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Zheng, C.

Publications and source records attributed to Zheng, C..

At least 55 records · Page 3Linked to original sources

Dual role of GABAB receptor in oligodendrocyte function and immune modulation in experimental multiple sclerosis

GABAB receptors (GABABR) mediate the actions of the inhibitory neurotransmitter GABA in the central nervous system, regulating key processes such as synaptic activity, interneuron communication and excitation-inhibition balance in the brain. Recent studies using the GABABR agonist baclofen have revealed a critical role for these receptors in promoting oligodendroglial differentiation in both health and disease, highlighting their potential as therapeutic targets in demyelinating diseases such as multiple sclerosis (MS). In this study, we identify a dual role for oligodendroglial GABABR in experimental autoimmune encephalomyelitis (EAE), an animal model of MS. Conditional deletion of the GABAB1 subunit in NG2+ cells ameliorates acute disease symptoms while inducing an immune-like phenotype in oligodendrocytes. This immunomodulatory role is further supported by pharmacological activation of GABABR in oligodendrocytes, which reduces the expression of MHC class II in these cells. Notably, baclofen treatment after EAE symptom onset attenuates the course of the disease while enhancing oligodendrocyte progenitor cell differentiation and suppressing T cell infiltration into demyelinating lesions. Moreover, prophylactic baclofen administration delays disease onset and further decreases immune cell recruitment into the spinal cord, underscoring its potent immunomodulatory effect. These data demonstrate that GABABR signaling exerts context-dependent effects on both oligodendrocyte lineage progression and neuroinflammatory responses. Importantly, these compelling findings validate baclofen, a drug already approved for MS-associated spasticity, as a promising candidate for therapies targeting both inflammation and remyelination, advancing our understanding of glial-immune interactions in demyelinating diseases and supporting the translational potential of GABABR modulation in MS.

neuroscience↗

A benchmark of the somatic mutation landscape using single-cell and single-molecule whole-genome sequencing

Accurate detection of somatic mutations in noncancerous cells is critical for studying somatic mosaicism, a process implicated in aging and multiple chronic diseases. However, single-cell and single-molecule DNA sequencing platforms differ in their error profiles, coverage biases, and sensitivity to specific mutation types, complicating cross-platform comparisons. Here, we present in vitro and in silico benchmarks to quantify true-positive and false-positive rates in single-cell whole-genome sequencing using Single-Cell Multiple Displacement Amplification, and in single-molecule sequencing using Nanorate Sequencing (NS) and whole-genome NS (WGNS). Using standard cell lines, we show that all three methods detect single-nucleotide variants (sSNVs) and small insertions and deletions (sINDELs) with high accuracy, but differ in genomic coverage and susceptibility to artifacts. Method-specific biases influence mutational signatures and hotspot detection. Applying results of the benchmark to IMR-90 fibroblasts, we estimate higher in vitro mutation rates using NS than expected from in vivo data, consistent with potential replication stress and culture-associated DNA damage. Overall, our study highlights the substantial impact of sequencing platform-specific biases on somatic mutation detection and interpretation, and lays the foundation for standardized, cross-platform-comparable analyses of somatic mosaicism in normal human tissues.

genomics↗

Environmental Heterogeneity Altered the Growth Fitness of Antibiotic-Resistant Mutants and the Resistance Prevalence in Escherichia coli populations

Antibiotic resistance is one of the most critical issues in public health. As antibiotic-resistant bacteria emerge under certain selection pressure, their further proliferation can facilitate the prevalence and maintenance of the antibiotic resistance. Here, we investigated which environmental factors could affect the growth fitness of antibiotic-resistant Escherichia coli strains using growth competition assays. We found that when antibiotic resistance selection pressure was removed, lower temperature and nutrient limitations, especially iron (Fe) deficiency, fostered a better fitness to resistant mutants when co-cultivated with the wild type, whereas changes in pH or salinity (Na or K) did not. Moreover, the growth advantage of resistant mutants under the selection pressure was reversed by suboptimal conditions like acidic/basic pH, high K salinity, and Fe limitation. These identified key environmental factors influencing the growth fitness of antibiotic-resistant mutants provide important insights into the prediction and mitigation of antibiotic resistance in heterogeneous environments.

