On the extent and role of the small proteome in the parasitic eukaryote Trypanosoma brucei

Background: Although technical advances in genomics and proteomics research have yielded a better understanding of the coding capacity of a genome, one major challenge remaining is the identification of all expressed proteins, especially those less than 100 amino acids in length. Such information can be particularly relevant to human pathogens, such as Trypanosoma brucei, the causative agent of African trypanosomiasis, since it will provide further insight into the parasite biology and life cycle. Results: Starting with 993 T. brucei transcripts, previously shown by RNA-Sequencing not to coincide with annotated coding sequences (CDS), homology searches revealed that 173 predicted short open reading frames in these transcripts are conserved across kinetoplastids with 13 also conserved in representative eukaryotes. Mining mass spectrometry data sets revealed 42 transcripts encoding at least one matching peptide. RNAi-induced down-regulation of these 42 transcripts revealed seven to be essential in insect-form trypanosomes with two also required for the bloodstream life cycle stage. To validate the specificity of the RNAi results, each lethal phenotype was rescued by co-expressing an RNAi-resistant construct of each corresponding CDS. These previously non-annotated essential small proteins localized to a variety of cell compartments, including the cell surface, mitochondria, nucleus and cytoplasm, inferring the diverse biological roles they are likely to play in T. brucei. We also provide evidence that one of these small proteins is required for replicating the kinetoplast (mitochondrial) DNA. Conclusions: Our studies highlight the presence and significance of small proteins in a protist and expose potential new targets to block the survival of trypanosomes in the insect vector and/or the mammalian host

Cell type-specific expression and localization of cytochrome P450 isoforms in tridimensional aggregating rat brain cell cultures.

Within the Predict-IV FP7 project a strategy for measurement of in vitro biokinetics was developed, requiring the characterization of the cellular model used, especially regarding biotransformation, which frequently depends on cytochrome P450 (CYP) activity. The extrahepatic in situ CYP-mediated metabolism is especially relevant in target organ toxicity. In this study, the constitutive mRNA levels and protein localization of different CYP isoforms were investigated in 3D aggregating brain cell cultures. CYP1A1, CYP2B1/B2, CYP2D2/4, CYP2E1 and CYP3A were expressed; CYP1A1 and 2B1 represented almost 80% of the total mRNA content. Double-immunolabeling revealed their presence in astrocytes, in neurons, and to a minor extent in oligodendrocytes, confirming the cell-specific localization of CYPs in the brain. These results together with the recently reported formation of an amiodarone metabolite following repeated exposure suggest that this cell culture system possesses some metabolic potential, most likely contributing to its high performance in neurotoxicological studies and support the use of this model in studying brain neurotoxicity involving mechanisms of toxication/detoxication

Cytochrome oxidase subunit VI of Trypanosoma brucei is imported without a cleaved presequence and is developmentally regulated at both RNA and protein levels

Mitochondrial respiration in the African trypanosome undergoes dramatic developmental stage regulation. This requires co-ordinated control of components encoded by both the nuclear genome and the kinetoplast, the unusual mitochondrial genome of these parasites. As a model for understanding the co-ordination of these genomes, we have examined the regulation and mitochondrial import of a nuclear-encoded component of the cytochrome oxidase complex, cytochrome oxidase subunit VI (COXVI). By generating transgenic trypanosomes expressing intact or mutant forms of this protein, we demonstrate that COXVI is not imported using a conventional cleaved presequence and show that sequences at the N-terminus of the protein are necessary for correct mitochondrial sorting. Analyses of endogenous and transgenic COXVI mRNA and protein expression in parasites undergoing developmental stage differentiation demonstrates a temporal order of control involving regulation in the abundance of, first, mRNA and then protein. This represents the first dissection of the regulation and import of a nuclear-encoded protein into the cytochrome oxidase complex in these organisms, which were among the earliest eukaryotes to possess a mitochondrion

