SEARCH · bioRxiv Science

Results for “Molecular Biology”

Search indexed bioRxiv preprints in genomics, neuroscience, cell biology and bioinformatics. Read source abstracts and check manuscript versions; preprints are not peer reviewed.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 1,081 records · Page 60Linked to original sources

bioRxiv2022

Dietary fibre controls blood pressure and cardiovascular risk by lowering large intestinal pH and activating the proton-sensing receptor GPR65

High blood pressure (BP) is the most common cause of death globally, due to increasing the risk of cardiovascular diseases. Dietary fiber regulates BP through gut microbial production of acidic metabolites known as short-chain fatty acids (SCFAs). The specific mechanisms of how SCFAs regulate BP are still emerging. In a phenome-wide association study, we identified that the proton-sensing G-protein-coupled receptor GPR65 gene is associated with hypertension and its associated end-organ damage phenotypes. We hypothesized that acidic metabolites produced from the gut microbiota may activate GPR65, thus conferring BP regulating effects. We found that dietary fiber levels determined the luminal and interstitial tissue pH in the large intestine through production of SCFAs by the gut microbiota. We identified that low pH produced by high fiber intake, acting via GPR65 signaling, increased cAMP production and phosphorylation of CREB, and restricted the production of hypertension-promoting inflammatory cytokines by CD8+ T cells. Gpr65-/- mice spontaneously developed higher BP, cardiac and renal hypertrophy and fibrosis. We showed that the benefits of a diet high in fiber, which prevented hypertension and associated end-organ damage, were decreased in Gpr65-/- mice. Finally, adoptive transfers revealed that GPR65 deficiency in CD8+ T cells causally explained this phenotype. In conclusion, we showed that pH sensing by GPR65 in CD8+ T cells mediates much of the cardiovascular benefits of dietary fiber. pH sensing represents a novel gene-by-environment interaction of gut microbiota-to-host biological effects and may form the basis for new therapeutic strategies for hypertension.

molecular biology↗

bioRxiv2023

Comparative Genomics and Directed Evolution Reveal Genetic Determinants of Extreme UVC Radiation Tolerance in Bacteria Recovered from the Stratosphere

Aerosolized microbes surviving transport to and in the stratosphere endure extremes of low temperature, atmospheric pressure, and relative humidity, and high shortwave ultraviolet radiation flux. However, the genetic determinants for traits enabling resistance to the combination of stresses experienced by microbes in the high atmosphere have not been systematically investigated. In this study, we examined Proteobacteria and Actinobacteria isolated from the stratosphere (18 to 29 km ASL) and that demonstrated high tolerance to desiccation (15-25% RH) and UVC radiation (UVCR; {lambda}= 254 nm). Closely related reference strains were more sensitive to UVCR than the stratospheric isolates, indicating that extreme resistance is not universally distributed in these phylogenetically related bacteria. Comparative genomic analyses revealed DNA repair and antioxidant defense genes in the isolates that are not possessed by the related reference strains, including genes encoding photolyase, DNA nucleases and helicases, and catalases. Directed evolution by repeated exposure to increasing doses of UVCR improved the LD90 in a sensitive reference strain by [~]3.5-fold. The mutations acquired in Curtobacterium flaccumfaciens pv. flaccumfaciens strain DSM 20129 incrementally increased its UVCR resistance, with the accumulation of 20 point mutations in protein coding genes increasing tolerance to a level approaching that of stratospheric isolate Curtobacterium sp. L6-1. The genetic basis for the increased UVCR tolerance phenotypes observed is discussed, with a specific emphasis on the role of genes involved in DNA repair and detoxification of reactive oxygen species. ImportanceUltraviolet radiation is omnipresent in sunlight and has important biological effects on organisms. The stratosphere is the only location on Earth where microbes receive natural exposure to highly mutagenic wavelengths (<280 nm) of ultraviolet radiation. Genetic studies of bacteria from an environment that selects for extreme ultraviolet radiation resistant phenotypes has expanded what is known from studies of model species (e.g., E. coli) and identified potentially novel protection and repair strategies. In addition to deepening understanding of ultraviolet radiation photobiology in atmospheric microbes and bacteria in general, these advancements are also highly relevant to astrobiology and space biology. The cold, dry, hypobaric, and high radiation environment of the stratosphere provides an earthly analog for thin extraterrestrial atmospheres (e.g., Mars) and is ideal for bioprospecting extremophile phenotypes that enable engineering of genetic stability and functionality in bio-based space life-support systems or any application where long-term persistence is desirable (e.g., biocontrol).

