Biology subjects

Lim, S.

Publications and source records attributed to Lim, S..

At least 55 records · Page 3Linked to original sources

bioRxiv2024

Comparative evaluation of cell-based assay technologies for scoring drug-induced condensation of SARS-CoV-2 nucleocapsid protein

Protein-nucleic acid phase separation has been implicated in many diseases such as viral infections, neurodegeneration, and cancer. There is great interest in identifying condensate modulators (CMODs), which are small molecules that alter the dynamics and functions of phase-separated condensates, as a potential therapeutic modality. Most CMODs were identified in cellular high-content screens (HCS) where micron-scale condensates were characterized by fluorescence microscopy. These approaches lack information on protein dynamics, are limited by microscope resolution, and are insensitive to subtle condensation phenotypes missed by overfit analysis pipelines. Here, we evaluate two alternative cell-based assays: high-throughput single molecule tracking (htSMT) and proximity-based condensate biosensors using NanoBIT (split luciferase) and NanoBRET (bioluminescence resonance energy transfer) technologies. We applied these methods to evaluate condensation of the SARS-CoV-2 nucleocapsid (N) protein under GSK3 inhibitor treatment, which we had previously identified in our HCS campaign to induce condensation with well-defined structure-activity relationships (SAR). Using htSMT, we observed robust changes in N protein diffusion as early as 3 hours post GSK3 inhibition. Proximity-based N biosensors also reliably reported on condensation, enabling the rapid assaying of large compound libraries with a readout independent of imaging. Both htSMT and proximity-based biosensors performed well in a screening format and provided information on CMOD activity that was complementary to HCS. We expect that this expanded toolkit for interrogating phase-separated proteins will accelerate the identification of CMODs for important therapeutic targets.

molecular biology↗

bioRxiv2024

Attention and Retention Effects of Culturally Targeted Billboard Messages: An Eye-Tracking Study Using Immersive Virtual Reality

Targeting, the creation of a match between message content and receiver characteristics, is a key strategy in communication message design. Cultural targeting, or adapting message characteristics to be congruent with a groups cultural knowledge, appearance, or beliefs of recipients, is used in practice and is a potentially effective strategy to boost the relevance of a message, affecting attention to messages and enhancing effects. However, many open questions remain regarding the mechanisms and consequences of targeting. This is partly due to methodological challenges in experimentally manipulating messages that match cultural recipient characteristics while simultaneously measuring effects and balancing experimental control and realism. Here, we used a novel VR-based paradigm in which participants drove along a virtual highway flanked by billboards with varying message designs. Specifically, we manipulated the message design to either match or mismatch peoples cultures of origin. We used unobtrusive eye tracking to assess participants attention (i.e., for how long and how often they look at matched vs. unmatched billboards). Results show a tendency of the participants to inspect culturally matched billboards more often and for longer. We further found that matched billboards produce better recall, indicating more efficient encoding and storage of the messages. Our results underscore the effectiveness of cultural targeting and demonstrate how researchers can rigorously manipulate relevant message factors using virtual environments. We discuss the implications of these findings regarding theories of cultural targeting and methodological perspectives for the objective measurement of exposure factors through eye tracking.

animal behavior and cognition↗

bioRxiv2024

Integrin-activating Yersinia protein Invasin sustains long-term expansion of primary epithelial cells as 2D organoid sheets

Matrigel/BME, a basement membrane-like preparation, supports long-term growth of epithelial 3D organoids from adult stem cells (ASC)1,2. Here, we show that interaction between Matrigels major component Laminin111 with epithelial 6{beta}1-integrin is crucial for this process. The outer membrane protein Invasin of Yersinia is known to activate multiple integrin-{beta}1 complexes, including integrin-6{beta}1. A C-terminal integrin-binding fragment of Invasin, coated on culture plates, mediated gut epithelial cell adhesion. Addition of organoid growth factors allowed multi-passage expansion in 2D. Polarization, junction formation and generation of enterocytes, goblet cells, Paneth cells, and enteroendocrine cells was stable over time. Sustained expansion of other human-, mouse-, and even snake epithelia was accomplished under comparable conditions. The 2D organoid sheet format holds advantages over the 3D in gel format in terms of imaging, accessibility of basal and apical domains and automation for high throughput screening. Invasin represents a fully defined, affordable, versatile, and animal-free complement to Matrigel/BME.

