Structural basis for the recognition of LDL-receptor family members by VSV glycoprotein

Jovan Nikolic,Pierre Legrand,Aurélie A Albertini,Yves Gaudin,Hélène Raux,Laura Belot

doi:10.1038/s41467-018-03432-4

Abstract

Vesicular stomatitis virus (VSV) is an oncolytic rhabdovirus and its glycoprotein G is widely used to pseudotype other viruses for gene therapy. Low-density lipoprotein receptor (LDL-R) serves as a major entry receptor for VSV. Here we report two crystal structures of VSV G in complex with two distinct cysteine-rich domains (CR2 and CR3) of LDL-R, showing that their binding sites on G are identical. We identify two basic residues on G, which are essential for its interaction with CR2 and CR3. Mutating these residues abolishes VSV infectivity even though VSV can use alternative receptors, indicating that all VSV receptors are members of the LDL-R family. Collectively, our data suggest that VSV G has specifically evolved to interact with receptor CR domains. These structural insights into the interaction between VSV G and host cell receptors provide a basis for the design of recombinant viruses with an altered tropism.

Highlights

Vesicular stomatitis virus (VSV) is an oncolytic rhabdovirus and its glycoprotein G is widely used to pseudotype other viruses for gene therapy
After 4 h of infection, BSR cells were labeled with an antibody directed against VSV nucleoprotein to visualize the infection and a GST-CRATTO550 to probe CR domain recognition by the surface displayed glycoprotein. f Labeling of G at the surface of BSR cells infected with VSV using fluorescent GST-CR1ATTO550, GST-CR2ATTO550, and GST-CR3ATTO550
Low-density lipoprotein receptor (LDL-R) has been demonstrated to be the major entry port of VSV and lentivirus pseudotyped by VSV G14

Summary

Results

LDL-R CR2 and CR3 bind G and neutralize viral infectivity. We have expressed individually each LDL-R CR domain in fusion with the glutathione S-transferase (GST) in Escherichia coli. After 20 min of incubation at 4 °C, the beads were washed and the associated proteins were analyzed by SDS/PAGE followed by Coomassie blue staining This revealed that only CR2 and CR3 domains are able to directly bind VSV G (Fig. 1b) at pH 8. The binding of Gth or VSV to fusion proteins GST-CR2 and GST-CR3 was pH dependent and no interaction was detected at pH 6. The cells expressing mutant G protein have a fusion phenotype similar to that of WT G (Fig. 5d). This confirms that the mutant glycoproteins are correctly folded and demonstrates a VSV Gmut

H LDLR CR3 88

Discussion

Methods