molecular biology↗

bioRxiv2024

Cell 3D Positioning by Optical encoding (C3PO) and its application to spatial transcriptomics

Current state-of-the-art spatial omics approaches suffer from the drawback that they are tissue section-based and thus inherently 2-dimensional. A full understanding of biological processes will only be possible when such data is available in 3-dimensions (3D). Here, we introduce Cell 3D Positioning by Optical encoding (C3PO) - the first technique capable of reconstructing the 3D positions of cells in a tissue, after they have been fully dissociated for single-cell omics analysis. It imposes a Cartesian coordinate system of positions on the tissue and cells of interest, before dissociation. This is created by multiple orthogonal spatial gradients of active fluorophores, carefully shaped by a 3D bleaching method, such that each position in the tissue is encoded by a unique fluorescent address. Upon dissociation of the tissue the fluorescent addresses of the cells can be read via an appropriate device (such as a FACS machine) to computationally reconstruct the tissue in 3D, before omics are performed downstream. Here, we show two proof-of-principle results for C3PO. First, pure C3PO without omics, to reconstruct the 3D geometry of a developing mouse limb bud. Second, an application of C3PO to spatial transcriptomics, revealing the expression patterns of 73 genes with interesting gene expression patterns in the developing limb.. C3PO is a genuinely novel approach to reconstruct the original 3D positions of cells in a tissue after dissociation. Combined with transcriptomics, it can play a significant role in the study of any tissue or organ in which 3D structure and geometry is important, such as developmental biology, cancer biology and neuroscience. It is not an omics technique per se, and in the future could be combined with the growing family of other omics technologies. One sentence summaryC3PO is a novel optical technique that can preserve the 3D positional coordinates of cells after tissue dissociation, enabling a radically new approach to spatial transcriptomics.

molecular biology↗

bioRxiv2023

Increased hippocampal epigenetic age in the Ts65Dn Mouse Model of Down Syndrome

Down syndrome (DS) is a segmental progeroid genetic disorder associated to multi-systemic precocious ageing phenotypes, which are particularly evident at the immune and nervous systems. Accordingly, people with DS show an increased biological age as measured by epigenetic clocks. Ts65Dn trisomic mouse, which harbors extra-numerary copies of Hsa21-syntenic regions, was shown to recapitulate several progeroid features of DS, but no biomarkers of age have been applied to it so far. Here we used a mouse specific epigenetic clock to measure epigenetic age of hippocampi from Ts65Dn and euploid mice at 20 weeks. Ts65Dn mice showed an increased hippocampal epigenetic age respect to controls, and the observed changes in DNA methylation partially recapitulated those observed in hippocampi from people with DS. Collectively, our results support the use of the Ts65Dn model to decipher the molecular mechanisms underlying the progeroid DS phenotypes.

