Biology subjects

Donati, G.

Publications and source records attributed to Donati, G..

14 recordsLinked to original sources

bioRxiv2026

Functional validation of allele-specific LMNB1 silencing in patient-derived astrocytes as a therapeutic option for Autosomal Dominant Leukodystrophy

Adult-onset Autosomal Dominant Leukodystrophy (ADLD) is a rare fatal leukodystrophy caused by increased LMNB1 gene dosage, most commonly resulting from duplication of the LMNB1 locus. Because ADLD is a gene dosage disorder, selective reduction of pathological LMNB1 expression represents a rational therapeutic strategy. Although allele-specific RNA interference has previously been shown to lower LMNB1 levels in patient-derived fibroblasts and directly reprogrammed neurons, its therapeutic effects have not been evaluated in disease-relevant human glial cells or using functional efficacy endpoints. Here, we established human induced pluripotent stem cell-derived astrocytes from ADLD patients as a human glial model in which to validate allele-specific LMNB1 silencing across molecular, cellular, and functional readouts. ADLD astrocytes recapitulated increased LMNB1 expression and characteristic nuclear abnormalities and displayed transcriptional alterations affecting extracellular matrix organization, calcium homeostasis, metabolism and RNA processing. Functionally, these cells also exhibited functional phenotypes suitable for therapeutic evaluation: astrocyte-conditioned medium impaired the viability of both murine and human oligodendroglial cultures, while conditioned-medium and direct astrocyte-seeding paradigms revealed impaired post-lesion myelin recovery in lysolecithin-treated cerebellar organotypic slices. Allele-specific LMNB1 silencing restored physiological LMNB1 levels, corrected nuclear abnormalities, attenuated astrocyte-mediated oligodendroglial toxicity, improved post-lesion myelin recovery, and was associated with selective transcriptional programs associated with extracellular support and cholesterol metabolism. Together, these findings provide molecular, cellular, and functional validation of allele-specific LMNB1 dosage correction in patient-derived human astrocytes and offer key support for LMNB1-lowering strategies in disease-relevant human glial cells.

neuroscience↗

bioRxiv2023

Spatial and single-cell transcriptomics illuminate bat immunity and barrier tissue evolution

The Egyptian fruit bat displays tolerance to lethal viruses and unique dietary adaptations, but the molecular basis for this is poorly understood. To this end, we generated detailed maps of bat gut, lung and blood cells using spatial and single-cell transcriptomics. We compared bat with mouse and human cells to reveal divergence in genetic programs associated with environmental interactions and immune responses. Complement system genes are transcriptionally divergent, uniquely expressed in bat lung and gut epithelium, and undergo rapid coding-sequence evolution. Specifically in the tip of the gut villus, bat enterocytes express evolutionarily young genes while lacking expression of genes related to specific nutrient absorption. Profiling immune stimulation of PBMCs revealed a monocyte subset with conserved cross-species interferon expression, suggesting strong constraints to avoid an excessive immune response. Our study thus uncovers conserved and divergent immune pathways in bat tissues, providing a unique resource to study bat immunity and evolution.

genomics↗

bioRxiv2023

A comparative analysis of the antiviral response in two bat species reveals conserved and divergent innate immune pathways

Bats host a range of viruses that cause severe disease in humans without displaying clinical symptoms to these infections. The mechanisms of bat adaptation to these viruses are a continuous source of interest but remain largely unknown. To understand the landscape of bat antiviral response in a comprehensive and comparative manner, we studied this response in two bat species - the Egyptian fruit bat and the insectivore Kuhls pipistrelle, representing the two major bat subordinal clades. We profiled the transcriptional response to dsRNA - that triggers a rapid innate immune response - in skin fibroblasts from a large cohort of replicates from each bat species, using RNA-sequencing, and compared bat response with responses in primates and rodents. Both bat species upregulate a similar set of genes, many of which are known to be involved in the antiviral response across mammals. However, a subset of these genes is transcriptionally divergent in response between the two bat species. These transcriptionally divergent genes also evolve rapidly in coding sequence across the bat clade and have particular regulatory and functional characteristics, including specific promoter architectures and association with expression programs thought to underlie tolerance and resistance in response to viral infection. In addition, using single-cell transcriptomics, we show that transcriptionally divergent genes display high expression variability between individual cells. A focused analysis of dsRNA-sensing pathways further points to significant differences between bat and human in basal expression of genes important for triggering antiviral responses. Finally, a survey of genes recently lost or duplicated in bats points to a limited set of antiviral genes that have undergone rapid gene loss or gain in bats, with the latter group resulting in paralogs displaying divergence in both coding sequence and expression in bat tissues. Our study reveals a largely conserved regulatory program of genes upregulated in response to viral infection across bats and other mammals, and points to a set of genes that evolved rapidly in bats through multiple evolutionary mechanisms. This divergence can contribute to bat adaptation to viral infection and provides directions to understanding the mechanisms behind it.

