Miracle-dz/UniFAR-FTUs huggingface dataset:
Single cell 3’UTR analysis identifies changes in alternative polyadenylation throughout neuronal differentiation and in autism

Autism spectrum disorder (ASD) is a neurodevelopmental disease affecting social behavior. Many of the high-confident ASD risk genes relate to mRNA translation. Specifically, many of these genes are involved in regulation of gene expression for subcellular compartmentalization of proteins1. Cis-regulatory motifs that often localize to 3’- and 5’-untranslated regions (UTRs) offer an additional path for posttranscriptional control of gene expression. Alternative cleavage and polyadenylation (APA) affect 3’UTR length thereby influencing the presence or absence of regulatory elements. However, APA has not yet been addressed in the context of neurodevelopmental disorders. Here we used single cell 3’end sequencing to examine changes in 3’UTRs along the differentiation from neural stem cells (NSCs) to neuroblasts within the adult brain. We identified many APA events in genes involved in neurodevelopment, many of them being high confidence ASD risk genes. Further, analysis of 3’UTR lengths in single cells from ASD and healthy individuals detected longer 3’UTRs in ASD patients. Motif analysis of modulated 3’UTRs in the mouse adult neurogenic lineage and ASD-patients revealed enrichment of the cytoplasmic and polyadenylation element (CPE). This motif is bound by CPE binding protein 4 (CPEB4). In human and mouse data sets we observed co-regulation of CPEB4 and the CPEB-binding synaptic adhesion molecule amyloid beta precursor-like protein 1 (APLP1). We show that mice deficient in APLP1 show aberrant regulation of APA, decreased number of neural stem cells, and autistic-like traits. Our findings indicate that APA is used for control of gene expression along neuronal differentiation and is altered in ASD patients.
Chronic Corticosterone Pretreatment Reverses Psilocybin’s Effects on Mouse Anxious and Hedonic Behaviors

Psilocybin has shown positive preliminary signals in small-scale clinical trials for psychiatric disorders that exhibit maladaptive stress responses as a major component of their presentation. However, there are relatively few assessments of whether an acute administration of psilocybin exhibits reproducible effects in rodent models useful for the study of stress-associated psychiatric disorders. Here, we measured the responses of male C57BL/6J mice to this compound in a battery of relevant behavioral tests. These tests included the open-field test, forced swim test, sucrose preference test, and novelty suppressed feeding test. Furthermore, these tests were presented in either the absence or presence of chronic corticosterone administration, as a chemically induced model of ongoing stress burden. Our results indicate that the effects of psilocybin within these tests are dependent on the chronic hormonal stress burden of the mice: psilocybin alone promotes anxiolytic and hedonic responses, but promotes anxiogenic and anhedonic responses when pre-treated with chronic corticosterone. This identified interaction between stress hormone burden and psilocybin behavioral effects in mice suggests the possibility of further developing rodent behavioral models that can assess additional context-dependent effects of psychedelic administration that are deemed clinically-relevant, but are otherwise difficult to control for, in human studies.
Proteomics reveals synergy in biomass conversion between fungal enzymes and inorganic Fenton chemistry in leaf-cutting ant colonies

The herbivorous symbiosis between leaf-cutting ants and fungal cultivars processes biomass via ant fecal fluid mixed with munched plant substrate before fungal degradation. Here we present a full proteome of the fecal fluid of Acromyrmex leaf-cutting ants, showing that most proteins function as biomass degrading enzymes and that ca. 80% are produced by the fungal cultivar and ingested, but not digested, by the ants. Hydrogen peroxide producing oxidoreductases were remarkably common in the fecal proteome, inspiring us to test a scenario in which hydrogen peroxide reacts with iron in the fecal fluid to form reactive oxygen radicals after which oxidized iron is reduced by other fecal-fluid enzymes. Our biochemical assays confirmed that these cyclical Fenton reactions do indeed take place in special substrate pellets, presumably to degrade recalcitrant lignocellulose. This implies that the symbiosis manages a combination of chemical and enzymatic degradation, an achievement that surpasses current human bioconversion technology.
Evaluation of Alginate-Based Bioinks for 3D Bioprinting, Mesenchymal Stromal Cell Osteogenesis, and Application for Patient-Specific Bone Grafts

