Biology subjects

Morales, A.

Publications and source records attributed to Morales, A..

At least 19 recordsLinked to original sources

bioRxiv2026

Emergence of tolerance and avoidance strategies from local and systemic responses to nitrogen: insights from modelling of auxin-mediated root plasticity

Understanding how root phenotypic plasticity enhances resource-use efficiency can help understand the outcomes of plant competition and identify suitable genotypes for medium- to low-input agricultural systems. Auxin regulates multiple root growth processes, including the root architectural responses to nitrogen (N). We explored the extent to which auxin-mediated local and systemic responses to external and internal N influences plant N uptake and use, using a functional-structural-plant (FSP) modelling approach. A simplified auxin module was developed at the level of the organ and integrated into an FSP model to represent physiological plastic root responses to N. Model performance was evaluated against experimental data. We then ran simulations under various N conditions with local or systemic responses enabled or disabled, to quantify their contribution to N uptake and use. Simulations showed that local auxin responses enhanced N uptake by distributing more roots towards deeper soil layers and increased N forage, thereby avoiding N stress. Systemic auxin responses reduced N uptake by distributing more roots near the top soil layers, which reduced plant size and N demand, thereby enhancing stress tolerance. This points to a trade-off in N uptake between the tolerance and avoidance strategies, which can be traced back to biomass and N investment resulting from source-sink relationships in the plant. Representing hormone-mediated root plasticity in an FSP model provides mechanistic insights into plant strategies for resource capture.

plant biology↗

bioRxiv2026

Quantifying the effect of cereal plant trait plasticity on weed suppression in intercrops

In cereal-legume intercrops, weed suppression is primarily driven by cereals, whose competitiveness is shaped by trait plasticity--morphological adjustments in response to the intercrop environment. However, how individual cereal traits respond plastically and contribute to system performance remains unclear, hampering improvements through breeding or system design. We combined field experiments with functional-structural plant modelling to quantify plastic responses of four cereal traits (tiller number, tiller angle, specific leaf area (SLA), and specific internode length (SIL)) and their effects on weed suppression and crop productivity. Field measurements revealed plasticity in tiller number, tiller angle, and SIL between sole crops and intercrops, while SLA showed minimal differences. Simulations showed that intermediate tiller numbers resulted in the strongest weed suppression and highest productivity, indicating an optimum, while more horizontal tillers suppressed weeds slightly better than vertical ones. Weed suppression increased with higher SLA values, while SIL showed a saturating response, increasing to intermediate SIL values and plateauing thereafter. In simulations with short-statured cereal phenotypes (low SIL), the reduction in cereal weed suppression was compensated by the legume component. This study demonstrates how FSP modelling can be used to investigate trait plasticity mechanisms and generate testable hypotheses about trait effects in complex intercrop systems. HighlightCereal trait plasticity shapes weed suppression in cereal-legume intercrops, with distinct response patterns per trait, while legumes can compensate for weakly competitive cereals, suggesting balanced competition over cereal dominance.

plant biology↗

bioRxiv2025

ScAPE: A lightweight multitask learning baseline method to predict transcriptomic responses to perturbations

Predicting biological responses to perturbations is essential for advancing drug development efforts. While high-throughput technologies have greatly enhanced our ability to study perturbation effects at the molecular level, accurately modeling these responses remains challenging due to the complex underlying molecular interactions, the vast dimensionality of cellular response, the limited amount of data, and technical limitations in data collection. Machine learning (ML) approaches have emerged as powerful tools for addressing these challenges. However, the observation in recent benchmarks that complex models do not consistently outperform simpler ones highlights the need for robust and efficient baselines for evaluating new methods. We present ScAPE (Single Cell Analysis of Perturbational Effects), an ML method for predicting drug-induced gene expression changes. ScAPE leverages only aggregated gene-level statistics across cell types and perturbations as input features, yet achieves good performance despite this simplicity. Its effectiveness was demonstrated in the Single-cell Perturbation Prediction NeurIPS 2023 challenge, where it ranked among the top-performing methods. We describe the core model and introduce extensions that enhance its utility as a baseline, including a multitask learning extension for simultaneous prediction of multiple transcriptional response metrics. A software API is provided to support adoption and integration into research pipelines. Benchmarking against other leading challenge entries and general-purpose ML approaches, using the original challenge metric, supports ScAPE as a simple, lightweight baseline for evaluating methods that predict perturbation effects at the gene-expression level. ScAPE is built using Keras 3, enabling support for different backends such as TensorFlow, JAX, or PyTorch, and freely available as an open-source package at github.com/scapeML/scape.

