Biology subjects

Do, S.

Publications and source records attributed to Do, S..

7 recordsLinked to original sources

bioRxiv2026

Stress granule fusion is a mitochondria-coordinated process for stress adaptation

Stress granules are cytoplasmic membraneless organelles assembled during stress to maintain cellular homeostasis. Although fusion is a hallmark of liquid-like behavior of these condensates, whether this process carries functional significance beyond its physical coalescence remains unclear. Here, we show that stress granule fusion is facilitated by mitochondrial dynamics and membrane potential. Intact mitochondria actively associate with stress granules, facilitating fusion and maturation. In contrast, loss of mitochondrial membrane potential, along with disrupted mitochondrial structure or motility, weakens these interactions and reduces fusion frequency. We find that impaired fusion leads to the accumulation of immature granules that retain fewer sequestered components, which correlates with premature cell death. Remarkably, restoring mitochondrial membrane potential rescues granule fusion and enlargement, and is accompanied by increased cell viability and a corresponding increase in granule-associated apoptotic factors. These results demonstrate that stress granule fusion is actively coordinated by mitochondria rather than driven solely by passive coalescence, reshaping how condensate dynamics are understood to integrate with organelle function during cellular stress response.

cell biology↗

bioRxiv2025

NIPBL-mediated 3D genome folding translates enhancer priming into gene activation and safeguards lineage fidelity during embryonic transitions

Precise gene control by complex regulatory landscapes is fundamental to embryo development, yet the instructive role of 3D genome architecture remains controversial. While acute cohesin depletion completely disrupts genome folding, it yields modest transcriptional impacts, but these findings are often confounded by cohesins essential roles in cell division and proliferation. Here, we resolve this discrepancy by decoupling architectural functions from cell-cycle roles using an acute NIPBL degron system. By integrating single-gene imaging with single-cell and bulk multi-omics during mouse pluripotency transitions and germ-layer specification, we show that NIPBL-mediated cohesin function is required for proper de novo activation of lineage-specifying genes. Mechanistically, NIPBL translates epigenetic priming into transcriptional outputs by physically bringing distal enhancers and target promoters into proximity. We further uncover a dual regulatory role: an acute requirement for establishing new enhancer-promoter interactions during cell state transitions and a long-term role in safeguarding transcriptional fidelity by preventing ectopic gene de-repression. Our findings demonstrate that NIPBL/cohesin-orchestrated genome folding facilitates the faithful execution of developmental gene expression programs. HighlightsO_LIAcute NIPBL depletion decouples the architectural functions of cohesin from its essential roles in chromosome segregation and cell cycle progression. C_LIO_LINIPBL-mediated loop extrusion is required to translate the epigenetic priming of distal enhancers into de novo gene activation during embryonic state transitions. C_LIO_LINIPBL is a "rate-limiting physical relay" required to bring distal enhancers and target promoters into proximity to initiate transcription. C_LIO_LI3D genome architecture serves a dual role: enabling acute enhancer-promoter communication and safeguarding long-term lineage fidelity by preventing ectopic gene de-repression. C_LI

genetics↗

bioRxiv2025

Two parallel lineage-committed progenitors contribute to the developing brain

The hindbrain is a life-sustaining brain region. In one model, a common neural progenitor generates all brain regions. Here our studies of mouse embryos and human pluripotent stem cells (hPSCs) support a different model: two parallel brain progenitors emerge simultaneously during gastrulation, anterior neural ectoderm (forebrain/midbrain progenitor) and posterior neural ectoderm (hindbrain progenitor). Not only are they lineage-committed to respectively form forebrain/midbrain vs. hindbrain in vitro, but they also have diverging chromatin landscapes foreshadowing future forebrain/midbrain vs. hindbrain identities. Leveraging these differences, we differentiated hPSCs into hindbrain rhombomere 5/6-specific motor neurons, hitherto difficult to generate in vitro. We postulate the brain is a composite organ emanating from two lineage-restricted progenitors; these dual progenitors may be evolutionarily conserved across 550 million years from hemichordates to mammals.

developmental biology↗

bioRxiv2024

Targeted protein degradation in lysosome utilizing naturally produced bifunctional antibodies with high levels of mannose 6-phosphate glycans

Novel antibodies have been created for targeted degradation of extracellular and membrane proteins in the lysosome. The mechanism of degradation of target proteins for these antibodies has involved either chemical conjugation of synthetic mannose 6-phosphate (M6P) or engineered bispecific antibodies. Currently, recombinant antibodies cannot be produced with naturally phosphorylated N-glycans. Here, we report the development of a novel platform technology for producing bifunctional therapeutic antibodies with high levels of M6P-bearing glycans directly from producing cells. The antibodies designated as phosphorylated N-glycosylated peptide chimeric antibodies (PNCA) maintain their affinity for antigens with concurrent high affinity binding to cell surface cation-independent mannose-6-phosphate receptors that facilitate internalization and delivery of antibody/antigen complexes to lysosomes for efficient degradation of both target extracellular soluble and membrane proteins. This PNCA approach provides a simple, scalable, and viable approach for producing naturally phosphorylated bifunctional antibodies from production cell lines for targeted protein degradation in lysosomes.

