Modelling Robust Feedback Control Mechanisms That Ensure Reliable Coordination of Histone Gene Expression with DNA Replication

Funding: Andrea Christopher was supported by the University of Aberdeen through a Milner Studentship. Heike Hameister was supported by a Postgraduate Research Studentship of the University of Aberdeen. Oliver Ebenhöh was supported by the University of Aberdeen and the Deutsche Forschungsgemeinschaft [Cluster of Excellence on Plant Sciences, CEPLAS (EXC 1028)]. Berndt Müller was supported by the University of Aberdeen. Ekkehard Ullner was supported by the Scottish Universities Life Sciences Alliance (SULSA). The funders provided support in the form of salaries for authors but did not have any additional role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript. The specific roles of these authors are articulated in the ‘author contributions’ section. Heike Hameister is currently employed by Merck Serono GmbH. Merck Serono GmbH did not provide any support for this work and did not have any role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript. This does not alter our adherence to PLOS ONE policies on sharing data and materials.Peer reviewedPublisher PD

The molecular aetiology of tRNA synthetase depletion : induction of a GCN4 amino acid starvation response despite homeostatic maintenance of charged tRNA levels

Biotechnology and Biological Sciences Research Council (BBSRC) [BB/I020926/1 to I.S., BB/N017161/1 to I.S., M.C.R.]; BBSRC PhD studentship awards [M108703G, C103817D to I.S. and M.C.R.]. Funding for open access charge: Biotechnology and Biological Sciences Research Council.Peer reviewedPreprintPublisher PD

Modelling autoregulation of histone gene expression.

<p>(A) Model structure. Expansion of the basic models to test the effect of additional H2B genes on the expression of endogenous H2B and H3 genes. (B) and (C). Panels describe the time evaluation of histone RNA (<i>R</i>), histone proteins (<i>H</i>), free histone binding sites on DNA (<i>D</i>), nucleosome packed DNA formed from endogenous histone H2B and H3 (<i>T</i>_<i>e</i>), formed from exogenous H2B and endogenous H3 (<i>T</i>_<i>p</i>) simulated by the mathematical models as a function of the external influence of SLBP (<i>S</i>) and DNA synthesis (<i>V</i>_<i>5</i>). Shown are results for the histone feedback loop (B) and DNA coupled (C) model.</p

Evidence for a feedback control mechanism that regulates histone gene expression.

<p>U2OS cells were transfected with plasmids pEGFP- H2B (H2B-GFP) or pEGFP (GFP) and subjected to antibiotic selection produce stable lines prior to FACS and protein analysis. (A, C) Analysis of H2B protein levels. Proteins were separated by SDS PAGE and analysed by Western blotting. Shown are H2B, H3 and GAPDH protein levels. Histone protein levels were standardised with respect to GAPDH protein, with H2B or H3 protein levels in cells transfected with pEGFP (GFP) defined as 1. Note that endogenous H2B levels (H2B) are significantly reduced in cells expressing H2B-GFP. (B, D) Analysis of H2B RNA levels. H2B RNA levels were analysed by Northern blotting. Shown are H2B and GAPDH RNA levels. The RNA levels were standardised with respect to GAPDH RNA with H2B RNA levels in cells transfected with pEGFP (GFP) defined as 1. Model predictions of the averaged H2B proteins (E, G) and RNAs (F, H) from the histone feedback loop (E, F) and DNA coupled (G, H) model. Endogenous H2B is in black, exogenous H2B in red. The different bars in each plot illustrate the effect of the strength of the promoter controlling exogenous H2B expression.</p

Model structures and equations.

<p>Histone feedback loop model and DNA coupled model. (A) Model structure. Dashed lines illustrate the links between free histone proteins and histone RNA synthesis and degradation (histone feedback loop model). The links between DNA replication and histone RNA are illustrated by dotted lines (DNA coupled model). Solid lines are common to both models. (B) and (C). Full set of equations for the histone feedback loop model (B) and the alternative fluxes <i>v</i>_<i>1</i> and <i>v</i>_<i>2</i> for the DNA coupled model (C). Other equations and fluxes are common to both models.</p

A comparison of histone RNA levels measured experimentally and predicted by the model.

<p>(A) U2OS cells were synchronised and released into S phase as described in Materials and Methods. Total RNA was prepared before release into S phase (0 min time point) and at regular intervals after that, and histone H2B RNA levels were subsequently analysed by Northern blotting. 28S rRNA levels were also measured and H2B RNA levels were standardised using 28S rRNA as a reference. (B) The graph shows the model prediction of histone RNA based on the analysis of DNA replication from two independent experiments (RNA simulation exp1 and 2, see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0165848#pone.0165848.s003" target="_blank">S2 File</a>). Also shown is the quantitation of the RNA analysis by Northern blotting from these two experiments (RNA exp 1 and 2). Note these data were scaled to match the maxima of the model predictions.</p

The histone feedback loop and DNA coupled models differ in their response to the inhibition of DNA synthesis but not in their response to transcription blocks.

<p>Shown variables and colour coding are as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0165848#pone.0165848.g002" target="_blank">Fig 2</a>. In (A), inhibition of DNA replication was implemented by setting <i>V</i>_<i>5</i> to 0 at 15000 s. In (B), inhibition of transcription was implemented by setting the transcription flux <i>v</i>_<i>1</i> (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0165848#pone.0165848.g001" target="_blank">Fig 1</a>) to 0.</p

Psychostimulants

A current survey and synthesis of the most important findings in our understanding of the neurobiological mechanisms of addiction are detailed in our Neurobiology of Addiction series, each volume addressing a specific area of addiction. Psychostimulants, Volume 2 in the series, explores the molecular and cellular systems in the brain responsible for psychostimulant addiction, including both direct/indirect sympathomimetics and nonsympathomimetics. This volume introduces the readers to the history of psychostimulant use. The authors clearly differentiate the neurobiological effects into three distinct stages of the addiction cycle: binge/intoxication, withdrawal/negative affect, and preoccupation/anticipation. © 2020 Elsevier Inc. All rights reserved