Biology subjects

Cha, H.-J.

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

15 recordsLinked to original sources

bioRxiv2026

FateDriver: Weakly supervised dual-view graph learning for transition-specific prioritization of candidate regulators in cellular reprogramming

Cellular reprogramming supports regenerative medicine, yet the endogenous regulators of a specified source-to-target transition remain incompletely defined. Existing computational methods rank candidate regulators from network topology, trajectory dynamics, or cell-state labels, leaving unused the factors experimentally delivered to induce the transition. We present FateDriver, a weakly supervised dual-view graph neural network that jointly represents cells and genes from single-cell transcriptomes, treats the delivered factors as sparse gene-level labels, and ranks the remaining genes for a specified transition under a positive--unlabeled formulation. Across two OSKM datasets and one GMT dataset, FateDriver ranked first or second among six methods in recovering externally curated reference genes, exceeded a supervision-matched non-graph model despite weaker co-expression with the delivered factors, and achieved the highest recovery of protein targets of pluripotency-inducing chemical cocktails in both OSKM datasets. Holding either the source or the endpoint fixed yielded distinct candidate programs, including a DDX58--interferon axis associated with cells lacking endogenous pluripotency markers. Applying the same framework to candidate source--target pairs prioritized distinct candidate origins for two in vivo partially reprogrammed intestinal populations. FateDriver converts sparse experimental knowledge into transition-specific hypotheses for regulator prioritization from single-cell trajectories.

bioinformatics↗

bioRxiv2026

Dysregulation of Hippo Signaling Pathway as a Convergent Mechanism Underlying Choroid Plexus Defects in Bipolar Disorder

Bipolar disorder (BD) is a prevalent and highly heritable psychiatric condition. Developmental mechanisms are implicated but the specific molecular origins remain unclear. The choroid plexus (ChP), which regulates cerebrospinal fluid (CSF) and brain homeostasis, has been implicated in BD, but its role is poorly understood. Here, we identify aberrant expansion of ChP in human cortical (hCOs) and ChP organoids (hChPOs) derived from, and in vivo structural magnetic resonance imaging (sMRI) scans of, individuals with BD compared to healthy comparison individuals. Single-cell transcriptomics revealed a lineage bias toward ChP fate in BD hCOs, accompanied by structural enlargement observed in both BD hChPOs and ChP in sMRI scans from individuals with BD. Comprehensive transcriptomic profiling implicates early hyperactivation of the Hippo signaling pathway in neural progenitor cells as a convergent mechanism driving ChP overgrowth. Whole-genome and genome-wide association study analyses highlight recurrent genetic variants in Hippo regulators, including STK4 and YAP1, suggesting genetic priming. BD hChPOs exhibit disrupted epithelial junctions and altered in vitro CSF secretion. These findings position the ChP as a genetically and developmentally predisposed to pathology in BD and highlight organoids as a platform for translational discovery in diagnostics and therapeutics.

developmental biology↗

bioRxiv2025

Enhanced liver regeneration via targeted mRNA delivery for partial in vivo reprogramming

Recent studies suggest that injury-induced dedifferentiation, which leads to the formation of injury-responsive cells, contributes significantly to tissue repair across various organs, including the liver. Utilizing Yamanaka factors (Oct4, Sox2, Klf4, and c-Myc: OSKM) for in vivo partial reprogramming generates injury-responsive cells in the intestine, mirroring those derived from injury-induced dedifferentiation. Thus, the transgene induction of OSKM or viral delivery of Oct4, Sox2, and Klf4 shows promise in facilitating tissue regeneration in the intestine, liver, skeletal muscle, and retina. Herein, we demonstrated that transient OSKM induction produces two distinct liver progenitor-like cell populations. One of these populations resembles liver progenitor-like cells (LPLCs) generated by acute acetaminophen (APAP) injury without triggering immune responses. To explore in vivo reprogramming as a viable strategy for tissue regeneration, we employed lipid nanoparticles (LNP) carrying OSKM mRNA (OSKM mRNA-LNP) to stimulate LPLCs formation. Notably, the production of Sox9+ LPLCs, and OSKM-induced dedifferentiation, was closely correlated with successful tissue regeneration in the liver post APAP injury. Thus, the OSKM mRNA-LNP approach represents a promising therapeutic intervention for the repair of acute liver injuries.

