Biology subjects

Tsai, H.-J.

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

5 recordsLinked to original sources

bioRxiv2026

Nanodisc-Mediated Visualization of Crowding-Induced Condensation and Membrane Reorganization in Two-Dimensional Membrane Environments

Although biological membranes exhibit complex and dynamic organization, the mechanistic role of molecular crowding in governing lateral membrane heterogeneity remains poorly characterized experimentally. Here, we provide direct experimental visualization showing that crowding-induced condensation of membrane-anchored macromolecules above the bilayer interface reorganizes membrane dynamics and gives rise to spatially heterogeneous lipid mobility. Both PEGylated lipids and membrane-anchored proteins undergo surface-density-driven condensation on supported lipid bilayers, forming immobile regions that constrain lipid diffusion. Notably, the condensation threshold varies inversely with PEG chain length and surface density, defining a quantitative relationship between molecular size and crowding strength. To directly visualize these crowding-induced structures, we employed nanodelivery using lipid-loaded nanodiscs, revealing a clear correspondence between condensed crowder regions and diffusion barriers reminiscent of the picket-fence model of live-cell membranes. Similar condensation behavior observed for protein-crowded SLBs demonstrates the generality of this crowding-driven mechanism. Together, these findings establish surface-density-driven crowding and condensation of membrane-anchored macromolecules as a key physical mechanism underlying lateral membrane inhomogeneity and position nanodelivery as a general approach for interrogating membrane organization across synthetic and biological systems.

biophysics↗

bioRxiv2024

Fungi activate Toll-1 dependent immune evasion to induce cell loss in the host brain

Fungi evolve within the host, ensuring their own nutrition and reproduction, at the expense of host health. They intervene in hosts brain function, to alter host behaviour and induce neurodegeneration. In humans, fungal infections are emerging as drivers of neuroinflammation, neurodegenerative diseases and psychiatric disorders. However, how fungi alter the host brain is unknown. Fungi trigger an innate immune response mediated by the Toll-1/TLR receptor, the adaptor MyD88 and the transcription factor Dif/NF{kappa}B, that induce the expression of antimicrobial peptides (AMPs). However, in the nervous system, Toll-1/TLR could also drive an alternative pathway involving the adaptor Sarm, which causes cell death instead. Sarm is the universal inhibitor of MyD88 and could drive immune evasion. The entomopathogenic fungus Beauveria bassiana is well-known to activate Toll-1 signalling in innate immunity in Drosophila. In fruit-flies, the adaptor Wek links Toll-1 to Sarm. Thus, here we asked whether B. bassiana could damage the Drosophila brain via Toll-1, Wek and Sarm. We show that exposure to B. bassiana reduced fly lifespan and impaired locomotion. B. bassiana entered the brain and induced the up-regulation of AMPs, as well as wek and sarm, within the brain. Exposure to B. bassiana caused neuronal and glial loss in the adult Drosophila brain. Importantly, RNAi knockdown of Toll-1, wek or sarm concomitantly with infection prevented B. bassiana induced cell loss. By contrast, over-expression of wek or sarm was sufficient to cause dopaminergic neuron loss in the absence of infection. These data show that B. bassiana caused cell loss in the host brain via Toll-1/Wek/Sarm signalling driving immune evasion. We conclude that pathogens can benefit from an innate immunity receptor to damage the host brain. A similar activation of Sarm downstream of TLRs in response to fungal infections could underlie psychiatric and neurodegenerative diseases in humans.

neuroscience↗

bioRxiv2022

Quantifying the mechanical properties of yeast Candida albicans using atomic force microscopy-based force spectroscopy