To realize the promise of 3D bioprinting, it is imperative to develop bioinks that possess the biological and rheological characteristics needed for the printing of cell-laden tissue grafts. Alginate is widely used as a bioink because its rheological properties can be modified through pre-crosslinking or the addition of thickening agents to increase printing resolution. However, modification of alginate’s physicochemical characteristics using common crosslinking agents can affect its cytocompatibility. Therefore, we evaluated the printability, physicochemical properties, and osteogenic potential of four common alginate bioinks: alginate-CaCl2 (alg-CaCl2), alginate-CaSO4 (alg-CaSO4), alginate-gelatin (alg-gel) and alginate-nanocellulose (alg-ncel) for the 3D bioprinting of patient-specific osteogenic grafts. While all bioinks possessed similar viscosity, printing fidelity was lower in the pre-crosslinked bioinks. When used to print geometrically defined constructs, alg-CaSO4 and alg-ncel exhibited higher mechanical properties and lower mesh size than those printed with alg-CaCl2 or alg-gel. The physical properties of these constructs affected the biological performance of encapsulated bone marrow-derived mesenchymal stromal cells (MSCs). Cell-laden constructs printed using alg-CaSO4 and alg-ncel exhibited greater cell apoptosis and contained fewer living cells 7 days post-printing. In addition, effective cell-matrix interactions were only observed in alg-CaCl2 printed constructs. When cultured in osteogenic media, MSCs in alg-CaCl2 constructs exhibited increased osteogenic differentiation compared to the other three bioinks. This bioink was then used to 3D print anatomically accurate cell-laden scaphoid bones that were capable of partial mineralization after 14 days of in vitro culture. These results highlight the importance of bioink properties to modulate cell behavior and the biofabrication of clinically relevant bone tissues.
Cell Ecosystem and Signaling Pathways of Primary and Metastatic Pediatric Posterior Fossa Ependymoma

Pediatric ependymoma is a devastating brain cancer marked by its relapsing pattern and lack of effective chemotherapies. This shortage of treatments is partially due to limited knowledge about ependymoma tumorigenic mechanisms. Although there is evidence that ependymoma originates in radial glia, the specific pathways underlying the progression and metastasis of these tumors are unknown. By means of single-cell transcriptomics, immunofluorescence, and in situ hybridization, we show that the expression profile of tumor cells from pediatric ependymomas in the posterior fossa is consistent with an origin in LGR5+ stem cells. Tumor stem cells recapitulate the developmental lineages of radial glia in neurogenic niches, promote an inflammatory microenvironment in cooperation with microglia, and upon metastatic progression initiate a mesenchymal program driven by reactive gliosis and hypoxia-related genes. Our results uncover the cell ecosystem of pediatric posterior fossa ependymoma and identify WNT/β-catenin and TGF-β signaling as major drivers of tumorigenesis for this cancer.
Transient Delivery of A-C/EBP Protein Perturbs Differentiation of 3T3-L1 Cells and Induces Preadipocyte Marker Genes

Transformation of committed 3T3-L1 preadipocytes to lipid-laden adipocytes involves timely appearance of numerous transcription factors (TFs), foremost among them C/EBPβ, is expressed during early phases of differentiation. Here we describe liposome-mediated protein transfection approach to rapidly downregulate C/EBPβ by A-C/EBP protein inhibitor. Signals from tagged A-C/EBP were observed in 3T3-L1 cells within 2hrs of protein inhibitor transfections whereas for gene transfection signals appeared in 48hrs. Following transient transfections, expression profiles of 24 marker genes belonging to pro- and anti-adipogenic, cell cycle, and preadipocytes pathways was analyzed. mRNA and protein expression profiles of adipocyte-marker genes showed lower expression in both A-C/EBP protein and gene transfected samples. Interestingly, for preadipocytes and cell fate determinant genes, striking differences were observed between protein and gene transfected samples. Preadipocyte differentiation factors Stat5a and Creb were downregulated in A-C/EBP protein samples. Five preadipocyte markers, namely, Pdgfrα, Pdgfrβ, Ly6A, CD34 Itgb1 showed high expression in protein samples whereas only Ly6A and CD34 were expressed in gene transfected samples. Pdgfrα and Pdgfrβ, two known cell fate markers were expressed in protein transfected samples 5-days post-differentiation suggesting a possible reversal of differentiation. Our study provides evidences for the robust and efficient knockdown of C/EBPβ protein to understand time-dependent gene regulation during adipogenesis.
Learning with uncertainty for biological discovery and design

