Biology subjects

Ge, M.

Publications and source records attributed to Ge, M..

18 recordsLinked to original sources

bioRxiv2026

Systematic Profiling and Functional Characterization of Alternative Splicing in C. elegans Olfactory Learning

Alternative splicing (AS) is prevalent in neuronal gene expression. However, its function in learning has not been systematically characterized. Here, by profiling pan-neuronal translatome changes during a C. elegans learning paradigm, we show that AS remodels expression of neuronal genes with critical function in learning at the genome-wide scale. Intriguingly, AS operates on a functionally distinct gene set from those showing transcript abundance changes, serving as a separate regulatory layer in response to experience. Specifically, a neuronally enriched worm ortholog of a mitochondrial DNA helicase twnk-1 displays significant learning-associated AS changes, with both isoform types acting in a pair of sensory neurons to regulate learning with distinct functions. AS of twnk-1 modulates a cell-nonautonomous signal from neuronal mitochondria to peripheral tissues to regulate physiological states critical for learning. These results establish AS as a systematic regulator of learning and mechanistically reveal how AS shapes physiological states to facilitate learning.

neuroscience↗

bioRxiv2026

Oxidized protein states define metastatic fitness in lung cancer

AbstractReactive oxygen species (ROS) are a pervasive feature of human cancers, yet the protein targets through which ROS-regulated cell states shape tumor biology remains poorly understood. Here, using cysteine chemical proteomics, we define signatures of protein states under distinct cellular ROS environments that capture protein oxidation and conformational changes. Quantifying these signatures in primary lung tumors and brain metastases revealed a surprising enrichment of oxidative states in metastasis. To determine how these states support fitness, we performed genome-wide CRISPR screens and identified the mitochondrial Complex I subunit NDUFA10 as a key oxidation-dependent vulnerability. Oxidation of NDUFA10*Cys253 supports Complex I function through a previously unrecognized nucleotide kinase activity that maintains mitochondrial DNA levels. Enforcing a reduced conformation in NDUFA10 disrupts brain metastatic colonization in vivo. These findings establish ROS regulated protein states as a functional layer of tumor fitness, providing a framework for identifying redox-dependent mechanisms that support cancer progression.

Cancer Biology↗

bioRxiv2025

Aversive Learning Induces Context-Gated Global Reorganization of Neural Dynamics in Caenorhabditis elegans

Learning generates experience-dependent changes to the brain. However, how neurons of diverse functions and connectivity reorganize and modulate their activities to generate coherent changes while preserving essential functions is not well understood. Here, we address this question using an aversive olfactory learning paradigm whereby Caenorhabditis elegans learns to reduce its olfactory preference for pathogenic bacteria. Using functional imaging during olfactory stimulation in naive and trained animals, we show that, at brain-wide scale, cell type-by-cell type, learning induces context-gated reconfiguration of the organization of neural activity throughout the brain to alter neural responses only during bacteria-discrimination task, while leaving intact bacteria-sensing functions. We found that the context-gated encoding of learning is globally distributed across layers of the nervous system, composed of neurons carrying information of context or learning experience. In particular, aversive training modulates multiple functional connections within the nervous system, including those between sensory neurons and interneurons, as well as those among interneurons, in a context-gated manner. At the systems level, training modulates correlated activity of neural populations; we found that low-dimensional temporal patterns of population activity correlate well with locomotory gaits that express olfactory preferences. Upon training, the rotation and contraction of the low-dimensional neural manifolds shifts the brain into predisposed states for the context-gated display of learned behavior. Because animals encounter unpredictable environments in life, efficient learning about relevant cues while maintaining other functions is essential for survival. Our findings uncover network-level mechanisms that help explain how the brain reorganizes its activity patterns to both encode new experience and preserve essential functions.

neuroscience↗

bioRxiv2024

TFE3 fusions direct an oncogenic transcriptional program that drives OXPHOS and unveils vulnerabilities in translocation renal cell carcinoma

