Biology subjects

Lai, Y.-C.

Publications and source records attributed to Lai, Y.-C..

16 recordsLinked to original sources

bioRxiv2026

Pathway-selective mitophagy regulates retinal physiology and neurogenic transitions in Muller glia

Mitochondrial quality control (MQC) is essential for retinal homeostasis, yet how distinct mitophagy pathways are coordinated within specialized retinal cell types remains poorly understood. Here, we show that Muller glia engage distinct mitophagy programmes that are differentially activated across physiological, metabolic stress, and differentiation contexts. Using pathway-resolved analyses supported by mouse and human single-cell transcriptomic datasets, we demonstrate that PINK1-dependent and receptor-mediated mitophagy pathways coexist within Muller glia and exhibit distinct functional and spatial regulation. To enable precise, time-resolved interrogation of these processes, we developed MQ-MG2, a spontaneously immortalised Muller glial model stably expressing the Mito-QC reporter while preserving endogenous mitophagy adaptors and metabolic features of primary Muller cells. Using this system, we identify context-dependent activation of mitophagy pathways with spatial relevance in vivo and reveal transient coordination of PINK1-dependent and receptor-associated mitophagy during Muller glial neurogenic differentiation. Suppression of fission-dependent mitophagy impaired the acquisition of complex neurite features in MQ-MG2, with a comparable phenotype observed following targeted PINK1 deletion in human neurogenic cells. Together, these findings position Muller glia as active integrators of mitochondrial quality control, capable of engaging distinct mitophagy programmes according to cellular context.

cell biology↗

bioRxiv2025

Deep learning-enabled scaffolding of spatial arrays of PfCSP epitopes

Malaria is a leading cause of disease in developing countries. The licensed malaria vaccine RTS,S/AS01 confers partial protection in part due to the elicitation of circumsporozoite protein (CSP) antibodies, of which those to the CSP repeat and junctional regions offer the most potent protection. Anti-repeat region antibodies, including the protective antibody L9, frequently develop mutations that promote inter-Fab contacts when bound to CSP in "spiral" quaternary structures. As a first step toward the design of immunogens that elicit L9-like antibodies, we utilized generative deep learning models to design epitope-scaffolds that incorporated up to three junctional repeat epitopes with structural conformations and relative spatial orientations matching those of the multivalent complex of CSP bound to three copies of L9. Affinity and structural studies demonstrated accurate scaffolding of two epitopes with the intended relative orientation, and less accurate positioning of the third epitope. This study demonstrates proof of principle for design of multi-epitope scaffolds with pre-determined relative epitope spatial positioning. The study also represents an initial step toward development of multi-epitope immunogens to elicit antibodies that utilize homotypic interactions to bind pathogen in multivalent clusters.

immunology↗

bioRxiv2025

Recombination alters the furin cleavage site in novel bat-borne HKU5-CoV-2 coronavirus

HKU5-CoV-2 is a new bat-infecting coronavirus phylogenetically related to MERS-CoV. It has recently been confirmed that HKU5-CoV-2 can enter human cells and organoids in vitro via the ACE2 receptor, raising concerns about its pandemic potential due to zoonotic spillover. Whether recombination has an influence on HKU5-CoV-2 is completely unclear to date. Here we report the first evidence of HKU5-CoV-2 viral recombination, in association with mutations at the receptor binding domain (RBD) and S1/S2 furin cleavage site (FCS) of the spike protein. Using linkage disequilibrium and haploblock analysis, we identified that 167 recombination hotspots and 27 haploblocks. SNP23016/23043/23064/23156/23193/23285 at the RBD and SNP23833/23847 at the FCS are recombinant hotspots. Our results suggest that recombination may lead to the substitution at RBD residue 498 (Thr498Val/Val498Thr, Thr498Ile/Ile498Thr), which Thr498 directly contacts the ACE2 receptor. Recombination also causes Ser723 deletion/insertion and Ser729Ala substitution at the FCS. These mutations could affect host tropism and change furin cleavage activity. Our results indicate that recombination has played a critical role in HKU5-CoV-2 evolution and infectivity.

