Biology subjects

Chen, X.

Publications and source records attributed to Chen, X..

At least 703 records · Page 39Linked to original sources

bioRxiv2024

PHOSPHATE-INDUCED 1 and its paralogs positively regulate cellulose biosynthesis in Arabidopsis.

Cellulose is not only the most abundant biopolymer on Earth but also an essential multi-purpose resource, primarily synthesized by cellulose synthase complexes (CSCs) located on plasma membrane in vascular plants. Although insights into the functions of CSCs have been provided by the identification of cellulose synthase catalytic subunits, our understanding of the regulation of CSC activity remains limited. In this study, combining molecular and genetic methods, we demonstrated that both PHI-1 and its paralogs residing in the plasma membrane are involved in the cellulose biosynthesis regulation as a positive regulatory component of CSCs by feeding developmental and environmental cues into their activities in Arabidopsis (Arabidopsis thaliana). Our finding reveals a mechanism for how plants regulate cellulose synthesis in response to developmental and environmental signals, and thus improves the likelihood that cellulose biosynthesis could be feasibly adapted for sustainable purposes.

plant biology↗

bioRxiv2024

Adult-Onset Deletion of ATP13A2 in Mice Induces Progressive Nigrostriatal Pathway Dopaminergic Degeneration and Lysosomal Abnormalities

Although most cases of Parkinsons disease (PD) are sporadic, mutations in over 20 genes are known to cause heritable forms of PD. A surprising number of familial PD-linked genes and PD risk genes are involved in intracellular trafficking and protein degradation. Recessive loss-of-function mutations in ATP13A2, a lysosomal transmembrane P5B-type ATPase and polyamine exporter, can cause early-onset familial PD. Familial ATP13A2 mutations are also linked to related neurodegenerative diseases, including Kufor-Rakeb syndrome (KRS), hereditary spastic paraplegias (HSPs), neuronal ceroid lipofuscinosis, and amyotrophic lateral sclerosis (ALS). Given the severe effects of ATP13A2 mutations in humans, it is surprising that ATP13A2 knockout (KO) mice fail to exhibit neurodegeneration even at advanced ages. This discrepancy between human subjects and rodents makes it challenging to study the neuropathological effects of ATP13A2 loss in vivo. Germline deletion of ATP13A2 in rodents may trigger the upregulation of compensatory pathways during embryonic development that mask the full neurotoxic effects of ATP13A2 loss in the brain. To explore this idea, we selectively deleted ATP13A2 in the adult mouse brain by the unilateral delivery of an AAV-Cre vector into the substantia nigra of young adult mice carrying conditional loxP-flanked ATP13A2 KO alleles. We observe a progressive loss of striatal dopaminergic nerve terminals at 3 and 10 months after AAV-Cre delivery. Cre-injected mice also exhibit robust dopaminergic neuronal degeneration in the substantia nigra at 10 months. Adult-onset ATP13A2 KO also recreates many of the phenotypes observed in aged germline ATP13A2 KO mice, including lysosomal abnormalities, p62-positive inclusions, and neuroinflammation. Our study demonstrates that the adult-onset homozygous deletion of ATP13A2 in the nigrostriatal pathway produces robust and progressive dopaminergic neurodegeneration that serves as a useful in vivo model of ATP13A2-related neurodegenerative diseases.

neuroscience↗

bioRxiv2024

A dual-gene reporter-amplifier architecture for enhancing the sensitivity of molecular MRI by water exchange

The development of genetic reporters for magnetic resonance imaging (MRI) is essential for investigating biological functions in intact animals. However, current MRI reporters have low sensitivity, making it challenging to create significant contrast against the tissue background, especially when only a small percentage of cells express the reporter. To overcome this limitation, we developed an approach that amplifies signals by co-expressing an MRI reporter gene, Oatp1b3, with a water channel, aquaporin-1 (Aqp1). We first show that the expression of Aqp1 amplifies the paramagnetic relaxation effect of Oatp1b3 by facilitating transmembrane water exchange. This mechanism provides Oatp1b3-expressing cells with access to a larger water pool compared with typical exchange-limited conditions. We further demonstrated that our methodology allows dual-labeled cells to be detected using approximately 10-fold lower concentrations of contrast agent than that in the Aqp1-free scenario. Finally, we show that our approach enables the imaging of mixed-cell populations containing a low fraction of Oatp1b3-labeled cells that are otherwise undetectable based on Oatp1b3 expression alone.

