DOCSLIB.ORG

Degradation Rate Uniformity Determines Success of Oscillations in Repressive Feedback Regulatory Net- Works

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Degradation Rate Uniformity Determines Success of Oscillations in Repressive Feedback Regulatory Net- Works Degradation rate uniformity determines success of oscillations in repressive feedback regulatory net- works Karen M. Page1 & Ruben Perez-Carrasco1 1 Department of Mathematics, University College London, Gower Street, WC1E 6BT, London, UK October 9, 2018 Summary the nuclear factor κB localization [Hoffmann et al., 2002]. Ring oscillators are biochemical circuits consisting Due to their range of utilities, different oscillatory of a ring of interactions capable of sustained oscil- gene regulatory circuits have been synthetically en- lations. The non-linear interactions between genes gineered [Purcell et al., 2010]. In particular, lot of hinder the analytical insight into their function, attention has been focused on the engineering of ring usually requiring computational exploration. Here oscillators consisting of a set of genes interacting with we show that, despite the apparent complexity, the each other sequentially and forming a repressive feed- stability of the unique steady state in an incoher- back loop. This work was initiated by the synthesis of ent feedback ring depends only on the degradation the 3-gene repressilator [Elowitz and Leibler, 2000], rates and a single parameter summarizing the feed- that has been further refined to improve its oscillation back of the circuit. Concretely, we show that the properties (e.g. [Stricker et al., 2008, Niederholtmeyer range of regulatory parameters that yield oscilla- et al., 2015]). Consequently, the theoretical and nu- tory behaviour, is maximized when the degradation merical analysis of the working of ring oscillators has rates are equal. Strikingly, this results holds inde- also received substantial attention. Such work was pio- pendently of the regulatory functions used or num- neered by Fraser and Tiwari [Fraser and Tiwari, 1974] ber of genes. We also derive properties of the oscil- who performed numerical simulations. Subsequent lations as a function of the degradation rates and analysis showed that for sufficiently strong repression, number of nodes forming the ring. Finally, we ex- oscillations arise due to a Hopf bifurcation, relating the plore the role of mRNA dynamics by applying the genetic oscillatory behaviour with dynamical systems generic results to the specific case with two natu- theory [Smith, 1987], which has lead to many different rally different degradation time scales. studies delving into dynamical properties of the oscilla- tions (e.g. [Buşe et al., 2009, Buşe et al., 2010,Müller et al., 2006,Garcia-Ojalvo et al., 2004,Pigolotti et al., Introduction 2007]) These analytical and numerical studies of biochem- Genetic regulatory networks (GRNs), consisting of the ical circuits require insight into a set of simultane- arXiv:1802.05946v1 [q-bio.MN] 16 Feb 2018 interactions between a set of genes, are core to the reg- ous non-linear feedback interactions between multiple ulation of the temporal genetic expression profiles re- genes usually analyzed as a set of ordinary differential quired for various cellular processes, ranging from cell equations (ODEs). Determining the role of different fate determination during embryogenesis to cellular parameters in the solutions to these equations poses homesotasis [Davidson and Levine, 2005,Levine and enormous analytical complexity that hinders quantita- Davidson, 2005, Panovska-Griffiths et al., 2013, Olson, tive studies. For this reason, computational and ana- 2006, Sauka-Spengler and Bronner-Fraser, 2008, De- lytical studies are often reduced to tackling relatively quéant et al., 2006]. GRNs are capable of many dy- small networks, and, even in such cases, to a reduced namical functions, including oscillatory gene expres- parameter set or certain simplified regulatory func- sion [Monk, 2003], as has been observed in somito- tions. This can constrain the range of application of the genesis [Hirata et al., 2002], circadian clocks [Reddy results found [Estrada et al., 2016]. Even in the case and Rey, 2014], the activity of the p53 tumor suppres- of the repressilator, the dynamical complexity can be sor [Bar-Or et al., 2000, Michael and Oren, 2003]) or huge [Potapov et al., 2015] and restrictive assumptions Degradation rate uniformity determines success of oscillations in repressive feedback regulatory networks within the quantitative model again become unavoid- able. This highlights the necessity to develop tools x_ = δ (f (x ) − x ) capable of understanding dynamical properties of the 1 1 1 3 1 system independently of the regulatory functions used. x_ 2 = δ2(f2(x1) − x2) A useful assumption, present in the vast majority of x_ 3 = δ3(f3(x2) − x3); studies, is that the degradation rates of proteins are where f ; f and f describe the repressive interactions identical for different genes. However, due to the high 1 2 3 between genes and are therefore decreasing, positive span of protein structures and mechanisms controlling functions. f can be thought of as the maximal ex- degradation