Quantitative analysis of competition in post-transcriptional regulation reveals a novel signature in target expression variation

When small RNAs are loaded onto Argonaute proteins they can form the RNA-induced silencing complexes (RISCs), which mediate RNA interference. RISC-formation is dependent on a shared pool of Argonaute proteins and RISC loading factors, and is thus susceptible to competition among small RNAs for loading. We present a mathematical model that aims to understand how small RNA competition for the PTR resources affects target gene repression. We discuss that small RNA activity is limited by RISC-formation, RISC-degradation and the availability of Argonautes. Together, these observations explain a number of PTR saturation effects encountered experimentally. We show that different competition conditions for RISC-loading result in different signatures of PTR activity determined also by the amount of RISC-recycling taking place. In particular, we find that the small RNAs less efficient at RISC-formation, using fewer resources of the PTR pathway, can perform in the low RISC-recycling range equally well as their more effective counterparts. Additionally, we predict a novel signature of PTR in target expression levels. Under conditions of low RISC-loading efficiency and high RISC-recycling, the variation in target levels increases linearly with the target transcription rate. Furthermore, we show that RISC-recycling determines the effect that Argonaute scarcity conditions have on target expression variation. Our observations taken together offer a framework of predictions which can be used in order to infer from experimental data the particular characteristics of underlying PTR activity.Comment: 23 pages, 3 Figures, accepted for publication to the Biophysical Journa

PTRE-seq reveals mechanism and interactions of RNA binding proteins and miRNAs

A large number of RNA binding proteins (RBPs) and miRNAs bind to the 3′ untranslated regions of mRNA, but methods to dissect their function and interactions are lacking. Here the authors introduce post-transcriptional regulatory element sequencing (PTRE-seq) to dissect sequence preferences, interactions and consequences of RBP and miRNA binding

Main memory in HPC: do we need more, or could we live with less?

An important aspect of High-Performance Computing (HPC) system design is the choice of main memory capacity. This choice becomes increasingly important now that 3D-stacked memories are entering the market. Compared with conventional Dual In-line Memory Modules (DIMMs), 3D memory chiplets provide better performance and energy efficiency but lower memory capacities. Therefore, the adoption of 3D-stacked memories in the HPC domain depends on whether we can find use cases that require much less memory than is available now. This study analyzes the memory capacity requirements of important HPC benchmarks and applications. We find that the High-Performance Conjugate Gradients (HPCG) benchmark could be an important success story for 3D-stacked memories in HPC, but High-Performance Linpack (HPL) is likely to be constrained by 3D memory capacity. The study also emphasizes that the analysis of memory footprints of production HPC applications is complex and that it requires an understanding of application scalability and target category, i.e., whether the users target capability or capacity computing. The results show that most of the HPC applications under study have per-core memory footprints in the range of hundreds of megabytes, but we also detect applications and use cases that require gigabytes per core. Overall, the study identifies the HPC applications and use cases with memory footprints that could be provided by 3D-stacked memory chiplets, making a first step toward adoption of this novel technology in the HPC domain.This work was supported by the Collaboration Agreement between Samsung Electronics Co., Ltd. and BSC, Spanish Government through Severo Ochoa programme (SEV-2015-0493), by the Spanish Ministry of Science and Technology through TIN2015-65316-P project and by the Generalitat de Catalunya (contracts 2014-SGR-1051 and 2014-SGR-1272). This work has also received funding from the European Union’s Horizon 2020 research and innovation programme under ExaNoDe project (grant agreement No 671578). Darko Zivanovic holds the Severo Ochoa grant (SVP-2014-068501) of the Ministry of Economy and Competitiveness of Spain. The authors thank Harald Servat from BSC and Vladimir Marjanovi´c from High Performance Computing Center Stuttgart for their technical support.Postprint (published version

Variation in the organization and subunit composition of the mammalian pyruvate dehydrogenase complex E2/E3BP core assembly

