.

    2016-01-01

    Roč. 8, č. 2 (2016), s. 199-210 ISSN 1662-811X R&D Projects: GA ČR GA14-27816S Institutional support: RVO:60077344 ; RVO:68378050 Keywords : chitinase-like proteins * imaginal disc growth factor * hemolymph clot Subject RIV: CE - Biochemistry; EB - Genetics ; Molecular Biology (UMG-J) Impact factor: 3.938, year: 2016 http://www.karger.com/Article/FullText/442351

  15. Interdependence of nutrient metabolism and the circadian clock system: Importance for metabolic health

    Science.gov (United States)

    Ribas-Latre, Aleix; Eckel-Mahan, Kristin

    2016-01-01

    Background While additional research is needed, a number of large epidemiological studies show an association between circadian disruption and metabolic disorders. Specifically, obesity, insulin resistance, cardiovascular disease, and other signs of metabolic syndrome all have been linked to circadian disruption in humans. Studies in other species support this association and generally reveal that feeding that is not in phase with the external light/dark cycle, as often occurs with night or rotating shift workers, is disadvantageous in terms of energy balance. As food is a strong driver of circadian rhythms in the periphery, understanding how nutrient metabolism drives clocks across the body is important for dissecting out why circadian misalignment may produce such metabolic effects. A number of circadian clock proteins as well as their accessory proteins (such as nuclear receptors) are highly sensitive to nutrient metabolism. Macronutrients and micronutrients can function as zeitgebers for the clock in a tissue-specific way and can thus impair synchrony between clocks across the body, or potentially restore synchrony in the case of circadian misalignment. Circadian nuclear receptors are particularly sensitive to nutrient metabolism and can alter tissue-specific rhythms in response to changes in the diet. Finally, SNPs in human clock genes appear to be correlated with diet-specific responses and along with chronotype eventually may provide valuable information from a clinical perspective on how to use diet and nutrition to treat metabolic disorders. Scope of review This article presents a background of the circadian clock components and their interrelated metabolic and transcriptional feedback loops, followed by a review of some recent studies in humans and rodents that address the effects of nutrient metabolism on the circadian clock and vice versa. We focus on studies in which results suggest that nutrients provide an opportunity to restore or, alternatively

  16. Biological clocks: riding the tides.

    Science.gov (United States)

    de la Iglesia, Horacio O; Johnson, Carl Hirschie

    2013-10-21

    Animals with habitats in the intertidal zone often display biological rhythms that coordinate with both the tidal and the daily environmental cycles. Two recent studies show that the molecular components of the biological clocks mediating tidal rhythms are likely different from the phylogenetically conserved components that mediate circadian (daily) rhythms. Copyright © 2013 Elsevier Ltd. All rights reserved.

  17. Influence of Quercetin in the Temporal Regulation of Redox Homeostasis in Drosophila melanogaster.

    Science.gov (United States)

    Subramanian, Perumal; Kaliyamoorthy, Kanimozhi; Jayapalan, Jaime Jacqueline; Abdul-Rahman, Puteri Shafinaz; Haji Hashim, Onn

    2017-01-01

    Numerous biological processes are governed by the biological clock. Studies using Drosophila melanogaster (L.) are valuable that could be of importance for their effective applications on rodent studies. In this study, the beneficial role of quercetin (a flavonoid) on H2O2 induced stress in D. melanogaster was investigated. D. melanogaster flies were divided into four groups (group I - control, group II - H2O2 (acute exposure), group III - quercetin, and group IV - quercetin + H2O2 treated). Negative geotaxis assay, oxidative stress indicators (protein carbonyls, thiobarbituric reactive substances [TBARS]), and antioxidants (superoxide dismutase [SOD], catalase [CAT], glutathione-S-transferase [GST], glutathione peroxidase, and reduced glutathione [GSH]) were measured at 4 h intervals over 24 h and temporal expression of heat shock protein-70 (Hsp70), Upd1 (homolog of IL-6 in Drosophila), and nitric oxide synthase (Nos) was analyzed by Western blotting. Groups II and IV showed altered biochemical rhythms (compared with controls). Decreased mesor values of negative geotaxis, SOD, CAT, GST, and GSH were noticed in H2O2, increased mesor of oxidative stress indicators (TBARS and protein carbonyl content) and a reversibility of the rhythmic characteristics were conspicuous after quercetin treatment. The expression levels of Hsp70, Upd1, and Nos were noticeably maximum at 04:00. Significant elevation of expression by H2O2 was nearly normalized by quercetin treatment. The possible mechanism by which quercetin modulates oxidant-antioxidant imbalance under oxidative stress could be ascribed to the modulation of the rhythmic properties. Our results will be helpful to understand the molecular interlink between circadian rhythm and oxidative stress mechanism. © The Author 2017. Published by Oxford University Press on behalf of the Entomological Society of America.

  18. A network of (autonomic) clock outputs

    NARCIS (Netherlands)

    Kalsbeek, A.; Perreau-Lenz, S.; Buijs, R. M.

    2006-01-01

    The circadian clock in the suprachiasmatic nuclei (SCN) is composed of thousands of oscillator neurons, each of which is dependent on the cell-autonomous action of a defined set of circadian clock genes. A major question is still how these individual oscillators are organized into a biological clock

  19. A network of (autonomic) clock outputs

    NARCIS (Netherlands)

    Kalsbeek, A.; Perreau-Lenz, S.; Buijs, R. M.

    2006-01-01

    The circadian clock in the suprachiasmatic nuclei (SCN) is composed of thousands of oscillator neurons, each dependent on the cell-autonomous action of a defined set of circadian clock genes. A major question is still how these individual oscillators are organized into a biological clock that

  20. Light and the human circadian clock

    NARCIS (Netherlands)

    Roenneberg, Till; Kantermann, Thomas; Juda, Myriam; Vetter, Céline; Allebrandt, Karla V

    2013-01-01

    The circadian clock can only reliably fulfil its function if it is stably entrained. Most clocks use the light-dark cycle as environmental signal (zeitgeber) for this active synchronisation. How we think about clock function and entrainment has been strongly influenced by the early concepts of the

  1. NF-1 Dependent Gene Regulation in Drosophila Melanogaster

    National Research Council Canada - National Science Library

    Zhong, Yi

    2004-01-01

    .... We have used an Affymetrix whole genome chip, containing all 13,500 genes of the fruit fly Drosophila, to identify 93 genes with altered expression patterns in flies that have no NF1 protein compared...

  2. Drosophila Syd-1, liprin-α, and protein phosphatase 2A B' subunit Wrd function in a linear pathway to prevent ectopic accumulation of synaptic materials in distal axons.

    Science.gov (United States)

    Li, Long; Tian, Xiaolin; Zhu, Mingwei; Bulgari, Dinara; Böhme, Mathias A; Goettfert, Fabian; Wichmann, Carolin; Sigrist, Stephan J; Levitan, Edwin S; Wu, Chunlai

    2014-06-18

    During synaptic development, presynaptic differentiation occurs as an intrinsic property of axons to form specialized areas of plasma membrane [active zones (AZs)] that regulate exocytosis and endocytosis of synaptic vesicles. Genetic and biochemical studies in vertebrate and invertebrate model systems have identified a number of proteins involved in AZ assembly. However, elucidating the molecular events of AZ assembly in a spatiotemporal manner remains a challenge. Syd-1 (synapse defective-1) and Liprin-α have been identified as two master organizers of AZ assembly. Genetic and imaging analyses in invertebrates show that Syd-1 works upstream of Liprin-α in synaptic assembly through undefined mechanisms. To understand molecular pathways downstream of Liprin-α, we performed a proteomic screen of Liprin-α-interacting proteins in Drosophila brains. We identify Drosophila protein phosphatase 2A (PP2A) regulatory subunit B' [Wrd (Well Rounded)] as a Liprin-α-interacting protein, and we demonstrate that it mediates the interaction of Liprin-α with PP2A holoenzyme and the Liprin-α-dependent synaptic localization of PP2A. Interestingly, loss of function in syd-1, liprin-α, or wrd shares a common defect in which a portion of synaptic vesicles, dense-core vesicles, and presynaptic cytomatrix proteins ectopically accumulate at the distal, but not proximal, region of motoneuron axons. Strong genetic data show that a linear syd-1/liprin-α/wrd pathway in the motoneuron antagonizes glycogen synthase kinase-3β kinase activity to prevent the ectopic accumulation of synaptic materials. Furthermore, we provide data suggesting that the syd-1/liprin-α/wrd pathway stabilizes AZ specification at the nerve terminal and that such a novel function is independent of the roles of syd-1/liprin-α in regulating the morphology of the T-bar structural protein BRP (Bruchpilot). Copyright © 2014 the authors 0270-6474/14/348474-14$15.00/0.

  3. Drosophila Syd-1, Liprin-α, and Protein Phosphatase 2A B′ Subunit Wrd Function in a Linear Pathway to Prevent Ectopic Accumulation of Synaptic Materials in Distal Axons

    Science.gov (United States)

    Li, Long; Tian, Xiaolin; Zhu, Mingwei; Bulgari, Dinara; Böhme, Mathias A.; Goettfert, Fabian; Wichmann, Carolin; Sigrist, Stephan J.; Levitan, Edwin S.

    2014-01-01

    During synaptic development, presynaptic differentiation occurs as an intrinsic property of axons to form specialized areas of plasma membrane [active zones (AZs)] that regulate exocytosis and endocytosis of synaptic vesicles. Genetic and biochemical studies in vertebrate and invertebrate model systems have identified a number of proteins involved in AZ assembly. However, elucidating the molecular events of AZ assembly in a spatiotemporal manner remains a challenge. Syd-1 (synapse defective-1) and Liprin-α have been identified as two master organizers of AZ assembly. Genetic and imaging analyses in invertebrates show that Syd-1 works upstream of Liprin-α in synaptic assembly through undefined mechanisms. To understand molecular pathways downstream of Liprin-α, we performed a proteomic screen of Liprin-α-interacting proteins in Drosophila brains. We identify Drosophila protein phosphatase 2A (PP2A) regulatory subunit B′ [Wrd (Well Rounded)] as a Liprin-α-interacting protein, and we demonstrate that it mediates the interaction of Liprin-α with PP2A holoenzyme and the Liprin-α-dependent synaptic localization of PP2A. Interestingly, loss of function in syd-1, liprin-α, or wrd shares a common defect in which a portion of synaptic vesicles, dense-core vesicles, and presynaptic cytomatrix proteins ectopically accumulate at the distal, but not proximal, region of motoneuron axons. Strong genetic data show that a linear syd-1/liprin-α/wrd pathway in the motoneuron antagonizes glycogen synthase kinase-3β kinase activity to prevent the ectopic accumulation of synaptic materials. Furthermore, we provide data suggesting that the syd-1/liprin-α/wrd pathway stabilizes AZ specification at the nerve terminal and that such a novel function is independent of the roles of syd-1/liprin-α in regulating the morphology of the T-bar structural protein BRP (Bruchpilot). PMID:24948803

  4. Clocks do not tick in unison: isolation of Clock and vrille shed new light on the clockwork model of the sand fly Lutzomyia longipalpis.

    Science.gov (United States)

    Gesto, João Silveira Moledo; Rivas, Gustavo Bueno da Silva; Pavan, Marcio Galvão; Meireles-Filho, Antonio Carlos Alves; Amoretty, Paulo Roberto de; Souza, Nataly Araújo de; Bruno, Rafaela Vieira; Peixoto, Alexandre Afranio

    2015-10-06

    Behavior rhythms of insect vectors directly interfere with the dynamics of pathogen transmission to humans. The sand fly Lutzomyia longipalpis is the main vector of visceral leishmaniasis in America and concentrates its activity around dusk. Despite the accumulation of behavioral data, very little is known about the molecular bases of the clock mechanism in this species. This study aims to characterize, within an evolutionary perspective, two important circadian clock genes, Clock and vrille. We have cloned and isolated the coding sequence of L. longipalpis' genes Clock and vrille. The former is structured in eight exons and encodes a protein of 696 amino acids, and the latter comprises three exons and translates to a protein of 469 amino acids. When compared to other insects' orthologues, L. longipalpis CLOCK shows a high degree of conservation in the functional domains bHLH and PAS, but a much shorter glutamine-rich (poly-Q) C-terminal region. As for L. longipalpis VRILLE, a high degree of conservation was found in the bZIP domain. To support these observations and provide an elegant view of the evolution of both genes in insects, phylogenetic analyses based on maximum-likelihood and Bayesian inferences were performed, corroborating the previously known insect systematics. The isolation and phylogenetic analyses of Clock and vrille orthologues in L. longipalpis bring novel and important data to characterize this species' circadian clock. Interestingly, the poly-Q shortening observed in CLOCK suggests that its transcription activity might be impaired and we speculate if this effect could be compensated by other clock factors such as CYCLE.

  5. Daily rhythmicity of clock gene transcripts in atlantic cod fast skeletal muscle.

    Directory of Open Access Journals (Sweden)

    Carlo C Lazado

    Full Text Available The classical notion of a centralized clock that governs circadian rhythmicity has been challenged with the discovery of peripheral oscillators that enable organisms to cope with daily changes in their environment. The present study aimed to identify the molecular clock components in Atlantic cod (Gadus morhua and to investigate their daily gene expression in fast skeletal muscle. Atlantic cod clock genes were closely related to their orthologs in teleosts and tetrapods. Synteny was conserved to varying degrees in the majority of the 18 clock genes examined. In particular, aryl hydrocarbon receptor nuclear translocator-like 2 (arntl2, RAR-related orphan receptor A (rora and timeless (tim displayed high degrees of conservation. Expression profiling during the early ontogenesis revealed that some transcripts were maternally transferred, namely arntl2, cryptochrome 1b and 2 (cry1b and cry2, and period 2a and 2b (per2a and per2b. Most clock genes were ubiquitously expressed in various tissues, suggesting the possible existence of multiple peripheral clock systems in Atlantic cod. In particular, they were all detected in fast skeletal muscle, with the exception of neuronal PAS (Per-Arnt-Single-minded domain-containing protein (npas1 and rora. Rhythmicity analysis revealed 8 clock genes with daily rhythmic expression, namely arntl2, circadian locomotor output cycles kaput (clock, npas2, cry2, cry3 per2a, nuclear receptor subfamily 1, group D, member 1 (nr1d1, and nr1d2a. Transcript levels of the myogenic genes myogenic factor 5 (myf5 and muscleblind-like 1 (mbnl1 strongly correlated with clock gene expression. This is the first study to unravel the molecular components of peripheral clocks in Atlantic cod. Taken together, our data suggest that the putative clock system in fast skeletal muscle of Atlantic cod has regulatory implications on muscle physiology, particularly in the expression of genes related to myogenesis.

  6. Clocking In Time to Gate Memory Processes: The Circadian Clock Is Part of the Ins and Outs of Memory

    Directory of Open Access Journals (Sweden)

    Oliver Rawashdeh

    2018-01-01

    Full Text Available Learning, memory consolidation, and retrieval are processes known to be modulated by the circadian (circa: about; dies: day system. The circadian regulation of memory performance is evolutionarily conserved, independent of the type and complexity of the learning paradigm tested, and not specific to crepuscular, nocturnal, or diurnal organisms. In mammals, long-term memory (LTM formation is tightly coupled to de novo gene expression of plasticity-related proteins and posttranslational modifications and relies on intact cAMP/protein kinase A (PKA/protein kinase C (PKC/mitogen-activated protein kinase (MAPK/cyclic adenosine monophosphate response element-binding protein (CREB signaling. These memory-essential signaling components cycle rhythmically in the hippocampus across the day and night and are clearly molded by an intricate interplay between the circadian system and memory. Important components of the circadian timing mechanism and its plasticity are members of the Period clock gene family (Per1, Per2. Interestingly, Per1 is rhythmically expressed in mouse hippocampus. Observations suggest important and largely unexplored roles of the clock gene protein PER1 in synaptic plasticity and in the daytime-dependent modulation of learning and memory. Here, we review the latest findings on the role of the clock gene Period 1 (Per1 as a candidate molecular and mechanistic blueprint for gating the daytime dependency of memory processing.

  7. Neuronal Cbl Controls Biosynthesis of Insulin-Like Peptides in Drosophila melanogaster

    OpenAIRE

    Yu, Yue; Sun, Ying; He, Shengqi; Yan, Cheng; Rui, Liangyou; Li, Wenjun; Liu, Yong

    2012-01-01

    The Cbl family proteins function as both E3 ubiquitin ligases and adaptor proteins to regulate various cellular signaling events, including the insulin/insulin-like growth factor 1 (IGF1) and epidermal growth factor (EGF) pathways. These pathways play essential roles in growth, development, metabolism, and survival. Here we show that in Drosophila melanogaster, Drosophila Cbl (dCbl) regulates longevity and carbohydrate metabolism through downregulating the production of Drosophila insulin-lik...

  8. A Member of the p38 Mitogen-Activated Protein Kinase Family Is Responsible for Transcriptional Induction of Dopa decarboxylase in the Epidermis of Drosophila melanogaster during the Innate Immune Response▿ †

    Science.gov (United States)

    Davis, Monica M.; Primrose, David A.; Hodgetts, Ross B.

    2008-01-01

    Drosophila innate immunity is controlled primarily by the activation of IMD (immune deficiency) or Toll signaling leading to the production of antimicrobial peptides (AMPs). IMD signaling also activates the JUN N-terminal kinase (JNK) cascade, which is responsible for immune induction of non-antimicrobial peptide immune gene transcription though the transcription factor AP-1. Transcription of the Dopa decarboxylase (Ddc) gene is induced in response to gram-negative and gram-positive septic injury, but not aseptic wounding. Transcription is induced throughout the epidermis and not specifically at the site of infection. Ddc transcripts are detectible within 2 h and remain high for several hours following infection with either gram-negative or gram-positive bacteria. Using Ddc-green fluorescent protein (GFP) reporter gene constructs, we show that a conserved consensus AP-1 binding site upstream of the Ddc transcription start site is required for induction. However, neither the Toll, IMD, nor JNK pathway is involved. Rather, Ddc transcription depends on a previously uncharacterized member of the p38 mitogen-activated protein kinase family, p38c. We propose that the involvement of DDC in a new pathway involved in Drosophila immunity increases the levels of dopamine, which is metabolized to produce reactive quinones that exert an antimicrobial effect on invading bacteria. PMID:18519585

  9. Complementary approaches to understanding the plant circadian clock

    Directory of Open Access Journals (Sweden)

    Ozgur E. Akman

    2010-02-01

    Full Text Available Circadian clocks are oscillatory genetic networks that help organisms adapt to the 24-hour day/night cycle. The clock of the green alga Ostreococcus tauri is the simplest plant clock discovered so far. Its many advantages as an experimental system facilitate the testing of computational predictions. We present a model of the Ostreococcus clock in the stochastic process algebra Bio-PEPA and exploit its mapping to different analysis techniques, such as ordinary differential equations, stochastic simulation algorithms and model-checking. The small number of molecules reported for this system tests the limits of the continuous approximation underlying differential equations. We investigate the difference between continuous-deterministic and discrete-stochastic approaches. Stochastic simulation and model-checking allow us to formulate new hypotheses on the system behaviour, such as the presence of self-sustained oscillations in single cells under constant light conditions. We investigate how to model the timing of dawn and dusk in the context of model-checking, which we use to compute how the probability distributions of key biochemical species change over time. These show that the relative variation in expression level is smallest at the time of peak expression, making peak time an optimal experimental phase marker. Building on these analyses, we use approaches from evolutionary systems biology to investigate how changes in the rate of mRNA degradation impacts the phase of a key protein likely to affect fitness. We explore how robust this circadian clock is towards such potential mutational changes in its underlying biochemistry. Our work shows that multiple approaches lead to a more complete understanding of the clock.

  10. Automatic control of clock duty cycle

    Science.gov (United States)

    Feng, Xiaoxin (Inventor); Roper, Weston (Inventor); Seefeldt, James D. (Inventor)

    2010-01-01

    In general, this disclosure is directed to a duty cycle correction (DCC) circuit that adjusts a falling edge of a clock signal to achieve a desired duty cycle. In some examples, the DCC circuit may generate a pulse in response to a falling edge of an input clock signal, delay the pulse based on a control voltage, adjust the falling edge of the input clock signal based on the delayed pulse to produce an output clock signal, and adjust the control voltage based on the difference between a duty cycle of the output clock signal and a desired duty cycle. Since the DCC circuit adjusts the falling edge of the clock cycle to achieve a desired duty cycle, the DCC may be incorporated into existing PLL control loops that adjust the rising edge of a clock signal without interfering with the operation of such PLL control loops.