cell biology↗

bioRxiv2024

Using VR and eye-tracking to study attention to and retention of AI-generated ads in outdoor advertising environments

In contemporary urban environments, advertisements are ubiquitous, capturing the attention of individuals navigating through cityscapes. This study simulates this situation by using VR as an advertising research tool and combining it with eye-tracking to rigorously assess attention to and retention of visual advertisements. Specifically, participants drove through a virtual city with 40 AI-generated, experimentally manipulated, and randomly assigned advertisements (20 commercial, 20 health) distributed throughout. Our results confirm theoretical predictions about the link between exposure, visual attention, and incidental memory. Specifically, we found that attended ads are likely to be recalled and recognized, and concurrent task demands (counting sales signs) decreased visual attention and subsequent recall and recognition of the ads. Finally, we identify the influence of ad placement in the city as a very important but previously hard-to-study factor influencing advertising effects. This paradigm offers great flexibility for biometric advertising research and can be adapted to varying contexts, including highways, airports, and subway stations, and theoretically manipulate other variables. Moreover, considering the metaverse as a next-generation advertising environment, our work demonstrates how causal mechanisms can be identified in ways that are of equally high interest to both theoretical as well as applied advertising research.

animal behavior and cognition↗

bioRxiv2024

Normalization of Single-cell RNA-seq Data UsingPartial Least Squares with Adaptive Fuzzy Weight

Normalization of single-cell RNA-seq (scRNA-seq) is a crucial step in downstream analysis, where raw data are adjusted to correct unwanted factors that prevent the direct comparison of expression measures. scRNA-seq data exhibits a multivariate relationship between transcript-specific expression and sequencing depth that a single scale factor cannot address. A partial least squares (PLS) regression was performed to accommodate the variability of gene expression in each condition, and upper and lower quantiles with adaptive fuzzy weights were utilized to correct unwanted biases in scRNA-seq data. The present approach was compared using real and simulated datasets across various state-of-the-art performance measures.

bioinformatics↗

bioRxiv2024

Extensively acquired antimicrobial resistant bacteria restructure the individual microbial community in post-antibiotic conditions

In recent years, the overuse of antibiotics has led to the emergence of antimicrobial resistant (AMR) bacteria. To evaluate the spread of AMR bacteria, the reservoir of AMR genes (resistome) has traditionally been identified from environmental samples, hospital environments, and human populations; however, the functional role of AMR bacteria in the human gut microbiome and their persistency within individuals has not been fully investigated. Here, we performed a strain-resolved in-depth analysis of the resistome changes by reconstructing a large number of metagenome-assembled genomes (MAGs) of antibiotics- treated individuals gut microbiome. Interestingly, we identified two bacterial populations with different resistome profiles, extensively acquired antimicrobial resistant bacteria (EARB) and sporadically acquired antimicrobial resistant bacteria (SARB), and found that EARB showed broader drug resistance and a significant functional role in shaping individual microbiome composition after antibiotic treatment. Furthermore, longitudinal strain analysis revealed that EARB bacteria were inherently carried by individuals and can reemerge through strain switching in the human gut microbiome. Our data on the presence of AMR bacteria in the human gut microbiome provides a new avenue for controlling the spread of AMR bacteria in the human community.

genomics↗

bioRxiv2024

Eyes on VR: Unpacking the Causal Chain Between Exposure, Reception, and Retention for Emotional Billboard Messages