molecular biology↗

bioRxiv2022

Developing a temperature-inducible transcriptional rheostat in Neurospora crassa

Heat shock protein (hsp) encoding genes, part of the highly conserved Heat Shock Response (HSR), are known to be induced by thermal stress in several organisms. In Neurospora crassa, three hsp genes, hsp30, hsp70, and hsp80, have been characterized; however, the role of defined cis-elements in their response to discrete changes in temperature remains largely unexplored. To fill this gap, while also aiming to obtain a reliable fungal heat-shock inducible system, we analyzed different sections of each hsp promoter, by assessing the expression of real-time transcriptional reporters. Whereas all three promoters, and their resected versions, were acutely induced by high temperatures, only hsp30 displayed a broad range of expression and high tunability amply exciding other inducible promoter systems existing in Neurospora, such as Quinic acid- or light-inducible ones. As proof of concept, we employed one of these promoters to control the expression of clr-2, which encodes for the master regulator of Neurospora cellulolytic capabilities. The resulting strain fails to grow on cellulose at 25{degrees}C, whereas it robustly grows if heat shock pulses are delivered daily. Additionally, we designed two hsp30 synthetic promoters and characterized these, as well as the native promoters, to a gradient of high temperatures, yielding a wide range of responses to thermal stimuli. Thus, Neurospora hsp30-based promoters represent a new set of modular elements that can be used as a transcriptional rheostat to adjust the expression of a gene of interest or for the implementation of regulated circuitries for synthetic biology and biotechnological strategies. ImportanceTimely and dynamic response to strong temperature rises is paramount for organismal biology. At the same time, inducible promoters are a powerful tool for fungal biotechnological and synthetic biology endeavors. In this work, we analyzed the activity of several N. crassa heat shock protein (hsp) promoters upon a wide range of temperatures, observing that hsp30 exhibits remarkable sensitivity and dynamic range of expression as we chartered the response of this promoter to subtle increases in temperature, while also building synthetic promoters based on hsp30 cis-elements. As proof of concept, we analyzed the ability of hsp30 to provide tight control of a central process such as cellulose degradation. While this study provides an unprecedented description of the regulation of the N. crassa hsp genes it also contributes with a noteworthy addition to the molecular toolset of transcriptional controllers in filamentous fungi.

molecular biology↗

bioRxiv2021

Early life stress affects the miRNA cargo in epididymal extracellular vesicles in mouse

Sperm RNA can be modified by environmental factors and has been implicated in communicating signals about changes in a fathers environment to the offspring. The RNA composition of sperm is influenced during its final stage of maturation in the epididymis by extracellular vesicles released by epididymal cells. We studied the effect of exposure to stress in postnatal life on the transcriptome of epididymal extracellular vesicles using a mouse model of transgenerational transmission. We found that the small RNA signature of epididymal extracellular vesicles, particularly miRNAs, is altered in adult males exposed to postnatal stress. miRNAs changes correlate with differences in the expression of their target genes in sperm and zygotes generated from that sperm. These results suggest that stressful experiences in early life can have persistent biological effects on the male reproductive tract that may in part be responsible for the transmission of the effects of exposure to the offspring. Summary SentencemiRNA cargo of extracellular vesicles in cauda epididymis is changed by paternal exposure to early life stress, which correlates with differences in the expression of their target genes in sperm and zygotes generated from that sperm

molecular biology↗

bioRxiv2024

Protein complex heterogeneity and topology revealed by electron capture charge reduction and surface induced dissociation

We illustrate the utility of native mass spectrometry (nMS) combined with a fast, tunable gas-phase charge reduction, electron capture charge reduction (ECCR), for the characterization of protein complex topology and glycoprotein heterogeneity. ECCR efficiently reduces the charge states of tetradecameric GroEL, illustrating Orbitrap m/z measurements to greater than 100,000 m/z. For pentameric C-reactive protein and tetradecameric GroEL, our novel device combining ECCR with surface induced dissociation (SID) reduces the charge states and yields more topologically informative fragmentation. This is the first demonstration that ECCR yields more native-like SID fragmentation. ECCR also significantly improved mass and glycan heterogeneity measurements of heavily glycosylated SARS-CoV-2 spike protein trimer and thyroglobulin dimer. Protein glycosylation is important for structural and functional properties and plays essential roles in many biological processes. The immense heterogeneity in glycosylation sites and glycan structure poses significant analytical challenges that hinder a mechanistic understanding of the biological role of glycosylation. Without ECCR, average mass determination of glycoprotein complexes is available only through charge detection mass spectrometry or mass photometry. With narrow m/z selection windows followed by ECCR, multiple glycoform m/z values are apparent, providing quick global glycoform profiling and providing a future path for glycan localization on individual intact glycoforms.