microbiology↗

bioRxiv2025

Neuromodulation in neural organoids with shell MEAs

Neural organoids (NOs) have emerged as important tissue engineering models for brain sciences and biocomputing. Establishing reliable relationships between stimulation and recording traces of electrical activity is essential to monitor the functionality of NOs, especially as it relates to realizing biocomputing paradigms such as reinforcement learning or stimulus discrimination. While researchers have demonstrated neuromodulation in NOs, they have primarily used 2D microelectrode arrays (MEAs) with limited access to the entire 3D contour of the NOs. Here, we report neuromodulation using tiny mimics of macroscale EEG caps or shell MEAs. Specifically, we observe that stimulating current within a specific range (20 to 30 {micro}A) induced a statistically significant increase in neuron firing rate when comparing the activity five seconds before and after stimulation. We observed neuromodulatory behavior using both three- and 16-electrode shells and could generate 3D spatiotemporal maps of neuromodulatory activity around the surface of the NO. Our studies demonstrate a methodology for investigating 3D spatiotemporal neuromodulation in organoids of broad relevance to biomedical engineering and biocomputing. One-Sentence SummaryNeuromodulation, an essential intelligence feature, was observed using 3D stimulation and recording from neural organoids.

bioengineering↗

bioRxiv2024

The N-terminus of Plasmodium falciparum Circumsporozoite Protein Contains Three Non-Overlapping Murine B-cell Epitope Regions

The generation of an anti-malarial vaccine that produces broad, potent, and durable responses is highly desirable to control the burden of Plasmodium falciparum disease. Current vaccines have offered modest efficacy ranging from 50%-70%, likely associated with antibody responses that are relatively short lived and strain specific. Currently approved malaria vaccines, RTS,S and R21, target the repeat region and C-terminal region of Plasmodium falciparum CSP, leaving the N-terminal region of CSP neglected as a target for protective immunogen design. Here, we isolate and express a panel of memory B-cell derived N-terminal CSP-specific monoclonal antibodies (mAbs) from mice immunized with an N-terminal CSP specific immunogen. The characterization of N-terminal specific mAbs including peptide walking and affinity experiments indicate that these antibodies target three distinct sites within the N-terminus of CSP. Site ntCSP-A contains the Region I (RI) cleavage site, which has been previously defined, whereas the remaining two sites are in previously undescribed locations upstream of RI, termed ntCSP-B and ntCSP-C.

immunology↗

bioRxiv2023

Effects of growth feedback on gene circuits: A dynamical understanding

The successful integration of engineered gene circuits into host cells remains a significant challenge in synthetic biology due to circuit-host interactions, such as growth feedback, where the circuit influences cell growth and vice versa. Understanding the dynamics of circuit failures and identifying topologies resilient to growth feedback are crucial for both fundamental and applied research. Utilizing transcriptional regulation circuits with adaptation as a paradigm, we systematically study more than four hundred topological structures and uncover various categories of failures. Three dynamical mechanisms of circuit failures are identified: continuous deformation of the response curve, strengthened or induced oscillations, and sudden switching to coexisting attractors. Our extensive computations also uncover a scaling law between a circuit robustness measure and the strength of growth feedback. Despite the negative effects of growth feedback on the majority of circuit topologies, we identify several circuits that maintain optimal performance as designed, a feature important for applications.

synthetic biology↗

bioRxiv2023

Discovering pathways through ribozyme fitness landscapes using information theoretic quantification of epistasis