  11. Patterns of mutation and selection at synonymous sites in Drosophila

    DEFF Research Database (Denmark)

    Singh, Nadia D; Bauer DuMont, Vanessa L; Hubisz, Melissa J

    2007-01-01

    , when applied to 18 coding sequences in 3 species of Drosophila, confirmed an earlier report that the Notch gene in Drosophila melanogaster was evolving under selection in favor of those codons defined as unpreferred in this species. This finding opened the possibility that synonymous sites may...... be subject to a variety of selective pressures beyond weak selection for increased frequencies of the codons currently defined as "preferred" in D. melanogaster. To further explore patterns of synonymous site evolution in Drosophila in a lineage-specific manner, we expanded the application of the maximum...... likelihood framework to 8,452 protein coding sequences with well-defined orthology in D. melanogaster, Drosophila sechellia, and Drosophila yakuba. Our analyses reveal intragenomic and interspecific variation in mutational patterns as well as in patterns and intensity of selection on synonymous sites. In D...

  12. Drosophila Studies on Autism Spectrum Disorders

    Institute of Scientific and Technical Information of China (English)

    Yao Tian; Zi Chao Zhang; Junhai Han

    2017-01-01

    In the past decade,numerous genes associated with autism spectrum disorders (ASDs) have been identified.These genes encode key regulators of synaptogenesis,synaptic function,and synaptic plasticity.Drosophila is a prominent model system for ASD studies to define novel genes linked to ASDs and decipher their molecular roles in synaptogenesis,synaptic function,synaptic plasticity,and neural circuit assembly and consolidation.Here,we review Drosophila studies on ASD genes that regulate synaptogenesis,synaptic function,and synaptic plasticity through modulating chromatin remodeling,transcription,protein synthesis and degradation,cytoskeleton dynamics,and synaptic scaffolding.

  13. Learning and memory deficits consequent to reduction of the fragile X mental retardation protein result from metabotropic glutamate receptor-mediated inhibition of cAMP signaling in Drosophila.

    Science.gov (United States)

    Kanellopoulos, Alexandros K; Semelidou, Ourania; Kotini, Andriana G; Anezaki, Maria; Skoulakis, Efthimios M C

    2012-09-19

    Loss of the RNA-binding fragile X protein [fragile X mental retardation protein (FMRP)] results in a spectrum of cognitive deficits, the fragile X syndrome (FXS), while aging individuals with decreased protein levels present with a subset of these symptoms and tremor. The broad range of behavioral deficits likely reflects the ubiquitous distribution and multiple functions of the protein. FMRP loss is expected to affect multiple neuronal proteins and intracellular signaling pathways, whose identity and interactions are essential in understanding and ameliorating FXS symptoms. We used heterozygous mutants and targeted RNA interference-mediated abrogation in Drosophila to uncover molecular pathways affected by FMRP reduction. We present evidence that FMRP loss results in excess metabotropic glutamate receptor (mGluR) activity, attributable at least in part to elevation of the protein in affected neurons. Using high-resolution behavioral, genetic, and biochemical analyses, we present evidence that excess mGluR upon FMRP attenuation is linked to the cAMP decrement reported in patients and models, and underlies olfactory associative learning and memory deficits. Furthermore, our data indicate positive transcriptional regulation of the fly fmr1 gene by cAMP, via protein kinase A, likely through the transcription factor CREB. Because the human Fmr1 gene also contains CREB binding sites, the interaction of mGluR excess and cAMP signaling defects we present suggests novel combinatorial pharmaceutical approaches to symptom amelioration upon FMRP attenuation.

  14. Hanle Detection for Optical Clocks

    Directory of Open Access Journals (Sweden)

    Xiaogang Zhang

    2015-01-01

    Full Text Available Considering the strong inhomogeneous spatial polarization and intensity distribution of spontaneous decay fluorescence due to the Hanle effect, we propose and demonstrate a universe Hanle detection configuration of electron-shelving method for optical clocks. Experimental results from Ca atomic beam optical frequency standard with electron-shelving method show that a designed Hanle detection geometry with optimized magnetic field direction, detection laser beam propagation and polarization direction, and detector position can improve the fluorescence collection rate by more than one order of magnitude comparing with that of inefficient geometry. With the fixed 423 nm fluorescence, the improved 657 nm optical frequency standard signal intensity is presented. The potential application of the Hanle detection geometry designed for facilitating the fluorescence collection for optical lattice clock with a limited solid angle of the fluorescence collection has been discussed. The Hanle detection geometry is also effective for ion detection in ion optical clock and quantum information experiments. Besides, a cylinder fluorescence collection structure is designed to increase the solid angle of the fluorescence collection in Ca atomic beam optical frequency standard.

  15. Altered Rhythm of Adrenal Clock Genes, StAR and Serum Corticosterone in VIP Receptor 2-Deficient Mice

    DEFF Research Database (Denmark)

    Fahrenkrug, Jan; Georg, Birgitte; Hannibal, Jens

    2012-01-01

    oscillator based on a group of clock genes and their protein products. Mice lacking the VPAC2 receptor display disrupted circadian rhythm of physiology and behaviour, and therefore, we using real-time RT-PCR quantified (1) the mRNAs for the clock genes Per1 and Bmal1 in the adrenal gland and SCN, (2......RNA expression and serum corticosterone concentration. Double immunohistochemistry showed that the PER1 protein and StAR were co-localised in the same steroidogenic cells. Circulating corticosterone plays a role in the circadian timing system and the misaligned corticosterone rhythm in the VPAC2 receptor......The circadian time-keeping system consists of clocks in the suprachiasmatic nucleus (SCN) and in peripheral organs including an adrenal clock linked to the rhythmic corticosteroid production by regulating steroidogenic acute regulatory protein (StAR). Clock cells contain an autonomous molecular...

  16. Discrete gene replication events drive coupling between the cell cycle and circadian clocks.

    Science.gov (United States)

    Paijmans, Joris; Bosman, Mark; Ten Wolde, Pieter Rein; Lubensky, David K

    2016-04-12

    Many organisms possess both a cell cycle to control DNA replication and a circadian clock to anticipate changes between day and night. In some cases, these two rhythmic systems are known to be coupled by specific, cross-regulatory interactions. Here, we use mathematical modeling to show that, additionally, the cell cycle generically influences circadian clocks in a nonspecific fashion: The regular, discrete jumps in gene-copy number arising from DNA replication during the cell cycle cause a periodic driving of the circadian clock, which can dramatically alter its behavior and impair its function. A clock built on negative transcriptional feedback either phase-locks to the cell cycle, so that the clock period tracks the cell division time, or exhibits erratic behavior. We argue that the cyanobacterium Synechococcus elongatus has evolved two features that protect its clock from such disturbances, both of which are needed to fully insulate it from the cell cycle and give it its observed robustness: a phosphorylation-based protein modification oscillator, together with its accompanying push-pull read-out circuit that responds primarily to the ratios of different phosphoform concentrations, makes the clock less susceptible to perturbations in protein synthesis; the presence of multiple, asynchronously replicating copies of the same chromosome diminishes the effect of replicating any single copy of a gene.

  17. Entanglement of quantum clocks through gravity.

    Science.gov (United States)

    Castro Ruiz, Esteban; Giacomini, Flaminia; Brukner, Časlav

    2017-03-21

    In general relativity, the picture of space-time assigns an ideal clock to each world line. Being ideal, gravitational effects due to these clocks are ignored and the flow of time according to one clock is not affected by the presence of clocks along nearby world lines. However, if time is defined operationally, as a pointer position of a physical clock that obeys the principles of general relativity and quantum mechanics, such a picture is, at most, a convenient fiction. Specifically, we show that the general relativistic mass-energy equivalence implies gravitational interaction between the clocks, whereas the quantum mechanical superposition of energy eigenstates leads to a nonfixed metric background. Based only on the assumption that both principles hold in this situation, we show that the clocks necessarily get entangled through time dilation effect, which eventually leads to a loss of coherence of a single clock. Hence, the time as measured by a single clock is not well defined. However, the general relativistic notion of time is recovered in the classical limit of clocks.

  18. A VMEbus clock system for accelerator control

    International Nuclear Information System (INIS)

    Beechy, D.G.; McClure, C.R.

    1992-01-01

    Because an accelerator has many systems which must operate with a high degree of synchronization, a clock signal is typically generated which carries timing information to the various accelerator components. This paper discusses two VMEbus modules designed to generate and receive this clock signal. Together they implement a clock system which can generate timing markers with 200 nanosecond resolution and can generate timing delays of over one hour with one microsecond resolution. The Clock Generator module contains both a time line generator programmed to produce clock events at specific times and eight programmable input channels to produce clock events when externally triggered. Additional clock events are generated directly from the VMEbus. Generators can be cascaded for added capability. The Clock Timer module receives the signal from the generator. It can be programmed to recognize specific clock events which act as triggers to the eight timing channels on the module. Each timing channel is programmed with a 32-bit delay value. The channels are clocked at 1 MHz. At the end of the delay period, a timer channel produces an output pulse and optionally can generate a bus interrupt

  19. Combination of Hypomorphic Mutations of the Drosophila Homologues of Aryl Hydrocarbon Receptor and Nucleosome Assembly Protein Family Genes Disrupts Morphogenesis, Memory and Detoxification

    OpenAIRE

    Kuzin, Boris A.; Nikitina, Ekaterina A.; Cherezov, Roman O.; Vorontsova, Julia E.; Slezinger, Mikhail S.; Zatsepina, Olga G.; Simonova, Olga B.; Enikolopov, Grigori N.; Savvateeva-Popova, Elena V.

    2014-01-01

    Aryl hydrocarbon receptor is essential for biological responses to endogenous and exogenous toxins in mammals. Its Drosophila homolog spineless plays an important role in fly morphogenesis. We have previously shown that during morphogenesis spineless genetically interacts with CG5017 gene, which encodes a nucleosome assembly factor and may affect cognitive function of the fly. We now demonstrate synergistic interactions of spineless and CG5017 in pathways controlling oxidative stress response...

  20. Automated analysis of long-term grooming behavior in Drosophila using a k-nearest neighbors classifier

    Science.gov (United States)

    Allen, Victoria W; Shirasu-Hiza, Mimi

    2018-01-01

    Despite being pervasive, the control of programmed grooming is poorly understood. We addressed this gap by developing a high-throughput platform that allows long-term detection of grooming in Drosophila melanogaster. In our method, a k-nearest neighbors algorithm automatically classifies fly behavior and finds grooming events with over 90% accuracy in diverse genotypes. Our data show that flies spend ~13% of their waking time grooming, driven largely by two major internal programs. One of these programs regulates the timing of grooming and involves the core circadian clock components cycle, clock, and period. The second program regulates the duration of grooming and, while dependent on cycle and clock, appears to be independent of period. This emerging dual control model in which one program controls timing and another controls duration, resembles the two-process regulatory model of sleep. Together, our quantitative approach presents the opportunity for further dissection of mechanisms controlling long-term grooming in Drosophila. PMID:29485401

  1. The Implementation of E1 Clock Recovery

    Directory of Open Access Journals (Sweden)

    Wang Ziyu

    2016-01-01

    Full Text Available Clock transform and recovery is of significant importance in microwave TDM service, and it is always extracted from the E1 line data stream in most cases. However, intrinsically uncertain delay and jitter caused by packet transmission of E1 data information, may lead to the indexes of the data recovery clock exceed the clock performance template. Through analysis of the E1 clock indexes and measuring methods, this paper proposes a new clock recovery method. The method employs two buffers, the first RAM is used as a buffer to deduct excess information, and the second FIFO is used as a buffer to recovery the clock and data. The first buffer has a feedback from the second one, and is able to actively respond to changes in the data link and requests from the second one. The test results validate the effectiveness of the method, and the corresponding scheme is also valuable for the other communication systems.

  2. The Square Light Clock and Special Relativity

    Science.gov (United States)

    Galli, J. Ronald; Amiri, Farhang

    2012-01-01

    A thought experiment that includes a square light clock is similar to the traditional vertical light beam and mirror clock, except it is made up of four mirrors placed at a 45[degree] angle at each corner of a square of length L[subscript 0], shown in Fig. 1. Here we have shown the events as measured in the rest frame of the square light clock. By…

  3. Space experiments with high stability clocks

    International Nuclear Information System (INIS)

    Vessot, R.F.C.

    1993-01-01

    Modern metrology depends increasingly on the accuracy and frequency stability of atomic clocks. Applications of such high-stability oscillators (or clocks) to experiments performed in space are described and estimates of the precision of these experiments are made in terms of clock performance. Methods using time-correlation to cancel localized disturbances in very long signal paths and a proposed space borne four station VLBI system are described. (TEC). 30 refs., 14 figs., 1 tab

  4. NONO couples the circadian clock to the cell cycle.

    Science.gov (United States)

    Kowalska, Elzbieta; Ripperger, Juergen A; Hoegger, Dominik C; Bruegger, Pascal; Buch, Thorsten; Birchler, Thomas; Mueller, Anke; Albrecht, Urs; Contaldo, Claudio; Brown, Steven A

    2013-01-29

    Mammalian circadian clocks restrict cell proliferation to defined time windows, but the mechanism and consequences of this interrelationship are not fully understood. Previously we identified the multifunctional nuclear protein NONO as a partner of circadian PERIOD (PER) proteins. Here we show that it also conveys circadian gating to the cell cycle, a connection surprisingly important for wound healing in mice. Specifically, although fibroblasts from NONO-deficient mice showed approximately normal circadian cycles, they displayed elevated cell doubling and lower cellular senescence. At a molecular level, NONO bound to the p16-Ink4A cell cycle checkpoint gene and potentiated its circadian activation in a PER protein-dependent fashion. Loss of either NONO or PER abolished this activation and circadian expression of p16-Ink4A and eliminated circadian cell cycle gating. In vivo, lack of NONO resulted in defective wound repair. Because wound healing defects were also seen in multiple circadian clock-deficient mouse lines, our results therefore suggest that coupling of the cell cycle to the circadian clock via NONO may be useful to segregate in temporal fashion cell proliferation from tissue organization.

  5. Metabolomic Studies in Drosophila.

    Science.gov (United States)

    Cox, James E; Thummel, Carl S; Tennessen, Jason M

    2017-07-01

    Metabolomic analysis provides a powerful new tool for studies of Drosophila physiology. This approach allows investigators to detect thousands of chemical compounds in a single sample, representing the combined contributions of gene expression, enzyme activity, and environmental context. Metabolomics has been used for a wide range of studies in Drosophila , often providing new insights into gene function and metabolic state that could not be obtained using any other approach. In this review, we survey the uses of metabolomic analysis since its entry into the field. We also cover the major methods used for metabolomic studies in Drosophila and highlight new directions for future research. Copyright © 2017 by the Genetics Society of America.

  6. Enhancing Nanos expression via the bacterial TomO protein is a conserved strategy used by the symbiont Wolbachia to fuel germ stem cell maintenance in infected Drosophila females.

    Science.gov (United States)

    Ote, Manabu; Yamamoto, Daisuke

    2018-04-27

    The toxic manipulator of oogenesis (TomO) protein has been identified in the wMel strain of Wolbachia that symbioses with the vinegar fly Drosophila melanogaster, as a protein that affects host reproduction. TomO protects germ stem cells (GSCs) from degeneration, which otherwise occurs in ovaries of host females that are mutant for the gene Sex-lethal (Sxl). We isolated the TomO homologs from wPip, a Wolbachia strain from the mosquito Culex quinquefasciatus. One of the homologs, TomO w Pip 1, exerted the GSC rescue activity in fly Sxl mutants when lacking its hydrophobic stretches. The GSC-rescuing action of the TomO w Pip 1 variant was ascribable to its abilities to associate with Nanos (nos) mRNA and to enhance Nos protein expression. The analysis of structure-activity relationships with TomO homologs and TomO deletion variants revealed distinct modules in the protein that are each dedicated to different functions, i.e., subcellular localization, nos mRNA binding or Nos expression enhancement. We propose that modular reshuffling is the basis for structural and functional diversification of TomO protein members. © 2018 Wiley Periodicals, Inc.

  7. CRY Drives Cyclic CK2-Mediated BMAL1 Phosphorylation to Control the Mammalian Circadian Clock.

    Directory of Open Access Journals (Sweden)

    Teruya Tamaru

    Full Text Available Intracellular circadian clocks, composed of clock genes that act in transcription-translation feedback loops, drive global rhythmic expression of the mammalian transcriptome and allow an organism to anticipate to the momentum of the day. Using a novel clock-perturbing peptide, we established a pivotal role for casein kinase (CK-2-mediated circadian BMAL1-Ser90 phosphorylation (BMAL1-P in regulating central and peripheral core clocks. Subsequent analysis of the underlying mechanism showed a novel role of CRY as a repressor for protein kinase. Co-immunoprecipitation experiments and real-time monitoring of protein-protein interactions revealed that CRY-mediated periodic binding of CK2β to BMAL1 inhibits BMAL1-Ser90 phosphorylation by CK2α. The FAD binding domain of CRY1, two C-terminal BMAL1 domains, and particularly BMAL1-Lys537 acetylation/deacetylation by CLOCK/SIRT1, were shown to be critical for CRY-mediated BMAL1-CK2β binding. Reciprocally, BMAL1-Ser90 phosphorylation is prerequisite for BMAL1-Lys537 acetylation. We propose a dual negative-feedback model in which a CRY-dependent CK2-driven posttranslational BMAL1-P-BMAL1 loop is an integral part of the core clock oscillator.

  8. Drosophila's contribution to stem cell research [version 2; referees: 2 approved

    Directory of Open Access Journals (Sweden)

    Gyanesh Singh

    2016-08-01

    Full Text Available The discovery of Drosophila stem cells with striking similarities to mammalian stem cells has brought new hope for stem cell research. Recent developments in Drosophila stem cell research is bringing wider opportunities for contemporary stem cell biologists. In this regard, Drosophila germ cells are becoming a popular model of stem cell research. In several cases, genes that controlled Drosophila stem cells were later discovered to have functional homologs in mammalian stem cells. Like mammals, Drosophila germline stem cells (GSCs are controlled by both intrinsic as well as external signals. Inside the Drosophila testes, germline and somatic stem cells form a cluster of cells (the hub. Hub cells depend on JAK-STAT signaling, and, in absence of this signal, they do not self-renew. In Drosophila, significant changes occur within the stem cell niche that contributes to a decline in stem cell number over time. In case of aging Drosophila, somatic niche cells show reduced DE-cadherin and unpaired (Upd proteins. Unpaired proteins are known to directly decrease stem cell number within the niches, and, overexpression of upd within niche cells restored GSCs in older males also . Stem cells in the midgut of Drosophila are also very promising. Reduced Notch signaling was found to increase the number of midgut progenitor cells. On the other hand, activation of the Notch pathway decreased proliferation of these cells. Further research in this area should lead to the discovery of additional factors that regulate stem and progenitor cells in Drosophila.

  9. Dynamical Analysis of bantam-Regulated Drosophila Circadian Rhythm Model

    Science.gov (United States)

    Li, Ying; Liu, Zengrong

    MicroRNAs (miRNAs) interact with 3‧untranslated region (UTR) elements of target genes to regulate mRNA stability or translation, and play a crucial role in regulating many different biological processes. bantam, a conserved miRNA, is involved in several functions, such as regulating Drosophila growth and circadian rhythm. Recently, it has been discovered that bantam plays a crucial role in the core circadian pacemaker. In this paper, based on experimental observations, a detailed dynamical model of bantam-regulated circadian clock system is developed to show the post-transcriptional behaviors in the modulation of Drosophila circadian rhythm, in which the regulation of bantam is incorporated into a classical model. The dynamical behaviors of the model are consistent with the experimental observations, which shows that bantam is an important regulator of Drosophila circadian rhythm. The sensitivity analysis of parameters demonstrates that with the regulation of bantam the system is more sensitive to perturbations, indicating that bantam regulation makes it easier for the organism to modulate its period against the environmental perturbations. The effectiveness in rescuing locomotor activity rhythms of mutated flies shows that bantam is necessary for strong and sustained rhythms. In addition, the biological mechanisms of bantam regulation are analyzed, which may help us more clearly understand Drosophila circadian rhythm regulated by other miRNAs.

