Biology subjects

Torres, M.

Publications and source records attributed to Torres, M..

51 records · Page 3Linked to original sources

bioRxiv2022

The activation of mGluR4 rescues parallel fiber LTP, motor learning and social behavior in a mouse model of Fragile X Syndrome

Fragile X patients and mice lacking the Fragile X Mental Retardation Protein (FMRP) suffer from multiple behavioral alterations, including deficits in motor learning. We found that enhanced synaptic vesicle (SV) docking in cerebellar parallel fiber to Purkinje cell Fmr1KO synapses was associated with enhanced asynchronous release, which not only occludes further potentiation, but it also compromises presynaptic parallel fiber long-term potentiation (PF-LTP). A reduction in extracellular Ca2+ restored the readily releasable pool (RRP) size, rescuing {beta} adrenergic receptor-mediated potentiation and parallel fiber LTP. Interestingly, VU 0155041, a selective positive allosteric modulator of mGluR4, also restored both the RRP size and parallel fiber LTP. Moreover, when injected into Fmr1KO mice, VU 0155041 improved motor learning in skilled reaching, classical eyeblink conditioning and vestibuloocular reflex (VOR) tests, as well as improving the social behavior of these mice. Thus, pharmacological activation of mGluRs may offer therapeutic relief for motor learning and social deficits in Fragile X Syndrome.

neuroscience↗

bioRxiv2021

Transcriptional response in a sepsis mouse model reflects transcriptional response in sepsis patients

Mortality due to sepsis remains unacceptably high, especially for septic shock patients. Murine models have been used to better understand pathophysiology mechanisms. However, the mouse model is still under debate. Here we investigated the transcriptional response of mice injected with lipopolysaccharide (LPS) and compared it to either human cells stimulated in vitro with LPS or to blood cells of septic patients. We identified a molecular signature composed of 2331 genes with an FDR median of 0%. This molecular signature is highly enriched in regulated genes in peritoneal macrophages stimulated with LPS. There is a significant enrichment in several inflammatory signaling pathways, and in disease terms, such as pneumonia, sepsis, systemic inflammatory response syndrome, severe sepsis, an inflammatory disorder, immune suppression, and septic shock. A significant overlap between the genes up-regulated in mouse and human cells stimulated with LPS has been demonstrated. Finally, genes up-regulated in mouse cells stimulated with LPS are enriched in genes up-regulated in human cells stimulated in vitro and in septic patients, who are at high risk of death. Our results support the hypothesis of common molecular and cellular mechanisms between mouse and human sepsis.

genomics↗

bioRxiv2021

Pseudo-dynamic analysis of heart tube formation in the mouse reveals strong regional variability and early left-right asymmetry

Understanding organ morphogenesis requires a precise geometrical description of the tissues involved in the process. In highly regulative embryos, like those of mammals, morphological variability hinders the quantitative analysis of morphogenesis. In particular, the study of early heart development in mammals remains a challenging problem, due to imaging limitations and innate complexity. Around embryonic day 7.5 (E7.5), the cardiac crescent folds in an intricate and coordinated manner to produce a pumping linear heart tube at E8.25, followed by heart looping at E8.5. In this work we provide a complete morphological description of this process based on detailed imaging of a temporally dense collection of embryonic heart morphologies. We apply new approaches for morphometric staging and quantification of local morphological variations between specimens at the same stage. We identify hot spots of regionalized variability and identify left-right asymmetry in the inflow region starting at the late cardiac crescent stage, which represents the earliest signs of organ left-right asymmetry in the mammalian embryo. Finally, we generate a 3D+t digital model that provides a framework suitable for co-representation of data from different sources and for the computer modelling of the process. SUMMARY STATEMENTWe provide the first complete atlas for morphometric analysis and visualization of heart tube morphogenesis, reporting morphological variability and early emergence of left-right asymmetry patterns.

developmental biology↗

bioRxiv2021

Unlike Chloroquine, mefloquine inhibits SARS-CoV-2 infection in physiologically relevant cells and does not induce viral variants.

