Biology subjects

Martin, S.

Publications and source records attributed to Martin, S..

At least 55 records · Page 3Linked to original sources

bioRxiv2024

Cross-species transmission of human hepatitis B virus to wild Neotropical primates

Hepatitis B virus (HBV) infects approximately one-third of the worlds human population and kills over one million people each year. HBV is also prevalent in Old World apes but not in New World primates. Human-to-primate transmission of HBV was suspected in zoo-captive monkeys and Mauritius macaques, but empirical data are scarce. Here, we collected blood and liver samples from 88 monkeys of 27 species in two areas of the Amazon, one with pristine forest and the other highly occupied and deforested by humans. A total of 17 (34.7%) out of 49 specimens from the human-occupied region tested positive for HBV. At this site, there was a positive relationship between human population density in the sampling location and the likelihood of primates being infected by HBV. Conversely, all 39 samples from the pristine forest tested negative for HBV. By sequencing a portion of the HBV S gene in five positive samples, each from a distinct primate genus, we found that four samples were closely related to the globally widespread human HBV-A strain, but not to the Americas-native HBV-F strain. The fifth sample aligned with the human HBV-D type, prevalent in the region where these samples were obtained. To our knowledge, this study represents the first reported cases of HBV in multiple wild New World primate species anywhere in the world. Our results suggest that primates were infected by strains brought into this part of Brazil by human immigrants, where HBV transmission may have been facilitated by the close contact between humans and monkeys due to high human occupation. This shows that the impact of human immigration, occupation and population growth in the Amazon extends beyond habitat loss; it also facilitates cross-species infections, potentially leading to the emergence of new, virulent viral strains that threaten both Amazonian biodiversity and human health. Author SummaryJean P. Boubli: Professor in Primate Ecology and Evolution, School of Science, Engineering and the Environment, University of Salford, Uk Hani R. El Bizri: PhD in Wildlife Conservation with a focus on Amazonian sustainable development. Analyst of Center for International Forestry Research (CIFOR), Bogor, Indonesia Luan F. Botelho-Souza: Masters and doctorate in experimental biology at the Universidade Federal de Rondonia/Brazil, Chrysoula Gubili: Researcher at the Fisheries Research Institute Nea Peramos, Kavala, Greece, specializing in population ecology and conservation genetics. Stephen J. Martin: Professor in Social Insects, School of Science, Engineering and the Environment, University of Salford, Uk Maisa da S. Araujo: Masters and doctorate in experimental biology, at the Universidade Federal de Rondonia/Brazil, Mariluce R. Messias: Professor of Zoology and curator of the Mammalogy Museum at the Universidade Federal de Rondonia/Brazil. Alcione de O. dos Santos: Masters and doctorate in experimental biology at the Universidade Federal de Rondonia/Brazil. Luiz S. Ozaki: Masters degree from Universidade de Brasilia and PhD in Physiological Sciences, Molecular Biology in Fukuoka, Kyushu, Japan. Andre V.C. Pereira: Bachelor in Biology the Universidade Federal de Rondonia/Brazil. Tony H. Katsuragawa: Masters and doctorate in experimental biology, at the Universidade Federal de Rondonia/Brazil. Ana Maisa Passos-Silva: Masters degree in experimental biology from the Federal University of Rondonia/Brazil. Luiz H. S. Gil: Masters in experimental biology, at the Universidade Federal de Rondonia/Brazil. Izeni P. Farias: Professor of Genetics, Universidade Federal do Amazonas, Brazil Juan M.V. Salcedo: Master in Tropical Medicine from the University of Brasilia/Brazil and PhD in Sciences from the University of Sao Paulo/Brazil. Tommy C. Burch: Masters in Biological Sciences and PhD candidate from the University of Salford, UK. Deusilene Vieira: Master in Tropical Medicine from the University of Brasilia/Brazil and PhD in Sciences from the University of Sao Paulo/Brazil.

pathology↗

bioRxiv2024

IRE1-dependent GOLIM4 expression controls protein secretion to modulate glioblastoma cell adhesion and migration