bioinformatics↗

bioRxiv2025

Quantitative Membrane Binding Assays Reveal an Inhibitory Role for the BRAF-Specific Region in CRD and Lipid Interaction

BRAF is a serine/threonine kinase and a central effector of the mitogen-activated protein kinase (MAPK) signaling pathway, frequently mutated in cancer. Its activation is tightly controlled by autoinhibitory mechanisms that regulate membrane recruitment and dimerization. The BRAF-specific region (BSR), located at the N-terminus, is known to promote isoform-preferred RAS binding and facilitate dimerization with kinase suppressor of RAS (KSR), yet its role in regulating lipid interaction has remained unexplored. Here, we identify the BSR as a previously unrecognized inhibitory module that attenuates lipid binding by the cysteine-rich domain (CRD). Using quantitative in vitro reconstitution with supported lipid bilayers and fluorescence microscopy, we demonstrate that the BRAF CRD exhibits high intrinsic affinity for phosphatidylserine-rich membranes, but the inclusion of the BSR markedly reduces the membrane binding. We further demonstrate that the inhibitory function of the BSR correlates with its global electrostatic properties rather than a single defined sequence motif. This inhibitory effect of BSR was corroborated in live cells by quantifying plasma membrane localization of BRAF constructs, including the full-length protein. When canonical autoinhibition of CRD--mediated by sequestration within the 14-3-3 dimer--is disrupted by oncogenic mutation or RAF inhibitor treatment, the BSR assumes a compensatory role in repressing CRD-lipid interaction. This additional regulatory layer provided by the BSR prevents RAS-independent membrane recruitment under both physiological and pathological conditions. Broad Impact StatementProtein-lipid interactions are a fundamental mechanism for regulating the localization and activity of signaling proteins. This study reveals that the N-terminal segment of BRAF acts as an inhibitory module that suppresses lipid engagement by CRD, particularly when canonical autoinhibition is disrupted by oncogenic mutation or inhibitor treatment. This additional layer of regulation provides new insight into BRAF membrane dynamics and may have implications in therapeutic intervention of dysregulated BRAF signaling.

biochemistry↗

bioRxiv2025

Gut microbial ecosystems differ across metabolic and obesity phenotypes

Obesity is a heterogeneous condition comprising a continuum of phenotypes with various metabolic and inflammatory profiles. Metabolically healthy obesity (MHO) identifies individuals with obesity but a relatively preserved metabolic state. However, the criteria defining MHO remain inconsistent, and little is known about the gut microbiome (GM) features underlying this intermediate phenotype. Here, we aim to describe microbial structures contributing to metabolic health and disease. To do so, we analyzed the GM of 959 individuals classified as metabolically healthy non-obese (MHNO), MHO, metabolically unhealthy non-obese (MUNO), and metabolically unhealthy obese (MUO), using stool shotgun metagenomics. MHO subjects display intermediate anthropometric and biochemical profiles, with a GM composition and diversity in an in-between state among MHNO and MUO individuals. Network science analyses reveal that metabolic health, rather than obesity, drives microbial connectivity: MHNO and MHO individuals harbor more robust and functionally cohesive microbial networks, whose most influential nodes are focused toward SCFA production. In contrast, MUO and MUNO communities exhibit a dysbiotic state with reduced connectivity and increased influence of low-abundance, ectopic and potentially pro-inflammatory species resulting in a damaged, unstable microbial community network. These findings suggest that metabolic disorders disrupt microbial ecology beyond compositional shifts, emphasizing the need for systems-level approaches. Our findings show differences in microbial connectivity and association patterns across metabolic and obesity phenotypes, shedding light on how distinct microbial structures may contribute to metabolic health and disease.

bioinformatics↗

bioRxiv2025

Positive cooperativity between RAS-binding and cysteine-rich domains regulates RAF membrane binding kinetics via lateral rebinding

RAF activation requires interactions with both RAS nanoclusters and membrane lipids, yet the molecular basis of this process remains unclear. Using a bottom-up reconstitution approach, we show how coordinated protein-protein and protein-lipid interactions regulate membrane binding dynamics of RAF to drive its multistep activation. Within membrane environments, the RAS-binding domain (RBD) and cysteine-rich domain (CRD) exhibit cooperativity, with CRD-mediated phosphatidylserine binding stabilizing the RBD:RAS complex. Importantly, RAF remains membrane-bound through lateral rebinding to RAS, where a weak CRD-lipid interaction plays an essential role. This lateral rebinding extends RAFs membrane dwell time under high RAS density conditions, which are found in RAS nanoclusters. This prolonged membrane residence likely facilitates kinetic proofreading of RAFs multistep activation within RAS nanoclusters, ensuring signaling specificity. Given the high abundance of weak multivalent membrane interactions, lateral rebinding may be a common mechanism for regulating the activity of signaling proteins through sustained membrane retention.