rates, such as ubiquitination [Bachmair i pression level of gene i multiplied by the probability et al., 1986,Dice, 1987], the turnover rate can range that its repressor is inactive. At any given steady state orders of magnitude in the proteome of a single sys- (x∗; x∗; x∗) given by x_ =x _ =x _ = 0, the repressi- tem [Belle et al., 2006, Christiano et al., 2014]. Since 1 2 3 1 2 3 lator follows the relationship x∗ = f (f (f (x∗))) ≡ oscillations in a network are generated by an ongoing 3 3 2 1 3 F (x∗), where the function F captures the overall neg- imbalance between the production and degradation of 3 ative feedback. Since F (x) is a decreasing, positive the different species, it is expected that degradation function, there is a unique possible value of x∗, which rates play a determinant role in the behavior of oscilla- 3 yields unique values x∗ = f (x∗) and x∗ = f (x∗). The tory circuits. Particuarly, simulations of a repressilator 1 1 3 2 2 1 stability of the protein levels dictated by this steady model showed that oscillations are favoured for com- state, can be computed through the eigenvalues of its parable values of the degradation of the protein and Jacobian matrix mRNA [Elowitz and Leibler, 2000], and, more gener- 0 0 ∗ 1 ally, a certain level of symmetry around the ring [Tuttle −δ1 0 δ1f1(x3) 0 ∗ et al., 2005]. Nevertheless, there is no analytical study J = @ δ2f2(x1) −δ2 0 A : (1) 0 ∗ that gives insight into the role of degradation rates for 0 δ3f3(x2) −δ3 general ring oscillators independent of the regulatory These eigenvalues λ satisfy the characteristic equa- functions used. tion To gain insight into the role of degradation rates in oscillatory networks, dynamical system theory and (λ + δ1)(λ + δ2)(λ + δ3) + Aδ1δ2δ3 = 0; (2) bifurcation theory have proven to be essential tools. where A ≡ −f 0 (x∗)f 0 (x∗)f 0 (x∗) = −F 0(x∗) is the These allow us to categorise different possible dynam- 1 3 2 1 3 2 3 modulus of the slope of the composite repression func- ical responses of oscillatory networks [Smith, 1987, tion at the steady state. Interestingly, the parameter Strelkowa and Barahona, 2010,K. et al., 2015,Monk, A contains all the details of the interactions of the 2003]. Using bifurcation theory we aim to obtain infor- network necessary to solve the characteristic equation mation on the role of degradation rates in oscillatory (2). This means that the eigenvalues of the character- networks, making these results as general as possible istic equation and so the stability of the steady protein and using minimal details of the regulatory functions. levels will depend only on A and on the degradation Specifically, we show how relevant information on the rates. This allows us to perform the stability analysis interactions between different genes can be captured without any further information on the explicit form with a single parameter. We show how this parame- of the repressive interactions. Concretely, since F (x) ter controls the appearance of oscillations through a is a monotonically decreasing function (A > 0) the Hopf bifurcation. First we develop our methodology product of the eigenvalues of J will always be negative, for the repressilator, expanding the theory in the fol- lowing sections to negative feedback rings oscillators λ λ λ = det J = −δ δ δ (1 + A) < 0: (3) with an arbitrary number of species. Finally we study 1 2 3 1 2 3 the case in which the species are categorized as mRNAs Therefore, the repressive ring forbids any eigenvalue to and proteins, which have distinct degradation rates, be zero. As a result, a change in stability of the steady giving insight in the role of mRNA dynamics in the state can only occur through a Hopf bifurcation, in performance of ring oscillators. which a pair of complex conjugate eigenvalues crosses the imaginary axis. We write this pair λe2 = iα and λe3 = −iα, where α is the angular velocity of the sus- Results tained oscillations that appear at the Hopf bifurcation. Following Equation(3), the other eigenvalue λ1 must Three-gene repressilator be real and negative, everywhere, and in particular at the Hopf bifurcation (λe1 < 0). The classic general form of the repressilator consists of Introducing the purely imaginary eigenvalues λe2 and three genes repressing each other sequentially [Elowitz λe3 in the characteristic equation (2), expressions for and Leibler, 2000] (Fig.1a). In the simple case in α and A~ (value of A at the bifurcation) are obtained which mRNA dynamics are considered fast compared that only depend on the degradation rates, with protein dynamics, the dynamical evolution of the p system can be described as a set of ODEs α = δ1δ2 + δ2δ3 + δ3δ1: (4) Page 2 of 17 Degradation rate uniformity determines success of oscillations in repressive feedback regulatory networks a) X c) X X X X X X X b) X 1000 gene gene t t expression expression expression expression 300 100 30 10 steady states steady gene expression ~ Figure 1: Oscillatory behaviour of the repressilator a) Schematic of the repressilator b) Bifurcation diagram schematic shows how the oscillations appear and disappear through a Hopf bifurcation depending on the magnitude A that summarizes the negative feedback strength of the circuit.
