Biology subjects

Wang, J.-H.

Publications and source records attributed to Wang, J.-H..

At least 19 recordsLinked to original sources

bioRxiv2026

A modular architecture of human cellular aging across organs and diseases

Aging is a heterogeneous biological process in which different cellular systems undergo molecular remodeling at distinct rates, yet whether human cellular aging follows an organized architecture across organs remains unclear. Here, we integrate a multi-organ human single-cell transcriptomic atlas with plasma proteomic profiles from approximately 50,000 participants to reconstruct cellular aging states at population scale. By projecting cell-type-enriched molecular signatures onto circulating proteins, we characterize aging patterns across 128 organ-cell type pairs and identify 14 cellular aging modules comprising conserved cross-organ programs and organ-specific aging states. These modules reveal cellular identity as a dominant organizing axis of human aging that transcends anatomical boundaries. Module-level aging states uncover substantial inter-individual heterogeneity, with 34% of individuals exhibiting extreme aging deviation in at least one cellular module. Cellular aging modules exhibit distinct temporal trajectories, with structural and tissue-resident modules showing earlier remodeling than immune lineages. The modular organization of cellular aging is reflected in disease susceptibility, with accelerated aging of specific modules, particularly epithelial aging, showing broad associations with disease burden and mortality. Longitudinal analyses further demonstrate the stability and clinical relevance of cellular aging states, whereas lifestyle, metabolic and pharmacological factors show selective relationships with individual aging programs. Together, our study establishes a modular framework for understanding human cellular aging and reveals an organization of biological aging that may help explain individual differences in healthspan.

cell biology↗

bioRxiv2026

Optimization of AAV tools to target M&uumlller glial cells for retinal gene therapy

Reprogramming of Muller glial (MG) cells into retinal neurons has the potential to treat vision loss by regenerating the retina. Development of efficient gene delivery systems to target the MG cells is critical. Adeno-associated virus (AAV) serotypes and promoter specificity are important factors that influence AAV transduction profile in the retina. However, studies that optimize these parameters to specifically target MG cells are limited, in particular in rats which are commonly used for eye research. Here we tested 4 AAV serotypes and 14 promoters to optimize gene delivery to human MG cells in vitro and/or rat MG cells in vivo. We showed that the combinatorial use of MG-specific serotypes and promoters achieved high specificity for MG cell targeting, with ShH10Y serotype and the GFAP (gfaABC1D) promoter as the best performing tool to target rat MG cells in vivo. We developed new AAV vectors using known and novel MG-specific promoters and engineered short promoter variants to improve the cargo capacity of AAV delivery. Our results highlighted a number of promoters that can target MG cells in vitro or in vivo. This study further expands the AAV toolbox to target MG cells, which has important implications for retinal gene therapy development.

bioengineering↗

bioRxiv2025

Optimization of AAV tools to target Muller glial cells for retinal gene therapy

molecular biology↗

bioRxiv2025

Modularity in the DCC extracellular domain elicits distinct effects on axon guidance

Complex neuronal circuits arise from a small set of cell-surface receptors that position neurons, promote axon extension, and define synaptic connections. A central receptor is Deleted in Colon Cancer (DCC), which mediates both short- and long-range axon guidance and confines migrating neurons to the central nervous system. DCCs versatility reflects its ability to interact at distinct sites of its extracellular domain with two ligands, Netrin-1 and Draxin, which also bind to each other. Alternative splicing further alters the Netrin-1 binding site and modulates affinity. By generating two mouse lines with mutations that selectively impair DCC binding to Netrin-1 and/or Draxin, we show that molecular modularity within the DCC extracellular domain is essential for precise circuit assembly. An eight-amino acid insertion in the DCClong isoform is required for Netrin-1-dependent long-range commissural axon guidance in the spinal cord. Conversely, isoleucine 372 in the Draxin binding site enables DCC clustering and is necessary for all known DCC functions, including axon guidance in the spinal cord and retina and neuronal migration in the brainstem. Draxin also supports long-range commissural guidance, but mutations in its binding site cause stronger defects. These results underscore how DCCs distinct modules drive specific developmental responses.

neuroscience↗

bioRxiv2025

KiF1a-regulated neuronal infrastructures in sensory and prefrontal cortices essential for fear memory and anxiety