physiology↗

bioRxiv2025

Mutation-Agnostic Base Editing of the Progerin Farnesylation Site Rescues Hutchinson-Gilford Progeria Syndrome Phenotypes in Neuromuscular Organoids

Hutchinson Gilford progeria syndrome (HGPS) is a fatal premature aging disorder caused by pathogenic farnesylated lamin A variants that disrupt nuclear architecture and DNA repair. Current therapies, including farnesyltransferase inhibitors, provide only modest survival benefits and lack molecular specificity, while mutation-specific genome-editing strategies cannot address atypical laminopathies. Here, we develop Farnesylation Amino acid Targeted Editing (FATE), a mutation-agnostic base-editing platform that selectively disrupts the LMNA farnesylation motif without affecting other farnesylated proteins. Using isogenic human pluripotent stem cell derived neuromuscular organoids (NMOs), we reveal muscle-specific progerin accumulation that sequesters 53BP1 and abolishes DNA damage foci formation. FATE eliminates perinuclear progerin, restores 53BP1 mobility, reconstitutes DNA repair foci, and normalizes heterochromatin architecture. Importantly, transient delivery of FATE mRNA conjugated with lipid nanoparticles to HGPS NMOs achieves efficient base editing and phenotypic rescue. These findings establish FATE as a mutation-independent therapeutic strategy targeting a fundamental pathogenic mechanism in HGPS and provide a proof-of-concept for RNA-based in situ genome editing in progeroid disease.

pathology↗

bioRxiv2025

N-Terminal Deleted Isoforms of E3 Ligase RNF220 (Isoform 4) Are Ubiquitously Expressed and Required for Mouse Muscle Differentiation.

Four isoform peptides of the novel E3 ligase RNF220 have been identified in humans. However, all of previous studies have predominantly focused on isoform 1, which consists of 566 amino acids (aa). Here, we show that a shorter isoform, isoform 4 (308 aa), lacking most of the N-terminus, is the predominant and ubiquitously expressed variant that warrants functional investigation. Both isoform 1 and isoform 4 are expressed in the brain; however, isoform 4 is the major isoform expressed in all other tissues in mice. Consistently, H3K4me3 ChIP-seq data from ENCODE reveal that the transcription start site for isoform 4 demonstrates broader and stronger activity across human tissues than that of isoform 1. Isoform 4 produces two peptides (4a and 4b) through alternative translation initiation, with isoform 4b displaying distinct subcellular localization and subnuclear structures. Notably, during embryonic stem cell differentiation into neural stem cells, isoform 1 expression increases, whereas isoform 4 expression decreases. In murine myoblasts, isoform 4 is the sole expressed isoform and is required for MyoD and myogenin expression, as well as for muscle differentiation. Our findings highlight isoform 4 as the ubiquitously and highly expressed variant, likely playing a fundamental role across tissues while exhibiting functional differences from isoform 1. These results emphasize the critical importance of isoform 4 in future studies investigating the biological functions of RNF220.

molecular biology↗

bioRxiv2023

Sequential genetic, epigenetic alterations in human pluripotent stem cells for recurrent abnormality

Human embryonic stem cells (hESCs) are naturally equipped to maintain genome integrity to minimize genetic mutations during early embryo development. However, genetic aberration risks and subsequent cellular changes in hESCs during in vitro culture pose a significant threat to stem cell therapy. While a few studies have reported specific somatic mutations and copy number variations (CNVs), the molecular mechanisms underlying culture-adapted phenotype acquisitions of hESCs are largely unknown. Therefore, we conducted comprehensive genomic, single-cell transcriptomic, and single-cell ATAC-seq analyses of an isogenic hESC model displaying definitive culture-adapted phenotypes. Notably, hPSCs with a copy number gain of 20q11.21 during early passage did not present culture-adapted phenotypes nor BCL2L1 induction. Single-cell RNAseq and ATACseq analyses revealed active transcriptional regulation at 20q11.21 loci at late-passaged hESCs with the induced BCL2L1 and TPX2 to trigger culture-adapted phenotypes was associated with epigenetic changes facilitating TEA domain (TEAD) binding. These results suggest that copy number 20q11.21 gain and additional epigenetic changes are necessary for expressing culture-adapted phenotypes by activating gene transcription at this specific locus.