Load more

Recommended publications
  • Design Principles of Genetic Circuits Justin Bois Caltech Spring, 2018
    BE 150: Design Principles of Genetic Circuits Justin Bois Caltech Spring, 2018 This document was prepared at Caltech with financial support from the Donna and Benjamin M. Rosen Bioengineering Center. © 2018 Justin Bois. All rights reserved except for figures taken from literature sources. 8 The repressilator Today we will talk about an oscillating genetic circuit developed by Elowitz and Leibler and published in 2000, called the repressilator. As we work through this example, we will learn a valuable technique for analyzing dynamical systems, including many of those we encounter in systems biology, linear stability analysis. 8.1 Design of the repressilator The repressilator consists of three repressors on a plasmid, as shown in Fig. 7. They are TetR, λ cI, and LacI. For our analysis here, the names are unimportant, and we will call them repressor 1, 2, and 3. Repressorletters 1 represses to production nature of repressor 2, which in turn represses production of repressor 3. Finally, repressor 3 represses production or repressor 1, completing the loop. a Repressilator Reporter 30 8C. At least 100 individual cell lineages in each of three micro- colonies were tracked manually, and their ¯uorescence intensity was P lac01 L quanti®ed. ampR tetR-lite The timecourse of the ¯uorescence of one such cell is shown in PLtet01 Fig. 2. Temporal oscillations (in this case superimposed on an kanR overall increase in ¯uorescence) occur with a period of around Tet R Tet R pSC101 gfp-aav 150 minutes, roughly threefold longer than the typical cell-division λP origin R time. The amplitude of oscillations is large compared with baseline λ cI LacI GFP levels of GFP ¯uorescence.
    [Show full text]
  • The Plant Circadian Clock and Chromatin Modifications
    G C A T T A C G G C A T genes Review The Plant Circadian Clock and Chromatin Modifications Ping Yang 1,2, Jianhao Wang 1,2, Fu-Yu Huang 3 , Songguang Yang 1,* and Keqiang Wu 3,* 1 Key Laboratory of South China Agricultural Plant Molecular Analysis and Genetic Improvement, South China Botanical Garden, Chinese Academy of Sciences, Guangzhou 510650, China; [email protected] (P.Y.); [email protected] (J.W.) 2 University of Chinese Academy of Sciences, Chinese Academy of Sciences, Beijing 100049, China 3 Institute of Plant Biology, National Taiwan University, Taipei 106, Taiwan; [email protected] * Correspondence: [email protected] (S.Y.); [email protected] (K.W.) Received: 3 October 2018; Accepted: 5 November 2018; Published: 20 November 2018 Abstract: The circadian clock is an endogenous timekeeping network that integrates environmental signals with internal cues to coordinate diverse physiological processes. The circadian function depends on the precise regulation of rhythmic gene expression at the core of the oscillators. In addition to the well-characterized transcriptional feedback regulation of several clock components, additional regulatory mechanisms, such as alternative splicing, regulation of protein stability, and chromatin modifications are beginning to emerge. In this review, we discuss recent findings in the regulation of the circadian clock function in Arabidopsis thaliana. The involvement of chromatin modifications in the regulation of the core circadian clock genes is also discussed. Keywords: circadian clock; oscillators; transcriptional and post-transcriptional regulation; chromatin modifications; Arabidopsis 1. Introduction Plants, like animals, exhibit rhythmic biological activity, with a periodicity of 24 h.