Stress in social activities induces fears. Cellular infrastructures and molecular profiles underlying fear-relevant psychosis remain elusive. By giving psychological stress to mice that watched fear scenes in the resident/intruder paradigm, we have studied the features of neuronal infrastructures in visual, auditory and medial prefrontal cortices correlated to fear-induced anxiety by approaches of behavior task, neural tracing, molecular biology and electrophysiology in vivo. This psychological stress causes observational mice fears specific to resident mouse and anxiety. This social phobia is associated with the interconnections newly emerged among visual, auditory and medial prefrontal cortices as well as among intramodal neurons. These cortical neurons receive the new synapse innervations from their interconnected neurons alongside the innate synapse innervations, and become to encode the stress signals including battle image and battle sound. The KiF1a knockdown in the medial prefrontal cortex precludes the formations of neuronal infrastructures and social phobia. KiF1a-mediated intracellular transportation in the medial prefrontal cortex is essential for the recruitment of associative memory neurons that encode fear scenes and anxiety. HighlightO_LIPsychological stress induces fears and interconnections among sensory and prefrontal cortices. C_LIO_LIAssociative memory neurons in these cortices are recruited to encode fear scenes and anxiety. C_LIO_LIKiF1a-mediated transportation essential for these cellular infrastructures relevant to learned fear. C_LI

neuroscience↗

bioRxiv2024

Do-it-yourself de novo antibody sequencing workflow that achieves complete accuracy of the variable regions

Antibodies are widely used as research tools or therapeutic agents. Knowing the sequences of the variable regions of an antibody--both the heavy chain and the light chain--is a prerequisite for the production of recombinant antibodies. Mass spectrometry-based de novo sequencing is a frequently used, and sometimes the only approach to gaining this information. Here, we describe a workflow that enables accurate sequence determination of monoclonal antibodies based on mass spectrometry data and freely available software tools. This workflow, which we developed using a homemade anti-FLAG monoclonal antibody as a reference sample, achieved 100% accuracy of the variable regions with clear distinction between leucine (L) and isoleucine (I). Using this workflow, we successfully decoded a monoclonal anti-HA antibody, for which we had no prior knowledge of its sequence. Based on the de novo sequencing result, we generated a recombinant anti-HA antibody, and demonstrated that it has the same specificity, sensitivity, and affinity as the commercial antibody. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=142 SRC="FIGDIR/small/630060v1_ufig1.gif" ALT="Figure 1"> View larger version (28K): org.highwire.dtl.DTLVardef@180c502org.highwire.dtl.DTLVardef@1064399org.highwire.dtl.DTLVardef@4005d3org.highwire.dtl.DTLVardef@13e65ed_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

bioRxiv2024

Characterisation of RNA editing and gene therapy with a compact CRISPR-Cas13 in the retina

CRISPR-Cas13 nucleases are programmable RNA-targeting effectors that can silence gene expression in a reversible manner. Recent iterations of Cas13 nucleases are compact for adeno-associated virus (AAV) delivery to achieve strong and persistent expression in various organs in a safe manner. Here, we report significant transcriptomic signatures of Cas13bt3 expression in retinal cells and show all-in-one AAV gene therapy with Cas13bt3 can effectively silence VEGFA mRNA in human retinal organoids and humanised VEGF transgenic mouse (trVEGF029, Kimba) models. Specifically, human embryonic stem cells (hESC)-derived retinal pigment epithelium cells show high expression of Cas13bt3 from virus delivery corresponding to a significant reduction of VEGFA mRNA. We further show that intravitreal delivery of Cas13bt3 can transduce mouse retinal cells efficiently, reaching the photoreceptors for specific knockdown of human VEGFA in the Kimba mouse. Our results reveal important considerations for assessing Cas13 activity and establish Cas13bt3 as a potential anti-VEGF agent that can achieve long-term control of VEGFA for the treatment of retinal neovascularization.

molecular biology↗

bioRxiv2023

Associative memory neurons of encoding multi-modal signals are recruited by neuroligin-3 mediated new synapse formation