molecular biology↗

bioRxiv2024

Dynamics of MicroRNA Secreted via Extracellular Vesicles During the Maturation of Embryonic Stem Cell-Derived Retinal Pigment Epithelium

Retinal pigment epithelium (RPE) cells are exclusive to the retina, critically multifunctional in maintaining the visual functions and health of photoreceptors and the retina. Despite their vital functions throughout lifetime, RPE cells lack regenerative capacity, rendering them vulnerable and central to degenerative retinal diseases. With advancements in stem cell technology enabling the differentiation of functional cells from pluripotent stem cells and leveraging the robust autocrine and paracrine functions of RPE cells, extracellular vesicles (EVs) secreted by RPE cells hold significant therapeutic potential in supplementing RPE cell activity. While previous research has primarily focused on the trophic factors secreted by RPE cells, there is a lack of studies investigating miRNA, which serves as a master regulator of gene expression. Profiling and defining the functional role of miRNA contained within RPE-secreted EVs is critical as it constitutes a necessary step in identifying the optimal phenotype of the EV secreting cell and understanding biological cargo of EVs to develop EV-based therapeutics. In this study, we present a comprehensive profile of miRNA in small extracellular vesicles (sEV) secreted during RPE maturation following differentiation from human embryonic stem cells (hESCs). This exploration is essential for ongoing efforts to develop and optimize EV-based intraocular therapeutics utilizing RPE-secreted EVs, which may significantly impact the function of dysfunctional RPE cells.

molecular biology↗

bioRxiv2024

Epigenetic age prediction drifts resulting from next-generation methylation arrays

BackgroundEpigenetic clocks based on DNA methylation data are routinely used to obtain surrogate measures of biological age and estimate epigenetic age acceleration rates. These tools are mathematical models that rely on the methylation state of specific sets of CpG islands quantified using microarrays. The set of CpG islands probed in the microarrays differed between the models. Thus, as new methylation microarrays are developed and older models are discontinued, existing epigenetic clocks might become obsolete. Here, we explored the effects of the changes introduced in the new DNA methylation array from Illumina (EPICv2) on existing epigenetic clocks. MethodsWe compiled a whole-blood DNA methylation dataset of 10835 samples to test the performance of four epigenetic clocks on the probe set of the EPICv2 array. We then used the same data to train a new epigenetic age prediction model compatible across the 450k, EPICv1 and EPICv2 microarrays. We compiled a validation dataset of 2095 samples to compare our model with a state-of-the-art epigenetic clock. Using two datasets with repeated samples from the same subjects, we computed an estimate of the contribution of technical noise and intra-subject variation to the variation of epigenetic age predictions from each of the models tested. We used a dataset of cancer survivors who had undergone different types of therapy, a dataset of breast cancer patients and controls, and a dataset from an exercise-based interventional study to test the ability of our model to detect alterations in epigenetic age acceleration. ResultsWe found that the results of the four epigenetic clocks tested are significantly distorted by the absence of specific probes in the EPICv2 microarray, causing an average difference of up to 25 years. We developed an epigenetic age prediction model compatible with the 450k, EPICv1 and EPICv2 microarrays. Our model produced highly accurate chronological age predictions that were comparable to those of a state-of-the-art epiclock. We obtained estimates for the variation of epigenetic age acceleration on normal, non-pathological populations associated with each of the models tested. These parameters provide thresholds to evaluate the relevance of epigenetic age alterations. In all cases, the estimated technical noise and intra-subject variability were smaller than the population-based epigenetic age prediction variability. Finally, we used our new models to reproduce previous results showing increased epigenetic age acceleration in cancer patients and in survivors who had been treated with radiation therapy, as well as a lack of changes as a result of exercise-based interventions. ConclusionOur work demonstrated that existing epigenetic clocks need to be updated to be applicable to data generated with the new EPICv2 microarray, which has phased out the 450k and EPICv1 models. To overcome this technical hurdle, we developed a new model that translates the capabilities of state-of-the-art epigenetic clocks to the new EPICv2 platform and is cross-compatible with the 450k and EPICv1 microarrays. Our characterization of the variation of epigenetic age predictions provides useful metrics to contextualize the biological relevance of epigenetic age alterations. The analysis of data from subjects influenced by radiation, cancer and exercise-based interventions shows that despite being good predictors of chronological age, neither a pathological state like breast cancer, a hazardous environmental factor (radiation) or exercise (a beneficial intervention) caused significant changes in the values of the "epigenetic age" determined by these first-generation models.