    [Show full text]
  • The Plant Circadian Oscillator
    biology Review The Plant Circadian Oscillator C. Robertson McClung Department of Biological Sciences, Dartmouth College, Hanover, NH 03755, USA; [email protected]; Tel.: +1-603-646-3940 Received: 19 December 2018; Accepted: 9 March 2019; Published: 12 March 2019 Abstract: It has been nearly 300 years since the first scientific demonstration of a self-sustaining circadian clock in plants. It has become clear that plants are richly rhythmic, and many aspects of plant biology, including photosynthetic light harvesting and carbon assimilation, resistance to abiotic stresses, pathogens, and pests, photoperiodic flower induction, petal movement, and floral fragrance emission, exhibit circadian rhythmicity in one or more plant species. Much experimental effort, primarily, but not exclusively in Arabidopsis thaliana, has been expended to characterize and understand the plant circadian oscillator, which has been revealed to be a highly complex network of interlocked transcriptional feedback loops. In addition, the plant circadian oscillator has employed a panoply of post-transcriptional regulatory mechanisms, including alternative splicing, adjustable rates of translation, and regulated protein activity and stability. This review focuses on our present understanding of the regulatory network that comprises the plant circadian oscillator. The complexity of this oscillatory network facilitates the maintenance of robust rhythmicity in response to environmental extremes and permits nuanced control of multiple clock outputs. Consistent with this view, the clock is emerging as a target of domestication and presents multiple targets for targeted breeding to improve crop performance. Keywords: circadian rhythms; circadian clock; transcriptional feedback loops; plant circadian clock; posttranscriptional; posttranslational; alternative splicing; protein stability 1. Introduction This special issue celebrates the 2017 Nobel Prize in Physiology awarded to Jeff Hall, Michael Rosbash, and Mike Young.
    [Show full text]
  • Building and Understanding Repressilators and Synchrony
    BUILDING AND UNDERSTANDING: REPRESSILATORS AND SYNCHRONY THREE CELLS IN A BUBBLE BATH Brenton Munson Clara Posner Jonathan Pekar October 23, 2016 UNIVERSITY OF CALIFORNIA SAN DIEGO DEPARTMENT OF BIOENGINNERING BENG 221 PROJECT i Contents ii CONTENTS 1 Introduction 1 1.1 Problem Statement . .2 2 Methods 2 2.1 Simplifying and Parameterization . .2 2.2 Numerical Method . .5 3 Results and Discussion6 References 14 4 Appendix 16 4.1 Code............................................. 16 4.1.1 Single Cell Repressilator Model . 16 4.1.2 Repressilator Model with Drug Sensing . 23 LISTOFFIGURES Figure 1 Repressilator System . .2 Figure 2 Repressilator with Drug Sensing . .4 Figure 3 Multicell Environment . .5 Figure 4 Starting a Repressilator . .7 Figure 5 Repressilator at Steady State . .8 Figure 6 Model Sensitivity to Transcription Rate . .8 Figure 7 Model Sensitivity to Translation Rate . .9 Figure 8 Model Sensitivity to Protein Degradation Rates . 10 Figure 9 Model Sensitivity to Hill Parameters . 11 Figure 10 Controlling a Repressilator with an External Stimulus . 11 Figure 11 Cell Synchronization . 12 Figure 12 Cell Synchronization . 12 INTRODUCTION 1 1 INTRODUCTION The demands placed upon modern medicine are rapidly driving scientists and engineers to inves- tigate the mechanisms of disease and biological function to ever-increasing complexity. Much of this frontier lies in modeling not only the outcome of individual genetic perturbations but also how the networks of genetic interactions coordinate cellular- and tissue-level phenotypes [1]. While progress has been made in describing living systems, these networks often become too complex and cumbersome to interrogate with the current tools of bioinformatics [2,3]. As such, the problem is often broken down into more discrete subunits, with which they can be modeled, perturbed, and hopefully understood on a more functional level [4].
