Biology subjects

Lee, K. Y.

Publications and source records attributed to Lee, K. Y..

15 recordsLinked to original sources

bioRxiv2026

Exercise engages a mechanically activated astrocyte state linking muscle activity to hippocampal plasticity

Physical exercise promotes brain health in part through muscle-derived factors that enter the brain, but how peripheral signals are translated into neural responses remain unclear. Here, we identify astrocyte contraction as a previously unrecognized physiological response to exercise signals that may contribute to adult hippocampal neurogenesis. In vivo, voluntary running rapidly induced nuclear localization of the mechanically sensitive transcriptional regulator Yes-associated protein (YAP), and increased non-muscle myosin II phosphorylation in hilar astrocytes in mice, consistent with acute contraction. Using an in vitro platform with ultrasensitive force sensors, we found that factors released by contracting skeletal muscles activated astrocytes which in turn increased contraction that was necessary and sufficient for their proliferation and expansion. Activated astrocytes subsequently released soluble factors that modulated neuronal network tension and promoted immature neuron abundance. These findings identify astrocyte contractility as a physiological transducer of exercise-derived muscle signals and establish cellular force generation as a potential mechanism regulating neuroplasticity.

neuroscience↗

bioRxiv2026

Reversing Pathophysiology in Fragile X Syndrome Mice by Promoting PGC-1α and Mitochondrial Functions

Fragile X syndrome (FXS) is the leading cause of intellectual disabilities and autism, but a disease-modifying strategy remains unavailable. Recent studies have suggested reduced mitochondrial functions in FXS. However, the mechanisms underlying mitochondrial defects and their impact on FXS pathophysiology remain largely unclear. Here, we reveal a reduction in the mitochondrial master regulator peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1) in the mouse model of FXS, the Fmr1 knockout (KO) mice. We show that this impairment is caused by the inactivity of the transcription factor cAMP-response element-binding protein (CREB) in Fmr1 KO mice. Using the small molecule ZLN005, which induces AMP-activated protein kinase (AMPK)- and CREB-dependent elevation of PGC-1 in Fmr1 KO mice, we observed significantly increased mitochondrial functions and dynamics in cultured neurons in vitro and in the hippocampus in vivo. Furthermore, ZLN005 elicited a wide range of beneficial effects in Fmr1 KO mice, including enhanced inhibitory synaptic transmission, reduced circuit hyperexcitability, improved hippocampal synaptic plasticity, reduced cortical gamma-band oscillations, and improved interhemispheric coherence. Most importantly, we observed improved cognition and reduced autism-like behaviors in ZLN005-treated Fmr1 KO mice. Together, our findings identify AMPK-CREB signaling and PGC-1 as promising and selective therapeutic targets for FXS and reveal the broad impact of restoring PGC-1 on FXS pathophysiology. One Sentence SummaryPromoting PGC-1 Reverses FXS Pathophysiology.

neuroscience↗

bioRxiv2025

Tension shapes memory: Computational insights into neural plasticity

Mechanical forces have recently emerged as critical modulators of neural communication, yet their role in high-level cognitive functions remains poorly understood. Here, we present a biologically inspired spiking neural network model that integrates mechanical tension, vesicle dynamics, and spike-timing-dependent plasticity to examine how tension influences learning, memory, and cognitive operations such as pattern completion, projection, and association. We find that increased tension enhances synaptic efficiency by accelerating vesicle clustering and recovery, resulting in a 67% improvement in memory recall speed and a 17% increase in inter-regional synchrony during projection relative to relaxed states. Conversely, a 20% reduction in tension leads to a 31% decline in memory association performance, highlighting the tension-sensitive accessibility of stored information. The model further reveals that networks with 20% inhibitory neurons achieve optimal spatial precision in memory encoding and recall. Together, these in silico findings position mechanical tension as a functional neuromodulator and suggest new directions for neuromorphic design and energy-efficient, living computing platforms. Author summaryOur brains are not only electrical and chemical systems but also mechanical ones. Neurons and their connections are constantly under tension, yet the role of these physical forces in shaping memory and thought remains poorly understood. In this work, we built a computer model of a brain-like network that allowed us to test how mechanical tension influences learning and memory. We found that higher tension improved the speed and accuracy of memory recall, strengthened communication between groups of neurons, and supported more reliable association of memories. In contrast, reducing tension made memories harder to access, though they could be recovered once tension was restored. We also discovered that having the right balance between excitatory and inhibitory neurons was crucial for both storing and recalling information. Our results suggest that tension is not just a background property of brain tissue but an active player in cognition, with potential implications for understanding memory disorders and designing new brain-inspired technologies.

