Adenovirus binding to its receptor visualised by cryo-EM

28-05-2019

Adenoviruses are to date the most commonly used vectors in human clinical trials using gene therapy for their ability to deliver therapeutic genes or to destroy cancer cells directly. The mechanism by which adenoviruses bind to the cell has just been elucidated by cryo-electron microscopy. This paves the way to next generation adenoviral vector development.

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More than 60 adenovirus (Ad) serotypes are known in humans. While they are able to cause different types of disease such as gastroenteritis or conjunctivitis, most of them have respiratory tropism. Although not strictly speaking a major public health problem, several serotypes such as Ad3, Ad7 Ad11 and Ad14 (subgroup B2-Ads) investigated in this study, were recently reported in an outbreak. Eleven out of the 35 young patients died of adenovirus infection in New Jersey in November 2018. Besides this, adenoviruses are the most commonly used vectors in human clinical trials, where they are used as oncolytic vectors.  To this goal, adenoviruses are modified to replicate only in the cancer cells. Numerous clinical trials are underway in the United States and Europe, offering great hope for new anti-tumour strategies.

Like all viruses, Ads must replicate in a cell and therefore their binding to a cellular receptor is a critical step of infection. Adenoviral binding involves the distal ‘knob’ domain of an elongated protein called the fibre (Figure 1). Work to identify the long elusive subgroup B2-Ads receptor lead to the successful identification of desmoglein 2 (DSG2) [1]. A comprehensive biochemical study using the PSB facilities (MALLS, AUC, mass spec. and electron microscopy) was then undertaken to map the minimal domain required for this interaction [2]. This led to a 96 kDa complex made of one trimeric adenoviral fibre-knob in complex with two out of the four cadherin domains of the DSG2 ectodomain. Attempts to solve the structure at the atomic level using X-ray crystallography were unsuccessful as diffraction never went below 6 angstrom.

Artist view of an adenoviral-like particle binding to its cellular receptor in red.

Figure 1. Artist view of an adenoviral-like particle binding to its cellular receptor in red. The distal trimeric globular ‘knob’ of the ‘antenna-like’ adenoviral fibre protein is critical for this interaction (adapted from Fender et al., Nat. Biotech. 15, 52-56, 1997).

Until recently, using cryo-electron microscopy (cryo-EM) to solve the atomic structure of such a small complex seemed inconceivable. Thanks to advances in technology, this complex has now been solved at the Titan Krios microscope CM01 using a phase-plate. This structure is one of the smallest, if not the smallest, complexes solved by cryo-EM to date.  In particular, the data revealed an unusual mode of binding in which two monomers of the trimeric fibre-knob (the distal part of the adenoviral protein interacting with the receptor) bind with two independent cadherin domains of DSG2 thus forming a non-symmetrical complex (Figure 2). Moreover, key amino-acids involved in the interaction were identified and it has been shown that a single amino acid substitution in the adenoviruses fibre head was sufficient to completely abolish the receptor binding.

Ribbon-structure of the trimeric adenovirus of type 3 fibre-knob (in gold, red and blue) bound to two cadherin domains of the DSG2 receptor (in orange).

Figure 2. Ribbon-structure of the trimeric adenovirus of type 3 fibre-knob (in gold, red and blue) bound to two cadherin domains of the DSG2 receptor (in orange). Note that two monomers of the fibre knob (gold and blue) interact with two different cadherin domains of DSG2.

This discovery opens the way to the rational design of adenoviral inhibitors and also to a retargeting of oncolytic adenoviral vectors to tumours. The latter project is already in progress thanks to the support of the ANR (‘Ad-Cadh’ funding - ANR-189-CE11-0001).

 

Principal publication and authors
CryoEM structure of adenovirus type 3 fibre with desmoglein 2 shows an unusual mode of receptor engagement, E. Vassal-Stermann (a), G. Effantin (a), C. Zubieta (b), W. Burmeister (a), F. Iseni (c), H. Wang (d), A Lieber (d), G. Schoehn (a) & P. Fender (a), Nature Communications 10, 1181 (2019); doi: 10.1038/s41467-019-09220-y.
(a) Institut de Biologie Structurale (IBS), Université Grenoble Alpes, CNRS, CEA, Grenoble (France)
(b) Laboratoire de Physiologie Cellulaire et Végétale, Biosciences and Biotechnology Institute of Grenoble, UMR5168, CNRS/CEA/INRA/UGA, Grenoble (France)
(c) Unité de Virologie, Institut de Recherche Biomédicale des Armées, Brétigny-sur-Orge (France)
(d) Department of Medicine, Division of Medical Genetics, University of Washington, Seattle (USA)

 

References
[1] H. Wang, Z.Y. Li, Y. Liu, J. Persson, et al., Nat. Med. 17, 96–104 (2011).
[2] E. Vassal-Stermann, M. Mottet, C. Ducournau, F. Iseni et al., Scientific Reports 8, 8381 (2018).