bioengineering↗

bioRxiv2024

Alzheimer's disease-linked risk alleles elevate microglial cGAS-associated senescence and neurodegeneration in a tauopathy model

The strongest risk factors for Alzheimers disease (AD) include the {chi}4 allele of apolipoprotein E (APOE), the R47H variant of triggering receptor expressed on myeloid cells 2 (TREM2), and female sex. Here, we combine APOE4 and TREM2R47H (R47H) in female P301S tauopathy mice to identify the pathways activated when AD risk is the strongest, thereby highlighting disease-causing mechanisms. We find that the R47H variant induces neurodegeneration in female APOE4 mice without impacting hippocampal tau load. The combination of APOE4 and R47H amplified tauopathy-induced cell-autonomous microglial cGAS-STING signaling and type-I interferon response, and interferon signaling converged across glial cell types in the hippocampus. APOE4-R47H microglia displayed cGAS- and BAX-dependent upregulation of senescence, showing association between neurotoxic signatures and implicating mitochondrial permeabilization in pathogenesis. By uncovering pathways enhanced by the strongest AD risk factors, our study points to cGAS-STING signaling and associated microglial senescence as potential drivers of AD risk.

neuroscience↗

bioRxiv2024

Integrative genomic analysis identifies unique immune environments associated with immunotherapy response in diffuse large B cell lymphoma

Most diffuse large B-cell lymphoma (DLBCL) patients treated with bispecific antibodies (BsAb) or chimeric antigen receptor (CAR) T cells fail to achieve durable treatment responses, underscoring the need for a deeper understanding of mechanisms that regulate the immune environment and response to treatment. Here, an integrative, multi-omic approach was employed to characterize DLBCL immune environments, which effectively segregated DLBCLs into four quadrants - termed DLBCL-immune quadrants (IQ) - defined by cell-of-origin and immune-related gene set expression scores. Recurrent genomic alterations were enriched in each IQ, suggesting that lymphoma cell-intrinsic alterations contribute to orchestrating unique DLBCL immune environments. In relapsed/refractory DLBCL patients, DLBCL-IQ assignment correlated significantly with clinical benefit with the CD20 x CD3 BsAb, mosunetuzumab, but not with CD19-directed CAR T cells. DLBCL-IQ provides a new framework to conceptualize the DLBCL immune landscape and uncovers the differential impact of the endogenous immune environment on outcomes to BsAb and CAR T cell treatment.

cancer biology↗

bioRxiv2024

Neutralizing gut-derived lipopolysaccharide as a novel therapeutic strategy for severe leptospirosis

Leptospirosis is an emerging infectious disease caused by pathogenic Leptospira spp. Humans and some mammals can develop severe forms of leptospirosis accompanied by a dysregulated inflammatory response, which often results in death. The gut microbiota has been increasingly recognized as a vital element in systemic health. However, the precise role of the gut microbiota in severe leptospirosis is still unknown. Here, we aimed to explore the function and potential mechanisms of the gut microbiota in a hamster model of severe leptospirosis. Our study showed that leptospires were able to multiply in the intestine, cause pathological injury, and induce intestinal and systemic inflammatory responses. 16S rRNA gene sequencing analysis revealed that Leptospira infection changed the composition of the gut microbiota of hamsters with an expansion of Proteobacteria. In addition, gut barrier permeability was increased after infection, as reflected by a decrease in the expression of tight junctions. Translocated Proteobacteria were found in the intestinal epithelium of moribund hamsters, as determined by fluorescence in situ hybridization, with elevated LPS levels in the serum. Moreover, gut microbiota depletion reduced the survival time, increased the leptospiral load, and promoted the expression of proinflammatory cytokines after Leptospira infection. Intriguingly, fecal filtration and serum from moribund hamsters both increased the transcription of TNF-, IL-1{beta}, IL-10, and TLR4 in macrophages compared with those from uninfected hamsters. These stimulating activities were inhibited by LPS neutralization using polymyxin B. Based on our findings, we identified an LPS neutralization therapy that significantly improved the survival rates in severe leptospirosis when used in combination with antibiotic therapy or polyclonal antibody therapy. In conclusion, our study not only uncovers the role of the gut microbiota in severe leptospirosis but also provides a therapeutic strategy for severe leptospirosis.