microbiology↗

Two Spag6 genes control sperm formation and male fertility in mice

Sperm-associated antigen 6 (SPAG6) is the mammalian orthologue of Chlamydomonas PF16, a central axonemal protein essential for flagellar motility. In mice, two homologous genes have been identified: the ancestral Spag6 on chromosome 2 and the evolutionary derived Spag6l on chromosome 16. Although Spag6 knockout mice (Spag6-/-) are phenotypically normal, the surviving Spag6l-/- males are infertile. To further investigate the roles of SPAG6 and SPAG6L, we generated compound mutants by crossing the two knockout lines. Compound heterozygous Spag6+/-; Spag6l+/- mice are fertile, while all Spag6-/-; Spag6l+/- males are infertile despite grossly normal appearance. Histological and ultrastructural analyses revealed defective spermiogenesis, including abnormal chromatin condensation, malformed acrosome and manchette, and disorganized mitochondrial and fibrous sheath. Both SPAG6 and SPAG6L bind to SPINK2, a key regulator of acrosome function, but SPAG6 has an approximately 10-fold higher binding affinity than SPAG6L. Moreover, SPAG6 modulates testicular AKAP4 and SPAG16L levels, which are critical components of the fibrous sheath and central apparatus respectively. Notably, SPAG6 suppresses tubulin acetylation, whereas SPAG6L enhances this post-translational modification, suggesting antagonistic roles in microtubule assembly. Overall, our findings demonstrate that SPAG6 and SPAG6L coordinately regulate sperm formation and male fertility during evolution.

developmental biology↗

Cytoplasmic tail composition modulates the G protein and arrestin-3 signaling bias of the adhesion GPCR LPHN2

The class B2 Adhesion GPCRs (aGPCRs) combine cell adhesion with GPCR signaling to control diverse developmental and physiological processes. How aGPCRs interact with and integrate distinct groups of effectors including G proteins, arrestins, and G protein receptor kinases (GRKs) remains unclear. Here, we find that diversity in the aGPCR C-terminal intracellular tail modulates G protein activation, arrestin-3 recruitment, and utilization of selective GRKs in LPHN2, a postsynaptic aGPCR essential for synapse formation. The C-terminal tail of LPHN2 is required for G protein activation and arrestin-3 recruitment. LPHN2 with an intact tail recruits arrestin-3 in the absence of G protein activation, suggesting constitutive arrestin-3-biased signaling. Alternative splicing of the LPHN2 tail modulates G protein activation and arrestin-3 binding independently, supporting that it controls G protein vs arrestin-3 bias. GRKs are important but not essential for arrestin-3 recruitment to LPHN2. Moreover, GRK2 increases arrestin-3 recruitment only in a subset of LPHN2 variants. Collectively, these results show that the mechanisms of the interactions of class B2 aGPCRs and arrestin are distinct from those of class A GPCRs and that splicing of the LPHN2 C-terminal tail determines G protein vs arrestin bias.

molecular biology↗

RWKV-IF: Efficient and Controllable RNA Inverse Folding via Attention-Free Language Modeling

We present RWKV-IF, an efficient and controllable framework for RNA inverse folding based on the attention-free RWKV language model. By treating structure-to-sequence generation as a conditional language modeling task, RWKV-IF captures long-range dependencies with linear complexity. We introduce a decoding strategy that integrates Top-k sampling, temperature control, and G-C content biasing to generate sequences that are both structurally accurate and biophysically meaningful. To overcome limitations of existing datasets, we construct a large-scale synthetic training set from randomly generated sequences and demonstrate strong generalization to real-world RNA structures. Experimental results show that RWKV-IF significantly outperforms traditional search-based baselines, achieving higher accuracy and full match rate while greatly reducing edit distance. Our approach highlights the potential of lightweight generative models in RNA design under structural and biochemical constraints. The code is available at https://github.com/Lyttr/RWKVInverseFolding.git

bioengineering↗

HSV-1 orchestrates host RAP80 ubiquitination by ICP0 and UL36USP to promote viral survival