  10. Effect of the gene transformer of Anastrepha on the somatic sexual development of Drosophila.

    Science.gov (United States)

    Ruiz, María-Fernanda; Sánchez, Lucas

    2010-01-01

    The gene transformer (tra) is the key regulatory memory device for sex determination in tephritid insects. The present manuscript addressed the question about the functional conservation of the tephritid Anastrepha Transformer protein to direct somatic sexual development in Drosophila (Drosophilidae). The transformer cDNA of Anastrepha encoding the putative full-length Tra protein was cloned in pUAST and introduced into Drosophila melanogaster. To express this protein, the GAL4-UAS system was used. The Anastrepha Tra protein induced the female-specific splicing of both dsx and fru pre-mRNAs in Drosophila XY male flies, so that these became transformed into females, though this transformation was incomplete (the sexually dimorphic foreleg basitarsus and the external terminalia were monitored). It was found that the degree of female transformation directly depended on the dose of Anastrepha tra and Drosophila transformer-2 (tra-2) genes, and that the Anastrepha Tra-Drosophila Tra2 complex is not as efficient as the Drosophila Tra-Tra2 complex at inducing the female-specific splicing of Drosophila dsx pre-mRNA. This can explain why the Anastrepha Tra protein cannot fully substitute for the endogenous Drosophila Tra protein.

  11. High Performance Clocks and Gravity Field Determination

    Science.gov (United States)

    Müller, J.; Dirkx, D.; Kopeikin, S. M.; Lion, G.; Panet, I.; Petit, G.; Visser, P. N. A. M.

    2018-02-01

    Time measured by an ideal clock crucially depends on the gravitational potential and velocity of the clock according to general relativity. Technological advances in manufacturing high-precision atomic clocks have rapidly improved their accuracy and stability over the last decade that approached the level of 10^{-18}. This notable achievement along with the direct sensitivity of clocks to the strength of the gravitational field make them practically important for various geodetic applications that are addressed in the present paper. Based on a fully relativistic description of the background gravitational physics, we discuss the impact of those highly-precise clocks on the realization of reference frames and time scales used in geodesy. We discuss the current definitions of basic geodetic concepts and come to the conclusion that the advances in clocks and other metrological technologies will soon require the re-definition of time scales or, at least, clarification to ensure their continuity and consistent use in practice. The relative frequency shift between two clocks is directly related to the difference in the values of the gravity potential at the points of clock's localization. According to general relativity the relative accuracy of clocks in 10^{-18} is equivalent to measuring the gravitational red shift effect between two clocks with the height difference amounting to 1 cm. This makes the clocks an indispensable tool in high-precision geodesy in addition to laser ranging and space geodetic techniques. We show how clock measurements can provide geopotential numbers for the realization of gravity-field-related height systems and can resolve discrepancies in classically-determined height systems as well as between national height systems. Another application of clocks is the direct use of observed potential differences for the improved recovery of regional gravity field solutions. Finally, clock measurements for space-borne gravimetry are analyzed along with

  12. Long-Term Clock Behavior of GPS IIR Satellites

    National Research Council Canada - National Science Library

    Epstein, Marvin; Dass, Todd; Rajan, John; Gilmour, Paul

    2007-01-01

    .... Rubidium clocks, as opposed to cesium clocks, have significant long-term drift. The current literature describes an initial model of drift aging for rubidium atomic clocks followed by a long-term characteristic...

  13. Females With a Mutation in a Nuclear-Encoded Mitochondrial Protein Pay a Higher Cost of Survival Than Do Males in Drosophila

    OpenAIRE

    Melvin, Richard G.; Ballard, J. William O.

    2011-01-01

    Males and females age at different rates in a variety of species, but the mechanisms underlying the difference is not understood. In this study, we investigated sex-specific costs of a naturally occurring mildly deleterious deletion (DTrp85, DVal86) in cytochrome c oxidase subunit 7A (cox7A) in Drosophila simulans. We observed that females and males homozygous for the mutation had 30% and 26% reduced Cox activity, respectively, compared with wild type. Furthermore, 4-day-old females had 34%–4...

  14. Mutants dissecting development and behaviour in drosophila

    International Nuclear Information System (INIS)

    Joshi, Adita; Chandrashekaran, Shanti; Sharma, R.P.

    2005-01-01

    We have traced in this paper the progress in Drosophila genetics research from the 1960s, at the IARI, spearheaded by the visionary insight of M. S. Swaminathan. The work started with the study of indirect effect of radiation and the synergistic interaction of physical and chemical mutagens on chromosomal and genetic changes. This paved the way for the study of single gene mutants in dissecting developmental and behavioural processes. New genes discovered by us have been shown to encode conserved cell signalling molecules controlling developmental and behavioural pathways. With the complete sequencing of the Drosophila genome, in the year 2000, mounting evidence for the homology between Drosophila and human genes controlling genetic disorders became available. This has led to the fly becoming an indispensable tool for studying human diseases as well as a model to test for drugs and pharmaceuticals against human diseases and complex behavioural processes. For example wingless in Drosophila belongs to the conserved Wnt gene family and aberrant WNT signalling is linked to a range of human diseases, most notably cancer. Inhibition as well as activation of WNT signalling form the basis of an effective therapy for some cancers as well as several other clinical conditions. Recent experiments have shown that WNTs might also normally participate in self-renewal, proliferation or differentiation of stem cells and altering WNT signalling might be beneficial to the use of stem cells for therapeutic means. Likewise, the stambhA mutant of Drosophila which was discovered for its temperature-dependent paralytic behaviour is the fly homologue of Phospholipase Cβ. Phospholipase C mediated G protein signalling plays a central role in vital processes controlling epilepsy, vision, taste, and olfaction in animals. Proteins of the G-signalling pathway are of intense research interest since many human diseases involve defects in G-protein signalling pathways. In fact, approximately 50

  15. A clock synchronization skeleton based on RTAI

    NARCIS (Netherlands)

    Huang, Y.; Visser, P.M.; Broenink, Johannes F.

    2006-01-01

    This paper presents a clock synchronization skeleton based on RTAI (Real Time Application Interface). The skeleton is a thin layer that provides unified but extendible interfaces to the underlying operating system, the synchronization algorithms and the upper level applications in need of clock

  16. The clock paradox as a cosmological problem

    International Nuclear Information System (INIS)

    Fu, K.Y.

    1975-01-01

    In this paper the clock paradox is discussed within the framework of the general theory of relativity. It is shown that in general the aging asymmetry exists. It is also argued that the clock paradox, according to Mach's principle, is essentially a cosmological problem. (author)

  17. Could Atomic clocks be affected by neutrinos?

    CERN Document Server

    Hanafi, Hanaa

    2016-01-01

    An atomic clock is a clock device that uses an electronic transition frequency of the electromagnetic spectrum of atoms as a frequency standard in order to derive a time standard since time is the reciprocal of frequency. If the electronic transition frequencies are in an "optical region", we are talking in this case about optical atomic clocks. If they are in an "microwave region" these atomic clocks are made of the metallic element cesium so they are called Cesium atomic clocks. Atomic clocks are the most accurate time and frequency standards known despite the different perturbations that can affect them, a lot of researches were made in this domain to show how the transitions can be different for different type of perturbations..Since atomic clocks are very sensitive devices, based on coherent states (A coherent state tends to loose coherence after interacting). One question can arise (from a lot of questions) which is why cosmic neutrinos are not affecting these clocks? The answer to this question requir...

  18. Fast Clock Recovery for Digital Communications

    Science.gov (United States)

    Tell, R. G.

    1985-01-01

    Circuit extracts clock signal from random non-return-to-zero data stream, locking onto clock within one bit period at 1-gigabitper-second data rate. Circuit used for synchronization in opticalfiber communications. Derives speed from very short response time of gallium arsenide metal/semiconductor field-effect transistors (MESFET's).

  19. Internal Clock Drift Estimation in Computer Clusters

    Directory of Open Access Journals (Sweden)

    Hicham Marouani

    2008-01-01

    Full Text Available Most computers have several high-resolution timing sources, from the programmable interrupt timer to the cycle counter. Yet, even at a precision of one cycle in ten millions, clocks may drift significantly in a single second at a clock frequency of several GHz. When tracing the low-level system events in computer clusters, such as packet sending or reception, each computer system records its own events using an internal clock. In order to properly understand the global system behavior and performance, as reported by the events recorded on each computer, it is important to estimate precisely the clock differences and drift between the different computers in the system. This article studies the clock precision and stability of several computer systems, with different architectures. It also studies the typical network delay characteristics, since time synchronization algorithms rely on the exchange of network packets and are dependent on the symmetry of the delays. A very precise clock, based on the atomic time provided by the GPS satellite network, was used as a reference to measure clock drifts and network delays. The results obtained are of immediate use to all applications which depend on computer clocks or network time synchronization accuracy.

  20. Processing of visually presented clock times.

    Science.gov (United States)

    Goolkasian, P; Park, D C

    1980-11-01

    The encoding and representation of visually presented clock times was investigated in three experiments utilizing a comparative judgment task. Experiment 1 explored the effects of comparing times presented in different formats (clock face, digit, or word), and Experiment 2 examined angular distance effects created by varying positions of the hands on clock faces. In Experiment 3, encoding and processing differences between clock faces and digitally presented times were directly measured. Same/different reactions to digitally presented times were faster than to times presented on a clock face, and this format effect was found to be a result of differences in processing that occurred after encoding. Angular separation also had a limited effect on processing. The findings are interpreted within the framework of theories that refer to the importance of representational codes. The applicability to the data of Bank's semantic-coding theory, Paivio's dual-coding theory, and the levels-of-processing view of memory are discussed.

  1. Global synchronization of parallel processors using clock pulse width modulation

    Science.gov (United States)

    Chen, Dong; Ellavsky, Matthew R.; Franke, Ross L.; Gara, Alan; Gooding, Thomas M.; Haring, Rudolf A.; Jeanson, Mark J.; Kopcsay, Gerard V.; Liebsch, Thomas A.; Littrell, Daniel; Ohmacht, Martin; Reed, Don D.; Schenck, Brandon E.; Swetz, Richard A.

    2013-04-02

    A circuit generates a global clock signal with a pulse width modification to synchronize processors in a parallel computing system. The circuit may include a hardware module and a clock splitter. The hardware module may generate a clock signal and performs a pulse width modification on the clock signal. The pulse width modification changes a pulse width within a clock period in the clock signal. The clock splitter may distribute the pulse width modified clock signal to a plurality of processors in the parallel computing system.

  2. Combination of hypomorphic mutations of the Drosophila homologues of aryl hydrocarbon receptor and nucleosome assembly protein family genes disrupts morphogenesis, memory and detoxification.

    Science.gov (United States)

    Kuzin, Boris A; Nikitina, Ekaterina A; Cherezov, Roman O; Vorontsova, Julia E; Slezinger, Mikhail S; Zatsepina, Olga G; Simonova, Olga B; Enikolopov, Grigori N; Savvateeva-Popova, Elena V

    2014-01-01

    Aryl hydrocarbon receptor is essential for biological responses to endogenous and exogenous toxins in mammals. Its Drosophila homolog spineless plays an important role in fly morphogenesis. We have previously shown that during morphogenesis spineless genetically interacts with CG5017 gene, which encodes a nucleosome assembly factor and may affect cognitive function of the fly. We now demonstrate synergistic interactions of spineless and CG5017 in pathways controlling oxidative stress response and long-term memory formation in Drosophila melanogaster. Oxidative stress was induced by low doses of X-ray irradiation of flies carrying hypomorphic mutation of spineless, mutation of CG5017, and their combination. To determine the sensitivity of these mutants to pharmacological modifiers of the irradiation effect, we irradiated flies growing on standard medium supplemented by radiosensitizer furazidin and radioprotector serotonin. The effects of irradiation were investigated by analyzing leg and antenna morphological structures and by using real-time PCR to measure mRNA expression levels for spineless, Cyp6g1 and Gst-theta genes. We also examined long-term memory in these mutants using conditioned courtship suppression paradigm. Our results show that the interaction of spineless and CG5017 is important for regulation of morphogenesis, long-term memory formation, and detoxification during oxidative stress. Since spineless and CG5017 are evolutionary conserved, these results must be considered when evaluating the risk of combining similar mutations in other organisms, including humans.

  3. Combination of hypomorphic mutations of the Drosophila homologues of aryl hydrocarbon receptor and nucleosome assembly protein family genes disrupts morphogenesis, memory and detoxification.

    Directory of Open Access Journals (Sweden)

    Boris A Kuzin

    Full Text Available Aryl hydrocarbon receptor is essential for biological responses to endogenous and exogenous toxins in mammals. Its Drosophila homolog spineless plays an important role in fly morphogenesis. We have previously shown that during morphogenesis spineless genetically interacts with CG5017 gene, which encodes a nucleosome assembly factor and may affect cognitive function of the fly. We now demonstrate synergistic interactions of spineless and CG5017 in pathways controlling oxidative stress response and long-term memory formation in Drosophila melanogaster. Oxidative stress was induced by low doses of X-ray irradiation of flies carrying hypomorphic mutation of spineless, mutation of CG5017, and their combination. To determine the sensitivity of these mutants to pharmacological modifiers of the irradiation effect, we irradiated flies growing on standard medium supplemented by radiosensitizer furazidin and radioprotector serotonin. The effects of irradiation were investigated by analyzing leg and antenna morphological structures and by using real-time PCR to measure mRNA expression levels for spineless, Cyp6g1 and Gst-theta genes. We also examined long-term memory in these mutants using conditioned courtship suppression paradigm. Our results show that the interaction of spineless and CG5017 is important for regulation of morphogenesis, long-term memory formation, and detoxification during oxidative stress. Since spineless and CG5017 are evolutionary conserved, these results must be considered when evaluating the risk of combining similar mutations in other organisms, including humans.

  4. Rhythms of mammalian body temperature can sustain peripheral circadian clocks.

    Science.gov (United States)

    Brown, Steven A; Zumbrunn, Gottlieb; Fleury-Olela, Fabienne; Preitner, Nicolas; Schibler, Ueli

    2002-09-17

    Low-amplitude temperature oscillations can entrain the phase of circadian rhythms in several unicellular and multicellular organisms, including Neurospora and Drosophila. Because mammalian body temperature is subject to circadian variations of 1 degrees C-4 degrees C, we wished to determine whether these temperature cycles could serve as a Zeitgeber for circadian gene expression in peripheral cell types. In RAT1 fibroblasts cultured in vitro, circadian gene expression could be established by a square wave temperature rhythm with a (Delta)T of 4 degrees C (12 hr 37 degrees C/12 hr 33 degrees C). To examine whether natural body temperature rhythms can also affect circadian gene expression, we first measured core body temperature cycles in the peritoneal cavities of mice by radiotelemetry. We then reproduced these rhythms with high precision in the liquid medium of cultured fibroblasts for several days by means of a homemade computer-driven incubator. While these "in vivo" temperature rhythms were incapable of establishing circadian gene expression de novo, they could maintain previously induced rhythms for multiple days; by contrast, the rhythms of control cells kept at constant temperature rapidly dampened. Moreover, circadian oscillations of environmental temperature could reentrain circadian clocks in the livers of mice, probably via the changes they imposed upon both body temperature and feeding behavior. Interestingly, these changes in ambient temperature did not affect the phase of the central circadian pacemaker in the suprachiasmatic nucleus (SCN) of the hypothalamus. We postulate that both endogenous and environmental temperature cycles can participate in the synchronization of peripheral clocks in mammals.

  5. Dilatation effect of ''quantum clocks''

    International Nuclear Information System (INIS)

    Chylinski, Z.

    1981-01-01

    The relativistic dilatation effect of the life-time of unstable microparticles combined with quantum symmetry of their description results in the ''quantum-dilatation'' dilemma. It is due to the classical character of the relativity theory which here reveals itself in the classical world-line of the clock necessary in order to deduce the dilatation effect from the Lorentz transformation. It is shown how to solve this dilemma, basing on the relation continuum C 4 . Two types of measurements of time intervals, the direct and indirect one, are analyzed. The former type corresponds to the external space-time continuum, where any direct measurement takes place, and the latter, to the internal relation continuum C 4 , where the internal structures of isolated micro-systems are sunk. (author)

  6. A Screening of UNF Targets Identifies Rnb, a Novel Regulator of Drosophila Circadian Rhythms.

    Science.gov (United States)

    Kozlov, Anatoly; Jaumouillé, Edouard; Machado Almeida, Pedro; Koch, Rafael; Rodriguez, Joseph; Abruzzi, Katharine C; Nagoshi, Emi

    2017-07-12

    Behavioral circadian rhythms are controlled by multioscillator networks comprising functionally different subgroups of clock neurons. Studies have demonstrated that molecular clocks in the fruit fly Drosophila melanogaster are regulated differently in clock neuron subclasses to support their specific functions (Lee et al., 2016; Top et al., 2016). The nuclear receptor unfulfilled ( unf ) represents a regulatory node that provides the small ventral lateral neurons (s-LNvs) unique characteristics as the master pacemaker (Beuchle et al., 2012). We previously showed that UNF interacts with the s-LNv molecular clocks by regulating transcription of the core clock gene period ( per ) (Jaumouillé et al., 2015). To gain more insight into the mechanisms by which UNF contributes to the functioning of the circadian master pacemaker, we identified UNF target genes using chromatin immunoprecipitation. Our data demonstrate that a previously uncharacterized gene CG7837 , which we termed R and B ( Rnb ), acts downstream of UNF to regulate the function of the s-LNvs as the master circadian pacemaker. Mutations and LNv-targeted adult-restricted knockdown of Rnb impair locomotor rhythms. RNB localizes to the nucleus, and its loss-of-function blunts the molecular rhythms and output rhythms of the s-LNvs, particularly the circadian rhythms in PDF accumulation and axonal arbor remodeling. These results establish a second pathway by which UNF interacts with the molecular clocks in the s-LNvs and highlight the mechanistic differences in the molecular clockwork within the pacemaker circuit. SIGNIFICANCE STATEMENT Circadian behavior is generated by a pacemaker circuit comprising diverse classes of pacemaker neurons, each of which contains a molecular clock. In addition to the anatomical and functional diversity, recent studies have shown the mechanistic differences in the molecular clockwork among the pacemaker neurons in Drosophila Here, we identified the molecular characteristics

  7. Divergence times in Caenorhabditis and Drosophila inferred from direct estimates of the neutral mutation rate.

    Science.gov (United States)

    Cutter, Asher D

    2008-04-01

    Accurate inference of the dates of common ancestry among species forms a central problem in understanding the evolutionary history of organisms. Molecular estimates of divergence time rely on the molecular evolutionary prediction that neutral mutations and substitutions occur at the same constant rate in genomes of related species. This underlies the notion of a molecular clock. Most implementations of this idea depend on paleontological calibration to infer dates of common ancestry, but taxa with poor fossil records must rely on external, potentially inappropriate, calibration with distantly related species. The classic biological models Caenorhabditis and Drosophila are examples of such problem taxa. Here, I illustrate internal calibration in these groups with direct estimates of the mutation rate from contemporary populations that are corrected for interfering effects of selection on the assumption of neutrality of substitutions. Divergence times are inferred among 6 species each of Caenorhabditis and Drosophila, based on thousands of orthologous groups of genes. I propose that the 2 closest known species of Caenorhabditis shared a common ancestor <24 MYA (Caenorhabditis briggsae and Caenorhabditis sp. 5) and that Caenorhabditis elegans diverged from its closest known relatives <30 MYA, assuming that these species pass through at least 6 generations per year; these estimates are much more recent than reported previously with molecular clock calibrations from non-nematode phyla. Dates inferred for the common ancestor of Drosophila melanogaster and Drosophila simulans are roughly concordant with previous studies. These revised dates have important implications for rates of genome evolution and the origin of self-fertilization in Caenorhabditis.

  8. Females with a mutation in a nuclear-encoded mitochondrial protein pay a higher cost of survival than do males in Drosophila.

    Science.gov (United States)

    Melvin, Richard G; Ballard, J William O

    2011-07-01

    Males and females age at different rates in a variety of species, but the mechanisms underlying the difference is not understood. In this study, we investigated sex-specific costs of a naturally occurring mildly deleterious deletion (DTrp85, DVal86) in cytochrome c oxidase subunit 7A (cox7A) in Drosophila simulans. We observed that females and males homozygous for the mutation had 30% and 26% reduced Cox activity, respectively, compared with wild type. Furthermore, 4-day-old females had 34%-42% greater physical activity than males. Greater physical activity in mutant females was correlated with a 19% lower 50% survival compared with wild-type females. Mutant and wild-type males had equal survival. These data suggest that females paid a higher cost of the mutation than did males. The data demonstrate linking population genetics and structural modeling to experimental manipulations that lead to functional predictions of mitochondrial bioenergetics and organism aging.