microbiology↗

bioRxiv2024

Genomes of Aegilops umbellulata provide new insights into unique structural variations and genetic diversity in the U-genome for wheat improvement

Aegilops spp. serve as an important reservoir for novel sources of resistance or tolerance to biotic and abiotic stresses. To harness this reservoir, we have generated a high-quality chromosome-level genome assembly of an Ae. umbellulata accession PI 554389 using a combination of PacBio HiFi, Oxford nanopore, and chromosome conformation capture (Hi-C) sequencing technologies and resequenced 20 Ae. umbellulata genomes using Illumina sequencing. We assembled a 4.20 Gb genome spanned over seven chromosomes, rich in repetitive elements ([~]84%), achieving a QV of 59.54 with 98.14% completeness. The phylogenetic analysis places the U-genome with D-lineage, but major and distinct rearrangements were revealed in the U-genome. Unique transposon landscape of diploid U-genome and complex chromosomal rearrangements, most prominently in 4U and 6U chromosomes uncovered a distinct evolutionary trajectory of Ae. umbellulata. Additionally, the resequencing of geographically and morphologically diverse Ae. umbellulata accessions revealed three distinctive evolutionary sub-populations. Resequencing also identified six new haplotypes for Lr9, the first leaf rust resistance gene introgressed and cloned from Ae. umbellulata. These genomics resources along with high levels of resistance in the resequenced accessions against five devastating wheat diseases affirmed the genetic potential of Ae. umbellulata for wheat improvement.

plant biology↗

bioRxiv2024

Brain and cancer associated binding domain mutations provide insight into CTCF's relationship with chromatin and its ability to act as a chromatin organizer

Here we used a series of CTCF mutations to explore CTCFs relationship with chromatin and its contribution to gene regulation. CTCFs impact depends on the genomic context of bound sites and the unique binding properties of WT and mutant CTCF proteins. Specifically, CTCFs signal strength is linked to changes in accessibility, and the ability to block cohesin is linked to its binding stability. Multivariate modelling reveals that both CTCF and accessibility contribute independently to cohesin binding and insulation, however CTCF signal strength has a stronger effect. CTCF and chromatin have a bidirectional relationship such that at CTCF sites, accessibility is reduced in a cohesin-dependent, mutant specific fashion. In addition, each mutant alters TF binding and accessibility in an indirect manner, changes which impart the most influence on rewiring transcriptional networks and the cells ability to differentiate. Collectively, the mutant perturbations provide a rich resource for determining CTCFs site-specific effects.

molecular biology↗

bioRxiv2024

Absolute quantitative and base-resolution sequencing reveals comprehensive landscape of pseudouridine across the human transcriptome

Pseudouridine ({Psi}) is one of the most abundant modifications in cellular RNA. However, its function remains elusive, mainly due to the lack of highly sensitive and accurate detection methods. To address this challenge, we introduced 2-bromoacrylamide-assisted cyclization sequencing (BACS) for quantitative profiling of {Psi} at single-base resolution. Based on novel bromoacrylamide cyclization chemistry, BACS enables a {Psi}-to-C transition. Compared to previous methods, BACS allowed the precise identification of {Psi} positions, especially in densely modified {Psi} regions and consecutive uridine sequences. BACS successfully detected all known {Psi} sites in human rRNA and spliceosomal snRNAs and generated the first quantitative {Psi} map of human snoRNA and tRNA. Furthermore, BACS simultaneously detected adenosine-to-inosine (A-to-I) editing sites and N1-methyladenosine (m1A). Depletion of three key pseudouridine synthases (PUS) enabled us to elucidate the targets and sequence motifs of TRUB1, PUS7, and PUS1 in HeLa cells. We further applied BACS to Epstein-Barr virus (EBV)-encoded small RNAs (EBERs) and identified a highly abundant {Psi}114 site in EBER2. Surprisingly, applying BACS to a panel of RNA viruses demonstrated the absence of {Psi} in their viral transcripts or genomes, shedding light on differences in pseudouridylation between virus families. We anticipate BACS to serve as a powerful tool to uncover the biological importance of {Psi} in future studies.

