Self-assembly of the general membrane-remodeling protein PVAP into sevenfold virus-associated pyramids

Abstract

Viruses have developed a wide range of strategies to escape from the host cells in which they replicate. For egress some archaeal viruses use a pyramidal structure with sevenfold rotational symmetry. Virus-associated pyramids (VAPs) assemble in the host cell membrane from the virus-encoded protein PVAP and open at the end of the infection cycle. We characterize this unusual supramolecular assembly using a combination of genetic, biochemical, and electron microscopic techniques. By whole-cell electron cryotomography, we monitored morphological changes in virus-infected host cells. Subtomogram averaging reveals the VAP structure. By heterologous expression of PVAP in cells from all three domains of life, we demonstrate that the protein integrates indiscriminately into virtually any biological membrane, where it forms sevenfold pyramids. We identify the protein domains essential for VAP formation in PVAP truncation mutants by their ability to remodel the cell membrane. Self-assembly of PVAP into pyramids requires at least two different, in-plane and out-of-plane, protein interactions. Our findings allow us to propose a model describing how PVAP arranges to form sevenfold pyramids and suggest how this small, robust protein may be used as a general membrane-remodeling system. archaea | archeovirus | viral egress

Structure and assembly mechanism of virus-associated pyramids

“…Towards the end of the infection cycle, when the virions inside the cell have matured, all VAPs open approximately at the same stage of infection, much like the petals of a flower ( Fig. 1) (Fu et al 2010;Quax et al 2011;Daum et al 2014). This results in apertures of~150 nm in diameter, through which the virions can leave the cell (Fig.…”

“…1). After releasing the virions, the VAPs remain open, thus causing complete lysis of host cell (Bize et al 2009;Brumfield et al 2009;Fu et al 2010;Daum et al 2014).…”

“…Indeed, overexpression of SIRV2 or STIV1-derived PVAP in the archaea Sulfolobus acidocaldarius, S. solfataricus, the bacterium E. coli and the eukaryote Saccharomyces cerevisiae resulted in VAPs in all membranes (Snyder et al 2011;Daum et al 2014). In S. solfataricus, opening of the VAPs occurred after heterologous expression of both STIV1-and SIRV2-derived PVAP.…”

“…In S. solfataricus, opening of the VAPs occurred after heterologous expression of both STIV1-and SIRV2-derived PVAP. However, VAPs that had been expressed heterologously in E. coli never opened, even after incubating of the cells for up to 2 weeks at room temperature (Daum et al 2014). This indicates that VAP opening is not a spontaneous, independent process and suggests that a specific factor may have to be present to trigger the unfolding mechanism.…”

“…The expression of a single viral membrane protein is the only requirement for the formation of these pyramids, which can reach hundreds of nanometres in diameter Snyder et al 2011). Overexpression of this protein in archaeal, bacterial and eukaryotic model organisms has shown that self-assembly of the pyramidal structures is independent of the origin and type of the lipid membrane (Daum et al 2014). As such, the VAPs represent a universal membrane remodelling system with potential nano-biotechnological applications.…”