Synthetic lethal screening in the mammalian central nervous system identifies Gpx6 as a modulator of Huntington’s disease

Huntington’s disease, the most common inherited neurodegenerative disease, is characterized by a dramatic loss of deep-layer cortical and striatal neurons, as well as morbidity in midlife. Human genetic studies led to the identification of the causative gene, huntingtin. Recent genomic advances have also led to the identification of hundreds of potential interacting partners for huntingtin protein and many hypotheses as to the molecular mechanisms whereby mutant huntingtin leads to cellular dysfunction and death. However, the multitude of possible interacting partners and cellular pathways affected by mutant huntingtin has complicated efforts to understand the etiology of this disease, and to date no curative therapeutic exists. To address the general problem of identifying the disease-phenotype contributing genes from a large number of correlative studies, here we develop a synthetic lethal screening methodology for the mammalian central nervous system, called SLIC, for synthetic lethal in the central nervous system. Applying SLIC to the study of Huntington’s disease, we identify the age-regulated glutathione peroxidase 6 (Gpx6) gene as a modulator of mutant huntingtin toxicity and show that overexpression of Gpx6 can dramatically alleviate both behavioral and molecular phenotypes associated with a mouse model of Huntington’s disease. SLIC can, in principle, be used in the study of any neurodegenerative disease for which a mouse model exists, promising to reveal modulators of neurodegenerative disease in an unbiased fashion, akin to screens in simpler model organisms.National Institute of Neurological Disorders and Stroke (U.S.) (Award R01NS085880)William N. and Bernice E. Bumpus Foundation (Early Career Investigator Innovation Award)JPB FoundationEuropean Molecular Biology Organization (Long-term Fellowship

ZFIRE: The Evolution of the Stellar Mass Tully-Fisher Relation to Redshift 2.0 < Z < 2.5 with MOSFIRE

Using observations made with MOSFIRE on Keck I as part of the ZFIRE survey, we present the stellar mass Tully-Fisher relation at 2.0 < z < 2.5. The sample was drawn from a stellar mass limited, Ks-band selected catalog from ZFOURGE over the CANDELS area in the COSMOS field. We model the shear of the Halpha emission line to derive rotational velocities at 2.2X the scale radius of an exponential disk (V2.2). We correct for the blurring effect of a two-dimensional PSF and the fact that the MOSFIRE PSF is better approximated by a Moffat than a Gaussian, which is more typically assumed for natural seeing. We find for the Tully-Fisher relation at 2.0 < z < 2.5 that logV2.2 =(2.18 +/- 0.051)+(0.193 +/- 0.108)(logM/Msun - 10) and infer an evolution of the zeropoint of Delta M/Msun = -0.25 +/- 0.16 dex or Delta M/Msun = -0.39 +/- 0.21 dex compared to z = 0 when adopting a fixed slope of 0.29 or 1/4.5, respectively. We also derive the alternative kinematic estimator S0.5, with a best-fit relation logS0.5 =(2.06 +/- 0.032)+(0.211 +/- 0.086)(logM/Msun - 10), and infer an evolution of Delta M/Msun= -0.45 +/- 0.13 dex compared to z < 1.2 if we adopt a fixed slope. We investigate and review various systematics, ranging from PSF effects, projection effects, systematics related to stellar mass derivation, selection biases and slope. We find that discrepancies between the various literature values are reduced when taking these into account. Our observations correspond well with the gradual evolution predicted by semi-analytic models.Comment: 21 pages, 14 figures, 1 appendix. Accepted for publication by Apj, February 28, 201

ZFOURGE: Using Composite Spectral Energy Distributions to Characterize Galaxy Populations at 1<z<4

We investigate the properties of galaxies as they shut off star formation over the 4 billion years surrounding peak cosmic star formation. To do this we categorize $\sim7000$ galaxies from $1<z<4$ into $90$ groups based on the shape of their spectral energy distributions (SEDs) and build composite SEDs with $R\sim 50$ resolution. These composite SEDs show a variety of spectral shapes and also show trends in parameters such as color, mass, star formation rate, and emission line equivalent width. Using emission line equivalent widths and strength of the 4000\AA\ break, $D(4000)$, we categorize the composite SEDs into five classes: extreme emission line, star-forming, transitioning, post-starburst, and quiescent galaxies. The transitioning population of galaxies show modest H$\alpha$ emission ($EW_{\rm REST}\sim40$\AA) compared to more typical star-forming composite SEDs at $\log_{10}(M/M_\odot)\sim10.5$ ($EW_{\rm REST}\sim80$\AA). Together with their smaller sizes (3 kpc vs. 4 kpc) and higher S\'ersic indices (2.7 vs. 1.5), this indicates that morphological changes initiate before the cessation of star formation. The transitional group shows a strong increase of over one dex in number density from $z\sim3$ to $z\sim1$, similar to the growth in the quiescent population, while post-starburst galaxies become rarer at $z\lesssim1.5$. We calculate average quenching timescales of 1.6 Gyr at $z\sim1.5$ and 0.9 Gyr at $z\sim2.5$ and conclude that a fast quenching mechanism producing post-starbursts dominated the quenching of galaxies at early times, while a slower process has become more common since $z\sim2$.Comment: Accepted for publication in The Astrophysical Journa

