Biology subjects

Zhuang, X.

Publications and source records attributed to Zhuang, X..

At least 37 records · Page 2Linked to original sources

bioRxiv2025

A Cell-Autonomous Role for the Vitamin B6 Metabolism Gene PNPO in Drosophila GABAergic Neurons

In animals, the enzyme pyridox(am)ine 5-phosphate oxidase (PNPO) is critical for synthesizing the active form of vitamin B6 (VB6), pyridoxal 5-phosphate (PLP), from inactive vitamers. PLP is a required cofactor for many enzymatic reactions, including the synthesis of GABA and the monoamines. PNPO disruption in humans is associated with an array of epilepsy syndromes, while Drosophila harboring mutations in the sole PNPO ortholog, sugarlethal (sgll), display spontaneous seizures and short lifespans. These phenotypes are suppressed by PLP supplementation and are exacerbated by restriction of dietary B6 vitamers. In the context of PNPO deficiency, it remains to be resolved what the specific contributions by cellular subpopulations in the nervous system are to neurological phenotypes. We addressed this question in sgll mutants by expressing human PNPO (hPNPO) cDNA in cholinergic, glutamatergic, and GABAergic neurons as well as glia and measuring changes in survival and seizure phenotypes. We found hPNPO expression in GABAergic neurons largely restored lifespan and attenuated seizure activity, while glial expression also improved sgll phenotypes albeit to a lesser degree. In contrast, hPNPO expression in either cholinergic or glutamatergic neurons, accounting for most neurons in the fly brain, did not appreciably alter sgll phenotypes. We contrasted these observations with changes in sgll mutants induced by feeding GABA receptor modulators. The GABAB agonist SKF-97541 reduced mortality, while GABA or GABAA receptor modulators did not improve survival. Together, our data establish a cell-autonomous role for PNPO in GABAergic neurons to support brain function, especially under VB6-restricted conditions.

neuroscience↗

bioRxiv2025

High-resolution spatial mapping of cell state and lineage dynamics in vivo with PEtracer

Charting the spatiotemporal dynamics of cell fate determination in development and disease is a long-standing objective in biology. Here we present the design, development, and extensive validation of PEtracer, a prime editing-based, evolving lineage tracing technology compatible with both single-cell sequencing and multimodal imaging methodologies to jointly profile cell state and lineage in dissociated cells or while preserving cellular context in tissues with high spatial resolution. Using PEtracer coupled with MERFISH spatial transcriptomic profiling in a syngeneic mouse model of tumor metastasis, we reconstruct the growth of individually-seeded tumors in vivo and uncover distinct modules of cell-intrinsic and cell-extrinsic factors that coordinate tumor growth. More generally, PEtracer enables systematic characterization of cell state and lineage relationships in intact tissues over biologically-relevant temporal and spatial scales.

molecular biology↗

bioRxiv2025

Molecular and Spatial Organization of the Primary Olfactory System and its Responses to Social Odors

The detection of olfactory cues is essential to signal food, predators, and social encounters. To determine how the sensory detection of physiologically relevant odors is systematically mapped into the mouse primary olfactory system, we used Multiplexed Error Robust Fluorescent In Situ Hybridization (MERFISH) to construct a molecular atlas of odorant receptor (OR) expression in the main olfactory epithelium (MOE) and olfactory bulb (OB). We comprehensively quantified the expression of the mouse OR repertoire and uncovered stereotypical gradients of sensory neuron distribution in the MOE along two, central-to-peripheral and basal-to-apical, axes. Projections of sensory neurons mirror MOE gradients along the dorsal-ventral and anterior-posterior axes of the OB, respectively. Integration with sequencing data revealed candidate signaling molecules underlying this spatial organization. Co-imaging OR and activity marker expression identified distinct spatial domains of sensory responses in the MOE and OB, providing a topographical basis for olfactory responses to ethologically relevant odors.

neuroscience↗

bioRxiv2025

Effect of Shufeng Xingbi Therapy on Th1/Th2 immune balance and intestinal flora in rats with allergic rhinitis