    [Show full text]
  • Synthetic Biology Approaches in Cancer Immunotherapy, Genetic
    Integrative Biology REVIEW ARTICLE Synthetic biology approaches in cancer immunotherapy, genetic network engineering, Downloaded from https://academic.oup.com/ib/article-abstract/8/4/504/5163774 by Columbia University user on 27 March 2020 Cite this: Integr. Biol., 2016, 8, 504 and genome editing Deboki Chakravarti,* Jang Hwan Cho,† Benjamin H. Weinberg,† Nicole M. Wong† and Wilson W. Wong* Investigations into cells and their contents have provided evolving insight into the emergence of complex biological behaviors. Capitalizing on this knowledge, synthetic biology seeks to manipulate the cellular machinery towards novel purposes, extending discoveries from basic science to new applications. While these developments have demonstrated the potential of building with biological parts, the complexity of cells can pose numerous challenges. In this review, we will highlight the broad Received 24th December 2015, and vital role that the synthetic biology approach has played in applying fundamental biological Accepted 31st March 2016 discoveries in receptors, genetic circuits, and genome-editing systems towards translation in the fields DOI: 10.1039/c5ib00325c of immunotherapy, biosensors, disease models and gene therapy. These examples are evidence of the strength of synthetic approaches, while also illustrating considerations that must be addressed when www.rsc.org/ibiology developing systems around living cells. Insight, innovation, integration Investigations into cells and their contents have provided evolving insight into the emergence of complex behaviors. Capitalizing on this knowledge, synthetic biology seeks to manipulate the cellular machinery towards novel purposes, extending basic discoveries from basic science to new applications. While these developments have demonstrated the potential of building with biological parts, the complexity of cells can pose numerous challenges.
    [Show full text]
  • Realizing the Potential of Synthetic Biology
    Nature Reviews Molecular Cell Biology | AOP, published online 12 March 2014; doi:10.1038/nrm3767 PERSPECTIVES computer-aided-design (CAD), safety sys- VIEWPOINT tems, integrating models, genome editing and accelerated evolution. Synthetic biology Realizing the potential of synthetic is less like highly modular (or ‘switch-like’) electrical engineering and computer science biology and more like civil and mechanical engineer- ing in its use of optimization of modelling of whole system-level stresses and traffic flow. George M. Church, Michael B. Elowitz, Christina D. Smolke, Christopher A. Voigt and Ron Weiss Michael B. Elowitz. At the most general level, synthetic biology expands the subject Abstract | Synthetic biology, despite still being in its infancy, is increasingly matter of biology from the (already enor- providing valuable information for applications in the clinic, the biotechnology mous) space of existing species and cellular industry and in basic molecular research. Both its unique potential and the systems that have evolved to the even larger challenges it presents have brought together the expertise of an eclectic group of space of non-natural, but feasible, species scientists, from cell biologists to engineers. In this Viewpoint article, five experts and systems. Although we started with circuits to carry out the simplest kinds of discuss their views on the future of synthetic biology, on its main achievements in dynamic behaviours, synthetic approaches basic and applied science, and on the bioethical issues that are associated with the can be applied broadly to all types of bio­ design of new biological systems. logical functions from metabolism to multi­ cellular development. Synthetic biology allows us to figure out what types of genetic An increasing number of publications For instance, insight gained from systems circuit designs are capable of implement- and institutions are dedicated to biology investigations of natural processes ing different cellular behaviours, and what synthetic biology.
    [Show full text]
  • Feedback Loops of the Mammalian Circadian Clock Constitute Repressilator
    RESEARCH ARTICLE Feedback Loops of the Mammalian Circadian Clock Constitute Repressilator J. Patrick Pett1, Anja Korenčič2, Felix Wesener1, Achim Kramer3, Hanspeter Herzel4* 1 Institute for Theoretical Biology, Humboldt-UniversitaÈt zu Berlin, Berlin, Germany, 2 Center for Functional Genomics and Biochips, Institute of Biochemistry, Faculty of Medicine, University of Ljubljana, Ljubljana, Slovenia, 3 Laboratory of Chronobiology, ChariteÂ-UniversitaÈtsmedizin Berlin, Berlin, Germany, 4 Institute for Theoretical Biology, ChariteÂ-UniversitaÈtsmedizin Berlin, Berlin, Germany * [email protected] a11111 Abstract Mammals evolved an endogenous timing system to coordinate their physiology and behav- iour to the 24h period of the solar day. While it is well accepted that circadian rhythms are generated by intracellular transcriptional feedback loops, it is still debated which network motifs are necessary and sufficient for generating self-sustained oscillations. Here, we sys- OPEN ACCESS tematically explore a data-based circadian oscillator model with multiple negative and posi- Citation: Pett JP, Korenčič A, Wesener F, Kramer A, tive feedback loops and identify a series of three subsequent inhibitions known as Herzel H (2016) Feedback Loops of the ªrepressilatorº as a core element of the mammalian circadian oscillator. The central role of Mammalian Circadian Clock Constitute the repressilator motif is consistent with time-resolved ChIP-seq experiments of circadian Repressilator. PLoS Comput Biol 12(12): clock transcription factors and loss of rhythmicity in core clock gene knockouts. e1005266. doi:10.1371/journal.pcbi.1005266 Editor: Denis Thieffry, Ecole Normale SupeÂrieure, FRANCE Received: August 23, 2016 Author Summary Accepted: November 27, 2016 Circadian clocks are endogenous oscillators that drive daily rhythms in physiology, Published: December 12, 2016 metabolism and behavior.
