Biology subjects

Jih, J.

Publications and source records attributed to Jih, J..

5 recordsLinked to original sources

bioRxiv2025

IsoNet2 determines cellular structures at submolecular resolution without averaging

AbstractWe introduce IsoNet2, an end-to-end self-supervised deep-learning method that directly reconstructs high-quality 3D densities from cryogenic electron tomography. A unified network simultaneously performs denoising, contrast transfer function correction, and missing-wedge restoration, achieving [~]20 [A] resolution without averaging. A feature-rich GUI enables rapid, dataset-specific fine-tuning for end-users. IsoNet2 resolves domain organization in HIV capsid proteins, tRNA occupancy in individual ribosomes, and in situ architectures of mitochondrial respiration-related complexes, enabling atomic-level interpretation of cellular environments.

cell biology↗

bioRxiv2025

Structure of a new capsid form and comparison with A-, B- and C-capsids clarify herpesvirus assembly and DNA translocation

Three capsid types have been recognized from the nuclei of herpesvirus-infected cells: empty A-capsids, scaffolding-containing B-capsids, and DNA-filled C-capsids. Despite progress in determining atomic structures of these capsids and extracellular virions in recent years, debate persists concerning the origins and temporal relationships among these capsids during capsid assembly and genome packaging. Here, we have imaged over 300,000 capsids of herpes simplex virus type 1 by cryogenic electron microscopy (cryoEM) and exhaustively classified them to characterize the structural heterogeneity of the DNA-translocating portal complex and their functional states. The resultant atomic structures reveal not only the expected A-, B-, and C-capsids, but also capsids with portal vertices similar to C-capsids but no resolvable genome in the capsid lumen, which we named D-capsids. The dodecameric dsDNA-translocating portal complex varies across these capsid types in their radial positions in icosahedral capsids and exhibits structural dynamics within each capsid type. In D-capsids, terminal DNA density exists in multiple conformations including one reminiscent to that in C-capsids, suggesting D-capsids are products of failed DNA retention. This interpretation is supported by varying amounts of DNA outside individual D-capsids and by correlation of capsid counts observed in situ of infected cell nuclei and those after purification. Additionally, an "anchoring" segment of the scaffold protein is resolved interacting with the portal baskets of A- and B-capsids but not D- and C-capsids. Taken together, our data indicate that A-capsids arise from failed DNA packaging and D-capsids from failed genome retention, clarifying the origins of empty capsids in herpesvirus assembly. SignificanceAs the prototypical herpesvirus, herpes simplex virus 1 (HSV-1) exhibits a global seroprevalence of 67% and approaching 90% in some localities. Herpesvirus infections can cause devastating cancers and birth defects, with HSV-1 infections leading to cold sores among the general population worldwide and blindness in developing nations. Here, we present atomic structures of the capsids sorted out from the nuclear isolates of HSV-1 infected cells, including the previously recognized A-, B-, and C-capsids, as well as the newly identified D-capsid. The structures show the details of protein-protein and protein-DNA interactions within each capsid type and the positional and interactional variability of the viral DNA-translocating portal vertex among these capsids. Importantly, our findings suggest that A-capsids are products of failed dsDNA packaging and D-capsids of failed genome retention. Together, the high-resolution 3D structures clarify the processes of genome packaging, maintenance, and ejection during capsid assembly, which are conserved across all herpesviruses.

microbiology↗

bioRxiv2025

Demonstration and structural basis of a therapeutic DNA aptamer for SARS-CoV-2 spike protein detection

At the onset of the COVID-19 pandemic, the absence of a rapid and highly specific diagnostic method for the SARS-CoV-2 virus led to significant delays in detection, adversely affecting clinical outcomes. This shortfall highlights the urgent need for adaptable, scalable, and reusable diagnostic technologies to improve future pandemic responses. To address this challenge, we developed a renewable electrochemical impedance biosensor device employing a synthetic nucleotide-based therapeutic aptamer (termed aptasensor) targeting the SARS-CoV-2 spike (S) protein receptor-binding domain (RBD). We demonstrate that our aptasensor can detect the Omicron BA.2 S protein within one hour and possesses concentration-dependent sensitivity at biologically relevant levels. Notably, the aptasensor is reusable after regeneration by a simple pH 2 buffer treatment. Aptamer binding to the S protein was confirmed by immunogold labeling and visualization by negative-stain electron microscopy. We used cryogenic electron microscopy (cryo-EM) to resolve high-resolution maps of the S protein in both the open and closed conformations and characterized aptamer binding to the up RBD in the open conformation. Taken together, these results establish the versatility and scalability of aptamer-based biosensors, presenting them as a potential transformative diagnostic platform for emerging pathogens. This combination of rapid detection, specificity, and renewable capabilities in a single diagnostic solution marks a significant advance in pandemic preparedness.