ObjectiveTo study the effect of Shufeng Xingbi Therapy (SFXBT) on Th1/Th2 immune balance and intestinal flora in allergic rhinitis (AR) rats. Methods32 SD rats were randomly divided into control group, OVA group, antibiotic + SFXBT group and acetic acid + SFXBT group. Evaluate the AR behavioral score, observe the pathological changes of nasal mucosa with H&E, detect the composition and content of colonic contents with 16S rDNA, the levels of serum IgE, IL-4 and short-chain fatty acid (SCFAs) with ELISA, the mRNA expressions of STAT5, STAT6 and GATA3 in nasal mucosa with RT-qPCR, and the protein expressions of IL-4, STAT5, STAT6 and GATA3 in nasal mucosa with Western Blot. ResultsCompared with the OVA group, the AR behavioral score in the antibiotic + SFXBT group and acetic acid + SFXBT group decreased (P < 0.01), and the pathological changes of nasal mucosa were alleviated. At the door level, the relative abundance of Firmicutes in feces increased significantly, while the relative abundance of Bacteroidetes decreased significantly. At the genus level, The relative abundance of fecal Lactobacillus, Romboutsia, Allobaculum and Dubosiella increased significantly, the levels of serum IgE and IL-4 decreased (P < 0.05), the content of SCFAs increased significantly (P < 0.05), and the expression levels of STAT5, STAT6 and GATA3 mRNA and protein in nasal mucosa decreased significantly (P < 0.05). ConclusionShufeng Xingbi Therapy can significantly improve the inflammatory symptoms of nasal mucosa in AR rats, and its mechanism may be closely related to regulating Th1/Th2 immune balance and intestinal flora.

immunology↗

bioRxiv2025

Overexpression of alpha synuclein in Midbrain Dopamine Neurons Reduces Dopamine Release Without Cell Loss and Drives Mild Motor Deficits in Mice

It has proven challenging to faithfully recapitulate the key pathological, physiological, and behavioral features of Parkinsons Disease (PD) in animals. Here we used adeno-associated virus (AAV) vectors to achieve cell type-specific overexpression of wild-type human -synuclein (syn) and a fluorophore (mCherry) in midbrain dopamine neurons to model PD in mice. We found that AAVs drove selective expression of both syn and mCherry in midbrain dopamine neurons. In conjunction with approximately 2-fold overexpression of syn, we found several histopathological markers of PD-like pathology, including progressive accumulation of phosphorylated and aggregated syn, ubiquitin, and a reduction in the expression of tyrosine hydroxylase, without overt cell loss. In parallel, syn overexpression drove a profound loss of evoked dopamine release, without a substantive change in the intrinsic properties of dopamine neurons, nor in striatal dopamine content. Finally, syn overexpression led to mild locomotor deficits. Together, these findings suggest that moderate syn overexpression can mimic some aspects of premotor and early symptomatic phases of PD, including markers of Lewy Body-like pathology and functional loss of evoked dopamine release. This model may be useful for investigating cellular and circuit mechanisms related to PD pathogenesis and progression.

neuroscience↗

bioRxiv2025

Genetic vitamin B6 deficiency and alcohol interaction in behavior and metabolism

Alcohol abuse is a leading cause of preventable deaths, affecting brain function and metabolism, including GABA transmission and vitamin B6 (VB6) levels. However, the interaction between genetic VB6 deficiency and alcohol consumption remains unexplored. Here, we utilized Drosophila models with mutations in pyridox(am)ine-5-phosphate oxidase (PNPO), a key enzyme in VB6 metabolism, to examine this interaction at behavioral and biochemical levels. Our findings demonstrate that PNPO deficiency reduces alcohol aversion, increases consumption, and alters locomotor behavior. Biochemically, PNPO deficiency and alcohol exposure converge on amino acid metabolism, elevating inhibitory neurotransmitters GABA and glycine. Moreover, both PNPO deficiency and alcohol exposure lead to lethality with significant interaction, which can be rescued by VB6 supplementation. These results highlight a functional interaction between genetic VB6 deficiency and alcohol, suggesting potential therapeutic strategies for alcohol-related behaviors.

animal behavior and cognition↗

bioRxiv2024

A platform for multimodal in vivo pooled genetic screens reveals regulators of liver function