  9. In vitro maturation of Drosophila melanogaster Spätzle protein with refolded Easter reveals a novel cleavage site within the prodomain.

    Science.gov (United States)

    Ursel, Christian; Fandrich, Uwe; Hoffmann, Anita; Sieg, Torsten; Ihling, Christian; Stubbs, Milton T

    2013-08-01

    Dorsoventral patterning during Drosophila melanogaster embryogenesis is mediated by a well-defined gradient of the mature NGF-like ligand Spätzle. Easter, the ultimate protease of a ventrally-restricted serine protease cascade, plays a key role in the regulation of the morphogenic gradient, catalyzing the activation cleavage of proSpätzle. As a result of alternative splicing, proSpätzle exists in multiple isoforms, almost all of which differ only in their prodomain. Although this domain is unstructured in isolation, it has a stabilizing influence on the mature cystine knot domain and is involved in the binding to the Toll receptor. Here, we report the expression and refolding of Easter, and show that the renatured enzyme performs the activation cleavage of two Spätzle isoforms. We determine the affinity of the prodomain for the cystine knot domain, and show that Easter performs a previously unknown secondary cleavage in each prodomain.

  10. Transmission delays in hardware clock synchronization

    Science.gov (United States)

    Shin, Kang G.; Ramanathan, P.

    1988-01-01

    Various methods, both with software and hardware, have been proposed to synchronize a set of physical clocks in a system. Software methods are very flexible and economical but suffer an excessive time overhead, whereas hardware methods require no time overhead but are unable to handle transmission delays in clock signals. The effects of nonzero transmission delays in synchronization have been studied extensively in the communication area in the absence of malicious or Byzantine faults. The authors show that it is easy to incorporate the ideas from the communication area into the existing hardware clock synchronization algorithms to take into account the presence of both malicious faults and nonzero transmission delays.

  11. Circadian clock components in the rat neocortex

    DEFF Research Database (Denmark)

    Rath, Martin Fredensborg; Rohde, Kristian; Fahrenkrug, Jan

    2013-01-01

    in the rat neocortex. Among these, Per1, Per2, Per3, Cry1, Bmal1, Nr1d1 and Dbp were found to exhibit daily rhythms. The amplitude of circadian oscillation in neocortical clock gene expression was damped and the peak delayed as compared with the SCN. Lesions of the SCN revealed that rhythmic clock gene...... expression in the neocortex is dependent on the SCN. In situ hybridization and immunohistochemistry showed that products of the canonical clock gene Per2 are located in perikarya throughout all areas of the neocortex. These findings show that local circadian oscillators driven by the SCN reside within...... neurons of the neocortex....

  12. Hearing regulates Drosophila aggression.

    Science.gov (United States)

    Versteven, Marijke; Vanden Broeck, Lies; Geurten, Bart; Zwarts, Liesbeth; Decraecker, Lisse; Beelen, Melissa; Göpfert, Martin C; Heinrich, Ralf; Callaerts, Patrick

    2017-02-21

    Aggression is a universal social behavior important for the acquisition of food, mates, territory, and social status. Aggression in Drosophila is context-dependent and can thus be expected to involve inputs from multiple sensory modalities. Here, we use mechanical disruption and genetic approaches in Drosophila melanogaster to identify hearing as an important sensory modality in the context of intermale aggressive behavior. We demonstrate that neuronal silencing and targeted knockdown of hearing genes in the fly's auditory organ elicit abnormal aggression. Further, we show that exposure to courtship or aggression song has opposite effects on aggression. Our data define the importance of hearing in the control of Drosophila intermale aggression and open perspectives to decipher how hearing and other sensory modalities are integrated at the neural circuit level.

  13. Regulation of circadian clock transcriptional output by CLOCK:BMAL1

    Science.gov (United States)

    Trott, Alexandra J.

    2018-01-01

    The mammalian circadian clock relies on the transcription factor CLOCK:BMAL1 to coordinate the rhythmic expression of 15% of the transcriptome and control the daily regulation of biological functions. The recent characterization of CLOCK:BMAL1 cistrome revealed that although CLOCK:BMAL1 binds synchronously to all of its target genes, its transcriptional output is highly heterogeneous. By performing a meta-analysis of several independent genome-wide datasets, we found that the binding of other transcription factors at CLOCK:BMAL1 enhancers likely contribute to the heterogeneity of CLOCK:BMAL1 transcriptional output. While CLOCK:BMAL1 rhythmic DNA binding promotes rhythmic nucleosome removal, it is not sufficient to generate transcriptionally active enhancers as assessed by H3K27ac signal, RNA Polymerase II recruitment, and eRNA expression. Instead, the transcriptional activity of CLOCK:BMAL1 enhancers appears to rely on the activity of ubiquitously expressed transcription factors, and not tissue-specific transcription factors, recruited at nearby binding sites. The contribution of other transcription factors is exemplified by how fasting, which effects several transcription factors but not CLOCK:BMAL1, either decreases or increases the amplitude of many rhythmically expressed CLOCK:BMAL1 target genes. Together, our analysis suggests that CLOCK:BMAL1 promotes a transcriptionally permissive chromatin landscape that primes its target genes for transcription activation rather than directly activating transcription, and provides a new framework to explain how environmental or pathological conditions can reprogram the rhythmic expression of clock-controlled genes. PMID:29300726

  14. Clock Drawing in Spatial Neglect: A Comprehensive Analysis of Clock Perimeter, Placement, and Accuracy

    Science.gov (United States)

    Chen, Peii; Goedert, Kelly M.

    2012-01-01

    Clock drawings produced by right-brain-damaged (RBD) individuals with spatial neglect often contain an abundance of empty space on the left while numbers and hands are placed on the right. However, the clock perimeter is rarely compromised in neglect patients’ drawings. By analyzing clock drawings produced by 71 RBD and 40 healthy adults, this study investigated whether the geometric characteristics of the clock perimeter reveal novel insights to understanding spatial neglect. Neglect participants drew smaller clocks than either healthy or non-neglect RBD participants. While healthy participants’ clock perimeter was close to circular, RBD participants drew radially extended ellipses. The mechanisms for these phenomena were investigated by examining the relation between clock-drawing characteristics and performance on six subtests of the Behavioral Inattention Test (BIT). The findings indicated that the clock shape was independent of any BIT subtest or the drawing placement on the test sheet and that the clock size was significantly predicted by one BIT subtest: the poorer the figure and shape copying, the smaller the clock perimeter. Further analyses revealed that in all participants, clocks decreased in size as they were placed farther from the center of the paper. However, even when neglect participants placed their clocks towards the center of the page, they were smaller than those produced by healthy or non-neglect RBD participants. These results suggest a neglect-specific reduction in the subjectively available workspace for graphic production from memory, consistent with the hypothesis that neglect patients are impaired in the ability to enlarge the attentional aperture. PMID:22390278

  15. Transcripts from the Circadian Clock: Telling Time and Season

    NARCIS (Netherlands)

    K. Brand (Karl)

    2011-01-01

    textabstractWe all know it when we wake mere moments before an alarm clock is scheduled to wake us: our body clock made the alarm clock redundant. This phenomenon is driven by an endogenous timer known as the biological, or circadian clock. Each revolution of the Earth about its own axis produces

  16. Effect of monochromatic light on circadian rhythmic expression of clock genes in the hypothalamus of chick.

    Science.gov (United States)

    Jiang, Nan; Wang, Zixu; Cao, Jing; Dong, Yulan; Chen, Yaoxing

    2017-08-01

    To clarify the effect of monochromatic light on circadian clock gene expression in chick hypothalamus, a total 240 newly hatched chickens were reared under blue light (BL), green light (GL), red light (RL) and white light (WL), respectively. On the post-hatched day 14, 24-h profiles of seven core clock genes (cClock, cBmal1, cBmal2, cCry1, cCry2, cPer2 and cPer3) were measured at six time points (CT 0, CT 4, CT 8, CT 12, CT 16, CT 20, circadian time). We found all these clock genes expressed with a significant rhythmicity in different light wavelength groups. Meanwhile, cClock and cBmal1 showed a high level under GL, and followed a corresponding high expression of cCry1. However, RL decreased the expression levels of these genes. Be consistent with the mRNA level, CLOCK and BMAL1 proteins also showed a high level under GL. The CLOCK-like immunoreactive neurons were observed not only in the SCN, but also in the non-SCN brain region such as the nucleus anterior medialis hypothalami, the periventricularis nucleus, the paraventricular nucleus and the median eminence. All these results are consistent with the auto-regulatory circadian feedback loop, and indicate that GL may play an important role on the circadian time generation and development in the chick hypothalamus. Our results also suggest that the circadian clock in the chick hypothalamus such as non-SCN brain region were involved in the regulation of photo information. Copyright © 2017 Elsevier B.V. All rights reserved.

  17. Pittendrigh: The Darwinian Clock-Watcher

    Indian Academy of Sciences (India)

    to our current understanding of how timing systems work in living organisms. .... to periodic factors in the geophysical environment. He postulated .... clocks against temperature, nutrition and light, while the latter needs maintenance of a stable.

  18. Programmable Clock Waveform Generation for CCD Readout

    Energy Technology Data Exchange (ETDEWEB)

    Vicente, J. de; Castilla, J.; Martinez, G.; Marin, J.

    2006-07-01

    Charge transfer efficiency in CCDs is closely related to the clock waveform. In this paper, an experimental framework to explore different FPGA based clock waveform generator designs is described. Two alternative design approaches for controlling the rise/fall edge times and pulse width of the CCD clock signal have been implemented: level-control and time-control. Both approaches provide similar characteristics regarding the edge linearity and noise. Nevertheless, dissimilarities have been found with respect to the area and frequency range of application. Thus, while the time-control approach consumes less area, the level control approach provides a wider range of clock frequencies since it does not suffer capacitor discharge effect. (Author) 8 refs.

  19. The Mechanics of Mechanical Watches and Clocks

    CERN Document Server

    Du, Ruxu

    2013-01-01

    "The Mechanics of Mechanical Watches and Clocks" presents historical views and mathematical models of mechanical watches and clocks. Although now over six hundred years old, mechanical watches and clocks are still popular luxury items that fascinate many people around the world. However few have examined the theory of how they work as presented in this book. The illustrations and computer animations are unique and have never been published before. It will be of significant interest to researchers in mechanical engineering, watchmakers and clockmakers, as well as people who have an engineering background and are interested in mechanical watches and clocks. It will also inspire people in other fields of science and technology, such as mechanical engineering and electronics engineering, to advance their designs. Professor Ruxu Du works at the Chinese University of Hong Kong, China. Assistant Professor Longhan Xie works at the South China University of Technology, China.

  20. Cellular Reprogramming–Turning the Clock Back

    Indian Academy of Sciences (India)

    Home; Journals; Resonance – Journal of Science Education; Volume 18; Issue 6. Cellular Reprogramming - Turning the Clock Back - Nobel Prize in Physiology or Medicine, 2012. Deepa Subramanyam. General Article Volume 18 Issue 6 June 2013 pp 514-521 ...

  1. A GPS Satellite Clock Offset Prediction Method Based on Fitting Clock Offset Rates Data

    Directory of Open Access Journals (Sweden)

    WANG Fuhong

    2016-12-01

    Full Text Available It is proposed that a satellite atomic clock offset prediction method based on fitting and modeling clock offset rates data. This method builds quadratic model or linear model combined with periodic terms to fit the time series of clock offset rates, and computes the model coefficients of trend with the best estimation. The clock offset precisely estimated at the initial prediction epoch is directly adopted to calculate the model coefficient of constant. The clock offsets in the rapid ephemeris (IGR provided by IGS are used as modeling data sets to perform certain experiments for different types of GPS satellite clocks. The results show that the clock prediction accuracies of the proposed method for 3, 6, 12 and 24 h achieve 0.43, 0.58, 0.90 and 1.47 ns respectively, which outperform the traditional prediction method based on fitting original clock offsets by 69.3%, 61.8%, 50.5% and 37.2%. Compared with the IGU real-time clock products provided by IGS, the prediction accuracies of the new method have improved about 15.7%, 23.7%, 27.4% and 34.4% respectively.

  2. Epigenetic and Posttranslational Modifications in Light Signal Transduction and the Circadian Clock in Neurospora crassa

    Directory of Open Access Journals (Sweden)

    Marco Proietto

    2015-07-01

    Full Text Available Blue light, a key abiotic signal, regulates a wide variety of physiological processes in many organisms. One of these phenomena is the circadian rhythm presents in organisms sensitive to the phase-setting effects of blue light and under control of the daily alternation of light and dark. Circadian clocks consist of autoregulatory alternating negative and positive feedback loops intimately connected with the cellular metabolism and biochemical processes. Neurospora crassa provides an excellent model for studying the molecular mechanisms involved in these phenomena. The White Collar Complex (WCC, a blue-light receptor and transcription factor of the circadian oscillator, and Frequency (FRQ, the circadian clock pacemaker, are at the core of the Neurospora circadian system. The eukaryotic circadian clock relies on transcriptional/translational feedback loops: some proteins rhythmically repress their own synthesis by inhibiting the activity of their transcriptional factors, generating self-sustained oscillations over a period of about 24 h. One of the basic mechanisms that perpetuate self-sustained oscillations is post translation modification (PTM. The acronym PTM generically indicates the addition of acetyl, methyl, sumoyl, or phosphoric groups to various types of proteins. The protein can be regulatory or enzymatic or a component of the chromatin. PTMs influence protein stability, interaction, localization, activity, and chromatin packaging. Chromatin modification and PTMs have been implicated in regulating circadian clock function in Neurospora. Research into the epigenetic control of transcription factors such as WCC has yielded new insights into the temporal modulation of light-dependent gene transcription. Here we report on epigenetic and protein PTMs in the regulation of the Neurospora crassa circadian clock. We also present a model that illustrates the molecular mechanisms at the basis of the blue light control of the circadian clock.

  3. Clock gene variation in Tachycineta swallows.

    Science.gov (United States)

    Dor, Roi; Cooper, Caren B; Lovette, Irby J; Massoni, Viviana; Bulit, Flor; Liljesthrom, Marcela; Winkler, David W

    2012-01-01

    Many animals use photoperiod cues to synchronize reproduction with environmental conditions and thereby improve their reproductive success. The circadian clock, which creates endogenous behavioral and physiological rhythms typically entrained to photoperiod, is well characterized at the molecular level. Recent work provided evidence for an association between Clock poly-Q length polymorphism and latitude and, within a population, an association with the date of laying and the length of the incubation period. Despite relatively high overall breeding synchrony, the timing of clutch initiation has a large impact on the fitness of swallows in the genus Tachycineta. We compared length polymorphism in the Clock poly-Q region among five populations from five different Tachycineta species that breed across a hemisphere-wide latitudinal gradient (Fig. 1). Clock poly-Q variation was not associated with latitude; however, there was an association between Clock poly-Q allele diversity and the degree of clutch size decline within breeding seasons. We did not find evidence for an association between Clock poly-Q variation and date of clutch initiation in for any of the five Tachycineta species, nor did we found a relationship between incubation duration and Clock genotype. Thus, there is no general association between latitude, breeding phenology, and Clock polymorphism in this clade of closely related birds.Figure 1Photos of Tachycineta swallows that were used in this study: A) T. bicolor from Ithaca, New York, B) T. leucorrhoa from Chascomús, Argentina, C) T. albilinea from Hill Bank, Belize, D) T. meyeni from Puerto Varas, Chile, and E) T. thalassina from Mono Lake, California, Photographers: B: Valentina Ferretti; A, C-E: David Winkler.

  4. Reduced Kalman Filters for Clock Ensembles

    Science.gov (United States)

    Greenhall, Charles A.

    2011-01-01

    This paper summarizes the author's work ontimescales based on Kalman filters that act upon the clock comparisons. The natural Kalman timescale algorithm tends to optimize long-term timescale stability at the expense of short-term stability. By subjecting each post-measurement error covariance matrix to a non-transparent reduction operation, one obtains corrected clocks with improved short-term stability and little sacrifice of long-term stability.

  5. Do Caucasian and Asian clocks tick differently?

    Directory of Open Access Journals (Sweden)

    A.A. Barbosa

    Full Text Available The Period 3 and Clock genes are important components of the mammalian molecular circadian system. Studies have shown association between polymorphisms in these clock genes and circadian phenotypes in different populations. Nevertheless, differences in the pattern of allele frequency and genotyping distribution are systematically observed in studies with different ethnic groups. To investigate and compare the pattern of distribution in a sample of Asian and Caucasian populations living in Brazil, we evaluated two well-studied polymorphisms in the clock genes: a variable number of tandem repeats (VNTR in PER3 and a single nucleotide polymorphism (SNP in CLOCK. The aim of this investigation was to search for clues about human evolutionary processes related to circadian rhythms. We selected 109 Asian and 135 Caucasian descendants. The frequencies of the shorter allele (4 repeats in the PER3 gene and the T allele in the CLOCK gene among Asians (0.86 and 0.84, respectively were significantly higher than among Caucasians (0.69 and 0.71, respectively. Our results directly confirmed the different distribution of these polymorphisms between the Asian and Caucasian ethnic groups. Given the genetic differences found between groups, two points became evident: first, ethnic variations may have implications for the interpretation of results in circadian rhythm association studies, and second, the question may be raised about which evolutionary conditions shaped these genetic clock variations.

  6. Optical lattice clock with Strontium atoms

    International Nuclear Information System (INIS)

    Baillard, X.

    2008-01-01

    This thesis presents the latest achievements regarding the optical lattice clock with Strontium atoms developed at LNE-SYRTE. After a review of the different types of optical clocks that are currently under development, we stress on the concept of optical lattice clock which was first imagined for Sr 87 using the 1 S 0 → 3 P 0 transition. We exhibit the features of this atom, in particular the concept of magic wavelength for the trap, and the achievable performances for this kind of clock. The second part presents the experimental aspects, insisting particularly on the ultra-stable laser used for the interrogation of the atoms which is a central part of the experiment. Among the latest improvements, an optical pumping phase and an interrogation phase using a magnetic field have been added in order to refine the evaluation of the Zeeman effect. Finally, the last part presents the experimental results. The last evaluation of the clock using Sr 87 atoms allowed us to reach a frequency accuracy of 2.6*10 -15 and a measurement in agreement with the one made at JILA (Tokyo university) at the 10 -15 level. On another hand, thanks to recent theoretical proposals, we made a measurement using the bosonic isotope Sr 88 by adapting the experimental setup. This measurement represents the first evaluation for this type of clock, with a frequency accuracy of 7*10 -14 . (author)

  7. Towards Self-Clocked Gated OCDMA Receiver

    Science.gov (United States)

    Idris, S.; Osadola, T.; Glesk, I.

    2013-02-01

    A novel incoherent OCDMA receiver with incorporated all-optical clock recovery for self-synchronization of a time gate for the multi access interferences (MAI) suppression and minimizing the effect of data time jitter in incoherent OCDMA system was successfully developed and demonstrated. The solution was implemented and tested in a multiuser environment in an out of the laboratory OCDMA testbed with two-dimensional wavelength-hopping time-spreading coding scheme and OC-48 (2.5 Gbp/s) data rate. The self-clocked all-optical time gate uses SOA-based fibre ring laser optical clock, recovered all-optically from the received OCDMA traffic to control its switching window for cleaning the autocorrelation peak from the surrounding MAI. A wider eye opening was achieved when the all-optically recovered clock from received data was used for synchronization if compared to a static approach with the RF clock being generated by a RF synthesizer. Clean eye diagram was also achieved when recovered clock is used to drive time gating.

  8. Synergistic regulation of the mouse orphan nuclear receptor SHP gene promoter by CLOCK-BMAL1 and LRH-1

    International Nuclear Information System (INIS)

    Oiwa, Ako; Kakizawa, Tomoko; Miyamoto, Takahide; Yamashita, Koh; Jiang, Wei; Takeda, Teiji; Suzuki, Satoru; Hashizume, Kiyoshi

    2007-01-01

    Small heterodimer partner (SHP; NR0B2) is an orphan nuclear receptor and acts as a repressor for wide variety of nuclear hormone receptors. We demonstrated here that mouse SHP mRNA showed a circadian expression pattern in the liver. Transient transfection of the mSHP promoter demonstrated that CLOCK-BMAL1, core circadian clock components, bound to E-box (CACGTG), and stimulated the promoter activity by 4-fold. Liver receptor homologue-1 (LRH-1; NR5A2) stimulated the mSHP promoter, and CLOCK-BMAL1 synergistically enhanced the LRH-1-mediated transactivation. Interestingly, SHP did not affect the CLOCK-BMAL1-mediated promoter activity, but strongly repressed the synergistic activation of CLOCK-BMAL1 and LRH-1. Furthermore, in vitro pull-down assays revealed the existence of direct protein-protein interaction between LRH-1 and CLOCK. In summary, this study shows that CLOCK-BMAL1, LRH-1 and SHP coordinately regulate the mSHP gene to generate the circadian oscillation. The cyclic expression of mSHP may affect daily activity of other nuclear receptors and contribute to circadian liver functions

  9. Diversity of Internal Sensory Neuron Axon Projection Patterns Is Controlled by the POU-Domain Protein Pdm3 in Drosophila Larvae.

