Cryo-electron tomography of mouse hepatitis virus: Insights into the structure of the coronavirion

Abstract

Coronaviruses are enveloped viruses containing the largest reported RNA genomes. As a result of their pleomorphic nature, our structural insight into the coronavirion is still rudimentary, and it is based mainly on 2D electron microscopy. Here we report the 3D virion structure of coronaviruses obtained by cryo-electron tomography. Our study focused primarily on the coronavirus prototype murine hepatitis virus (MHV). MHV particles have a distinctly spherical shape and a relatively homogenous size (Ϸ85 nm envelope diameter). The viral envelope exhibits an unusual thickness (7.8 ؎ 0.7 nm), almost twice that of a typical biological membrane. Focal pairs revealed the existence of an extra internal layer, most likely formed by the C-terminal domains of the major envelope protein M. In the interior of the particles, coiled structures and tubular shapes are observed, consistent with a helical nucleocapsid model. Our reconstructions provide no evidence of a shelled core. Instead, the ribonucleoprotein seems to be extensively folded onto itself, assuming a compact structure that tends to closely follow the envelope at a distance of Ϸ4 nm. Focal contact points and thread-like densities connecting the envelope and the ribonucleoprotein are revealed in the tomograms. Transmissible gastroenteritis coronavirion tomograms confirm all the general features and global architecture observed for MHV. We propose a general model for the structure of the coronavirion in which our own and published observations are combined.coronaviruses ͉ enveloped viruses ͉ plus-stranded RNA viruses ͉ transmissible gastroenteritis coronavirus

Replication of coronavirus

“…Farklı yapısal özellikleri inceleyen çalışmalara göre coronovirüsler farklı büyüklüklerde (80-120 nm) ve şekillerde küreye benzer şekilde pleomorfik (farklı morfolojik biçimlerde) partiküller oluşturmaktadırlar (5,22,24,25).…”

“…S proteinleri homotrimerler oluşturarak Coronavirüslere ismini veren güneş şeklindeki morfolojilerin oluşmasını sağlarlar (19,24,28). S proteinleri C-terminal transmembran bölgeleri aracılığıyla virion zarına bağlanırlar ve aynı zamanda M proteinleriyle de etkileşim kurarlar (25).…”

Three‐Dimensional Imaging of Biomaterials with Electron Tomography

“…The allowable electron dose greatly depends on the type of sample and type of fixation. For a cryoimmobilized frozen-hydrated specimen, the allowable dose is seriously limited by radiation damage [1]. For a chemically 36.1 Introduction 1305 An image taken with a TEM is essentially a two-dimensional projection of the specimen in the direction of the illuminating electron beam.…”

“…Coronaviruses (Figure 36.3) are an example of pleomorphic viruses: they are not symmetrical, contain a lipid membrane, and show variations in size. To investigate their structure in 3D, cryo-ET is the methodology of choice [1]. This is also the case for many other viruses, such as herpes simplex virus, human immunodeficiency virus (HIV), and influenza [81][82][83].…”

Unraveling the Packaging Mechanism of Coronavirus Ribonucleocapsid

“…The pleomorphic nature of the coronavirus particle has hampered the effort to obtain high-resolution virion image at atomic resolution. The inability to observe a well-structured RNP layer inside the SARS-CoV particle and only short coiled fragments of RNP in MHV in the cryo-EM reconstructions strongly suggests that the helical nucleocapsid is a very flexible structure that extensively twists and folds upon itself [3]. Extensive biophysical studies suggest that all CoV N proteins share the same modular organization containing two structured domains, the N-terminal domain (NTD) and the C-terminal domain (CTD), surrounded by three intrinsically disordered regions (IDR) called the N-terminal arm (N-arm), the central linker region (LKR), and the C-terminal tail (C-tail), respectively [4].…”