The Membrane-Proximal Region (MPR) of Herpes Simplex Virus gB Regulates Association of the Fusion Loops with Lipid Membranes

ABSTRACT Glycoprotein B (gB), gD, and gH/gL constitute the fusion machinery of herpes simplex virus (HSV). Prior studies indicated that fusion occurs in a stepwise fashion whereby the gD/receptor complex activates the entire process, while gH/gL regulates the fusion reaction carried out by gB. Trime...

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Autores principales:	Spencer S. Shelly, Tina M. Cairns, J. Charles Whitbeck, Huan Lou, Claude Krummenacher, Gary H. Cohen, Roselyn J. Eisenberg
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2012
Materias:	Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/7d861036838f4b77a65ff46f03295d0d
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spelling	oai:doaj.org-article:7d861036838f4b77a65ff46f03295d0d2021-11-15T15:39:11ZThe Membrane-Proximal Region (MPR) of Herpes Simplex Virus gB Regulates Association of the Fusion Loops with Lipid Membranes10.1128/mBio.00429-122150-7511https://doaj.org/article/7d861036838f4b77a65ff46f03295d0d2012-12-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.00429-12https://doaj.org/toc/2150-7511ABSTRACT Glycoprotein B (gB), gD, and gH/gL constitute the fusion machinery of herpes simplex virus (HSV). Prior studies indicated that fusion occurs in a stepwise fashion whereby the gD/receptor complex activates the entire process, while gH/gL regulates the fusion reaction carried out by gB. Trimeric gB is a class III fusion protein. Its ectodomain of 773 amino acids contains a membrane-proximal region (MPR) (residues 731 to 773) and two fusion loops (FLs) per protomer. We hypothesized that the highly hydrophobic MPR interacts with the FLs, thereby masking them on virions until fusion begins. To test this hypothesis, we made a series of deletion, truncation, and point mutants of the gB MPR. Although the full-length deletion mutants were expressed in transfected cells, they were not transported to the cell surface, suggesting that removal of even small stretches of the MPR was highly detrimental to gB folding. To circumvent this limitation, we used a baculovirus expression system to generate four soluble proteins, each lacking the transmembrane region and cytoplasmic tail. All retained the FLs and decreasing portions of the MPR [gB(773t) (gB truncated at amino acid 773), gB(759t), gB(749t), and gB(739t)]. Despite the presence of the FLs, all were compromised in their ability to bind liposomes compared to the control, gB(730t), which lacks the MPR. We conclude that residues 731 to 739 are sufficient to mask the FLs, thereby preventing liposome association. Importantly, mutation of two aromatic residues (F732 and F738) to alanine restored the ability of gB(739t) to bind liposomes. Our data suggest that the MPR is important for modulating the association of gB FLs with target membranes. IMPORTANCE To successfully cause disease, a virus must infect host cells. Viral infection is a highly regulated, multistep process. For herpesviruses, genetic material transfers from the virus to the target cell through fusion of the viral and host cell lipid membranes. Here, we provide evidence that the ability of the herpes simplex virus (HSV) glycoprotein B (gB) fusion protein to interact with the host membrane is regulated by its membrane-proximal region (MPR), which serves to cover or shield its lipid-associating moieties (fusion loops). This in turn prevents the premature binding of gB with host cells and provides a level of regulation to the fusion process. These findings provide important insight into the complex regulatory steps required for successful herpesvirus infection.Spencer S. ShellyTina M. CairnsJ. Charles WhitbeckHuan LouClaude KrummenacherGary H. CohenRoselyn J. EisenbergAmerican Society for MicrobiologyarticleMicrobiologyQR1-502ENmBio, Vol 3, Iss 6 (2012)
institution	DOAJ
collection	DOAJ
language	EN
topic	Microbiology QR1-502