molecular biology↗

bioRxiv2024

Coordinated molecular and ecological adaptations underlie a highly successful parasitoid

The success of an organism depends on the molecular and ecological adaptations that promote its beneficial fitness. Parasitoids are valuable biocontrol agents for successfully managing agricultural pests, and they have evolved diversified strategies to adapt to both the physiological condition of hosts and the competition of other parasitoids. Here, we deconstructed the parasitic strategies in a highly successful parasitoid, Trichopria drosophilae, which parasitizes a broad range of Drosophila hosts, including the globally invasive species D. suzukii. We found that T. drosophilae had developed specialized venom proteins that arrest host development to obtain more nutrients via secreting tissue inhibitors of metalloproteinases (TIMPs), as well as a unique type of cell-- teratocytes--that digest host tissues for feeding by releasing trypsin proteins. In addition to the molecular adaptations that optimize nutritional uptake, this pupal parasitoid has evolved ecologically adaptive strategies including the conditional tolerance of intraspecific competition to enhance parasitic success in older hosts and the obligate avoidance of interspecific competition with larval parasitoids. Our study not only demystifies how parasitoids weaponize themselves to colonize formidable hosts but also provided empirical evidence of the intricate coordination between the molecular and ecological adaptations that drive evolutionary success.

ecology↗

bioRxiv2024

Signaling events at TMEM doorways provide potential targets for inhibiting breast cancer dissemination

Tumor cell intravasation is essential for metastatic dissemination, but its exact mechanism is incompletely understood. We have previously shown that in breast cancer, the direct and stable association of a tumor cell expressing Mena, a Tie2hi/VEGFhi macrophage, and a vascular endothelial cell, creates an intravasation portal, called a "tumor microenvironment of metastasis" (TMEM) doorway, for tumor cell intravasation, leading to dissemination to distant sites. The density of TMEM doorways, also called TMEM doorway score, is a clinically validated prognostic marker of distant metastasis in breast cancer patients. Although we know that tumor cells utilize TMEM doorway-associated transient vascular openings to intravasate, the precise signaling mechanisms involved in TMEM doorway function are only partially understood. Using two mouse models of breast cancer and an in vitro assay of intravasation, we report that CSF-1 secreted by the TMEM doorway tumor cell stimulates local secretion of VEGF-A from the Tie2hi TMEM doorway macrophage, leading to the dissociation of endothelial junctions between TMEM doorway associated endothelial cells, supporting tumor cell intravasation. Acute blockade of CSF-1R signaling decreases macrophage VEGF-A secretion as well as TMEM doorway-associated vascular opening, tumor cell trans-endothelial migration, and dissemination. These new insights into signaling events regulating TMEM doorway function should be explored further as treatment strategies for metastatic disease.

cancer biology↗

bioRxiv2024

TDP-43 pathology links innate and adaptive immunity in amyotrophic lateral sclerosis

Amyotrophic lateral sclerosis is the most common fatal motor neuron disease. Approximately 90% of ALS patients exhibit pathology of the master RNA regulator, Transactive Response DNA Binding protein (TDP-43). Despite the prevalence TDP-43 pathology in ALS motor neurons, recent findings suggest immune dysfunction is a determinant of disease progression in patients. Whether TDP-43 pathology elicits disease-modifying immune responses in ALS remains underexplored. In this study, we demonstrate that TDP-43 pathology is internalized by antigen presenting cells, causes vesicle rupture, and leads to innate and adaptive immune cell activation. Using a multiplex imaging platform, we observed interactions between innate and adaptive immune cells near TDP-43 pathological lesions in ALS brain. We used a mass cytometry-based whole-blood stimulation assay to provide evidence that ALS patient peripheral immune cells exhibit responses to TDP-43 aggregates. Taken together, this study provides a novel link between TDP-43 pathology and ALS immune dysfunction, and further highlights the translational and diagnostic implications of monitoring and manipulating the ALS immune response.

