RNA Unwinding by the Trf4/Air2/Mtr4 Polyadenylation (TRAMP) Complex

Many RNA-processing events in the cell nucleus involve the Trf4/Air2/Mtr4 polyadenylation (TRAMP) complex, which contains the poly(A) polymerase Trf4p, the Zn-knuckle protein Air2p, and the RNA helicase Mtr4p. TRAMP polyadenylates RNAs designated for processing by the nuclear exosome. In addition, TRAMP functions as an exosome cofactor during RNA degradation, and it has been speculated that this role involves disruption of RNA secondary structure. However, it is unknown whether TRAMP displays RNA unwinding activity. It is also not clear how unwinding would be coordinated with polyadenylation and the function of the RNA helicase Mtr4p in modulating poly(A) addition. Here, we show that TRAMP robustly unwinds RNA duplexes. The unwinding activity of Mtr4p is significantly stimulated by Trf4p/Air2p, but the stimulation of Mtr4p does not depend on ongoing polyadenylation. Nonetheless, polyadenylation enables TRAMP to unwind RNA substrates that it otherwise cannot separate. Moreover, TRAMP displays optimal unwinding activity on substrates with a minimal Mtr4p binding site comprised of adenylates. Our results suggest a model for coordination between unwinding and polyadenylation activities by TRAMP that reveals remarkable synergy between helicase and poly(A) polymerase

Degradation of Hypomodified tRNA\u3csub\u3ei\u3c/sub\u3e\u3csup\u3eMet\u3c/sup\u3e in vivo Involves RNA-dependent ATPase Activity of the DExH Helicase Mtr4p

Effective turnover of many incorrectly processed RNAs in yeast, including hypomodified tRNAi Met, requires the TRAMP complex, which appends a short poly(A) tail to RNA designated for decay. The poly(A) tail stimulates degradation by the exosome. The TRAMP complex contains the poly(A) polymerase Trf4p, the RNA-binding protein Air2p, and the DExH RNA helicase Mtr4p. The role of Mtr4p in RNA degradation processes involving the TRAMP complex has been unclear. Here we show through a genetic analysis that MTR4 is required for degradation but not for polyadenylation of hypomodified tRNAi Met. A suppressor of the trm6-504 mutation in the tRNA m1A58 methyltransferase (Trm6p/Trm61p), which causes a reduced level of tRNAi Met, was mapped to MTR4. This mtr4-20 mutation changed a single amino acid in the conserved helicase motif VI of Mtr4p. The mutation stabilizes hypomodified tRNAi Met in vivo but has no effect on TRAMP complex stability or polyadenylation activity in vivo or in vitro. We further show that purified recombinant Mtr4p displays RNA-dependent ATPase activity and unwinds RNA duplexes with a 3′-to-5′ polarity in an ATP-dependent fashion. Unwinding and RNA-stimulated ATPase activities are strongly reduced in the recombinant mutant Mtr4-20p, suggesting that these activities of Mtr4p are critical for degradation of polyadenylated hypomodified tRNAi Met

Probing Convolutional Neural Networks for Event Reconstruction in {\gamma}-Ray Astronomy with Cherenkov Telescopes

A dramatic progress in the field of computer vision has been made in recent years by applying deep learning techniques. State-of-the-art performance in image recognition is thereby reached with Convolutional Neural Networks (CNNs). CNNs are a powerful class of artificial neural networks, characterized by requiring fewer connections and free parameters than traditional neural networks and exploiting spatial symmetries in the input data. Moreover, CNNs have the ability to automatically extract general characteristic features from data sets and create abstract data representations which can perform very robust predictions. This suggests that experiments using Cherenkov telescopes could harness these powerful machine learning algorithms to improve the analysis of particle-induced air-showers, where the properties of primary shower particles are reconstructed from shower images recorded by the telescopes. In this work, we present initial results of a CNN-based analysis for background rejection and shower reconstruction, utilizing simulation data from the H.E.S.S. experiment. We concentrate on supervised training methods and outline the influence of image sampling on the performance of the CNN-model predictions.Comment: 8 pages, 4 figures, Proceedings of the 35th International Cosmic Ray Conference (ICRC 2017), Busan, Kore

Stock-catch analysis of carp recreational fisheries in Czech reservoirs: Insights into fish survival, and impact of extreme events