neuroscience↗

bioRxiv2024

High-Complexity Barcoded Rabies Virus for Scalable Circuit Mapping Using Single-Cell and Single-Nucleus Sequencing

Single cell genomics has revolutionized our understanding of neuronal cell types. However, scalable technologies for probing single-cell connectivity are lacking, and we are just beginning to understand how molecularly defined cell types are organized into functional circuits. Here, we describe a protocol to generate high-complexity barcoded rabies virus (RV) for scalable circuit mapping from tens of thousands of individual starter cells in parallel. In addition, we introduce a strategy for targeting RV-encoded barcode transcripts to the nucleus so that they can be read out using single-nucleus RNA sequencing (snRNA-seq). We apply this tool in organotypic slice cultures of the developing human cerebral cortex, which reveals the emergence of cell type- specific circuit motifs in midgestation. By leveraging the power and throughput of single cell genomics for mapping synaptic connectivity, we chart a path forward for scalable circuit mapping of molecularly-defined cell types in healthy and disease states.

neuroscience↗

bioRxiv2024

The Discovery of Small Molecule Inhibitors of cFLIP that Sensitise Tumour Cells to TRAIL

The TNF-related apoptosis-inducing ligand (TRAIL) has potential as a therapeutic agent as it has previously been shown to induce apoptosis in triple-negative breast cancer. Recombinant human TRAIL has shown promise in pre-clinical studies of breast cancer. TRAIL exhibits specificity for triple-negative and treatment-resistant disease subsets. However, several studies have demonstrated that patient tumours exhibit resistance to TRAIL and TRAIL-receptor agonists. We have previously demonstrated that suppression of the TRAIL-receptor inhibitor cFLIP can sensitise breast cancer stem cells to apoptosis inducers, but development of pharmacological inhibitors of cFLIP have been impeded by concerns over structural similarities between cFLIP and the pro-apoptotic procaspase-8. We used molecular dynamics to model the interactions between cFLIP, procaspase-8 and the TRAIL-receptor Death Inducing Signalling Complex (TRAIL-DISC), followed by virtual pharmacophore screening and in-cell viability assays to identify a small-molecule (OH14, 3) that selectively inhibited cFLIP binding to the DISC and promoted TRAIL-mediated apoptosis in breast cancer cell lines. When used in combination with TRAIL, OH14 significantly impaired breast cancer cell viability in primary derived and established cell culture. Given the relatively low (micromolar) potency of the initial hit compound inhibitor OH14 (3), limiting its utility as a preclinical development candidate, we carried out structure-activity relationship studies to find a cFLIP inhibitor with more potent cellular activity. Our findings confirm the proof-of-principle that selective pharmacological inhibition of cFLIP can be used to target a vulnerability in breast cancer cells.

cancer biology↗

bioRxiv2023

Synapses without tension fail to fire in an in vitro network of hippocampal neurons.

Neurons in the brain communicate with each other at their synapses. It has long been understood that this communication occurs through biochemical processes. Here, we reveal a previously unrecognized paradigm wherein mechanical tension in neurons is essential for communication. Using in vitro rat hippocampal neurons, we find that (1) neurons become tout/tensed after forming synapses resulting in a contractile neural network, and (2) without this contractility, neurons fail to fire. To measure time evolution of network contractility in 3D (not 2D) extracellular matrix, we developed an ultra-sensitive force sensor with 1 nN resolution. We employed Multi-Electrode Array (MEA) and iGluSnFR, a glutamate sensor, to quantify neuronal firing at the network and at the single synapse scale, respectively. When neuron contractility is relaxed, both techniques show significantly reduced firing. Firing resumes when contractility is restored. Neural contractility may play a crucial role in memory, learning, cognition, and various neuropathologies.

