Biology subjects

Lee, J.-A.

Publications and source records attributed to Lee, J.-A..

4 recordsLinked to original sources

bioRxiv2026

Hyperactivation of the AXL-ICD/SIRT2 axis by Amyloid-β impairs astrocytic autophagic flux and exacerbates neuroinflammation

Autophagy dysfunction and neuroinflammation are central to Alzheimer's disease (AD), yet how extracellular amyloid-{beta} (A{beta}) couples to impaired autophagic flux and heightened neuroinflammation remains unknown. Here, we identify the TAM receptor AXL as a molecular transducer that couples A{beta} sensing to the regulation of autophagy and neuroinflammation in astrocytes. A{beta} induces {gamma}-secretase-dependent cleavage of AXL, generating a nuclear intracellular domain (AXL-ICD) that forms phase-separated condensates and activates autophagy gene transcription through SIRT2-mediated recruitment of the RUVBL1/2-INO80 chromatin-remodeling complex. This axis is activated in astrocytes of postmortem AD brains. Concurrently, AXL-ICD binds to the SIRT2 catalytic domain and suppresses its deacetylase activity, increasing -tubulin acetylation and altering microtubule dynamics. While moderate AXL-ICD levels promote autophagic flux, excessive elevation paradoxically triggers microtubule hyperstabilization, thereby impairing autophagosome-lysosome fusion and causing pathological accumulation of autophagosomes and H2O2. The inhibitory peptide AxSBiP disrupts the AXL-ICD/SIRT2 interaction, restores autophagic flux, reduces plaque burden, and normalizes A{beta}-induced H2O2 production and astrogliosis in APP/PS1 mice. We propose the AXL-ICD/SIRT2 axis as an effective therapeutic target to reduce A{beta} burden and neuroinflammation in AD

neuroscience↗

bioRxiv2026

FORGE-KI: A Modular Framework for Endogenous Knock-In Engineering Across HDR and PITCh/MMEJ Repair Pathways

Targeted knock-in technologies have enabled precise insertion of reporters, affinity tags, degrons, and other functional payloads into endogenous genomic loci. Over the past decade, a diverse collection of genome engineering strategies has emerged, including approaches based on homology-directed repair (HDR), microhomology-mediated end joining (MMEJ), homology-mediated end joining (HMEJ), and related methodologies. While these advances have greatly expanded the capabilities of endogenous genome engineering, they have also increased the complexity of donor design, assembly, and validation. Here, we describe FORGE-KI (Functional Oncology Research Genetic Engineering - Knock in), a pathway-matched design workflow for endogenous knock-in engineering that aligns the assembly strategy with the underlying repair mechanism. For large-cargo insertions, we use a modular five-component framework that separates gene-specific targeting arms from reusable functional modules, allowing rapid assembly of HDR donor constructs targeting AHR, IRF1, and FOSL1 from a shared reagent collection. For MMEJ/PITCh applications, where short targeting elements permit rapid fabrication, we developed a streamlined one-step pipeline in which the entire donor and selection payload is synthesized as a single continuous fragment for direct cloning, compressing the design-to-reagent cycle time. This MMEJ workflow is paired with a dual-promoter nuclease vector (pForge-KI-MMEJ-Cas9-DualGuide) that drives the PITCh-release and locus-specific guides from distinct promoters, a design intended to reduce the repeated-promoter instability associated with some dual-guide vectors. We also established a standardized workflow for donor assembly, generation of knock-in cell populations, molecular validation, and selectable-cassette removal, and we demonstrate it by generating a functional, selection-marker-free, cytokine-inducible IRF1 HDR reporter line and an inducible IRF1 PITCh/MMEJ reporter pool with confirmed junction enrichment. In parallel, we developed forgeKI, an R package that automates C-terminal reporter knock-in design across both HDR and PITCh/MMEJ repair pathways, including guide selection, target-biology validation, targeting-arm design, domestication, donor-assembly planning, and generation of synthesis-ready constructs. Together, the reagents and software provide a practical system for endogenous knock-in engineering that supports multiple payloads, selection strategies, and repair pathways within a shared donor organization. Rather than replacing existing knock-in technologies, this framework provides a modular foundation for incorporating, extending, and automating the published knock-in methods.

molecular biology↗

bioRxiv2025

Targeted degradation of pathogenic TDP-43 proteins in amyotrophic lateral sclerosis using the AUTOTAC platform

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease characterized by the progressive loss of motor neurons and the cytoplasmic aggregation of misfolded proteins in the spinal cord, including TAR DNA-binding protein-43 (TDP-43). More than 97% of ALS cases exhibit pathological TDP-43 inclusions, yet therapeutic strategies that can selectively eliminate these aggregates remain yet to be developed. Here, we employed the AUTOTAC (Autophagy-Targeting Chimera) to degrade TDP-43 aggregates via macroautophagy mediated by the N-recognin p62/SQSTM1 of the N-degron pathway. The AUTOTAC degraders ATC141 and ATC142 were designed to bind and link the oligomeric species of misfolded TDP-43 to p62, which induces the targeting of TDP-43 cargoes to phagophores for lysosomal co-degradation, while sparing monomeric TDP-43. ATC142 induced the degradation of pathological TDP-43 A315T species and its cleaved variant, TDP-25, with DC50 values of 1.25-9.6 nM. In ALS model mice expressing TDP-43 A315T in the spinal cord, oral administration of 10 mg/kg ATC141 with 24 doses reduced TDP-43 aggregates as well as GFAP+ astrocytes and Iba1+ microglia. ATC141 also exerted disease-modifying efficacy to reverse the disease progression in neuromuscular coordination and cognitive function. This oral drug is under Phase 1 clinical trials in South Korea with 76 healthy volunteers aiming to treat ALS, Alzheimers diseases (AD), and progressive supranuclear palsy (PSP). We suggest that AUTOTAC provides a novel strategy to treat a broad range of neurodegenerative diseases. TeaserAUTOTAC degraders induce lysosomal degradation of pathogenic TDP-43 aggregates in a mouse model of amyotrophic lateral sclerosis.

neuroscience↗

bioRxiv2021

Modeling lung cell development using human pluripotent stem cells

Human pluripotent stem cells (hPSC) differentiations can capture developmental phenotypes and processes. They are useful for studying fundamental biological mechanisms driving tissue morphogenesis and cell lineage development. Here, we show temporal development of lung cell lineages using hPSC that recapitulate developmental milestones observed in primary tissue, the generation of renewable fetal lung epithelial spheroids, and the functional utility of the lung models at different differentiation stages for cystic fibrosis disease modeling. We first show the presence of hPSC-derived lung progenitor cells reminiscent of early trimester lung development and containing basal stem cells that generate renewable airway spheroids. Maturation and polarization in air liquid interface (ALI) generates additional epithelial cell lineages found in adult airways, including pulmonary neuroendocrine, brush, mature basal, ciliated and secretory cell types. Finally, pseudotime and RNA velocity analyses of the integrated datasets from fetal and ALI stages reveal both previously identified and new cell lineage relationships. Overall, hPSC differentiation can capture aspects of human lung development and potentially provide important insight into congenital causes of diseases.

developmental biology↗