The joint storage and reciprocal retrieval of leant associated signals are presumably encoded by associative memory cells. In the accumulation and enrichment of memory contents in lifespan, a signal often becomes a core signal associatively shared for other signals. One specific group of associative memory neurons that encode this core signal likely interconnects multiple groups of associative memory neurons that encode these other signals for their joint storage and reciprocal retrieval. We have examined this hypothesis in a mouse model of associative learning by pairing the whisker tactile signal sequentially with the olfactory signal, the gustatory signal and the tail-heating signal. Mice experienced this associative learning show the whisker fluctuation induced by olfactory, gustatory and tail-heating signals, or the other way around, i.e., memories to multi-modal associated signals featured by their reciprocal retrievals. Barrel cortical neurons in these mice become able to encode olfactory, gustatory and tail-heating signals alongside the whisker signal. Barrel cortical neurons interconnect piriform, S1-Tr and gustatory cortical neurons. With the barrel cortex as the hub, the indirect activation occurs among piriform, gustatory and S1-Tr cortices for the second-order associative memory. These associative memory neurons recruited to encode multi-modal signals in the barrel cortex for associative memory are downregulated by neuroligin-3 knockdown. Thus, associative memory neurons can be recruited as the core cellular substrate to memorize multiple associated signals for the first-order and the second-order of associative memories by neuroligin-3-mediated synapse formation, which constitutes neuronal substrates of cognitive activities in the field of memoriology. HighlightO_LIThe coactivity of cerebral cortices during associative learning induces their interconnections. C_LIO_LIInterconnections endorse the first order and the second order of associative memory. C_LIO_LIAssociative memory cells in cerebral cortices are recruited by mutual synapse innervations. C_LIO_LINeuroligin-3 mediates the recruitment of associative memory cells for associative memory. C_LI

neuroscience↗

bioRxiv2022

Knockout of AMD-associated gene POLDIP2 reduces mitochondrial superoxide in human retinal pigment epithelial cells

Genetic and epidemiologic studies have significantly advanced our understanding of the genetic factors contributing to age-related macular degeneration (AMD). In particular, recent expression quantitative trait loci (eQTL) studies have highlighted POLDIP2 as a significant gene that confers risk of developing AMD. However, the role of POLDIP2 in retinal cells such as retinal pigment epithelium (RPE) and how it contributes to AMD pathology are unknown. Here we report the generation of a stable human RPE cell line with POLDIP2 knockout using CRISPR/Cas, providing an in vitro model to investigate the functions of POLDIP2. We conducted functional studies on the POLDIP2 knockout cell line and showed that they retained normal levels of cell proliferation, cell viability, phagocytosis and autophagy. Also, we performed RNA sequencing to profile the transcriptome of POLDIP2 knockout cells. Our results highlighted significant changes in genes involved in immune response, complement activation, oxidative damage and vascular development. We showed that loss of POLDIP2 causes a reduction in mitochondrial superoxide levels, which is consistent with the upregulation of the mitochondrial superoxide dismutase SOD2. In conclusion, this study demonstrates a novel link between POLDIP2 and SOD2, which supports a potential role of POLDIP2 in regulating oxidative stress in AMD pathology.

neuroscience↗

bioRxiv2022

Preferential cross-linking of the stereospecific complex over the encounter complexes by DOPA2, a faster cross-linker than DSS

Transient protein-protein interactions are fundamental aspects of many biochemical reactions, but they are technically challenging to study. Chemical cross-linking of proteins coupled with mass spectrometry (CXMS) analysis is a powerful tool to facilitate the analysis of transient interactions. Central to this technology are chemical cross-linkers. Here, using two transient heterodimeric complexes--EIN/HPr with a KD of 7 M and EIIAGlc/EIIBGlc with a KD of 25 M--as model systems, we compared the effects of two amine-specific homo-bifunctional cross-linkers of different cross-linking speeds. Protein cross-linking by DOPA2, a di-ortho-phthalaldehyde cross-linker, is 60-120 times faster than that by DSS, an N-hydroxysuccinimide ester cross-linker. We analyzed the differences in the number of cross-links identified that reflected the stereospecific complex (SC), the final lowest-energy conformational state, and that of cross-links that reflected the encounter complexes (ECs), an ensemble of short-lived intermediate conformations mediated by nonspecific electrostatic interactions. We found that the faster DOPA2 cross-linking favored the SC whereas the slower DSS cross-linking favored the ECs. We propose a mechanistic model for this intriguing observation. This study suggests that it is feasible to probe the dynamics of protein-protein interaction using cross-linkers of different cross-linking speeds.