cell biology↗

bioRxiv2024

Human cell surface-AAV interactomes identify LRP6 as blood-brain-barrier transcytosis receptor and immune cytokine IL3 as AAV9 binder

Adeno-associated viruses (AAVs) are foundational gene delivery tools for basic science and clinical therapeutics. However, lack of mechanistic insight, especially for engineered vectors created by directed evolution, can hamper their application. Here, we adapted an unbiased human cell microarray platform to determine the extracellular and cell surface interactomes of natural and engineered AAVs. We identified a naturally-evolved and serotype-specific interaction between the AAV9 capsid and human interleukin 3 (IL3), with possible roles in host immune modulation, as well as lab-evolved low-density-lipoprotein-receptor-related-protein 6 (LRP6) interactions specific to engineered capsids that cross the blood-brain barrier in non-human primates after intravenous administration. The unbiased cell microarray screening approach also allowed us to identify off-target tissue binding interactions of engineered brain-enriched AAV capsids that may inform vectors peripheral organ tropism and side effects. These results allow confident application of engineered AAVs in diverse organisms and unlock future target-informed engineering of improved viral and non-viral vectors for non-invasive therapeutic delivery to the brain.

bioengineering↗

bioRxiv2024

Suppression of epileptic seizures by transcranial activation of K+-selective channelrhodopsin

Optogenetics is a valuable tool for studying the mechanisms of neurological diseases and is now being developed for therapeutic applications. In rodents and macaques, improved channelrhodopsins have been applied to achieve transcranial optogenetic stimulation. While transcranial photoexcitation of neurons has been achieved, noninvasive optogenetic inhibition for treating hyperexcitability-induced neurological disorders has remained elusive. There is a critical need for effective inhibitory optogenetic tools that are highly light-sensitive and capable of suppressing neuronal activity in deep brain tissue. In this study, we developed a highly sensitive K+-conductive channelrhodopsin (hsKCR) by molecular engineering of the recently discovered Hyphochytrium catenoides kalium (potassium) channelrhodopsin 1. Transcranial activation of hsKCR significantly prolongs the time to the first seizure, increases survival, and decreases seizure activity in several mouse epileptic models. Our approach for transcranial optogenetic inhibition of neural hyperactivity may be adapted for cell type-specific neuromodulation in both basic and preclinical settings.

neuroscience↗

bioRxiv2024

A PACAP-activated network for secretion requires coordination of calcium influx and calcium mobilization

Chromaffin cells of the adrenal medulla transduce sympathetic nerve activity into stress hormone secretion. The two neurotransmitters principally responsible for coupling cell stimulation to secretion are acetylcholine and pituitary adenylate activating polypeptide (PACAP). In contrast to acetylcholine, PACAP evokes a persistent secretory response from chromaffin cells. However, the mechanisms by which PACAP acts are poorly understood. Here, it is shown that PACAP induces sustained increases in cytosolic Ca2+ which are disrupted when Ca2+ influx through L-type channels is blocked or internal Ca2+ stores are depleted. PACAP liberates stored Ca2+ via inositol trisphosphate receptors (IP3Rs) on the endoplasmic reticulum (ER), thereby functionally coupling Ca2+ mobilization to Ca2+ influx and supporting Ca2+-induced Ca2+-release. These Ca2+ influx and mobilization pathways are unified by an absolute dependence on phospholipase C epsilon (PLC{varepsilon}) activity. Thus, the persistent secretory response that is a defining feature of PACAP activity, in situ, is regulated by a signaling network that promotes sustained elevations in intracellular Ca2+ through multiple pathways.

cell biology↗

bioRxiv2023

Spatial genomics of AAVs reveals mechanism of transcriptional crosstalk that enables targeted delivery of large genetic cargo