cell biology↗

bioRxiv2023

Ampk activation by glycogen expenditure primes the exit of naive pluripotency

Embryonic and epiblast stem cells in pre-and post-implantation embryos are characterized by their naive and primed states, respectively, which represent distinct phases of pluripotency. Thus, the cellular transition from naive to primed pluripotency recapitulates a drastic metabolic and cellular remodeling after implantation to adapt to changes in extracellular conditions. Here, we found that inhibition of Ampk occurred during naive transition with two conventional inhibitors (2i) of the Mek1 and Gsk3{beta} pathways. The accumulation of glycogen due to the inhibition of Gsk3{beta} was responsible for Ampk inhibition, which accounted for high de novo fatty acid synthesis in naive embryonic stem cells (ESCs). The knockout of glycogen synthase 1 (Gys1) in naive ESCs (GKO), resulting in a drastic glycogen loss, led to a robust Ampk activation and lowered the level of fatty acids. GKO lost the cellular characteristics of naive ESCs and rapidly transitioned to a primed state, as evidenced by a decrease in pluripotency markers in teratoma from GKO. The characteristics of GKO were restored by the simultaneous knockout of Ampk. These findings suggest that glycogen in naive ESCs within the blastocyst may act as a signaling molecule for the timely activation of Ampk, thus ultimately contributing to the transition to the epiblast stage. Graphical Summary O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=108 SRC="FIGDIR/small/541467v1_ufig1.gif" ALT="Figure 1"> View larger version (30K): org.highwire.dtl.DTLVardef@118ea81org.highwire.dtl.DTLVardef@cbe4bforg.highwire.dtl.DTLVardef@16d5c1org.highwire.dtl.DTLVardef@12b825c_HPS_FORMAT_FIGEXP M_FIG C_FIG

developmental biology↗

bioRxiv2023

Partial in vivo reprogramming enables injury-free intestinal regeneration via autonomous Ptgs1 induction

Tissue regeneration after injury involves the dedifferentiation of somatic cells, a natural adaptive reprogramming process that leads to the emergence of injury-responsive cells with fetal-like characteristics in the intestinal epithelium. However, there is no direct evidence that adaptive reprogramming involves a shared molecular mechanism with direct cellular reprogramming. Here, we induced dedifferentiation of intestinal epithelial cells through forced partial reprogramming in vivo using "Yamanaka factors" (Oct4, Sox2, Klf4, and c-Myc: OSKM). The OSKM-induced dedifferentiation showed similar molecular features of intestinal regeneration, including a rapid transition from homeostatic cell types to injury-responsive-like cell types. These injury-responsive-like cells, sharing a gene signature of revival stem cells and atrophy-induced villus epithelial cells, actively assisted tissue regeneration following ionizing radiation-induced acute tissue damage. In contrast to normal intestinal regeneration, which involves epi-mesenchymal crosstalk through induction of Ptgs2 (encoding Cox2) upon injury, the OSKM expression promotes the autonomous production of prostaglandin E2 via epithelial Ptgs1 (encoding Cox1) expression. These results indicate that prostaglandin synthesis is a common mechanism for intestine epithelial regeneration but involves a different enzyme (Ptgs1 for Cox1) when partial reprogramming is directly applied to the intestinal epithelium.

cell biology↗

bioRxiv2022

Identification of a novel HASPIN inhibitor and its synergism with the PLK1 inhibitor

BackgroundHASPIN, a mitotic kinase for Histone H3, is a promising target for anti-cancer therapy. However, as HASPIN is an atypical kinase with low similarity to eukaryotic protein kinases, development of a HASPIN inhibitor from the conventional pharmacophore of kinase inhibitors would be technically challenging. MethodsChemical modifications of a cytotoxic 4-thioadenosine analogue with high genotoxicity and multiple kinomescan profiles were performed to produce a novel non-genotoxic kinase inhibitor, LJ4827. The mode of action of this inhibitor with clear anti-cancer activity was inferred based on transcriptomic and chemical similarity to known drugs. ResultsThe specificity and potency of LJ4827 as a HASPIN inhibitor were validated by in vitro kinase screening and subsequent X-ray crystallography. As predicted, LJ4827 treatment delayed mitosis by clear inhibition of the recruitment of Aurora B at the centromere in cancer cells, without a genotoxic response. Through transcriptome analysis of lung cancer patients, PLK1 was predicted as a druggable synergistic partner to complement HASPIN inhibition. Cotreatment with the PLK1 inhibitor BI2536 and LJ4827 led to pronounced cytotoxicity of lung cancers in vitro and in vivo. ConclusionSimultaneous inhibition of both HASPIN and PLK1 is a promising therapeutic strategy for lung cancers.