molecular biology↗

bioRxiv2023

Cyanobacterial α-carboxysome carbonic anhydrase is allosterically regulated by the Rubisco substrate RuBP

Cyanobacterial CO2 concentrating mechanisms (CCMs) sequester a globally significant proportion of carbon into the biosphere. Proteinaceous microcompartments, called carboxysomes, play a critical role in CCM function, housing two enzymes to enhance CO2 fixation: carbonic anhydrase (CA) and Rubisco. Despite its importance, our current understanding of the carboxysomal CAs found in [a]-cyanobacteria, CsoSCA, remains limited, particularly regarding the regulation of its activity. Here, we present the first structural and biochemical study of CsoSCA from the cyanobacterium Cyanobium PCC7001. Our results show that the Cyanobium CsoSCA is allosterically activated by the Rubisco substrate ribulose-1,5-bisphosphate (RuBP), and forms a hexameric trimer of dimers. Comprehensive phylogenetic and mutational analyses are consistent with this regulation appearing exclusively in cyanobacterial [a]-carboxysome CAs. These findings clarify the biologically relevant oligomeric state of -carboxysomal CAs and advance our understanding of the regulation of photosynthesis in this globally dominant lineage. One-Sentence SummaryThe carboxysomal carbonic anhydrase, CsoSCA, is allosterically activated by the Rubisco substrate RuBP, revealing a novel mechanism controlling key enzyme activity in cyanobacterial -carboxysomes.

molecular biology↗

bioRxiv2017

Interplay between mitochondria and diet mediates pathogen and stress resistance in C. elegans

Diet is a crucial determinant of organismal biology. Here we demonstrate the dramatic impact of a subtle shift in diet on the ability of Caenorhabditis elegans to survive pathogenic or abiotic stress. Interestingly, this shift occurs independently of canonical host defense pathways, arising instead from improvements in mitochondrial health. Using a variety of assays, we reveal that the most common C. elegans food source (E. coli OP50) results in a vitamin B12 deficiency that compromises mitochondrial homeostasis. Increasing B12 supply by feeding on E. coli HT115 or by supplementing bacterial media with methylcobalamin restored mitochondrial function, even if the bacteria were dead. B12 supplementation also efficiently increased host health without adversely affecting lifespan. Our study forges a molecular link between a dietary deficiency (nutrition/microbiota) and a physiological consequence (host sensitivity), using the host-microbiota-diet framework. The ubiquity of B12 deficiency (~10-40% of US adults) highlights the importance of our findings.

molecular biology↗

bioRxiv2020

RAS-inhibiting biologics identify and probe druggable pockets including an SII-α3 allosteric site.

RAS mutations are the most common oncogenic drivers across human cancers, but there remains a paucity of clinically-validated pharmacological inhibitors of RAS, as druggable pockets have proven difficult to identify. We have identified two RAS-binding Affimer proteins, K3 and K6, that inhibit nucleotide exchange and downstream signalling pathways with distinct isoform and mutant profiles. Affimer K6 is the first biologic to bind in the SI/SII pocket, whilst Affimer K3 is the first non-covalent inhibitor of the SII region, revealing a novel RAS conformer with a large, druggable SII/3 pocket. This work demonstrates the potential of using biologics with small interface surfaces to select novel druggable conformations in conjunction with pharmacophore identification for hard-to-drug proteins.

molecular biology↗

bioRxiv2021

Identification of the receptor-binding protein of Clostridium difficile phage CDHS-1 reveals a new class of receptor-binding domains.

