Biology subjects

Cho, T.

Publications and source records attributed to Cho, T..

11 recordsLinked to original sources

bioRxiv2026

Transfer Learning Enables Drug-Target Interaction Prediction in Data-Scarce One-Carbon Metabolism

Predicting drug-target interactions (DTIs) with deep learning offers opportunities to accelerate drug discovery, yet performance is constrained by the scarcity of target-specific training data. This is a particular challenge for mitochondrial one-carbon (1C) pathway enzymes, which are attractive therapeutic targets but remain pharmacologically understudied. Mitochondrial 1C metabolism supplies glycine, reducing equivalents, and 1C units critical for nucleotide synthesis, and has emerged as a key pathway in cancer and fibrosis. SHMT2 and MTHFD2, two key 1C enzymes, support collagen production in fibroblasts, blocking either prevents TGF-{beta}-induced glycine and collagen accumulation. Here, we developed transfer learning-based deep learning models to predict interactions between approved drugs and SHMT2 or MTHFD2 despite minimal target-specific training data, pre-training on large datasets from related enzymes before fine-tuning to these targets. Virtual screening of the DrugBank library identified six candidates, three of which, Carbimazole, Crizotinib, and GSK2018682 reduced TGF-{beta}-induced collagen production and glycine accumulation in human lung fibroblasts, demonstrating transfer learning as a strategy for repurposable drug identification in data-scarce metabolic targets.

bioinformatics↗

bioRxiv2025

De novo Design of a Peptide Modulator to Reverse Sodium Channel Dysfunction Linked to Cardiac Arrhythmias and Epilepsy

Ion channels orchestrate electrical signaling in excitable cells. In nature, ion channel function is customized by modulatory proteins that have evolved to fulfill distinct physiological needs. Yet, engineering synthetic modulators that precisely tune ion channel function is challenging. One example involves the voltage-gated sodium (NaV) channel that initiates the action potential, and whose dysfunction amplifies late/persistent sodium current (INaL), a commonality that underlies various human diseases including cardiac arrhythmias and epilepsy. Here, using a computational protein design platform, we engineered a de novo peptide modulator, ELIXIR, that binds NaV channels with submicromolar affinity. Functional analysis revealed an unexpected selectivity in inhibiting pathogenic INaL and confirmed its effectiveness in reversing NaV dysfunction linked to both cardiac arrhythmias and epilepsy in cellular and murine models. These findings exemplify the efficacy of de novo protein design for engineering synthetic ion channel modulators and sets the stage for rational design of future therapeutic approaches.

biophysics↗

bioRxiv2024

Role of Arginine and its Metabolism in TGF-β-Induced Activation of Lung Fibroblasts

Arginine is a conditionally essential amino acid with known roles in protein production, nitric oxide synthesis, biosynthesis of proline and polyamines, and regulation of intracellular signaling pathways. Arginine biosynthesis and catabolism have been linked to TGF-{beta}-induced activation of fibroblasts in the context of pulmonary fibrosis; however, a thorough study on the metabolic and signaling roles of arginine in the process of fibroblast activation has not been conducted. Here, we used metabolic dropouts and labeling strategies to determine how activated fibroblasts utilize arginine. We found that arginine limitation leads to activation of GCN2 while inhibiting TGF-{beta}-induced mTORC1 activation and collagen protein production. Extracellular citrulline could rescue the effect of arginine deprivation in an ASS1-dependent manner. Using metabolic tracers of arginine and its precursors, we found little evidence of arginine synthesis or catabolism in lung fibroblasts treated with TGF-{beta}. Extracellular ornithine or glutamine were the primary sources of ornithine and polyamines, not arginine. Our findings suggest that the major role for arginine in lung fibroblasts is for charging of arginyl-tRNAs and for promotion of mTOR signaling. Highlights- Arginine depletion inhibits TGF-{beta}-induced transcription in human lung fibroblasts (HLFs). - Arginine is not significantly catabolized in HLFs either through NOS or ARG dependent pathways. - Extracellular glutamine and ornithine are the primary sources of polyamines in lung fibroblasts. - The primary role of arginine in lung fibroblasts is for signaling through mTOR and GNC2.

