EPHRIN-B2: A RECEPTOR FOR HENIPAVIRUSES

Ephrin-B2 is a 40 kDa transmembrane protein that belongs to the Ephrin ligand family and binds the receptors EphB2, B3, and B4. The involvement of Ephrin-B2 in a range of physiological systems has been described, including vascular, lymphatic, neuronal, and renal development, neurotransmission, synaptic plasticity, and tumor metastasis.^1-5 Ephrin-B2 and its cognate receptors are expressed in complementary fashion on adjacent cells. Upon ligation, both Ephrin-B2 and Eph receptors initiate signals in their respective cells. The combination of forward and reverse signaling is central to the tissue development and remodeling functions of Ephrin and Eph proteins.⁶

The Hendra and Nipah viruses (HeV and NiV) are emergent members of the Paramyxoviridae family. The first cases were detected in Australia in 1994-95 and Malaysia in 1998-99.^7,8 The Henipavirus genus was defined for these two viruses which are more closely related to each other than to other paramyxoviruses.⁹ Henipaviruses have been transmitted to humans from a variety of infected animals as well as from other humans.¹⁰ Henipavirus infection produces a high mortality rate in humans due to severe respiratory disease and febrile encephalitis.^7,8 The primary targets of Henipaviruses are endothelial cells, which undergo cell-cell fusion and syncytia formation.¹¹ Henipavirus interactions with target cells are mediated by its 75 kDa attachment (G) and 70 kDa fusion (F) proteins, both of which are required for membrane fusion.^12,13

Recently Ephrin-B2 has been identified as a functional cellular receptor for Hendra and Nipah viruses (Figure 1). Bonaparte et al. 14 compared the gene expression profile of two cell lines that are differentially permissive to infection by HeV and NiV. They selected 21 candidate genes that were expressed at significantly higher levels in the permissive compared to the nonpermissive cells. Transfection of ten of these genes into nonpermissive cells identified which candidates conferred susceptibility to viral fusion. Only those cells transfected with Ephrin-B2 became fusion competent. Fusion was completely inhibited by soluble Ephrin-B2 and a peptide derived from NiV F protein. Direct interaction of Ephrin-B2 and G protein from both HeV and NiV was demonstrated by ELISA, co-immunoprecipitation from cell lysates, and surface plasmon resonance (Kd = 1 nM). Soluble Ephrin-B2 also completely blocked intact HeV and NiV infection of African green monkey kidney (Vero) cells.

Figure 1. HeV and NiV interact with Ephrin-B2 on endothelial cells, leading to the formation of multinucleated giant cells in a process that requires Viral attachment (G) and Fusion (F) proteins. After infection, viral components are present in cytoplasmic inclusion bodies and as extracellular infective virions.

Negrete et al. 15 arrived at the same conclusion by searching for binding interactions between the NiV G protein with target cell lines. Co-immunoprecipitation experiments identifed a binding protein found in extracts of permissive, but not nonpermissive cells. Tryptic digestion and mass spectrometry led to identification of the binding protein as Ephrin-B2. Specificity of NiV G protein for Ephrin-B2 was demonstrated by ELISA and by competition on permissive cells for soluble NiV G protein binding. In addition, transfection of Ephrin-B2 but not Ephrin-B1 into nonpermissive cells conferred NiV infection sensitivity. Fusion was blocked by soluble Ephrin-B2 or EphB4 but not by Ephrin-B1. Viral infectivity was neutralized by a polyclonal antibody raised against NiV F and G proteins and by soluble Ephrin-B2 but not by Ephrin-B1.

It is possible that other molecules may contribute to HeV and NiV fusion permissiveness and that other cell types may use different or additional receptors. However, there is a strong correlation between Ephrin-B2 expression patterns and the tropism of Henipaviruses. Ephrin-B2 is also very highly conserved between species, which is consistent with the broad range of animals susceptible to infection. This recent work highlights a novel function for Ephrin-B2 and contributes to the understanding of HeV and NiV pathogenesis.

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