Assembly and architecture of the EBV B cell entry triggering complex.

Karthik Sathiyamoorthy,Wei Jiang,Cynthia L Rowe,Jia Chen,Jiansen Jiang,Britta S Möhl,Elizabeth D Mellins,Richard Longnecker,Yao Xiong Hu,Theodore S Jardetzky,Z Hong Zhou

doi:10.1371/journal.ppat.1004309

Abstract

Epstein-Barr Virus (EBV) is an enveloped double-stranded DNA virus of the gammaherpesvirinae sub-family that predominantly infects humans through epithelial cells and B cells. Three EBV glycoproteins, gH, gL and gp42, form a complex that targets EBV infection of B cells. Human leukocyte antigen (HLA) class II molecules expressed on B cells serve as the receptor for gp42, triggering membrane fusion and virus entry. The mechanistic role of gHgL in herpesvirus entry has been largely unresolved, but it is thought to regulate the activation of the virally-encoded gB protein, which acts as the primary fusogen. Here we study the assembly and function of the reconstituted B cell entry complex comprised of gHgL, gp42 and HLA class II. The structure from negative-stain electron microscopy provides a detailed snapshot of an intermediate state in EBV entry and highlights the potential for the triggering complex to bring the two membrane bilayers into proximity. Furthermore, gHgL interacts with a previously identified, functionally important hydrophobic pocket on gp42, defining the overall architecture of the complex and playing a critical role in membrane fusion activation. We propose a macroscopic model of the initiating events in EBV B cell fusion centered on the formation of the triggering complex in the context of both viral and host membranes. This model suggests how the triggering complex may bridge the two membrane bilayers, orienting critical regions of the N- and C- terminal ends of gHgL to promote the activation of gB and efficient membrane fusion.

Highlights

Epstein Barr Virus (EBV) or Human Herpesvirus 4 (HHV-4) is a gammaherpesvirus that is ubiquitous in humans and predominantly infects host epithelial and B cells, in which it establishes long-term latency
Our experiments show that the gp42 hydrophobic pocket (HP) interacts with gHgL and that mutations of the predicted HP contact residues on gHgL are detrimental for fusion
Constraints imposed by the triggering complex architecture relative to its predicted membrane anchors highlight a close approach and potential deformation of both viral and host membranes affected by human leukocyte antigen (HLA) receptor binding as a prerequisite to viral entry

Summary

Introduction

Epstein Barr Virus (EBV) or Human Herpesvirus 4 (HHV-4) is a gammaherpesvirus that is ubiquitous in humans and predominantly infects host epithelial and B cells, in which it establishes long-term latency. It is an oncogenic virus associated with a wide array of human tumors including epithelial cell tumors such as nasopharyngeal and gastric carcinomas, and lymphoid malignancies like Hodgkin and Burkitt lymphoma. Viral membrane fusion is a requisite step for infection for all lipid bilayer encased viruses, such as the herpesviruses, and requires one or several virus-encoded glycoproteins that orchestrate the merging of viral and host membranes in a step-wise manner [2]. The gp protein forms stable, high affinity complexes with the gHgL complex [12], and binds to human leukocyte antigen (HLA) class II [13] which acts as the triggering receptor for EBV entry into B cells [4]

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Conclusion