Project description:The most common genetic cause of both frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS) is a G4C2 repeat expansion in intron 1 of the C9orf72 gene. This repeat expansion undergoes bidirectional transcription to produce sense and antisense repeat RNA species. Both sense and antisense-derived repeat RNAs undergo repeat-associated non-AUG translation in all reading frames to generate five distinct dipeptide repeat proteins (DPRs). Importantly, toxicity has been associated with both sense and antisense repeat-derived RNA and DPRs. This suggests targeting both sense and antisense repeat RNA may provide the most effective therapeutic strategy. The RNA-targeting CRISPR-Cas13 systems offer a promising avenue for simultaneous targeting of multiple RNA transcripts, as they mature their own guide arrays, thus allowing targeting of more than one RNA species from a single construct. We show that CRISPR-Cas13d originating from Ruminococcus flavefaciens (CasRx) can successfully reduce C9orf72 sense and antisense repeat transcripts and DPRs to background levels in HEK cells overexpressing C9orf72 repeats. CRISPR-CasRx also markedly reduced the endogenous sense and antisense repeat RNAs and DPRs in three independent C9orf72 patient-derived iPSC-neuron lines, without detectable off-target effects. To determine whether CRISPR-CasRx is effective in vivo, we treated two distinct C9orf72 repeat mouse models using AAV delivery and observed a significant reduction in both sense and antisense repeat-containing transcripts. Taken together this work highlights the potential for RNA-targeting CRISPR systems as therapeutics for C9orf72 ALS/FTD.
Project description:Hdac4 has been found to modulate symptoms in Huntington's Disease (HD) mouse models through an uknown mechanism unrelated to any enzymatic activity. We investigated the protein-protein interactions to gain insight into the role of Hdac4 in HD.
Project description:Large White and Meishan pigs were either non-treated or injected with mammalian 1-24 ACTH (Immediate Synachten, Novartis France) at the dose of 250 µg per animal. Pigs were sacrificed either immediately after capture from their home cage (non-treated animals) or 1 hour following ACTH injection. Adrenal glands were immediately collected from pigs and frozen on dry ice and then stored at -80°C until RNA isolation. Keywords: stress response, adrenal, gene expression, pig
Project description:BACKGROUND:In animal breeding, identification of causative genetic variants is of major importance and high economical value. Usually, the number of candidate variants exceeds the number of variants that can be validated. One way of prioritizing probable candidates is by evaluating their potential to have a deleterious effect, e.g. by predicting their consequence. Due to experimental difficulties to evaluate variants that do not cause an amino-acid substitution, other prioritization methods are needed. For human genomes, the prediction of deleterious genomic variants has taken a step forward with the introduction of the combined annotation dependent depletion (CADD) method. In theory, this approach can be applied to any species. Here, we present pCADD (p for pig), a model to score single nucleotide variants (SNVs) in pig genomes. RESULTS:To evaluate whether pCADD captures sites with biological meaning, we used transcripts from miRNAs and introns, sequences from genes that are specific for a particular tissue, and the different sites of codons, to test how well pCADD scores differentiate between functional and non-functional elements. Furthermore, we conducted an assessment of examples of non-coding and coding SNVs, which are causal for changes in phenotypes. Our results show that pCADD scores discriminate between functional and non-functional sequences and prioritize functional SNVs, and that pCADD is able to score the different positions in a codon relative to their redundancy. Taken together, these results indicate that based on pCADD scores, regions with biological relevance can be identified and distinguished according to their rate of adaptation. CONCLUSIONS:We present the ability of pCADD to prioritize SNVs in the pig genome with respect to their putative deleteriousness, in accordance to the biological significance of the region in which they are located. We created scores for all possible SNVs, coding and non-coding, for all autosomes and the X chromosome of the pig reference sequence Sscrofa11.1, proposing a toolbox to prioritize variants and evaluate sequences to highlight new sites of interest to explain biological functions that are relevant to animal breeding.
Project description:We develop a CRISPR-Assisted RNA-Protein Interaction Detection method (CARPID), which leverages CRISPR/CasRx-based RNA targeting and proximity labeling to identify binding proteins of specific lncRNA in the native cellular context.
Project description:Large White and Meishan pigs were either non-treated or injected with mammalian 1-24 ACTH (Immediate Synachten, Novartis France) at the dose of 250 µg per animal. Pigs were sacrificed either immediately after capture from their home cage (non-treated animals) or 1 hour following ACTH injection. Adrenal glands were immediately collected from pigs and frozen on dry ice and then stored at -80°C until RNA isolation. Keywords: stress response, adrenal, gene expression, pig 47 samples
Project description:Fecal samples (N = 10) from 6- to 8-week-old wild boar piglets (Sus scrofa), collected from an animal park in Hungary in April 2011, were analyzed using viral metagenomics and complete genome sequencing. Kobuvirus (genus Kobuvirus, family Picornaviridae) was detected in all (100 %) specimens, with the closest nucleotide (89 %) and amino acid (94 %) sequence identity of the strain wild boar/WB1-HUN/2011/HUN (JX177612) to the prototype porcine kobuvirus S-1-HUN (EU787450). This study suggests that genetically highly similar (practically the same geno-/serotype) porcine kobuvirus circulate in wild boars, the wildlife counterparts of domestic pigs. Wild boars could be an important host and reservoir for kobuvirus.
Project description:Contrary to spontaneous yawning-an ancient phenomenon common to vertebrates-contagious yawning (elicited by others' yawns) has been found only in highly social species and may reflect an emotional inter-individual connection. We investigated yawn contagion in the domestic pig, Sus scrofa. Owing to the complex socio-emotional and cognitive abilities of Sus scrofa, we posited that yawn contagion could be present in this species (Prediction 1) and influenced by individual/social factors (Prediction 2). In June-November 2018, on 104 semi-free ranging adolescent/adult pigs, 224 videos were recorded for video analysis on yawning. Kinship information was refined via genetic analyses. Statistical elaboration was conducted via GLMMs and non-parametric/randomization/cross-tabulation tests. We found yawn contagion in Sus scrofa, as it was more likely that pigs yawned when perceiving rather than not perceiving (yawning/control condition) others' yawns (response peak in the first out of three minutes). Yawn contagion was more likely: (1) in response to males' yawns; (2) as the age increased; (3) within short distance (1 m); (4) between full siblings, with no significant association between kinship and distance. The influence of kinship suggests that-as also hypothesized for Homo sapiens-yawn contagion might be linked with emotional communication and possibly contagion.