Herpes simplex virus 1 (HSV-1) is a widespread human pathogen that establishes lifelong infections, ubiquitination is crucial for HSV - 1 to interact with the host and regulate its life activities. But the mechanisms of how HSV-1 dynamically modulates host proteins at different infection stages to promote its own survival remains challenging. Here we identified DNA damage response protein RAP80 as a critical target of HSV-1 ubiquitination enzymes ICP0 and UL36USP throughout HSV-1s infection cycle. In early stages, RAP80 inhibits HSV-1 transcription by directly binding to the viral genome, blocking viral protein ICP4 binding to transcription factors. This interaction is regulated by phase separation via an intricate interplay between RAP80 and ICP0/ICP4. As infection progresses, the viral E3 ligase ICP0 degrades RAP80, dissolving phase separation, and allowing the formation of a mature viral replication compartment. In late infection stages, RAP80 is deubiquitinated by the viral deubiquitinase UL36USP, which restores cellular homeostasis and promotes HSV-1 survival. This process involves modulating the R-loop-cGAS-apoptosis pathway. These findings underscore the dynamic interplay between viral and host factors and the complex mechanisms used by HSV-1 to subvert host defenses, offering practical implications for the future development of antiviral strategies.

molecular biology↗

Spatial metabolic gradients in the liver and small intestine

Mammalian organs are composed of cells, whose properties differ depending upon their spatial location. Gene expression in liver varies between periportal and pericentral hepatocytes1-3, and in intestine from crypts to villus tips4,5. A key element of tissue spatial organization is likely metabolic, but direct assessments of spatial metabolism remain limited. Here we map spatial metabolic gradients in murine liver and intestine. We developed an integrated experimental-computational workflow using MALDI imaging mass spectrometry, isotope tracing, and deep-learning artificial intelligence. Most measured metabolites (> 95%) showed significant spatial concentration gradients in liver lobules and intestinal villi. In the liver, tricarboxylic acid (TCA)-cycle metabolites and their labeling from both glutamine and lactate localized periportally. Energy-stress metabolites including adenosine monophosphate (AMP) also localized periportally, consistent with high periportal energy demand. In intestine, the TCA intermediates malate (tip) and citrate (crypt) showed opposite spatial patterns, which aligned with higher glutamine catabolism in tips and lactate oxidation in crypts based on isotope tracing. Finally, we mapped the fate of the obesogenic dietary sugar fructose. In the intestine, oral fructose was catabolized faster in the villus bottom than the tips. In the liver, fructose-derived carbon accumulated pericentrally as fructose-1-phosphate and triggered pericentral adenosine triphosphate (ATP) depletion. Thus, we both provide foundational knowledge regarding intestine and liver metabolic organization and identify fructose-induced focal derangements in liver metabolism.

biochemistry↗

stTrace: Detecting Spatial-Temporal Domains from spatial transcriptome to Trace Developmental Path

Development is essential for the growth and functional maintenance of organisms. Investigating the development process is vital for uncovering the formation of complex biological systems. However, current approaches to studying development from gene expression rely primarily on single-cell gene expression data to infer developmental trajectories, neglecting the spatial distribution of cells within tissues and their interactions. Although spatial transcriptomics provides spatial context for gene expression, existing algorithms focus mainly on identifying spatial regions without further exploring their developmental connections. In this study, we propose an algorithm for detecting spatial-temporal domains in tissue to trace developmental path (stTrace) using spatial transcriptomics. stTrace integrates the degree of cell development, gene expression, and spatial location to identify "spatial-temporal domains", regions where cells share similar functions and developmental stages within the tissue. Moreover, hierarchical relationships exist among these regions, reflecting developmental connections between cells in the tissue. Our experiments on mouse embryo and human breast cancer data revealed that stTrace can detect more refined regions than traditional spatial domain identification algorithms. Furthermore, the directions of developmental paths inferred from hierarchical relationships are consistent with the dynamic trajectories derived from single-cell velocity. Key pointsO_LIWe proposed spatial-temporal domain, which refers to spatially continuous regions where cells exhibit similar gene expression patterns, related functions, and close developmental levels. C_LIO_LIWe present stTrace, a computational framework for identifying spatial-temporal domains by iteratively optimizing two objectives: minimizing Structure Entropy to capture functional similarity and hierarchy, and maximizing the Silhouette Coefficient to ensure temporal compactness. C_LIO_LIIn both mouse embryo and human breast cancer datasets, spatial-temporal domains align better with H&E-stained images and and have better compactness in developmental level. C_LIO_LIstTrace detected developmental paths that are well-mapped with cells temporal information like single-cell velocity and trajectories and application show the usage in studying embryo development and cancer invasion. C_LI