The identification of catalytic RNAs is typically achieved through primarily experimental means. However, only a small fraction of sequence space can be analyzed even with high-throughput techniques. Methods to extrapolate from a limited data set to predict additional ribozyme sequences, particularly in a human-interpretable fashion, could be useful both for designing new functional RNAs and for generating greater understanding about a ribozyme fitness landscape. Using information theory, we express the effects of epistasis (i.e., deviations from additivity) on a ribozyme. This representation was incorporated into a simple model of the epistatic fitness landscape, which identified potentially exploitable combinations of mutations. We used this model to theoretically predict mutants of high activity for a self-aminoacylating ribozyme, identifying potentially active triple and quadruple mutants beyond the experimental data set of single and double mutants. The predictions were validated experimentally, with nine out of nine sequences being accurately predicted to have high activity. This set of sequences included mutants that form a previously unknown evolutionary bridge between two ribozyme families that share a common motif. Individual steps in the method could be examined, understood, and guided by a human, combining interpretability and performance in a simple model to predict ribozyme sequences by extrapolation.

systems biology↗

bioRxiv2022

The Role of UBE2-Conjugating Enzymes in the Mechanism of MuRF1 Ubiquitylation

MuRF1 (Muscle-specific RING finger protein 1; gene name TRIM63) is a ubiquitin E3 ligase, associated with the progression of muscle atrophy. As a RING (Really Interesting New Gene)-type E3 ligase, its unique activity of ubiquitylation is driven by a specific interaction with UBE2 (ubiquitin conjugating enzyme) to ubiquitylate its substrate protein. The understanding of MuRF1 function remains unclear as candidate UBE2 has not been elucidated and thus the mechanism of ubiquitylation is inconclusive. In the present study, we screened human ubiquitin dependent E2s using in-vitro ubiquitylation assays. We found that MuRF1 engages in ubiquitylation/auto-ubiquitylation with UBE2D, UBE2E, UBE2N/V families and UBE2W. Our result indicated that MuRF1 can cause mono-ubiquitylation, K48, and K63 specific poly-ubiquitin chains in a UBE2-dependent manner. Interestingly, we identified a two-step UBE2-dependent mechanism by which UBE2W allows MuRF1 to mono-ubiquitylate which then acts as an anchor for UBE2N/V and UBE2D generated poly-ubiquitin chain formation. Furthermore, MuRF1 was shown to cooperate with the identified interacting UBE2s to directly ubiquitylate substrates Titin (A168-A170), Desmin, and MYLPF (Myosin Light Chain, Phosphorylatable, Fast Skeletal Muscle; also called Myosin Light Regulatory Chain 2). Our work presents a novel insight into the mechanisms that underpin MuRF1 activity by highlighting the diversity of MuRF1 ubiquitylation enabled by different UBE2s.

molecular biology↗

bioRxiv2022

Arabidopsis histone H3 lysine 9 methyltransferases KYP/SUVH5/6 are involved in leaf development by interacting with AS1-AS2 to repress KNAT1 and KNAT2

Arabidopsis KRYPTONITE/SUPPRESSOR OF VARIEGATION 3-9 HOMOLOG 4 (KYP/SUVH4), SUVH5 and SUVH6 are H3K9 methyltransferases and they are redundantly involved in silencing of transposable elements (TEs). A recent study indicated that KYP/SUVH5/6 can directly interact with the histone deacetylase HDA6 to synergistically regulate TE expression. However, the function of KYP/SUVH5/6 in plant development is still unclear. The ASYMMETRIC LEAVES1 (AS1) and AS2 form a transcription complex, which is involved in leaf development by repressing the homeobox genes KNOTTED-LIKE FROM ARABIDOPSIS THALIANA 1 (KNAT1) and KNAT2. In this study, we found that KYP and SUVH5/6 directly interact with AS1-AS2 to repress KNAT1 and KNAT2 by altering histone H3 acetylation and H3K9 dimethylation levels. In addition, KYP can directly target on the promoters of KNAT1 and KNAT2, and the binding of KYP is dependent on AS1. Furthermore, the genome-wide occupancy profile of KYP indicated that KYP is enriched in the promoter regions of coding genes, and the binding of KYP is positively correlated with that of AS1 and HDA6. Together, these results indicate that Arabidopsis H3K9 methyltransferases KYP/SUVH5/6 are involved in leaf development by interacting with AS1-AS2 to alter histone H3 acetylation and H3K9 dimethylation from the KNAT1 and KNAT2 loci.

