The Amphipathic Helix of Adenovirus Capsid Protein VI Contributes to Penton Release and Postentry Sorting

Abstract

Nuclear delivery of the adenoviral genome requires that the capsid cross the limiting membrane of the endocytic compartment and traverse the cytosol to reach the nucleus. This endosomal escape is initiated upon internalization and involves a highly coordinated process of partial disassembly of the entering capsid to release the membrane lytic internal capsid protein VI. Using wild-type and protein VI-mutated human adenovirus serotype 5 (HAdV-C5), we show that capsid stability and membrane rupture are major determinants of entry-related sorting of incoming adenovirus virions. Furthermore, by using electron cryomicroscopy, as well as penton-and protein VI-specific antibodies, we show that the amphipathic helix of protein VI contributes to capsid stability by preventing premature disassembly and deployment of pentons and protein VI. Thus, the helix has a dual function in maintaining the metastable state of the capsid by preventing premature disassembly and mediating efficient membrane lysis to evade lysosomal targeting. Based on these findings and structural data from cryo-electron microscopy, we suggest a refined disassembly mechanism upon entry. IMPORTANCEIn this study, we show the intricate connection of adenovirus particle stability and the entry-dependent release of the membrane-lytic capsid protein VI required for endosomal escape. We show that the amphipathic helix of the adenovirus internal protein VI is required to stabilize pentons in the particle while coinciding with penton release upon entry and that release of protein VI mediates membrane lysis, thereby preventing lysosomal sorting. We suggest that this dual functionality of protein VI ensures an optimal disassembly process by balancing the metastable state of the mature adenovirus particle.A denoviruses (AdVs) are nonenveloped, double-stranded DNA viruses that assemble in the nuclei of productively infected cells and are released at the end of the infection cycle into the extracellular milieu. Productive infection of new cells requires that the capsid follow a stepwise disassembly process upon entry that, if perfectly executed, results in highly efficient genome transfer to the nucleus (1, 2).The AdV virion is composed of 13 different polypeptides that form an icosahedral capsid encompassing the genome-containing viral core. The capsid is mainly composed of trimeric hexons building up the facets of the capsid. Pentons and fibers are located at each of the 12 vertices, where they form a pentameric penton base from which the trimeric fiber molecule elongates (3-5). In addition the capsid is stabilized via the cement proteins IIIa, VI, VIII, and IX. The capsid encloses the viral core, with the viral genome organized into chromatin through association with the major core protein VII and proteins V, X, TP, and IVa2 (3-5). Following (or concomitant with) capsid assembly, several of the virion proteins undergo proteolytic processing by the virion-incorporated adenoviral proteinase (AVP) (6). This process of virus maturation is essential to render ne...

Imaging the adenovirus infection cycle

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Model for the architecture of caveolae based on a flexible, net-like assembly of Cavin1 and Caveolin discs

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Roles of secretory glycoproteins in particle formation of Flaviviridae viruses

“…Amphipathic helices of the influenza virus matrix protein M1 are crucial for the organization of virion structures, 61 whereas the amphipathic helix of adenovirus capsid protein 4 plays an important role in the release of viral capsids via endosomal membrane lysis. 62 These data suggest that some structural proteins other than E rns are involved in particle formation and transport via amphipathic helices. By contrast, the viral secretory glycoprotein of reovirus, FAST, is known to affect cell fusion by altering membrane curvature.…”

“…In other viruses, amphipathic helices of viral proteins are also reported to be involved in the formation or release of viral particles. Amphipathic helices of the influenza virus matrix protein M1 are crucial for the organization of virion structures, whereas the amphipathic helix of adenovirus capsid protein 4 plays an important role in the release of viral capsids via endosomal membrane lysis . These data suggest that some structural proteins other than E rns are involved in particle formation and transport via amphipathic helices.…”