pharmacology and toxicology↗

bioRxiv2022

MSH2 and MSH6 as size dependent cellular determinants for prime editing in human embryonic stem cells

Potential applications of precise genome editing in human pluripotent stem cells (hPSCs), not only for isogenic disease modeling but also for ex vivo stem cell therapy, have urged the application of diverse genome editing tools in hPSCs. However, unlike differentiated somatic cells, the unique cellular properties of hPSCs (e.g., high susceptibility to DNA damage and active DNA repair) largely determine the overall efficiency of editing tools. Considering high demand of prime editors (PE), mostly due to its broad editing coverage compared to base editors, it is important to characterize the key molecular determinants of PE efficiency in hPSCs. Herein, we showed that MSH2 and MSH6, two main components of the MutS complex of mismatch repair (MMR), are highly expressed in hPSCs and determine PE efficiency in an editing size-dependent manner. Importantly, loss of MSH2, which disrupts both MutS and MutS{beta} complexes, was found to dramatically improve the efficiency of PE from one base to 10 bases, up to 50 folds. In contrast, genetic perturbation of MSH6, which solely abrogates MutS activity, marginally improved the editing efficiency up to 3 base pairs. The size dependent effect of MSH2 or MSH6 on prime editing in hPSCs not only implies MMR is a major determinant of PE efficiency in hPSCs but also highlights the distinct roles of MutS and MutS{beta} in the outcome of genome editing.

molecular biology↗

bioRxiv2021

TPX2 Amplification-Driven Aberrant Mitosis in Long-Term Cultured Human Embryonic Stem Cells

Although human embryonic stem cells (hESCs) are equipped with highly effective machinery for the maintenance of genome integrity, the frequency of genetic aberrations during long-term in vitro hESC culture has been a serious issue that raises concerns over their safety in future clinical applications. By passaging hESCs over a broad range of timepoints, we found that mitotic aberrations, such as the delay of mitosis, multipolar centrosomes, and chromosome mis-segregation, were increased in the late-passaged hESCs (LP-hESCs) in parallel with polyploidy compared to early-passaged hESCs (EP-hESCs). Through high-resolution genome-wide approaches and by following transcriptome analysis, we found that LP-hESCs with a minimal amplicon in chromosome 20q11.21 highly expressed TPX2 (targeting protein for Xklp2), a key protein for governing spindle assembly and cancer malignancy. Consistent with these findings, the inducible expression of TPX2 in EP-hESCs reproduced aberrant mitotic events, such as the delay of mitotic progression, spindle stability, misaligned chromosomes, and polyploidy. This data suggests that the amplification and increased transcription of the TPX2 gene at 20q11.21 could contribute to an increase in aberrant mitosis due to altered spindle dynamics.

cell biology↗

bioRxiv2020

Efficient GNE myopathy disease modeling with mutation specific phenotypes in human pluripotent stem cells by base editors

Despite the great potential of disease modeling with the isogenic pairs of human pluripotent stem cells (hPSCs), the extremely low efficiency of precise gene editing in hPSCs remains a technical hurdle for this approach. Herein, we took advantage of currently available base editors (BEs) to epitomize the isogenic disease model from hPSCs. Using this method, we established 14 hPSCs that harbor point mutations on the GNE gene, including four different mutations found in GNE myopathy patients. Due to lesser activation of p53 by BEs than Cas9, a higher editing efficiency with BEs was achieved. Four different mutations in the epimerase or kinase domains of GNE revealed mutation-specific hyposialylation, which was closely correlated to pathological clinical phenotypes. These mutation-specific hyposialylation patterns were evident in GNE protein structure modeling. Furthermore, treatment with a drug candidate currently under clinical trials showed a mutation-specific drug response in GNE myopathy disease models. These data suggest that isogenic disease models from hPSCs using BEs could serve as a useful tool for mimicking the pathophysiology of GNE myopathy and for predicting drug responses.