Translocation renal cell carcinoma (tRCC) is an aggressive subtype of kidney cancer driven by TFE3 gene fusions, which act via poorly characterized downstream mechanisms. Here we report that TFE3 fusions transcriptionally rewire tRCCs toward oxidative phosphorylation (OXPHOS), contrasting with the highly glycolytic metabolism of most other renal cancers. This TFE3 fusion-driven OXPHOS program, together with heightened glutathione levels found in renal cancers, renders tRCCs sensitive to reductive stress - a metabolic stress state induced by an imbalance of reducing equivalents. Genome-scale CRISPR screening identifies tRCC-selective vulnerabilities linked to this metabolic state, including EGLN1, which hydroxylates HIF-1 and targets it for proteolysis. Inhibition of EGLN1 compromises tRCC cell growth by stabilizing HIF-1a and promoting metabolic reprogramming away from OXPHOS, thus representing a vulnerability to OXPHOS-dependent tRCC cells. Our study defines a distinctive tRCC-essential metabolic program driven by TFE3 fusions and nominates EGLN1 inhibition as a therapeutic strategy to counteract fusion-induced metabolic rewiring.

cancer biology↗

bioRxiv2024

Systematic Targeting of Protein Complexes with Molecular COUPLrs

Molecular glues that engage protein complexes have transformed the study of cell biology and have had a direct impact on clinical oncology. However, the identification of new glue classes and their corresponding protein complexes has remained largely serendipitous. To overcome this challenge, we report the development of molecular COUPLrs, elaborated small molecules flanked by two cysteine-reactive warheads, as well as CONNECT, an integrated chemical proteomic platform for target deconvolution. By profiling a library of molecular COUPLrs across 13 cancer cell lines, we uncovered hundreds of proteins that can be coupled together, including in some cases in mutant selective fashions. We develop an advanced COUPLr for the oncogene EML4-ALK, which engages the fusion outside of its kinase domain, restricts protein dynamics, and disrupts EML4-ALK signaling. Collectively, molecular COUPLrs substantially expand the scope of proteins that can be chemically connected, providing an unbiased approach to identify small molecules that target protein complexes.

systems biology↗

bioRxiv2023

DrugMap: A quantitative pan-cancer analysis of cysteine ligandability

Cysteine-focused chemical proteomic platforms have accelerated the clinical development of covalent inhibitors of a wide-range of targets in cancer. However, how different oncogenic contexts influence cysteine targeting remains unknown. To address this question, we have developed DrugMap, an atlas of cysteine ligandability compiled across 416 cancer cell lines. We unexpectedly find that cysteine ligandability varies across cancer cell lines, and we attribute this to differences in cellular redox states, protein conformational changes, and genetic mutations. Leveraging these findings, we identify actionable cysteines in NF{kappa}B1 and SOX10 and develop corresponding covalent ligands that block the activity of these transcription factors. We demonstrate that the NF{kappa}B1 probe blocks DNA binding, whereas the SOX10 ligand increases SOX10-SOX10 interactions and disrupts melanoma transcriptional signaling. Our findings reveal heterogeneity in cysteine ligandability across cancers, pinpoint cell-intrinsic features driving cysteine targeting, and illustrate the use of covalent probes to disrupt oncogenic transcription factor activity.

systems biology↗

bioRxiv2023

CH4 transport in wetland plants under controlled environmental conditions: untangling the impacts of phenology

O_LIMethane (CH4) fluxes at plant surfaces are the net result of transport of soil-produced CH4 and within-plant CH4 production and consumption, yet factors and processes controlling these fluxes remain unclear. C_LIO_LIWe conducted high-frequency automated CH4 flux measurements from shoots of Carex rostrata (sedge), Menyanthes trifoliata (forb) and shrubs (Betula nana, Salix lapponum) during early, high and late summer in a climate-controlled environment to assess the effects of environmental variables, seasonality and CH4 cycling microbes in the CH4 flux. Measurements were conducted from intact plant-soil samples collected throughout growing seasons 2020 and 2021 from Lompolojankkafen, northern Finland. C_LIO_LIAll studied species showed seasonal variability in CH4 fluxes. The CH4 fluxes were not impacted by light level, while out of the studied species, porewater CH4 concentration increased fluxes from all but B. nana. Air temperature only and negatively affected CH4 flux from C. rostrata. Both methanogens and methanotrophs were detected in aboveground parts of S. lapponum and M. trifoliata, methanotrophs in B. nana, while neither were detected in C. rostrata. C_LIO_LIOur study demonstrates that the seasonal phase of the plants regulates CH4 flux they mediate across species, which was not observed in the field. The detection of methanogens and methanotrophs in herbs and shrubs suggests that microbial processes may contribute to their CH4 flux. C_LI