One of the main glioblastoma (GB) features is the diffuse migration of the tumor cells within the surrounding brain parenchyma, rendering almost impossible the complete tumor resection and irradiation, leading to inexorable lethal relapse of the disease. In the past years, we demonstrated that IRE1 (hereafter IRE1), one of the Endoplasmic Reticulum (ER) stress sensors, plays a key role in GB biology by impacting on immune infiltration, angiogenesis and tumor cell migration/invasion, all these features being linked to an alteration of protein secretion. In the present study, we investigated if and how IRE1 could regulate the functionality of the secretory machinery in GB cells and identified GOLIM4, a Golgi-associated molecule whose expression is regulated downstream of IRE1 through the transcription of XBP1s. Interestingly, GOLIM4 silencing led to decreased surface expression of multiple molecules including MHC class I molecules, growth factor receptors (PDGFRA and IL13RA2) and proteins involved in cell-cell adhesion (CD44, CD54, NCAM1), adhesion to matrix (ITGB1) or cell migration (CD90) without alteration of their encoding transcripts expression levels. Moreover, GOLIM4 silencing phenotypically affected GB cell-cell adhesion and cell migration in multiple models. Overall, we have described a novel IRE1/XBP1s/GOLIM4 operon that controls the secretion of specific proteins and impacts the tumor aggressiveness. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=100 SRC="FIGDIR/small/619629v1_ufig1.gif" ALT="Figure 1"> View larger version (24K): org.highwire.dtl.DTLVardef@511636org.highwire.dtl.DTLVardef@18971f6org.highwire.dtl.DTLVardef@98ff81org.highwire.dtl.DTLVardef@ae7189_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2024

A coregulatory influence map of glioblastoma heterogeneity and plasticity - From cells in vitro to tumors and back again

We present GBM-cRegMap, an online resource providing a comprehensive coregulatory influence network perspective on glioblastoma (GBM) heterogeneity and plasticity. Using representation learning algorithms, we derived two components of this resource: GBM-CoRegNet, a highly specific coregulatory network of tumor cells, and GBM-CoRegMap, a unified network influence map based on 1612 tumors from 16 studies. As a widely applicable closed-loop system connecting cellular models and tumors, GBM-cRegMap will provide the GBM research community with an easy-to-use web tool (https://gbm.cregmap.com) that maps any existing or newly generated transcriptomic "query" data to a reference coregulatory network and a large-scale manifold of disease heterogeneity. Using GBM-cRegMap, we demonstrated the synergy between the two components by refining the molecular classification of GBM, identifying potential key regulators, and aligning the transcriptional profiles of tumors and in vitro models. Through the amalgamation of a vast dataset, we validated the proneural (PN)-mesenchymal (MES) axis and identified three subclasses of classical (CL) tumors: astrocyte-like (CL-A), epithelial basal-like (CL-B), and cilium-rich (CL-C). We revealed the CL-C subclass, an intermediate state demonstrating the plasticity of GBM cells along the PN-MES axis under chemotherapy. We identified key regulators, such as PAX8, and NKX2.5, involved in TMZ resistance. Notably, NKX2.5, more expressed in higher-grade gliomas, negatively impacts patient survival and regulates genes involved in glucose metabolism. O_FIG O_LINKSMALLFIG WIDTH=152 HEIGHT=200 SRC="FIGDIR/small/609303v1_ufig1.gif" ALT="Figure 1"> View larger version (54K): org.highwire.dtl.DTLVardef@c7a401org.highwire.dtl.DTLVardef@e0681corg.highwire.dtl.DTLVardef@d31f55org.highwire.dtl.DTLVardef@ae350f_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2024

High-fat diet exacerbates the loss of bone fracture toughness in aging for C57BL/6JN mice

The elderly are at increased risk of bone fracture and more often consume poor quality diets, such as high-fat diet (HFD). We hypothesized that HFD would exacerbate the loss of bone fracture resistance in aging. To test this hypothesis, we fed 5-month and 22-month C57BL/6JN mice of both sexes HFD or low-fat diet for 8 weeks. Aging and HFD lowered bone fracture toughness, while only aging reduced bone strength. At the tissue-scale, aging increased bone mineral maturity, while HFD altered bone mineralization and maturity as well as collagen structure. Cortical bone metabolism was also dysregulated with aging and HFD. Dysregulated pathways in aging and HFD were related to cellular function and viability, or glucose regulation, respectively. Aging with HFD also impacted osteoclast and adipocyte abundance and osteocyte viability. Together, these data demonstrate that HFD may exacerbate the loss of bone matrix quality and fracture resistance in aging.