cell biology↗

bioRxiv2023

Mitochondrial One-Carbon Metabolism is Required for TGF-β-Induced Glycine Synthesis and Collagen Protein Production

A hallmark of Idiopathic Pulmonary Fibrosis is the TGF-{beta}-dependent activation of lung fibroblasts, leading to excessive deposition of collagen proteins and progressive scarring. We have previously shown that synthesis of collagen by lung fibroblasts requires de novo synthesis of glycine, the most abundant amino acid in collagen protein. TGF-{beta} upregulates the expression of the enzymes of the de novo serine/glycine synthesis pathway in lung fibroblasts through mTORC1 and ATF4- dependent transcriptional programs. SHMT2, the final enzyme of the de novo serine/glycine synthesis pathway, transfers a one-carbon unit from serine to tetrahydrofolate (THF), producing glycine and 5,10-methylene-THF (meTHF). meTHF is converted back to THF in the mitochondrial one-carbon (1C) pathway through the sequential actions of MTHFD2 (which converts meTHF to 10-formyl-THF), and either MTHFD1L, which produces formate, or ALDH1L2, which produces CO2. It is unknown how the mitochondrial 1C pathway contributes to glycine biosynthesis or collagen protein production in fibroblasts, or fibrosis in vivo. Here, we demonstrate that TGF-{beta} induces the expression of MTHFD2, MTHFD1L, and ALDH1L2 in human lung fibroblasts. MTHFD2 expression was required for TGF-{beta}-induced cellular glycine accumulation and collagen protein production. Combined knockdown of both MTHFD1L and ALDH1L2 also inhibited glycine accumulation and collagen protein production downstream of TGF-{beta}; however knockdown of either protein alone had no inhibitory effect, suggesting that lung fibroblasts can utilize either enzyme to regenerate THF. Pharmacologic inhibition of MTHFD2 recapitulated the effects of MTHFD2 knockdown in lung fibroblasts and ameliorated fibrotic responses after intratracheal bleomycin instillation in vivo. Our results provide insight into the metabolic requirements of lung fibroblasts and provide support for continued development of MTHFD2 inhibitors for the treatment of IPF and other fibrotic diseases.

cell biology↗

bioRxiv2023

Chemogenetic profiling of ubiquitin-like modifier pathways identifies NFATC2IP as a mediator of SUMO-dependent genome integrity

The post-translational modification of proteins by ubiquitin and ubiquitin-like polypeptides controls multiple cellular processes including the abundance of a large fraction of the proteome. We applied genome-scale CRISPR/Cas9 screens to elucidate the genetic architecture of the response to inhibition of ubiquitin, NEDD8 and SUMO conjugation pathways as well as inhibition of the p97/VCP segregase. This effort identified 395 genes whose disruption alters the fitness of human cells when faced with perturbations in these pathways. We validated that the TMED2 and TMED10 proteins, which are localized to the secretory pathway, promote resistance to p97/VCP inhibition and also characterized NFATC2IP, an evolutionarily conserved protein harboring SUMO-like domains as a major player in promoting genomic integrity when SUMOylation is inhibited. We propose that NFATC2IP acts in interphase cells to promote the SUMO-dependent E3 ligase activity of the SMC5/SMC6 complex, which is critical for SUMO-dependent genome integrity.

molecular biology↗

bioRxiv2022

Genome-wide CRISPR screens identify novel regulators of wild-type and mutant p53 stability

Tumour suppressor p53 (TP53) is the most frequently mutated gene in cancer. Several hotspot p53 mutants not only lose tumour suppressive capabilities, but also function in a dominant-negative manner, suppressing canonical wild-type p53 function. Furthermore, some hotspot p53 mutants promote oncogenesis by gain-of-function mechanisms. Levels of p53 are regulated predominantly through regulation of protein stability and while wild-type p53 is normally kept at very low levels at steady-state, p53 mutants are often stabilized in tumours, which may be vital for their oncogenic properties. Here, we systematically profiled the factors that regulate protein stability of wild-type and mutant p53 using marker-based genome-wide CRISPR screens. We found that most proteins that regulate wild-type p53 also regulate a subset of p53 mutants with the exception of p53 R337H regulators, which are largely private to this mutant. Mechanistically, we identified FBXO42 as a novel positive regulator of a subset of p53 mutants comprising R273H, R248Q and R248W. We show that FBXO42 acts together with CCDC6 to regulate USP28-mediated p53 stabilization. Our work also identifies C16orf72 as a negative regulator of the stability of wild-type p53 and of all p53 mutants tested. C16orf72 is amplified in breast cancer, and we show that C16orf72 regulates p53 levels in mammary epithelium of mice and its overexpression results in accelerated breast cancer with reduced p53 levels. Together, this work provides a network view of the processes that regulate p53 stability, which might provide clues for reinforcing wild-type p53 or targeting mutant p53 in cancer.