Repositioning of clinical approved drugs could represent the fastest way to identify therapeutic options during public health emergencies, the majority of drugs explored for repurposing as antivirals for 2019 coronavirus disease (COVID-19) have failed to demonstrate clinical benefit. Without specific antivirals, the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic continues to cause major global mortality. Antimalarial drugs, such as chloroquine (CQ)/hydroxychloroquine (HCQ) and mefloquine have emerged as potential anti-SARS-CoV-2 antivirals. CQ/HCQ entered the Solidarity and RECOVERY clinical trials against COVID-19 and showed lack of efficacy. Importantly, mefloquine is not a 4-aminoquinoline like CQ and HCQ and has been previously repurposed for other respiratory diseases. Unlike the 4-aminoquinolines that accumulate in the high pH of intracellular lysosomes of the lung, the high respiratory tract penetration of mefloquine is driven by its high lipophilicity. While CQ and HCQ exhibit activity in Vero E6 cells, their activity is obviated in TMPRSS2-expressing cells, such as Calu-3 cells, which more accurately recapitulate in vivo entry mechanisms for SARS-CoV-2. Accordingly, here we report the anti-SARS-CoV-2 activity of mefloquine in Calu-3 type II pneumocytes and primary human monocytes. Mefloquine inhibited SARS-CoV-2 replication in Calu-3 cells with low cytotoxicity and EC50 and EC90 values of 1.2 and 5.3 {micro}M, respectively. In addition, mefloquine reduced up to 68% the SARS-CoV-2 RNA levels in infected monocytes, reducing viral-induced inflammation. Mefloquine blocked early steps of the SARS-CoV-2 replicative cycle and was less prone than CQ to induce drug-associated viral mutations and synergized with RNA polymerase inhibitor. The pharmacological parameters of mefloquine are consistent with its plasma exposure in humans and its tissue-to-plasma predicted coefficient points that this drug may accumulate in the lungs. These data indicate that mefloquine could represent an orally available clinically approved drug option against COVID-19 and should not be neglected on the basis of the failure of CQ and HCQ.

microbiology↗

bioRxiv2021

Genetic architecture of natural variations of cardiac performance in flies

Deciphering the genetic architecture of human cardiac disorders is of fundamental importance but their underlying complexity is a major hurdle. We investigated the natural variation of cardiac performance in the sequenced inbred lines of the Drosophila Genetic Reference Panel (DGRP). Genome Wide Associations Studies (GWAS) identified genetic networks associated with natural variation of cardiac traits which were used to gain insights as to the molecular and cellular processes affected. Non-coding variants that we identified were used to map potential regulatory non-coding regions, which in turn were employed to predict Transcription Factors (TFs) binding sites. Cognate TFs, many of which themselves bear polymorphisms associated with variations of cardiac performance, were also validated by heart specific knockdown. Additionally, we showed that the natural variations associated with variability in cardiac performance affect a set of genes overlapping those associated with average traits but through different variants in the same genes. Furthermore, we showed that phenotypic variability was also associated with natural variation of gene regulatory networks. More importantly, we documented correlations between genes associated with cardiac phenotypes in both flies and humans, which supports a conserved genetic architecture regulating adult cardiac function from arthropods to mammals. Specifically, roles for PAX9 and EGR2 in the regulation of the cardiac rhythm were established in both models, illustrating that the characteristics of natural variations in cardiac function identified in Drosophila can accelerate discovery in humans.

genetics↗

bioRxiv2021

Predisposition to Proinsulin Misfolding as a Genetic Risk to Diet-Induced Diabetes

Throughout evolution, proinsulin has exhibited significant sequence variation in both C-peptide and insulin moieties. As the proinsulin coding sequence evolves, the gene product continues to be under selection pressure both for ultimate insulin bioactivity and for the ability of proinsulin to be folded for export through the secretory pathway of pancreatic {beta}-cells. The substitution proinsulin-R(B22)E is known to yield a bioactive insulin, although R(B22)Q has been reported as a mutation that falls within the spectrum of Mutant INS-gene induced Diabetes of Youth (MIDY). Here we have studied mice expressing heterozygous (or homozygous) proinsulin-R(B22)E knocked into the Ins2 locus. Neither females nor males bearing the heterozygous mutation develop diabetes at any age examined, but subtle evidence of increased proinsulin misfolding in the endoplasmic reticulum is demonstrable in isolated islets from the heterozygotes. Moreover, males have indications of glucose intolerance and within a few week exposure to a high-fat diet, they develop frank diabetes. Diabetes is more severe in homozygotes, and the development of disease parallels a progressive heterogeneity of {beta}-cells with increasing fractions of proinsulin-rich/insulin-poor cells, as well as glucagon-positive cells. Evidently, sub-threshold predisposition to proinsulin misfolding can go undetected, but provides genetic susceptibility to diet-induced {beta}-cell failure.