bioengineering↗

bioRxiv2024

Computational modeling of drug response identifies mutant-specific constraints for dosing panRAF and MEK inhibitors in melanoma

Purpose: This study explores the potential of preclinical in vitro cell line response data and computational modeling in identifying optimal dosage requirements of pan-RAF (Belvarafenib) and MEK (Cobimetinib) inhibitors in melanoma treatment. Our research is motivated by the critical role of drug combinations in enhancing anti-cancer responses and the need to close the knowledge gap around selecting effective dosing strategies to maximize their potential. Results: In a drug combination screen of 43 melanoma cell lines, we identified unique dosage landscapes of panRAF and MEK inhibitors for NRAS vs BRAF mutant melanomas. Both experienced benefits, but with a notably more synergistic and narrow dosage range for NRAS mutant melanoma. Computational modeling and molecular experiments attributed the difference to a mechanism of adaptive resistance by negative feedback. We validated in vivo translatability of in vitro dose-response maps by accurately predicting tumor growth in xenografts. Then, we analyzed pharmacokinetic and tumor growth data from Phase 1 clinical trials of Belvarafenib with Cobimetinib to show that the synergy requirement imposes stricter precision dose constraints in NRAS mutant melanoma patients. Conclusion: Leveraging pre-clinical data and computational modeling, our approach proposes dosage strategies that can optimize synergy in drug combinations, while also bringing forth the real-world challenges of staying within a precise dose range. Simple SummaryCombining drugs is crucial for enhancing anti-cancer responses. However, the potential of pre-clinical data in identifying suitable combinations and dosage is often underutilized. In this study, we leverage preclinical in vitro cell line drug response data and computational modeling of signal transduction and of pharmacokinetics to elucidate distinct dose requirements for the combination of pan-RAF and MEK inhibitors in melanoma. Our findings reveal a more synergistic, but narrower dosing landscape in NRAS vs BRAF mutant melanoma, which we linked to a mechanism of adaptive resistance through negative feedback. Further, our analysis suggests the importance of drug dosing strategies to optimize synergy based on mutational context, yet highlights the real-world challenges of maintaining a narrow dose range. This approach establishes a framework for translational investigation of drug responses in the refinement of combination therapy, balancing the potential for synergy and practical feasibility in cancer treatment planning.

systems biology↗

bioRxiv2024

Limited cognitive resources reduce the language predictability benefit across the adult lifespan

Humans routinely anticipate upcoming language, but whether such predictions come at a cognitive cost remains debated. In this study, we demonstrate the resource-dependent nature of predictive mechanisms in language comprehension across the lifespan: Experimentally limiting executive resources through a concurrent task reduces the effect of language predictability on reading time. Participants (N=175, replication N=96) read short articles presented word-by-word while completing a secondary font colour n-back task, thus varying cognitive demand. Language predictability was indexed by word surprisal as derived from a pre-trained large language model (GPT-2). Across two independent samples, our findings reveal that language predictions are not cost-free: They draw on executive control resources, and this dependency becomes more pronounced with age (18-85 years). These results help resolve the debate over cognitive demands in language comprehension and highlight prediction as a dynamic, resource-dependent process across the lifespan.

neuroscience↗

bioRxiv2024

Clearing method adapted to FFPE tissues for 3D imaging of nerve fibers, B cells, and tertiary lymphoid structures

Biomedical samples are commonly used for histological examination after their inclusion in paraffin (Formalin-Fixed Paraffin-Embedded (FFPE) biopsy). However, they provide minimal information about the interactions between different tissue components such as blood vessels, nerves, and cellular aggregates. Here we present a modified iDISCO tissue-clearing method that we term miDISCO+. miDISCO+ can be used for analyzing FFPE samples, requires the use of only one single FFPE sample, and allows to acquire a detailed image of the 3D tissue/organ architecture and the relevant cellular interactions. The addition of an antigen retrieval step to the protocol enables to use antibodies whose binding is sensitive to formalin fixation due to antigen masking. This method enabled us to detect CD20+ B cell follicles and to show that they are in close contact with TH+ sympathetic nerve fibers in FFPE biopsies of human palatine tonsils, and to detect CD20+ B cells in lung tumors and observe their 3D organization within tertiary lymphoid structures. Thus, miDISCO+ could be a potent tool for clinicians to refine diagnosis and to select optimal personalized treatment by giving an integrated 3D view of the tissue structures and cellular interactions in single FFPE samples.