bioinformatics↗

RABEP1 amplifies front signaling in neutrophil migration

Neutrophils are the first responders of our innate immune system, crucial for defense against various infections. The intricate regulation of neutrophil migration is essential for neutrophil function. However, a complete mechanistic understanding is missing. We performed a miRNA overexpression screen and identified miR-190 as a potent suppressor of neutrophil migration in zebrafish. Through a second round of small-scale screening using neutrophil-specific knockouts of putative miR-190 targets, we identified that rabep1 (Rabaptin-5, RAB GTPase binding effector protein 1) is essential for neutrophil motility and chemotaxis in zebrafish. Re-expressing full-length RABEP1 in the knockout, but not a truncated form lacking the Rab4/Rab5 binding domain, rescued cell motility. Knocking down RABEP1 in human dHL-60 cells consistently reduced cell motility. RABEP1-deficient dHL-60 cells accumulate excessive members inside the cells. The Rab5 GTP level is unaffected, but the RABEP1 knockdown cells displayed reduced PAK phosphorylation and overall reduced actin polymerization, but still appropriately polarized upon chemokine stimulation. Overexpression of dominant-negative Rab4 or Rab5 similarly inhibits neutrophil migration. Our data suggests that RABEP1 drives endosomal recycling, Rac activation, and leading-edge actin polymerization, providing significant insights into the role of the endocytic pathway in neutrophil motility.

cell biology↗

VGLL1 contributes to both the transcriptome and epigenome of the developing trophoblast compartment

The trophectoderm (TE), the first lineage specified during mammalian development, initiates implantation and gives rise to placental trophoblasts. While animal models have elucidated key conserved signaling pathways involved in early TE specification, including BMP, WNT, and HIPPO, species-specific differences during early development emphasize the need for human-specific models. We previously identified VGLL1, a coactivator of TEAD transcription factors, as a human-specific placental marker. In this study, we employed a pluripotent stem cell (PSC)-based model of TE induction by BMP4 to investigate chromatin remodeling and transcriptional dynamics during TE formation. BMP4-induced chromatin accessibility changes promoted a trophoblast gene expression program, while mesoderm lineage markers were only transiently expressed upon canonical WNT activation. We found that VGLL1 was expressed downstream of key TE transcription factors (GATA2/3, TFAP2A/C) but was essential for establishment of full trophoblast identity by up-regulating EGFR and reinforcing GATA3 expression through positive feedback. Notably, VGLL1 enhanced canonical WNT signaling via direct regulation of WNT receptors and effectors. We also identified KDM6B, a histone demethylase that removes H3K27me3 repressive marks, as a direct VGLL1 target. KDM6B facilitated activation of bivalent promoters associated with TE markers, linking epigenetic regulation to lineage identity. Our findings establish a mechanistic framework positioning VGLL1 as a central regulator that integrates HIPPO, BMP, and WNT signaling pathways to drive establishment of human TE. Statement of SignificanceEarly development of the human placenta is essential for pregnancy success, yet the mechanisms that guide placental lineage specification remain poorly defined. Using human stem cells, we show how signaling pathways and chromatin remodeling programs work together to direct formation of the trophectoderm, the earliest placental cell type. We identify VGLL1 as a key regulator linking multiple signaling networks to gene expression and epigenetic control. Our findings reveal a species-specific mechanism of placental initiation with broad implications for understanding reproductive disorders, pregnancy loss, and advancing stem cell-based models to study and potentially treat human placental disease.