evolutionary biology↗

bioRxiv2023

Lifelong tissue memory relies on spatially organised dedicated progenitors located distally from the injury

It is believed epithelial cells that have participated in a wound repair elicit a more efficient but locally restricted response to future injuries. However here we show that the cell adaptation resulting from a localised tissue damage has a wide spatial impact at a scale not previously noticed. We demonstrate that away from injured site, after a first injury a specific epithelial stem cell population gives rise to long term wound-memory progenitors residing in their own niche of origin. Notably these progenitors have not taken part in the first wound healing but become pre-activated through priming. This adaptation differs from classical features of trained immunity previously shown to be adopted by other epithelial stem cells. Our newly identified wound-distal memory cells display a cell-autonomous transcriptional pre-activated state leading to an enhanced wound repair ability that can be partially recapitulated through epigenetic perturbation even in absence of an injury. Importantly, the harmful consequences of wound repair, such as exacerbated tumorigenesis, occur within these primed cells and follow their spatial distribution. Overall, we show that sub-organ scale adaptation of an injury relies on spatially organised and memory-dedicated progenitors, characterised by an epigenetic actionable cell state, that predisposes to tumour onset.

cell biology↗

bioRxiv2023

HuR modulation with tanshinone mimics impairs LPS response in murine macrophages

Lipopolysaccharide exposure to macrophages induces an inflammatory response that is heavily regulated at the transcriptional and post-transcriptional levels. HuR (ELAVL1) is an RNA binding protein that binds and regulates the maturation and half-life of AU/U rich elements (ARE) containing cytokines and chemokines transcripts, mediating the LPS-induced response. Here we investigated how and to what extent small molecule tanshinone mimics (TMs) inhibiting HuR-RNA interaction counteract LPS stimulus in macrophages. We show TMs exist in solution in keto-enolic tautomerism and that, by molecular dynamic calculations, the orto quinone form is the bioactive species interacting with HuR and inhibiting its binding mode vs mRNA targets. A chemical blockage of the diphenolic, reduced form as a diacetate caused the loss of activity of TMs in vitro but resulted to prodrug-like activity in vivo. The murine macrophage cell line RAW264.7 was treated with LPS and TMs, and the modulation of cellular LPS-induced response was monitored by RNA and Ribonucleoprotein immunoprecipitation sequencing. Correlation analyses indicated that LPS induced a strong coupling between differentially expressed genes and HuR-bound genes, and that TMs reduced such interactions. Functional annotation addressed a specific set of genes involved in chemotaxis and immune response, such as Cxcl10, Il1b, Cd40, and Fas, with a decreased association with HuR, a reduction of their expression and protein secretion. The same effect was observed in primary murine bone marrow-derived macrophages, and in vivo in an LPS induced peritonitis model, in which the serum level of Cxcl10 and Il1b was strongly reduced, endowing TMs such as TM7nox with remarkable anti-inflammatory properties in vivo.

molecular biology↗

bioRxiv2022

Molecular and spatial design of early skin development

A wealth of specialized cell populations within the skin facilitates its hair producing, protective, sensory and thermoregulatory functions, but how the vast cell-type diversity and tissue architecture develops is largely unexplored. Here, with single-cell transcriptomics, spatial cell-type assignment and cell-lineage tracing we deconstruct early embryonic mouse skin during the key transitions from seemingly uniform developmental precursor states to a multilayered, multilineage epithelium and complex dermal identity. We reveal the spatiotemporal emergence of hair-follicle-inducing, muscle-supportive, and fascia-forming fibroblasts. We also uncover the formation of the panniculus carnosus muscle, sprouting blood vessels without pericyte coverage, and the earliest residence of mast and dendritic immune cells in skin. Finally, we reveal an unexpected epithelial heterogeneity within the early single-layered epidermis and a signaling-rich periderm layer. Overall, this cellular blueprint of early skin development establishes histological landmarks - essential for placing cells in their spatial tissue context - and highlights unprecedented dynamical interactions among skin cells.

developmental biology↗

bioRxiv2022

Oxidative stress potentiates the therapeutic action of a mitochondrial complex I inhibitor in MYC-driven B-cell lymphoma