ecology↗

bioRxiv2023

Loss of Mitochondrial Enoyl CoA Reductase causes elevated ceramide levels and impairs iron metabolism

In most eukaryotic cells fatty acid synthesis occurs in the cytoplasm as well as in mitochondria. However, the relative contribution of mitochondrial fatty acid synthesis (mtFAS) to the cellular lipidome of metazoans is ill-defined. Hence, we studied the function of the fly Mitochondria enoyl CoA reductase (Mecr), the enzyme required for the last step of mtFAS. Loss of mecr causes lethality while neuronal loss leads to progressive neurological defects. We observe an elevated level of ceramides, a defect in Fe-S cluster biogenesis and increased iron levels in mecr mutants. Reducing the levels of either iron or ceramide suppresses the neurodegenerative phenotypes indicating that increased ceramides and iron metabolism are interrelated and play an important role in the pathogenesis. Mutations in human MECR cause pediatric-onset neurodegeneration and patient-derived fibroblasts display similar elevated ceramide levels and impaired iron homeostasis. In summary, this study shows an as-yet-unidentified role of mecr/MECR in ceramide and iron metabolism providing a mechanistic link between mtFAS and neurodegeneration.

neuroscience↗

bioRxiv2023

Multi-omics and Pharmacological Characterization of Patient-derived Glioma Cell Lines

Glioblastoma (GBM) is the most common brain tumor and is currently incurable. Primary GBM cultures are widely used tools for screening potentially therapeutic drugs; however, there is a lack of genomic and pharmacological characterization of these primary GBM cultures. Here, we collected 52 patient-derived glioma cell (PDGC) lines and characterized them through whole- genome sequencing (WGS), RNA-seq, and drug response screening. We identified three molecular subtypes among PDGCs: mesenchymal (MES), proneural (PN), and oxidative phosphorylation (OXPHOS). Upon profiling the responses of PDGCs to 214 drugs, we found that the PN subtype PDGCs were sensitive to tyrosine kinase inhibitors, whereas the OXPHOS subtype PDGCs were sensitive to histone deacetylase inhibitors, oxidative phosphorylation inhibitors, and HMG-CoA reductase inhibitors. PN and OXPHOS subtype PDGCs stably formed tumors in vivo upon intracranial transplantation into immunodeficient mice, while most MES subtype PDGCs were incapable of tumorigenesis in vivo. In addition, profiling and follow-up investigations showed that the serum-free culture system used for PDGCs enriched and propagated rare MYC/MYCN- amplified glioma cells. Our study provides a resource for understanding primary glioma cell cultures and aiding clinical translation. SignificanceOur study provides a resource for patient-derived glioma cell lines (PDGCs) on transcriptome, genome, drug response, and tumorigenic abilities. PDGCs are categorized into PN, MES, and OXPHOS subtypes, with MES-subtype PDGCs incapable of tumorigenesis in vivo. Notably, the serum-free culture system for PDGCs enriches glioma cells with MYC/MYCN amplification.

cancer biology↗

bioRxiv2023

Comprehensive characterization of tumor microenvironment in colorectal cancer via histopathology-molecular analysis