The causal chain from message exposure to reception to effects is widely accepted as the basic explanatory model for communication outcomes. Problematically, the chains links are often studied in isolation, leaving measurement gaps that compromise the ecological validity and practical utility of experimental research. Here we introduce a VR-based paradigm that encompasses a realistic message reception context, i.e., a simulated car ride on a highway flanked by billboards. We varied attentional message factors (emotional content) as well as contextual task distractions (trash-counting). VR-integrated eye trackers were used to capture participants incidental message exposure dependent on their actual gaze behavior. Consistent with our predictions, results show that 1. exposure gates all subsequent effects; 2. distraction impacts likelihood of exposure; 3. both the manipulation of emotional content and distraction affect retention. This comprehensive analysis of the exposure-reception-retention chain can be broadly applied to a variety of message reception contexts that will be discussed.

animal behavior and cognition↗

bioRxiv2024

Modulation of Ideonella sakaiensis PETase active site flexibility and activity on morphologically distinct substrates by surface charge engineering

Enzymatic hydrolysis of polyethylene terephthalate (PET) waste is a compelling strategy for environmentally friendly recycling of a major pollutant. Here, we investigate the effects of surface charge point mutations both proximal and distal to the active site of the mesophilic PET-degrading enzyme from Ideonella sakaienses (IsPETase) and an engineered thermostable variant with superior activity, STAR PETase. The vicinal K95A mutation significantly inhibited IsPETase activity on mechanically processed PET powder. Conversely, this mutation significantly increased hydrolysis of PET powder in the STAR PETase background. Activity of both enzymes on PET film was inhibited by the K95A mutation, highlighting complex interplay between mutation context and substrate morphology. Further installing the distal R132N and R280A surface charge mutations potentiated activity of STAR on all substrates tested. This variant afforded 100% degradation of bottle-grade PET powder in 3 days at 40{degrees}C reaction temperature, a 3-fold improvement over IsPETase. Molecular dynamics simulations reveal modulation of active site flexibility in mutants, which differentially impacts both hydrolysis of morphologically distinct PET substrates and the concentration-dependent inhibition phenomenon observed for PETase.

bioengineering↗

bioRxiv2024

Characterization of the murine spine for spaceflight studies

Rodents provide a useful analog for understanding the effects of spaceflight on the human body, offering opportunities for investigations into the relationship between microgravity and the musculoskeletal system. In particular, rodents have often been utilized to improve our understanding of the effects of spaceflight on the spine, including intervertebral disc and vertebral body health. However, there are a number of experimental factors that differ between existing works, including mission duration, animal housing, and anatomical location of interest, making it difficult to draw holistic conclusions. Additionally, the quadrupedal nature of the murine spine results in different biomechanical loading than in a bipedal organism. Thus, the objective of this study was to more fully define the bulk properties of the murine lumbar spine model after 28 days of spaceflight. Additionally, the proximal tibia was analyzed to provide insight into the skeletal site-specificity of gravitational unloading in space. Results indicated that the effects of spaceflight on vertebral body bone microarchitecture, intervertebral disc biochemistry, and intervertebral disc joint mechanics were statistically insignificant, while large and significant bone loss was observed in the proximal tibia of the same animals. We hypothesize that this may be due to site-specific loading changes in space. Specifically, vigorous ambulatory behaviors observed in this experiment after initial acclimation to spaceflight may increase axial load-bearing in the lumbar spine, while maintaining microgravity induced mechanical unloading in the tibia. In total, this work shows that the rodent spine, unlike the weight bearing tibia in the same mice, is not affected by gravitational unloading, suggesting the tissue degenerative effects of spaceflight are site- and load-specific and not systemic. This study also highlights the importance of considering experimental variables such as habitat acclimation, physical activity, and experiment duration as key factors in determining musculoskeletal and spine health outcomes during spaceflight.

bioengineering↗

bioRxiv2024

The eyes have it: Inter-subject correlations of pupillary responses for audience response measurement in VR