plant biology↗

bioRxiv2021

Double-edged role of resource competition in gene expression noise and control

Despite extensive investigation demonstrating that resource competition can significantly alter the circuits deterministic behaviors, a fundamental issue is how resource competition contributes to the gene expression noise and how the noise can be controlled. Utilizing a two-gene circuit as a prototypical system, we uncover a surprising double-edged role of resource competition in gene expression noise: the competition decreases noise through a resource constraint but generates its own type of noise which we name as "resource competitive noise." Utilization of orthogonal resources enables retaining the noise reduction conferred by resource constraint while removing the added resource competitive noise. The noise reduction effects are studied using three negative feedback controller types: negatively competitive regulation (NCR), local, and global controllers, each having four placement architectures in the protein biosynthesis pathway (mRNA or protein inhibition on transcription or translation). Our results show that both local and NCR controllers with mRNA-mediated inhibition are efficacious at reducing noise, with NCR controllers demonstrating a superior noise-reduction capability. We also find that combining negative feedback controllers with orthogonal resources can improve the local controllers. This work provides deep insights into the origin of stochasticity in gene circuits with resource competition and guidance for developing effective noise control strategies.

synthetic biology↗

bioRxiv2021

Ubiquitin E3 ligases Atrogin-1 and MuRF1 protein contents are differentially regulated in the rapamycin-sensitive mTOR-S6K1 signaling pathway in C2C12 myotubes

Muscle-specific ubiquitin E3 ligases, Atrogin-1 and MuRF1, are highly expressed in multiple conditions of skeletal muscle atrophy. The PI3K/Akt/FoxO signaling pathway is well known to regulate Atrogin-1 and MuRF1 gene expressions. Evidence supporting this is largely based on stimuli by insulin and IGF-1, that activate anabolic signaling, including Akt and Akt-dependent transcription factors. However, Akt activation also activates the mammalian target of rapamycin complex 1 (mTORC1) which induces skeletal muscle hypertrophy. However, whether mTORC1-dependent signaling has a role in regulating Atrogin-1 and/or MuRF1 gene and protein expression is currently unclear. In this study, we confirmed that activation of insulin-mediated Akt signaling suppresses both Atrogin-1 and MuRF1 protein content and that inhibition of Akt increases both Atrogin-1 and MuRF1 protein content in C2C12 myotubes. Interestingly, inhibition of mTORC1 using a specific mTORC1 inhibitor, rapamycin, increased Atrogin-1, but not MuRF1, protein content. Furthermore, activation of AMP-activated protein kinase (AMPK), a negative regulator of the mTORC1 signaling pathway, also showed distinct time-dependent changes between Atrogin-1 and MuRF1 protein content, suggesting differential regulatory mechanisms between Atrogin-1 and MuRF1 protein content. To further explore the downstream of mTORC1 signaling, we employed a specific S6K1 inhibitor, PF-4708671, and found that Atrogin-1 protein content was dose-dependently increased with PF-4708671 treatment, whereas MuRF1 protein content was not significantly altered. Overall, our results indicate that Atrogin-1 and MuRF1 protein contents are regulated by different mechanisms, the downstream of Akt, and that Atrogin-1 protein content can be regulated by rapamycin-sensitive mTOR-S6K1 dependent signaling pathway.

