A04 The role of splicing factor SRSF6 in incomplete splicing of the HTT transcript

Background Huntington’s disease (HD) is caused by an expanded CAG repeat in exon 1 of the HTT gene. In models of HD, an expanded CAG repeat in HTT causes premature termination of HTT RNA during transcription; this occurs by a process called incomplete splicing. Incompletely spliced HTT (HTTexon1) includes exon 1 of the coding region of HTT, as well as a 5’ region of intron 1, which is non-coding. HTTexon1 encodes a truncated exon 1 HTT protein, which is implicated in HD pathogenesis. Although the precise RNA processing mechanism of Httexon1 is unknown, splicing factor SRSF6 has been shown to co-precipitate with transcripts containing Htt intron 1 in HD mice. Aim To elucidate the role of splicing factor SRSF6 in incomplete splicing of Htt in HD mice. Methods Heterozygous Srsf6 knock-out (KO) mice (Srsf6±) were generated by CRISPR/Cas9. Characterisation of Srsf6± mice was undertaken by quantitative RT-PCR and western blotting. Viability of homozygous Srsf6 KO (Srsf6-/-) mice was examined by inbreeding of Srsf6± mice. To assess the modulation of incomplete splicing by decreasing SRSF6, Srsf6± mice were bred to HD knock in mice (zQ175) and tissues were analysed. Levels of Httexon1 were measured by Quantigene, a gene expression assay. Results Srsf6-/- homozygotes were embryonic lethal, limiting us to the use of Srsf6± mice only. In Srsf6± heterozygotes, Srsf6 mRNA was decreased by 50% in brain and peripheral regions, and SRSF6 protein was decreased by 70% in brain compared to wild type mice. However, heterozygosity for Srsf6 knock out did not modulate the level on incomplete splicing in zQ175 mice. Conclusion Ablation of a single Srsf6 allele did not reduce levels of incomplete splicing in HD mice and therefore, further Srsf6 knock down may be required. Accordingly, mouse embryonic fibroblasts (MEFs) have been generated and will be used to measure Httexon1 levels after further Srsf6 knockdown by RNA interference. This work is supported by the CHDI foundation

Modelling the natural history of Huntington's disease progression.

BACKGROUND: The lack of reliable biomarkers to track disease progression is a major problem in clinical research of chronic neurological disorders. Using Huntington's disease (HD) as an example, we describe a novel approach to model HD and show that the progression of a neurological disorder can be predicted for individual patients. METHODS: Starting with an initial cohort of 343 patients with HD that we have followed since 1995, we used data from 68 patients that satisfied our filtering criteria to model disease progression, based on the Unified Huntington's Disease Rating Scale (UHDRS), a measure that is routinely used in HD clinics worldwide. RESULTS: Our model was validated by: (A) extrapolating our equation to model the age of disease onset, (B) testing it on a second patient data set by loosening our filtering criteria, (C) cross-validating with a repeated random subsampling approach and (D) holdout validating with the latest clinical assessment data from the same cohort of patients. With UHDRS scores from the past four clinical visits (over a minimum span of 2 years), our model predicts disease progression of individual patients over the next 2 years with an accuracy of 89-91%. We have also provided evidence that patients with similar baseline clinical profiles can exhibit very different trajectories of disease progression. CONCLUSIONS: This new model therefore has important implications for HD research, most obviously in the development of potential disease-modifying therapies. We believe that a similar approach can also be adapted to model disease progression in other chronic neurological disorders.This study was supported by the Cotswold Trust, the Rosetrees Trust, donations to the Huntington’s disease clinic in the John van Geest Centre for Brain Repair, and NIHR award of the Biomedical Research Centre - Cambridge University NHS Foundation Trust. This project was also supported by EPSRC through projects EP/I03210X/1 and EP/G066477/1.This article has been accepted for publication in Journal of Neurology, Neurosurgery, and Psychiatry, following peer review. The definitive copyedited, typeset version J Neurol Neurosurg Psychiatry doi:10.1136/jnnp-2014-308153 is available online at: http://jnnp.bmj.com/content/early/2014/12/16/jnnp-2014-308153.long

Extensive Expression Analysis of Htt Transcripts in Brain Regions from the zQ175 HD Mouse Model Using a QuantiGene Multiplex Assay

Huntington’s disease (HD) is an inherited neurodegenerative disorder caused by a CAG repeat expansion within exon 1 of the huntingtin (HTT) gene. HTT mRNA contains 67 exons and does not always splice between exon 1 and exon 2 leading to the production of a small polyadenylated HTTexon1 transcript, and the full-length HTT mRNA has three 3′UTR isoforms. We have developed a QuantiGene multiplex panel for the simultaneous detection of all of these mouse Htt transcripts directly from tissue lysates and demonstrate that this can replace the more work-intensive Taqman qPCR assays. We have applied this to the analysis of brain regions from the zQ175 HD mouse model and wild type littermates at two months of age. We show that the incomplete splicing of Htt occurs throughout the brain and confirm that this originates from the mutant and not endogenous Htt allele. Given that HTTexon1 encodes the highly pathogenic exon 1 HTT protein, it is essential that the levels of all Htt transcripts can be monitored when evaluating HTT lowering approaches. Our QuantiGene panel will allow the rapid comparative assessment of all Htt transcripts in cell lysates and mouse tissues without the need to first extract RNA

The heat shock response, determined by QuantiGene multiplex, is impaired in HD mouse models and not caused by HSF1 reduction.