developmental biology↗

Maternal Effects on Postembryonic Neuroblast Migration in C. elegans

Maternal effect genes mostly regulate early embryogenesis as their mRNAs or proteins are deposited into the oocytes to function during early embryonic development before the onset of zygotic transcription. Here, we report a case where a maternal effect gene regulates postembryonic neuroblast migration long after the early embryonic stages. We found that the defects of the Q neuroblast migration in C. elegans mannosyltransferase dpy-19 mutants can be rescued by a maternal copy of the gene. Maternal dpy-19 mRNAs are deposited into the oocytes and persist throughout embryonic development into the Q cells to regulate their migration in early larval stages. These mRNAs appeared to be remarkably stable, since long-term developmental arrest, changing the 3UTR sequence, and mutations in genes involved in RNA binding and modification all had weak effects on the maternal rescue of the neuroblast migration defects. Since the defects can also be rescued by a zygotic copy of dpy-19(+), our results suggest that postembryonic neurodevelopment is redundantly regulated by maternal and zygotic copies of the same gene.

developmental biology↗

Intranasal AAV Vaccination of SARS-CoV-2 Induce Strong and Sustained Neutralizing Antibodies in Mice

The COVID-19 pandemic continues to pose significant health challenges, despite existing vaccines. This study evaluates the immunogenicity of recombinant adeno-associated viruses (AAV) expressing SARS-CoV-2 spike proteins, administered intramuscularly and intranasally in mice. Both delivery methods of AAV5-spike, AAV5-spike stabilized trimer as well as AAV44.9-spike elicited robust serum anti-spike antibodies within 8-12 weeks, with high levels of anti-spike antibodies sustained for over a year. Comparison of mouse serum antibodies 16 weeks post intramuscular or intranasal AAV5 administration demonstrated similar SARS-CoV-2 spike binding neutralizing activity in vitro. Analysis of changes in cellular immunity by ELISpot at 12 weeks post-AAV spike transduction revealed interferon-{gamma} induction in response to peptide challenge. Despite a decline in AAV vector DNA at the injection site, the persistence of anti-spike antibodies demonstrated that AAV-vectors can elicit lasting immune responses, highlighting nasal AAV administration as a potential strategy to block respiratory virus infections.

immunology↗

Inhibition of ice recrystallization with designed twistless helical repeat proteins

Given the repetitive structure of crystalline ice, its unsurprising that highly active ice-binding proteins, often with beta-roll structures, also have repeating motifs. Here, we introduce a de novo designed family of ice-binding twistless alpha-helical repeat (iTHR) proteins. Each iTHR protein comprises two planar layers of parallel alpha-helices connected by loops--a structural topology not seen in native ice-binding proteins. The ice-binding helices feature an ordered array of TXXXAXXXAXX motifs, spaced to match the pyramidal {201} and secondary prism plane {110} ice lattice facets, and engineered with a 98.2{degrees} turn angle per residue to orient threonines uniformly towards the ice surface. iTHR proteins show high solubility, thermostability, and produce varied ice crystal morphologies depending on their intended target facet. Crucially, iTHRs exhibit ice recrystallization inhibition (IRI) at critical concentrations comparable to those of many native globular ice-binding proteins. Extensive site-specific mutagenesis shows that ice-binding activity in iTHR proteins is robust, remaining largely unaffected by changes in chemical composition. Variation in the repeat number reveals a non-monotonic relationship to IRI activity. X-ray crystal structures of two designs confirm the intended orientation of threonines, uniformly pointing toward the ice surface. The iTHR family provides a versatile platform to systematically investigate the complex structure-activity relationships underlying protein-ice interactions.

biophysics↗

Inwardly rectifying potassium channels regulate membrane potential polarization and direction sensing during neutrophil chemotaxis

Potassium channels regulate membrane potential and diverse physiological processes, including cell migration. However, the specific function of the inwardly rectifying potassium channels in immune cell chemotaxis is unknown. Here, we identified that the inwardly rectifying potassium channel Kir7.1 (KCNJ13) maintains the resting membrane potential and is required for directional sensing during neutrophil chemotaxis. Pharmacological or genetic inhibition of Kir7.1 in neutrophils impaired direction sensing toward various chemoattractants without affecting cell polarization in multiple neutrophil models. Using genetically encoded voltage indicators, we observed oscillating depolarization of the membrane potential in protrusions in zebrafish neutrophils, and Kir7.1 is required for polarized depolarization towards the chemokine source. Focal depolarization with optogenetic tools biases pseudopod selection and induces de novo protrusions. Global hyperpolarizing neutrophils stalled cell migration. Furthermore, Kir7.1 regulates GPCR signaling activation. This work adds membrane potential to the intricate feedforward mechanism, coupling the adaptive and excitable network required to steer immune cells in complex tissue environments.