cancer biology↗

bioRxiv2021

Multiple Sclerosis Patients have an Altered Gut Mycobiome and Increased Fungal to Bacterial Richness Running Title: RRMS Patients have Gut Fungal Dysbiosis

Trillions of microbes such as bacteria, fungi, and viruses exist in the healthy human gut microbiome. Although gut bacterial dysbiosis has been extensively studied in multiple sclerosis (MS), the significance of the fungal microbiome (mycobiome) is an understudied and neglected part of the intestinal microbiome in MS. The aim of this study was to characterize the gut mycobiome of patients with relapsing-remitting multiple sclerosis (RRMS), compare it to healthy controls, and examine its association with changes in the bacterial microbiome. We characterized and compared the mycobiome of 20 RRMS patients and 33 healthy controls (HC) using Internal Transcribed Spacer 2 (ITS2) and compared mycobiome interactions with the bacterial microbiome using 16S rRNA sequencing. Our results demonstrate an altered mycobiome in RRMS patients compared with HC. RRMS patients showed an increased abundance of Basidiomycota and decreased Ascomycota at the phylum level with an increased abundance of Candida and Epicoccum genera along with a decreased abundance of Saccharomyces compared to HC. We also observed an increased ITS2/16S ratio, altered fungal and bacterial associations, and altered fungal functional profiles in MS patients compared to HC. This study demonstrates that RRMS patients had a distinct mycobiome with associated changes in the bacterial microbiome compared to HC. There is an increased fungal to bacterial ratio as well as more diverse fungal-bacterial interactions in RRMS patients compared to HC. Our study is the first step towards future studies in delineating the mechanisms through which the fungal microbiome can influence MS disease.

neuroscience↗

bioRxiv2020

Enhanced virus detection and metagenomic sequencing in patients with meningitis and encephalitis

Meningitis and encephalitis are leading causes of central nervous system (CNS) disease and often result in severe neurological compromise or death. Traditional diagnostic workflows largely rely on pathogen-specific diagnostic tests, sometimes over days to weeks. Metagenomic next-generation sequencing (mNGS) is a high-throughput platform that profiles all nucleic acid in a sample. We prospectively enrolled 68 patients from New England with known or suspected CNS infection and performed mNGS from both RNA and DNA to identify potential pathogens. Using a computational metagenomic classification pipeline based on KrakenUniq and BLAST, we detected pathogen nucleic acid in cerebrospinal fluid (CSF) from 22 subjects. This included some pathogens traditionally diagnosed by serology or not typically identified in CSF, including three transmitted by Ixodes scapularis ticks (Powassan virus, Borrelia burgdorferi, Anaplasma phagocytophilum). Among 24 subjects with no clinical diagnosis, we detected enterovirus in two subjects and Epstein Barr virus in one subject. We also evaluated two methods to enhance detection of viral nucleic acid, hybrid capture and methylated DNA depletion. Hybrid capture nearly universally increased viral read recovery. Although results for methylated DNA depletion were mixed, it allowed detection of varicella zoster virus DNA in two samples that were negative by standard mNGS. Overall, mNGS is a promising approach that can test for multiple pathogens simultaneously, with similar efficacy to pathogen-specific tests, and can uncover geographically relevant infectious CNS disease, such as tick-borne infections in New England. With further laboratory and computational enhancements, mNGS may become a mainstay of workup for encephalitis and meningitis. ImportanceMeningitis and encephalitis are leading global causes of central nervous system (CNS) disability and mortality. Current diagnostic workflows remain inefficient, requiring costly pathogen-specific assays and sometimes invasive surgical procedures. Despite intensive diagnostic efforts, 40-60% of people with meningitis or encephalitis have no clear cause of their CNS disease identified. As diagnostic uncertainty often leads to costly inappropriate therapies, the need for novel pathogen detection methods is paramount. Metagenomic next-generation sequencing (mNGS) offers the unique opportunity to circumvent these challenges using unbiased laboratory and computational methods. Here, we performed comprehensive mNGS from 68 patients with suspected CNS infection, and define enhanced methods to improve the detection of CNS pathogens, including those not traditionally identified in the CNS by nucleic acid detection. Overall, our work helps elucidate how mNGS can become a mainstay in the diagnostic toolkit for CNS infections.