biochemistry↗

bioRxiv2022

The majority of microorganisms in gas hydrate bearing subseafloor sediments ferment macromolecules

BackgroundGas hydrate-bearing subseafloor sediments harbor a large number of microorganisms. Sedimentary organic matter and upward methane fluids represent two important sources of carbon and energy for deep biosphere. However, which metabolism dominates the deep subseafloor of gas hydrate zone is poorly constrained. Here we studied the microbial communities in gas-hydrate rich sediments up to 49 meters below seafloor recovered by drilling in the South China Sea. We focused on distinct geochemical conditions, and performed metagenomic and metatranscriptomic analyses to characterize microbial diversity and their role in carbon mineralization. ResultsComparative microbial community analysis revealed that samples above and in sulfate-methane interface (SMI) zones clearly distinguish from those below the SMI. Chloroflexota, are most abundant above the SMI, whereas Caldatribacteriota dominate below the SMI. Verrucomicrobiota, Bathyarchaeia and Hadarchaeota were similarly present in both types of sediment. The genomic inventory and transcriptional activity suggest roles in fermentation of macromolecule. By contrast, sulfate reducers and methanogens, organisms that catalyze the consumption or production of commonly observed chemical compounds in sediments are rare. Methanotrophs of the ANME-1 group thrived in or slightly below the current sulfate methane interface. Rare members from Heimdallarchaeia were identified to encode the potential for anaerobic oxidation of short-chain hydrocarbons. ConclusionsThese findings indicate that fermentation of macromolecules is the predominant energy source for microorganisms in deep subseafloor sediments that are experiencing upward methane fluxes.

microbiology↗

bioRxiv2022

Subtyping of major SARS-CoV-2 variants reveals different transmission dynamics

SARS-CoV-2 continues to evolve, causing waves of the pandemic. Up to March 2022, eight million genome sequences have accumulated, which are classified into five major variants of concern. With the growing number of sequenced genomes, analysis of the big dataset has become increasingly challenging. Here we developed systematic approaches for comprehensive subtyping and pattern recognition for transmission dynamics. By analyzing the first two million viral genomes as of July 2021, we found that different subtypes of the same variant exhibited distinct temporal trajectories. For example, some Delta subtypes did not spread rapidly, while others did. We identified sets of characteristic single nucleotide variations (SNVs) that appeared to enhance transmission or decrease efficacy of antibodies for some subtypes of the Delta and Alpha variants. We also identified a set of SNVs that appeared to suppress transmission or increase viral sensitivity to antibodies. These findings are later confirmed in an analysis of six million genomes as of December 2021. For the Omicron variant, the dominant type in the world, we identified the subtypes with enhanced and suppressed transmission in an analysis of seven million genomes as of January 2022 and further confirmed the findings in a later analysis of eight million genomes as of March 2022. While the "enhancer" SNVs exhibited an enriched presence on the spike protein, the "suppressor" SNVs are mainly elsewhere. Disruption of the SNV correlation largely destroyed the enhancer-suppressor phenomena. These results suggest the importance of fine subtyping of variants, and point to potential complex interactions among SNVs.

bioinformatics↗

bioRxiv2022

The endoplasmic reticulum membrane protein complex (EMC) negatively regulates intestinal homeostasis through the Hippo signaling pathway

Disruption of tissue homeostasis often results in many diseases. Balanced control of stem cell proliferation and differentiation underlines tissue homeostasis. However, how endogenous factors influence the proliferation and differentiation of intestinal stem cells (ISCs) under physiological conditions remains not fully understood. Here, we find that the endoplasmic reticulum membrane protein complex (EMC) negatively regulates ISC proliferation in adult Drosophila midgut. Compromising EMC function in progenitors leads to excessive ISC proliferation and intestinal homeostasis disruption. Mechanistically, the EMC complex associates with and stabilizes Hippo (Hpo), the key component of the Hpo signaling pathway. In the absence of the EMC complex, Yki (Yorkie) is activated to promote ISC proliferation. Furthermore, the role of the EMC complex in stem cell proliferation control is evolutionarily conserved. Thus, our study uncovers the molecular mechanism of the EMC complex in controlling stem cell proliferation. Our results provide new insight into the underlying mechanisms of how stem cell proliferation is properly controlled under physiological conditions.

