Unique structures in a tumor herpesvirus revealed by cryo-electron tomography and microscopy

Abstract

Gammaherpesviruses, including the human pathogens Epstein-Barr virus and Kaposi's sarcomaassociated herpesvirus, are causative agents of lymphomas and other malignancies. The structural characterization of these viruses has been limited due to difficulties in obtaining adequate amount of virion particles. Here we report the first three-dimensional structural characterization of a whole gammaherpesvirus virion by an emerging integrated approach of the technique of cryo-electron tomography combined with single-particle cryo-electron microscopy, using murine gammaherpesvirus-68 (MHV-68) as a model system. We found that the MHV-68 virion consists of distinctive envelope and tegument compartments, and a highly conserved nucleocapsid. Two layers of tegument are identified: an inner tegument layer tethered to the underlying capsid and an outer, flexible tegument layer conforming to the overlying, pleomorphic envelope, consistent with the sequential viral tegumentation process inside host cells. Surprisingly, comparison of the MHV-68 virion and capsid reconstructions shows that the interactions between the capsid and inner tegument proteins are completely different from those observed in alpha and betaherpesviruses. These observations support the notion that inner layer tegument across different subfamilies of herpesviruses have evolved significantly to confer specific characteristics related to viral-host interactions, in contrast to a highly conserved capsid for genome encapsidation and protection.

Organization of Capsid-Associated Tegument Components in Kaposi's Sarcoma-Associated Herpesvirus

“…1C). The weak intensity of these tegument densities explains why we failed to detect them previously in a low-resolution cryo-EM reconstruction of murine herpesvirus 68 (MHV-68) virion with only a few hundred particles (5).…”

“…Like all herpesviruses, the KSHV virion is composed of a double-stranded, linear DNA genome encased within an icosahedral capsid, which in turn is wrapped in a proteinaceous layer, called the tegument and, last, a lipid bilayer membrane called the envelope. Although the tegument compartment of herpesvirus is pleomorphic and has no distinguishable substructures, it is roughly divided into outer tegument and inner tegument layers (5,6). Upon cell entry, the outer tegument proteins are released into the host cell cytoplasm where some of them perform functions needed early in the infection process, while the inner tegument proteins remain associated with the nucleocapsid and participate in its transport to the nucleus via the microtubule/dynein motor system (7)(8)(9)(10)(11)(12).…”

Role of tegument proteins in herpesvirus assembly and egress

“…1). An infectious virion contains a doublestranded DNA genome, an icosahedral capsid shell, a thick, proteinaceous tegument compartment, and a lipid bilayer envelope spiked with glycoproteins (Rixon, 1993;Roizman and Pellett, 2001;Liu and Zhou, 2006;Dai et al, 2008). As a unique structure of herpesviruses, the tegument plays important roles in multiple aspects of the viral life cycle, including translocation of nucleocapsids into the nucleus as well as virion assembly and egress (Subak-Sharpe and Dargan, 1998;Fuchs et al, 2002a).…”

Role of RRV ORF52 in Virion Maturation

“…This ordering is significant enough to be further divided into "inner" and "outer" tegument layers, which are seen with cryo-electron tomography ( Figure 1-4) (62). While the research does not strictly define the terms "inner" and "outer," the former generally refers to tegument proteins more tightly associated with the capsid and more resistant to Triton X-100 detergent treatment (336 (239), and I will discuss them separately at the end of this section.…”

“…Earlier studies divide the tegument into "inner" and "outer" portions, referring to biochemical or structural evidence describing the relative affinity and proximity of specific sets of proteins to the surface of the capsid (62,239,336,356,357). When one of these tegument proteins is missing, it affects the ability of other proteins to be added or perhaps remain stably attached to the particle.…”