The eye is the vanguard of the reception process, constituting the point where visual information arrives and is transformed into neural signals. While we view dynamic media contents, a fine-tuned interplay of mechanisms causes our pupils to dilate and constrict over time - and putatively similarly across audience members exposed to the same messages. Research that once pioneered pupillometry did actually use dynamic media as stimuli, but this trend then stalled, and pupillometry remained underdeveloped in the study of naturalistic media stimuli. Here, we introduce a VR-based approach to capture audience members pupillary responses during media consumption and suggest an innovative analytic framework. Specifically, we expose audiences to a set of 30 different video messages and compute the cross-receiver similarity of pupillometric responses. Based on this data, we identify the specific video an individual is watching. Our results show that this pupil-pulse-tracking enables highly accurate decoding of video identity. Moreover, we demonstrate that the decoding is relatively robust to manipulations of video size and distractor presence. Finally, we examine the relationship between pupillary responses and subsequent memory. Theoretical implications for objectively quantifying exposure and states of audience engagement are discussed. Practically, we anticipate that this pupillary audience response measurement approach could find application in media measurement across contexts, ranging from traditional screen-based media (commercials, movies) to social media (e.g., TikTok and YouTube), and to next-generation virtual media environments (e.g., Metaverse, gaming).

physiology↗

bioRxiv2023

Tardigrade secretory proteins protect biological structures from desiccation

Tardigrades, microscopic animals that survive a broad range of environmental stresses, express a unique set of proteins termed tardigrade-specific intrinsically disordered proteins (TDPs). TDPs are often expressed at high levels in tardigrades upon desiccation, and appear to mediate stress adaptation. Here, we focused on the proteins belonging to the secretory family of tardigrade proteins termed secreted-abundant heat soluble ("SAHS") proteins, and investigated their ability to protect diverse biological structures. Recombinantly expressed SAHS proteins prevented desiccated liposomes from fusion, and enhanced desiccation tolerance of E. coli and Rhizobium tropici upon extracellular application. Molecular dynamics simulation and comparative structural analysis suggest a model by which SAHS proteins may undergo a structural transition upon desiccation, in which removal of water and solutes from a large internal cavity in SAHS proteins destabilizes the beta-sheet structure. These results highlight the potential application of SAHS proteins as stabilizing molecules for preservation of cells.

molecular biology↗

bioRxiv2023

Sensory-memory interactions via modular structure explain errors in visual working memory

Errors in stimulus estimation reveal how stimulus representation changes during cognitive processes. Repulsive bias and minimum variance observed near cardinal axes are well-known error patterns typically associated with visual orientation perception. Recent experiments suggest that these errors continuously evolve during working memory, posing a challenge that neither static sensory models nor traditional memory models can address. Here, we demonstrate that these evolving errors, maintaining characteristic shapes, require network interaction between two distinct modules. Each module fulfills efficient sensory encoding and memory maintenance, which cannot be achieved simultaneously in a single-module network. The sensory module exhibits heterogeneous tuning with strong inhibitory modulation reflecting natural orientation statistics. While the memory module, operating alone, supports homogeneous representation via continuous attractor dynamics, the fully connected network forms discrete attractors with moderate drift speed and nonuniform diffusion processes. Together, our work underscores the significance of sensory-memory interaction in continuously shaping stimulus representation during working memory.

neuroscience↗

bioRxiv2023

Slow conduction and spatial dispersion of repolarization are intrinsic properties of cardiomyocyte electrophysiology that contribute to proarrhythmia in an iPSC model of hypertrophic cardiomyopathy

Hypertrophic cardiomyopathy (HCM) is an inherited heart muscle disease; characterised by left ventricular wall thickening, cardiomyocyte disarray, and fibrosis, and is associated with arrhythmias, heart failure and sudden death. However, it is unclear to what extent the electrophysiological disturbances that lead to sudden death occur secondary to the structural changes in the myocardium, or as a result of intrinsic properties of the HCM cardiomyocyte. In this study, we used an induced pluripotent stem cell model of the Arg403Gln variant in myosin heavy chain 7 (MYH7) to study tissue level electrophysiological properties of HCM cardiomyocytes. For the first time, we show significant slowing of conduction velocity and an increase in local spatial dispersion of repolarisation - both well-established substrates for arrhythmia - in monolayers of HCM cardiomyocytes. Analysis of rhythmonome protein expression in R403Q cardiomyocytes revealed dramatically reduced connexin-43, sodium channels, and inward rectifier channels - a three-way hit that combines to reduce electrotonic coupling between HCM cardiomyocytes and slow cardiac conduction. Our data therefore represent a novel, biophysical basis for arrhythmia in HCM, that is intrinsic to cardiomyocyte electrophysiology. Later in the progression of the disease, these proarrhythmic electrical phenotypes may be accentuated by fibrosis and myocyte disarray to contribute to sudden death in HCM patients.