cell biology↗

bioRxiv2020

The Fc-mediated effector functions of a potent SARS-CoV-2 neutralizing antibody, SC31, isolated from an early convalescent COVID-19 patient, are essential for the optimal therapeutic efficacy of the antibody

SARS-CoV-2-neutralizing antibodies are promising therapeutics for COVID-19. However, little is known about the mechanisms of action of these antibodies or their effective dosing windows. We report the discovery and development of SC31, a potent SARS-CoV-2 neutralizing IgG1 antibody, originally isolated from a convalescent patient at day 27 after the onset of symptoms. Neutralization occurs via a binding epitope that maps within the ACE2 interface of the SARS-CoV-2 Spike protein, conserved across all common circulating SARS-CoV-2 mutants. In SARS-CoV-2 infected K18-human ACE2 transgenic mice, SC31 demonstrated potent survival benefit by dramatically reducing viral load concomitant with attenuated pro-inflammatory responses linked to severe systemic disease, such as IL-6. Comparison with a Fc-null LALA variant of SC31 demonstrated that optimal therapeutic efficacy of SC31 requires intact Fc-mediated effector functions that can further induce an IFN{gamma}-driven anti-viral immune response. Dose-dependent efficacy for SC31 was observed down to 5mg/kg when dosed before the activation of lung inflammatory responses. Importantly, despite Fc{gamma}R binding, no evidence of antibody dependent enhancement was observed with the Fc-competent SC31 even at sub-therapeutic doses. Therapeutic efficacy was confirmed in SARS-CoV-2-infected hamsters, where SC31 again significantly reduced viral load, decreased lung lesions and inhibited progression to severe disease manifestations. This study underlines the potential for significant COVID-19 patient benefit for the SC31 antibody that justifies rapid advancement to the clinic, as well as highlighting the importance of appropriate mechanistic and functional studies during development. One Sentence SummaryAnti-SARS-CoV-2 IgG1 antibody SC31 controls infection in vivo by blocking SP:ACE2 binding and triggering a Fc-mediated anti-viral response.

microbiology↗

bioRxiv2020

A Sequence determinant in 3'UTR of mRNAs for Nuclear Retention by Paraspeckles

Paraspeckles are nuclear membraneless structures composed of a long non-coding RNA, Nuclear-Enriched-Abundant-Transcript-1 and RNA binding proteins, which associate numerous mRNAs. It is therefore believed that their cellular function is to sequester in the nucleus their associated proteins and/or target mRNAs. However, little is known about the molecular determinant in mRNA targets that allow their association to paraspeckles except that inverted repeats of Alu sequences (IRAlu) present in 3UTR of mRNAs may allow this association. While in a previous study we established the list of paraspeckle target RNAs in a rat pituitary cell line, we didnt find however, inverted repeated SINEs, the rat equivalent of primate IRAlus in 3UTR of these RNAs. By developing a candidate gene strategy, we selected a paraspeckle target gene, namely calreticulin mRNA, and we searched for other potential RNA recruitment elements in its 3UTR, since 3UTRs usually contain the sequence recognition for nuclear localization. We found a 15-nucleotide sequence, present as a tandem repeat in 3UTR of this mRNA, which is involved in the nuclear retention by paraspeckles. While being not overrepresented in the 3UTR of the paraspeckle target RNAs, this recruitment element was present in near 30% of all 3UTR mRNAs. In addition, since an oligonucleotide containing this sequence binds the paraspeckle protein component HNRNPK, it is proposed that HNRNPK may constitute a bridging protein between paraspeckles and target mRNAs.