PurposeTo explain how the tumor microenvironment (TME) contributes to biological and clinical heterogeneity of colorectal cancer (CRC). MethodsUsing multi-omics analysis, single cell transcriptomic sequencing analysis and artificial intelligence-enabled spatial analysis of whole-slide images, we performed a comprehensive characterization of TME in colorectal cancer (CCCRC). ResultsCRC samples were classified into four CCCRC subtypes with distinct TME features, namely, C1 as the proliferative subtype with low immunogenicity; C2 as the immunosuppressed subtype with the terminally exhausted immune characteristics; C3 as the immune-excluded subtype with the distinct upregulation of stromal components and a lack of T cell infiltration in tumor core; and C4 as the immunomodulatory subtype with the remarkable upregulation of anti-tumor immune components. The four CCCRC subtypes had distinct histopathological and molecular characteristics, therapeutic efficacy, and prognosis. The C1 subtype was more sensitive to chemotherapy, the C2 and C3 subtypes were more sensitive to WNT pathway inhibitor SB216763 and Hedgehog pathway inhibitor vismodegib, and the C4 subtype was suitable for ICB treatment. Finally, we established a single-sample gene classifier for identifying the CCCRC subtypes. ConclusionsOur integrative analyses ultimately established a holistic framework to thoroughly dissect the TME of CRC, and the CCCRC classification system with high biological interpretability might facilitate biomarker discoveries and clinical treatment decisions in the future.

cancer biology↗

bioRxiv2023

Single-cell sequencing for the molecular identity involved in zinc therapy for spinal cord injury

Spinal cord injury (SCI) is a severe traumatic neurological condition often caused by car accidents, violent impacts, or unintentional falls and can have devastating consequences for the patient. SCI include primary and secondary injuries. Followed by primary injury, and secondary injuries including inflammatory responses and oxidative stress, which exacerbate the disease process of SCI. Therefore, the treatment of SCI, especially the suppression of secondary injuries, is a major focus of attention in the field of neuroscience. In previous studies, we have demonstrated that zinc therapy can exert neuroprotective effects in mice after acute SCI, including reducing the inflammatory response in the central nervous system, decreasing the neuronal apoptosis and downregulating oxidative stress at the region of injury. However, little is known about how zinc therapy systematically alleviates SCI. Here, we have systematically analysed and mapped the single-cell atlas of the spinal cord in mice with SCI treated by zinc therapy, which provides a systematic analysis of the transcriptome of individual cells in the spinal cord. Among the results, we found that zinc therapy-induced alterations in the IL-17 inflammatory pathway and produced immune heterogeneity in microglia which are the inherent immune cells in the central nervous system. By cell subpopulations clustering analysis, we defined seven microglia subpopulations in zinc-therapy spinal cord tissue and identified the presence of novel VEGFA+ microglia. This finding presents that the microglia subpopulations affected by zinc therapy may have the potential to promote angiogenesis, which is a potential mechanism for the treatment of SCI. In conclusion, this study maps/reveals the transcriptomic of zinc therapy for the treatment of acute SCI with unprecedented resolution and provides a molecular basis for the clinical application.

genomics↗

bioRxiv2022

Distinct super-enhancer elements differentially control Il2ra gene expression in a cell-type specific fashion

The IL-2 receptor -chain (IL-2R/CD25) is constitutively expressed on DN2/DN3 thymocytes and Treg cells but induced by IL-2 on mature T and NK cells. Il2ra expression is regulated by a super-enhancer extensively bound by STAT5 in mature T cells. Here, we demonstrate that STAT5 cooperates with Notch to induce/maintain Il2ra/CD25 expression in DN2/DN3 cells. Moreover, we systematically investigated CD25 regulation using a series of mice with deletions spanning STAT5 binding elements. Deleting the upstream super-enhancer region mainly affected constitutive CD25 expression on DN2/DN3 thymocytes and Tregs, whereas deleting an intronic region primarily decreased IL-2-induced CD25 on peripheral T and NK cells. Thus, distinct elements preferentially control constitutive versus inducible expression in a cell-type-specific manner, with the MED1 coactivator co-localizing with specific STAT5 binding sites. Moreover, the intronic region was a dominant element whose deletion altered the structure throughout the super-enhancer in mature T cells. These results demonstrate differential functions for distinct super-enhancer elements, thereby indicating ways to manipulate CD25 expression in a cell-type specific fashion.