biochemistry↗

bioRxiv2025

Isolation and primary culture of Octopus vulgaris cells: Assessment of their proliferative capacity

The octopus cell culture system represents a useful tool in various fields, such as biological studies and sustainable octopus production. In this paper, we describe the isolation procedure and culture protocols, metabolic activity, and the proliferative capacity assessment of the common octopus (Octopus vulgaris) cells, at three different life stages: embryo, paralarvae and adult. Isolation of cells was carried out with minimum essential medium (MEM) or Leibovitz L-15 medium, using 0.4 % type I collagenase for 2 (paralarvae and embryo) or 3 h (adult) at 25 {degrees}C. Isolated cells showed significant metabolic activity, demonstrating their viability for in vitro studies. The culture of isolated cells in MEM or L-15 supplemented with different sera (4 % fetal bovine serum (FBS), 10 % FBS and 5 % haemolymph (HEMO)) was viable for 168 h, although the survival of different cell types was variable. A detachment protocol using trypsin was also successfully developed. Finally, isolated muscle cells from both paralarval and adult specimens were marked with the mitotic indicator PHH3. Results from flow cytometry analysis showed that all developmental stages examined contained cells in mitotic phase, with embryos XII-XIII showing the highest proportion. This study provides novel guidelines for the in vitro culture of O. vulgaris cells, which are useful for the future development of cellular aquaculture, understood as the production of lab-grown meat for this species. Moreover, the results reported in this communication could contribute to reduce the use of animals for scientific purposes as well as providing a new option for human consumption of cephalopods.

cell biology↗

bioRxiv2024

Comparative single-cell analyses reveal evolutionary repurposing of a conserved gene program in bat wing development

Bats are the only mammals capable of self-powered flight, an evolutionary innovation based on the transformation of forelimbs into wings. The bat wing is characterized by an extreme elongation of the second to fifth digits and a wing membrane called chiropatagium connecting them. Here we investigated the developmental and cellular origin of this structure by comparing bat and mouse limbs using omics tools and single-cell analyses. Despite the substantial morphological differences between the species, we observed an overall conservation of cell populations and gene expression patterns including interdigital apoptosis. Single-cell analyses of micro-dissected embryonic chiropatagium identified a specific fibroblast population, independent of apoptosis-associated interdigital cells, as the origin of this tissue. These distal cells express a conserved gene program including the transcription factors MEIS2 and TBX3, which are commonly known to specify and pattern the early proximal limb. Transgenic ectopic expression of MEIS2 and TBX3 in mouse distal limb cells resulted in the activation of genes expressed during wing development and phenotypic changes related to wing morphology, such as the fusion of digits. Our results elucidate fundamental molecular mechanisms of bat wing development and illustrate how drastic morphological changes can be achieved through repurposing of existing developmental programs during evolution.

evolutionary biology↗

bioRxiv2024

The Virtual Plant Laboratory: a modern plant modeling framework in Julia

The Virtual Plant Laboratory (VPL) is a novel software for building, simulating, and visualizing functional- structural plant (FSP) models. FSP models focus on the interactions between plant structure, internal physiological processes, and the biotic and abiotic environment. VPL is built in the Julia programming language and is designed to be a flexible and extensible platform for FSP modeling. Using Julia brings the advantage that only one programming language is required for the whole modeling cycle as Julia is as fast as compiled languages but also dynamic as interpreted languages. VPL provides a graph rewriting system for building dynamic models of plant growth and development, an interactive 3D visualization system and a Monte Carlo ray tracer for simulating radiation interception by plant canopies. In this paper, we introduce VPL, highlighting the main components, modeling paradigms, and design decisions behind it, as well as a future roadmap for further development. We also present a short case study of a model for intercropping of legumes and cereals that was built fully with VPL, as an example of what can be built with this software. VPL is fully open source and available in all common computing platforms for anyone to use. Full documentation and tutorials are available at https://virtualplantlab.com.