immunology↗

bioRxiv2024

Contrasting dynamics of two incursions of low pathogenicity avian influenza virus into Australia

The current panzootic of high pathogenicity avian influenza virus H5N1 demonstrates how viral incursions can have major ramifications for wildlife and domestic animals. Herein, we describe the recent incursion into Australia of two low pathogenicity avian influenza virus subtypes, H4 and H10, that exhibited contrasting evolutionary dynamics. Viruses detected from national surveillance and disease investigations between 2020-2022 revealed 27 genomes, 24 of which have at least one segment more closely related to Eurasian or North American avian influenza lineages than those already circulating in Australia. Phylogenetic analysis revealed that H4 viruses circulating in shorebirds represent a recent incursion from Asia that is distinct from those circulating concurrently in Australian waterfowl. Analysis of the internal segments further demonstrates exclusive, persistent circulation in shorebirds. This contrasts with H10, where a novel lineage has emerged in wild waterfowl, poultry and captive birds across Australia, and has likely replaced previously circulating H10 lineages through competitive exclusion. Elucidating different dynamics for avian influenza incursions supports effective disease risk identification and communication that better informs disease preparedness and response.

microbiology↗

bioRxiv2024

Systematic perturbation screens decode regulators of inflammatory macrophage states and identify a role for TNF mRNA m6A modification

Macrophages adopt dynamic cell states with distinct effector functions to maintain tissue homeostasis and respond to environmental challenges. During chronic inflammation, macrophage polarization is subverted towards sustained inflammatory states which contribute to disease, but there is limited understanding of the regulatory mechanisms underlying these disease-associated states. Here, we describe a systematic functional genomics approach that combines genome-wide phenotypic screening in primary murine macrophages with transcriptional and cytokine profiling of genetic perturbations in primary human monocyte-derived macrophages (hMDMs) to uncover regulatory circuits of inflammatory macrophage states. This process identifies regulators of five distinct inflammatory states associated with key features of macrophage function. Among these, the mRNA m6A writer components emerge as novel inhibitors of a TNF-driven cell state associated with multiple inflammatory pathologies. Loss of m6A writer components in hMDMs enhances TNF transcript stability, thereby elevating macrophage TNF production. A PheWAS on SNPs predicted to impact m6A installation on TNF revealed an association with cystic kidney disease, implicating an m6A-mediated regulatory mechanism in human disease. Thus, systematic phenotypic characterization of primary human macrophages describes the regulatory circuits underlying distinct inflammatory states, revealing post-transcriptional control of TNF mRNA stability as an immunosuppressive mechanism in innate immunity.

immunology↗

bioRxiv2024

Vascular dysfunction is at the onset of oxaliplatin-induced peripheral neuropathy symptoms in mice

Oxaliplatin-induced peripheral neuropathy (OIPN) is an adverse side effect of this chemotherapy used for gastrointestinal cancers. The continuous pain experienced by OIPN patients often result in the reduction or discontinuation of chemotherapy, thereby affecting patient survival. Several pathogenic mechanisms involving sensory neurons were shown to participate in the occurrence of OIPN symptoms. However, the dysfunction of the blood-nerve barrier as a source of nerve alteration had not been thoroughly explored. To characterise the vascular contribution to OIPN symptoms, we undertook two comparative transcriptomic analyses from mouse purified brain and sciatic nerve blood vessels (BVs), and nerve BVs after oxaliplatin or control administration. These analyses reveal distinct molecular landscapes between brain and nerve BVs and the upregulation of transcripts involved in vascular contraction after oxaliplatin treatment. Anatomical examination of the nerve yet shows the preservation of BV architecture in acute OIPN mouse model, although treated mice exhibit both neuropathic symptoms and enhanced vasoconstriction reflected by hypoxia. Moreover, vasodilators significantly reduce oxaliplatin-induced neuropathic symptoms and endoneurial hypoxia, establishing the key involvement of nerve blood flow in OIPN.