Integrating cell type-specific regulatory elements (e.g. enhancers) with recombinant adeno-associated viruses (AAVs) can provide broad and efficient genetic access to specific cell types. However, the packaging capacity of AAVs restricts the size of both the enhancers and the cargo that can be delivered. Transcriptional crosstalk offers a novel paradigm for cell type-specific expression of large cargo, by separating distally-acting regulatory elements into a second AAV genome. Here, we identify and profile transcriptional crosstalk in AAV genomes carrying 11 different enhancers active in mouse brain. To understand transcriptional crosstalk, we develop spatial genomics methods to identify and localize AAV genomes and their concatemeric forms in cultured cells and in tissue. Using these methods, we construct detailed views of the dynamics of AAV transduction and demonstrate that transcriptional crosstalk is dependent upon concatemer formation. Finally, we leverage transcriptional crosstalk to drive expression of a large Cas9 cargo in a cell type-specific manner with systemically-administered engineered AAVs and demonstrate AAV-delivered, minimally-invasive, cell type-specific gene editing in wildtype animals that recapitulates known disease phenotypes. HighlightsO_LITranscriptional crosstalk between enhancers and promoters delivered in trans by AAVs is a generalized phenomenon. C_LIO_LISpatial genomics techniques, AAV-Zombie and SpECTr, reveal that AAV genome concatemerization facilitates transcriptional crosstalk. C_LIO_LITranscriptional crosstalk can be leveraged for minimally-invasive, targeted AAV delivery of large cargo, including machinery for CRISPR-based gene editing and manipulation. C_LIO_LITranscriptional crosstalk enables cell-type specific gene disruption in wildtype animals, recapitulating behavioural phenotypes of genetic knockouts. C_LI

bioengineering↗

bioRxiv2023

A direct estrogenic involvement in the expression of human hypocretin

BackgroundHypocretin-1 may play an important role in depression, which was increased in female depression in LHA in our previous study. Estrogen acts through its nuclear transcriptional receptors. MethodsWe studied the possibility of a direct action of estrogen receptors on the expression of human hypocretin. To explore the activation of ERs by hypocretin, we first investigated the potential presence of co-localization of the estrogen receptors (ERs) in hypocretin-immunoreactive neurons in the lateral hypothalamic area (LHA) in human and nuclear ERs in hypocretin immunoreactive neuroblastoma SK-N-SH cells. We investigated the relationship between E2 and hypocretin in female rats. After we explored the regulating role of exogenous estrogen on hypocretin gene expression via ERs through Chromatin immunoprecipitation, Electrophoretic mobility shift assay and Luciferase assay. ResultsWe found that hypocretin-1 plasma level was significantly higher in female depression. Estrogen receptors (ER and ER{beta}) and hypocretin are co-localized in female depression LHA, PC12 and SK-N-SH cell lines. The estrogen receptor response elements (EREs) existing in the hypocretin promoter region may directly regulate the expression of hypocretin, the synchronicity of change of hypocretin and estradiol both in hypothalamus and plasma was verified in female rats. With the presence of estradiol, there is specific binding between human ER and the Hypocretin-ERE. Expression of ER combined with estradiol repressed hypocretin promoter activity through the ERE. Conclusionswe defined that estradiol may affect hypocretin neurons in the human hypothalamus via ER binding to the Hypocretin-ERE directly, which may thus lead to the sex-specific pathogenesis of depression.

neuroscience↗

bioRxiv2023

ARBUR: a Machine Learning-based Analysis System for Relating Behaviors and Ultrasonic Vocalizations of Rats

Deciphering how different types of behavior and ultrasonic vocalizations (USVs) of rats interact can yield insights into the neural basis of social interaction. However, the behavior-vocalization interplay of rats remains elusive because of the challenges of relating the two communication media in complex social contexts. Here, we propose a machine learning-based analysis system (ARBUR) that can cluster without bias both non-step (continuous) and step USVs, hierarchically detect eight types of behavior of two freely behaving rats with high accuracy, and locate the vocal rat in 3-D space. By simultaneously recording the video and ultrasonic streams of two freely behaving rats, ARBUR reveals that rats communicate via distinct USVs when engaging in different types of social behavior. Moreover, we show that ARBUR can not only automatically reveal the well-understood behavior-associated vocalizations that were carefully concluded by other behavioral researchers, but also hold the promise to indicate novel findings that can be hardly found by manual analysis, especially regarding step USVs and the active/passive rat-associated USVs during easy-to-confuse social behaviors. This work highlights the potential of machine learning algorithms in automatic animal behavioral and acoustic analysis and could help mechanistically understand the interactive influence between the behaviors and USVs of rats.

animal behavior and cognition↗