cell biology↗

bioRxiv2021

SARS-CoV-2-Triggered Mast Cell Rapid Degranulation Induces Alveolar Epithelial Inflammation and Lung Injury

SARS-CoV-2 infection-induced hyper-inflammation links to the acute lung injury and COVID-19 severity. Identifying the primary mediators that initiate the uncontrolled hypercytokinemia is essential for treatments. Mast cells (MCs) are strategically located at the mucosa and beneficially or detrimentally regulate immune inflammations. Here we showed that SARS-CoV-2-triggeed MC degranulation initiated alveolar epithelial inflammation and lung injury. SARS-CoV-2 challenge induced MC degranulation in ACE-2 humanized mice and rhesus macaques, and a rapid MC degranulation could be recapitulated with Spike-RBD binding to ACE2 in cells; MC degranulation alterred various signaling pathways in alveolar epithelial cells, particularly, led to the production of pro-inflammatory factors and consequential disruption of tight junctions. Importantly, the administration of clinical MC stabilizers for blocking degranulation dampened SARS-CoV-2-induced production of pro-inflammatory factors and prevented lung injury. These findings uncover a novel mechanism for SARS-CoV-2 initiating lung inflammation, and suggest an off-label use of MC stabilizer as immunomodulators for COVID-19 treatments. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=135 SRC="FIGDIR/small/449680v1_ufig1.gif" ALT="Figure 1"> View larger version (29K): org.highwire.dtl.DTLVardef@899996org.highwire.dtl.DTLVardef@1c26c0eorg.highwire.dtl.DTLVardef@1442cdcorg.highwire.dtl.DTLVardef@dd4204_HPS_FORMAT_FIGEXP M_FIG C_FIG In BriefSARS-CoV-2 triggers an immediate mast cell (MC) degranulation, which initiates the alveolar epithelial inflammation and disrupts the tight junction. MC stabilizers that block degranulation reduce virus-induced lung inflammation and injury. HighlightsO_LIThe binding of RBD of Spike protein of SARS-CoV-2-to ACE2 receptor protein triggers an immediate MC degranulation C_LIO_LIMC degranulation induces transcriptomic changes include an upregulated inflammatory signaling and a downregulated cell-junction signaling C_LIO_LIMC degranulation leads to alveolar epithelial inflammation and disruption of tight junctions C_LIO_LIMC stabilizer that inhibits degranulation reduces SARS-CoV-2-induced lung inflammation and injury in vivo C_LI

microbiology↗

bioRxiv2021

Single-cell RNA sequencing uncovers the excitatory/inhibitory synaptic unbalance in the retrosplenial cortex after peripheral nerve injury

Nerve injury in the somatosensory pathway may induce maladaptive changes at the transcriptional or protein level, contributing to the development and maintenance of neuropathic pain. In contrast to the retrosplenial cortex (RSC), which processes nociceptive information and exhibits structural and molecular changes after nerve injury, detailed transcriptional changes in the RSC are not yet known. Here we confirm the involvement of the RSC in regulating pain sensation and observe that the same peripheral stimulation activates more retrosplenial neurons after nerve injury; reducing the activities of CaMKII+ splenial cells relieves peripheral pain hypersensitivity after nerve injury. Using a single-cell RNA sequencing (scRNA-seq) approach, we identified cell-type-specific gene expression changes after nerve injury, and the gene set enrichment analysis results revealed suppressed ion homeostasis in CaMKII+ neurons. Furthermore, examination of the expression of genes encoding ligand-gated ion channels showed a decrease in Gabar1a but an increase in Gria1 in CaMKII+ neurons; consistently, we confirmed the unbalanced excitatory/inhibitory synaptic transmission by using the electrophysiological recording approach. Moreover, micro-infusion of 1-Naphthyl acetyl spermine in the RSC to reduce excitatory synaptic transmission alleviated peripheral pain hypersensitivity. Our data confirm the involvement of the RSC in pain regulation and provide information on cell type-dependent transcriptomic changes after nerve injury, which will contribute to the understanding of the mechanisms mediating neuropathic pain.