molecular biology↗

bioRxiv2020

Structural basis of αE-catenin-F-actin catch bond behavior

Cell-cell and cell-matrix junctions transmit mechanical forces during tissue morphogenesis and homeostasis. -Catenin links cell-cell adhesion complexes to the actin cytoskeleton, and mechanical load strengthens its binding to F-actin in a direction-sensitive manner. This so-called catch bond behavior is described by a model in which force promotes a transition between weak and strong actin-bound states. We describe the cryo-electron microscopy structure of the F-actin-bound E-catenin actin-binding domain, which in solution forms a 5-helix bundle. Upon binding to actin, the first helix of the bundle dissociates and the remaining four helices and connecting loops rearrange to form the interface with actin. Deletion of the N-terminal helix produces strong actin binding in the absence of force. Our analysis explains how mechanical force applied to E-catenin or its homolog vinculin favors the strongly bound state, and the dependence of catch bond strength on the direction of applied force.

biophysics↗

bioRxiv2020

Semi-automated annotation of known and novel cancer long noncoding RNAs with the Cancer LncRNA Census 2 (CLC2)

Long noncoding RNAs play key roles in cancer and are at the vanguard of precision therapeutic development. These efforts depend on large and high-confidence collections of cancer lncRNAs. Here we present the Cancer LncRNA Census 2 (CLC2): at 492 cancer lncRNAs, it is 4-fold greater than its predecessor, without compromising on strict criteria of confident functional / genetic roles and inclusion in the GENCODE annotation scheme. This increase was enabled by leveraging high-throughput transposon insertional mutagenesis (TIM) screening data, yielding 95 novel cancer lncRNAs. CLC2 makes a valuable addition to existing collections: it is amongst the largest, holds the greatest number of unique genes, and carries functional labels (oncogene / tumour suppressor). Analysis of this dataset reveals that cancer lncRNAs are impacted by germline variants, somatic mutations, and changes in expression consistent with inferred disease functions. Furthermore, we show how clinical / genomic features can be used to vet prospective gene sets from high-throughput sources. The combination of size and quality makes CLC2 a foundation for precision medicine, demonstrating cancer lncRNAs evolutionary and clinical significance.

cancer biology↗

bioRxiv2020

Axial skeleton anterior-posterior patterning is regulated through feedback regulation between Meis transcription factors and retinoic acid

Vertebrate axial skeletal patterning is controlled by coordinated collinear expression of Hox genes and axial level-dependent activity of Hox protein combinations. Transcription factors of the Meis family act as cofactors of Hox proteins and profusely bind to Hox complex DNA, however their roles in mammalian axial patterning have not been established. Similarly, retinoic acid (RA) is known to regulate axial skeletal element identity through the transcriptional activity of its receptors, however, whether this role is related to Meis/Hox activity in axial patterning remains unknown. Here we study the role of Meis in axial skeleton formation and its relationship to the RA pathway by characterizing Meis1, Meis2 and Raldh2 mutant mice. Meis elimination produces axial skeleton defects without affecting Hox gene transcription, including vertebral homeotic transformations and rib mis-patterning associated to defects in the hypaxial myotome. While Raldh2 and Meis positively regulate each other, Raldh2 elimination largely recapitulates the defects associated to Meis-deficiency and Meis overexpression rescues the axial skeletal defects in Raldh2 mutants. We propose a Meis-RA positive feedback loop whose output is Meis levels and is essential to establish anterior-posterior identities and pattern of the vertebrate axial skeleton.

developmental biology↗

bioRxiv2020

Repeated colonization of caves leads to phenotypic convergence in catfishes (Siluriformes: Trichomycterus) at a small geographical scale

Across various animal groups, adaptation to the extreme conditions of cave environments has resulted in convergent evolution of morphological, physiological, and behavioral traits. We document a Neotropical cave fish system with ample potential to study questions related to convergent adaptation to cave environments at the population level. In the karstic region of the Andes of Santander, Colombia, cave-dwelling catfishes in the genus Trichomycterus exhibit variable levels of reduction of eyes and body pigmentation relative to surface congeners. We tested whether cave-dwelling, eye reduced, depigmented Trichomycterus from separate caves in Santander were the result of a single event of cave colonization and subsequent dispersal, or of multiple colonizations to caves by surface ancestors followed by phenotypic convergence. Using mitochondrial DNA sequences to reconstruct phylogenetic relationships of Trichomycterus from Santander, we found that caves in this region have been colonized independently by two separate clades. Additional events of cave colonization -and possibly recolonization of surface streams- may have occurred in one of the clades, where surface and cave-dwelling populations exhibit shallow mtDNA differentiation, suggesting recent divergence or divergence in the face of gene flow. We also identified various taxonomic challenges including both a considerable number of potentially undescribed species and likely problems with the circumscription of named taxa. The system appears especially promising for studies on a wide range of ecological and evolutionary questions.