immunology↗

bioRxiv2022

Empagliflozin reduces renal lipotoxicity in experimental Alport syndrome

Sodium-glucose cotransporter-2 inhibitors (SGLT2i) are anti-hyperglycemic agents that prevent glucose reabsorption in proximal tubular cells. SGLT2i improves renal outcomes in both diabetic and non-diabetic patients, indicating it may have beneficial effects beyond glycemic control. Here, we demonstrate that SGLT2i affects energy metabolism and renal lipotoxicity in experimental Alport syndrome (AS). In vitro, we found that SGLT2 protein expression levels in human and mouse podocytes were similar to tubular cells. Newly established immortalized podocytes from Col4a3 knockout mice (AS podocytes) accumulate lipid droplets along with increased apoptosis when compared to wildtype podocytes. Treatment with SGLT2i empagliflozin reduces lipid droplet accumulation and apoptosis in AS podocytes. Empagliflozin inhibits the utilization of glucose/pyruvate as a metabolic substrate in AS podocytes. In vivo, we demonstrate that empagliflozin reduces albuminuria and prolongs the survival of AS mice. Empagliflozin-treated AS mice show decreased serum blood urea nitrogen and creatinine levels in association with reduced triglyceride and cholesterol ester content in kidney cortices when compared to AS mice. Lipid accumulation in kidney cortices correlates with the decline in renal function. In summary, empagliflozin reduces renal lipotoxicity and improves kidney function in experimental AS in association with the energy substrates switch from glucose to fatty acids in podocytes.

cell biology↗

bioRxiv2022

NRF2 activation induces NADH-reductive stress providing a metabolic vulnerability in lung cancer

Multiple cancers regulate oxidative stress by activating the transcription factor NRF2 through mutation of its negative regulator KEAP1. NRF2 has been studied extensively in KEAP1-mutant cancers, however the role of this pathway in cancers with wildtype KEAP1 remains poorly understood. To answer this question, we induced NRF2 via pharmacological inactivation of KEAP1 in a panel of 50+ non-small lung cancer cell lines. Unexpectedly, marked decreases in viability were observed in >13% of the cell lines--an effect that was completely rescued by NRF2 ablation. Genome-wide and targeted CRISPR screens revealed that NRF2 induces NADH-reductive stress, through the upregulation of the NAD+-consuming enzyme ALDH3A1. Leveraging these findings, we show that cells treated with KEAP1 inhibitors or those with endogenous KEAP1 mutations are selectively vulnerable to Complex I inhibition, which impairs NADH oxidation capacity and potentiates reductive stress. Thus, we identify reductive stress as a metabolic vulnerability in NRF2-activated lung cancers.

molecular biology↗

bioRxiv2021

An expanded toolkit for Drosophila gene tagging using synthesized homology donor constructs for CRISPR mediated homologous recombination

Previously, we described a large collection of Drosophila strains that each carry an artificial exon containing a T2AGAL4 cassette inserted in an intron of a target gene based on CRISPR-mediated homologous recombination (Lee et al., 2018). These alleles permit numerous applications and have proven to be very useful. Initially, the homologous recombination-based donor constructs had long homology arms (>500 bps) to promote precise integration of large constructs (>5kb). Recently, we showed that in vivo linearization of the donor constructs enables insertion of large artificial exons in introns using short homology arms (100-200 bps) (Kanca et al., 2019a). Shorter homology arms make it feasible to commercially synthesize homology donors and minimize the cloning steps for donor construct generation. Unfortunately, about 50% of Drosophila genes lack suitable coding introns for integration of artificial exons. Here, we report the development of new set of constructs that allow the replacement of the coding region of genes that lack suitable introns with a KozakGAL4 cassette, generating a knock-out/knock-in allele that expresses GAL4 similarly as the targeted gene. We also developed custom vector backbones to further facilitate and improve transgenesis. Synthesis of homology donor constructs in custom plasmid backbones that contain the target gene sgRNA obviates the need to inject a separate sgRNA plasmid and significantly increases the transgenesis efficiency. These upgrades will enable the targeting of nearly every fly gene, regardless of exon-intron structure, with a 70-80% success rate.