MYC is a key oncogenic driver and an adverse prognostic factor in multiple types of cancer, including diffuse large B-cell lymphoma (DLBCL). Yet, MYC activation also endows cancer cells with a series of metabolic dependencies, which can provide strategic points for targeted pharmacological intervention. We recently reported that targeting the mitochondrial electron transport chain (ETC) complex I with the small molecule inhibitor IACS-010759 selectively killed MYC-overexpressing lymphoid cells. Here, we unravel the mechanistic basis for this synthetic-lethal interaction and exploit it to improve the anti-tumoral effects of ETC inhibition. In a mouse B-cell line, MYC hyperactivation and IACS-010759 treatment added up to induce oxidative stress, with consequent depletion of reduced glutathione and lethal disruption of redox homeostasis. This effect could be enhanced by targeted pharmacological intervention, with either inhibitors of NADPH production through the pentose phosphate pathway, or with ascorbate (vitamin C), known to contribute pro-oxidant effects when administered at high doses. In these conditions, ascorbate synergized with IACS-010759 to kill MYC-overexpressing cells in vitro and reinforced its therapeutic action against human B-cell lymphoma xenografts. Hence, ETC inhibition and high-dose ascorbate might improve the outcome of patients affected by high-grade lymphomas and other MYC-driven cancers. Key point #1MYC and the ETC complex I inhibitor IACS-010759 elicit different reactive oxygen species (ROS) that cooperate to disrupt redox homeostasis. Key point #2Further boosting of oxidative stress with high doses of ascorbate increases the killing of MYC-driven lymphoma xenografts by IACS-010759.

cancer biology↗

bioRxiv2021

Temporal pattern recognition in the human brain: a dual simultaneous processing

Pattern recognition is a major scientific topic. Strikingly, while machine learning algorithms are constantly refined, the human brain emerges as an ancestral biological example of such complex procedure. However, how it transforms sequences of single objects into meaningful temporal patterns remains elusive. Using magnetoencephalography (MEG) and magnetic resonance imaging (MRI), we discovered and mathematically modelled an inedited dual simultaneous processing responsible for pattern recognition in the brain. Indeed, while the objects of the temporal pattern were independently elaborated by a local, rapid brain processing, their combination into a meaningful superordinate pattern depended on a concurrent global, slower processing involving a widespread network of sequentially active brain areas. Expanding the established knowledge of neural information flow from low- to high-order brain areas, we revealed a novel brain mechanism based on simultaneous activity in different frequency bands within the same brain regions, highlighting its crucial role underlying complex cognitive functions.

neuroscience↗

bioRxiv2020

Rapid encoding of temporal sequences discovered in brain dynamics

Information encoding has received a wide neuroscientific attention, but the underlying rapid spatiotemporal brain dynamics remain largely unknown. Here, we investigated the rapid brain mechanisms for encoding and prediction of sounds forming a complex temporal sequence. Specifically, we used magnetoencephalography (MEG) to record the brain activity of 68 participants while they listened to a highly structured musical prelude. Advanced analysis of the phase synchronisation and graph theoretical measures showed the rapid transition of brain activity from primary auditory cortex to higher order association areas including insula and superior temporal pole within a whole-brain network, occurring during the first 220 ms of the encoding process. We discovered individual differences, revealing the rapid unfolding of brain network dynamics responsible for the processing of the current sounds and the prediction of the forthcoming events of the sequence. This provides a first glimpse of the general mechanisms underlying pattern encoding in the human brain.

neuroscience↗

bioRxiv2020

Targeting mitochondrial respiration and the BCL2 family in MYC-associated B-cell lymphoma

Multiple molecular features, such as activation of specific oncogenes (e. g. MYC, BCL2) or a variety of gene expression signatures, have been associated with disease course in diffuse large B-cell lymphoma (DLBCL). Understanding the relationships between these features and their possible exploitation toward disease classification and therapy remains a major priority in the field. Here, we report that MYC activity in DLBCL is closely correlated with - and most likely a driver of - gene signatures related to Oxidative Phosphorylation (OxPhos). On this basis, we hypothesized that enzymes involved in Oxidative Phosphorylation, and in particular electron-transport chain (ETC) complexes, might constitute tractable therapeutic targets in MYC-associated lymphoma. Indeed, our data show that MYC sensitizes B-cells to IACS-010759, a selective inhibitor of ETC complex I. Mechanistically, IACS-010759 activates an ATF4-driven Integrated Stress Response (ISR), engaging the intrinsic apoptosis pathway through the transcription factor CHOP. In line with these findings, IACS-010759 shows synergy with the BCL2 inhibitor venetoclax against double-hit lymphoma (DHL), a high-grade form of DLBCL with concurrent activation of MYC and BCL2. Similarly, in BCL2-negative lymphoma cell lines, inhibition of the BCL2-related protein Mcl-1 potentiates killing by IACS-010759. Altogether, ETC complex I inhibition engages the ISR to lower the apoptotic threshold in MYC-driven lymphomas and, in combination with select BCL2-family inhibitors, provides a novel therapeutic principle against this aggressive DLBCL subset. Statement of significanceThis work points to OxPhos as a key MYC-activated process and a tractable therapeutic target toward personalized treatment of high-grade DLBCL, providing strong context-dependent cooperation with BH3-mimetic compounds.