Organ function requires coordinated activities of thousands of genes in distinct, spatially organized cell types. Understanding the basis of emergent tissue function requires approaches to dissect the genetic control of diverse cellular and tissue phenotypes in vivo. Here, we develop paired imaging and sequencing methods to construct large-scale, multi-modal genotype-phenotypes maps in tissue with pooled genetic perturbations. Using imaging, we identify genetic perturbations in individual cells while simultaneously measuring their gene expression and subcellular morphology. Using single-cell sequencing, we measure transcriptomic responses to the same genetic perturbations. We apply this approach to study hundreds of genetic perturbations in the mouse liver. Our study reveals regulators of hepatocyte zonation and liver unfolded protein response, as well as distinct pathways that cause hepatocyte steatosis. Our approach enables new ways of interrogating the genetic basis of complex cellular and organismal physiology and provides crucial training data for emerging machine-learning models of cellular function.

genomics↗

bioRxiv2024

Multi-purpose controllable protein generation via prompted language models

Deep learning is increasingly powerful for designing proteins that meet structural and functional requirements. However, most existing methods follow a conventional pipeline: first defining a backbone structure and then generating sequences consistent with it. This approach, which encodes all design goals indirectly through structures, restricts flexibility and struggles to address multiple, complex design objectives simultaneously. We present PROPEND, a multi-purpose protein sequence design method based on the "pre-train and prompt" framework. We show PROPENDs broad utility and accuracy both in silico and in vitro by directly controlling multiple properties through the prompt of backbones, blueprints, functional tags, and their combinations. For the five sequences tested with in vitro experiments, PROPEND achieved a maximum functional recovery of 105.2%, significantly outperforming the classical design pipelines 50.8%.

synthetic biology↗

bioRxiv2024

Biomolecular condensation of ERC1 recruits ATG8 and NBR1 to drive autophagosome formation for plant heat tolerance

Macroautophagy (hereafter autophagy) is essential for cells to respond to nutrient stress by delivering cytosolic contents to vacuoles for degradation via the formation of a multi-layer vesicle named autophagosome. A set of autophagy-related (ATG) regulators are recruited to the phagophore assembly site for the initiation of phagophore, as well as its expansion and closure and subsequent delivery into the vacuole. However, it remains elusive that how the phagophore assembly is regulated under different stress conditions. Here, we described an unknown Arabidopsis (Arabidopsis thaliana) cytosolic ATG8-interaction protein family (ERC1/2), that binds ATG8 and NBR1 to promote autophagy. ERC1 proteins translocate to the phagophore membrane and develop into classical ring-like autophagosomes upon autophagic induction. However, ERC1 proteins form large droplets together with ATG8e proteins when in the absence of ATG8 lipidation activity. We described the property of these structures as phase-separated membraneless condensates by solving the in vivo organization with spatial and temporal resolution. Moreover, ERC1 condensates elicits a strong recruitment of the autophagic receptor NBR1. Loss of ERC1 suppressed NBR1 turnover and attenuated plant tolerance to heat stress condition. This work provides novel insights into the mechanical principle of phagophore initiation via an unreported ERC1-mediated biomolecular condensation for heat tolerance in Arabidopsis.

plant biology↗

bioRxiv2024

YTHDF1 mediates translational control by m6A mRNA methylation in adaptation to environmental challenges