genetics↗

bioRxiv2021

Escape Steering by Cholecystokinin Peptidergic Signaling

Escape is an evolutionarily conserved and essential avoidance response. Considered to be innate, most studies on escape responses focused on hard-wired circuits. We report here that peptidergic signaling is an integral and necessary component of the Caenorhabditis elegans escape circuit. Combining genetic screening, electrophysiology and calcium imaging, we reveal that a neuropeptide NLP-18 and its cholecystokinin receptor CKR-1 enable the escape circuit to execute a full omega ({Omega}) turn, the last motor step where the animal robustly steers away from its original trajectory. We demonstrate in vivo and in vitro that CKR-1 is a Gq protein coupled receptor for NLP-18. in vivo, NLP-18 is mainly secreted by the gustatory sensory neuron (ASI) to activate CKR-1 in the head motor neuron (SMD) and the turn-initiating interneuron (AIB). Removal of NLP-18, removal of CKR-1, or specific knockdown of CKR-1 in SMD or AIB neurons lead to shallower turns hence less robust escape steering. Consistently, elevation of head motor neuron (SMD)s Ca2+ transients during escape steering is attenuated upon the removal of NLP-18 or CKR-1. in vitro, synthetic NLP-18 directly evokes CKR-1-dependent currents in oocytes and CKR-1-dependent Ca2+ transients in SMD. Thus, cholecystokinin signaling modulates an escape circuit to generate robust escape steering.

neuroscience↗

bioRxiv2021

A T-cell independent universal cellular therapy strategy through antigen depletion

CD19-targeting chimeric antigen receptor (CAR) T-cell therapeutics is a revolutionary, novel and successful treatment for B-cell malignancies. However, while CD19-CAR-T therapy can obtain high rates of complete responses in these patients, a significant fraction of patients may experience CD19-negative relapse. Moreover, the dependency on T-cell mediated cytotoxicity restricts CAR-T therapy as a patient-specific individualized therapy with severe side effects such as cytokine-release syndrome (CRS). Whether CAR-T therapy can be substituted by a non-T-cell based universal cellular therapy is largely unknown. Surprisingly, we have demonstrated here that T-lymphocytic cells, as well as non-lymphocytic cells, can cause CD19 internalization and subsequent depletion when they are armed with a CD19-recognizing moiety. This CD19 antigen depletion can efficiently induce T-cell independent apoptosis in target cancer cells whose survival is dependent upon CD19 expression, suggesting that CD19 antigen depletion constitutes a crucial tumor destroying mechanism for CD19-CAR-T, especially for its long-term efficacy. We therefore proposed a universal strategy for CRS-free cellular therapeutics, utilizing artificial antigen-recognizing cells (AARC), which can be manufactured universally and standardly as "off-the-shelf" mesenchymal stromal cells (MSCs) or other types of non-autologous cell expressing anergic CARs. Our results not only uncovered an unrecognized mechanism for CAR-T cytotoxicity and antigen loss, but also shed new insight into a shift in cellular therapeutics from unique patient-specific autologous therapeutics, to universal and standardized allogeneic treatment.

cancer biology↗

bioRxiv2019

An efficient CRISPR-based strategy to insert small and large fragments of DNA using short homology arms

We previously reported a CRISPR-mediated knock-in strategy into introns of Drosophila genes, generating an attP-FRT-SA-T2A-GAL4-polyA-3XP3-EGFP-FRT-attP transgenic library for multiple uses (Lee et al., 2018b). The method relied on double stranded DNA (dsDNA) homology donors with [~]1 kb homology arms. Here, we describe three new simpler ways to edit genes in flies. We create single stranded DNA (ssDNA) donors using PCR and add 100 nt of homology on each side of an integration cassette, followed by enzymatic removal of one strand. Using this method, we generated GFP-tagged proteins that mark organelles in S2 cells. We then describe two dsDNA methods using cheap synthesized donors flanked by 100 nt homology arms and gRNA target sites cloned into a plasmid. Upon injection, donor DNA (1 to 5 kb) is released from the plasmid by Cas9. The cassette integrates efficiently and precisely in vivo. The approach is fast, cheap, and scalable.

genomics↗