developmental biology↗

bioRxiv2020

Systematic identification of a nuclear receptor-enriched predictive signature for erastin-induced ferroptosis

Erastin, which has been initially identified as a synthetic lethal compound against cancer expressing an RAS oncogene, inhibits cystine/glutamate antiporters and causes ferroptic cell death in various cell types, including therapy-resistant mesenchymal cancer cells. However, despite recent emerging evidence for the mechanisms underlying ferroptosis, molecular biomarkers associated with erastin-dependent ferroptosis have not yet been identified. In the present study, we employed isogenic lung cancer cell models with therapy-resistant mesenchymal properties to show that a redox imbalance leads to glutathione depletion and ferroptotic cell death. Subsequent gene expression analysis of pan-cancer cell lines revealed that the activity of transcription factors, including nuclear factor erythroid 2-related factor 2 (NRF2) and aryl hydrocarbon receptor (AhR), serve as important markers of erastin resistance. Based on the integrated expression of genes in the nuclear receptor meta-pathway (NRM), we constructed an NRM model and validated its robustness using an independent pharmacogenomics dataset. The NRM model was further evaluated by employing it in the sensitivity testing of nine cancer cell lines for which erastin sensitivities had not yet been undetermined. Our pharmacogenomics approach has the potential to pave the way for the efficient classification of patients for therapeutic intervention using erastin or erastin analogs.

systems biology↗

bioRxiv2019

Scarless Enriched selection of Genome edited Human Pluripotent Stem Cells Using Induced Drug Resistance

An efficient gene editing technique for use in human pluripotent stem cells (hPSCs) would have great potential value in regenerative medicine, as well as in drug discovery based on isogenic human disease models. However, the extremely low efficiency of gene editing in hPSCs is a major technical hurdle that remains to be resolved. Previously, we demonstrated that YM155, a survivin inhibitor developed as an anti-cancer drug, induces highly selective cell death in undifferentiated hPSCs. In this study, we demonstrated that the high cytotoxicity of YM155 in hPSCs, which is mediated by selective cellular uptake of the drug, is due to high expression of SLC35F2 in these cells. Consistent with this, knockout of SLC35F2 with CRISPR-Cas9 or depletion with siRNAs made hPSCs highly resistant to YM155. Simultaneous gene editing of a gene of interest and transient knockdown of SLC35F2 following YM155 treatment enabled genome-edited hPSCs to survive because YM155 resistance was temporarily induced, thereby achieving enriched selection of genome-edited clonal populations. This precise and efficient genome editing approach took as little as 3 weeks without cell sorting or introduction of additional genes.

developmental biology↗

bioRxiv2018

In silico Drug Repositioning of bortezomib to reverse metastatic effect of GALNT14 in lung cancer

Although many molecular targets for cancer therapy have been discovered, they often show poor druggability, which is a major obstacle to develop targeted drugs. As an alternative route to drug discovery, we adopted an in silico drug repositioning (in silico DR) approach based on large-scale gene expression signatures, with the goal of identifying inhibitors of lung cancer metastasis. Our analysis of clinicogenomic data identified GALNT14, an enzyme involved in O-linked N-acetyl galactosamine glycosylation, as a putative driver of lung cancer metastasis leading to poor survival. To overcome the poor druggability of GALNT14, we leveraged Connectivity Map approach, an in silico screening for drugs that are likely to revert the metastatic expression patterns. It leads to identification of bortezomib (BTZ) as a potent metastatic inhibitor, bypassing direct inhibition of poorly druggable target, GALNT14. The anti-metastatic effect of BTZ was verified in vitro and in vivo. Notably, both BTZ treatment and GALNT14 knockdown attenuated TGF{beta}-mediated gene expression and suppressed TGF{beta}-dependent metastatic genes, suggesting that BTZ acts by modulating TGF{beta} signalingTaken together, these results demonstrate that our in silico DR approach is a viable strategy to identify a candidate drug for undruggable targets, and to uncover its underlying mechanisms.

cancer biology↗