cell biology↗

bioRxiv2021

Effects of stochasticity on the length and behaviour of ecological transients

There is a growing recognition that ecological systems can spend extended periods of time far away from an asymptotic state, and that ecological understanding will therefore require a deeper appreciation for how long ecological transients arise. Recent work has defined classes of deterministic mechanisms that can lead to long transients. Given the ubiquity of stochasticity in ecological systems, a similar systematic treatment of transients that includes the influence of stochasticity is important. Stochasticity can of course promote the appearance of transient dynamics by preventing systems from settling permanently near their asymptotic state, but stochasticity also interacts with deterministic features to create qualitatively new dynamics. As such, stochasticity may shorten, extend, or fundamentally change a systems transient dynamics. Here, we describe a general framework that is developing for understanding the range of possible outcomes when random processes impact the dynamics of ecological systems over realistic time scales. We emphasize that we can understand the ways in which stochasticity can either extend or reduce the lifetime of transients by studying the interactions between the stochastic and deterministic processes present, and we summarize both the current state of knowledge and avenues for future advances.

ecology↗

bioRxiv2020

Molecular Survey for Selected Viral Pathogens in Wild Leopard Cats (Prionailurus bengalensis) in Taiwan with an Emphasis on the Spatial and Temporal Dynamics of Carnivore Protoparvovirus 1

The leopard cat (Prionailurus bengalensis) has been listed as an endangered species under the Wildlife Conservation Act in Taiwan since 2009. In this study, we targeted viral pathogens, included carnivore protoparvovirus 1 (CPPV-1), feline leukemia virus (FeLV), feline immunodeficiency virus (FIV), coronavirus (CoV), and canine morbillivirus (CMV), using molecular screening. The spatial and temporal dynamics of the target pathogens were evaluated. Through sequencing and phylogenetic analysis, we aimed to clarify the phylogenetic relationship of isolated viral pathogens between leopard cats and domestic carnivores. Samples from 23 and 29 leopard cats that were live-trapped and found dead, respectively, were collected from Miaoli County from 2015 to 2019 in northwestern Taiwan. CPPV-1 and coronavirus were detected in leopard cats. The prevalence (95% confidence interval) of CPPV-1, and CoV was 63.5% (50.4%-76.6%) and 8.8% (0%-18.4%), respectively. The majority of sequences of each CPPV-1 strain amplified from Taiwanese leopard cats and domestic carnivores were identical. All the amplified CoV sequences from leopard cats were identified as feline coronavirus. The spatial and temporal aggregation of CPPV-1 infection in leopard cats was not determined in the sampling area, which indicated a wide distribution of CPPV-1 in the leopard cat habitat. We consider sympatric domestic carnivores to be the probable primary reservoir for the pathogens identified. We strongly recommend establishing efforts to manage CPPV-1 and FCoV in the leopard cat habitat, with an emphasis on vaccination programs and population control measures for free-roaming dogs and cats. IMPORTANCEThe leopard cat (Prionailurus bengalensis) is an endangered species in Taiwan. The effects of infectious diseases on the wildlife population have increasingly been recognized. In this study, we targeted highly pathogenic viral pathogens in wild cat species, included carnivore protoparvovirus 1 (CPPV-1), feline leukemia virus (FeLV), feline immunodeficiency virus (FIV), coronavirus (CoV), and canine morbillivirus (CMV), using molecular screening. Furthermore, we collected the epidemiological and phylogenetic data to understand the spatial and temporal dynamics of the target pathogens in the wild leopard cat population and identified the possible origin of target pathogens. Based on our study, we consider sympatric domestic carnivores to be the probable primary reservoir for the pathogens identified. Our study provides a deeper understanding related to the distribution of target viral pathogens in the wild leopard cats. The information is essential for leopard cat conservation and pathogen management.

ecology↗

bioRxiv2019

PINK1-dependent phosphorylation of Serine111 within the SF3 motif of Rab GTPases impairs effector interactions and LRRK2 mediated phosphorylation at Threonine72