    Science.gov (United States)

    Qian, Cheng Sam; Kaplow, Margarita; Lee, Jennifer K; Grueber, Wesley B

    2018-02-21

    Internal sensory neurons innervate body organs and provide information about internal state to the CNS to maintain physiological homeostasis. Despite their conservation across species, the anatomy, circuitry, and development of internal sensory systems are still relatively poorly understood. A largely unstudied population of larval Drosophila sensory neurons, termed tracheal dendrite (td) neurons, innervate internal respiratory organs and may serve as a model for understanding the sensing of internal states. Here, we characterize the peripheral anatomy, central axon projection, and diversity of td sensory neurons. We provide evidence for prominent expression of specific gustatory receptor genes in distinct populations of td neurons, suggesting novel chemosensory functions. We identify two anatomically distinct classes of td neurons. The axons of one class project to the subesophageal zone (SEZ) in the brain, whereas the other terminates in the ventral nerve cord (VNC). We identify expression and a developmental role of the POU-homeodomain transcription factor Pdm3 in regulating the axon extension and terminal targeting of SEZ-projecting td neurons. Remarkably, ectopic Pdm3 expression is alone sufficient to switch VNC-targeting axons to SEZ targets, and to induce the formation of putative synapses in these ectopic target zones. Our data thus define distinct classes of td neurons, and identify a molecular factor that contributes to diversification of axon targeting. These results introduce a tractable model to elucidate molecular and circuit mechanisms underlying sensory processing of internal body status and physiological homeostasis. SIGNIFICANCE STATEMENT How interoceptive sensory circuits develop, including how sensory neurons diversify and target distinct central regions, is still poorly understood, despite the importance of these sensory systems for maintaining physiological homeostasis. Here, we characterize classes of Drosophila internal sensory neurons (td

  10. Whole genome phylogenies for multiple Drosophila species

    Directory of Open Access Journals (Sweden)

    Seetharam Arun

    2012-12-01

    Full Text Available Abstract Background Reconstructing the evolutionary history of organisms using traditional phylogenetic methods may suffer from inaccurate sequence alignment. An alternative approach, particularly effective when whole genome sequences are available, is to employ methods that don’t use explicit sequence alignments. We extend a novel phylogenetic method based on Singular Value Decomposition (SVD to reconstruct the phylogeny of 12 sequenced Drosophila species. SVD analysis provides accurate comparisons for a high fraction of sequences within whole genomes without the prior identification of orthologs or homologous sites. With this method all protein sequences are converted to peptide frequency vectors within a matrix that is decomposed to provide simplified vector representations for each protein of the genome in a reduced dimensional space. These vectors are summed together to provide a vector representation for each species, and the angle between these vectors provides distance measures that are used to construct species trees. Results An unfiltered whole genome analysis (193,622 predicted proteins strongly supports the currently accepted phylogeny for 12 Drosophila species at higher dimensions except for the generally accepted but difficult to discern sister relationship between D. erecta and D. yakuba. Also, in accordance with previous studies, many sequences appear to support alternative phylogenies. In this case, we observed grouping of D. erecta with D. sechellia when approximately 55% to 95% of the proteins were removed using a filter based on projection values or by reducing resolution by using fewer dimensions. Similar results were obtained when just the melanogaster subgroup was analyzed. Conclusions These results indicate that using our novel phylogenetic method, it is possible to consult and interpret all predicted protein sequences within multiple whole genomes to produce accurate phylogenetic estimations of relatedness between

  11. Cancer in Drosophila

    DEFF Research Database (Denmark)

    Herranz, Héctor; Eichenlaub, Teresa; Cohen, Stephen M

    2016-01-01

    Cancer genomics has greatly increased our understanding of the complexity of the genetic and epigenetic changes found in human tumors. Understanding the functional relationships among these elements calls for the use of flexible genetic models. We discuss the use of Drosophila models to study...

  12. The translation factors of Drosophila melanogaster.

    Science.gov (United States)

    Marygold, Steven J; Attrill, Helen; Lasko, Paul

    2017-01-02

    Synthesis of polypeptides from mRNA (translation) is a fundamental cellular process that is coordinated and catalyzed by a set of canonical 'translation factors'. Surprisingly, the translation factors of Drosophila melanogaster have not yet been systematically identified, leading to inconsistencies in their nomenclature and shortcomings in functional (Gene Ontology, GO) annotations. Here, we describe the complete set of translation factors in D. melanogaster, applying nomenclature already in widespread use in other species, and revising their functional annotation. The collection comprises 43 initiation factors, 12 elongation factors, 3 release factors and 6 recycling factors, totaling 64 of which 55 are cytoplasmic and 9 are mitochondrial. We also provide an overview of notable findings and particular insights derived from Drosophila about these factors. This catalog, together with the incorporation of the improved nomenclature and GO annotation into FlyBase, will greatly facilitate access to information about the functional roles of these important proteins.

  13. Poly(ADP-ribose) Glycohydrolase and Poly(ADP-ribose)-interacting Protein Hrp38 Regulate Pattern Formation during Drosophila Eye Development

    Science.gov (United States)

    Ji, Yingbiao; Jarnik, Michael; Tulin, Alexei V.

    2013-01-01

    Drosophila Hrp38, a homolog of human hnRNP A1, has been shown to regulate splicing, but its function can be modified by poly(ADP-ribosyl)ation. Notwithstanding such findings, our understanding of the roles of poly(ADP-ribosyl)ated Hrp38 on development is limited. Here, we have demonstrated that Hrp38 is essential for fly eye development based on a rough-eye phenotype with disorganized ommatidia observed in adult escapers of the hrp38 mutant. We also observed that Poly(ADP-ribose) Glycohydrolase (Parg) loss-of-function, which caused increased Hrp38 poly(ADP-ribosyl)ation, also resulted in the rough-eye phenotype with disrupted ommatidial lattice and reduced number of photoreceptor cells. In addition, ectopic expression of DE-cadherin, which is required for retinal morphogenesis, fully rescued the rough-eye phenotype of the hrp38 mutant. Similarly, Parg mutant eye clones had decreased expression level of DE-cadherin with orientation defects, which is reminiscent of DE-cadherin mutant eye phenotype. Therefore, our results suggest that Hrp38 poly(ADP-ribosyl)ation controls eye pattern formation via regulation of DE-cadherin expression, a finding which has implications for understanding the pathogenic mechanisms of Hrp38-related Fragile X syndrome and PARP1-related retinal degeneration diseases. PMID:23711619

  14. Purification, isolation, crystallization, and preliminary X-ray diffraction study of the BTB domain of the centrosomal protein 190 from Drosophila melanogaster

    Science.gov (United States)

    Boyko, K. M.; Nikolaeva, A. Yu.; Kachalova, G. S.; Bonchuk, A. N.; Popov, V. O.

    2017-11-01

    The spatial organization of the genome is controlled by a special class of architectural proteins, including proteins containing BTB domains that are able to dimerize or multimerize. The centrosomal protein 190 is one of such architectural proteins. The purification, crystallization, and preliminary X-ray diffraction study of the BTB domain of the centrosomal protein 190 are reported. The crystallization conditions were found by the vapor-diffusion technique. The crystals diffracted to 1.5 Å resolution and belonged to sp. gr. P3221. The structure was solved by the molecular replacement method. The structure refinement is currently underway.

  15. The expression of the clock gene cycle has rhythmic pattern and is affected by photoperiod in the moth Sesamia nonagrioides.

    Science.gov (United States)

    Kontogiannatos, Dimitrios; Gkouvitsas, Theodoros; Kourti, Anna

    2017-06-01

    To obtain clues to the link between the molecular mechanism of circadian and photoperiod clocks, we have cloned the circadian clock gene cycle (Sncyc) in the corn stalk borer, Sesamia nonagrioides, which undergoes facultative diapause controlled by photoperiod. Sequence analysis revealed a high degree of conservation among insects for this gene. SnCYC consists of 667 amino acids and structural analysis showed that it contains a BCTR domain in its C-terminal in addition to the common domains found in Drosophila CYC, i.e. bHLH, PAS-A, PAS-B domains. The results revealed that the sequence of Sncyc showed a similarity to that of its mammalian orthologue, Bmal1. We also investigated the expression patterns of Sncyc in the brain of larvae growing under long-day 16L: 8D (LD), constant darkness (DD) and short-day 10L: 14D (SD) conditions using qRT-PCR assays. The mRNAs of Sncyc expression was rhythmic in LD, DD and SD cycles. Also, it is remarkable that the photoperiodic conditions affect the expression patterns and/or amplitudes of circadian clock gene Sncyc. This gene is associated with diapause in S. nonagrioides, because under SD (diapause conditions) the photoperiodic signal altered mRNA accumulation. Sequence and expression analysis of cyc in S. nonagrioides shows interesting differences compared to Drosophila where this gene does not oscillate or change in expression patterns in response to photoperiod, suggesting that this species is an interesting new model to study the molecular control of insect circadian and photoperiodic clocks. Copyright © 2017 Elsevier Inc. All rights reserved.

  16. Modeling Human Cancers in Drosophila.

    Science.gov (United States)

    Sonoshita, M; Cagan, R L

    2017-01-01

    Cancer is a complex disease that affects multiple organs. Whole-body animal models provide important insights into oncology that can lead to clinical impact. Here, we review novel concepts that Drosophila studies have established for cancer biology, drug discovery, and patient therapy. Genetic studies using Drosophila have explored the roles of oncogenes and tumor-suppressor genes that when dysregulated promote cancer formation, making Drosophila a useful model to study multiple aspects of transformation. Not limited to mechanism analyses, Drosophila has recently been showing its value in facilitating drug development. Flies offer rapid, efficient platforms by which novel classes of drugs can be identified as candidate anticancer leads. Further, we discuss the use of Drosophila as a platform to develop therapies for individual patients by modeling the tumor's genetic complexity. Drosophila provides both a classical and a novel tool to identify new therapeutics, complementing other more traditional cancer tools. © 2017 Elsevier Inc. All rights reserved.

  17. A comparison of Frost expression among species and life stages of Drosophila.

    Science.gov (United States)

    Bing, X; Zhang, J; Sinclair, Brent J

    2012-02-01

    Frost (Fst) is a gene associated with cold exposure in Drosophila melanogaster. We used real-time PCR to assess whether cold exposure induces expression of Fst in 10 different life stages of D. melanogaster, and adults of seven other Drosophila species. We exposed groups of individuals to 0 °C (2 h), followed by 1 h recovery (22 °C). Frost was significantly upregulated in response to cold in eggs, third instar larvae, and 2- and 5-day-old male and female adults in D. melanogaster. Life stages in which cold did not upregulate Fst had high constitutive expression. Frost is located on the opposite strand of an intron of Diuretic hormone (DH), but cold exposure did not upregulate DH. Frost orthologues were identified in six other species within the Melanogaster group (Drosophila sechellia, Drosophila simulans, Drosophila yakuba, Drosophila erecta, Drosophila ananassae and Drosophila mauritiana). Frost orthologues were upregulated in response to cold exposure in both sexes in adults of all of these species. The predicted structure of a putative Frost consensus protein shows highly conserved tandem repeats of motifs involved in cell signalling (PEST and TRAF2), suggesting that Fst might encode an adaptor protein involved in acute stress or apoptosis signalling in vivo. © 2011 The Authors. Insect Molecular Biology © 2011 The Royal Entomological Society.

  18. Acetylcholine receptors and cholinergic ligands: biochemical and genetic aspects in Torpedo californica and Drosophila melanogaster

    International Nuclear Information System (INIS)

    Rosenthal, L.S.

    1987-01-01

    This study evaluates the biochemical and genetic aspects of the acetylcholine receptor proteins and cholinergic ligands in Drosophila melanogaster and Torpedo californica. Included are (1) a comparative study of nicotinic ligand-induced cation release from acetylcholine receptors isolated from Torpedo californica and from Drosophila melanogaster, (2) solution studies of the cholinergic ligands, nikethamide and ethamivan, aimed at measuring internal molecular rotational barriers in solvents of different polarity; and (3) the isolation and characterization of the gene(s) for the acetylcholine receptor in Drosophila melasogaster. Acetylcholine receptor proteins isolated from Drosphila melanogaster heads were found to behave kinetically similar (with regards to cholinergic ligand-induced 155 Eu: 3+ displacement from prelabeled proteins) to receptor proteins isolated from Torpedo californica electric tissue, providing additional biochemical evidence for the existence of a Drosophila acetylcholine receptor

  19. Survival rate and expression of Heat-shock protein 70 and Frost genes after temperature stress in Drosophila melanogaster lines that are selected for recovery time from temperature coma.

    Science.gov (United States)

    Udaka, Hiroko; Ueda, Chiaki; Goto, Shin G

    2010-12-01

    In this study, we investigated the physiological mechanisms underlying temperature tolerance using Drosophila melanogaster lines with rapid, intermediate, or slow recovery from heat or chill coma that were established by artificial selection or by free recombination without selection. Specifically, we focused on the relationships among their recovery from heat or chill coma, survival after severe heat or cold, and survival enhanced by rapid cold hardening (RCH) or heat hardening. The recovery time from heat coma was not related to the survival rate after severe heat. The line with rapid recovery from chill coma showed a higher survival rate after severe cold exposure, and therefore the same mechanisms are likely to underlie these phenotypes. The recovery time from chill coma and survival rate after severe cold were unrelated to RCH-enhanced survival. We also examined the expression of two genes, Heat-shock protein 70 (Hsp70) and Frost, in these lines to understand the contribution of these stress-inducible genes to intraspecific variation in recovery from temperature coma. The line showing rapid recovery from heat coma did not exhibit higher expression of Hsp70 and Frost. In addition, Hsp70 and Frost transcription levels were not correlated with the recovery time from chill coma. Thus, Hsp70 and Frost transcriptional regulation was not involved in the intraspecific variation in recovery from temperature coma. Copyright © 2010 Elsevier Ltd. All rights reserved.

  20. dFOXO Activates Large and Small Heat Shock Protein Genes in Response to Oxidative Stress to Maintain Proteostasis in Drosophila.

    Science.gov (United States)

    Donovan, Marissa R; Marr, Michael T

    2016-09-02

    Maintaining protein homeostasis is critical for survival at the cellular and organismal level (Morimoto, R. I. (2011) Cold Spring Harb. Symp. Quant. Biol. 76, 91-99). Cells express a family of molecular chaperones, the heat shock proteins, during times of oxidative stress to protect against proteotoxicity. We have identified a second stress responsive transcription factor, dFOXO, that works alongside the heat shock transcription factor to activate transcription of both the small heat shock protein and the large heat shock protein genes. This expression likely protects cells from protein misfolding associated with oxidative stress. Here we identify the regions of the Hsp70 promoter essential for FOXO-dependent transcription using in vitro methods and find a physiological role for FOXO-dependent expression of heat shock proteins in vivo. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  1. The PTK7-Related Transmembrane Proteins Off-track and Off-track 2 Are Co-receptors for Drosophila Wnt2 Required for Male Fertility

    OpenAIRE

    Linnemannstöns, Karen; Ripp, Caroline; Honemann-Capito, Mona; Brechtel-Curth, Katja; Hedderich, Marie; Wodarz, Andreas

    2014-01-01

    Wnt proteins regulate many developmental processes and are required for tissue homeostasis in adult animals. The cellular responses to Wnts are manifold and are determined by the respective Wnt ligand and its specific receptor complex in the plasma membrane. Wnt receptor complexes contain a member of the Frizzled family of serpentine receptors and a co-receptor, which commonly is a single-pass transmembrane protein. Vertebrate protein tyrosine kinase 7 (PTK7) was identified as a Wnt co-recept...

  2. A model of guarded recursion with clock synchronisation

    DEFF Research Database (Denmark)

    Bizjak, Aleš; Møgelberg, Rasmus Ejlers

    2015-01-01

    productivity to be captured in types. The calculus uses clocks representing time streams and clock quantifiers which allow limited and controlled elimination of modalities. The calculus has since been extended to dependent types by Møgelberg. Both works give denotational semantics but no rewrite semantics....... In previous versions of this calculus, different clocks represented separate time streams and clock synchronisation was prohibited. In this paper we show that allowing clock synchronisation is safe by constructing a new model of guarded recursion and clocks. This result will greatly simplify the type theory...... by removing freshness restrictions from typing rules, and is a necessary step towards defining rewrite semantics, and ultimately implementing the calculus....

  3. A new tagging system for production of recombinant proteins in Drosophila S2 cells using the third domain of the urokinase receptor

    DEFF Research Database (Denmark)

    Gårdsvoll, Henrik; Hansen, Line V; Jørgensen, Thomas J D

    2007-01-01

    The use of protein fusion tag technology greatly facilitates detection, expression and purification of recombinant proteins, and the demands for new and more effective systems are therefore expanding. We have used a soluble truncated form of the third domain of the urokinase receptor...... as a convenient C-terminal fusion partner for various recombinant extracellular human proteins used in basic cancer research. The stability of this cystein-rich domain, which structure adopts a three-finger fold, provides an important asset for its applicability as a fusion tag for expression of recombinant...... chromatography procedure using the immobilized anti-uPAR monoclonal antibody R2. An optional enterokinase cleavage site is included between the various recombinant proteins and the linker region of the tag, which enables generation of highly pure preparations of tag-free recombinant proteins. Using this system...

  4. Sugars, the clock and transition to flowering

    Directory of Open Access Journals (Sweden)

    Mohammad Reza eBolouri Moghaddam

    2013-02-01

    Full Text Available Sugars do not only act as source of energy, but they also act as signals in plants. This mini review summarizes the emerging links between sucrose-mediated signaling and the cellular networks involved in flowering time control and defense. Cross-talks with gibberellin (GA and jasmonate (JA signaling pathways are highlighted. The circadian clock fulfills a crucial role at the heart of cellular networks and the bilateral relation between sugar signaling and the clock is discussed. It is proposed that important factors controlling plant growth (DELLAs, PIFs, invertases and trehalose- 6-phosphate or T6P might fulfill central roles in the transition to flowering as well. The emerging concept of ‘sweet immunity’, modulated by the clock, might at least partly rely on a sucrose-specific signaling pathway that needs further exploration.

  5. Robustness from flexibility in the fungal circadian clock

    Directory of Open Access Journals (Sweden)

    Akman Ozgur E

    2010-06-01

    Full Text Available Abstract Background Robustness is a central property of living systems, enabling function to be maintained against environmental perturbations. A key challenge is to identify the structures in biological circuits that confer system-level properties such as robustness. Circadian clocks allow organisms to adapt to the predictable changes of the 24-hour day/night cycle by generating endogenous rhythms that can be entrained to the external cycle. In all organisms, the clock circuits typically comprise multiple interlocked feedback loops controlling the rhythmic expression of key genes. Previously, we showed that such architectures increase the flexibility of the clock's rhythmic behaviour. We now test the relationship between flexibility and robustness, using a mathematical model of the circuit controlling conidiation in the fungus Neurospora crassa. Results The circuit modelled in this work consists of a central negative feedback loop, in which the frequency (frq gene inhibits its transcriptional activator white collar-1 (wc-1, interlocked with a positive feedback loop in which FRQ protein upregulates WC-1 production. Importantly, our model reproduces the observed entrainment of this circuit under light/dark cycles with varying photoperiod and cycle duration. Our simulations show that whilst the level of frq mRNA is driven directly by the light input, the falling phase of FRQ protein, a molecular correlate of conidiation, maintains a constant phase that is uncoupled from the times of dawn and dusk. The model predicts the behaviour of mutants that uncouple WC-1 production from FRQ's positive feedback, and shows that the positive loop enhances the buffering of conidiation phase against seasonal photoperiod changes. This property is quantified using Kitano's measure for the overall robustness of a regulated system output. Further analysis demonstrates that this functional robustness is a consequence of the greater evolutionary flexibility conferred on

  6. The Drosophila gene CG9918 codes for a pyrokinin-1 receptor

    DEFF Research Database (Denmark)

    Cazzamali, Giuseppe; Torp, Malene; Hauser, Frank

    2005-01-01

    The database from the Drosophila Genome Project contains a gene, CG9918, annotated to code for a G protein-coupled receptor. We cloned the cDNA of this gene and functionally expressed it in Chinese hamster ovary cells. We tested a library of about 25 Drosophila and other insect neuropeptides......, and seven insect biogenic amines on the expressed receptor and found that it was activated by low concentrations of the Drosophila neuropeptide, pyrokinin-1 (TGPSASSGLWFGPRLamide; EC50, 5 x 10(-8) M). The receptor was also activated by other Drosophila neuropeptides, terminating with the sequence PRLamide...... (Hug-gamma, ecdysis-triggering-hormone-1, pyrokinin-2), but in these cases about six to eight times higher concentrations were needed. The receptor was not activated by Drosophila neuropeptides, containing a C-terminal PRIamide sequence (such as ecdysis-triggering-hormone-2), or PRVamide (such as capa...