plant biology↗

bioRxiv2024

Oxydifficidin, a potent Neisseria gonorrhoeae antibiotic due to DedA assisted uptake and ribosomal protein RplL sensitivity

Gonorrhea, which is caused by Neisseria gonorrhoeae, is the second most reported sexually transmitted infection worldwide. The increasing appearance of isolates that are resistant to approved therapeutics raises the concern that gonorrhea may become untreatable. Here, we serendipitously identified oxydifficidin as a potent N. gonorrhoeae antibiotic through the observation of a Bacillus amyloliquefaciens contaminant in a lawn of N. gonorrhoeae. Oxydifficidin is active against both wild- type and multidrug-resistant N. gonorrhoeae. Its potent activity results from a combination of DedA- assisted uptake into the cytoplasm and the presence of an oxydifficidin-sensitive ribosomal protein L7/L12 (RplL). Our data indicates that oxydifficidin binds to the ribosome at a site that is distinct from other antibiotics and that L7/L12 is uniquely associated with its mode of action. This study opens a potential new avenue for addressing antibiotic resistant gonorrhea and underscores the possibility of identifying overlooked natural products from cultured bacteria, particularly those with activity against previously understudied pathogens.

microbiology↗

bioRxiv2024

Microbiome-induced Increases and Decreases in Bone Tissue Strength can be Initiated After Skeletal Maturity

Recent studies in mice have indicated that the gut microbiome can regulate bone tissue strength. However, prior work involved modifications to the gut microbiome in growing animals and it is unclear if the same changes in the microbiome, applied later in life, would change matrix strength. Here we changed the composition of the gut microbiome before and/or after skeletal maturity (16 weeks of age) using oral antibiotics (ampicillin + neomycin). Male and female mice (n=143 total, n=12-17/group/sex) were allocated into five study groups:1) Unaltered, 2) Continuous (dosing 4-24 weeks of age), 3) Delayed (dosing only 16-24 weeks of age), 4) Initial (dosing 4-16 weeks of age, suspended at 16 weeks), and 5) Reconstituted (dosing from 4-16 weeks following by fecal microbiota transplant from Unaltered donors). Animals were euthanized at 24 weeks of age. In males, bone matrix strength in the femur was 25-35% less than expected from geometry in mice from the Continuous (p= 0.001), Delayed (p= 0.005), and Initial (p=0.040) groups as compared to Unaltered. Reconstitution of the gut microbiota, however, led to a bone matrix strength similar to Unaltered animals (p=0.929). In females, microbiome-induced changes in bone matrix strength followed the same trend as males but were not significantly different, demonstrating sex-related differences in the response of bone matrix to the gut microbiota. Minor differences in chemical composition of bone matrix were observed (Raman spectroscopy). Our findings indicate that microbiome-induced impairment of bone matrix in males can be initiated and/or reversed after skeletal maturity. The portion of the femoral cortical bone formed after skeletal maturity (16 weeks) is small; however, this suggests that microbiome-induced changes in bone matrix occur without osteoblast/osteoclast turnover using an, as of yet unidentified mechanism. These findings add to evidence that the mechanical properties of bone matrix can be altered in the adult skeleton.

bioengineering↗

bioRxiv2023

A Parasite Odyssey: An RNA virus concealed in Toxoplasma gondii

We are entering a "Platinum Age of Virus Discovery", an era marked by exponential growth in the discovery of virus biodiversity, and driven by advances in metagenomics and computational analysis. In the ecosystem of a human (or any animal) there are more species of viruses than simply those directly infecting the animal cells. Viruses can infect all organisms constituting the microbiome, including bacteria, fungi, and unicellular parasites. Thus the complexity of possible interactions between host, microbe, and viruses is unfathomable. To understand this interaction network we must employ computationally-assisted virology as a means of analyzing and interpreting the millions of available samples to make inferences about the ways in which viruses may intersect human health. From a computational viral screen of human neuronal datasets, we identified a novel narnavirus Apocryptovirus odysseus (Ao) which likely infects the neurotropic parasite Toxoplasma gondii. Previously, several parasitic protozoan viruses (PPVs) have been mechanistically established as triggers of host innate responses, and here we present in silico evidence that Ao is a plausible pro-inflammatory factor in human and mouse cells infected by T. gondii. T. gondii infects billions of people worldwide, yet the prognosis of toxoplasmosis disease is highly variable, and PPVs like Ao could function as a hitherto undescribed hypervirulence factor. In a broader screen of over 7.6 million samples, we explored phylogenetically-proximal viruses to Ao and discovered 19 Apocryptovirus species, all found in libraries annotated as vertebrate transcriptome or metatranscriptomes. While the Narnavirus samples making up this genus-like clade are derived from sheep, goat, bat, rabbit, chicken, and pigeon samples, the presence of virus is strongly predictive of parasitic (Apicomplexa) nucleic acid co-occurrence, supporting that these are a clade of parasite-infecting viruses. This is a computational proof-of-concept study in which we rapidly analyze millions of datasets from which we distilled a mechanistically, ecologically, and phylogenetically refined hypothesis. We predict this highly diverged Ao RNA virus is biologically a T. gondii infection, and that Ao, and other viruses like it, will modulate this disease which afflicts billions worldwide.

