Benchmarking sequencing methods and tools that facilitate the study of alternative polyadenylation

Background Alternative cleavage and polyadenylation (APA), an RNA processing event, occurs in over 70% of human protein-coding genes. APA results in mRNA transcripts with distinct 3′ ends. Most APA occurs within 3′ UTRs, which harbor regulatory elements that can impact mRNA stability, translation, and localization. Results APA can be profiled using a number of established computational tools that infer polyadenylation sites from standard, short-read RNA-seq datasets. Here, we benchmarked a number of such tools—TAPAS, QAPA, DaPars2, GETUTR, and APATrap— against 3′-Seq, a specialized RNA-seq protocol that enriches for reads at the 3′ ends of genes, and Iso-Seq, a Pacific Biosciences (PacBio) single-molecule full-length RNA-seq method in their ability to identify polyadenylation sites and quantify polyadenylation site usage. We demonstrate that 3′-Seq and Iso-Seq are able to identify and quantify the usage of polyadenylation sites more reliably than computational tools that take short-read RNA-seq as input. However, we find that running one such tool, QAPA, with a set of polyadenylation site annotations derived from small quantities of 3′-Seq or Iso-Seq can reliably quantify variation in APA across conditions, such asacross genotypes, as demonstrated by the successful mapping of alternative polyadenylation quantitative trait loci (apaQTL). Conclusions We envisage that our analyses will shed light on the advantages of studying APA with more specialized sequencing protocols, such as 3′-Seq or Iso-Seq, and the limitations of studying APA with short-read RNA-seq. We provide a computational pipeline to aid in the identification of polyadenylation sites and quantification of polyadenylation site usages using Iso-Seq data as input.

Cascade Spiking Neuron Network For Event-based Image Classification In Noisy Environment

<div> <div> <div> <div> <p>Spiking Neuron Network (SNN) has shown advantages in processing event-based data for image classification. However, the classification accuracy of SNNs decreases in noisy environment. The cascade spiking neuron network (cascade-SNN) was proposed to solve this problem in this letter. We used spiking convolutional spiking neuron network (SCNN) for features extraction and liquid state machine (LSM) for read out. Compared with early works on ANNs, this network achieved the state-of-the-art classification accuracy in DVS-CIFAR10 dataset and DVS-Gesture dataset, which are both challenging dataset because of noisy environment. We conducted ablation experiments to verify the proposed structure is effective and analyzed the influence of different hyper-parameters. </p> </div> </div> </div> </div>

Cascade Spiking Neuron Network For Event-based Image Classification In Noisy Environment

<div> <div> <div> <div> <p>Spiking Neuron Network (SNN) has shown advantages in processing event-based data for image classification. However, the classification accuracy of SNNs decreases in noisy environment. The cascade spiking neuron network (cascade-SNN) was proposed to solve this problem in this letter. We used spiking convolutional spiking neuron network (SCNN) for features extraction and liquid state machine (LSM) for read out. Compared with early works on ANNs, this network achieved the state-of-the-art classification accuracy in DVS-CIFAR10 dataset and DVS-Gesture dataset, which are both challenging dataset because of noisy environment. We conducted ablation experiments to verify the proposed structure is effective and analyzed the influence of different hyper-parameters. </p> </div> </div> </div> </div>

PPR-DYW Protein EMP17 Is Required for Mitochondrial RNA Editing, Complex III Biogenesis, and Seed Development in Maize

The conversion of cytidines to uridines (C-to-U) at specific sites in mitochondrial and plastid transcripts is a post-transcriptional processing event that is important to the expression of organellar genes. Pentatricopeptide repeat (PPR) proteins are involved in this process. In this study, we report the function of a previously uncharacterized PPR-DYW protein, Empty pericarp17 (EMP17), in the C-to-U editing and kernel development in maize. EMP17 is targeted to mitochondria. The loss-function of EMP17 arrests maize kernel development, abolishes the editing at ccmFC-799 and nad2-677 sites, and reduces the editing at ccmFC-906 and -966 sites. The absence of editing causes amino acid residue changes in CcmFC-267 (Ser to Pro) and Nad2-226 (Phe to Ser), respectively. As CcmFC functions in cytochrome c (Cytc) maturation, the amount of Cytc and Cytc1 protein is drastically reduced in emp17, suggesting that the CcmFC-267 (Ser to Pro) change impairs the CcmFC function. As a result, the assembly of complex III is strikingly decreased in emp17. In contrast, the assembly of complex I appears less affected, suggesting that the Nad2-226 (Phe to Ser) change may have less impact on Nad2 function. Together, these results indicate that EMP17 is required for the C-to-U editing at several sites in mitochondrial transcripts, complex III biogenesis, and seed development in maize.