The distribution of satellites around massive galaxies at 1<z<3 in ZFOURGE/CANDELS: dependence on star formation activity

We study the statistical distribution of satellites around star-forming and quiescent central galaxies at 1<z<3 using imaging from the FourStar Galaxy Evolution Survey (ZFOURGE) and the Cosmic Assembly Near-IR Deep Extragalactic Legacy Survey (CANDELS). The deep near-IR data select satellites down to $\log(M/M_\odot)>9$ at z<3. The radial satellite distribution around centrals is consistent with a projected NFW profile. Massive quiescent centrals, $\log(M/M_\odot)>10.78$, have $\sim$2 times the number of satellites compared to star-forming centrals with a significance of 2.7$\sigma$ even after accounting for differences in the centrals' stellar-mass distributions. We find no statistical difference in the satellite distributions of intermediate-mass quiescent and star-forming centrals, $10.48<\log(M/M_\odot)<10.78$. Comparing to the Guo2011 semi-analytic model, the excess number of satellites indicates that quiescent centrals have halo masses 0.3 dex larger than star-forming centrals, even when the stellar-mass distributions are fixed. We use a simple toy model that relates halo mass and quenching, which roughly reproduces the observed quenched fractions and the differences in halo mass between star-forming and quenched galaxies only if galaxies have a quenching probability that increases with halo mass from $\sim$0 for $\log(M_h/M_\odot)\sim$11 to $\sim$1 for $\log(M_h/M_\odot)\sim$13.5. A single halo-mass quenching threshold is unable to reproduce the quiescent fraction and satellite distribution of centrals. Therefore, while halo quenching may be an important mechanism, it is unlikely to be the only factor driving quenching. It remains unclear why a high fraction of centrals remain star-forming even in relatively massive halos.Comment: 19 pages, 17 figures, accepted by ApJ. Information on ZFOURGE can be found at http://zfourge.tamu.ed

Core Circadian Clock Genes Regulate Leukemia Stem Cells in AML

Leukemia stem cells (LSCs) have the capacity to self-renew and propagate disease upon serial transplantation in animal models, and elimination of this cell population is required for curative therapies. Here, we describe a series of pooled, in vivo RNAi screens to identify essential transcription factors (TFs) in a murine model of acute myeloid leukemia (AML) with genetically and phenotypically defined LSCs. These screens reveal the heterodimeric, circadian rhythm TFs Clock and Bmal1 as genes required for the growth of AML cells in vitro and in vivo. Disruption of canonical circadian pathway components produces anti-leukemic effects, including impaired proliferation, enhanced myeloid differentiation, and depletion of LSCs. We find that both normal and malignant hematopoietic cells harbor an intact clock with robust circadian oscillations, and genetic knockout models reveal a leukemia-specific dependence on the pathway. Our findings establish a role for the core circadian clock genes in AML.National Institutes of Health (U.S.) (Grant P01 CA066996)National Institutes of Health (U.S.) (Grant R01 HL082945)National Cancer Institute (U.S.) (Grant P30-CA14051

PIK3CA mutant tumors depend on oxoglutarate dehydrogenase

Oncogenic PIK3CA mutations are found in a significant fraction of human cancers, but therapeutic inhibition of PI3K has only shown limited success in clinical trials. To understand how mutant PIK3CA contributes to cancer cell proliferation, we used genome scale loss-of-function screening in a large number of genomically annotated cancer cell lines. As expected, we found that PIK3CA mutant cancer cells require PIK3CA but also require the expression of the TCA cycle enzyme 2-oxoglutarate dehydrogenase (OGDH). To understand the relationship between oncogenic PIK3CA and OGDH function, we interrogated metabolic requirements and found an increased reliance on glucose metabolism to sustain PIK3CA mutant cell proliferation. Functional metabolic studies revealed that OGDH suppression increased levels of the metabolite 2-oxoglutarate (2OG). We found that this increase in 2OG levels, either by OGDH suppression or exogenous 2OG treatment, resulted in aspartate depletion that was specifically manifested as auxotrophy within PIK3CA mutant cells. Reduced levels of aspartate deregulated the malate-aspartate shuttle, which is important for cytoplasmic NAD + regeneration that sustains rapid glucose breakdown through glycolysis. Consequently, because PIK3CA mutant cells exhibit a profound reliance on glucose metabolism, malate-aspartate shuttle deregulation leads to a specific proliferative block due to the inability to maintain NAD + /NADH homeostasis. Together these observations define a precise metabolic vulnerability imposed by a recurrently mutated oncogene. Keyword: PIK3CA; 2OG; OGDH; TCA cycle; glycolysisDamon Runyon Cancer Research Foundation (HHMI Fellowship