Animals adapt to environmental challenges with long-term changes at the behavioral, circuit, cellular, and synaptic levels which often require new protein synthesis. The discovery of reversible N6-methyladenosine (m6A) modifications of mRNA has revealed an important layer of post-transcriptional regulation which affects almost every phase of mRNA metabolism and therefore translational control. Many in vitro and in vivo studies have demonstrated the significant role of m6A in cell differentiation and survival, but its role in adult neurons is understudied. We used cell-type specific gene deletion of Mettl14, which encodes one of the subunits of the m6A methyltransferase, and Ythdf1, which encodes one of the cytoplasmic m6A reader proteins, in dopamine D1 receptor expressing or D2 receptor expressing neurons. Mettl14 or Ythdf1 deficiency blunted responses to environmental challenges at the behavioral, cellular, and molecular levels. In three different behavioral paradigms, gene deletion of either Mettl14 or Ythdf1 in D1 neurons impaired D1-dependent learning, whereas gene deletion of either Mettl14 or Ythdf1 in D2 neurons impaired D2-dependent learning. At the cellular level, modulation of D1 and D2 neuron firing in response to changes in environments was blunted in all three behavioral paradigms in mutant mice. Ythdf1 deletion resembled impairment caused by Mettl14 deletion in a cell type-specific manner, suggesting YTHDF1 is the main mediator of the functional consequences of m6A mRNA methylation in the striatum. At the molecular level, while striatal neurons in control mice responded to elevated cAMP by increasing de novo protein synthesis, striatal neurons in Ythdf1 knockout mice didnt. Finally, boosting dopamine release by cocaine drastically increased YTHDF1 binding to many mRNA targets in the striatum, especially those that encode structural proteins, suggesting the initiation of long-term neuronal and/or synaptic structural changes. While the m6A-YTHDF1 pathway has similar functional significance at cellular level, its cell type specific deficiency in D1 and D2 neurons often resulted in contrasting behavioral phenotypes, allowing us to cleanly dissociate the opposing yet cooperative roles of D1 and D2 neurons.

neuroscience↗

bioRxiv2024

Aging limits stemness and tumorigenesis in the lung by reprogramming iron homeostasis

Aging is associated with a decline in the number and fitness of adult stem cells1-4. Aging-associated loss of stemness is posited to suppress tumorigenesis5,6, but this hypothesis has not been tested in vivo. Here, using physiologically aged autochthonous genetically engineered mouse models and primary cells7,8, we demonstrate aging suppresses lung cancer initiation and progression by degrading stemness of the alveolar cell of origin. This phenotype is underpinned by aging-associated induction of the transcription factor NUPR1 and its downstream target lipocalin-2 in the cell of origin in mice and humans, leading to a functional iron insufficiency in the aged cells. Genetic inactivation of the NUPR1--lipocalin-2 axis or iron supplementation rescue stemness and promote tumorigenic potential of aged alveolar cells. Conversely, targeting the NUPR1-- lipocalin-2 axis is detrimental to young alveolar cells via induction of ferroptosis. We find that aging-associated DNA hypomethylation at specific enhancer sites associates with elevated NUPR1 expression, which is recapitulated in young alveolar cells by inhibition of DNA methylation. We uncover that aging drives a functional iron insufficiency, which leads to loss of stemness and tumorigenesis, but promotes resistance to ferroptosis. These findings have significant implications for the therapeutic modulation of cellular iron homeostasis in regenerative medicine and in cancer prevention. Furthermore, our findings are consistent with a model whereby most human cancers initiate in young individuals, revealing a critical window for such cancer prevention efforts.

cancer biology↗

bioRxiv2024

Multiplexed optical barcoding and sequencing for spatial omics

Spatial omics has brought a fundamental change in the way that we study cell and tissue biology in health and disease. Among various spatial omics methods, genome-scale imaging allows transcriptomic, 3D-genomic, and epigenomic profiling of individual cells with high spatial (subcellular) resolution but often requires a preselection of targeted genes or genomic loci. On the other hand, spatially dependent barcoding of molecules followed by sequencing provides untargeted, genome-wide profiling but typically lacks single-cell resolution. Here, we report a spatial omics method that could potentially combine the power of the two approaches by optically controlled spatial barcoding followed by sequencing. Specifically, we utilize patterned light to encode the locations of cells in tissues using oligonucleotide-based barcodes and then identify the barcoded molecular content, such as mRNAs, by sequencing. This optical barcoding method is designed with multiplexing and error-correction capacity and achieved by a light-directed ligation chemistry that attaches distinct nucleic-acid sequences to the reverse transcribed cDNA products at different locations. As a proof of principle for this method, we demonstrated high-efficiency in situ light-directed ligation, spatially dependent barcoding with multiplexed light-controlled ligations, and high-accuracy detection of spatially barcoded mRNAs in cells.

genomics↗

bioRxiv2024

Diverse origins of near-identical antifreeze proteins in unrelated fish lineages provide insights into evolutionary mechanisms of new gene birth and protein sequence convergence