neuroscience↗

bioRxiv2019

A genetic map of the response to DNA damage in human cells

The response to DNA damage is critical for cellular homeostasis, tumor suppression, immunity and gametogenesis. In order to provide an unbiased and global view of the DNA damage response in human cells, we undertook 28 CRISPR/Cas9 screens against 25 genotoxic agents in the retinal pigment epithelium-1 (RPE1) cell line. These screens identified 840 genes whose loss causes either sensitivity or resistance to DNA damaging agents. Mining this dataset, we uncovered that ERCC6L2, which is mutated in a bone-marrow failure syndrome, codes for a canonical non-homologous end-joining pathway factor; that the RNA polymerase II component ELOF1 modulates the response to transcription-blocking agents and that the cytotoxicity of the G-quadruplex ligand pyridostatin involves trapping topoisomerase II on DNA. This map of the DNA damage response provides a rich resource to study this fundamental cellular system and has implications for the development and use of genotoxic agents in cancer therapy.

cancer biology↗

bioRxiv2019

A consensus set of genetic vulnerabilities to ATR inhibition

The response to DNA replication stress in eukaryotes is under the control of the ataxia-telangiectasia and Rad3-related (ATR) kinase. ATR responds to single-stranded (ss) DNA to stabilize distressed DNA replication forks, modulate DNA replication firing and prevent cells with damaged DNA or incomplete DNA replication from entering into mitosis. Furthermore, inhibitors of ATR are currently in clinical development either as monotherapies or in combination with agents that perturb DNA replication. To gain a genetic view of the cellular pathways requiring ATR kinase function, we mapped genes whose mutation causes hypersensitivity to ATR inhibitors with genome-scale CRISPR/Cas9 screens. We delineate a consensus set of 117 genes enriched in DNA replication, DNA repair and cell cycle regulators that promote survival when ATR kinase activity is suppressed. We validate 14 genes from this set and report genes not previously described to modulate response to ATR inhibitors. In particular we found that the loss of the POLE3/POLE4 proteins, which are DNA polymerase e accessory subunits, results in marked hypersensitivity to ATR inhibition. We anticipate that this 117-gene set will be useful for the identification of genes involved in the regulation of genome integrity, the characterization of new biological processes involving ATR, and may reveal biomarkers of ATR inhibitor response in the clinic.

molecular biology↗

bioRxiv2017

LARGE, an AMPA receptor interactor, plays a large role in long-term memory formation by driving homeostatic scaling-down

Dynamic trafficking of AMPA-type glutamate receptor (AMPA-R) in neuronal cells is a key cellular mechanism for learning and memory in the brain, which is regulated by AMPA-R interacting proteins. LARGE, a protein associated with intellectual disability, was found to be a novel component of the AMPA-R protein complex in our proteomic study. Here, our functional study of LARGE showed that during homeostatic scaling-down, increased LARGE expression at the Golgi apparatus (Golgi) negatively controlled AMPA-R trafficking from the Golgi to the plasma membrane, leading to downregulated surface and synaptic AMPA-R targeting. In LARGE knockdown mice, long-term potentiation (LTP) was occluded by synaptic AMPA-R overloading, resulting in impaired long-term memory formation. These findings indicate that the fine-tuning of AMPA-R trafficking by LARGE at the Golgi is critical for memory stability in the brain. Our study thus provides novel insights into the pathophysiology of brain disorders associated with intellectual disability.

neuroscience↗