Machine learning that generates biological hypotheses has transformative potential, but most learning algorithms are susceptible to pathological failure when exploring regimes beyond the training data distribution. A solution is to quantify prediction uncertainty so that algorithms can gracefully handle novel phenomena that confound standard methods. Here, we demonstrate the broad utility of robust uncertainty prediction in biological discovery. By leveraging Gaussian process-based uncertainty prediction on modern pretrained features, we train a model on just 72 compounds to make predictions over a 10,833-compound library, identifying and experimentally validating compounds with nanomolar affinity for diverse kinases and whole-cell growth inhibition of Mycobacterium tuberculosis. We show how uncertainty facilitates a tight iterative loop between computation and experimentation, improves the generative design of novel biochemical structures, and generalizes across disparate biological domains. More broadly, our work demonstrates that uncertainty should play a key role in the increasing adoption of machine learning algorithms into the experimental lifecycle.
The phase of Theta oscillations modulates successful memory formation at encoding

Several past studies have shown that attention and perception can depend upon the phase of ongoing neural oscillations at stimulus onset. Here, we extend this idea to the memory domain. We tested the hypothesis that ongoing fluctuations in neural activity have an impact on memory encoding using a picture paired-associates task to gauge episodic memory performance. Experiment 1 capitalized on the principle of phase reset. We tested if subsequent memory performance fluctuates rhythmically, time-locked to a reset cue presented before the to-be-remembered pairs. We found indication that behavioral performance was periodically and selectively modulated at theta frequency (∼4 Hz). In Experiment 2 we focused on prestimulus ongoing activity using scalp EEG recorded while participants performed the pair-associate task. We analyzed subsequent memory performance as a function of theta and alpha activity around the presentation of the to-be-remembered pairs. The results of the pre-registered analyses, using large electrode clusters and generic spectral ranges, returned null results of prestimulus phase-behavior correlation. However, we found that post-stimulus theta-power modulations in left frontal scalp predicted subsequent memory performance. This post-stimulus effect in theta power was used to guide a post-hoc prestimulus phase analysis, narrowed down to more precise scalp location and frequency. This analysis returned a correlation between prestimulus theta phase and subsequent memory. Altogether, these results suggest that the prestimulus theta activity at encoding has an impact on later memory performance.
A chlorzoxazone-baclofen combination improves cerebellar impairment in spinocerebellar ataxia type 1

Background A combination of central muscle relaxants, chlorzoxazone and baclofen (chlorzoxazone-baclofen), has been proposed for treatment of cerebellar symptoms in human spinocerebellar ataxia (SCA). However, central muscle relaxants can worsen balance. The optimal dose for target engagement without toxicity remains unknown. Objectives Using the genetically precise Atxn1154Q/2Q model of SCA1, we determine the role of cerebellar dysfunction in motor impairment. We also aim to identify appropriate concentrations of chlorzoxazone-baclofen needed for target engagement without toxicity to plan for human clinical trials. Methods We use patch clamp electrophysiology in acute cerebellar slices and immunostaining to identify the specific ion channels targeted by chlorzoxazone-baclofen. Behavioral assays for coordination and grip strength are used to determine specificity of chlorzoxazone-baclofen for improving cerebellar dysfunction without off-target effects in Atxn1154Q/2Q mice. Results We identify irregular Purkinje neuron firing in association with reduced expression of the ion channels Kcnma1 and Cacna1g in Atxn1154Q/2Q mice. Using in vitro electrophysiology in brain slices, we identify concentrations of chlorzoxazone-baclofen that improve Purkinje neuron spike regularity without reducing firing frequency. At a disease stage in Atxn1154Q/2Q mice when motor impairment is due to cerebellar dysfunction, orally administered chlorzoxazone-baclofen improves motor performance without affecting muscle strength. Conclusion We identify a tight relationship between baclofen-chlorzoxazone concentrations needed to engage target, and levels above which cerebellar function will be compromised. We propose to use this information for a novel clinical trial design, using sequential dose escalation within each subject, to identify dose levels that are likely to improve ataxia symptoms while minimizing toxicity.
Single-molecule view of coordination in a multi-functional DNA polymerase