neuroscience↗

bioRxiv2024

Experimental evolution of an RNA virus in Caenorhabditis elegans

The discovery of Orsay virus (OrV), the first virus infecting wild populations of Caenorhabditis elegans, has boosted studies of viral immunity pathways in this nematode. Considering the many advantages that C. elegans offers for fundamental research in host-pathogen interactions, this pathosystem has high potential to become a model system for experimental virus evolution studies. However, the evolutionary constraints operating in this pathosystem have barely been explored. Here we describe for the first time an evolution experiment of two different OrV strains in C. elegans. After 10 serial passages of evolution, we report slight changes in infectivity and non-synonymous mutations fixed in the evolved viral populations. In addition, we observed numerous minor variants emerging in the viral population. These minor variants were not randomly distributed along the genome but concentrated in polymorphic genomic regions. Overall, our work established the grounds for future experimental virus evolution studies using Caenorhabditis nematodes. HIGHLIGHTSO_LICaenorhabditis elegans-Orsay virus is a convenient pathosystem to study virus evolution. C_LIO_LIThe approach used to test the viral strains may interfere with the infection phenotypes observed. C_LIO_LIThere may be specific genomic hotspots regions of nucleotide diversity important for the evolution of Orsay virus. C_LIO_LIThe substitution rate observed for Orsay virus was low, suggesting that the two strains studied might be already well adapted to laboratory conditions. C_LI

evolutionary biology↗

bioRxiv2024

Striatal Dopamine Modulates Temporal Surprise P3a

Dopamine is vital in forming mental models of "what" and "when" sensory events occur that essentially guide goal-directed behaviour. However, it remains largely unknown how variations in temporal predictability are incorporated into such mental models. A better understanding of the underlying mechanisms is important, considering dopaminergic depletion in diseases such as Parkinsons Disease and Schizophrenia, where abnormal temporal processing is observed. Some electroencephalographic (EEG) studies indicate that noradrenergic mechanisms, as reflected in the P3b event-related potential, are modulated by temporal predictability, whereas others indicate that dopaminergic mechanisms as reflected in the P3a, underlie surprise. In this study, resting-state and task-dependent EEG was recorded from 24 healthy participants who were administered a selective D2 agonist or antagonist before they performed a pure tone auditory "oddball" task. Two oddball sequences included either partially predictable, with inter-stimulus intervals (ISIs) of 400ms/1200ms; or fully temporally predictable tones, with a consistent ISI of 600ms. Tones following 400ms ISIs were perceived as surprising, or "early", as shown in an enhanced P3a response; tones following a 1200ms ISIs showed a much reduced P3a response ("late"). The agonist accentuated the "late" effect, demonstrating that drugs targeting D2 receptors modulate temporal prediction. These findings differentiate the role of the dopaminergic system in temporal processing and model-based auditory predictions.

neuroscience↗

bioRxiv2023

A chromosome-level, haplotype-resolved genome assembly and annotation for the Eurasian minnow (Leuciscidae - Phoxinus phoxinus) provide evidence of haplotype diversity

AO_SCPLOWBSTRACTC_SCPLOWIn this study we present an in-depth analysis of the Eurasian minnow (Phoxinus phoxinus) genome, highlighting its genetic diversity, structural variations, and evolutionary adaptations. We generated an annotated haplotype-phased, chromosome-level genome assembly (2n = 50) by integrating high-fidelity (HiFi) long reads and chromosome conformation capture data (Hi-C). We achieved a haploid size of 940 Megabase pairs (Mbp) for haplome one and 929 Mbp for haplome two with high scaffold N50 values of 36.4 Mb and 36.6 Mb and BUSCO scores of 96.9% and 97.2%, respectively, indicating a highly complete genome assembly. We detected notable heterozygosity (1.43%) and a high repeat content (approximately 54%), primarily consisting of DNA transposons, which contribute to genome rearrangements and variations. We found substantial structural variations within the genome, including insertions, deletions, inversions, and translocations. These variations affect genes enriched in functions such as dephosphorylation, developmental pigmentation, phagocytosis, immunity, and stress response. In the annotation of protein-coding genes, 30,980 mRNAs and 23,497 protein-coding genes were identified with a high completeness score, which further underpins the high contiguity of our genome assemblies. We performed a gene family evolution analysis by comparing our proteome to ten other teleost species, which identified immune system gene families that prioritise histone-based disease prevention over NLR-based immune responses. Additionally, demographic analysis indicates historical fluctuations in the effective population size of P. phoxinus, likely correlating with past climatic changes. This annotated, phased reference genome provides a crucial resource for resolving the taxonomic complexity within the genus Phoxinus and highlights the importance of haplotype-phased assemblies in understanding haplotype diversity in species characterised by high heterozygosity.