In culture-based fisheries, managers strive for high stocking efficiency, the ratio between the total weight of caught and stocked fish. Here we present a new time series approach to examine the dependence of reported anglers. catches on stocking and external events, using data on carp ('Cyprinus carpio' L.) from 14 reservoirs in the Czech Republic. Average stocking efficiency varied between 0.25 and 2.2, with values close to unity in most reservoirs. The lowest efficiencies occurred in three reservoirs receiving cold hypoxic water from a large upstream reservoir, while the highest efficiencis were found in two shallow, highly productive reservoirs. Analyses further indicate that stocked carp are typically caught during the year of release or the year after; but also that the mean time lag- between stocking and capture increases with reservoir area. External events can be important: major floods in the years 2002 and 2006 were in many cases followed by large, up to 10-fold, increases in catches in subsequent years; we attribute the surplus catch to carp washed down from upstream aquaculture and river stretches. In contrast, the "Velvet REvolution" (demise of the communist regime in 1989) had no discernible effect on catches in subsequent years. In conclusion, the proposed method can simultaneously estimate the likely mean survival time of stocked carp and identify the impact of major environmental and societal events on recreational fisheries. The approach thus sheds light on the performance of current stocking practices at individual reservoirs, and could be used to monitor and improve stocking strategies and management of culture-based recreational fisheries

TARGET: A Digitizing And Trigger ASIC For The Cherenkov Telescope Array

The future ground-based gamma-ray observatory Cherenkov Telescope Array (CTA) will feature multiple types of imaging atmospheric Cherenkov telescopes, each with thousands of pixels. To be affordable, camera concepts for these telescopes have to feature low cost per channel and at the same time meet the requirements for CTA in order to achieve the desired scientific goals. We present the concept of the TeV Array Readout Electronics with GSa/s sampling and Event Trigger (TARGET) Application Specific Circuit (ASIC), envisaged to be used in the cameras of various CTA telescopes, e.g. the Gamma-ray Cherenkov Telescope (GCT), a proposed 2-Mirror Small-Sized Telescope, and the Schwarzschild-Couder Telescope (SCT), a proposed Medium-Sized Telescope. In the latest version of this readout concept the sampling and trigger parts are split into dedicated ASICs, TARGET C and T5TEA, both providing 16 parallel input channels. TARGET C features a tunable sampling rate (usually 1 GSa/s), a 16k sample deep buffer for each channel and on-demand digitization and transmission of waveforms with typical spans of ~100 ns. The trigger ASIC, T5TEA, provides 4 low voltage differential signal (LVDS) trigger outputs and can generate a pedestal voltage independently for each channel. Trigger signals are generated by T5TEA based on the analog sum of the input in four independent groups of four adjacent channels and compared to a threshold set by the user. Thus, T5TEA generates four LVDS trigger outputs, as well as 16 pedestal voltages fed to TARGET C independently for each channel. We show preliminary results of the characterization and testing of TARGET C and T5TEA.Comment: 6 pages, 8 figures, Proceedings of the 6th International Symposium on High-Energy Gamma-Ray Astronomy (Gamma2016

Rodent Aβ Modulates the Solubility and Distribution of Amyloid Deposits in Transgenic Mice

The amino acid sequence of amyloid precursor protein (APP) is highly conserved, and age-related Abeta aggregates have been described in a variety of vertebrate animals, with the notable exception of mice and rats. Three amino acid substitutions distinguish mouse and human Abeta that might contribute to their differing properties in vivo. To examine the amyloidogenic potential of mouse Abeta, we studied several lines of transgenic mice overexpressing wild-type mouse amyloid precursor protein (moAPP) either alone or in conjunction with mutant PS1 (PS1dE9). Neither overexpression of moAPP alone nor co-expression with PS1dE9 caused mice to develop Alzheimer-type amyloid pathology by 24 months of age. We further tested whether mouse Abeta could accelerate the deposition of human Abeta by crossing the moAPP transgenic mice to a bigenic line expressing human APPswe with PS1dE9. The triple transgenic animals (moAPP x APPswe/PS1dE9) produced 20% more Abeta but formed amyloid deposits no faster and to no greater extent than APPswe/PS1dE9 siblings. Instead, the additional mouse Abeta increased the detergent solubility of accumulated amyloid and exacerbated amyloid deposition in the vasculature. These findings suggest that, although mouse Abeta does not influence the rate of amyloid formation, the incorporation of Abeta peptides with differing sequences alters the solubility and localization of the resulting aggregates