neuroscience↗

bioRxiv2021

TAK1: a potential therapeutic target for retinal neovascularization

Retinal neovascularization, or pathological angiogenesis in the retina, is a leading cause of blindness in developed countries. Transforming growth factor-{beta}-activated kinase 1 (TAK1) is a mitogen-activated protein kinase kinase kinase (MAPKKK) activated by TGF-{beta}1 and other pro-inflammatory cytokines. TAK1 is also a key mediator of inflammation, innate immune responses, apoptosis and tissue homeostasis and plays an important role in physiological angiogenesis. Its role in pathological angiogenesis, particularly in retinal neovascularization, remains unclear. We investigated the regulatory role of TAK1 in pathological angiogenesis in the retina. Transcriptome analysis of human retina featuring retinal neovascularization revealed enrichment of known TAK1-mediated signaling pathways. Selective inhibition of TAK1 activation by 5Z-7-oxozeaenol attenuated aberrant retinal angiogenesis in rats following oxygen-induced retinopathy. Transcriptome profiling revealed that TAK1 activation in human microvascular endothelial cells under TNF stimulation led to increase the gene expression related to cytokines and leukocyte-endothelial interaction, mainly through nuclear factor kappa B (NF{kappa}B) signaling pathways. These results reveal that inhibition of TAK1 signaling may have therapeutic value for the treatment of pathological angiogenesis in the retina.

biochemistry↗

bioRxiv2020

A Fast Lysine Cross-linker DOPA Enables Mass Spectrometry Analyses of Protein Unfolding and Weak Protein-protein Interactions

Chemical cross-linking of proteins coupled with mass spectrometry analysis (CXMS) has become a widely used method for protein structure analysis. Central to this technology are chemical cross-linkers. The most popular cross-linkers are N-hydroxysuccinimide (NHS) esters, which react with protein amino groups relatively slowly over 10 minutes or more while in competition with the hydrolysis reaction of NHS esters. To improve the speed of cross-linking, we developed a new class of amine-selective and non-hydrolyzable di-ortho-phthalaldehyde (DOPA) cross-linkers. DOPA can cross-link proteins in 10 seconds under near physiological conditions, which is 60 times faster than the NHS ester cross-linker DSS. DOPA also works at low pH, low temperature, or in the presence of high concentrations of denaturants such as 8 M urea or 6 M guanidine hydrochloride. Further, DOPA-mediated pulse cross-linking captured the dynamic conformational changes associated with RNase A unfolding. Lastly, DOPA outperformed DSS at capturing weak but specific protein-protein interactions.

biochemistry↗

bioRxiv2020

Deep sea sediments associated with cold seeps are a subsurface reservoir of viral diversity

In marine ecosystems, viruses exert control on the composition and metabolism of microbial communities, thus influencing overall biogeochemical cycling. Deep sea sediments associated with cold seeps are known to host taxonomically diverse microbial communities, but little is known about viruses infecting these microorganisms. Here, we probed metagenomes from seven geographically diverse cold seeps across global oceans, to assess viral diversity, virus-host interaction, and virus-encoded auxiliary metabolic genes (AMGs). Gene-sharing network comparisons with viruses inhabiting other ecosystems reveal that cold seep sediments harbour considerable unexplored viral diversity. Most cold seep viruses display high degrees of endemism with seep fluid flux being one of the main drivers of viral community composition. In silico predictions linked 14.2% of the viruses to microbial host populations, with many belonging to poorly understood candidate bacterial and archaeal phyla. Lysis was predicted to be a predominant viral lifestyle based on lineage-specific virus/host abundance ratios. Metabolic predictions of prokaryotic host genomes and viral AMGs suggest that viruses influence microbial hydrocarbon biodegradation at cold seeps, as well as other carbon, sulfur and nitrogen cycling via virus-induced mortality and/or metabolic augmentation. Overall, these findings reveal the global diversity and biogeography of cold seep viruses and indicate how viruses may manipulate seep microbial ecology and biogeochemistry.

microbiology↗