genomics↗

bioRxiv2023

DNA methylation by CcrM contributes to genome maintenance in the Agrobacterium tumefaciens plant pathogen

The cell cycle-regulated DNA methyltransferase CcrM is conserved in most Alphaproteobacteria, but its role in bacteria with complex or multicentric genomes remains unexplored. Here, we compare the methylome, the transcriptome and the phenotypes of wild-type and CcrM-depleted Agrobacterium tumefaciens cells with a dicentric genome with two essential replication origins. We find that DNA methylation has a pleiotropic impact on motility, biofilm formation and viability. Remarkably, CcrM promotes the expression of the repABCCh2 operon, encoding proteins required for replication initiation/partitioning at ori2, and inhibits gcrA, encoding a conserved global cell cycle regulator. Imaging ori1 and ori2 in live cells, we show that replication from ori2 is often delayed in cells with a hypo-methylated genome, while ori2 over-initiates in cells with a hyper-methylated genome. We thus propose that methylation by CcrM stimulates RepABC-dependent chromosomal origins, uncovering a novel and original connection between CcrM-dependent DNA methylation and genome maintenance in an Alphaproteobacterial pathogen.

microbiology↗

bioRxiv2023

Hypothesis Generation For Rare and Undiagnosed Diseases Through Clustering and Classifying Time-Versioned Biological Ontologies

Rare diseases affect 1-in-10 people in the United States and despite increased genetic testing, up to half never receive a diagnosis. Even when using advanced genome sequencing platforms to discover variants, if there is no connection between the variants found in the patients genome and their phe-notypes in the literature, then the patient will remain undiagnosed. When a direct variant-phenotype connection is not known, putting a patients information in the larger context of phenotype relation-ships and protein-protein-interactions may provide an opportunity to find an indirect explanation. Databases such as STRING contain millions of protein-protein-interactions and HPO contains the relations of thousands of phenotypes. By integrating these networks and clustering the entities within we can potentially discover latent gene-to-phenotype connections. The historical records for STRING and HPO provide a unique opportunity to create a network time series for evaluating the cluster sig-nificance. Most excitingly, working with Childrens Hospital Colorado we provide promising hy-potheses about latent gene-to-phenotype connections for 38 patients with undiagnosed diseases. We also provide potential answers for 14 patients listed on MyGene2. Clusters our tool finds significant harbor 2.35 to 8.72 times as many gene-to-phenotypes edges inferred from known drug interactions than clusters find to be insignificant. Our tool, BOCC, is available as a web app and command line tool.

bioinformatics↗

bioRxiv2023

Algebraic invariants for inferring 4-leaf semi-directed phylogenetic networks

A core goal of phylogenomics is to determine the evolutionary history of a set of species from biological sequence data. Phylogenetic networks are able to describe more complex evolutionary phenomena than phylogenetic trees but are more difficult to accurately reconstruct. Recently, there has been growing interest in developing methods to infer semi-directed phylogenetic networks. As computing such networks can be computationally intensive, one approach to building such networks is to puzzle together smaller networks. Thus, it is essential to have robust methods for inferring semi-directed phylogenetic networks on small numbers of taxa. In this paper, we investigate an algebraic method for performing phylogenetic network inference from nucleotide sequence data on 4-leaf semi-directed phylogenetic networks by analysing the distribution of leaf-pattern probabilities. On simulated data, we found that we can correctly identify with high accuracy the undirected phylogenetic network for sequences of length at least 10kbp. We found that identifying the semi-directed network is more challenging and requires sequences of length approaching 10Mbp. We are also able to use our approach to identify tree-like evolution and determine the underlying tree. Finally, we employ our method on a real dataset from Xiphophorus species and use the results to build a phylogenetic network.

evolutionary biology↗

bioRxiv2023

ATP13A3 Variants Promote Pulmonary Arterial Hypertension by Disrupting Polyamine Transport