cancer biology↗

bioRxiv2020

Strong functional divergence of tropical reef fish assemblages along the global diversity gradient

Coral reefs are experiencing declines due to climate change and local human impacts. While at local scale biodiversity loss induces shifts in community structure, previous biogeographical analyses recorded consistent taxonomic structure of fish communities across global coral reefs. This suggest that regional communities represent a random subset of the global species and traits pool, whatever their species richness. Using distributional data on 3,586 fish species and latest advances in species distribution models we show that the global distribution of reef fishes is influenced by two major traits (body size and diet) and produces a strong divergence in the trait structure of assemblages across the biodiversity gradient. This divergence is best explained by the isolation of reefs during past unfavorable climatic conditions and highlights the risk of a global community re-organization if the ongoing climate-induced reef fragmentation is not halted.

ecology↗

bioRxiv2020

Subepidermal blisters are healed by wedge-shaped keratinocytes of hair follicle origin

Injury in adult tissue generally reactivates developmental programs to foster regeneration, but it is not known whether this paradigm applies to growing tissue. Here, by employing blisters, we show that epidermal wounds heal at the expense of skin development. The regenerated epidermis suppresses the expression of tissue morphogenesis genes accompanied by delayed hair follicle (HF) growth. Lineage tracing experiments, cell proliferation dynamics, and mathematical modeling reveal that the progeny of HF junctional zone stem cells, which undergo a morphological transformation, repair the blisters while not promoting HF development. In contrast, the contribution of interfollicular stem cell progeny to blister healing is small. These findings demonstrate that tissue development can be sacrificed for the sake of wound regeneration and suggest that tissue repair does not coincide with the reactivation of developmental programs in all regenerative contexts. Our study elucidates the key cellular mechanism of wound healing in skin blistering diseases.

cell biology↗

bioRxiv2017

Epidermal Wnt signalling regulates transcriptome heterogeneity and proliferative fate in neighbouring cells

Canonical Wnt/beta-catenin signalling regulates self-renewal and lineage selection within the mouse epidermis. Although the transcriptional response of keratinocytes that receive a Wnt signal is well characterised, little is known about the mechanism by which keratinocytes in proximity to the Wntreceiving cell are co-opted to undergo a change in cell fate. To address this, we performed single-cell mRNA-Seq on mouse keratinocytes co-cultured with and without the presence of beta-catenin activated neighbouring cells. We identified seven distinct cell states in cultures that had not been exposed to the beta-catenin stimulus and show that the stimulus redistributes wild type subpopulation proportions. Using temporal single-cell analysis we reconstruct the cell fate changes induced by neighbour Wnt activation. Gene expression heterogeneity was reduced in neighbouring cells and this effect was most dramatic for protein synthesis associated genes. The changes in gene expression were accompanied by a shift from a quiescent to a more proliferative stem cell state. By integrating imaging and reconstructed sequential gene expression changes during the state transition we identified transcription factors, including Smad4 and Bcl3, that were responsible for effecting the transition in a contact-dependent manner. Our data indicate that non cell-autonomous Wnt/beta-catenin signalling decreases transcriptional heterogeneity and further our understanding of how epidermal Wnt signalling orchestrates regeneration and self-renewal.

genomics↗

bioRxiv2017

A Balance Between Regulatory Constraints And Pathogen Pressure Shapes The Evolution Of Innate Immunity

As the first line of defence against pathogens, cells mount an innate immune response, which is highly variable from cell to cell. The response must be potent yet carefully controlled to avoid self-damage. How these constraints have shaped the evolution of innate immunity remains poorly understood. Here, we characterise this programmes transcriptional divergence between species and expression variability across cells. Using bulk and single-cell transcriptomics in primate and rodent fibroblasts challenged with an immune stimulus, we reveal a striking architecture of the innate immune response. Rapidly diverging genes, including cytokines and chemokines, also vary across cells and have distinct promoter structures. Conversely, genes involved in response regulation, such as transcription factors and kinases, are conserved between species and display low cell-to-cell variability. We suggest that this unique expression pattern, observed across species and conditions, has evolved as a mechanism for fine-tuned regulation, to achieve an effective but balanced response.

evolutionary biology↗