neuroscience↗

bioRxiv2023

An isoform-resolution transcriptomic atlas of colorectal cancer from long-read single-cell sequencing

Colorectal cancer (CRC) is the second leading cause of cancer death worldwide. In recent years, short-read single-cell RNA sequencing (scRNA-seq) has been instrumental in deciphering tumor cell heterogeneities. However, these studies only enable gene-level expression quantification but neglect alterations in transcript structures, which arise from alternative end processing or splicing, and are frequently observed in cancer. In this study, we integrated short- and long-read scRNA-seq of CRC patient samples to build the first isoform-resolution CRC transcriptomic atlas. We identified 394 dysregulated transcript structures in tumor epithelial cells, including 299 resulting from various combinations of multiple splicing events. Secondly, we characterized genes and isoforms associated with epithelial lineages and subpopulations that exhibit distinct prognoses. Finally, we built an algorithm that integrated novel peptides derived from predicted ORFs of recurrent tumor-specific transcripts with mass spectrometry data and identified a panel of recurring neoepitopes that may aid the development of neoantigen-based cancer vaccines.

cancer biology↗

bioRxiv2022

Astrocyte-mediated transduction of muscle fiber contractions synchronizes hippocampal neuronal network development

Exercise supports brain health in part through enhancing hippocampal function. The leading hypothesis is that muscles release factors when they contract (e.g., lactate, myokines, growth factors) that enter circulation and reach the brain where they enhance plasticity (e.g., increase neurogenesis and synaptogenesis). However, it remains unknown how the muscle signals are transduced by the hippocampal cells to modulate network activity and synaptic development. Thus, we established an in vitro model in which the media from contracting primary muscle cells (CM) is applied to developing primary hippocampal cell cultures on a microelectrode array. We found that the hippocampal neuronal network matures more rapidly (as indicated by synapse development and synchronous neuronal activity) when exposed to CM than regular media (RM). This was accompanied by a 1.4-fold and 4.4-fold increase in the proliferation of neurons and astrocytes, respectively. Further, experiments established that the astrocytes release factors that inhibit neuronal excitability and facilitate network development. Results provide new insight into how exercise may support hippocampal function through regulating astrocyte proliferation and subsequent taming of neuronal activity into an integrated network. HighlightsO_LIContracting muscle conditioned media enhances neuronal activity. C_LIO_LIContracting muscle conditioned media expedites neuronal maturation and accumulation of filamentous actin at presynaptic terminals. C_LIO_LIContracting muscle conditioned media induces significant neuron and astrocyte proliferation. C_LIO_LIAstrocytes release factors that inhibit muscle media-induced neuronal activity. C_LI

neuroscience↗

bioRxiv2021

Structure-Function in Helical Cardiac Musculature Using Additive Textile Manufacturing

For more than fifty years, it has been hypothesized that the helical alignment of the heart gives rise to its mechanical function. Testing this hypothesis in an engineered environment is difficult, as the fine spatial features and complex three-dimensional (3D) structures of the cardiac musculature are challenging to reproduce using current biofabrication techniques. Addressing this, here we report a new form of additive textile manufacturing, Focused Rotary Jet Spinning (FRJS). FRJS allows for the rapid manufacturing of micro/nanofibers with controlled alignments. Using this method, we manufacture 3D models of the left ventricle, showing that helically aligned scaffolds display increased strain uniformity, axial shortening, cardiac output, and ejection fractions as compared to circumferential models. We then demonstrate how FRJS can enable the assembly of a full-sized model of the human hearts musculature. This work experimentally confirms that ventricular alignment plays a critical role in ensuring healthy cardiac performance.

bioengineering↗

bioRxiv2020

Targeted CRISPR screening identifies PRMT5 as synthetic lethality combinatorial target with gemcitabine in pancreatic cancer cells