    [Show full text]
  • Robustness of Synthetic Oscillators in Growing and Dividing Cells
    Robustness of synthetic oscillators in growing and dividing cells Joris Paijmans,1 David K. Lubensky,2 and Pieter Rein ten Wolde1 1AMOLF, Science Park 104, 1098 XG Amsterdam, The Netherlands 2Department of Physics, University of Michigan, Ann Arbor, MI 48109-1040, USA Abstract Synthetic biology sets out to implement new functions in cells, and to develop a deeper under- standing of biological design principles. In 2000, Elowitz and Leibler showed that by rational design of the reaction network, and using existing biological components, they could create a network that exhibits periodic gene expression, dubbed the repressilator (Elowitz and Leibler, Nature, 2000). More recently, Stricker et al. presented another synthetic oscillator, called the dual-feedback oscil- lator (Stricker et al., 2008), which is more stable. How the stability of these oscillators is affected by the intrinsic noise of the interactions between the components and the stochastic expression of their genes, has been studied in considerable detail. However, as all biological oscillators reside in growing and dividing cells, an important question is how these oscillators are perturbed by the cell cycle. In previous work we showed that the periodic doubling of the gene copy numbers due to DNA replication can couple not only natural, circadian oscillators to the cell cycle (Paijmans et al., PNAS, 113, 4063, (2016)), but also these synthetic oscillators. Here we expand this study. We find that the strength of the locking between oscillators depends not only on the positions of the genes on the chromosome, but also on the noise in the timing of gene replication: noise tends to weaken the coupling.
    [Show full text]
  • Repressilator - Wikipedia, the Free Encyclopedia Repressilator from Wikipedia, the Free Encyclopedia
    3/3/2016 Repressilator - Wikipedia, the free encyclopedia Repressilator From Wikipedia, the free encyclopedia The repressilator is a synthetic genetic regulatory network consisting of a ring­oscillator with three genes, each expressing a protein that represses the next gene in the loop.[1] Contents 1 Discovery 2 Impact 3 References 4 External links Discovery Repressilators were first reported in a paper[2] by Michael Elowitz and Stanislas Leibler in 2000. This network was designed from scratch to exhibit a stable oscillation which is reported via the expression of green fluorescent protein, and hence acts like an electrical oscillator system with fixed time periods. The network was implemented in Escherichia coli using standard molecular biology methods and observations were performed that verify that the engineered colonies do indeed exhibit the desired oscillatory behavior. The repressilator consists of three genes connected in a feedback loop, such that each gene represses the next gene in the loop, and is repressed by the previous gene. In addition, green fluorescent protein is used as a reporter so that the behavior of the network can be observed using fluorescence microscopy. The repressilator genetic regulatory network. The design of the repressilator was guided by two simple mathematical models, one continuous and deterministic and the other discrete and stochastic. https://en.wikipedia.org/wiki/Repressilator 1/4 3/3/2016 Repressilator - Wikipedia, the free encyclopedia A continuous simulation of the repressilator. These models were analyzed to determine the values for the various rates which would yield a sustained oscillation. It was found that these oscillations were favoured by strong promoters A discrete and stochastic simulation of the repressilator.