  7. Nematocytes: Discovery and characterization of a novel anculeate hemocyte in Drosophila falleni and Drosophila phalerata.

    Directory of Open Access Journals (Sweden)

    Julianna Bozler

    Full Text Available Immune challenges, such as parasitism, can be so pervasive and deleterious that they constitute an existential threat to a species' survival. In response to these ecological pressures, organisms have developed a wide array of novel behavioral, cellular, and molecular adaptations. Research into these immune defenses in model systems has resulted in a revolutionary understanding of evolution and functional biology. As the field has expanded beyond the limited number of model organisms our appreciation of evolutionary innovation and unique biology has widened as well. With this in mind, we have surveyed the hemolymph of several non-model species of Drosophila. Here we identify and describe a novel hemocyte, type-II nematocytes, found in larval stages of numerous Drosophila species. Examined in detail in Drosophila falleni and Drosophila phalerata, we find that these remarkable cells are distinct from previously described hemocytes due to their anucleate state (lacking a nucleus and unusual morphology. Type-II nematocytes are long, narrow cells with spindle-like projections extending from a cell body with high densities of mitochondria and microtubules, and exhibit the ability to synthesize proteins. These properties are unexpected for enucleated cells, and together with our additional characterization, we demonstrate that these type-II nematocytes represent a biological novelty. Surprisingly, despite the absence of a nucleus, we observe through live cell imaging that these cells remain motile with a highly dynamic cellular shape. Furthermore, these cells demonstrate the ability to form multicellular structures, which we suggest may be a component of the innate immune response to macro-parasites. In addition, live cell imaging points to a large nucleated hemocyte, type-I nematocyte, as the progenitor cell, leading to enucleation through a budding or asymmetrical division process rather than nuclear ejection: This study is the first to report such a

  8. Tolerance in Drosophila

    OpenAIRE

    Atkinson, Nigel S.

    2009-01-01

    The set of genes that underlie ethanol tolerance (inducible resistance) are likely to overlap with the set of genes responsible for ethanol addiction. Whereas addiction is difficult to recognize in simple model systems, behavioral tolerance is readily identifiable and can be induced in large populations of animals. Thus, tolerance lends itself to analysis in model systems with powerful genetics. Drosophila melanogaster has been used by a variety of laboratories for the identification of genes...

  9. Crime clocks and target performance maps

    CSIR Research Space (South Africa)

    Cooper, Antony K

    1999-12-01

    Full Text Available the period of analysis. Each segment of a pie chart represents a selected part of the day (eg: a two- or three-hour period) or a day of the week. The first and last segments in the day or week are then adjacent, ensuring that there is no artificial break... clocks We have also used crime clocks to map the proportion of crimes that occur during normal police working hours (07:00 to 16:00, Monday to Friday, in the case of the Johannesburg Area), against those that occur outside these hours. 3. Target...

  10. The Fermilab D0 Master Clock System

    International Nuclear Information System (INIS)

    Rotolo, C.; Fachin, M.; Chappa, S.; Rauch, M.; Needles, C.; Dyer, A.

    1991-11-01

    The Clock System provides bunch crossing related timing signals to various detector subsystems. Accelerator synchronization and monitoring as well as timing signal generation and distribution are discussed. The system is built using three module types implemented in Eurostandard hardware with a VME communications interface. The first two types of modules are used to facilitate synchronization with the accelerator and to generate 23 timing signals that are programmable with one RF bucket (18.8 ns) resolution and 1 ns accuracy. Fifty-four of the third module type are used to distribute the timing signals and two synchronous 53 MHz and 106 MHz clocks to various detector subsystems. 6 refs., 5 figs

  11. Clocking Scheme for Switched-Capacitor Circuits

    DEFF Research Database (Denmark)

    Steensgaard-Madsen, Jesper

    1998-01-01

    A novel clocking scheme for switched-capacitor (SC) circuits is presented. It can enhance the understanding of SC circuits and the errors caused by MOSFET (MOS) switches. Charge errors, and techniques to make SC circuits less sensitive to them are discussed.......A novel clocking scheme for switched-capacitor (SC) circuits is presented. It can enhance the understanding of SC circuits and the errors caused by MOSFET (MOS) switches. Charge errors, and techniques to make SC circuits less sensitive to them are discussed....

  12. A high-quality catalog of the Drosophila melanogaster proteome

    DEFF Research Database (Denmark)

    Brunner, Erich; Ahrens, Christian H.; Mohanty, Sonaly

    2007-01-01

    % of the predicted Drosophila melanogaster proteome by detecting 9,124 proteins from 498,000 redundant and 72,281 distinct peptide identifications. This unprecedented high proteome coverage for a complex eukaryote was achieved by combining sample diversity, multidimensional biochemical fractionation and analysis...

  13. Heat shock protection against cold stress of Drosophila melanogaster

    OpenAIRE

    Burton, Vicky; Mitchell, Herschel K.; Young, Patricia; Petersen, Nancy S.

    1988-01-01

    Heat shock protein synthesis can be induced during recovery from cold treatment of Drosophila melanogaster larvae. Survival of larvae after a cold treatment is dramatically improved by a mild heat shock just before the cold shock. The conditions which induce tolerance to cold are similar to those which confer tolerance to heat.

  14. Circadian rhythmicity of active GSK3 isoforms modulates molecular clock gene rhythms in the suprachiasmatic nucleus.

    Science.gov (United States)

    Besing, Rachel C; Paul, Jodi R; Hablitz, Lauren M; Rogers, Courtney O; Johnson, Russell L; Young, Martin E; Gamble, Karen L

    2015-04-01

    The suprachiasmatic nucleus (SCN) drives and synchronizes daily rhythms at the cellular level via transcriptional-translational feedback loops comprising clock genes such as Bmal1 and Period (Per). Glycogen synthase kinase 3 (GSK3), a serine/threonine kinase, phosphorylates at least 5 core clock proteins and shows diurnal variation in phosphorylation state (inactivation) of the GSK3β isoform. Whether phosphorylation of the other primary isoform (GSK3α) varies across the subjective day-night cycle is unknown. The purpose of this study was to determine if the endogenous rhythm of GSK3 (α and β) phosphorylation is critical for rhythmic BMAL1 expression and normal amplitude and periodicity of the molecular clock in the SCN. Significant circadian rhythmicity of phosphorylated GSK3 (α and β) was observed in the SCN from wild-type mice housed in constant darkness for 2 weeks. Importantly, chronic activation of both GSK3 isoforms impaired rhythmicity of the GSK3 target BMAL1. Furthermore, chronic pharmacological inhibition of GSK3 with 20 µM CHIR-99021 enhanced the amplitude and shortened the period of PER2::luciferase rhythms in organotypic SCN slice cultures. These results support the model that GSK3 activity status is regulated by the circadian clock and that GSK3 feeds back to regulate the molecular clock amplitude in the SCN. © 2015 The Author(s).

  15. Crystal Structure of the CLOCK Transactivation Domain Exon19 in Complex with a Repressor

    Energy Technology Data Exchange (ETDEWEB)

    Hou, Zhiqiang; Su, Lijing; Pei, Jimin; Grishin, Nick V.; Zhang, Hong (UTSMC)

    2017-08-01

    In the canonical clock model, CLOCK:BMAL1-mediated transcriptional activation is feedback regulated by its repressors CRY and PER and, in association with other coregulators, ultimately generates oscillatory gene expression patterns. How CLOCK:BMAL1 interacts with coregulator(s) is not well understood. Here we report the crystal structures of the mouse CLOCK transactivating domain Exon19 in complex with CIPC, a potent circadian repressor that functions independently of CRY and PER. The Exon19:CIPC complex adopts a three-helical coiled-coil bundle conformation containing two Exon19 helices and one CIPC. Unique to Exon19:CIPC, three highly conserved polar residues, Asn341 of CIPC and Gln544 of the two Exon19 helices, are located at the mid-section of the coiled-coil bundle interior and form hydrogen bonds with each other. Combining results from protein database search, sequence analysis, and mutagenesis studies, we discovered for the first time that CLOCK Exon19:CIPC interaction is a conserved transcription regulatory mechanism among mammals, fish, flies, and other invertebrates.

  16. Centriole Remodeling during Spermiogenesis in Drosophila.

    Science.gov (United States)

    Khire, Atul; Jo, Kyoung H; Kong, Dong; Akhshi, Tara; Blachon, Stephanie; Cekic, Anthony R; Hynek, Sarah; Ha, Andrew; Loncarek, Jadranka; Mennella, Vito; Avidor-Reiss, Tomer

    2016-12-05

    The first cell of an animal (zygote) requires centrosomes that are assembled from paternally inherited centrioles and maternally inherited pericentriolar material (PCM) [1]. In some animals, sperm centrioles with typical ultrastructure are the origin of the first centrosomes in the zygote [2-4]. In other animals, however, sperm centrioles lose their proteins and are thought to be degenerated and non-functional during spermiogenesis [5, 6]. Here, we show that the two sperm centrioles (the giant centriole [GC] and the proximal centriole-like structure [PCL]) in Drosophila melanogaster are remodeled during spermiogenesis through protein enrichment and ultrastructure modification in parallel to previously described centrosomal reduction [7]. We found that the ultrastructure of the matured sperm (spermatozoa) centrioles is modified dramatically and that the PCL does not resemble a typical centriole. We also describe a new phenomenon of Poc1 enrichment of the atypical centrioles in the spermatozoa. Using various mutants, protein expression during spermiogenesis, and RNAi knockdown of paternal Poc1, we found that paternal Poc1 enrichment is essential for the formation of centrioles during spermiogenesis and for the formation of centrosomes after fertilization in the zygote. Altogether, these findings demonstrate that the sperm centrioles are remodeled both in their protein composition and in ultrastructure, yet they are functional and are essential for normal embryogenesis in Drosophila. Copyright © 2016 Elsevier Ltd. All rights reserved.

  17. Or47b receptor neurons mediate sociosexual interactions in the fruit fly Drosophila melanogaster.

    Science.gov (United States)

    Lone, Shahnaz Rahman; Sharma, Vijay Kumar

    2012-04-01

    In the fruit fly Drosophila melanogaster, social interactions especially among heterosexual couples have been shown to have significant impact on the circadian timing system. Olfaction plays a major role in such interactions; however, we do not know yet specifically which receptor(s) are involved. Further, the role of circadian clock neurons in the rhythmic regulation of such sociosexual interactions (SSIs) is not fully understood. Here, we report the results of our study in which we assayed the locomotor activity and sleep-wake behaviors of male-male (MM), female-female (FF), and male-female (MF) couples from several wild-type and mutant strains of Drosophila with an aim to identify specific olfactory receptor(s) and circadian clock neurons involved in the rhythmic regulation of SSI. The results indicate that Or47b receptor neurons are necessary for SSI, as ablation or silencing of these neurons has a severe impact on SSI. Further, the neuropeptide pigment dispersing factor (PDF) and PDF-positive ventral lateral (LN(v)) clock neurons appear to be dispensable for the regulation of SSI; however, dorsal neurons may be involved.

  18. Effect of Spaceflight on the Circadian Rhythm, Lifespan and Gene Expression of Drosophila melanogaster

    Science.gov (United States)

    Xu, Kanyan

    2015-01-01

    Space travelers are reported to experience circadian rhythm disruption during spaceflight. However, how the space environment affects circadian rhythm is yet to be determined. The major focus of this study was to investigate the effect of spaceflight on the Drosophila circadian clock at both the behavioral and molecular level. We used China’s Shenzhou-9 spaceship to carry Drosophila. After 13 days of spaceflight, behavior tests showed that the flies maintained normal locomotor activity rhythm and sleep pattern. The expression level and rhythm of major clock genes were also unaffected. However, expression profiling showed differentially regulated output genes of the circadian clock system between space flown and control flies, suggesting that spaceflight affected the circadian output pathway. We also investigated other physiological effects of spaceflight such as lipid metabolism and lifespan, and searched genes significantly affected by spaceflight using microarray analysis. These results provide new information on the effects of spaceflight on circadian rhythm, lipid metabolism and lifespan. Furthermore, we showed that studying the effect of spaceflight on gene expression using samples collected at different Zeitgeber time could obtain different results, suggesting the importance of appropriate sampling procedures in studies on the effects of spaceflight. PMID:25798821

  19. Effect of spaceflight on the circadian rhythm, lifespan and gene expression of Drosophila melanogaster.

    Directory of Open Access Journals (Sweden)

    Lingling Ma

    Full Text Available Space travelers are reported to experience circadian rhythm disruption during spaceflight. However, how the space environment affects circadian rhythm is yet to be determined. The major focus of this study was to investigate the effect of spaceflight on the Drosophila circadian clock at both the behavioral and molecular level. We used China's Shenzhou-9 spaceship to carry Drosophila. After 13 days of spaceflight, behavior tests showed that the flies maintained normal locomotor activity rhythm and sleep pattern. The expression level and rhythm of major clock genes were also unaffected. However, expression profiling showed differentially regulated output genes of the circadian clock system between space flown and control flies, suggesting that spaceflight affected the circadian output pathway. We also investigated other physiological effects of spaceflight such as lipid metabolism and lifespan, and searched genes significantly affected by spaceflight using microarray analysis. These results provide new information on the effects of spaceflight on circadian rhythm, lipid metabolism and lifespan. Furthermore, we showed that studying the effect of spaceflight on gene expression using samples collected at different Zeitgeber time could obtain different results, suggesting the importance of appropriate sampling procedures in studies on the effects of spaceflight.

  20. Circadian Clocks: Unexpected Biochemical Cogs.

    Science.gov (United States)

    Mori, Tetsuya; Mchaourab, Hassane; Johnson, Carl Hirschie

    2015-10-05

    A circadian oscillation can be reconstituted in vitro from three proteins that cycles with a period of ∼ 24 h. Two recent studies provide surprising biochemical answers to why this remarkable oscillator has such a long time constant and how it can switch effortlessly between alternating enzymatic modes. Copyright © 2015 Elsevier Ltd. All rights reserved.

  1. Circadian Clocks: Unexpected Biochemical Cogs

    OpenAIRE

    Mori, Tetsuya; Mchaourab, Hassane; Johnson, Carl Hirschie

    2015-01-01

    A circadian oscillation can be reconstituted in vitro from three proteins that cycles with a period of ~24 h. Two recent studies provide surprising biochemical answers to why this remarkable oscillator has such a long time constant and how it can switch effortlessly between alternating enzymatic modes.

  2. A review of neurohormone GPCRs present in the fruitfly Drosophila melanogaster and the honey bee Apis mellifera

    DEFF Research Database (Denmark)

    Hauser, Frank; Cazzamali, Giuseppe; Williamson, Michael

    2006-01-01

    in the recently sequenced genome from the honey bee Apis mellifera. We found 35 neuropeptide receptor genes in the honey bee (44 in Drosophila) and two genes, coding for leucine-rich repeats-containing protein hormone GPCRs (4 in Drosophila). In addition, the honey bee has 19 biogenic amine receptor genes (21...

  3. Behavioral Teratogenesis in Drosophila melanogaster.

    Science.gov (United States)

    Mishra, Monalisa; Barik, Bedanta Kumar

    2018-01-01

    Developmental biology is a fascinating branch of science which helps us to understand the mechanism of development, thus the findings are used in various therapeutic approach. Drosophila melanogaster served as a model to find the key molecules that initiate and regulate the mechanism of development. Various genes, transcription factors, and signaling pathways helping in development are identified in Drosophila. Many toxic compounds, which can affect the development, are also recognized using Drosophila model. These compounds, which can affect the development, are named as a teratogen. Many teratogens identified using Drosophila may also act as a teratogen for a human being since 75% of conservation exist between the disease genes present in Drosophila and human. There are certain teratogens, which do not cause developmental defect if exposed during pregnancy, however; behavioral defect appears in later part of development. Such compounds are named as a behavioral teratogen. Thus, it is worthy to identify the potential behavioral teratogen using Drosophila model. Drosophila behavior is well studied in various developmental stages. This chapter describes various methods which can be employed to test behavioral teratogenesis in Drosophila.

  4. Novel transcriptional networks regulated by CLOCK in human neurons.

    Science.gov (United States)

    Fontenot, Miles R; Berto, Stefano; Liu, Yuxiang; Werthmann, Gordon; Douglas, Connor; Usui, Noriyoshi; Gleason, Kelly; Tamminga, Carol A; Takahashi, Joseph S; Konopka, Genevieve

    2017-11-01

    The molecular mechanisms underlying human brain evolution are not fully understood; however, previous work suggested that expression of the transcription factor CLOCK in the human cortex might be relevant to human cognition and disease. In this study, we investigated this novel transcriptional role for CLOCK in human neurons by performing chromatin immunoprecipitation sequencing for endogenous CLOCK in adult neocortices and RNA sequencing following CLOCK knockdown in differentiated human neurons in vitro. These data suggested that CLOCK regulates the expression of genes involved in neuronal migration, and a functional assay showed that CLOCK knockdown increased neuronal migratory distance. Furthermore, dysregulation of CLOCK disrupts coexpressed networks of genes implicated in neuropsychiatric disorders, and the expression of these networks is driven by hub genes with human-specific patterns of expression. These data support a role for CLOCK-regulated transcriptional cascades involved in human brain evolution and function. © 2017 Fontenot et al.; Published by Cold Spring Harbor Laboratory Press.

  5. Susceptibility of Redundant Versus Singular Clock Domains Implemented in SRAM-Based FPGA TMR Designs

    Science.gov (United States)

    Berg, Melanie D.; LaBel, Kenneth A.; Pellish, Jonathan

    2016-01-01

    We present the challenges that arise when using redundant clock domains due to their clock-skew. Radiation data show that a singular clock domain (DTMR) provides an improved TMR methodology for SRAM-based FPGAs over redundant clocks.

  6. The Drosophila melanogaster host model

    Science.gov (United States)

    Igboin, Christina O.; Griffen, Ann L.; Leys, Eugene J.

    2012-01-01

    The deleterious and sometimes fatal outcomes of bacterial infectious diseases are the net result of the interactions between the pathogen and the host, and the genetically tractable fruit fly, Drosophila melanogaster, has emerged as a valuable tool for modeling the pathogen–host interactions of a wide variety of bacteria. These studies have revealed that there is a remarkable conservation of bacterial pathogenesis and host defence mechanisms between higher host organisms and Drosophila. This review presents an in-depth discussion of the Drosophila immune response, the Drosophila killing model, and the use of the model to examine bacterial–host interactions. The recent introduction of the Drosophila model into the oral microbiology field is discussed, specifically the use of the model to examine Porphyromonas gingivalis–host interactions, and finally the potential uses of this powerful model system to further elucidate oral bacterial-host interactions are addressed. PMID:22368770

  7. The Drosophila melanogaster host model

    Directory of Open Access Journals (Sweden)

    Christina O. Igboin

    2012-02-01

    Full Text Available The deleterious and sometimes fatal outcomes of bacterial infectious diseases are the net result of the interactions between the pathogen and the host, and the genetically tractable fruit fly, Drosophila melanogaster, has emerged as a valuable tool for modeling the pathogen–host interactions of a wide variety of bacteria. These studies have revealed that there is a remarkable conservation of bacterial pathogenesis and host defence mechanisms between higher host organisms and Drosophila. This review presents an in-depth discussion of the Drosophila immune response, the Drosophila killing model, and the use of the model to examine bacterial–host interactions. The recent introduction of the Drosophila model into the oral microbiology field is discussed, specifically the use of the model to examine Porphyromonas gingivalis–host interactions, and finally the potential uses of this powerful model system to further elucidate oral bacterial-host interactions are addressed.