Replication and repair of genomic DNA requires the action of multiple enzymatic functions that must be coordinated in order to ensure efficient and accurate product formation. Here we have used single-molecule FRET microscopy to investigate the physical basis of functional coordination in DNA polymerase I (Pol I) from E. coli, a key enzyme involved in lagging-strand replication and base excision repair. Pol I contains active sites for template-directed DNA polymerization and 5’ flap processing in separate domains. We show that a DNA substrate can spontaneously transfer between polymerase (pol) and 5’ nuclease (5’ nuc) domains during a single encounter with Pol I. Additionally, we show that the flexibly tethered 5’ nuc domain adopts different positions within Pol I-DNA complexes, depending on the nature of the DNA substrate. Our results reveal the structural dynamics that underlie functional coordination in Pol I and are likely relevant to other multi-functional DNA polymerases.
ATF3 drives senescence by reconstructing accessible chromatin profiles

Chromatin architecture and gene expression profile undergo tremendous reestablishment during senescence. However, the regulatory mechanism between chromatin reconstruction and gene expression in senescence remain elusive. The chromatin accessibility is an excellent perspective to reveal the latent regulatory elements. Thus, we depicted the landscapes of chromatin accessibility and gene expression during HUVECs senescence. We found that chromatin accessibilities are re-distributed during senescence. The senescence related increased accessible regions (IARs) and the decreased accessible regions (DARs) are mainly distributed in distal intergenic regions. The DARs are correlated with the function declines caused by senescence, whereas the IARs are involved in the regulation for senescence program. Moreover, the heterochromatin contributes most of IARs in senescent cells. We identified that the AP-1 transcription factors, especially ATF3 is responsible for driving chromatin accessibility reconstruction in IARs. In particular, DNA methylation is negatively correlated with chromatin accessibility during senescence. AP-1 motifs with low DNA methylation may improve their binding affinity in IARs and further opens the chromatin nearby. Our results described a dynamic landscape of chromatin accessibility whose remodeling contributes to the senescence program. And we identified a cellular senescence regulator, AP-1, which promotes senescence through organizing the accessibility profile in IARs.
“Supersessioning”: A hardware/software system for electrophysiology spanning multiple sessions in marmosets

We introduce a straightforward, robust method for recording and analyzing spiking activity over timeframes longer than a single session, with primary application to the marmoset (Callithrix jacchus). Although in theory the marmoset’s smooth brain allows for broad deployment of powerful tools in primate cortex, in practice marmosets do not typically engage in long experimental sessions akin to rhesus monkeys. This potentially limits their value for detailed, quantitative neurophysiological study. Here we describe chronically-implanted arrays with a 3D arrangement of electrodes yielding stable single and multi-unit responses, and an analytic method for creating “supersessions” combining that array data across multiple experiments. We could match units across different recording sessions over several weeks, demonstrating the feasibility of pooling data over sessions. This could be a key tool for extending the viability of marmosets for dissecting neural computations in primate cortex.
Culture volume influences the dynamics of adaptation under long-term stationary phase