The final version of this article is available at the link below.Crucial to glucose homoeostasis in humans, the hPDC (human pyruvate dehydrogenase complex) is a massive molecular machine comprising multiple copies of three distinct enzymes (E1–E3) and an accessory subunit, E3BP (E3-binding protein). Its icosahedral E2/E3BP 60-meric ‘core’ provides the central structural and mechanistic framework ensuring favourable E1 and E3 positioning and enzyme co-operativity. Current core models indicate either a 48E2+12E3BP or a 40E2+20E3BP subunit composition. In the present study, we demonstrate clear differences in subunit content and organization between the recombinant hPDC core (rhPDC; 40E2+20E3BP), generated under defined conditions where E3BP is produced in excess, and its native bovine (48E2+12E3BP) counterpart. The results of the present study provide a rational basis for resolving apparent differences between previous models, both obtained using rhE2/E3BP core assemblies where no account was taken of relative E2 and E3BP expression levels. Mathematical modelling predicts that an ‘average’ 48E2+12E3BP core arrangement allows maximum flexibility in assembly, while providing the appropriate balance of bound E1 and E3 enzymes for optimal catalytic efficiency and regulatory fine-tuning. We also show that the rhE2/E3BP and bovine E2/E3BP cores bind E3s with a 2:1 stoichiometry, and propose that mammalian PDC comprises a heterogeneous population of assemblies incorporating a network of E3 (and possibly E1) cross-bridges above the core surface.This work was partly supported by EPSRC (under grants GR/R99393/01 and EP/C015452/1)

Competitive titration in living sea urchin embryos of regulatory factors required for expression of the CyIIIa actin gene

Previous studies have located some twenty distinct sites within the 2.3 kb 5' regulatory domain of the sea urchin CyIIIa cytoskeletal actin gene, where there occur in vitro high-specificity interactions with nuclear DNA-binding proteins of the embryo. This gene is activated in late cleavage, exclusively in cells of the aboral ectoderm cell lineages. In this study, we investigate the functional importance in vivo of these sites of DNA-protein interaction. Sea urchin eggs were coinjected with a fusion gene construct in which the bacterial chloramphenicol acetyltransferase (CAT) reporter gene is under the control of the entire CyIIIa regulatory domain, together with molar excesses of one of ten nonoverlapping competitor subfragments of this domain, each of which contains one or a few specific site(s) of interaction. The exogenous excess binding sites competitively titrate the available regulatory factors away from the respective sites associated with the CyIIIa.CAT reporter gene. This provides a method for detecting in vivo sites within the regulatory domain that are required for normal levels of expression, without disturbing the structure of the regulatory domain. We thus identify five nonoverlapping regions of the regulatory DNA that apparently function as binding sites for positively acting transcriptional regulatory factors. Competition with a subfragment bearing an octamer site results in embryonic lethality. We find that three other sites display no quantitative competitive interference with CyIIIa.CAT expression, though as shown in the accompanying paper, two of these sites are required for control of spatial expression. We conclude that the complex CyIIIa regulatory domain must assess the state of many distinct and individually necessary interactions in order to properly regulate CyIIIa transcriptional activity in development

Higher Representations Duals

We uncover novel solutions of the 't Hooft anomaly matching conditions for scalarless gauge theories with matter transforming according to higher dimensional representations of the underlying gauge group. We argue that, if the duals exist, they are gauge theories with fermions transforming according to the defining representation of the dual gauge group. The resulting conformal windows match the one stemming from the all-orders beta function results when taking the anomalous dimension of the fermion mass to be unity which are also very close to the ones obtained using the Schwinger-Dyson approximation. We use the solutions to gain useful insight on the conformal window of the associated electric theory. A consistent picture emerges corroborating previous results obtained via different analytic methods and in agreement with first principle lattice explorations.Comment: RevTeX, 23 pages, 3 figure

A framework for interrogating social media images to reveal an emergent archive of war