As natural bacterial predators, bacteriophages have the potential to be developed to tackle antimicrobial resistance, but our exploitation of them is limited by understanding their vast uncharacterised genetic diversity1,2. Fascinatingly, this genetic diversity reflects many ways that phages can make proteins, performing similar functions that together form the familiar phage particle. Critical to infection are phage receptor-binding proteins (RBPs) that bind bacterial receptors and initiate bacterial entry3. Here we identified and characterised Gp22, a novel RBP for phage CDHS-1 that infects pathogenic C. difficile, but that had no recognisable RBPs. We showed that Gp22 antibodies neutralised CDHS-1 infection and used immunogold-labelling and transmission electron microscopy to identify their location on the capsid. The Gp22 three-dimensional structure was resolved by X-ray crystallography revealing a new RBP class with an N-terminal L-shaped -helical superhelix domain and a C-terminal Mg2+-binding domain. The findings provide novel insights into C. difficile phage biology and phage-host interactions. This will facilitate optimal phage development and future engineering strategies4,5. Furthermore, the AlphaFold2-predicted Gp22 structure, which was strikingly accurate, paves the way for a structurome based transformation and guidance of future phage studies where many proteins lack sequence homology but have recognisable protein structures.

molecular biology↗

bioRxiv2021

Global fitting for high-accuracy multi-channel single-molecule localization

Multi-channel detection in single-molecule localization microscopy (SMLM) greatly increases information content for various biological applications. Here, we present globLoc, a graphics processing unit (GPU) based global fitting algorithm with flexible PSF modeling and parameter sharing, to extract maximum information from multi-channel single molecule data. We show, both in simulations and experiments, that global fitting can substantially improve the 3D localization precision for biplane and 4Pi SMLM and color assignment for ratiometric multicolor imaging.

molecular biology↗

bioRxiv2024

Human lymphoid-neutrophil/monocyte restriction co-ordinately activates increased proliferation despite parallel heterogeneity in transcriptional changes

Recent studies indicate the human lympho-myeloid restriction process to be a different and more heterogeneous one than historically inferred. Here we describe the development of bulk and clonal culture systems that efficiently support early B-lymphoid differentiation and their use to identify biological and molecular changes that accompany their initial restriction from subsets of CD34+ human cord blood cells with lympho-myeloid-limited potential. Analyses of the changes observed revealed the acquisition of B-lymphoid- and neutrophil/monocyte (NM)-restricted properties are accompanied by a concomitantly accelerated and lineage-shared cell cycling activity and loss of self-renewal properties. Parallel, single-cell transcriptome analysis identified reduced expression of multiple self-renewal-associated genes and an accompanying heterogeneous activation of lineage-regulatory modules during the production of B, NM and dendritic cell precursors. These results uncover a connected regulation of lineage-shared proliferation control with persistent heterogeneity in the biological and transcriptional changes in the same cells undergoing B and NM lineage restriction.

immunology↗

bioRxiv2024

Differential biological effect of low doses of ionizing radiation depending on the radiosensitivity in a cell line model

PurposeExposure to low doses (LD) of ionizing radiation (IR), such as the ones employed in computed tomography (CT) examination, can be associated with cancer risk. However, not all individuals respond the same to IR, and cancer development could depend on the individual radiosensitivity. Notably, inter-individual differences in the response to IR have been very well studied for high and medium doses, but not for LD. In the present study, we wanted to evaluate the differences in the response to a CT-scan radiation dose of 20 mGy in two lymphoblastoid cell lines with different radiosensitivity. Materials and MethodsSeveral parameters were studied: gene expression, DNA damage, and its repair (by analyzing gamma-H2AX foci, chromosome breaks, and sister chromatid exchange), as well as cell viability, proliferation, and death. ResultsAfter 20 mGy of IR, the radiosensitive (RS) cell line showed an increase in DNA damage, and higher cell proliferation and apoptosis, whereas the radioresistant (RR) cell line was insensitive to this LD. Interestingly, gene expression analysis showed a higher expression of an antioxidant gene in the RR cell line, which could be used by the cells as a protective mechanism. After a dose of 500 mGy, both cell lines were affected by IR but with significant differences. The RS cells presented an increase in DNA damage and apoptosis, but a decrease in cell proliferation and cell viability, as well as less antioxidant response. ConclusionsA differential biological effect was observed between two cell lines with different radiosensitivity, and these differences are especially interesting after a CT scan dose. If this is confirmed by further studies, one could think that individuals with radiosensitivity-related genetic variants may be more vulnerable to long-term effects of IR, potentially increasing cancer risk after LD exposure.

