# In-vitro analysis of HSV-1 membrane fusion mechanism

> **NIH NIH R21** · TUFTS UNIVERSITY BOSTON · 2021 · $254,790

## Abstract

PROJECT SUMMARY/ABSTRACT
Herpes Simplex virus type 1 (HSV-1) requires four glycoproteins for cell entry and membrane fusion – gB, gD,
gH, and gL – in addition to a cellular receptor. A thorough knowledge of membrane fusion mechanisms is
essential for understanding how herpesviruses penetrate cells and finding ways to inhibit this process. Due to
the complexity of this process, many fundamental questions endure despite decades of research. Moreover,
traditional entry assays rely on downstream reporters and do not measure fusion directly. Studies of cell-cell
fusion of uninfected receptor-bearing cells expressing the four essential HSV entry glycoproteins have yielded
much of the current mechanistic knowledge of membrane fusion, including the prevalent regulatory cascade
model. However, the cell-cell fusion system, while informative, fails to capture the proper context of virus-cell
fusion. Accurate, systematic dissection of the HSV-mediated membrane fusion mechanism thus requires an
experimentally tractable system that enables direct visualization and kinetic measurements of the viral fusion.
The goal of this exploratory proposal is to reconstitute HSV-1 fusion in vitro and to characterize it at a single-
virion level by imaging fusion of individual virions with fluid, supported lipid bilayers using total internal reflection
microscopy. This approach will be used to visualize different stages in fusion, measure their kinetic
parameters, identify kinetic intermediates, and correlate them with structural rearrangements in gB. In addition
to HSV-1, the proposed studies will employ Vesicular Stomatitis Virus (VSV) virions lacking the native fusogen
G and pseudotyped with HSV-1 entry glycoproteins gB, gD, gH, and gL (VSVDG-BHLD). The use of this
simplified, “bare-bones” system will benefit this work by excluding the potential effects of the 12 other envelope
proteins. In-vitro reconstitution of fusion and its characterization at a single particle level will address the
lingering questions in HSV-mediated membrane fusion mechanism, with the ultimate goal of reconstructing the
HSV-1-mediated fusion pathway more fully.

## Key facts

- **NIH application ID:** 10230779
- **Project number:** 1R21AI160821-01
- **Recipient organization:** TUFTS UNIVERSITY BOSTON
- **Principal Investigator:** Ekaterina Heldwein
- **Activity code:** R21 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $254,790
- **Award type:** 1
- **Project period:** 2021-03-17 → 2023-02-28

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10230779

## Citation

> US National Institutes of Health, RePORTER application 10230779, In-vitro analysis of HSV-1 membrane fusion mechanism (1R21AI160821-01). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10230779. Licensed CC0.

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