  8. The Drosophila melanogaster host model.

    Science.gov (United States)

    Igboin, Christina O; Griffen, Ann L; Leys, Eugene J

    2012-01-01

    The deleterious and sometimes fatal outcomes of bacterial infectious diseases are the net result of the interactions between the pathogen and the host, and the genetically tractable fruit fly, Drosophila melanogaster, has emerged as a valuable tool for modeling the pathogen-host interactions of a wide variety of bacteria. These studies have revealed that there is a remarkable conservation of bacterial pathogenesis and host defence mechanisms between higher host organisms and Drosophila. This review presents an in-depth discussion of the Drosophila immune response, the Drosophila killing model, and the use of the model to examine bacterial-host interactions. The recent introduction of the Drosophila model into the oral microbiology field is discussed, specifically the use of the model to examine Porphyromonas gingivalis-host interactions, and finally the potential uses of this powerful model system to further elucidate oral bacterial-host interactions are addressed.

  9. RNA editing in Drosophila melanogaster: new targets and functionalconsequences

    Energy Technology Data Exchange (ETDEWEB)

    Stapleton, Mark; Carlson, Joseph W.; Celniker, Susan E.

    2006-09-05

    Adenosine deaminases that act on RNA (ADARs) catalyze the site-specific conversion of adenosine to inosine in primary mRNA transcripts. These re-coding events affect coding potential, splice-sites, and stability of mature mRNAs. ADAR is an essential gene and studies in mouse, C. elegans, and Drosophila suggest its primary function is to modify adult behavior by altering signaling components in the nervous system. By comparing the sequence of isogenic cDNAs to genomic DNA, we have identified and experimentally verified 27 new targets of Drosophila ADAR. Our analyses lead us to identify new classes of genes whose transcripts are targets of ADAR including components of the actin cytoskeleton, and genes involved in ion homeostasis and signal transduction. Our results indicate that editing in Drosophila increases the diversity of the proteome, and does so in a manner that has direct functional consequences on protein function.

  10. big bang gene modulates gut immune tolerance in Drosophila.

    Science.gov (United States)

    Bonnay, François; Cohen-Berros, Eva; Hoffmann, Martine; Kim, Sabrina Y; Boulianne, Gabrielle L; Hoffmann, Jules A; Matt, Nicolas; Reichhart, Jean-Marc

    2013-02-19

    Chronic inflammation of the intestine is detrimental to mammals. Similarly, constant activation of the immune response in the gut by the endogenous flora is suspected to be harmful to Drosophila. Therefore, the innate immune response in the gut of Drosophila melanogaster is tightly balanced to simultaneously prevent infections by pathogenic microorganisms and tolerate the endogenous flora. Here we describe the role of the big bang (bbg) gene, encoding multiple membrane-associated PDZ (PSD-95, Discs-large, ZO-1) domain-containing protein isoforms, in the modulation of the gut immune response. We show that in the adult Drosophila midgut, BBG is present at the level of the septate junctions, on the apical side of the enterocytes. In the absence of BBG, these junctions become loose, enabling the intestinal flora to trigger a constitutive activation of the anterior midgut immune response. This chronic epithelial inflammation leads to a reduced lifespan of bbg mutant flies. Clearing the commensal flora by antibiotics prevents the abnormal activation of the gut immune response and restores a normal lifespan. We now provide genetic evidence that Drosophila septate junctions are part of the gut immune barrier, a function that is evolutionarily conserved in mammals. Collectively, our data suggest that septate junctions are required to maintain the subtle balance between immune tolerance and immune response in the Drosophila gut, which represents a powerful model to study inflammatory bowel diseases.

  11. Clock Synchronization for Multihop Wireless Sensor Networks

    Science.gov (United States)

    Solis Robles, Roberto

    2009-01-01

    In wireless sensor networks, more so generally than in other types of distributed systems, clock synchronization is crucial since by having this service available, several applications such as media access protocols, object tracking, or data fusion, would improve their performance. In this dissertation, we propose a set of algorithms to achieve…

  12. Food at work around the clock

    DEFF Research Database (Denmark)

    Dahl Lassen, Anne; Beck, Anne Marie; Thorsen, Anne Vibeke

    This report brings together 12 invited presentations and outcomes of a workshop on food and meals for employees working irregular hours “around the clock”. The workshop, “Food at work around the clock – The Nordic Model”, was hosted by the National Food Institute at the Technical University...

  13. Hands Together! An Analog Clock Problem

    Science.gov (United States)

    Earnest, Darrell; Radtke, Susan; Scott, Siri

    2017-01-01

    In this article, the authors first present the Hands Together! task. The mathematics in this problem concerns the relationship of hour and minute durations as reflected in the oft-overlooked proportional movements of the two hands of an analog clock. The authors go on to discuss the importance of problem solving in general. They then consider…

  14. The mammalian retina as a clock

    Science.gov (United States)

    Tosini, Gianluca; Fukuhara, Chiaki

    2002-01-01

    Many physiological, cellular, and biochemical parameters in the retina of vertebrates show daily rhythms that, in many cases, also persist under constant conditions. This demonstrates that they are driven by a circadian pacemaker. The presence of an autonomous circadian clock in the retina of vertebrates was first demonstrated in Xenopus laevis and then, several years later, in mammals. In X. laevis and in chicken, the retinal circadian pacemaker has been localized in the photoreceptor layer, whereas in mammals, such information is not yet available. Recent advances in molecular techniques have led to the identification of a group of genes that are believed to constitute the molecular core of the circadian clock. These genes are expressed in the retina, although with a slightly different 24-h profile from that observed in the central circadian pacemaker. This result suggests that some difference (at the molecular level) may exist between the retinal clock and the clock located in the suprachiasmatic nuclei of hypothalamus. The present review will focus on the current knowledge of the retinal rhythmicity and the mechanisms responsible for its control.

  15. Analytic clock frequency selection for global DVFS

    NARCIS (Netherlands)

    Gerards, Marco Egbertus Theodorus; Hurink, Johann L.; Holzenspies, P.K.F.; Kuper, Jan; Smit, Gerardus Johannes Maria

    2014-01-01

    Computers can reduce their power consumption by decreasing their speed using Dynamic Voltage and Frequency Scaling (DVFS). A form of DVFS for multicore processors is global DVFS, where the voltage and clock frequency is shared among all processor cores. Because global DVFS is efficient and cheap to

  16. Intergeneric complementation of a circadian rhythmicity defect : phylogenetic conservation of structure and function of the clock gene frequency

    NARCIS (Netherlands)

    Merrow, Martha W.; Dunlap, Jay C.; Dover, G.

    1994-01-01

    The Neurospora crassa frequency locus encodes a 989 amino acid protein that is a central component, a state variable, of the circadian biological clock. We have determined the sequence of all or part of this protein and surrounding regulatory regions from additional fungi representing three genera

  17. CNTNAP2 and NRXN1 are mutated in autosomal-recessive Pitt-Hopkins-like mental retardation and determine the level of a common synaptic protein in Drosophila

    DEFF Research Database (Denmark)

    Zweier, Christiane; de Jong, Eiko K; Zweier, Markus

    2009-01-01

    , phenotypically overlapping with Pitt-Hopkins syndrome. With a frequency of at least 1% in our cohort of 179 patients, recessive defects in CNTNAP2 appear to significantly contribute to severe MR. Whereas the established synaptic role of NRXN1 suggests that synaptic defects contribute to the associated...... protein can reorganize synaptic morphology and induce increased density of active zones, the synaptic domains of neurotransmitter release. Moreover, both Nrx-I and Nrx-IV determine the level of the presynaptic active-zone protein bruchpilot, indicating a possible common molecular mechanism in Nrx...

  18. "In vivo" toxicity of a truncated version of the Drosophila Rst-IrreC protein is dependent on the presence of a glutamine-rich region in its intracellular domain

    Directory of Open Access Journals (Sweden)

    RICARDO C. MACHADO

    2002-06-01

    Full Text Available The roughest-irregular chiasm C ( rst-irreC gene of Drosophila melanogaster encodes a transmembrane glycoprotein containing five immunoglobulin-like domains in its extracellular portion and an intracytoplasmic tail rich in serine and threonine as well some conserved motifs suggesting signal transduction activity. In the compound eye, loss-of-function rst-irreC mutants lack the characteristic wave of programmed cell death happening in early pupa and which is essential for the elimination of the surplus interommatidial cells. Here we report an investigation on the role played by the Rst-irreC molecule in triggering programmed cell death. "In vivo" transient expression assays showed that deletion of the last 80 amino acids of the carboxyl terminus produces a form of the protein that is highly toxic to larvae. This toxicity is suppressed if an additional 47 amino acid long, glutamine-rich region ("opa-like domain", is also removed from the protein. The results suggest the possibility that the opa-like domain and the carboxyl terminus act in concert to modulate rst-irreC function in apoptosis, and we discuss this implication in the context of the general mechanisms causing glutamine-rich neurodegenerative diseases in humans.O gene roughest-irregular chiasm C ( rst-irreC de Drosophila melanogaster, codifica uma glicoproteína transmembranar contendo cinco domínios semelhantes a imunoglobulina em sua porção extracelular e uma cauda intracitoplasmática rica em serina e treonina, assim como alguns sequências conservadas que sugerem atividade transdutora de sinais. No olho composto, mutantes rst-irreC de perda de função não apresentam uma característica "onda" de morte celular programada que ocorre no início do período pupal e que é essencial para a eliminação de células interomatidiais em excesso. Aqui descrevemos uma investigação sobre o papel desempenhado pela molécula Rst-IrreC no disparo da morte celular programada. Ensaios de

  19. A role for clock genes in sleep homeostasis.

    Science.gov (United States)

    Franken, Paul

    2013-10-01

    The timing and quality of both sleep and wakefulness are thought to be regulated by the interaction of two processes. One of these two processes keeps track of the prior sleep-wake history and controls the homeostatic need for sleep while the other sets the time-of-day that sleep preferably occurs. The molecular pathways underlying the latter, circadian process have been studied in detail and their key role in physiological time-keeping has been well established. Analyses of sleep in mice and flies lacking core circadian clock gene proteins have demonstrated, however, that besides disrupting circadian rhythms, also sleep homeostatic processes were affected. Subsequent studies revealed that sleep loss alters both the mRNA levels and the specific DNA-binding of the key circadian transcriptional regulators to their target sequences in the mouse brain. The fact that sleep loss impinges on the very core of the molecular circadian circuitry might explain why both inadequate sleep and disrupted circadian rhythms can similarly lead to metabolic pathology. The evidence for a role for clock genes in sleep homeostasis will be reviewed here. Copyright © 2013 Elsevier Ltd. All rights reserved.

  20. The developmental transcriptome of Drosophila melanogaster

    Energy Technology Data Exchange (ETDEWEB)

    University of Connecticut; Graveley, Brenton R.; Brooks, Angela N.; Carlson, Joseph W.; Duff, Michael O.; Landolin, Jane M.; Yang, Li; Artieri, Carlo G.; van Baren, Marijke J.; Boley, Nathan; Booth, Benjamin W.; Brown, James B.; Cherbas, Lucy; Davis, Carrie A.; Dobin, Alex; Li, Renhua; Lin, Wei; Malone, John H.; Mattiuzzo, Nicolas R.; Miller, David; Sturgill, David; Tuch, Brian B.; Zaleski, Chris; Zhang, Dayu; Blanchette, Marco; Dudoit, Sandrine; Eads, Brian; Green, Richard E.; Hammonds, Ann; Jiang, Lichun; Kapranov, Phil; Langton, Laura; Perrimon, Norbert; Sandler, Jeremy E.; Wan, Kenneth H.; Willingham, Aarron; Zhang, Yu; Zou, Yi; Andrews, Justen; Bicke, Peter J.; Brenner, Steven E.; Brent, Michael R.; Cherbas, Peter; Gingeras, Thomas R.; Hoskins, Roger A.; Kaufman, Thomas C.; Oliver, Brian; Celniker, Susan E.

    2010-12-02

    Drosophila melanogaster is one of the most well studied genetic model organisms; nonetheless, its genome still contains unannotated coding and non-coding genes, transcripts, exons and RNA editing sites. Full discovery and annotation are pre-requisites for understanding how the regulation of transcription, splicing and RNA editing directs the development of this complex organism. Here we used RNA-Seq, tiling microarrays and cDNA sequencing to explore the transcriptome in 30 distinct developmental stages. We identified 111,195 new elements, including thousands of genes, coding and non-coding transcripts, exons, splicing and editing events, and inferred protein isoforms that previously eluded discovery using established experimental, prediction and conservation-based approaches. These data substantially expand the number of known transcribed elements in the Drosophila genome and provide a high-resolution view of transcriptome dynamics throughout development. Drosophila melanogaster is an important non-mammalian model system that has had a critical role in basic biological discoveries, such as identifying chromosomes as the carriers of genetic information and uncovering the role of genes in development. Because it shares a substantial genic content with humans, Drosophila is increasingly used as a translational model for human development, homeostasis and disease. High-quality maps are needed for all functional genomic elements. Previous studies demonstrated that a rich collection of genes is deployed during the life cycle of the fly. Although expression profiling using microarrays has revealed the expression of, 13,000 annotated genes, it is difficult to map splice junctions and individual base modifications generated by RNA editing using such approaches. Single-base resolution is essential to define precisely the elements that comprise the Drosophila transcriptome. Estimates of the number of transcript isoforms are less accurate than estimates of the number of genes

  1. Epigenetic telomere protection by Drosophila DNA damage response pathways.

    Science.gov (United States)

    Oikemus, Sarah R; Queiroz-Machado, Joana; Lai, KuanJu; McGinnis, Nadine; Sunkel, Claudio; Brodsky, Michael H

    2006-05-01

    Analysis of terminal deletion chromosomes indicates that a sequence-independent mechanism regulates protection of Drosophila telomeres. Mutations in Drosophila DNA damage response genes such as atm/tefu, mre11, or rad50 disrupt telomere protection and localization of the telomere-associated proteins HP1 and HOAP, suggesting that recognition of chromosome ends contributes to telomere protection. However, the partial telomere protection phenotype of these mutations limits the ability to test if they act in the epigenetic telomere protection mechanism. We examined the roles of the Drosophila atm and atr-atrip DNA damage response pathways and the nbs homolog in DNA damage responses and telomere protection. As in other organisms, the atm and atr-atrip pathways act in parallel to promote telomere protection. Cells lacking both pathways exhibit severe defects in telomere protection and fail to localize the protection protein HOAP to telomeres. Drosophila nbs is required for both atm- and atr-dependent DNA damage responses and acts in these pathways during DNA repair. The telomere fusion phenotype of nbs is consistent with defects in each of these activities. Cells defective in both the atm and atr pathways were used to examine if DNA damage response pathways regulate telomere protection without affecting telomere specific sequences. In these cells, chromosome fusion sites retain telomere-specific sequences, demonstrating that loss of these sequences is not responsible for loss of protection. Furthermore, terminally deleted chromosomes also fuse in these cells, directly implicating DNA damage response pathways in the epigenetic protection of telomeres. We propose that recognition of chromosome ends and recruitment of HP1 and HOAP by DNA damage response proteins is essential for the epigenetic protection of Drosophila telomeres. Given the conserved roles of DNA damage response proteins in telomere function, related mechanisms may act at the telomeres of other organisms.

  2. Mini Screening of Kinase Inhibitors Affecting Period-length of Mammalian Cellular Circadian Clock

    International Nuclear Information System (INIS)

    Yagita, Kazuhiro; Yamanaka, Iori; Koinuma, Satoshi; Shigeyoshi, Yasufumi; Uchiyama, Yasuo

    2009-01-01

    In mammalian circadian rhythms, the transcriptional-translational feedback loop (TTFL) consisting of a set of clock genes is believed to elicit the circadian clock oscillation. The TTFL model explains that the accumulation and degradation of mPER and mCRY proteins control the period-length (tau) of the circadian clock. Although recent studies revealed that the Casein Kinase Iεδ (CKIεδ) regurates the phosphorylation of mPER proteins and the circadian period-length, other kinases are also likely to contribute the phosphorylation of mPER. Here, we performed small scale screening using 84 chemical compounds known as kinase inhibitors to identify candidates possibly affecting the circadian period-length in mammalian cells. Screening by this high-throughput real-time bioluminescence monitoring system revealed that the several chemical compounds apparently lengthened the cellular circadian clock oscillation. These compounds are known as inhibitors against kinases such as Casein Kinase II (CKII), PI3-kinase (PI3K) and c-Jun N-terminal Kinase (JNK) in addition to CKIεδ. Although these kinase inhibitors may have some non-specific effects on other factors, our mini screening identified new candidates contributing to period-length control in mammalian cells

  3. Gypsy transposition correlates with the production of a retroviral envelope-like protein under the tissue-specific control of the Drosophila flamenco gene.

    OpenAIRE

    Pélisson, A; Song, S U; Prud'homme, N; Smith, P A; Bucheton, A; Corces, V G

    1994-01-01

    Gypsy displays striking similarities to vertebrate retroviruses, including the presence of a yet uncharacterized additional open reading frame (ORF3) and the recent evidence for infectivity. It is mobilized with high frequency in the germline of the progeny of females homozygous for the flamenco permissive mutation. We report the characterization of a gypsy subgenomic ORF3 RNA encoding typical retroviral envelope proteins. In females, env expression is strongly repressed by one copy of the no...

  4. [Ulysses retrotransposon aspartate proteinase (Drosophila virilis)].

    Science.gov (United States)

    Volkov, D A; Savvateeva, L V; Dergousova, N I; Rumsh, L D

    2002-01-01

    Retrotransposones are mobile genetic elements occurring in genomes of bacteria, plants or animals. Retrotransposones were found to contain nucleotide sequences encoding proteins which are homological to retroviral aspartic proteinases. Our research has been focused on Ulysses which is mobile genetic element found in Drosophila virilis. We suggested a primary structure of Ulysses proteinase using comparative analysis of amino acid sequences of retroviral proteinases and proteinases from retrotransposones. The appropriate cDNA fragment has been cloned and expressed in E. coli. The purification of recombinant protein (12 kD) has been carried out by affinity chromatography using pepstatine-agarose. The obtained protein has proteolytic activity at optimum pH 5.5 like the majority of aspartic proteinases.

  5. Crystallization of Spätzle, a cystine-knot protein involved in embryonic development and innate immunity in Drosophila melanogaster

    Energy Technology Data Exchange (ETDEWEB)

    Hoffmann, Anita; Neumann, Piotr [Institut für Biochemie und Biotechnologie, Martin-Luther-Universität Halle-Wittenberg, Abteilung Physikalische Biotechnologie, Kurt-Mothes-Strasse 3, 06120 Halle (Saale) (Germany); Schierhorn, Angelika [Max-Planck-Institut für Proteinfaltung, Abteilung Massenspektrometrie, Kurt-Mothes-Strasse 3, 06120 Halle (Saale) (Germany); Stubbs, Milton T., E-mail: stubbs@biochemtech.uni-halle.de [Institut für Biochemie und Biotechnologie, Martin-Luther-Universität Halle-Wittenberg, Abteilung Physikalische Biotechnologie, Kurt-Mothes-Strasse 3, 06120 Halle (Saale) (Germany); Mitteldeutsches Zentrum für Struktur und Dynamik der Proteine (MZP), Martin-Luther-Universität Halle-Wittenberg (Germany)

    2008-08-01

    Crystallization of the cystine-knot protein Spätzle occurred following serendipitous limited degradation of the pro-Spätzle propeptide during the crystallization experiment. The Spätzle protein is involved in both the definition of the dorsal–ventral axis during embryonic development and in the adult innate immune response. The disulfide-linked dimeric cystine-knot protein has been expressed as a proprotein in inclusion bodies in Escherichia coli and refolded in vitro by rapid dilution. Initial orthorhombic crystals that diffracted to 7 Å resolution were obtained after three months by the sitting-drop vapour-diffusion method. Optimization of the crystallization conditions resulted in orthorhombic crystals (space group P2{sub 1}2{sub 1}2{sub 1}, with unit-cell parameters a = 53.0, b = 59.2, c = 62.5 Å) that diffracted to 2.8 Å resolution in-house. The small volume of the asymmetric unit indicated that it was not possible for the crystals to contain the complete pro-Spätzle dimer. Mass spectrometry, N-terminal sequencing and Western-blot analysis revealed that the crystals contained the C-terminal disulfide-linked cystine-knot dimer. Comparison of various crystallization experiments indicated that degradation of the N-terminal prodomain was dependent on the buffer conditions.