microbiology↗

bioRxiv2022

Octopamine metabolically reprograms astrocytes to confer neuroprotection against α-synuclein

Octopamine is a well-established invertebrate neurotransmitter involved in fight-or-flight responses. In mammals, its function was replaced by norepinephrine. Nevertheless, it is present at trace amounts and can modulate the release of monoamine neurotransmitters by a yet unidentified mechanism. Here, through a multidisciplinary approach utilizing in vitro and in vivo models of -synucleinopathy, we uncovered an unprecedented role for octopamine in driving the conversion from toxic to neuroprotective astrocytes in the cerebral cortex by fostering aerobic glycolysis. Physiological levels of neuron-derived octopamine act on astrocytes via a TAAR1-Orai1-Ca2+-calcineurin-mediated signaling pathway to stimulate lactate secretion. Lactate uptake in neurons via the MCT2-calcineurin-dependent pathway increases ATP and prevents neurodegeneration. Pathological increases of octopamine caused by -synuclein halts lactate production in astrocytes and short-circuits the metabolic communication to neurons. Our work provides a novel function of octopamine as a modulator of astrocyte metabolism and subsequent neuroprotection with implications to -synucleinopathies.

neuroscience↗

bioRxiv2022

Bat pluripotent stem cells reveal unique entanglement between host and viruses

Bats have evolved features unique amongst mammals, including flight, laryngeal echolocation, and certain species have been shown to have a unique immune response that may enable them to tolerate viruses such as SARS-CoVs, MERS-CoVs, Nipah, and Marburg viruses. Robust cellular models have yet to be developed for bats, hindering our ability to further understand their special biology and handling of viral pathogens. To establish bats as new model study species, we generated induced pluripotent stem cells (iPSCs) from a wild greater horseshoe bat (Rhinolophus ferrumequinum) using a modified Yamanaka protocol. Rhinolophids are amongst the longest living bat species and are asymptomatic carriers of coronaviruses, including one of the viruses most closely related to SARS-CoV-2. Bat induced pluripotent stem (BiPS) cells were stable in culture, readily differentiated into all three germ layers, and formed complex embryoid bodies, including organoids. The BiPS cells were found to have a core pluripotency gene expression program similar to that of other species, but it also resembled that of cells attacked by viruses. The BiPS cells produced a rich set of diverse endogenized viral sequences and in particular retroviruses. We further validated our protocol by developing iPS cells from an evolutionary distant bat species Myotis myotis (greater mouse-eared bat) non-lethally sampled in the wild, which exhibited similar attributes to the greater horseshoe bat iPS cells, suggesting that this unique pluripotent state evolved in the ancestral bat lineage. Although previous studies have suggested that bats have developed powerful strategies to tame their inflammatory response, our results argue that they have also evolved mechanisms to accommodate a substantial load of endogenous viral sequences and suggest that the natural history of bats and viruses is more profoundly intertwined than previously thought. Further study of bat iPS cells and their differentiated progeny should advance our understanding of the role bats play as virus hosts, provide a novel method of disease surveillance, and enable the functional studies required to ascertain the molecular basis of bats unique traits.

developmental biology↗

bioRxiv2022

Integrating gene annotation with orthology inference at scale

Annotating coding genes and inferring orthologs are two classical challenges in genomics and evolutionary biology that have traditionally been approached separately, limiting scalability. We present TOGA, a method that integrates structural gene annotation and orthology inference. TOGA implements a different paradigm to infer orthologous loci, improves ortholog detection and annotation of conserved genes compared to state-of-the-art methods, and handles even highly-fragmented assemblies. TOGA scales to hundreds of genomes, which we demonstrate by applying it to 488 placental mammal and 501 bird assemblies, creating the largest comparative gene resources so far. Additionally, TOGA detects gene losses, enables selection screens, and automatically provides a superior measure of mammalian genome quality. Together, TOGA is a powerful and scalable method to annotate and compare genes in the genomic era.

