Biology subjects

Hecksel, C. W.

Publications and source records attributed to Hecksel, C. W..

3 recordsLinked to original sources

bioRxiv2025

Cryogenic Electron Tomography Redefines Herpesvirus Capsid Assembly Intermediates Inside the Cell Nucleus.

Herpesviruses encapsulate their double-stranded DNA (dsDNA) genomes within an icosahedral nucleocapsid formed in the infected cell nucleus. Four biochemically purified nucleocapsids have been characterized, but their roles in herpesvirus replication remain controversial. The status of the capsid vertex-specific component (CVSC), essential for capsid stability and dsDNA packaging and retention, is also unclear. By integrating cryogenic focused ion beam milling with electron tomography and subtomogram averaging, we derived atomic models for all protein components, including the CVSC, across different herpesvirus capsid types within infected cell nuclei. Focused classification of pentonal vertex densities revealed differences in CVSC occupancy between genome-filled capsids and capsids lacking dsDNA, highlighting structural heterogeneity and providing insight into distinct capsid assembly stages in situ. These intra-nuclear findings redefine the maturation model of herpesvirus capsid assembly, advancing the understanding of herpesvirus replication, and demonstrate the effectiveness of in situ electron imaging by studying virus assembly within host cells.

microbiology↗

bioRxiv2020

Assembly intermediates of orthoreovirus captured in the cell

Traditionally molecular assembly pathways for viruses have been inferred from high resolution structures of stable intermediates purified in vitro, and from low resolution images of cell sections as well as from genetic approaches including conditionally lethal mutants. Here, we directly visualise a previously unsuspected single shelled icosahedral intermediate for a mammalian orthoreovirus, in addition to the expected virions, in cryo-preserved infected cells by cryo-electron tomography of cellular lamellae1,2. Particle classification and averaging yielded structures at resolutions as high as 5.6 [A], sufficient to identify secondary structural elements and place known molecular structures, allowing us to produce an atomic model of the intermediate, comprising 120 copies of protein {lambda}1 and 120 copies of {sigma}2. This {lambda}1 shell is in a collapsed form compared to the mature virions, with the molecules pushed inwards at the icosahedral 5-folds by ~100 [A]. This grossly indented shell, although produced by a mammalian reovirus, is reminiscent of the first assembly intermediate of prokaryotic dsRNA viruses belonging to a different virus family3, adding weight to the supposition that these diverse viruses share a common ancestor, and suggesting mechanisms for the assembly of viruses of the Reoviridae. Such methodology holds enormous promise for the dissection of the replication cycle of many viruses.

microbiology↗

bioRxiv2020

Multi-Scale 3D Cryo-Correlative Microscopy for Vitrified Cells

Three-dimensional (3D) visualization of vitrified cells can uncover structures of subcellular complexes without chemical fixation or staining. Here, we present a pipeline integrating three imaging modalities to visualize the same specimen at cryogenic temperature at different scales: cryo-fluorescence confocal microscopy, volume cryo-focused ion beam scanning electron microscopy, and transmission cryo-electron tomography. Our proof-of-concept benchmark revealed the 3D distribution of organelles and subcellular structures in whole heat-shocked yeast cells, including the ultrastructure of protein inclusions that recruit fluorescently-labelled chaperone Hsp104. Since our workflow efficiently integrates imaging at three different scales and can be applied to other types of cells, it could be used for large-scale phenotypic studies of frozen-hydrated specimens in a variety of healthy and diseased conditions with and without treatments.

cell biology↗