bioengineering↗

bioRxiv2021

Native structure of the RhopH complex, a key determinant of malaria parasite nutrient acquisition

The RhopH complex is implicated in malaria parasites ability to invade and create new permeability pathways in host erythrocytes, but its mechanisms remain poorly understood. Here we enrich the endogenous RhopH complex in a native soluble form, comprising RhopH2, CLAG3.1 and RhopH3, directly from parasite cell lysates and determine its atomic structure using cryo electron microscopy, mass spectrometry, and the cryoID program. This first direct observation of an exported P. falciparum transmembrane protein--in a soluble, trafficking state and with atomic details of buried putative membrane-insertion helices--offers insights into assembly and trafficking of RhopH and other parasite-derived complexes to the erythrocyte membrane. Our study demonstrates the potential endogenous structural proteomics approach holds for elucidating the molecular mechanisms of hard-to-isolate complexes in their native, functional forms.

biophysics↗

bioRxiv2018

Different capsid-binding patterns of the β-herpesvirus-specific tegument protein pp150 (pM32/pUL32) in murine and human cytomegaloviruses

The phosphoprotein pp150 is a structurally, immunogenically, and regulatorily important capsid-associated tegument protein abundant in {beta}-herpesviruses including cytomegaloviruses (CMV), but absent in -herpesviruses and {Gamma}-herpesviruses. In human CMV (HCMV), bridging across each triplex and three adjacent major capsid proteins (MCPs) is a group of three pp150 subunits in a \"{bigtriangleup}\"-shaped fortifying configuration, 320 of which encase and stabilize the genome-containing capsid. Because murine CMV (MCMV) has been used as a model for HCMV pathogenesis and therapeutic studies, one might expect that pp150 and the capsid in MCMV and HCMV have similar structures. Here, by cryoEM and sub-particle reconstructions, we have obtained structures of MCMV capsid and pp150 at near atomic resolutions and built their atomic models. Surprisingly, the capsid-binding patterns of pp150 differ between HCMV and MCMV despite their highly similar capsid structures. In MCMV, pp150 is absent on triplex Tc and exists as a \"{Lambda}\"-shaped dimer on other triplexes, leading to only 260 groups of two pp150 subunits per capsid in contrast to 320 groups of three pp150 subunits encasing each HCMV capsid. Many more amino acids contribute to pp150-pp150 interactions in MCMV than in HCMV, making MCMV pp150 dimer inflexible thus incompatible to instigate triplex Tc-binding as observed in HCMV. While pp150 is essential in HCMV, pp150-deleted MCMV mutants remained viable though with attenuated infectivity and exhibiting defects in retaining viral genome. These results support targeting capsid proteins, but invalidate targeting pp150, when using MCMV as a model for HCMV pathogenesis and therapeutic studies.\n\nImportanceCMV infection is a leading viral cause of congenital birth defects and often responsible for life-threating complications in immunocompromised individuals like AIDS and post-organ transplantation patients. Absence of effective vaccines and potent drugs against CMV infections has motivated animal-based studies, mostly based on the mouse model with MCMV, both for understanding pathogenesis of CMV infections and for developing therapeutic strategies. Here, we present the first atomic structures of MCMV and show that the organization patterns of capsid-associated tegument protein pp150 between human and mouse CMV are different despite their highly similar capsid structures. Our functional studies demonstrate that deleting pp150 does not eliminate MCMV infection in contrast to pp150s essential role in HCMV infections. These results thus establish the validity to target capsid proteins, but raise concerns to target pp150, when using MCMV as HCMV model for pathogenesis and therapeutic studies.

microbiology↗