spellingShingle	Microbiology QR1-502 Spencer S. Shelly Tina M. Cairns J. Charles Whitbeck Huan Lou Claude Krummenacher Gary H. Cohen Roselyn J. Eisenberg The Membrane-Proximal Region (MPR) of Herpes Simplex Virus gB Regulates Association of the Fusion Loops with Lipid Membranes
description	ABSTRACT Glycoprotein B (gB), gD, and gH/gL constitute the fusion machinery of herpes simplex virus (HSV). Prior studies indicated that fusion occurs in a stepwise fashion whereby the gD/receptor complex activates the entire process, while gH/gL regulates the fusion reaction carried out by gB. Trimeric gB is a class III fusion protein. Its ectodomain of 773 amino acids contains a membrane-proximal region (MPR) (residues 731 to 773) and two fusion loops (FLs) per protomer. We hypothesized that the highly hydrophobic MPR interacts with the FLs, thereby masking them on virions until fusion begins. To test this hypothesis, we made a series of deletion, truncation, and point mutants of the gB MPR. Although the full-length deletion mutants were expressed in transfected cells, they were not transported to the cell surface, suggesting that removal of even small stretches of the MPR was highly detrimental to gB folding. To circumvent this limitation, we used a baculovirus expression system to generate four soluble proteins, each lacking the transmembrane region and cytoplasmic tail. All retained the FLs and decreasing portions of the MPR [gB(773t) (gB truncated at amino acid 773), gB(759t), gB(749t), and gB(739t)]. Despite the presence of the FLs, all were compromised in their ability to bind liposomes compared to the control, gB(730t), which lacks the MPR. We conclude that residues 731 to 739 are sufficient to mask the FLs, thereby preventing liposome association. Importantly, mutation of two aromatic residues (F732 and F738) to alanine restored the ability of gB(739t) to bind liposomes. Our data suggest that the MPR is important for modulating the association of gB FLs with target membranes. IMPORTANCE To successfully cause disease, a virus must infect host cells. Viral infection is a highly regulated, multistep process. For herpesviruses, genetic material transfers from the virus to the target cell through fusion of the viral and host cell lipid membranes. Here, we provide evidence that the ability of the herpes simplex virus (HSV) glycoprotein B (gB) fusion protein to interact with the host membrane is regulated by its membrane-proximal region (MPR), which serves to cover or shield its lipid-associating moieties (fusion loops). This in turn prevents the premature binding of gB with host cells and provides a level of regulation to the fusion process. These findings provide important insight into the complex regulatory steps required for successful herpesvirus infection.
format	article
author	Spencer S. Shelly Tina M. Cairns J. Charles Whitbeck Huan Lou Claude Krummenacher Gary H. Cohen Roselyn J. Eisenberg
author_facet	Spencer S. Shelly Tina M. Cairns J. Charles Whitbeck Huan Lou Claude Krummenacher Gary H. Cohen Roselyn J. Eisenberg
author_sort	Spencer S. Shelly
title	The Membrane-Proximal Region (MPR) of Herpes Simplex Virus gB Regulates Association of the Fusion Loops with Lipid Membranes
title_short	The Membrane-Proximal Region (MPR) of Herpes Simplex Virus gB Regulates Association of the Fusion Loops with Lipid Membranes
title_full	The Membrane-Proximal Region (MPR) of Herpes Simplex Virus gB Regulates Association of the Fusion Loops with Lipid Membranes
title_fullStr	The Membrane-Proximal Region (MPR) of Herpes Simplex Virus gB Regulates Association of the Fusion Loops with Lipid Membranes
title_full_unstemmed	The Membrane-Proximal Region (MPR) of Herpes Simplex Virus gB Regulates Association of the Fusion Loops with Lipid Membranes
title_sort	membrane-proximal region (mpr) of herpes simplex virus gb regulates association of the fusion loops with lipid membranes
publisher	American Society for Microbiology
publishDate	2012
url	https://doaj.org/article/7d861036838f4b77a65ff46f03295d0d
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The Membrane-Proximal Region (MPR) of Herpes Simplex Virus gB Regulates Association of the Fusion Loops with Lipid Membranes

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