Loss of function mutations in the PINK1 kinase are causal for autosomal recessive Parkinson disease (PD) whilst gain of function mutations in the LRRK2 kinase cause autosomal dominant PD. PINK1 indirectly regulates the phosphorylation of a subset of Rab GTPases at a conserved Serine111 (Ser111) residue within the SF3 motif. Using genetic code expansion technologies we have produced stoichiometric Ser111-phosphorylated Rab8A revealing impaired interactions with its cognate guanine nucleotide exchange factor (GEF) and GTPase activating protein (GAP). In a screen for Rab8A kinases we identify TAK1 and MST3 kinases that can efficiently phosphorylate the Switch II residue Threonine72 (Thr72) in a similar manner as LRRK2. Strikingly we demonstrate that Ser111 phosphorylation negatively regulates the ability of LRRK2 but not MST3 or TAK1 to phosphorylate Thr72 in vitro and demonstrate an interplay of PINK1- and LRRK2-mediated phosphorylation of Rab8A in cells. Finally, we present the crystal structure of Ser111-phosphorylated Rab8A and NMR structure of Ser111-phosphorylated Rab1B that does not demonstrate any major changes suggesting that the phosphorylated SF3 motif may disrupt effector-Switch II interactions. Overall, we demonstrate antagonistic regulation between PINK1-dependent Ser111 phosphorylation and LRRK2-mediated Thr72 phosphorylation of Rab8A suggesting that small molecule activators of PINK1 may have therapeutic potential in patients harbouring LRRK2 mutations.

biochemistry↗

bioRxiv2019

Nicotinamide riboside augments the human skeletal muscle NAD+ metabolome and induces transcriptomic and anti-inflammatory signatures in aged subjects: a placebo-controlled, randomized trial

NAD+ is modulated by conditions of metabolic stress and has been reported to decline with aging, but human data are sparse. Nicotinamide riboside (NR) supplementation ameliorates metabolic dysfunction in rodents. We aimed to establish whether oral NR supplementation in aged participants can increase the skeletal muscle NAD+ metabolome, and questioned if tissue NAD+ levels are depressed with aging. We supplemented 12 aged men with NR 1g per day for 21-days in a placebo-controlled, randomized, double-blind, crossover trial. Targeted metabolomics showed that NR elevated the muscle NAD+ metabolome, evident by increased nicotinic acid adenine dinucleotide and nicotinamide clearance products. Muscle RNA sequencing revealed NR-mediated downregulation of energy metabolism and mitochondria pathways. NR also depressed levels of circulating inflammatory cytokines. In an additional study, 31P magnetic resonance spectroscopy-based NAD+ measurement in muscle and brain showed no difference between young and aged individuals. Our data establish that oral NR is available to aged human muscle and identify anti-inflammatory effects of NR, while suggesting that NAD+ decline is not associated with chronological aging per se in human muscle or brain.

clinical trials↗

bioRxiv2018

Coding advantage of grid cell orientation under noisy conditions

Grid cells constitute a crucial component of the \"GPS\" in the mammalian brain. Recent experiments revealed that grid cell activity is anchored to environmental boundaries. More specifically, these results revealed a slight yet consistent offset of 8 degrees relative to boundaries of a square environment. The causes and possible functional roles of this orientation are still unclear. Here we propose that this phenomenon maximizes the spatial information conveyed by grid cells. Computer simulations of the grid cell network reveal that the universal grid orientation at 8 degrees optimizes spatial coding specifically in the presence of noise. Our model also predicts the minimum number of grid cells in each module. In addition, analytical results and a dynamical reinforcement learning model reveal the mechanism underlying the noise-induced orientation preference at 8 degrees. Together, these results suggest that the experimentally observed common orientation of grid cells serves to maximize spatial information in the presence of noise.\n\nAuthor summarySpatial navigation depends on several specialized cell types including place and grid cells. Grid cells have multiple firing fields that are arranged in a regular hexagonal pattern. The axes of this pattern are anchored to environmental boundaries at a universal angle of 8{degrees}. Here, we combine computer simulations of the grid cell network with analytical derivations and a reinforcement learning model to explain the functional relevance of this universal grid cell orientation. We show that spatial information provided by grid cells is maximized at the experimentally observed grid orientation within a broad parameter range. This relationship occurs only in the presence of noise. The model allows for several experimentally testable predictions including the number of grid cells.

neuroscience↗