physiology↗

bioRxiv2023

Modeling the seasonal variation of windborne transmission of porcine reproductive and respiratory syndrome virus between swine farms

Modeling windborne transmission of aerosolized pathogens is challenging. We adapted an atmospheric dispersion model named Hybrid Single-Particle Lagrangian Integrated Trajectory model (HYSPLIT) to simulate windborne dispersion of porcine reproductive and respiratory syndrome virus (PRRSv) between swine farms and incorporated the findings into an outbreak investigation. The risk was estimated semi-quantitatively based on the cumulative daily deposition of windborne particles, and the distance to closest emitting farm with an ongoing outbreak. Five years of data (2014 : 2018) were used to study seasonal differences of deposition thresholds of the airborne particles containing PRRSv and to evaluate model in relation to risk prediction and barn air filtration. When considered the 14-day cumulative deposition, in Winter, above threshold particle depositions would reach up to 30 km from emitting farms with 84% of them being within 10km. Long-distance pathogen transmission was highest in Winter and Fall, lower in Spring, and least in Summer. The model successfully replicated the observed seasonality of PRRSv where Fall and Winter posing a higher risk for outbreaks. Reaching the humidity and temperature thresholds tolerated by the virus in Spring and Summer reduced the survival and infectivity of aerosols beyond 10 -20 km. Within in the data limitations of voluntary participation, when assumed wind as the sole route of PRRSv transmission, the predictive performance of the model was fair with >0.64 AUC. Barn air filtration was associated with fewer outbreaks, particularly when exposed to high levels of viral particles. The study confirms the usefulness of HYSPLIT models as a tool when determining seasonal effects, distances, and inform near real-time risk of windborne PRRSv transmission that can be useful in future outbreak investigations and implementing timely control measures.

evolutionary biology↗

bioRxiv2023

Decision-consistent bias mediated by drift dynamics of human visual working memory

Sensory evidence tends to be fleeting, often unavailable when we categorize or estimate world features. To overcome this, our brains sustain sensory information in working memory. Although keeping that information accurate while acting on it is vital, humans display two canonical biases: estimates are biased toward a few stimuli ("stimulus-specific bias") and prior decisions ("decision-consistent bias"). Integrative--especially neural mechanistic--accounts of these biases remain scarce. Here, we identify drift dynamics toward discrete attractors as a common source of both biases in orientation estimation, with decisions further steering memory states. Behavior and neuroimaging data reveal how these biases co-evolve through the decision-steered attractor dynamics. Task-optimized recurrent neural networks suggest neural mechanisms that enable categorical decisions to emerge from working memory for continuous stimuli while updating its trajectory, warping decision-consistent biases under stimulus-specific drift.

neuroscience↗

bioRxiv2023

Spatiotemporal Transcriptomes of Pig Hearts Reveal Midkine-Mediated Vascularization in a Chronic Myocardial Infarcted Model

Ischemic heart disease is the most prevalent cause of death globally. Regenerative cardiology using stem cell-based therapy is a potential approach to replace infarcted myocardial (MI) heart tissue. We used cardiovascular progenitors (CVPs) derived from human pluripotent embryonic stem cells differentiated to cardiomyocyte progenitors on a laminin 521+221 matrix and transplanted them into acute and chronic MI pig hearts (AMI and CMI). We performed time-series spatial transcriptomics to characterize these human cells at AMI 1- and 2- and at CMI 1-, 4- and 12 weeks post-transplantation. Both models showed high transcriptional reproducibility in the replicates. Furthermore, the human grafts engrafted well, matured, and expressed metabolic, ribosomal, T-tubule, and channel-related genes in the human graft over time. Cell-cell communication analysis revealed Midkine (MDK) signaling as a key pathway that may lead to increased angiogenesis of collaterals in the human graft.