A comprehensive platform for highly multiplexed mammalian functional genetic screens

<p>Abstract</p> <p>Background</p> <p>Genome-wide screening in human and mouse cells using RNA interference and open reading frame over-expression libraries is rapidly becoming a viable experimental approach for many research labs. There are a variety of gene expression modulation libraries commercially available, however, detailed and validated protocols as well as the reagents necessary for deconvolving genome-scale gene screens using these libraries are lacking. As a solution, we designed a comprehensive platform for highly multiplexed functional genetic screens in human, mouse and yeast cells using popular, commercially available gene modulation libraries. The Gene Modulation Array Platform (GMAP) is a single microarray-based detection solution for deconvolution of loss and gain-of-function pooled screens.</p> <p>Results</p> <p>Experiments with specially constructed lentiviral-based plasmid pools containing ~78,000 shRNAs demonstrated that the GMAP is capable of deconvolving genome-wide shRNA "dropout" screens. Further experiments with a larger, ~90,000 shRNA pool demonstrate that equivalent results are obtained from plasmid pools and from genomic DNA derived from lentivirus infected cells. Parallel testing of large shRNA pools using GMAP and next-generation sequencing methods revealed that the two methods provide valid and complementary approaches to deconvolution of genome-wide shRNA screens. Additional experiments demonstrated that GMAP is equivalent to similar microarray-based products when used for deconvolution of open reading frame over-expression screens.</p> <p>Conclusion</p> <p>Herein, we demonstrate four major applications for the GMAP resource, including deconvolution of pooled RNAi screens in cells with at least 90,000 distinct shRNAs. We also provide detailed methodologies for pooled shRNA screen readout using GMAP and compare next-generation sequencing to GMAP (i.e. microarray) based deconvolution methods.</p

The receptor TREML4 amplifies TLR7-mediated signaling during antiviral responses and autoimmunity

The genes and pathways that fine-tune TLR7-mediated innate inflammatory responses remain to be fully elucidated. Using an unbiased genome-scale shRNA screen, we identified the receptor TREML4 as an essential positive regulator of TLR7 signaling. Macrophages from Treml4–/– mice were hyporesponsive to TLR7 agonists and failed to produce type I interferon due to impaired phosphorylation of the transcription factor STAT1 by the MAP kinase p38 and decreased recruitment of MyD88 to TLR7. TREML4 deficiency reduced production of inflammatory cytokines and autoantibodies in SLE-prone MRL/lpr mice and inhibited the antiviral immune response to influenza. Our data identify TREML4 as a positive regulator of TLR7 signaling and provide insight into the molecular mechanisms that control antiviral immunity and the development of autoimmunity

THE SFR– M * RELATION AND EMPIRICAL STAR FORMATION HISTORIES FROM ZFOURGE AT 0.5 < z < 4

We explore star formation histories (SFHs) of galaxies based on the evolution of the star formation rate stellar mass relation (SFR-M∗). Using data from the FourStar Galaxy Evolution Survey (ZFOURGE) in combination with far-IR imaging from the Spitzer and Herschel observatories we measure the SFR-M∗ relation at 0.5 &lt; z &lt; 4. Similar to recent works we find that the average infrared spectral energy distributions of galaxies are roughly consistent with a single infrared template across a broad range of redshifts and stellar masses, with evidence for only weak deviations. We find that the SFR-M∗ relation is not consistent with a single power law of the form at any redshift; it has a power law slope of α ∼ 1 at low masses, and becomes shallower above a turnover mass (M0) that ranges from 109.5 to 1010.8 M⊙, with evidence that M0 increases with redshift. We compare our measurements to results from state-of-the-art cosmological simulations, and find general agreement in the slope of the SFR-M∗ relation albeit with systematic offsets. We use the evolving SFR-M∗ sequence to generate SFHs, finding that typical SFRs of individual galaxies rise at early times and decline after reaching a peak. This peak occurs earlier for more massive galaxies. We integrate these SFHs to generate mass growth histories and compare to the implied mass growth from the evolution of the stellar mass function (SMF). We find that these two estimates are in broad qualitative agreement, but that there is room for improvement at a more detailed level. At early times the SFHs suggest mass growth rates that are as much as 10× higher than inferred from the SMF. However, at later times the SFHs under-predict the inferred evolution, as is expected in the case of additional growth due to mergers