genomics↗

bioRxiv2021

Effects of sub-lethal single, simultaneous, and sequential abiotic stresses on phenotypic traits of Arabidopsis thaliana

Plant responses to abiotic stresses are complex and dynamic, and involve changes in different traits, either as the direct consequence of the stress, or as an active acclimatory response. Abiotic stresses frequently occur simultaneously or in succession, rather than in isolation. Despite this, most studies have focused on a single stress and single or few plant traits. To address this gap, our study comprehensively and categorically quantified the individual and combined effects of three major abiotic stresses associated with climate change (flooding, progressive drought and high temperature) on 12 phenotypic traits related to morphology, development, growth and fitness, at different developmental stages in four Arabidopsis thaliana accessions. Combined sub-lethal stresses were applied either simultaneously (high temperature and drought) or sequentially (flooding followed by drought). In total, we analyzed the phenotypic responses of 1782 individuals across these stresses and different developmental stages. Overall, abiotic stresses and their combinations resulted in distinct patterns of effects across the traits analyzed, with both quantitative and qualitative differences across accessions. Stress combinations had additive effects on some traits, whereas clear positive and negative interactions were observed for other traits: 9 out of 12 traits for high temperature and drought, 6 out of 12 traits for post-submergence and drought showed significant interactions. In many cases where the stresses interacted, the strength of interactions varied across accessions. Hence, our results indicated a general pattern of response in most phenotypic traits to the different stresses and stress combinations, but it also indicated a natural genetic variation in the strength of these responses. Overall, our study provides a rich characterization of trait responses of Arabidopsis plants to sub-lethal abiotic stresses at the phenotypic level and can serve as starting point for further in-depth physiological research and plant modelling efforts.

plant biology↗

bioRxiv2021

Gene losses in the common vampire bat illuminate molecular adaptations to blood feeding

Feeding exclusively on blood, vampire bats represent the only obligate sanguivorous lineage among mammals. To uncover genomic changes associated with adaptations to this unique dietary specialization, we generated a new haplotype-resolved reference-quality genome of the common vampire bat (Desmodus rotundus) and screened 26 bat species for genes that were specifically lost in the vampire bat lineage. We discovered previously-unknown gene losses that relate to metabolic and physiological changes, such as reduced insulin secretion (FFAR1, SLC30A8), limited glycogen stores (PPP1R3E), and a distinct gastric physiology (CTSE). Other gene losses likely reflect the biased nutrient composition (ERN2, CTRL) and distinct pathogen diversity of blood (RNASE7). Interestingly, the loss of REP15 likely helped vampire bats to adapt to high dietary iron levels by enhancing iron excretion and the loss of the 24S-hydroxycholesterol metabolizing enzyme CYP39A1 could contribute to their exceptional cognitive abilities. Finally, losses of key cone phototransduction genes (PDE6H, PDE6C) suggest that these strictly-nocturnal bats completely lack cone-based vision. These findings enhance our understanding of vampire bat biology and the genomic underpinnings of adaptations to sanguivory.

evolutionary biology↗

bioRxiv2021

Defining dysfunction due to loss of MECP2 in Rett Patient Brain

Rett Syndrome is characterized by a postnatal loss of neurophysiological function and regression of childhood development. Because the syndrome is X-linked and males with MECP2 mutations generally do not survive birth, the study of this syndrome has been complicated by the fact that in female brain, a portion of neurons express wild type MECP2, and another portion express a non-functional allele of MECP2. Therefore, bulk-RNA-sequencing of Rett brain is confounded by the presence of chimerism of neurons for functional MECP2 in neurons. We developed an approach that allows for single-nuclei transcriptional profiling of individual neurons and a direct comparison between neurons that express functional MECP2 with those that express the disease-causing allele. We found that mutant neurons from Rett brain show patterns of aberrant expression of synaptic and metabolic genes, both of which can be detected in in vitro models of Rett Syndrome. We used these resources to identify a role for POU2F1/OCT1 transcription factor in mediating the response to stress due to loss of MECP2, highlighting a potential key molecular regulator of stress in Rett neurons. Together, our new sorting approach enables us to highlight defective molecular and metabolic pathways in Rett brain neurons and suggests that in vitro models could serve as valuable tools to further study this syndrome and potentially for development of novel therapeutics.

molecular biology↗