The helicase HAGE prevents interferon-a-induced PML expression in ABCB5+ malignant melanoma-initiating cells by promoting the expression of SOCS1

The tumour suppressor PML (promyelocytic leukaemia protein) regulates several cellular pathways involving cell growth, apoptosis, differentiation and senescence. PML also has an important role in the regulation of stem cell proliferation and differentiation. Here, we show the involvement of the helicase HAGE in the transcriptional repression of PML expression in ABCB5 + malignant melanoma-initiating cells (ABCB5 + MMICs), a population of cancer stem cells which are responsible for melanoma growth, progression and resistance to drug-based therapy. HAGE prevents PML gene expression by inhibiting the activation of the JAK-STAT (janus kinase-signal transducers and activators of transcription) pathway in a mechanism which implicates the suppressor of cytokine signalling 1 (SOCS1). Knockdown of HAGE led to a significant decrease in SOCS1 protein expression, activation of the JAK-STAT signalling cascade and a consequent increase of PML expression. To confirm that the reduction in SOCS1 expression was dependent on the HAGE helicase activity, we showed that SOCS1, effectively silenced by small interfering RNA, could be rescued by re-introduction of HAGE into cells lacking HAGE. Furthermore, we provide a mechanism by which HAGE promotes SOCS1 mRNA unwinding and protein expression in vitro

The helicase Ded1p controls use of near-cognate translation initiation codons in 5' UTRs.

The conserved and essential DEAD-box RNA helicase Ded1p from yeast and its mammalian orthologue DDX3 are critical for the initiation of translation1. Mutations in DDX3 are linked to tumorigenesis2-4 and intellectual disability5, and the enzyme is targeted by a range of viruses6. How Ded1p and its orthologues engage RNAs during the initiation of translation is unknown. Here we show, by integrating transcriptome-wide analyses of translation, RNA structure and Ded1p-RNA binding, that the effects of Ded1p on the initiation of translation are connected to near-cognate initiation codons in 5' untranslated regions. Ded1p associates with the translation pre-initiation complex at the mRNA entry channel and repressing the activity of Ded1p leads to the accumulation of RNA structure in 5' untranslated regions, the initiation of translation from near-cognate start codons immediately upstream of these structures and decreased protein synthesis from the corresponding main open reading frames. The data reveal a program for the regulation of translation that links Ded1p, the activation of near-cognate start codons and mRNA structure. This program has a role in meiosis, in which a marked decrease in the levels of Ded1p is accompanied by the activation of the alternative translation initiation sites that are seen when the activity of Ded1p is repressed. Our observations indicate that Ded1p affects translation initiation by controlling the use of near-cognate initiation codons that are proximal to mRNA structure in 5' untranslated regions

Environmental Enrichment Mitigates Cognitive Deficits in a Mouse Model of Alzheimer's Disease

Epidemiological studies suggest that individuals with greater education or more cognitively demanding occupations have diminished risk of developing dementia. We wanted to test whether this effect could be recapitulated in rodents using environmental enrichment, a paradigm well documented to attenuate behavioral deficits induced by various pathological insults. Here, we demonstrate that learning and memory deficits observed in a transgenic mouse model of Alzheimer's disease can be ameliorated by enrichment. Female transgenic mice overexpressing amyloid precursor protein and/or presenilin-1 and nontransgenic controls were placed into enriched or standard cages at 2 months of age and tested for cognitive behavior after 6 months of differential housing. Enrichment significantly improved performance of all genotypes in the radial water maze and in the classic and repeated-reversal versions of the Morris water maze. However, enrichment did not benefit all genotypes equally. Mice overproducing amyloid-β (Aβ), particularly those with amyloid deposits, showed weaker memory for the platform location in the classic Morris water maze and learned new platform positions in the repeated-reversals task less quickly than their nontransgenic cagemates. Nonetheless, enrichment normalized the performance of Aβ-overproducing mice to the level of standard-housed nontransgenic mice. Moreover, this functional preservation occurred despite increased neuritic plaque burden in the hippocampus of double-transgenic animals and elevated steady-state Aβ levels, because both endogenous and transgene-derived Aβ are increased in enriched animals. These results demonstrate that the generation of Aβ in vivo and its impact on the function of the nervous system can be strongly modulated by environmental factors