  6. Drosophila's contribution to stem cell research [v1; ref status: indexed, http://f1000r.es/5h7

    Directory of Open Access Journals (Sweden)

    Gyanesh Singh

    2015-06-01

    Full Text Available The discovery of Drosophila stem cells with striking similarities to mammalian stem cells has brought new hope for stem cell research. A recent development in Drosophila stem cell research is bringing wider opportunities for contemporary stem cell biologists. In this regard, Drosophila germ cells are becoming a popular model of stem cell research. In several cases, genes that controlled Drosophila stem cells were later discovered to have functional homologs in mammalian stem cells. Like mammals, Drosophila germline stem cells (GSCs are controlled by both intrinsic as well as external signals. Inside the Drosophila testes, germline and somatic stem cells form a cluster of cells (the hub. Hub cells depend on JAK-STAT signaling, and, in absence of this signal, they do not self-renew. In Drosophila, significant changes occur within the stem cell niche that contributes to a decline in stem cell number over time. In case of aging Drosophila, somatic niche cells show reduced DE-cadherin and unpaired (Upd proteins. Unpaired proteins are known to directly decrease stem cell number within the niches, and, overexpression of upd within niche cells restored GSCs in older males also . Stem cells in the midgut of Drosophila are also very promising. Reduced Notch signaling was found to increase the number of midgut progenitor cells. On the other hand, activation of the Notch pathway decreased proliferation of these cells. Further research in this area should lead to the discovery of additional factors that regulate stem and progenitor cells in Drosophila.

  7. Olfactory memory traces in Drosophila.

    Science.gov (United States)

    Berry, Jacob; Krause, William C; Davis, Ronald L

    2008-01-01

    In Drosophila, the fruit fly, coincident exposure to an odor and an aversive electric shock can produce robust behavioral memory. This behavioral memory is thought to be regulated by cellular memory traces within the central nervous system of the fly. These molecular, physiological, or structural changes in neurons, induced by pairing odor and shock, regulate behavior by altering the neurons' response to the learned environment. Recently, novel in vivo functional imaging techniques have allowed researchers to observe cellular memory traces in intact animals. These investigations have revealed interesting temporal and spatial dynamics of cellular memory traces. First, a short-term cellular memory trace was discovered that exists in the antennal lobe, an early site of olfactory processing. This trace represents the recruitment of new synaptic activity into the odor representation and forms for only a short period of time just after training. Second, an intermediate-term cellular memory trace was found in the dorsal paired medial neuron, a neuron thought to play a role in stabilizing olfactory memories. Finally, a long-term protein synthesis-dependent cellular memory trace was discovered in the mushroom bodies, a structure long implicated in olfactory learning and memory. Therefore, it appears that aversive olfactory associations are encoded by multiple cellular memory traces that occur in different regions of the brain with different temporal domains.

  8. Identification of genes associated with resilience/vulnerability to sleep deprivation and starvation in Drosophila.

    Science.gov (United States)

    Thimgan, Matthew S; Seugnet, Laurent; Turk, John; Shaw, Paul J

    2015-05-01

    Flies mutant for the canonical clock protein cycle (cyc(01)) exhibit a sleep rebound that is ∼10 times larger than wild-type flies and die after only 10 h of sleep deprivation. Surprisingly, when starved, cyc(01) mutants can remain awake for 28 h without demonstrating negative outcomes. Thus, we hypothesized that identifying transcripts that are differentially regulated between waking induced by sleep deprivation and waking induced by starvation would identify genes that underlie the deleterious effects of sleep deprivation and/or protect flies from the negative consequences of waking. We used partial complementary DNA microarrays to identify transcripts that are differentially expressed between cyc(01) mutants that had been sleep deprived or starved for 7 h. We then used genetics to determine whether disrupting genes involved in lipid metabolism would exhibit alterations in their response to sleep deprivation. Laboratory. Drosophila melanogaster. Sleep deprivation and starvation. We identified 84 genes with transcript levels that were differentially modulated by 7 h of sleep deprivation and starvation in cyc(01) mutants and were confirmed in independent samples using quantitative polymerase chain reaction. Several of these genes were predicted to be lipid metabolism genes, including bubblegum, cueball, and CG4500, which based on our data we have renamed heimdall (hll). Using lipidomics we confirmed that knockdown of hll using RNA interference significantly decreased lipid stores. Importantly, genetically modifying bubblegum, cueball, or hll resulted in sleep rebound alterations following sleep deprivation compared to genetic background controls. We have identified a set of genes that may confer resilience/vulnerability to sleep deprivation and demonstrate that genes involved in lipid metabolism modulate sleep homeostasis. © 2015 Associated Professional Sleep Societies, LLC.

  9. A mighty small heart: the cardiac proteome of adult Drosophila melanogaster.

    Directory of Open Access Journals (Sweden)

    Anthony Cammarato

    2011-04-01

    Full Text Available Drosophila melanogaster is emerging as a powerful model system for the study of cardiac disease. Establishing peptide and protein maps of the Drosophila heart is central to implementation of protein network studies that will allow us to assess the hallmarks of Drosophila heart pathogenesis and gauge the degree of conservation with human disease mechanisms on a systems level. Using a gel-LC-MS/MS approach, we identified 1228 protein clusters from 145 dissected adult fly hearts. Contractile, cytostructural and mitochondrial proteins were most abundant consistent with electron micrographs of the Drosophila cardiac tube. Functional/Ontological enrichment analysis further showed that proteins involved in glycolysis, Ca(2+-binding, redox, and G-protein signaling, among other processes, are also over-represented. Comparison with a mouse heart proteome revealed conservation at the level of molecular function, biological processes and cellular components. The subsisting peptidome encompassed 5169 distinct heart-associated peptides, of which 1293 (25% had not been identified in a recent Drosophila peptide compendium. PeptideClassifier analysis was further used to map peptides to specific gene-models. 1872 peptides provide valuable information about protein isoform groups whereas a further 3112 uniquely identify specific protein isoforms and may be used as a heart-associated peptide resource for quantitative proteomic approaches based on multiple-reaction monitoring. In summary, identification of excitation-contraction protein landmarks, orthologues of proteins associated with cardiovascular defects, and conservation of protein ontologies, provides testimony to the heart-like character of the Drosophila cardiac tube and to the utility of proteomics as a complement to the power of genetics in this growing model of human heart disease.

  10. A Statistically Representative Atlas for Mapping Neuronal Circuits in the Drosophila Adult Brain.

    Science.gov (United States)

    Arganda-Carreras, Ignacio; Manoliu, Tudor; Mazuras, Nicolas; Schulze, Florian; Iglesias, Juan E; Bühler, Katja; Jenett, Arnim; Rouyer, François; Andrey, Philippe

    2018-01-01

    Imaging the expression patterns of reporter constructs is a powerful tool to dissect the neuronal circuits of perception and behavior in the adult brain of Drosophila , one of the major models for studying brain functions. To date, several Drosophila brain templates and digital atlases have been built to automatically analyze and compare collections of expression pattern images. However, there has been no systematic comparison of performances between alternative atlasing strategies and registration algorithms. Here, we objectively evaluated the performance of different strategies for building adult Drosophila brain templates and atlases. In addition, we used state-of-the-art registration algorithms to generate a new group-wise inter-sex atlas. Our results highlight the benefit of statistical atlases over individual ones and show that the newly proposed inter-sex atlas outperformed existing solutions for automated registration and annotation of expression patterns. Over 3,000 images from the Janelia Farm FlyLight collection were registered using the proposed strategy. These registered expression patterns can be searched and compared with a new version of the BrainBaseWeb system and BrainGazer software. We illustrate the validity of our methodology and brain atlas with registration-based predictions of expression patterns in a subset of clock neurons. The described registration framework should benefit to brain studies in Drosophila and other insect species.

  11. A Statistically Representative Atlas for Mapping Neuronal Circuits in the Drosophila Adult Brain

    Directory of Open Access Journals (Sweden)

    Ignacio Arganda-Carreras

    2018-03-01

    Full Text Available Imaging the expression patterns of reporter constructs is a powerful tool to dissect the neuronal circuits of perception and behavior in the adult brain of Drosophila, one of the major models for studying brain functions. To date, several Drosophila brain templates and digital atlases have been built to automatically analyze and compare collections of expression pattern images. However, there has been no systematic comparison of performances between alternative atlasing strategies and registration algorithms. Here, we objectively evaluated the performance of different strategies for building adult Drosophila brain templates and atlases. In addition, we used state-of-the-art registration algorithms to generate a new group-wise inter-sex atlas. Our results highlight the benefit of statistical atlases over individual ones and show that the newly proposed inter-sex atlas outperformed existing solutions for automated registration and annotation of expression patterns. Over 3,000 images from the Janelia Farm FlyLight collection were registered using the proposed strategy. These registered expression patterns can be searched and compared with a new version of the BrainBaseWeb system and BrainGazer software. We illustrate the validity of our methodology and brain atlas with registration-based predictions of expression patterns in a subset of clock neurons. The described registration framework should benefit to brain studies in Drosophila and other insect species.

  12. A damped oscillator imposes temporal order on posterior gap gene expression in Drosophila

    Science.gov (United States)

    Verd, Berta; Clark, Erik; Wotton, Karl R.; Janssens, Hilde; Jiménez-Guri, Eva; Crombach, Anton

    2018-01-01

    Insects determine their body segments in two different ways. Short-germband insects, such as the flour beetle Tribolium castaneum, use a molecular clock to establish segments sequentially. In contrast, long-germband insects, such as the vinegar fly Drosophila melanogaster, determine all segments simultaneously through a hierarchical cascade of gene regulation. Gap genes constitute the first layer of the Drosophila segmentation gene hierarchy, downstream of maternal gradients such as that of Caudal (Cad). We use data-driven mathematical modelling and phase space analysis to show that shifting gap domains in the posterior half of the Drosophila embryo are an emergent property of a robust damped oscillator mechanism, suggesting that the regulatory dynamics underlying long- and short-germband segmentation are much more similar than previously thought. In Tribolium, Cad has been proposed to modulate the frequency of the segmentation oscillator. Surprisingly, our simulations and experiments show that the shift rate of posterior gap domains is independent of maternal Cad levels in Drosophila. Our results suggest a novel evolutionary scenario for the short- to long-germband transition and help explain why this transition occurred convergently multiple times during the radiation of the holometabolan insects. PMID:29451884

  13. Myoblast fusion in Drosophila

    Energy Technology Data Exchange (ETDEWEB)

    Haralalka, Shruti [Stowers Institute for Medical Research, Kansas City, MO 64110 (United States); Abmayr, Susan M., E-mail: sma@stowers.org [Stowers Institute for Medical Research, Kansas City, MO 64110 (United States); Department of Anatomy and Cell Biology, University of Kansas Medical Center, Kansas City, MO 66160 (United States)

    2010-11-01

    The body wall musculature of a Drosophila larva is composed of an intricate pattern of 30 segmentally repeated muscle fibers in each abdominal hemisegment. Each muscle fiber has unique spatial and behavioral characteristics that include its location, orientation, epidermal attachment, size and pattern of innervation. Many, if not all, of these properties are dictated by founder cells, which determine the muscle pattern and seed the fusion process. Myofibers are then derived from fusion between a specific founder cell and several fusion competent myoblasts (FCMs) fusing with as few as 3-5 FCMs in the small muscles on the most ventral side of the embryo and as many as 30 FCMs in the larger muscles on the dorsal side of the embryo. The focus of the present review is the formation of the larval muscles in the developing embryo, summarizing the major issues and players in this process. We have attempted to emphasize experimentally-validated details of the mechanism of myoblast fusion and distinguish these from the theoretically possible details that have not yet been confirmed experimentally. We also direct the interested reader to other recent reviews that discuss myoblast fusion in Drosophila, each with their own perspective on the process . With apologies, we use gene nomenclature as specified by Flybase (http://flybase.org) but provide Table 1 with alternative names and references.

  14. Myoblast fusion in Drosophila

    International Nuclear Information System (INIS)

    Haralalka, Shruti; Abmayr, Susan M.

    2010-01-01

    The body wall musculature of a Drosophila larva is composed of an intricate pattern of 30 segmentally repeated muscle fibers in each abdominal hemisegment. Each muscle fiber has unique spatial and behavioral characteristics that include its location, orientation, epidermal attachment, size and pattern of innervation. Many, if not all, of these properties are dictated by founder cells, which determine the muscle pattern and seed the fusion process. Myofibers are then derived from fusion between a specific founder cell and several fusion competent myoblasts (FCMs) fusing with as few as 3-5 FCMs in the small muscles on the most ventral side of the embryo and as many as 30 FCMs in the larger muscles on the dorsal side of the embryo. The focus of the present review is the formation of the larval muscles in the developing embryo, summarizing the major issues and players in this process. We have attempted to emphasize experimentally-validated details of the mechanism of myoblast fusion and distinguish these from the theoretically possible details that have not yet been confirmed experimentally. We also direct the interested reader to other recent reviews that discuss myoblast fusion in Drosophila, each with their own perspective on the process . With apologies, we use gene nomenclature as specified by Flybase (http://flybase.org) but provide Table 1 with alternative names and references.

  15. The Drosophila rolled locus encodes a MAP kinase required in the sevenless signal transduction pathway.

    OpenAIRE

    Biggs, W H; Zavitz, K H; Dickson, B; van der Straten, A; Brunner, D; Hafen, E; Zipursky, S L

    1994-01-01

    Mitogen-activated protein (MAP) kinases have been proposed to play a critical role in receptor tyrosine kinase (RTK)-mediated signal transduction pathways. Although genetic and biochemical studies of RTK pathways in Caenorhabditis elegans, Drosophila melanogaster and mammals have revealed remarkable similarities, a genetic requirement for MAP kinases in RTK signaling has not been established. During retinal development in Drosophila, the sevenless (Sev) RTK is required for development of the ...

  16. Does exercise training impact clock genes in patients with coronary artery disease and type 2 diabetes mellitus?

    Science.gov (United States)

    Steidle-Kloc, Eva; Schönfelder, Martin; Müller, Edith; Sixt, Sebastian; Schuler, Gerhard; Patsch, Wolfgang; Niebauer, Josef

    2016-09-01

    Recent findings revealed negative effects of deregulated molecular circadian rhythm in coronary artery disease (CAD) and type 2 diabetes mellitus (T2DM). Physical exercise training (ET) has been shown to promote anti-diabetic and anti-atherogenic responses in skeletal muscle of these patients, but the role of the circadian clock-machinery remains unknown. This study investigated whether mRNA expression of clock genes in skeletal muscle of CAD and T2DM patients is influenced by physical ET intervention. Nineteen patients with CAD and T2DM (age 64 ± 5 years) were randomised to either six months of ET (four weeks of in-hospital ET followed by a five-month ambulatory programme) or usual care. At the beginning of the study, after four weeks and after six months parameters of metabolic and cardiovascular risk factors, and physical exercise capacity were assessed. Gene expression was measured in skeletal muscle biopsies by quantitative real-time polymerase chain reaction (PCR). A selection of clock genes and associated components (circadian locomoter output cycle kaput protein (CLOCK), period (PER) 1, cryptochrome (CRY) 2 and aminolevulinate-deltA-synthase-1 (ALAS1)) was reliably measured and used for further analysis. A time-dependent effect in gene expression was observed in CLOCK (p = 0.013) and a significant interaction between time and intervention was observed for ALAS1 (p = 0.032; p = 0.014) as a result of ET. This is the first study to analyse clock gene expression in skeletal muscles of patients with CAD and T2DM participating in a long-lasting exercise intervention. ET, as one of the cornerstones in prevention and rehabilitation of CAD and T2DM, exerts no effects on CLOCK genes but meaningful effects on the clock-associated gene ALAS1. © The European Society of Cardiology 2016.

  17. Gene Expression Associated with Early and Late Chronotypes in Drosophila melanogaster

    Directory of Open Access Journals (Sweden)

    Mirko ePegoraro

    2015-05-01

    Full Text Available The circadian clock provides the temporal framework for rhythmic behavioural and metabolic functions. In the modern era of industrialization, work and social pressures, the clock function is often jeopardized, resulting in adverse and chronic effects on health. Understanding circadian clock function, particularly individual variation in diurnal phase preference (chronotype, and the molecular mechanisms underlying such chronotypes may lead to interventions that could abrogate clock dysfunction and improve human (and animal health and welfare. Our preliminary studies suggested that fruitflies, like humans, can be classified as early rising ‘larks’ or late rising ‘owls’, providing a convenient model system for these types of studies. We have identified strains of flies showing increased preference for morning emergence (Early or E from the pupal case, or more pronounced preference for evening emergence (Late or L. We have sampled pupae the day before eclosion (4th day after pupariation at 4 h intervals in the E and L strains, and examined differences in gene expression by RNAseq. We have identified differentially expressed transcripts between the E and L strains which provide candidate genes for studies of Drosophila chronotypes and their human orthologues.

  18. Molecular clock on a neutral network.

    Science.gov (United States)

    Raval, Alpan

    2007-09-28

    The number of fixed mutations accumulated in an evolving population often displays a variance that is significantly larger than the mean (the overdispersed molecular clock). By examining a generic evolutionary process on a neutral network of high-fitness genotypes, we establish a formalism for computing all cumulants of the full probability distribution of accumulated mutations in terms of graph properties of the neutral network, and use the formalism to prove overdispersion of the molecular clock. We further show that significant overdispersion arises naturally in evolution when the neutral network is highly sparse, exhibits large global fluctuations in neutrality, and small local fluctuations in neutrality. The results are also relevant for elucidating aspects of neutral network topology from empirical measurements of the substitution process.

  19. Supporting Family Awareness with the Whereabouts Clock

    Science.gov (United States)

    Sellen, Abigail; Taylor, Alex S.; Kaye, Joseph ‘Jofish'; Brown, Barry; Izadi, Shahram

    We report the results of a field trial of a situated awareness device for families called the “Whereabouts Clock”. The Clock displays the location of family members using cellphone data as one of four privacy-preserving, deliberately coarse-grained categories ( HOME, WORK, SCHOOL or ELSEWHERE). The results show that awareness of others through the Clock supports not only family communication and coordination but also more emotive aspects of family life such as reassurance, connectedness, identity and social touch. We discuss how the term “awareness” means many things in practice and highlight the importance of designing not just for family activities, but in order to support the emotional, social and even moral aspects of family life.

  20. Quantifying fluctuations in reversible enzymatic cycles and clocks

    Science.gov (United States)

    Wierenga, Harmen; ten Wolde, Pieter Rein; Becker, Nils B.

    2018-04-01

    Biochemical reactions are fundamentally noisy at a molecular scale. This limits the precision of reaction networks, but it also allows fluctuation measurements that may reveal the structure and dynamics of the underlying biochemical network. Here, we study nonequilibrium reaction cycles, such as the mechanochemical cycle of molecular motors, the phosphorylation cycle of circadian clock proteins, or the transition state cycle of enzymes. Fluctuations in such cycles may be measured using either of two classical definitions of the randomness parameter, which we show to be equivalent in general microscopically reversible cycles. We define a stochastic period for reversible cycles and present analytical solutions for its moments. Furthermore, we associate the two forms of the randomness parameter with the thermodynamic uncertainty relation, which sets limits on the timing precision of the cycle in terms of thermodynamic quantities. Our results should prove useful also for the study of temporal fluctuations in more general networks.

  1. Circadian modulation of short-term memory in Drosophila.

    Science.gov (United States)

    Lyons, Lisa C; Roman, Gregg

    2009-01-01

    Endogenous biological clocks are widespread regulators of behavior and physiology, allowing for a more efficient allocation of efforts and resources over the course of a day. The extent that different processes are regulated by circadian oscillators, however, is not fully understood. We investigated the role of the circadian clock on short-term associative memory formation using a negatively reinforced olfactory-learning paradigm in Drosophila melanogaster. We found that memory formation was regulated in a circadian manner. The peak performance in short-term memory (STM) occurred during the early subjective night with a twofold performance amplitude after a single pairing of conditioned and unconditioned stimuli. This rhythm in memory is eliminated in both timeless and period mutants and is absent during constant light conditions. Circadian gating of sensory perception does not appear to underlie the rhythm in short-term memory as evidenced by the nonrhythmic shock avoid