Penton-Dodecahedral Particles Trigger Opening of Intercellular Junctions and Facilitate Viral Spread during Adenovirus Serotype 3 Infection of Epithelial Cells

Zhuo-Zhuang Lu,Hongjie Wang,Pascal Fender,Zongyi Li,André Lieber,Hua Cao,Yiyi Zhang,Michael J Imperiale

doi:10.1371/journal.ppat.1003718

Abstract

Human adenovirus serotypes Ad3, Ad7, Ad11, and Ad14 use the epithelial junction protein desmoglein 2 (DSG2) as a receptor for infection. During Ad infection, the fiber and penton base capsid proteins are produced in vast excess and form hetero-oligomers, called pentons. It has been shown for Ad3 that pentons self-assemble into penton-dodecahedra (PtDd). Our previous studies with recombinant purified Ad3 PtDd (produced in insect cells) showed that PtDd bind to DSG2 and trigger intracellular signaling resulting in the transient opening of junctions between epithelial cells. So far, a definitive proof for a function of Ad3 PtDd in the viral life cycle is elusive. Based on the recently published 3D structure of recombinant Ad3 PtDd, we generated a penton base mutant Ad3 vector (mu-Ad3GFP). mu-Ad3GFP is identical to its wild-type counterpart (wt-Ad3GFP) in the efficiency of progeny virus production; however, it is disabled in the production of PtDd. For infection studies we used polarized epithelial cancer cells or cell spheroids. We showed that in wt-Ad3GFP infected cultures, PtDd were released from cells before viral cytolysis and triggered the restructuring of epithelial junctions. This in turn facilitated lateral viral spread of de novo produced virions. These events were nearly absent in mu-Ad3GFP infected cultures. Our in vitro findings were consolidated in mice carrying xenograft tumors derived from human epithelial cancer cells. Furthermore, we provide first evidence that PtDd are also formed by another DSG2-interacting Ad serotype, the newly emerged, highly pathogenic Ad14 strain (Ad14p1). The central finding of this study is that a subgroup of Ads has evolved to generate PtDd as a strategy to achieve penetration into and dissemination in epithelial tissues. Our findings are relevant for basic and applied virology, specifically for cancer virotherapy.

Highlights

The main structural proteins of the icosahedral capsids of adenoviruses (Ads) are the hexon and penton base
We demonstrate that PtDd trigger the opening of epithelial junctions and support the lateral spread of Ad3 progeny virus in epithelial tissues
It has been reported that Ad2 penton base and fiber self-assemble into pentons

Summary

Introduction

The main structural proteins of the icosahedral capsids of adenoviruses (Ads) are the hexon and penton base. The C-terminal part of the fibers, the fiber knob, mediates the high affinity binding to a cellular receptor, while the RGD containing loops within the penton base interact with cellular integrins, a step that mediates cell entry of virions, except species B Ads. Most human Ad serotypes use CAR as a primary attachment receptor. We have shown that complete inhibition of Ad3 binding and infection requires the physical linkage and, most likely, a specific spatial constellation of at least two fiber knobs [1]. This specific mode of Ad3-fiber knobDSG2 interaction is functionally relevant for opening of junctions between epithelial cells [1,2]. The ability to open epithelial junctions appears to be important for Ad3 penetration into and spread within epithelial tissues [1,2,3]

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Conclusion