Determining the origins of novel genes and the genetic mechanisms underlying the emergence of new functions is challenging yet crucial for understanding evolutionary innovations. The convergently evolved fish antifreeze proteins provide excellent opportunities to investigate evolutionary origins and pathways of new genes. Particularly notable is the near-identical type I antifreeze proteins (AFPI) in four phylogenetically divergent fish taxa. This study tested the hypothesis of protein sequence convergence beyond functional convergence in three unrelated AFPI-bearing fish lineages, revealing different paths by which a similar protein arose from diverse genomic resources. Comprehensive comparative analyses of de novo sequenced genome of the winter flounder and grubby sculpin, available high-quality genome of the cunner and 14 other relevant species found that the near-identical AFPI originated from a distinct genetic precursor in each lineage. Each independently evolved a coding region for the novel ice-binding protein while retaining sequence identity in the regulatory regions with their respective ancestor. The deduced evolutionary processes and molecular mechanisms are consistent with the Innovation-Amplification-Divergence (IAD) model applicable to AFPI formation in all three lineages, a new Duplication-Degeneration-Divergence (DDD) model we propose for the sculpin lineage, and a DDD model with gene fission for the cunner lineage. This investigation illustrates the multiple ways by which a novel functional gene with sequence convergence at the protein level could evolve across divergent species, advancing our understanding of the mechanistic intricacies in new gene formation.

evolutionary biology↗

bioRxiv2024

Opposing Motor Memories in the Direct and Indirect Pathways of the Basal Ganglia

Loss of dopamine neurons causes motor deterioration in Parkinsons disease patients. We have previously reported that in addition to acute motor impairment, the impaired motor behavior is encoded into long-term memory in an experience-dependent and task-specific manner, a phenomenon we refer to as aberrant inhibitory motor learning. Although normal motor learning and aberrant inhibitory learning oppose each other and this is manifested in apparent motor performance, in the present study, we found that normal motor memory acquired prior to aberrant inhibitory learning remains preserved in the brain, suggesting the existence of independent storage. To investigate the neuronal circuits underlying these two opposing memories, we took advantage of the RNA-binding protein YTHDF1, an m6A RNA methylation reader involved in the regulation of protein synthesis and learning/memory. Conditional deletion of Ythdf1 in either D1 or D2 receptor-expressing neurons revealed that normal motor memory is stored in the D1 (direct) pathway of the basal ganglia, while inhibitory memory is stored in the D2 (indirect) pathway. Furthermore, fiber photometry recordings of GCaMP signals from striatal D1 (dSPN) and D2 (iSPN) receptor-expressing neurons support the preservation of normal memory in the direct pathway after aberrant inhibitory learning, with activities of dSPN predictive of motor performance. Finally, a computational model based on activities of motor cortical neurons, dSPN and iSPN neurons, and their interactions through the basal ganglia loops supports the above observations. These findings have important implications for novel approaches in treating Parkinsons disease by reactivating preserved normal memory, and in treating hyperkinetic movement disorders such as chorea or tics by erasing aberrant motor memories.

neuroscience↗

bioRxiv2024

Inhibition of Indirect Pathway Activity Causes Abnormal Decision-Making In a Mouse Model of Impulse Control Disorder in Parkinson's Disease

Healthy action selection relies on the coordinated activity of striatal direct and indirect pathway neurons. In Parkinsons disease (PD), in which loss of midbrain dopamine neurons is associated with progressive motor and cognitive deficits, this coordination is disrupted. Dopamine replacement therapy can remediate motor symptoms, but can also cause impulse control disorder (ICD), which is characterized by pathological gambling, hypersexuality, and/or compulsive shopping. The cellular and circuit mechanisms of ICD remain unknown. Here we developed a mouse model of PD/ICD, in which ICD-like behavior was assayed with a delay discounting task. We found that in parkinsonian mice, the dopamine agonist pramipexole drove more pronounced delay discounting, as well as disrupted firing in both direct and indirect pathway neurons. We found that chemogenetic inhibition of indirect pathway neurons in parkinsonian mice drove similar phenotypes. Together, these findings provide a new mouse model and insights into ICD pathophysiology.