cell biology↗

Functional Interrogation of Neuronal Subtypes via Intersectional Expression of Optogenetic Actuator Reveals Non-linear Components in a Linear Circuit

Investigating signal integration in a neural circuit is oftentimes challenging when the circuit contains neuronal subtypes that are transcriptomically similar, due to the lack of tools to express optogenetic actuators with high cellular specificity and to deliver light with high spatiotemporal accuracy. Here, we demonstrate the use of a split GAL4-based genetic "AND" gate to express Chrimson in specific touch receptor neuron (TRN) subtypes in the C. elegans touch response circuit. Combining this intersectional strategy for transgene expression with high-throughput optical targeting and behavioral quantification, we optogenetically interrogated the role of each TRN subtype in mediating the mechanosensor-induced escape response and in integrating signals that trigger the opposite motor output. Surprisingly, we found that although the response of the overall circuit linearly combines the competing anterior and posterior stimuli, this linearity is comprised of antagonistic non-linear contributions from the anterior and posterior sensors, which conspire to generate a linear response.

neuroscience↗

Branched chain fatty acid-rich diet promotes lipid droplet enlargement and impacts organismal health in Caenorhabditis elegans

Considerable amounts of branched chain fatty acids (BCFAs) are present in the human diet from beef and dairy products. BCFAs can also be produced by the human gut microbiota and synthesized from branched chain amino acids. However, the physiological impact of a BCFA-rich diet on lipid metabolism and organismal health is unclear. In this study, by screening a collection of dietary bacteria, we find that the BCFA-rich Microbacterium diet causes the formation of supersized LDs and delays development, reduces brood size, and shortens lifespan of C. elegans. The high-BCFA diet downregulates argk-1/creatine kinase to inhibit the AMPK pathway and {beta}-oxidation and upregulates fat-7/desaturase to promote the accumulation of PUFA, which enhances lipogenesis and LD expansion. We also isolate a gain-of-function mutation in scav-4/CD36, which enhances BCFA absorption and exacerbates BCFA-induced LD enlargement, demonstrating that host genetic variation in a fatty acid transporter could influence the susceptibility to a high-BCFA diet.

cell biology↗

Protocol for assessing DNA damage levels in vivo in rodent testicular germ cells in the Alkaline Comet Assay

Few protocols are available for retrieving male germ cell-specific information on DNA damage dynamics or assessing the potential harmful effects of environmental contaminants, drugs, or lifestyle factors related to genotoxicity. Here, we present a protocol for evaluating testicular germ cell genotoxicity using a modified version of the alkaline comet assay in rodent testicular germ cells. The protocol includes experimental design, preparation of testicular cell suspensions, comet analysis, germ cell-specific scoring, and data curation methods to collect information on testicular male germ cells, specifically spermatids and primary spermatocytes. For complete details on the use of this protocol please refer to Olsen et al., in preparation HighlightsO_LIDNA damage levels can be specifically measured in testicular germ cells using the developed revised version of the alkaline comet assay C_LIO_LI1C spermatids as well as 4C primary spermatocytes can be assessed C_LIO_LIBoth manual- and modeling-based approaches were developed that facilitate user-friendly protocols to select 1C spermatid comets. C_LIO_LIThis protocol expands the limited methodologies available to study germ cell DNA damage dynamics C_LI O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/624648v7_ufig1.gif" ALT="Figure 1"> View larger version (43K): org.highwire.dtl.DTLVardef@1c1e54eorg.highwire.dtl.DTLVardef@b569e9org.highwire.dtl.DTLVardef@3c74adorg.highwire.dtl.DTLVardef@110122e_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical abstractC_FLOATNO C_FIG

pharmacology and toxicology↗