Biology subjects

Hung, S.-H.

Publications and source records attributed to Hung, S.-H..

5 recordsLinked to original sources

bioRxiv2025

The conserved bridging domain on HCV E1E2 glycoprotein complex is targeted by neutralizing antibodies from diverse lineages

The induction of potent and cross-reactive neutralizing antibody (nAb) responses remains a challenge in vaccine development against antigenically diverse viruses such as hepatitis C virus (HCV). The HCV E1E2 glycoprotein complex contains two major neutralizing sites: the neutralizing face (NF) and the less explored bridging domain (BD). Here, we characterized 25 BD-targeting nAbs isolated from infection or immunization. These antibodies arise from diverse B cell lineages but share convergent CDRH3 features. Epitope mapping by alanine scanning and negative-stain electron microscopy revealed overlapping epitopes on BD spanning antigenic regions AR4 and AR5, with variable back layer engagement. The crystal structure of a non-human primate BD nAb RM3-26 in complex with E2 uncovered a back layer-directed recognition mode analogous to that of the human nAb hcab40. Together, BD- and NF-directed nAbs exhibited additivity in their neutralization, highlighting BD as a conserved site of vulnerability on HCV and a valuable target for rational vaccine design.

immunology↗

bioRxiv2025

Native-like soluble E1E2 glycoprotein heterodimers on self-assembling protein nanoparticles for hepatitis C virus vaccine design

Hepatitis C virus (HCV) is a leading cause of chronic liver disease, cirrhosis, and hepatocellular carcinoma worldwide. E1E2-based HCV vaccine development has been hindered by the challenge of producing a soluble E1E2 (sE1E2) antigen that faithfully recapitulates the native glycoprotein heterodimer found on virions. Based on available cryo-electron microscopy (cryo-EM) structures, we rationally engineered sE1E2 for genotype 1a H77 by truncating the E1 and E2 stems (Cut1), removing a putative fusion peptide (pFP)-containing region in E1 (Cut2), and stabilizing the E1-E2 interface with diverse heterodimeric scaffolds. All H77 sE1E2.Cut1+2 scaffolds showed native-like E1-E2 association and robust binding to the broadly neutralizing antibody (bNAb) AR4A. A genotype 1a HCV-1 sE1E2.Cut1+2 variant scaffolded by a modified SpyTag/SpyCatcher (SPY{Delta}N) was selected for in vitro, structural, and immunogenic characterization. The structure of this sE1E2 scaffold in complex with bNAbs was analyzed by cryo-EM and negative-stain EM (nsEM), with an nsEM-based approach developed for antibody epitope mapping. HCV-1 sE1E2.Cut1+2.SPY{Delta}N was displayed on self-assembling protein nanoparticles (SApNPs) to enhance immunogenicity. HCV-1 sE1E2.Cut1+2.SPY{Delta}N heterodimer and SApNPs with wildtype and modified glycans were tested in mice, revealing the beneficial effects of multivalent display and oligomannose enrichment. Our study provides a rigorous foundation for next-generation HCV vaccine development. ONE-SENTENCE SUMMARYRational design, characterization, and in vivo assessment of HCV soluble E1E2 heterodimer and nanoparticles will inform vaccine development.

microbiology↗

bioRxiv2024

Multifaceted roles of H2B mono-ubiquitylation in D-loop metabolism during homologous recombination repair

Repairing DNA double-strand breaks is crucial for maintaining genome integrity, which occurs primarily through homologous recombination (HR) in S. cerevisiae. Nucleosomes, composed of DNA wrapped around a histone octamer, present a natural barrier to end-resection to initiate HR, but the impact on the downstream HR steps of homology search, DNA strand invasion and repair synthesis remain to be determined. Displacement loops (D-loops) play a pivotal role in HR, yet the influence of chromatin dynamics on D-loop metabolism remains unclear. Using the physical D-loop capture (DLC) and D-loop extension (DLE) assays to track HR intermediates, we employed genetic analysis to reveal that H2B mono-ubiquitylation (H2Bubi) affects multiple steps during HR repair. We infer that H2Bubi modulates chromatin structure, not only promoting histone degradation for nascent D-loop formation but also stabilizing extended D-loops through nucleosome assembly. Furthermore, H2Bubi regulates DNA resection via Rad9 recruitment to suppress a feedback control mechanism that dampens D-loop formation and extension at hyper-resected ends. Through physical and genetic assays to determine repair outcomes, we demonstrate that H2Bubi plays a crucial role in preventing break-induced replication and thus promoting genomic stability. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/612919v1_ufig1.gif" ALT="Figure 1"> View larger version (74K): org.highwire.dtl.DTLVardef@48b9edorg.highwire.dtl.DTLVardef@a6b23forg.highwire.dtl.DTLVardef@707dorg.highwire.dtl.DTLVardef@d93e58_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LIH2Bubi is epistatic to H2A.Z and INO80 in promoting homology search and D-loop formation C_LIO_LIH2Bubi stabilizes extended D-loop C_LIO_LIExcessive resection counteracts D-loop formation and extension C_LIO_LIH2Bubi promotes crossover events and limits the frequency of break-induced replication outcomes in HR repair C_LI

genetics↗

bioRxiv2023

Multi-Step Control of Homologous Recombination by Mec1/ATR Ensures Robust Suppression of Gross Chromosomal Rearrangements

The Mec1/ATR kinase is crucial for genome stability, yet the mechanism by which it prevents gross chromosomal rearrangements (GCRs) remains unknown. Here we find that in cells with deficient Mec1 signaling, GCRs accumulate due to the deregulation of multiple steps in homologous recombination (HR). Mec1 primarily suppresses GCRs through its role in activating the canonical checkpoint kinase Rad53, which ensures the proper control of DNA end resection. Upon loss of Rad53 signaling and resection control, Mec1 becomes hyperactivated and triggers a salvage pathway in which the Sgs1 helicase is recruited to sites of DNA lesions via the 911-Dpb11 scaffolds to favor heteroduplex rejection and limit HR-driven GCR accumulation. Fusing an ssDNA recognition domain to Sgs1 bypasses the requirement of Mec1 signaling for GCR suppression and nearly eliminates D-loop formation, thus preventing non-allelic recombination events. We propose that Mec1 regulates multiple steps of HR to prevent GCRs while ensuring balanced HR usage when needed for promoting tolerance to replication stress.

genetics↗

bioRxiv2022

Nitrogen competition is the general mechanism underlying cnidarian-Symbiodiniaceae symbioses

Symbiotic associations with Symbiodiniaceae have evolved independently across a diverse range of cnidarian taxa including reef-building corals, anemones and jellyfish, yet the molecular mechanisms underlying their regulation and repeated evolution are still elusive. Here we show that despite their independent evolution, cnidarian hosts employ the same mechanism of symbiont control in which symbiont-derived glucose is used to assimilate nitrogenous waste via amino acid biosynthesis to limit the availability of nitrogen to the symbionts. In this metabolic interaction, glucose significantly reduces symbiont density while ammonium promotes symbiont proliferation. We show that glucose-derived 13C and ammonium-derived 15N are co-incorporated into amino acids by the hosts. Metabolic differences between the hosts further suggest that corals are more susceptible to environmental stress and symbiosis breakdown due to their increased energy demands to satisfy calcification. Our results reveal the general metabolic interaction underlying these symbioses and provide a parsimonious explanation for their repeated evolution.

ecology↗