Escherichia coli and many other bacterial species, which are incapable of sporulation, can nevertheless survive within resource exhausted media by entering a state termed long-term stationary phase (LTSP). We have previously shown that E. coli populations adapt genetically under LTSP in an extremely convergent manner. Here we examine how the dynamics of LTSP genetic adaptation are influenced by varying a single parameter of the experiment - culture volume. We find that culture volume affects survival under LTSP, with viable counts decreasing as volumes increase. Across all volumes, mutations accumulate with time, and the majority of mutations accumulated demonstrate signals of being adaptive. However, positive selection appears to affect mutation accumulation more strongly at higher, compared to lower volumes. Finally, we find that several similar genes are likely involved in adaptation across volumes. However, the specific mutations within these genes that contribute to adaptation can vary in a consistent manner. Combined, our results demonstrate how varying a single parameter of an evolutionary experiment can substantially influence the dynamics of observed adaptation.
Cell-Scale Biophysical Cues from Collagen Fiber Architecture Instruct Cell Behavior and the Propagation of Mechanosensory Signals

Mechanosensory cues from the extracellular matrix underpin numerous cellular behaviors including tumor cell migration yet are influenced by the local structure and organization of the matrix in unknown ways. To investigate mechanical cues with respect to local collagen organization, we used a combination of intravital imaging of the mammary tumor microenvironment and 3D collagen gel systems with both migratory MDA-MB-231 cells and acellular pNIPAAm beads. We identified that fiber organization directs a bias in cell response along the axis of alignment. Using innovative methodology, we determined that local collagen alignment resulted in a 30-fold difference in directional cell-scale stiffness and also dramatically altered the rate at which cell-induced fiber displacements decayed over distance. Our results reveal differential mechanical properties across orthogonal directions in aligned matrices that provide sizeable cues to the cell and have important implications for cellular mechanosensing and cell-cell communication within the tissue microenvironment.
Levels of Moodle Use to Support University Face-to-Face Teaching

Educational platforms such as Moodle have become ubiquitous tools in universities around the world. They are often the technological basis for online and mixed teaching-learning processes, as well as a key tool in supporting face-to-face teaching-learning processes. This is the context for this paper. The aim is to find the levels of Moodle use in undergraduate face-to-face teaching at the University of Deusto, and to analyze its relationship with other factors such as the faculty, teaching experience, previous experience with Moodle, and specialist training received. A questionnaire was designed and sent to a sample of 100 lecturers from five different faculties. Five levels of lecturers’ use of Moodle were obtained by using a quantitative methodology and applying cluster analysis. The results showed a relatively limited use, since more than two-third of lecturers reported they used Moodle to distribute teaching materials, send messages to the news forum, and manage tasks without feedback, all of which were at the individual student level and not involving student groups.
RNAcmap: A Fully Automatic Method for Predicting Contact Maps of RNAs by Evolutionary Coupling Analysis

Motivation The accuracy of RNA secondary and tertiary structure prediction can be significantly improved by using structural restraints derived from evolutionary or direct coupling analysis. Currently, these coupling analyses relied on manually curated multiple sequence alignments collected in the Rfam database, which contains 3016 families. By comparison, millions of non-coding RNA sequences are known. Here, we established RNAcmap, a fully automatic method that enables evolutionary coupling analysis for any RNA sequences. The homology search was based on the covariance model built by Infernal according to two secondary structure predictors: a folding-based algorithm RNAfold and the latest deep-learning method SPOT-RNA. Results We show that the performance of RNAcmap is less dependent on the specific evolutionary coupling tool but is more dependent on the accuracy of secondary structure predictor with the best performance given by RNAcmap (SPOT-RNA). The performance of RNAcmap (SPOT-RNA) is comparable to that based on Rfam-supplied alignment and consistent for those sequences that are not in Rfam collections. Further improvement can be made with a simple meta predictor RNAcmap (SPOT-RNA/RNAfold) depending on which secondary structure predictor can find more homologous sequences. Reliable base-pairing information generated from RNAcmap, for RNAs with high effective homologous sequences, in particular, will be useful for aiding RNA structure prediction. Availability and implementation RNAcmap is available as a web server at https://sparks-lab.org/server/rnacmap/) and as a standalone application along with the datasets at https://github.com/sparks-lab-org/RNAcmap.