The visual image has long been central to how war is seen, contested and legitimised, remembered and forgotten. Archives are pivotal to these ends as is their ownership and access, from state and other official repositories through to the countless photographs scattered and hidden from a collective understanding of what war looks like in individual collections and dusty attics. With the advent and rapid development of social media, however, the amateur and the professional, the illicit and the sanctioned, the personal and the official, and the past and the present, all seem to inhabit the same connected and chaotic space.However, to even begin to render intelligible the complexity, scale and volume of what war looks like in social media archives is a considerable task, given the limitations of any traditional human-based method of collection and analysis. We thus propose the production of a series of ‘snapshots’, using computer-aided extraction and identification techniques to try to offer an experimental way in to conceiving a new imaginary of war. We were particularly interested in testing to see if twentieth century wars, obviously initially captured via pre-digital means, had become more ‘settled’ over time in terms of their remediated presence today through their visual representations and connections on social media, compared with wars fought in digital media ecologies (i.e. those fought and initially represented amidst the volume and pervasiveness of social media images).To this end, we developed a framework for automatically extracting and analysing war images that appear in social media, using both the features of the images themselves, and the text and metadata associated with each image. The framework utilises a workflow comprising four core stages: (1) information retrieval, (2) data pre-processing, (3) feature extraction, and (4) machine learning. Our corpus was drawn from the social media platforms Facebook and Flickr

The C-Terminal Domain of the Arabinosyltransferase Mycobacterium tuberculosis EmbC Is a Lectin-Like Carbohydrate Binding Module

The D-arabinan-containing polymers arabinogalactan (AG) and lipoarabinomannan (LAM) are essential components of the unique cell envelope of the pathogen Mycobacterium tuberculosis. Biosynthesis of AG and LAM involves a series of membrane-embedded arabinofuranosyl (Araf) transferases whose structures are largely uncharacterised, despite the fact that several of them are pharmacological targets of ethambutol, a frontline drug in tuberculosis therapy. Herein, we present the crystal structure of the C-terminal hydrophilic domain of the ethambutol-sensitive Araf transferase M. tuberculosis EmbC, which is essential for LAM synthesis. The structure of the C-terminal domain of EmbC (EmbCCT) encompasses two sub-domains of different folds, of which subdomain II shows distinct similarity to lectin-like carbohydrate-binding modules (CBM). Co-crystallisation with a cell wall-derived di-arabinoside acceptor analogue and structural comparison with ligand-bound CBMs suggest that EmbCCT contains two separate carbohydrate binding sites, associated with subdomains I and II, respectively. Single-residue substitution of conserved tryptophan residues (Trp868, Trp985) at these respective sites inhibited EmbC-catalysed extension of LAM. The same substitutions differentially abrogated binding of di- and penta-arabinofuranoside acceptor analogues to EmbCCT, linking the loss of activity to compromised acceptor substrate binding, indicating the presence of two separate carbohydrate binding sites, and demonstrating that subdomain II indeed functions as a carbohydrate-binding module. This work provides the first step towards unravelling the structure and function of a GT-C-type glycosyltransferase that is essential in M. tuberculosis. Author Summary Top Tuberculosis (TB), an infectious disease caused by the bacillus Mycobacterium tuberculosis, burdens large swaths of the world population. Treatment of active TB typically requires administration of an antibiotic cocktail over several months that includes the drug ethambutol. This front line compound inhibits a set of arabinosyltransferase enzymes, called EmbA, EmbB and EmbC, which are critical for the synthesis of arabinan, a vital polysaccharide in the pathogen's unique cell envelope. How precisely ethambutol inhibits arabinosyltransferase activity is not clear, in part because structural information of its pharmacological targets has been elusive. Here, we report the high-resolution structure of the C-terminal domain of the ethambutol-target EmbC, a 390-amino acid fragment responsible for acceptor substrate recognition. Combining the X-ray crystallographic analysis with structural comparisons, site-directed mutagenesis, activity and ligand binding assays, we identified two regions in the C-terminal domain of EmbC that are capable of binding acceptor substrate mimics and are critical for activity of the full-length enzyme. Our results begin to define structure-function relationships in a family of structurally uncharacterised membrane-embedded glycosyltransferases, which are an important target for tuberculosis therapy