molecular biology↗

bioRxiv2018

Toxoplasma gondii ROP18 Inhibits Human Glioblastoma Cell Apoptosis through Mitochondrial Pathway by Targeting Host Cell P2X1

It is known that Toxoplasma gondii infection both initiates and inhibits host cell apoptosis through different proapoptotic signaling cascades, but the parasitic factors involved in these processes remain unclear. T. gondii virulence factor ROP18 has been reported to regulate host cell apoptosis, but the results of this regulation are few reported and contradictory. In this study, we found that immune or neuro cells infected by any one of the T. gondii strains (RH-type I, ME49-type II, and VEG-type III) showed a significantly lower apoptosis index than their uninfected controls when apoptosis was induced by staurosporine (STS). We further found that ROP18 of RH strain inhibited ATP induced apoptosis in human glioblastoma cells (SF268) with endogenous expression of human proapoptotic protein purinergic receptor 1 (P2X1), but had no effects on the immune cells of RAW264.7 and THP-1 without detectable P2X1 expression, which may indicate that ROP18s inhibition of host cell apoptosis is related to P2X1. Interestingly, we further identified that ROP18 (RH strain) interacted with P2X1, and over-expression of ROP18 in COS-7 cells inhibited the cell apoptosis mediated by P2X1. We also found that ROP18 of RH strain inhibited P2X1-mediated Ca2+ influx, translocation of cytochrome C from mitochondria to cytoplasm, and 1 ATP-triggered caspases activation. Collectively, these findings supported that ROP18 inhibited the host cell apoptosis through the intrinsic mitochondria pathway by targeting host cell P2X1, thereby suggesting a sensor role of the host proapoptotic protein P2X1 in this process\n\nAuthor summaryThe obligate intracellular protozoan Toxoplasma gondii has been shown to modulate cell apoptosis through different apoptotic pathways. However, the consequences are various and even contradictory, and the parasite effectors and the precise biological mechanisms remain unclear. Herein we showed that T. gondii of type I, II, and III strains could inhibit the apoptosis of neuro cells and immune cells. Toxoplasma gondii ROP18 (RH strain) inhibited apoptosis of human glioblastoma cell SF268 by targeting C terminal of host cell P2X1 protein, but not through proteasome-dependent degradation of P2X1.

molecular biology↗

bioRxiv2020

Amyloid beta acts synergistically as a pro-inflammatory cytokine

The amyloid beta (A{beta}) peptide is believed to play a central role in Alzheimers disease (AD), the most common age-related neurodegenerative disorder. However, the natural, evolutionarily-selected functions of A{beta} are incompletely understood. Here, we report that nanomolar concentrations of A{beta} act synergistically with known cytokines to promote pro-inflammatory activation in primary human astrocytes (a cell type increasingly implicated in brain aging and AD). Using transcriptomics (RNA-seq), we show that A{beta} can directly substitute for the complement component C1q in a cytokine cocktail previously shown to induce astrocyte immune activation. Furthermore, we show that astrocytes synergistically activated by A{beta} have a transcriptional signature similar to neurotoxic "A1" astrocytes known to accumulate with age and in AD. Interestingly, we find that this biological action of A{beta} at low concentrations is distinct from the transcriptome changes induced by the high/supraphysiological doses of A{beta} often used in in vitro studies. Collectively, our results suggest an important, cytokine-like function for A{beta} and a novel mechanism by which it may directly contribute to the neuroinflammation associated with brain aging and AD.

molecular biology↗