evolutionary biology↗

bioRxiv2019

Engineering Phage Host-Range and Suppressing Bacterial Resistance Through Phage Tail Fiber Mutagenesis

The rapid emergence of antibiotic-resistant infections is prompting increased interest in phage-based antimicrobials. However, acquisition of resistance by bacteria is a major issue in the successful development of phage therapies. Through natural evolution and structural modeling, we identified host-range determining regions (HRDR) in the T3 phage tail fiber protein and developed a high-throughput strategy to genetically engineer these regions through site-directed mutagenesis. Inspired by antibody specificity engineering, this approach generates deep functional diversity (>107 different members), while minimizing disruptions to the overall protein structure, resulting in synthetic \"phagebodies\". We showed that mutating HRDRs yields phagebodies with altered host-ranges. Select phagebodies enable long-term suppression of bacterial growth by preventing the appearance of resistance in vitro and are functional in vivo using a mouse skin infection model. We anticipate this approach may facilitate the creation of next-generation antimicrobials that slow resistance development and could be extended to other viral scaffolds for a broad range of applications.\n\nHighlightsO_LIVastly diverse phagebody libraries containing 107 different members were created.\nC_LIO_LIStructure-informed engineering of viral tail fibers efficiently generated host-range alterations.\nC_LIO_LIPhagebodies prevented the development of bacterial resistance across long timescales in vitro and are functional in vivo.\nC_LI

synthetic biology↗

bioRxiv2018

Myc is dispensable for cardiac development in the mouse but can rescue Mycn-deficient hearts through functional replacement and cell competition

Myc is considered an essential transcription factor for heart development, but cardiac defects have only been studied in global Myc loss of function models. Here, we eliminated Myc by recombining a Myc floxed allele with the Nkx2.5Cre driver. We observed no anatomical, cellular or functional alterations in either fetuses or adult cardiac Myc-deficient mice. We re-examined Myc expression during development and found no expression in developing cardiomyocytes. In contrast, we confirmed that Mycn is essential for cardiomyocyte proliferation and cardiogenesis. Mosaic Myc overexpression in a Mycn-deficient background, shows that Myc can replace Mycn function, recovering heart development. We further show that this recovery involves the elimination of Mycn-deficient cells by Cell Competition. Our results indicate that Myc is dispensable during cardiogenesis and adult heart homeostasis and Mycn is exclusively responsible for cardiomyocyte proliferation during heart development. Nonetheless, our results show that Myc can functionally replace Mycn. We also show that cardiomyocytes compete according to their overall Myc+Mycn levels and that Cell Competition eliminates flawed cardiomyocytes, suggesting its relevance as a quality control mechanism in cardiac development.

developmental biology↗

bioRxiv2017

Live imaging of heart tube development in mouse reveals alternating phases of cardiac differentiation and morphogenesis

During vertebrate heart development two progenitor populations, first and second heart fields (FHF, SHF), sequentially contribute to longitudinal subdivisions of the heart tube (HT), with the FHF contributing the left ventricle and part of the atria, and the SHF the rest of the heart. Here we study the dynamics of cardiac differentiation and morphogenesis by tracking individual cells in live analysis of mouse embryos. We report that during an initial phase, FHF precursors differentiate rapidly to form a cardiac crescent, while limited morphogenesis takes place. In a second phase, no differentiation occurs while extensive morphogenesis, including splanchnic mesoderm sliding over the endoderm, results in HT formation. In a third phase, cardiac precursor differentiation resumes and contributes to SHF-derived regions and the dorsal closure of the HT. These results reveal tissue-level coordination between morphogenesis and differentiation during HT formation and provide a new framework to understand heart development.

developmental biology↗