Pancreatic ductal adenocarcinoma (PDAC) remains one of the most challenging cancer to treat. Due to the asymptomatic nature of the disease and ineffective drug treatment modalities, the survival rate of PDAC patients remains one of the lowest. The recurrent genetic alterations in PDAC are yet to be targeted; therefore, identifying effective therapeutic combinations is desperately needed. Here, we performed an in vivo CRISPR screening in a clinically relevant patient-derived xenograft (PDX) model system to identify synergistic drug combinations for PDAC treatment. Our approach revealed protein arginine methyltransferase gene 5 (PRMT5) as a promising druggable candidate whose inhibition creates synergistic vulnerability of PDAC cells to gemcitabine. Genetic and pharmacological inhibition results indicate that of PRMT5 depletion results in synergistic cytotoxicity with Gem due to depleted replication protein A (RPA) levels and an impaired non-homology end joining (NHEJ) DNA repair. Thus, the novel combination creates conditional lethality through the accumulation of excessive DNA damage and cell death, both in vitro and in vivo. The findings demonstrate that unbiased genetic screenings combined with a clinically relevant model system is an effective approach in identifying synthetic lethal drug combinations for cancer treatment. STATEMENT of SIGNIFICANCEIdentify synergistic drug combinations for PDAC is a significant unmet need. Through CRISPR screening, we discovered and validated that PRMT5 depletion creates synergistic vulnerability of PDAC cells to gemcitabine. Mechanistically, the combination impairs DNA repair, synergistic accumulation of DNA damage and cell death in vitro and in vivo.

cancer biology↗

bioRxiv2019

Excitatory neurons are more disinhibited than inhibitory neurons by chloride dysregulation in the spinal dorsal horn

Neuropathic pain is a debilitating condition caused by the abnormal processing of somatosensory input. Synaptic inhibition in the spinal dorsal horn plays a key role in that processing. Mechanical allodynia - the misperception of light touch as painful - occurs when inhibition is compromised. Disinhibition is due primarily to chloride dysregulation caused by hypofunction of the potassium-chloride co-transporter KCC2. Here we show, in rats, that excitatory neurons are disproportionately affected. This is not because chloride is differentially dysregulated in excitatory and inhibitory neurons, but, rather, because excitatory neurons rely more heavily on inhibition to counterbalance strong excitation. Receptive fields in both cell types have a center-surround organization but disinhibition unmasks more excitatory input to excitatory neurons. Differences in intrinsic excitability also affect how chloride dysregulation affects spiking. These results deepen understanding of how excitation and inhibition are normally balanced in the spinal dorsal horn, and how their imbalance disrupts somatosensory processing.

neuroscience↗

bioRxiv2019

An essential RNA-binding lysine residue in the Nab3 RRM domain undergoes mono and tri-methylation

The Nrd1-Nab3-Sen1 (NNS) complex integrates molecular inputs to direct termination of noncoding transcription in budding yeast. NNS is positively regulated by methylation of histone H3 lysine-4 as well as through Nrd1 binding to the initiating form of RNA PolII. These cues collaborate with Nrd1 and Nab3 binding to target RNA sequences in nascent transcripts through their RRM RNA recognition motifs. In this study, we identify nine lysine residues distributed amongst Nrd1, Nab3, and Sen1 that are mono-, di-, or trimethylated, suggesting novel molecular inputs for NNS regulation. One of these methylated residues, Nab3 lysine-363 (K363), resides within its RRM, and is known to physically contact target RNA. Although mutation of Nab3-K363 to arginine (Nab3-K363R) causes a severe growth defect, it nevertheless produces a stable protein that is incorporated into the NNS complex, suggesting that RNA binding through Nab3-K363 is crucial for NNS function. Consistent with this hypothesis, K363R mutation decreases RNA binding of the Nab3 RRM in vitro and causes transcription termination defects in vivo. These findings reveal crucial roles for Nab3-K363 and suggest that methylation of this residue may modulate NNS activity through its impact on Nab3 RNA binding.

molecular biology↗

bioRxiv2018

Combinatorial Genetic Control of Rpd3S through histone H3K4 and H3K36 Methylation in Budding Yeast