Land and cryosphere products from Suomi NPP VIIRS: overview and status

[1] The Visible Infrared Imaging Radiometer Suite (VIIRS) instrument was launched in October 2011 as part of the Suomi National Polar-Orbiting Partnership (S-NPP). The VIIRS instrument was designed to improve upon the capabilities of the operational Advanced Very High Resolution Radiometer and provide observation continuity with NASA's Earth Observing System's Moderate Resolution Imaging Spectroradiometer (MODIS). Since the VIIRS first-light images were received in November 2011, NASA- and NOAA-funded scientists have been working to evaluate the instrument performance and generate land and cryosphere products to meet the needs of the NOAA operational users and the NASA science community. NOAA's focus has been on refining a suite of operational products known as Environmental Data Records (EDRs), which were developed according to project specifications under the National Polar-Orbiting Environmental Satellite System. The NASA S-NPP Science Team has focused on evaluating the EDRs for science use, developing and testing additional products to meet science data needs, and providing MODIS data product continuity. This paper presents to-date findings of the NASA Science Team's evaluation of the VIIRS land and cryosphere EDRs, specifically Surface Reflectance, Land Surface Temperature, Surface Albedo, Vegetation Indices, Surface Type, Active Fires, Snow Cover, Ice Surface Temperature, and Sea Ice Characterization. The study concludes that, for MODIS data product continuity and earth system science, an enhanced suite of land and cryosphere products and associated data system capabilities are needed beyond the EDRs currently available from the VIIRS

Workgroup Report: Incorporating In Vitro Alternative Methods for Developmental Neurotoxicity into International Hazard and Risk Assessment Strategies

This is the report of the first workshop on Incorporating In Vitro Alternative Methods for Developmental Neurotoxicity (DNT) Testing into International Hazard and Risk Assessment Strategies, held in Ispra, Italy, on 19–21 April 2005. The workshop was hosted by the European Centre for the Validation of Alternative Methods (ECVAM) and jointly organized by ECVAM, the European Chemical Industry Council, and the Johns Hopkins University Center for Alternatives to Animal Testing. The primary aim of the workshop was to identify and catalog potential methods that could be used to assess how data from in vitro alternative methods could help to predict and identify DNT hazards. Working groups focused on two different aspects: a) details on the science available in the field of DNT, including discussions on the models available to capture the critical DNT mechanisms and processes, and b) policy and strategy aspects to assess the integration of alternative methods in a regulatory framework. This report summarizes these discussions and details the recommendations and priorities for future work

Quality of life after gamma knife radiosurgery treatment in patients with a vestibular schwannoma: the patient’s perspective

This study evaluates the impact of gamma knife radiosurgery (GKRS) on the quality of life (QOL) of patients with a sporadic vestibular schwannoma (VS). This study pertains to 108 VS patients who had GKRS in the years 2003 through 2007. Two different QOL questionnaires were used: medical outcome study short form 36 (SF36) and Glasgow benefit inventory (GBI). Radiosurgery was performed using a Leksell 4C gamma knife. The results of the QOL questionnaires in relation to prospectively and retrospectively gathered data of the VS patients treated by GKRS. Eventually, 97 patients could be included in the study. Their mean tumor size was 17 mm (range 6–39 mm); the mean maximum dose on the tumor was 19.9 Gy (range 16–25.5 Gy) and the mean marginal dose on the tumor was 11.1 (range 9.3–12.5 Gy). SF36 scores showed results comparable to those for a normal Dutch population. GBI showed a marginal decline in QOL. No correlation was found between QOL and gender, age, tumor size, or radiation dose. Increased audiovestibular symptoms after GKRS were correlated with a decreased GBI score, and decreased symptoms were correlated with a higher QOL post-GKRS. In this study shows that GKRS for VS has little impact on the general QOL of the VS patient. However, there is a wide range in individual QOL results. Individual QOL was influenced by the audiovestibular symptoms. No predictive patient, tumor, or treatment factors for QOL outcome after GKRS could be determined. Comparison with microsurgery is difficult because of intra group variability