biochemistry↗

bioRxiv2024

Genetic variation modulates susceptibility to aberrant DNA hypomethylation and imprint deregulation in naive pluripotent stem cells

Naive pluripotent stem cells (nPSC) frequently undergo pathological and not readily reversible loss of DNA methylation marks at imprinted gene loci. This abnormality poses a hurdle for using pluripotent cell lines in biomedical applications and underscores the need to identify the causes of imprint instability in these cells. We show that nPSCs from inbred mouse strains exhibit pronounced strain-specific susceptibility to locus-specific deregulation of imprinting marks during reprogramming to pluripotency and upon culture with MAP kinase inhibitors, a common approach to maintain naive pluripotency. Analysis of genetically highly diverse nPSCs from the Diversity Outbred (DO) stock confirms that genetic variation is a major determinant of epigenome stability in pluripotent cells. We leverage the variable DNA hypomethylation in DO lines to identify several trans-acting quantitative trait loci (QTLs) that determine epigenome stability at either specific target loci or genome-wide. Candidate factors encoded by two multi-target QTLs on chromosomes 4 and 17 suggest specific transcriptional regulators that contribute to DNA methylation maintenance in nPSCs. We propose that genetic variants represent candidate biomarkers to identify pluripotent cell lines with desirable properties and might serve as entry points for the targeted engineering of nPSCs with stable epigenomes. HighlightsO_LINaive pluripotent stem cells from distinct inbred mouse strains exhibit variable DNA methylation levels at imprinted gene loci. C_LIO_LIThe vulnerability of pluripotent stem cells to loss of genomic imprinting caused by MAP kinase inhibition strongly differs between inbred mouse strains. C_LIO_LIGenetically diverse pluripotent stem cell lines from Diversity Outbred mouse stock allow the identification of quantitative trait loci controlling DNA methylation stability. C_LIO_LIGenetic variants may serve as biomarkers to identify naive pluripotent stem cell lines that are epigenetically stable in specific culture conditions. C_LI

developmental biology↗

bioRxiv2024

Loop-mediated Isothermal Amplification (LAMP) assay for reliable detection of Xanthomonas axonopodis pv. vasculorum

Xanthomonas axonopodis pv. vasculorum (Xav), the causative agent of sugarcane gumming disease, represents a significant threat to global sugarcane production due to its systemic and destructive nature. Despite the economic implications, a field-deployable, Xav-specific diagnostic tool has not been developed. This resulted in a loop-mediated isothermal amplification (LAMP) assay targeting the pelL gene, unique to Xav strains, as a rapid and precise diagnostic assay. The selection of the pelL gene was informed by comprehensive in silico analyses of Xav genomes and related Xanthomonas species and other close relatives. Validation against the NCBI GenBank database and internally sequenced genomes confirmed the genes exclusivity to Xav. Subsequent primers for both endpoint PCR and LAMP assays were designed using the pelL gene region. The LAMP assay underwent extensive testing against inclusivity and exclusivity panels. Use of exclusivity panel, comprising 81 strains from related species, other bacterial genera, and host genomes, demonstrated the assays specificity with no false positives. The assay exhibited a detection limit of 1 pg, and its effectiveness was unimpeded by crude host lysate (sugarcane). Further validation through multi-device and multi-operator testing underscored the assays 100% reproducibility and robustness. Application to infected plant samples resulted in the detection of all infected specimens without any false positives or negatives. This novel LAMP assay is accurate and reliable tool for Xav detection, with promising applications in routine diagnostics, biosecurity measures, microbial forensics, and epidemiological research.

plant biology↗

bioRxiv2023

Engineering Programmable Material-To-Cell Pathways Via Synthetic Notch Receptors To Spatially Control Cellular Phenotypes In Multi-Cellular Constructs

Synthetic Notch (synNotch) receptors are modular synthetic components that are genetically engineered into mammalian cells to detect signals presented by neighboring cells and respond by activating prescribed transcriptional programs. To date, synNotch has been used to program therapeutic cells and pattern morphogenesis in multicellular systems. However, cell-presented ligands have limited versatility for applications that require spatial precision, such as tissue engineering. To address this, we developed a suite of materials to activate synNotch receptors and serve as generalizable platforms for generating user-defined material-to-cell signaling pathways. First, we demonstrate that synNotch ligands, such as GFP, can be conjugated to cell- generated ECM proteins via genetic engineering of fibronectin produced by fibroblasts. We then used enzymatic or click chemistry to covalently link synNotch ligands to gelatin polymers to activate synNotch receptors in cells grown on or within a hydrogel. To achieve microscale control over synNotch activation in cell monolayers, we microcontact printed synNotch ligands onto a surface. We also patterned tissues comprising cells with up to three distinct phenotypes by engineering cells with two distinct synthetic pathways and culturing them on surfaces microfluidically patterned with two synNotch ligands. We showcase this technology by co-transdifferentiating fibroblasts into skeletal muscle or endothelial cell precursors in user-defined spatial patterns towards the engineering of muscle tissue with prescribed vascular networks. Collectively, this suite of approaches extends the synNotch toolkit and provides novel avenues for spatially controlling cellular phenotypes in mammalian multicellular systems, with many broad applications in developmental biology, synthetic morphogenesis, human tissue modeling, and regenerative medicine.

synthetic biology↗