bioinformatics↗

bioRxiv2023

The VR Billboard Paradigm: Using VR and Eye-tracking to Examine the Exposure-Reception-Retention Link in Realistic Communication Environments

Exposure is the cornerstone of media and message effects research. If a health, political, or commercial message is not noticed, no effects can ensue. Yet, existing research in communication, advertising, and related disciplines often fails to measure exposure and demonstrate the causal link between quantified exposure to outcomes because actual exposure (i.e., whether recipients were not only exposed to messages but also took notice of them) is difficult to capture. Here, we harness Virtual Reality (VR) technology integrated with eye tracking to overcome this challenge. While eye-tracking technology alone can capture whether people attend to messages in their communication environment, most eye-tracking research is bound by laboratory-based screen-reading paradigms that are not representative of the broader communication environments in which messages are encountered. Emerging eye-tracking field research suffers from an inability to control and experimentally manipulate key variables. Our solution is to measure eye-tracking within an immersive environment in VR that resembles a realistic message reception context. Specifically, we simulate driving down a highway alongside which billboards are placed and use VR-integrated eye-tracking to measure whether the drivers look at individual billboard messages. This allows us to rigorously quantify the nexus between exposure and reception, and to link our measures to subsequent memory, i.e., whether messages were remembered, forgotten, or not even encoded. We further demonstrate that manipulating drivers attention directly impacts gaze behavior and memory. We discuss the large potential of this paradigm to quantify exposure and message reception in realistic communication environments and the equally promising applications in new media contexts (e.g., the Metaverse).

animal behavior and cognition↗

bioRxiv2023

Comparison of histological delineations of medial temporal lobe cortices by four independent neuroanatomy laboratories

The medial temporal lobe (MTL) cortex, located adjacent to the hippocampus, is crucial for memory and prone to the accumulation of certain neuropathologies such as Alzheimers disease neurofibrillary tau tangles. The MTL cortex is composed of several subregions which differ in their functional and cytoarchitectonic features. As neuroanatomical schools rely on different cytoarchitectonic definitions of these subregions, it is unclear to what extent their delineations of MTL cortex subregions overlap. Here, we provide an overview of cytoarchitectonic definitions of the cortices that make up the parahippocampal gyrus (entorhinal and parahippocampal cortices) and the adjacent Brodmann areas (BA) 35 and 36, as provided by four neuroanatomists from different laboratories, aiming to identify the rationale for overlapping and diverging delineations. Nissl-stained series were acquired from the temporal lobes of three human specimens (two right and one left hemisphere). Slices (50 {micro}m thick) were prepared perpendicular to the long axis of the hippocampus spanning the entire longitudinal extent of the MTL cortex. Four neuroanatomists annotated MTL cortex subregions on digitized (20X resolution) slices with 5 mm spacing. Parcellations, terminology, and border placement were compared among neuroanatomists. Cytoarchitectonic features of each subregion are described in detail. Qualitative analysis of the annotations showed higher agreement in the definitions of the entorhinal cortex and BA35, while definitions of BA36 and the parahippocampal cortex exhibited less overlap among neuroanatomists. The degree of overlap of cytoarchitectonic definitions was partially reflected in the neuroanatomists agreement on the respective delineations. Lower agreement in annotations was observed in transitional zones between structures where seminal cytoarchitectonic features are expressed more gradually. The results highlight that definitions and parcellations of the MTL cortex differ among neuroanatomical schools and thereby increase understanding of why these differences may arise. This work sets a crucial foundation to further advance anatomically-informed human neuroimaging research on the MTL cortex.

neuroscience↗