    [Show full text]
  • Synchronous Long-Term Oscillations in a Synthetic Gene Circuit
    View metadata, citation and similar papers at core.ac.uk brought to you by CORE provided by Apollo Noise-proofing the repressilator: synchronous long-term oscillations in a synthetic gene circuit Laurent Potvin-Trottier†‡ Nathan D. Lord† Glenn Vinnicombe§ Johan Paulsson† Synthetically engineered genetic circuits can perform a error propagation from the reporter system, from core cellular 55 wide range of tasks but generally with lower accuracy than processes, and from within the circuit itself. 5 natural systems. Here we revisited the first synthetic ge- netic oscillator, the repressilator1, and modified it based The repressilator consists of three genes – tetR from the Tn10 transposon, cI from bacteriophage λ and lacI from the lac- on principles from stochastic chemistry in single cells. 20 Specifically, we sought to reduce error propagation and in- tose operon – and each repressor has a C-terminal ssrA tag formation losses, not by adding control loops, but by sim- that targets it for degradation (Fig. 1A). The whole circuit was 60 encoded on a low-copy pSC101 plasmid in an Escherichia coli 10 ply removing existing features. This created highly reg- ular and robust oscillations. Some streamlined circuits strain lacking lacI, and a second high-copy ColE1 reporter plas- mid encoded GFP under the control of TetR with a modified kept 14 generation periods over a range of growth con- 1,21 ditions and kept phase for hundreds of generations in sin- degradation tag . We first reevaluated this circuit using a gle cells, allowing cells in flasks and colonies to oscillate microfluidic device in which cells are trapped in short chan- 65 nels and newborn cells are washed away by fresh medium22,23 15 synchronously without any coupling between them.
    [Show full text]
  • 1 Short Title: Monocot ELF3 Functions in Dicots 1 2 3 4 5 6 Corresponding
    bioRxiv preprint doi: https://doi.org/10.1101/131185; this version posted May 1, 2017. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license. 1 Short Title: Monocot ELF3 functions in dicots 2 3 4 5 6 7 Corresponding Author: 8 9 Dmitri A. Nusinow, Ph.D. 10 Assistant Member 11 Danforth Plant Science Center 12 975 N. Warson Rd.- Room 3030 13 St. Louis, MO 63132 14 (314)587-1489 15 [email protected] 16 1 bioRxiv preprint doi: https://doi.org/10.1101/131185; this version posted May 1, 2017. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license. 17 Cross-species complementation reveals conserved functions for EARLY FLOWERING 3 18 between monocots and dicots 19 20 He Huang, Malia A. Gehan, Sarah E. Huss, Sophie Alvarez2, Cesar Lizarraga, Ellen L. 21 Gruebbling, John Gierer, Michael J. Naldrett2, Rebecca K. Bindbeutel, Bradley S. Evans, Todd 22 C. Mockler, and Dmitri A. Nusinow* 23 24 Donald Danforth Plant Science Center, St. Louis, MO, 63132, USA (H.H., M.A.G., S.A., C.L., 25 J.G., M.J.N., R.K.B., B.S.E., T.C.M., D.A.N.) 26 Webster University, Webster Groves, MO, 63119, USA (S.E.H.) 27 Saint Louis University, St.
    [Show full text]
  • Systems Level Generation of Mammalian Circadian Rhythms and Its Connection to Liver Metabolism
    Systems level generation of mammalian circadian rhythms and its connection to liver metabolism DISSERTATION zur Erlangung des akademischen Grades Doctor rerum naturalium (Dr. rer. nat.) eingereicht an der Lebenswissenschaftlichen Fakultät der Humboldt-Universität zu Berlin von M.Sc., Jan Patrick, PETT Präsidentin der Humboldt-Universität zu Berlin: Prof. Dr.-Ing. Dr. Sabine Kunst Dekan der Lebenswissenschaftlichen Fakultät der Humboldt-Universität zu Berlin: Prof. Dr. Bernhard Grimm Gutachter/innen: 1. Prof. Dr. Hanspeter Herzel 2. Prof. Dr. Nils Blüthgen 3. Prof. Dr. Didier Gonze Tag der mündlichen Prüfung: 30.4.2019 HUMBOLDT-UNIVERSITÄT ZU BERLIN Abstract Lebenswissenschaftlichen Fakultät Institute for Theoretical Biology Doctor rerum naturalium Systems level generation of mammalian circadian rhythms and its connection to liver metabolism by Jan Patrick, PETT Circadian clocks are endogenous oscillators that generate 24-hour rhythms. They allow many organisms to synchronize their physiology and behaviour with daily changes of the environment. In mammals such clocks are based on transcriptional- translational feedbacks and exist in almost every cell. Within an organism different clocks are distinguished by their localization in different organs, which is connected with differences in the functions they modulate in a daily manner. One of the key physiological functions of circadian clocks in various organs seems to be the tempo- ral alignment of metabolic processes. The general mechanism of feedback regulation that generates circadian rhyth- micity has been uncovered, but it is not fully understood which feedback loops con- tribute to rhythm generation. A group of so called core clock genes that appear to be relevant for generating rhythmicity is connected in a complex network that com- prises a multitude of such loops.
    [Show full text]