ESPRESSO: The next European exoplanet hunter

The acronym ESPRESSO stems for Echelle SPectrograph for Rocky Exoplanets and Stable Spectroscopic Observations; this instrument will be the next VLT high resolution spectrograph. The spectrograph will be installed at the Combined-Coud\'e Laboratory of the VLT and linked to the four 8.2 m Unit Telescopes (UT) through four optical Coud\'e trains. ESPRESSO will combine efficiency and extreme spectroscopic precision. ESPRESSO is foreseen to achieve a gain of two magnitudes with respect to its predecessor HARPS, and to improve the instrumental radial-velocity precision to reach the 10 cm/s level. It can be operated either with a single UT or with up to four UTs, enabling an additional gain in the latter mode. The incoherent combination of four telescopes and the extreme precision requirements called for many innovative design solutions while ensuring the technical heritage of the successful HARPS experience. ESPRESSO will allow to explore new frontiers in most domains of astrophysics that require precision and sensitivity. The main scientific drivers are the search and characterization of rocky exoplanets in the habitable zone of quiet, nearby G to M-dwarfs and the analysis of the variability of fundamental physical constants. The project passed the final design review in May 2013 and entered the manufacturing phase. ESPRESSO will be installed at the Paranal Observatory in 2016 and its operation is planned to start by the end of the same year.Comment: 12 pages, figures included, accepted for publication in Astron. Nach

A major genetic locus in <i>Trypanosoma brucei</i> is a determinant of host pathology

The progression and variation of pathology during infections can be due to components from both host or pathogen, and/or the interaction between them. The influence of host genetic variation on disease pathology during infections with trypanosomes has been well studied in recent years, but the role of parasite genetic variation has not been extensively studied. We have shown that there is parasite strain-specific variation in the level of splenomegaly and hepatomegaly in infected mice and used a forward genetic approach to identify the parasite loci that determine this variation. This approach allowed us to dissect and identify the parasite loci that determine the complex phenotypes induced by infection. Using the available trypanosome genetic map, a major quantitative trait locus (QTL) was identified on T. brucei chromosome 3 (LOD = 7.2) that accounted for approximately two thirds of the variance observed in each of two correlated phenotypes, splenomegaly and hepatomegaly, in the infected mice (named &lt;i&gt;TbOrg1&lt;/i&gt;). In addition, a second locus was identified that contributed to splenomegaly, hepatomegaly and reticulocytosis (&lt;i&gt;TbOrg2&lt;/i&gt;). This is the first use of quantitative trait locus mapping in a diploid protozoan and shows that there are trypanosome genes that directly contribute to the progression of pathology during infections and, therefore, that parasite genetic variation can be a critical factor in disease outcome. The identification of parasite loci is a first step towards identifying the genes that are responsible for these important traits and shows the power of genetic analysis as a tool for dissecting complex quantitative phenotypic traits

HARPS3 for a roboticized Isaac Newton telescope

This is the author accepted manuscript. The final version is available from the publisher via the DOI in this record.We present a description of a new instrument development, HARPS3, planned to be installed on an upgraded and roboticized Isaac Newton Telescope by end-2018. HARPS3 will be a high resolution (R = 115,000) echelle spectrograph with a wavelength range from 380-690 nm. It is being built as part of the Terra Hunting Experiment - a future 10 year radial velocity measurement programme to discover Earth-like exoplanets. The instrument design is based on the successful HARPS spectrograph on the 3.6m ESO telescope and HARPS-N on the TNG telescope. The main changes to the design in HARPS3 will be: a customised fibre adapter at the Cassegrain focus providing a stabilised beam feed and on-sky fibre diameter ~ 1.4 arcsec, the implementation of a new continuous flow cryostat to keep the CCD temperature very stable, detailed characterisation of the HARPS3 CCD to map the effective pixel positions and thus provide an improved accuracy wavelength solution, an optimised integrated polarimeter and the instrument integrated into a robotic operation. The robotic operation will optimise our programme which requires our target stars to be measured on a nightly basis. We present an overview of the entire project, including a description of our anticipated robotic operation.R.H. acknowledges the Science and Technologies Facilities Council (STFC) for his PhD studentship award (2015).J.I.G.H. acknowledges financial support from the Spanish Ministry of Economy and Competitiveness (MINECO) under the 2013 Ram´on y Cajal program MINECO RYC-2013-14875.J.I.G.H., R.R., and S.S.T. also acknowledge the Spanish ministry project MINECO AYA2014-56359-P.NP and ES are grateful to Knut and Alice Wallenberg Foundation for a generous support of the Swedish contribution to the THE project.AD acknowledges the support from Russian Foundation for Basic Research as part of research grant 15-52-12371