Fungi can adapt to a wide range of environmental stress in the wild and host milieu by employing their plastic genome and great diversity in morphology. Among different adaptive strategies, mechanical stimuli, such as changes in osmotic pressure, surface remodelling, hyphal formation, and cell divisions, could guide the physical cues into physiological responses through complex signalling network. While fungal pathogens require a pressure-driven force to expand and penetrate host tissues, quantitatively studying the biophysical properties at the host-fungal interface is critical to understand the development of fungal diseases. Microscopy-based techniques have enabled researchers to monitor the dynamic mechanics on fungal cell surface in responses to the host stress and antifungal drugs. Here, we describe a label-free, high-resolution method based on atomic force microscopy, with a step-by-step protocol to measure the physical properties in human fungal pathogen Candida albicans.

microbiology↗

bioRxiv2021

Neutralizing IFNL3 Autoantibodies in Severe COVID-19 Identified Using Molecular Indexing of Proteins by Self-Assembly

Unbiased antibody profiling can identify the targets of an immune reaction. A number of likely pathogenic autoreactive antibodies have been associated with life-threatening SARS-CoV-2 infection; yet, many additional autoantibodies likely remain unknown. Here we present Molecular Indexing of Proteins by Self Assembly (MIPSA), a technique that produces ORFeome-scale libraries of proteins covalently coupled to uniquely identifying DNA barcodes for analysis by sequencing. We used MIPSA to profile circulating autoantibodies from 55 patients with severe COVID-19 against 11,076 DNA-barcoded proteins of the human ORFeome library. MIPSA identified previously known autoreactivities, and also detected undescribed neutralizing interferon lambda 3 (IFN-{lambda}3) autoantibodies. At-risk individuals with anti-IFN-{lambda}3 antibodies may benefit from interferon supplementation therapies, such as those currently undergoing clinical evaluation. One-Sentence SummaryMolecular Indexing of Proteins by Self Assembly (MIPSA) identifies neutralizing IFNL3 autoantibodies in patients with severe COVID-19. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=144 SRC="FIGDIR/small/432977v1_ufig1.gif" ALT="Figure 1"> View larger version (25K): org.highwire.dtl.DTLVardef@a3c55aorg.highwire.dtl.DTLVardef@1f1c840org.highwire.dtl.DTLVardef@920bc7org.highwire.dtl.DTLVardef@43633e_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2019

Autonomously replicating linear plasmids facilitate the analysis of replication origin function in Candida albicans

The ability to generate autonomously replicating plasmids has been elusive in Candida albicans, a prevalent human fungal commensal and pathogen. Instead, plasmids generally integrate into the genome. Here, we assessed plasmid and transformant properties, including plasmid geometry, transformant colony size, four selectable markers, and potential origins of replication for their ability to drive autonomous plasmid maintenance. Importantly, linear plasmids with terminal telomere repeats yielded many more autonomous transformants than circular plasmids with the identical sequences. Furthermore, we could distinguish by colony size, transient, autonomously replicating and chromosomally integrated transformants (tiny, medium and large, respectively). Candida albicans URA3 and a heterologous marker, ARG4, yielded many transient transformants indicative of weak origin activity; replication of plasmid carrying heterologous LEU2 marker was highly dependent upon the addition of a bona fide origin sequence. Several bona fide chromosomal origins, with an origin fragment of ~100 bp as well as a heterologous origin, panARS, from Kluyveromyces lactis drove autonomous replication, yielding moderate transformation efficiency and plasmid stability. Thus, C. albicans maintains linear plasmids that yield high transformation efficiency and are maintained autonomously in an origin-dependent manner. ImportanceCircular plasmids are important tools for molecular manipulation in model fungi such as bakers yeast, yet, in Candida albicans, an important yeast pathogen of humans, prior studies were not able to generate circular plasmids that were autonomous (duplicated without inserting themselves into the chromosome). Here, we found that linearizing circular plasmids with sequences from telomeres, the chromosome ends, allows the plasmids to duplicate and segregate in C. albicans. We used this system to identify chromosomal sequences that facilitate the initiation of plasmid replication (origins) and to show that a ~100 bp fragment of a C. albicans origin, as well as an origin sequence from a distantly related yeast, can both function as origins in C. albicans. Thus, the requirements for plasmid geometry, but not necessarily for origin sequences, differ between C. albicans and bakers yeast.

molecular biology↗