Chemogenetics-a transformational and translational platform

Neurologic disorders are frequently a result of inappropriate electrical and/or chemical signaling of neurons and glia. Ultimate remediation would necessitate reprogramming these signals. Historically, correcting neuronal and glial signaling is accomplished via drug therapy/administration, although they frequently fail to effectively and fully treat the underlying disorder. Developments in basic research have produced several new classes of potential therapeutics to directly and precisely control neuron activity at the single-cell level. We review one such technology, Designer Receptors Exclusively Activated by Designer Drugs, and suggest its potential as a powerful tool for augmenting neuronal and glial signaling and activity for basic and translational applications. Copyright 2015 American Medical Association. All rights reserved

The E-ELT Multi-Object Spectrograph: latest news from MOSAIC

There are 8000 galaxies, including 1600 at z larger than 1.6, which could be simultaneously observed in an E-ELT field of view of 40 sq. arcmin. A considerable fraction of astrophysical discoveries require large statistical samples, which can only be obtained with multi-object spectrographs (MOS). MOSAIC will provide a vast discovery space, enabled by a multiplex of 200 and spectral resolving powers of R=5000 and 20000. MOSAIC will also offer the unique capability of more than 10 "high-definition" (multi-object adaptive optics, MOAO) integral-field units, optimised to investigate the physics of the sources of reionization. The combination of these modes will make MOSAIC the world-leading MOS facility, contributing to all fields of contemporary astronomy, from extra-solar planets, to the study of the halo of the Milky Way and its satellites, and from resolved stellar populations in nearby galaxies out to observations of the earliest "first-light" structures in the Universe. It will also study the distribution of the dark and ordinary matter at all scales and epochs of the Universe. Recent studies of critical technical issues such as sky-background subtraction and MOAO have demonstrated that such a MOS is feasible with state-of-the-art technology and techniques. Current studies of the MOSAIC team include further trade-offs on the wavelength coverage, a solution for compensating for the non-telecentric new design of the telescope, and tests of the saturation of skylines especially in the near-IR bands. In the 2020s the E-ELT will become the world's largest optical/IR telescope, and we argue that it has to be equipped as soon as possible with a MOS to provide the most efficient, and likely the best way to follow-up on James Webb Space Telescope (JWST) observations.Comment: 10 pages, 3 Figures, in Ground-based and Airborne Instrumentation for Astronomy VI, 2016, Proc. SPI

Gene expression patterns unveil a new level of molecular heterogeneity in colorectal cancer.

The recognition that colorectal cancer (CRC) is a heterogeneous disease in terms of clinical behaviour and response to therapy translates into an urgent need for robust molecular disease subclassifiers that can explain this heterogeneity beyond current parameters (MSI, KRAS, BRAF). Attempts to fill this gap are emerging. The Cancer Genome Atlas (TGCA) reported two main CRC groups, based on the incidence and spectrum of mutated genes, and another paper reported an EMT expression signature defined subgroup. We performed a prior free analysis of CRC heterogeneity on 1113 CRC gene expression profiles and confronted our findings to established molecular determinants and clinical, histopathological and survival data. Unsupervised clustering based on gene modules allowed us to distinguish at least five different gene expression CRC subtypes, which we call surface crypt-like, lower crypt-like, CIMP-H-like, mesenchymal and mixed. A gene set enrichment analysis combined with literature search of gene module members identified distinct biological motifs in different subtypes. The subtypes, which were not derived based on outcome, nonetheless showed differences in prognosis. Known gene copy number variations and mutations in key cancer-associated genes differed between subtypes, but the subtypes provided molecular information beyond that contained in these variables. Morphological features significantly differed between subtypes. The objective existence of the subtypes and their clinical and molecular characteristics were validated in an independent set of 720 CRC expression profiles. Our subtypes provide a novel perspective on the heterogeneity of CRC. The proposed subtypes should be further explored retrospectively on existing clinical trial datasets and, when sufficiently robust, be prospectively assessed for clinical relevance in terms of prognosis and treatment response predictive capacity. Original microarray data were uploaded to the ArrayExpress database (http://www.ebi.ac.uk/arrayexpress/) under Accession Nos E-MTAB-990 and E-MTAB-1026. © 2013 Swiss Institute of Bioinformatics. Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland

VEGAS as a Platform for Facile Directed Evolution in Mammalian Cells

Directed evolution, artificial selection toward designed objectives, is routinely used to develop new molecular tools and therapeutics. Successful directed molecular evolution campaigns repeatedly test diverse sequences with a designed selective pressure. Unicellular organisms and their viral pathogens are exceptional for this purpose and have been used for decades. However, many desirable targets of directed evolution perform poorly or unnaturally in unicellular backgrounds. Here, we present a system for facile directed evolution in mammalian cells. Using the RNA alphavirus Sindbis as a vector for heredity and diversity, we achieved 24-h selection cycles surpassing 10−3 mutations per base. Selection is achieved through genetically actuated sequences internal to the host cell, thus the system's name: viral evolution of genetically actuating sequences, or “VEGAS.” Using VEGAS, we evolve transcription factors, GPCRs, and allosteric nanobodies toward functional signaling endpoints each in less than 1 weeks’ time. © 2019 Elsevier Inc.The VEGAS system is a platform for directed evolution, a method for engineering DNA sequences, in mammalian cells. The system is highly mutagenic, facile, and self-contained, requiring no in vitro handling during evolution cycles. As a result, robust evolution campaigns can be run within the context of a mammalian cell signaling environment. We perform three such campaigns as a proof-of-concept: evolving a transcription factor, a G-protein coupled receptor, and llama-derived nanobodies toward specific in vivo activities. © 2019 Elsevier Inc

Superconductivity in the two dimensional Hubbard Model.