ALS-linked FUS mutants affect the localization of U7 snRNP and replication-dependent histone gene expression in human cells

Genes encoding replication-dependent histones lack introns, and the mRNAs produced are a unique class of RNA polymerase II transcripts in eukaryotic cells that do not end in a polyadenylated tail. Mature mRNAs are thus formed by a single endonucleolytic cleavage that releases the pre-mRNA from the DNA and is the only processing event necessary. U7 snRNP is one of the key factors that determines the cleavage site within the 3ʹUTR of replication-dependent histone pre-mRNAs. We have previously showed that the FUS protein interacts with U7 snRNA/snRNP and regulates the expression of histone genes by stimulating transcription and 3ʹ end maturation. Mutations in the FUS gene first identified in patients with amyotrophic lateral sclerosis (ALS) lead to the accumulation of the FUS protein in cytoplasmic inclusions. Here, we report that mutations in FUS lead to disruption of the transcriptional activity of FUS and mislocalization of U7 snRNA/snRNP in cytoplasmic aggregates in cellular models and primary neurons. As a consequence, decreased transcriptional efficiency and aberrant 3ʹ end processing of histone pre-mRNAs were observed. This study highlights for the first time the deregulation of replication-dependent histone gene expression and its involvement in ALS.

The MAGOH paralogs - MAGOH, MAGOHB and their multiple isoforms

A central processing event in eukaryotic gene expression is splicing. Concurrent with splicing, the core-EJC proteins, eIF4A3 and RBM8A-MAGOH heterodimer are deposited 24 bases upstream of newly formed exon-exon junctions. One of the core-EJC proteins, MAGOH contains a paralog MAGOHB, and this paralog pair is conserved across vertebrates. Upon analysis of the splice variants of MAGOH-paralogs, we have found the presence of alternate protein isoforms which are also evolutionarily conserved. Further, comparison of the amino acid sequence of the principal and alternate protein isoforms has revealed absence of key amino acid residues in the alternate isoforms. The conservation of principal and alternate isoforms correlates to the importance of MAGOH and MAGOHB across vertebrates.

Pre-mRNA processing entropy in a mouse model of trauma

PurposeNext generation sequencing has expanded our understanding of many disease processes, including trauma and critical illness. Many studies focus identifying a small set of genes or proteins that are aberrantly expressed. Our objective was to determine whether global differences in pre-mRNA processing entropy, or disorder, could offer novel insights in the setting of critical illness.MethodsWe used an established murine model of trauma that consisted of hemorrhagic shock and cecal ligation and puncture. In our first experiment mice exposed to trauma were compared to controls. In our second experiment, survival 14 days after exposure to trauma was studied. Using deep RNA sequencing we determined entropy values for every pre-mRNA processing event identified. We then used principal component analysis (PCA) to conduct unsupervised classification of the data.ResultsMice exposed to trauma separated from controls using PCA. Similarly, mice that did not survive 14 days post exposure clustered closely together on PCA.ConclusionOur results suggest that there is a substantial difference in global pre-mRNA processing entropy in mice exposed to trauma vs. controls, and that pre-mRNA processing entropy may be helpful in predicting mortality. The method introduced here is easily transferrable to other disease processes and samples.

The effect of empathy and context on face-processing ERPs

The investigation of emotional face processing has largely used faces devoid of context, and does not account for within-perceiver differences in empathy. The importance of context in face perception has become apparent in recent years. This study examined the interaction of the contextual factors of facial expression, knowledge of a person’s character, and within-perceiver empathy levels on face processing event-related potentials (ERPs). Forty-two adult participants learned background information about six individuals’ character. Three types of character were described, in which the character was depicted as deliberately causing harm to others, accidentally causing harm to others, or undertaking neutral actions. Subsequently, EEG was recorded while participants viewed the characters’ faces displaying neutral or emotional expressions. Participants’ empathy was assessed using the Empathy Quotient survey. Results showed a significant interaction of character type and empathy on the early posterior negativity (EPN) ERP component. These results suggested that for those with either low or high empathy, more attention was paid to the face stimuli, with more distinction between the different characters. In contrast, those in the middle range of empathy tended to produce smaller EPN with less distinction between character types. Findings highlight the importance of trait empathy in accounting for how faces in context are perceived.

The CMS DAQ Pinball Machine

We present an interactive game for up to seven players that demonstrates the challenges of on-line event selection at the Compact Muon Solenoid (CMS) experiment to the public. The game - in the shape of a popular classic pinball machine - was conceived and prototyped by an interdisciplinary team of graphic designers, physicists and engineers at the CMS Create hackathon in 2016. Having won the competition, the prototype was turned into a fully working machine that is now exhibited on the CMS visitors’ path. Teams of 2-7 visitors can compete with one another to collect as many interesting events as possible within a simulated LHC fill. In a fun and engaging way, the game conveys concepts such as multi-level triggering, pipelined processing, event building, the importance of purity in event selection and more subtle details such as dead time. The multi-player character of the game corresponds to the distributed nature of the actual trigger and data acquisition system of the experiment. We present the concept of the game, its design and its technical implementation centered around an Arduino micro-controller controlling 700 RGB LEDs and a sound subsystem running on a Mac mini.