How does the herpes virus hijack our cells? Cryo-SXT can show us…

Estimates suggest that around two-thirds of the global population is infected with herpes simplex virus type 1 (HSV-1), the virus that causes cold sores and genital herpes. After the initial infection, the symptoms normally abate. However, the virus hides in the body in a latent form and can reactivate at a later date. 

The modus operandi of any virus is to penetrate the cells of another organism and take them over - turning them into factories that churn out more viral particles. HSV-1 and humans have evolved together for millions of years, so this particular virus is very good at invading and modifying our cells. A better understanding of how the virus works would enable us to develop better treatments and bring us closer to a vaccine that can prevent infection and transmission. We might also become better at re-purposing the virus as a cancer treatment or for gene therapy. An HSV-1 based therapy (T-VEC) was the first licensed anti-cancer virus, against melanoma, and it's also being developed as a gene delivery vector.

Herpes simplex virus infection. Wellcome Collection. CC0 1.0 Universal

HSV-1 is a large, enveloped DNA virus. It interacts with the nucleus and organelles - the machinery inside our cells - to produce virus particles in a complex multi-step process. Researchers commonly use transmission electron microscopy and fluorescence microscopy to study viruses like HSV-1, but these produce images that are only in 2D or have limited resolution. High-resolution 3D images would greatly improve our understanding of the changes the virus makes to the cell's components. Transmission electron microscopy and fluorescence microscopy imaging techniques can also require time-consuming sample processing - such as sectioning, staining and fixing - that can themselves cause changes to the cell.

Researchers from the University of Cambridge have been working closely with Diamond's B24 beamline to develop a new technique for 3D imaging of virus-infected cells. The first author on this paper, Kamal Nahas, is a joint PhD student with Diamond and the University of Cambridge.

The Cambridge team works with a modified version of HSV-1, engineered to fluoresce at different stages of the infection process, referred to as a "timestamp" reporter virus¹. That means they can capture a snapshot of what the cell looks like at different times in the infection and replication process. The cells are then flash frozen in liquid ethane, which preserves them in a near-native state.

B24 is a dedicated biology beamline with state-of-the-art equipment for Cryo Soft X-ray Tomography (Cryo-SXT), a technique ideally suited for high-resolution 3D imaging of cell structure. As soft X-rays pass through the sample cell, they are preferentially absorbed by carbon-rich biological structures. Gradually changing the angle at which the X-rays pass through the cell creates a series of images with structural information captured at different angles. These images are then reconstructed into tomograms - like a medical CT scan, but on a much smaller scale - to produce a 3D image volume.

For Dr Colin Crump, from the University of Cambridge, these experiments were his first experience of working at a synchrotron. He said

These experiments were proof of concept, demonstrating that high-resolution 3D images of cellular compartments within infected cells can be captured in a near-native state using Cryo-SXT.

Mitochondria are often referred to as the "powerhouses" of the cell, and the results showed marked changes during infection, with the mitochondria forming a network across the cell. A more short-lived effect is seen in the distribution of lipid droplets in the cell, which temporarily change their size during infection.

As Dr Stephen Graham from the University of Cambridge explains, one of the benefits of this technique is that it can quickly capture large amounts of data. He said:

To find out more about the B24 beamline or discuss potential applications, please contact Principal Beamline Scientist Maria Harkiolaki: [email protected].

1. Scherer KM et al. A fluorescent reporter system enables spatiotemporal analysis of host cell modification during herpes simplex virus-1 replication. Journal of Biological Chemistry 296:100236 (2021).