Quasiparticle bands of the two-dimensional Hubbard model are calculated using the Roth two-pole approximation to the one particle Green's function. Excellent agreement is obtained with recent Monte Carlo calculations, including an anomalous volume of the Fermi surface near half-filling, which can possibly be explained in terms of a breakdown of Fermi liquid theory. The calculated bands are very flat around the (pi,0) points of the Brillouin zone in agreement with photoemission measurements of cuprate superconductors. With doping there is a shift in spectral weight from the upper band to the lower band. The Roth method is extended to deal with superconductivity within a four-pole approximation allowing electron-hole mixing. It is shown that triplet p-wave pairing never occurs. Singlet d_{x^2-y^2}-wave pairing is strongly favoured and optimal doping occurs when the van Hove singularity, corresponding to the flat band part, lies at the Fermi level. Nearest neighbour antiferromagnetic correlations play an important role in flattening the bands near the Fermi level and in favouring superconductivity. However the mechanism for superconductivity is a local one, in contrast to spin fluctuation exchange models. For reasonable values of the hopping parameter the transition temperature T_c is in the range 10-100K. The optimum doping delta_c lies between 0.14 and 0.25, depending on the ratio U/t. The gap equation has a BCS-like form and (2*Delta_{max})/(kT_c) ~ 4.Comment: REVTeX, 35 pages, including 19 PostScript figures numbered 1a to 11. Uses epsf.sty (included). Everything in uuencoded gz-compressed .tar file, (self-unpacking, see header). Submitted to Phys. Rev. B (24-2-95

Erratum: VEGAS as a Platform for Facile Directed Evolution in Mammalian Cells (Cell (2019) 178(3) (748–761.e17), (S0092867419306221), (10.1016/j.cell.2019.05.051))

(Cell 178, 748–761.e1–e17; July 25, 2019) In our recent article reporting a platform for directed evolution in mammalian cells, we inadvertently failed to cite a paper that also reports a method for evolving biomolecules in mammalian cells (Berman et al., 2018). We have corrected the online version of our paper to cite this work, and we apologize for the omission. © 2019 Elsevier Inc

TRUPATH, an open-source biosensor platform for interrogating the GPCR transducerome

G-protein-coupled receptors (GPCRs) remain major drug targets, despite our incomplete understanding of how they signal through 16 non-visual G-protein signal transducers (collectively named the transducerome) to exert their actions. To address this gap, we have developed an open-source suite of 14 optimized bioluminescence resonance energy transfer (BRET) Gαβγ biosensors (named TRUPATH) to interrogate the transducerome with single pathway resolution in cells. Generated through exhaustive protein engineering and empirical testing, the TRUPATH suite of Gαβγ biosensors includes the first Gα15 and GαGustducin probes. In head-to-head studies, TRUPATH biosensors outperformed first-generation sensors at multiple GPCRs and in different cell lines. Benchmarking studies with TRUPATH biosensors recapitulated previously documented signaling bias and revealed new coupling preferences for prototypic and understudied GPCRs with potential in vivo relevance. To enable a greater understanding of GPCR molecular pharmacology by the scientific community, we have made TRUPATH biosensors easily accessible as a kit through Addgene

Constraints on Dark Matter Annihilation in Clusters of Galaxies with the Fermi Large Area Telescope

Nearby clusters and groups of galaxies are potentially bright sources of high-energy gamma-ray emission resulting from the pair-annihilation of dark matter particles. However, no significant gamma-ray emission has been detected so far from clusters in the first 11 months of observations with the Fermi Large Area Telescope. We interpret this non-detection in terms of constraints on dark matter particle properties. In particular for leptonic annihilation final states and particle masses greater than ~200 GeV, gamma-ray emission from inverse Compton scattering of CMB photons is expected to dominate the dark matter annihilation signal from clusters, and our gamma-ray limits exclude large regions of the parameter space that would give a good fit to the recent anomalous Pamela and Fermi-LAT electron-positron measurements. We also present constraints on the annihilation of more standard dark matter candidates, such as the lightest neutralino of supersymmetric models. The constraints are particularly strong when including the fact that clusters are known to contain substructure at least on galaxy scales, increasing the expected gamma-ray flux by a factor of ~5 over a smooth-halo assumption. We also explore the effect of uncertainties in cluster dark matter density profiles, finding a systematic uncertainty in the constraints of roughly a factor of two, but similar overall conclusions. In this work, we focus on deriving limits on dark matter models; a more general consideration of the Fermi-LAT data on clusters and clusters as gamma-ray sources is forthcoming.Comment: accepted to JCAP, Corresponding authors: T.E. Jeltema and S. Profumo, minor revisions to be consistent with accepted versio

Measurement of W Polarisation at LEP

The three different helicity states of W bosons produced in the reaction e+ e- -> W+ W- -> l nu q q~ at LEP are studied using leptonic and hadronic W decays. Data at centre-of-mass energies \sqrt s = 183-209 GeV are used to measure the polarisation of W bosons, and its dependence on the W boson production angle. The fraction of longitudinally polarised W bosons is measured to be 0.218 \pm 0.027 \pm 0.016 where the first uncertainty is statistical and the second systematic, in agreement with the Standard Model expectation