neuroscience↗

bioRxiv2024

Science Education for the Youth (SEFTY): A Neuroscience Outreach Program for High School Students in Southern Nevada During the COVID-19 Pandemic

Laboratory outreach programs for K-12 students in the United States from 2020-2022 were suspended or delayed due to COVID-19 restrictions. While Southern Nevada also observed similar closures for onsite programs, we and others hypothesized that in-person laboratory activities could be prioritized after increasing vaccine doses were available to the public and masking was encouraged. Here, we describe how the Laboratory of Neurogenetics and Precision Medicine at the University of Nevada Las Vegas (UNLV) collaborated with administrators from a local school district to conduct training activities for high school students during the COVID-19 pandemic. The Science Education for the Youth (SEFTY) programs curriculum was constructed to incorporate experiential learning, fostering collaboration and peer-to-peer knowledge exchange. Leveraging neuroscience tools from our UNLV laboratory, we engaged with 117 high school applicants from 2021-2022. Our recruitment efforts yielded a diverse cohort, with >41% Pacific Islander and Asian students, >9% African American students, and >12% multiracial students. We assessed the impact of the SEFTY program through pre- and post-assessment student evaluations, revealing a significant improvement of 20.3% in science proficiency (p<0.001) after participating in the program. Collectively, our laboratory curriculum offers valuable insights into the capacity of an outreach program to actively foster diversity and cultivate opportunities for academic excellence, even in the challenging context of a global pandemic. Significance StatementThe Science Education for the Youth (SEFTY) program at UNLV successfully engaged 117 diverse high school students in neuroscience-based experiential learning, demonstrating the viability of in-person education during a pandemic. Significant improvements in science proficiency (20.3% increase) underscore the programs effectiveness in fostering academic excellence and diversity. This initiative potentially serves as a model for maintaining high-quality, inclusive science education in challenging times.

scientific communication and education↗

bioRxiv2023

Cell-type-specific 3D-genome organization and transcription regulation in the brain

3D organization of the genome plays a critical role in regulating gene expression. However, it remains unclear how chromatin organization differs among different cell types in the brain. Here we used genome-scale DNA and RNA imaging to investigate 3D-genome organization in transcriptionally distinct cell types in the primary motor cortex of the mouse brain. We uncovered a wide spectrum of differences in the nuclear architecture and 3D-genome organization among different cell types, ranging from the physical size of the cell nucleus to the active-inactive chromatin compartmentalization and radial positioning of chromatin loci within the nucleus. These cell-type-dependent variations in nuclear architecture and chromatin organization exhibited strong correlation with both total transcriptional activity of the cell and transcriptional regulation of cell-type-specific marker genes. Moreover, we found that the methylated-DNA-binding protein MeCP2 regulates transcription in a divergent manner, depending on the nuclear radial positions of chromatin loci, through modulating active-inactive chromatin compartmentalization.

biophysics↗

bioRxiv2023

Hypoxia inducible factors inhibit respiratory syncytial virus infection by modulation of nucleolin expression

Respiratory syncytial virus (RSV) is a global healthcare problem, causing respiratory illness in young children and elderly individuals. However, our knowledge of the host pathways that define susceptibility to RSV infection and disease severity is limited. Hypoxia inducible factors (HIFs) define cellular metabolic processes in response to low oxygen and are recognised to play a key role in regulating inflammatory responses in the lower respiratory tract. We demonstrate a role for hypoxia inducible factors (HIFs) to suppress RSV entry and RNA replication. We show that hypoxia and HIF prolyl-hydroxylase inhibitors significantly reduce expression of the RSV entry receptor nucleolin and inhibit viral driven cell-cell fusion. We identify a HIF regulated microRNA, miR-494, that regulates nucleolin expression. In RSV-infected mice, treatment with the clinically approved HIF prolyl-hydroxylase inhibitor, Daprodustat, reduced the level of infectious virus and infiltrating monocytes and neutrophils in the lung. This study highlights a role for HIF-signalling to limit multiple aspects of RSV infection and associated inflammation and informs future therapeutic approaches for treating this respiratory pathogen.

molecular biology↗