Much of euchromatin regulation occurs through reversible methylation of histone H3 lysine-4 and lysine-36 (H3K4me and H3K36me). Using the budding yeast Saccharomyces cerevisiae, we previously found that levels of H3K4me modulated temperature sensitive alleles of the transcriptional elongation complex Spt6-Spn1 through an unknown H3K4me effector pathway. Here we identify the Rpd3S histone deacetylase complex as the H3K4me effector underlying these Spt6-Spn1 genetic interactions. Exploiting these Spt6-Spn1 genetic interactions, we show that H3K4me and H3K36me collaboratively impact Rpd3S function in an opposing manner. H3K36me is deposited by the histone methyltransferase Set2 and is known to promote Rpd3S function at RNA PolII transcribed open reading frames. Using genetic epistasis experiments, we find that mutations perturbing the Set2-H3K36me-Rpd3S pathway suppress the growth defects caused by temperature sensitive alleles of SPT6 and SPN1, illuminating that this pathway antagonizes Spt6-Spn1. Using these sensitive genetic assays, we also identify a role for H3K4me in antagonizing Rpd3S that functions through the Rpd3S subunit Rco1, which is known to bind H3 N-terminal tails in a manner that is prevented by H3K4me. Further genetic experiments reveal that the H3K4 and H3K36 demethylases JHD2 and RPH1 mediate this combinatorial control of Rpd3S. Finally, our studies also show that the Rpd3L complex, which acts at promoter-proximal regions of PolII transcribed genes, counters Rpd3S for genetic modulation of Spt6-Spn1, and that these two Rpd3 complexes balance the activities of each other. Our findings present the first evidence that H3K4me and H3K36me act combinatorially to control Rpd3S.

genetics↗

bioRxiv2018

H3K4 methylation dependent and independent chromatin regulation by JHD2 and SET1 in budding yeast

Set1 and Jhd2 regulate the methylation state of histone H3 lysine-4 (H3K4me) through their opposing methyltransferase and demethylase activities in the budding yeast Saccharomyces cerevisiae. H3K4me associates with actively transcribed genes and, like both SET1 and JHD2 themselves, is known to regulate gene expression diversely. It remains unclear, however, if Set1 and Jhd2 act solely through H3K4me. Relevantly, Set1 methylates lysine residues in the kinetochore protein Dam1 while genetic studies of the S. pombe SET1 ortholog suggest the existence of non-H3K4 Set1 targets relevant to gene regulation. We interrogated genetic interactions of JHD2 and SET1 with essential genes involved in varied aspects of the transcription cycle. Our findings implicate JHD2 in genetic inhibition of the histone chaperone complexes Spt16-Pob3 (FACT) and Spt6-Spn1. This targeted screen also revealed that JHD2 inhibits the Nrd1-Nab3-Sen1 (NNS) transcription termination complex. We find that while Jhd2s impact on these transcription regulatory complexes likely acts via H3K4me, Set1 governs the roles of FACT and NNS through opposing H3K4-dependent and -independent functions. We also identify diametrically opposing consequences for mutation of H3K4 to alanine or arginine, illuminating that caution must be taken in interpreting histone mutation studies. Unlike FACT and NNS, detailed genetic studies suggest an H3K4me-centric mode of Spt6-Spn1 regulation by JHD2 and SET1. Chromatin immunoprecipitation and transcript quantification experiments show that Jhd2 opposes the positioning of a Spt6-deposited nucleosome near the transcription start site of SER3, a Spt6-Spn1 regulated gene, leading to hyper-induction of SER3. In addition to confirming and extending an emerging role for Jhd2 in the control of nucleosome occupancy near transcription start sites, our findings suggest some of the chromatin regulatory functions of Set1 are independent of H3K4 methylation.

genetics↗

bioRxiv2018

A polymer probe-based system for high density, long-lasting electrophysiological recordings across distributed neuronal circuits

The brain is a massive neuronal network, organized into anatomically distributed sub-circuits, with functionally relevant activity occurring at timescales ranging from milliseconds to months. Current methods to monitor neural activity, however, lack the necessary conjunction of anatomical spatial coverage, temporal resolution, and long-term stability to measure this distributed activity. Here we introduce a large-scale, multi-site recording platform that integrates polymer electrodes with a modular stacking headstage design supporting up to 1024 recording channels in freely behaving rats. This system can support months-long recordings from hundreds of well-isolated units across multiple brain regions. Moreover, these recordings are stable enough to track 25% of single units for over a week. This platform enables large-scale electrophysiological interrogation of the fast dynamics and long-timescale evolution of anatomically distributed circuits, and thereby provides a new tool for understanding brain activity.

neuroscience↗