Huntington's disease (HD) is a devastating neurodegenerative disorder, caused by a CAG/polyglutamine repeat expansion, that results in the aggregation of the huntingtin protein, culminating in the deposition of inclusion bodies in HD patient brains. We have previously shown that the heat shock response becomes impaired with disease progression in mouse models of HD. The disruption of this inducible arm of the proteostasis network is likely to exacerbate the pathogenesis of this protein-folding disease. To allow a rapid and more comprehensive analysis of the heat shock response, we have developed, and validated, a 16-plex QuantiGene assay that allows the expression of Hsf1 and nine heat shock genes, to be measured directly, and simultaneously, from mouse tissue. We used this QuantiGene assay to show that, following pharmacological activation in vivo, the heat shock response impairment in tibialis anterior, brain hemispheres and striatum was comparable between zQ175 and R6/2 mice. In contrast, although a heat shock impairment could be detected in R6/2 cortex, this was not apparent in the cortex from zQ175 mice. Whilst the mechanism underlying this impairment remains unknown, our data indicated that it is not caused by a reduction in HSF1 levels, as had been reported

GP-SUM. Gaussian Processes Filtering of non-Gaussian Beliefs

This work studies the problem of stochastic dynamic filtering and state propagation with complex beliefs. The main contribution is GP-SUM, a filtering algorithm tailored to dynamic systems and observation models expressed as Gaussian Processes (GP), and to states represented as a weighted sum of Gaussians. The key attribute of GP-SUM is that it does not rely on linearizations of the dynamic or observation models, or on unimodal Gaussian approximations of the belief, hence enables tracking complex state distributions. The algorithm can be seen as a combination of a sampling-based filter with a probabilistic Bayes filter. On the one hand, GP-SUM operates by sampling the state distribution and propagating each sample through the dynamic system and observation models. On the other hand, it achieves effective sampling and accurate probabilistic propagation by relying on the GP form of the system, and the sum-of-Gaussian form of the belief. We show that GP-SUM outperforms several GP-Bayes and Particle Filters on a standard benchmark. We also demonstrate its use in a pushing task, predicting with experimental accuracy the naturally occurring non-Gaussian distributions.Comment: WAFR 2018, 16 pages, 7 figure

Spin chirality on a two-dimensional frustrated lattice

The collective behavior of interacting magnetic moments can be strongly influenced by the topology of the underlying lattice. In geometrically frustrated spin systems, interesting chiral correlations may develop that are related to the spin arrangement on triangular plaquettes. We report a study of the spin chirality on a two-dimensional geometrically frustrated lattice. Our new chemical synthesis methods allow us to produce large single crystal samples of KFe3(OH)6(SO4)2, an ideal Kagome lattice antiferromagnet. Combined thermodynamic and neutron scattering measurements reveal that the phase transition to the ordered ground-state is unusual. At low temperatures, application of a magnetic field induces a transition between states with different non-trivial spin-textures.Comment: 7 pages, 4 figure

The effect of ultrasound pretreatment on some selected physicochemical properties of black cumin (Nigella Sativa)

Background In the present study, the effects of ultrasound pretreatment parameters including irradiation time and power on the quantity of the extracted phenolic compounds quantity as well as on some selected physicochemical properties of the extracted oils including oil extraction efficiency, acidity and peroxide values, color, and refractive index of the extracted oil of black cumin seeds with the use of cold press have been studied. Methods For each parameter, three different levels (30, 60, and 90 W) for the ultrasound power and (30, 45, and 60 min) and for the ultrasound irradiation time were studied. Each experiment was performed in three replications. Results The achieved results revealed that, with enhancements in the applied ultrasound power, the oil extraction efficiency, acidity value, total phenolic content, peroxide value, and color parameters increased significantly (P 0.05). Conclusions In summary, it could be mentioned that the application of ultrasound pretreatment in the oil extraction might improve the oil extraction efficiency, the extracted oil’s quality, and the extracted phenolic compounds content.info:eu-repo/semantics/publishedVersio

The Thermal Design, Characterization, and Performance of the SPIDER Long-Duration Balloon Cryostat

We describe the SPIDER flight cryostat, which is designed to cool six millimeter-wavelength telescopes during an Antarctic long-duration balloon flight. The cryostat, one of the largest to have flown on a stratospheric payload, uses liquid helium-4 to deliver cooling power to stages at 4.2 and 1.6 K. Stainless steel capillaries facilitate a high flow impedance connection between the main liquid helium tank and a smaller superfluid tank, allowing the latter to operate at 1.6 K as long as there is liquid in the 4.2 K main tank. Each telescope houses a closed cycle helium-3 adsorption refrigerator that further cools the focal planes down to 300 mK. Liquid helium vapor from the main tank is routed through heat exchangers that cool radiation shields, providing negative thermal feedback. The system performed successfully during a 17 day flight in the 2014-2015 Antarctic summer. The cryostat had a total hold time of 16.8 days, with 15.9 days occurring during flight.Comment: 15 pgs, 17 fig

Contact Manifolds, Contact Instantons, and Twistor Geometry

Recently, Kallen and Zabzine computed the partition function of a twisted supersymmetric Yang-Mills theory on the five-dimensional sphere using localisation techniques. Key to their construction is a five-dimensional generalisation of the instanton equation to which they refer as the contact instanton equation. Subject of this article is the twistor construction of this equation when formulated on K-contact manifolds and the discussion of its integrability properties. We also present certain extensions to higher dimensions and supersymmetric generalisations.Comment: v3: 28 pages, clarifications and references added, version to appear in JHE