AimsPotential loss-of-function variants of ATP13A3, the gene encoding a P5B-type transport ATPase of undefined function, were recently identified in pulmonary arterial hypertension (PAH) patients. ATP13A3 is implicated in polyamine transport but its function has not been fully elucidated. Here, we sought to determine the biological function of ATP13A3 in vascular endothelial cells and how PAH-associated mutations may contribute to disease pathogenesis. We also generated mice harbouring an Atp13a3 variant analogous to a human disease-associated variant to establish whether these mice develop PAH. Methods and ResultsWe studied the impact of ATP13A3 deficiency and overexpression in endothelial cell (EC) models (human pulmonary ECs, blood outgrowth ECs (BOECs) and HMEC-1 cells), including a PAH patient-derived BOEC line harbouring an ATP13A3 variant (LK726X). ATP13A3 localised to the recycling endosomes of human ECs. Knockdown of ATP13A3 in ECs generally reduced the basal polyamine content, consistently reduced putrescine uptake, and altered the expression of enzymes involved in polyamine metabolism. Conversely, overexpression of wild-type ATP13A3 increased polyamine uptake, with an overall preference of putrescine > spermidine > spermine. Functionally, loss of ATP13A3 was associated with reduced EC proliferation, increased apoptosis in serum starvation and increased monolayer permeability to thrombin. Assessment of five PAH-associated missense ATP13A3 variants (L675V, M850I, V855M, R858H, L956P) confirmed loss-of-function phenotypes represented by impaired polyamine transport and dysregulated EC function. Furthermore, mice carrying a heterozygous germ-line Atp13a3 frameshift variant representing a human mutation spontaneously developed a PAH phenotype, with increased pulmonary pressures, right ventricular remodelling and muscularisation of pulmonary vessels. ConclusionWe identify ATP13A3 as a polyamine transporter, deficiency of which leads to EC dysfunction and predisposes to PAH. This suggests a need for targeted therapies to alleviate the imbalances in polyamine homeostasis and EC dysfunction in PAH. Translational perspectiveRare missense ATP13A3 disease-associated variants have been identified in patients with pulmonary arterial hypertension (PAH), though their pathogenicity has not been confirmed as the function of ATP13A3 is not known. We have identified ATP13A3 as a polyamine transporter, showing that ATP13A3 deficiency impaired polyamine homeostasis and uptake, and drove endothelial dysfunction. Conversely, overexpression increased polyamine uptake and rescued the proapoptotic phenotype of cells harbouring a disease-associate variant. Mice heterozygous for a disease-associated Atp13a3 mutation spontaneously develop PAH. These findings support the rationale for exploring dysregulated polyamine homeostasis in PAH and suggest a potential for therapeutic targeting of polyamine pathways in PAH.

cell biology↗

bioRxiv2023

The lipid flippase ATP10B enables cellular lipid uptake under stress conditions

Pathogenic ATP10B variants have been described in patients with Parkinsons disease and dementia with Lewy body disease, and we previously established ATP10B as a late endo-/lysosomal lipid flippase transporting both phosphatidylcholine (PC) and glucosylceramide (GluCer) from the lysosomal exoplasmic to cytoplasmic membrane leaflet. Since several other lipid flippases regulate cellular lipid uptake, we here examined whether also ATP10B impacts cellular lipid uptake. Transient co-expression of ATP10B with its obligatory subunit CDC50A stimulated the uptake of fluorescently (NBD-) labeled PC in HeLa cells. This uptake is dependent on the transport function of ATP10B, is impaired by disease-associated variants and appears specific for NBD-PC. Uptake of non-ATP10B substrates, such as NBD-sphingomyelin or NBD-phosphatidylethanolamine is not increased. Remarkably, in stable cell lines co-expressing ATP10B/CDC50A we only observed increased NBD-PC uptake following treatment with rotenone, a mitochondrial complex I inhibitor that induces transport-dependent ATP10B phenotypes. Conversely, Im95m and WM-115 cells with endogenous ATP10B expression, present a decreased NBD-PC uptake following ATP10B knockdown, an effect that is exacerbated under rotenone stress. Our data show that the endo-/lysosomal lipid flippase ATP10B contributes to cellular PC uptake under specific cell stress conditions.

cell biology↗